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TABLE OF CONTENTS

For VoLuME XXXIX, Number 4, October 1920

Micro-Technique. Suggestions for Methods and Apparatus, with five figures,

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TABLE OF CONTENTS

FOR VOLUME XXXIX, NUMBER 1, January 1920

Glaridacris catostomi gen. nov., sp. nov.: A Cestodarian Parasite, with Plates I

SUILCM OE Rat Diya RSC GODT aq acne ci Aaa MOON ai sal a ne RIG ahem Le SIRI SEARS

The Genera of the Enchytraeidae (Oligochaeta), by Paul S. Welch.............

An Ecological Study of the Algae of Some Sandhill Lakes, by Emma N. Andersen

aoe ani Wal en ete in Amey TONED AU Ui umn mrbene ya AN repie sm Mine EA URE Ae

Notes and Reviews: Leeches considered as Oligochaeta Modified for a Predatory

nile Hrevile wean bye PaySMIE Ls rae ecie ie Mente he) GUIS Leelee cra a ene RU ner NS) Sea

Wea uibesior the St NMouis vi cetiney yng kee L312 ly hale ture WN gal be Laat

RPE pOn On the Neu storbamiay aia unt ON ean tae tra Ahh alt SRT hh cag en Mies MPRA

ReHOn orien PECASUPCN cc. 4 meat e is Tirta sha tey by eget Nile) ahs A eMail LAGAN aia

as ie

Pe eae

eey 4)

TRANSACTIONS

American Microscopical Society

(Published in Quarterly Instalments)

Vol. XXXTX JANUARY, 1920 No. 1

GLARIDACRIS CATOSTOMI GEN. NOV., SP. NOV.:

A CESTODARIAN PARASITE

By A. R. CooPER

INTRODUCTION

In a preliminary paper Ward (1911) stated that he had found in

fish from the Illinois River a cestodarian tapeworm which showed

certain features common to the well known European genera, Caryo-

phyllaeus and Archigetes. ‘It resembles the former in the absence

of a caudal appendage and in the location chosen by the adult para-

site, viz., the intestine of a fish, whereas, so far as known in Europe,

Archigetes always possesses a tail and has been found only in the

body cavity of tubificid worms. In general appearance and structure

the American form resembles the European Archigetes very strongly.

It has a scolex of fixed form with prominent suckers or phyllidea and

also the musculature of Archigetes. The general arrangement of the

reproductive organs, especially the two rows of testes in the central

field, and the genital pores, correspond also closely to conditions in

Archigetes.”’ Much later the same writer (Ward and Whipple, 1918)

merely stated in his key to the Cestodaria that, as regards Archigetes,

“a form which undoubtedly belongs here has been described to me

as found in native earthworms.” Neither under Archigetes nor under

Caryophyllaeus does he make any further mention of the above form,

and concerning Amphilina says only: ‘Not yet reported from North

America but present.’”? Nor have I been able to locate any other

reference to members of the Cestodaria, sensu latu, having been

found on this continent up to date.

Before proceeding with the detailed account it mHeuld be men-

tioned as a matter of introduction that, apart from being evidently

6 A. R. COOPER

the first member of the group to be described from America, the spe-

cies to be dealt with here is of special interest in that it seems to

stand intermediate in the family, Caryophyllaeidae Liihe 1910,

between Archigetes and Caryophyllaeus. Excepting for the scolex,

however, which is quite similar at least in outward appearance to that

of Archigetes brachyurus Mrazek, it closely resembles the species of

Caryophyllaeus, of which three, namely, C. laticeps (Pallas), C. tuba

(Wagener) and C. fennicus Schneider, have been found in Europe,

and one, C. syrdarjensis Skrjabin, in Asia (Turkestan).

MATERIAL

The material for the present study was obtained at the Douglas

Lake Biological Station of the University of Michigan during the

summer of 1917 while the writer was paying particular attention to

the bothriocephalid cestodes of fishes. In all thirty-six specimens of

Catostomus commersonii (Lacépéde), the host species, were examined.

These fell into two lots as regards size: ten younger ones ranging in

length from 90 to 115mm. and twenty-six adults from 250 to 325mm.

The latter were caught in the trammel and fyke nets used in the lake

proper, while the former were seined out of Maple River which drains

the lake. No parasites belonging to the species described here were

met with in the younger hosts, but from two to at least sixty-three

were found in the stomachs and intestines of eleven of the adults.

The table shown on page 7 gives their number, distribution and kind

in nine of the hosts, the exact numbers not having been recorded

for the other two fish.

From this it is seen that the degree of infestation of the host is

comparatively small. Whereas the number of adults met with was

quite limited, larvae were very plentiful when present at all. In

situ all of the adults and most of the larvae were found free in the

stomach or intestine, but many larvae—forty-one in the case of

the third fish in the table—were attached to the bottoms of deep

pits in the mucosa of the pyloric region of the stomach. These pits

were not mere depressions of the wall of the stomach but actual

cavities, as shown in figure 7, bordered by a pronounced annular

thickening of the mucous membrane and as much as 2mm. in dia-

meter. Larvae ranging in size from almost the smallest met with to

those near the adult stage in development were tightly crowded into

these pits and at the same time strongly contracted longitudinally.

GLARIDACRIS CATOSTOMI GEN. NOV., SP. NOV. § 7

STOMACH INTESTINE

Length of host Number Kind Number Kind

Arey ATCA ee Re Yat. HOO LE ae 9 A UltS Wire ha ee ieee neta abled cise

PSO eaboals Rory eerree rieneiiet iret ie 4 AGAUILES TPMT Carat Rie ce aaron [tal tes ed shave ellen

14 GAVE NN cara) Cslape Vey reg ares ees een ae op stare ae atte

PT S\ Tato 2 RS oleae ee eo 63 Orb aveg: Kew) PADRE RIN Phets Aneta Ea AtAh omib ete iat

BLS} Teak sR Selle yest a ey Adee | eheoreT eG A ier Aaa] [CO TAA ina AL 2 Larvae

IX) TIN GO ho Ren aoe MAA oe 9 Larvae 52 Larvae

PADIS) Saale hw ee RIN US OD a IRS IDE EA AN IS Le GET ek Re UL 2 Adult

1 Larva

DOSWIMNIN< iio ek his ke 7 Larvae 21 Larvae

ZEON SUS sec ke tote let ane [ata ee Ole meee oe eee os 20+ Larvae

DADS Sr aaa als, on pa dey TRIS eal RE Se eR esl (Ae teat arene eo kaernee 12+ Larvae

EXTERNAL FEATURES

On account of its possessing a well developed musculature for its

size this species exhibits considerable differences in degree of contrac-

tion and elongation on fixation. If no care is taken in applying the

fixing reagent nor in slightly manipulating the specimen, it usually

contracts to such an extent that it becomes almost useless, at least

for the making of toto preparations. However, the adults, which

are here considered to be those whose uteri may be seen in toto

preparations to contain a few or many eggs, may be said to range in

length from about 5 to 25mm. and from 0.4 to 1.0mm. in maximum

breadth.

In immature individuals the scolex, when not strongly contracted,

has somewhat the form of a truncated rectangular pyramid with

the longer diameter in the transverse direction. As shown in figures

1 and 2, the edges of the base and the apex protrude markedly, in the

latter case forming a terminal disc comparable to that of many of

the bothriocephalid cestodes. The dorsal and ventral faces of the

organ are each divided by two ridges converging towards the apex

into three sucking grooves or loculi, of which the middle is best devel-

oped and most efficacious during life. It is also the last to become

smoothed out with strong contraction of the whole scolex. The

lateral loculi are, furthermore, not in the same plane with the medial

one but inclined towards the corresponding ones of the opposite

surface so that the edges of the scolex, especially just behind the

8 A. R. COOPER

terminal disc, are often not much thicker than the ridges between

the loculi. As regards these features the organ consequently resem-

bles that of Archigetes brachyurus Mrazek 1908, which is here repro-

duced (Fig. 5) for the sake of comparison. In adults, on the other

hand, the edges of the terminal disc are usually found in preserved

material to be contracted to the point of obliteration, so that the

whole organ is shaped more like a wedge or chisel with oftentimes

rather thick margins (Figs. 3, 4 and 9). As a matter of fact the

scolex of this form assumes a greater variety of shapes than that

of any other tapeworm I have yet examined, in which respect it is

comparable to the leaf-like anterior end of Caryophyllaeus. The

dimensions of the organ are as follows: Length, 0.30 to 0.45mm.;

width (posteriorly), 0.45 to 1.10mm.; depth (posteriorly), 0.50 to

0.75mm.

Behind the scolex the strobila narrows down for a short distance

and then much more gradually enlarges again to the region of maxi-

mum diameter, which is usually behind the genital openings. Yet

in many specimens, especially the more relaxed ones, the whole

strobila is all but uniform in width thruout its length. The region

between the scolex and formost vitelline follicles, which includes

the narrowest portion of the strobila and is consequently called

the neck, varies from 1.5 to 2.5mm. in length. Finally the posterior

end, as shown in figure 6, is somewhat triangular in outline with a

slightly indented tip where the excretory vessels open to the exterior,

but bears nothing in the nature of an appendix such as it present in

Archigetes.

CUTICULA, SUBCUTICULA AND PARENCHYMA

The cuticula, which varies in thickness from 7 to 11, is bounded

on the inside by a comparatively heavy basement membrane, about

one-sixth of the thickness of the whole layer, and on the outside by a

smooth membrane about one-half as thick as the basement mem-

brane. The remainder of the tissue has the appearance of a reticulum

enclosing numerous distinct granules. This reticulum is in reality

a meshwork of fine canaliculi which freely pierce both limiting

membranes, thus giving them the appearance in tangential sections

of fine sieves. Nowhere is the cuticula modified to form spinelets

nor distinct cirri, altho over the scolex it is considerably folded and

GLARIDACRIS CATOSTOMI GEN. NOV., SP. NOV. 9

irregular, the outer membrane being all but absent, especially within

the suckers. For Caryophyllaeus laticeps Will (1893) described a

cuticula 5 to 6m in thickness and composed of only two layers, an

outer showing radial striping, as if formed by fine bristle-like hairs,

and an inner, more deeply staining stratum, comparable to the

basement membrane of this form. He saw no distinct pores in the

cuticula, and thought that perhaps the striations might represent

prolongations of the subcuticula.

The subcuticula is made up of large flask-shaped cells, closely

crowded together and provided with comparatively large nuclei.

Whereas the individual cells are not distinctly separated from one

another, the whole layer, from 90 to 100, in thickness, is clearly

marked off from the underlying parenchyma owing to the very

granular nature of its components. The nuclei, which are spherical

to oval in shape and provided with distinct spherical nucleoli, vary

from 16 to 18 in greatest diameter. They are located at different

levels, so that the whole layer has a pseudostratified appearance.

The enlarged central ends of the cells are usually rounded off towards

the parenchyma, which feature is clearly indicated by their character-

istically large granules. On the whole the subcuticula is not very

different from that of C. laticeps as described by Will.

The parenchymatous cells form an open reticulum showing only

a very few nuclei. They are in strong contrast with the subcuticular

cells on account of their clear, non-granular cytoplasm. .Posteriorly

the whole tissue is much limited in amount by the large reproductive

organs which are imbedded in it. No such “fibrous strands” of

modified parenchymatous cells, as described by Will for C. laticeps

and by Skrjabin for C. syrdarjensis, were seen in thisform. In the base

of the scolex and in the neck region, however, the medulla is occupied

by a more or less X-shaped mass of cells (Fig. 10) containing large

nuclei with numerous large granules which have a great affinity for

the counterstain. They are probably glandular in their nature since

they send long processes, especially in the diagonal direction, to the

cuticula covering the scolex, between the cells of the subcuticular

layer. Furthermore, no evidence of the presence of calcareous

bodies in the parenchyma was met with in an examination of both

fresh and preserved material.

10 A. R. COOPER

MUSCULATURE

The musculature is comparable to that of the cestodes proper in

that it is composed of two sets of fibres, the parenchymatous and the

cuticular. The former consists of sagittal (dorsoventral), frontal

(transverse) and two sets of longitudinal fibres, of which the latter

are much the strongest. Whereas both sagittal and frontal fibres are

few in number, they are not equally so, for the sagittal are somewhat

larger and more numerous. Both kinds tend to course slightly

obliquely where they are greatly interfered with by the reproductive

organs. The main or inner longitudinal fibres are, on the other hand,

comparatively large and arranged in thick bundles (Figs. 11, 12

and 13). They are situated among the central ends of the subcuticu-

lar or just within them, the cortical parenchyma being thus consider-

ably restricted in amount. Posteriorly the fasciculi are very unequal

in size and quite numerous. As they are followed forward, however,

their numbers diminish while their size increases, until at the base of

the scolex there are only eight large bundles arranged as in figure 10.

This is brought about by the fusion of the smaller bundles and the

passage of the fibres from one fasciculus to another. In longitudinal

sections the bundles are irregularly striated owing to there being a

considerable amount of myoplasm in the middle of each fibre around

the remains of the original myoblastic nucleus. Nevertheless, no

distinct nuclei such as described and figured by Will for C. laticeps

were seen. In the posterior end of the worm many of these longi-

tudinal muscles terminate in the walls of the excretory invagination

or run alongside of it to the extremity of the strobila. The outer

longitudinal group (Fig. 10) consists of a large number of bundles,

smaller but more uniform in size than those of the inner group,

situated among the peripheral ends of the subcuticular cells just

outside of their nuclei or from 15 to 30u from the cuticula. Posterior-

ly only a few of them pass beyond the anterior end of the excretory

invagination, but anteriorly they are very pronounced and continue

into the scolex. Similar fibres in C. laticeps were considered by Will

to belong to the cuticular instead of to the parenchymatous series.

The cuticular muscles consists of an outer stratum of circular

fibres lying close to the inside of the cuticula and an inner of longi-

tudinal fibres situated close within that. The longitudinal fibres,

which in some places intermingle slightly with some of the outermost

GLARIDACRIS CATOSTOMI GEN. NOV., SP. NOV. 11

members of the outer longitudinal parenchymatous group, are

arranged in small bundles, each containing at most only about ten

or a dozen fibres. In the posterior end of the worm they proceed

farther back than the latter, after being closely associated with them

opposite the excretory invagination. The same may be said of the

circular cuticular muscles, excepting that they are not distinctly

arranged in bundles.

In the scolex the cuticular muscles are much less pronounced over

the sucking-grooves than on the lateral faces. As shown in figure 9,

the eight large bundles of inner longitudinal muscles, mentioned

above, are arranged so that four form two sagittal pairs situated

towards the lateral faces, while the other four, somewhat larger ones

form two other sagittal pairs, each about half way between the nerve

trunk and the median line. These are distributed in a radiating

manner to the corresponding portions of the tip of the scolex, the

median pairs going to the ridges between the loculi and the neighbor-

ing parts of the latter. On the whole their attachment is similar to

that of the main longitudinal group in C. tuba and C. laticeps, as

described respectively by Monticelli (1892) and Will. The outer

longitudinal muscles are more numerous on the lateral surfaces of the

scolex than opposite the suckers, to the cuticula of which they are

easily traced. The loculi are also provided with a few scattered

radiating fibres, lying in both the longitudinal and the transverse

directions, and comparable to those used in the Pseudophyllidea for

the enlargement of the bothria. They are, however, of much less

functional importance in that connection than the sagittal and

transverse fibres, which are somewhat larger and more numerous

than in the middle of the worm. In fine, the musculature of the

scolex is poorly developed as compared with that of Bothriocephalus,

s. str., for example, which fact is shown in the great diversity of

shapes of the organ in preserved material. In fact it might be

considered to represent an intermediate stage between that of the

anterior end of Caryophyllaeus and that of the typical bothriocephalid

scolex. But the comparative inefficiency of the individual sucking-

grooves is compensated for by their number and by their manner

of attachment to the host’s alimentary tract, namely at the bottom

of the spacious pits described above.

12 A. R. COOPER

NERVOUS SYSTEM

The nervous system consists of a pair of ill-defined longitudinal

trunks and two equally indistinct and diffuse terminal ganglia situated

in the scolex, into which they pass. The main strands can be fol-

lowed more or less easily in material not especially treated to demon-

strate them only in the neck region. There, as shown in figure 10,

they are situated symmetrically in the median frontal plane within

the trapezium formed by the two pairs of main longitudinal muscle

bundles, much closer, however, to the lateral pair than to the more

median pair. They supply these muscles with large branches.

Whereas in the neck they are fairly uniform in diameter—which varies

from 18 to 30u—behind the most anterior vitelline follicles they

become quite irregular in transection, all but disappearing in places.

In the middle of the worm and posteriorly they seem to break up

into a diffuse plexus lying just within the subcuticular cells, that is,

among the numerous bundles of the inner longitudinal muscles. No

collateral strands such as the eight described by Will for C. laticeps

were seen in this form.

In the base of the scolex these chief nerve strands expand con-

siderably in the dorsoventral direction and become united by a few

transverse fibrils. Farther towards the tip, however, each of these

enlargements divides into two parts sagittally, and each of the latter

unites with its fellow of the opposite side by a loose strand of trans-

verse fibrils, so that two anteriorly directed loops are thus formed.

On the whole the nervous system is comparatively poorly developed,

since not only the chief strands but also their connections in the

scolex are composed of very fine, indistinct and loosely arranged

fibrils.

EXCRETORY SYSTEM

Thruout most of the length of the worm the excretory system

consists of a single layer of comparatively large and much coiled

longitudinal vessels situated just outside of the inner longitudinal

muscles among the central ends of the subcuticular cells. Whereas the

number of these vessels cannot be stated definitely, owing to many

transverse connecting channels, there is a tendency, especially in the

anterior regions, for eight of them to take the courses indicated in

figure 11. Three are located on each surface and one in the median

frontal plane at each side. In the anterior part of the neck region

GLARIDACRIS CATOSTOMI GEN. NOV., SP. NOV. 13

the number increases, and the courses of these vessels become

irregular, that is, the plexus becomes more diffuse. There they invade

all parts of the subcuticula and the periphery of the cortical paren-

chyma (Fig. 10). From 1 to 1.5mm. behind the tip of the scolex two

branches leave the plexus above and below the nerve cord on each

side (Fig. 10) and unite on the medial side of the latter to form one

vessel. In these positions the two vessels thus formed pursue spiral

courses forward and apparently unite close behind the nerve com-

misures mentioned above.. For C. laticeps Fraipont (1880) and Will

described an excretory system consisting in brief of four ‘ascending

canals” and ten ‘descending canals,’’ connected in the mobile anterior

end of the worm with each other and posteriorly with the so-called

excretory vesicle. Thus it is seen that as regards the main channels

of the excretory system at least this species is somewhat less com-

plicated in structure than the European species in question. In the

posterior end of the former the plexus just described converges

towards the centre of the medulla, as the vessels diminish in size,

and unites by several openings with the terminal receptacle. The

latter, as pointed out for C. laticeps by Steudener (1877), is merely

an invagination of the hinder end of the worm, about 0.25mm. in

length by about 0.05 in diameter. Its wall is composed of only a

lining of cuticula continuous with that covering the posterior end of

the worm and also traceable for some distance into the larger branches

leading from the plexus into the invagination. In the sections made

it was also seen to be quite vacuolated and granular and poorly pro-

vided with cuticular muscles, thus indicating that the whole structure

is not a true pulsating vesicle.

Nowhere in any of the sections studied was I able to find the

typical terminal organs of the excretory system, namely, the flame-

cells, which according to Fraipont are present in C. laticeps with the

same structure as those in trematodes. But in their place there

appeared much less specialized cells which are, nevertheless, com-

parable in some respects to the ciliated funnels of other cestodes.

As shown in figure 8, each consists of a large cell provided with a

large nucleus with a distinct spherical nucleolus but much vacuolated

cytoplasm. The cytoplasm is aggregated close around the nucleus,

and from this mass numerous strands pass to the wall of the cell.

The latter is directly continuous with one or more canaliculi which

lead off from the structure and connect up with the larger vessels

14 A. R. COOPER

to form the plexus. The whole has the appearance of an enlarge-

ment of the terminal vessel, enclosing an amoeboid cell which is sus-

pended in the centre of the vesicle by its pseudopodia. Thus the

vacuolated space which surrounds the cytoplasmic mass and is

continuous with the cavity of the canaliculi is comparable in part at

least to the funnel which accommodates the “flame” in the typical

flame-cell. . These terminal organs are situated close around the

canals in the periphery of the cortex or even farther out among the

inner ends of the subcuticular cells. Furthermore, they are much

more numerous in the neck region than elsewhere. The only reference

I have been able to find to structures at all comparable to these

peculiar cells is that by Wright and Macallum (1887) on Sphyranura

oslert. For this form, a monogenetic trematode, they described as

the terminal renal organs peculiar elongated, club-shaped cells

which are situated in close proximity to the vitelline follicles and

the principal groups of muscles. The cytoplasm of the cell is divided

into a number of coarse, granular trabeculae radiating from the

nucleus to the wall, thus leaving a system of communicating spaces,

“empty in the fixed, but often unobserved in the fresh, condition.

. . . Each cell has a process at one pole, with an axial wavy channel

connected with one of the neighbouring excretory capillaries . . . ,

the wall of which passes insensibly into the membrane of the cell.”

Perhaps also certain large amoeboid cells with nuclei filling up almost

the whole of the cell and large nucleoli surrounded by clear areas,

found by Will in specimens of C. Jaticeps fixed in Flemming’s solution

and crude acetic acid and described under the nervous system, may

rightly belong to this category of peculiar excretory cells.

REPRODUCTIVE ORGANS

On the whole the reproductive organs of this species (Fig. 6)

closely resemble those of the species of Caryophyllaeus. In the longi-

tudinal direction they extend from 1.5 to 2.5mm. behind the scolex,

where the foremost vitelline follicles are situated, to the posterior end

of the worm. The openings and the central connections of the ducts

are located, however, near the posterior end, the former, in fact,

only from 1.5 to 2.8mm. from the tip, depending on the degree of

contraction of the specimen. Excepting Skrjabin, the European

writers emphasize in their descriptions of the species of Caryophyl-

laeus the fraction of the whole length of the worm occupied by the

GLARIDACRIS CATOSTOMI GEN. NOV., SP. NOV. 15

organs behind the opening of the cirrus. For C. tuba the latter opens

at the beginning of the last quarter of the body, for C. laticeps at the

beginning of the last fifth, and for C. fennicus in the last fifth. Skrja-

bin says only that in C. syrdarjensis the ovary is situated in the

posterior third of the body. Owing to very considerable differences

in degree of contraction and elongation it seems to me that, at least so

far as the present species is concerned, these proportions are not of

specific value. On account of the greater development of the muscu-

lature anteriorly that portion of the body ahead of the genital

openings is much more variable in length than that behind the

apertures—hence the above measurements for the latter only.

The genital openings are situated in the midline on the ventral

surface from 0.5 to 1.0mm. apart. The cirrus-opening is somewhat

transversely elongated and about 0.15mm. in diameter. The opening

of the female atrium has the form of a shallow, transverse, crescentic

groove, about 0.35mm. in width, with its concave side directed

anteriorly. Both apertures are so close together in most of the

specimens at hand that they are located at the bottom of a common

depression; or, the slight depression accommodating the male opening.

runs insensibly into the crescentic female atrium.

Male genitalia.—The testes (Fig. 11) are not entirely surrounded

by the vitelline follicles as in C. laticeps and C. syrdarjensis. Anter-

iorly they begin at the same level as do the latter, and posteriorly

they extend to the cirrus-sac or in some cases slightly beyond its

anterior border. They are irregularly ellipsoidal in shape, and have

lengths, widths and depths of from 0.135 to 0.227, 0.100 to 0.145

and 0.127 to 0.181mm., respectively. Their number as determined

by direct count and by calculation from the average number in

longitudinal and transverse sections varies from 150 to 160. They

are especially noteworthy on account of their showing the various

Stages of spermatogenesis with almost diagrammatic clearness, a

fact which was also noted by Monticelli in the case of C. tuba and by

Skrjabin in his description of C. syrdarjensis. Nevertheless in none of

the series of sections cut were any spermatozoa seen in any part of

the vas deferens, altho the uteri were in the same preparations well

filled with eggs. This would seem to indicate that contrary to the

usual procedure among cestodes the female genital organs develop

before the male organs and that self-fertilization does not take place.

16 A. R. COOPER

The vas deferens forms a loose and somewhat triangular mass of

coils about 0.32, 0.28 and 0.36mm. in length, width and depth,

respectively and situated immediately ahead of the cirrus-sac. Just

before entering the latter it expands into a muscular vesicula seminalis

having a diameter of from 65 to 90 and a length of about 0.30mm.;

but at its beginning it has no seminal reservoir like that attributed to

C. laticeps by Will. The wall of the duct consists of a lacerated or

pseudociliated, syncitial epithelium, provided with widely separated

nuclei—excepting in the seminal vesicle where they are fairly numer-

ous—and resting on a basement membrane. The musculature of the

vesicle consists of numerous circular fibres with a few oblique fibres

distributed among them.

Entering the cirrus-sac anterodorsally with a diameter of 30u,

the vas deferens expands in the dorsal third of the latter to form

a sort of secondary, but doubtless only temporary, seminal vesicle

averaging 60 in diameter. After taking several turns it gradually

diminishes to about 35m in the mid-region of the sac and passes

insensibly into the cirrus proper. The structure of the wall of the

duct within the sac up to this point is the same as that of the seminal

vesicle just outside of the sac. The cirrus, which occupies the lower

half of the cirrus-pouch, is a comparatively large closely coiled tube

with a diameter of 60 to 65yu. Its wall, which is much cleft and

folded on account of the length of the organ, is similar in structure to

that of the vas deferens, excepting that the number of circular muscu-

lar fibres is much greater and that the imperfect epithelium of the

latter is replaced (in the transitional region) by smooth cuticula,

continuous with that of the ventral surface of the worm as in the

cestodes proper. Altho in the material at hand there were no cases

of extruded cirrus, its structure and disposition within the sac is such

as to lead one to believe that when it is evaginated it is a compara-

tively long and stout organ.

The cirrus-sac (Fig. 12) is ellipsoidal in shape and occupies the

whole of the medulla of the region dorsoventrally and almost all of

it laterally. Its length, width and depth are, respectively, 0.40

to 0.50, 0.50 and 0.50 to 0.60mm. Its wall is composed of mus-

cular fibres running in all directions and not sharply separated from

the retractor muscles within the organ. A few dorsoventral fibres

pass from the top of the sac to the dorsal body-wall and a few from

GLARIDACRIS CATOSTOMI GEN. NOV., SP. NOV. 17

the equatorial region to the ventral body-wall. The contents of the

Sac are composed of numerous and very compactly arranged retrac-

tor muscles, their myoblastic nuclei and a small amount of parenchy-

matous tissues.

Female genitalia.—Into the dorsal portion of the female genital

atrium, which is about 0.25mm. in depth and lined with a much

lacerated continuation of the cuticula from the ventral surface

of the worm, the vagina empties slightly to one side of the median

line, the other side accomodating the opening of the uterus. From

the atrium it passes backward in the median line (Fig. 6) beneath,

or at some levels almost surrounded by, the coils of the uterus.

Its diameter near the opening varies from 50 to S5yu, but half way

along its course this is reduced to 30y. Thruout its length its wall

is composed of a lining of cuticula 5u in thickness and surrounded

by numerous circular muscles only, the myoblastic nuclei of which

form a rather distinct stratum about 10 distant from the fibres.

At the level of the posterior end of the ovary it opens into the oviduct

with a diameter of 84 and a much reduced cuticular lining and

layer of circular muscles. Unlike that of C. laticeps, as described

by Will, it is nowhere enlarged to form a receptaculum seminis.

The ovary is situated usually half way between the genital

openings and the posterior end of the animal (Fig. 6). It is from

0.8 to 0.9mm. in length and consists of a stout almost spherical

isthmus, about 0.4mm. in diameter, from which numerous, irregular

and thick lobules pass upward and slightly forward to enclose a capa-

cious generative space. In the latter respect this form resembles

not only the species of Caryophyllaeus but also Cyathocephalus and

Bothrimonus as described elsewhere by the writer (Cooper, 1919).

As shown in figure 13, the lobules lie in the periphery of the medulla,

close to the main longitudinal muscles. Ova near the beginning

of the oviduct average 15y in diameter in sections, and are composed

almost entirely of the nucleus, there being very little cytoplasm.

A distinct and almost spherical nucleolus taking the counterstain

very readily is to be seen in each nucleus.

The oviduct begins at the posterior end of the isthmus and

somewhat ventrolaterally in an oocapt, 254 in diameter by 20u

in length and provided with only a few circular muscles. About

125 from the oocapt it is joined by the vagina. This first portion

18 . A. R. COOPER

of the oviduct is 25 to 30u in diameter, and takes a dorsal course.

Its walls are composed of a thin but uniform layer of circular muscu-

lar fibres on the outside, and on the inside of a comparatively thick

layer of epithelium, the cells of which are not clearly separated

from each other but contain relatively large and deeply staining

nuclei. After passing backward and upward about 404 beyond

the point of union with the vagina the oviduct receives the common

vitelline duct.

As in the species of Caryophyllaeus the vitelline follicles are located

in the medulla in two distinct and separate regions: a large one

extending from 1.5 to 2.5mm. behind the tip of the scolex to the

cirrus sac, and a much smaller one in the more or less conical posterior

end of the worm behind the coils of the uterus (Fig. 6). In the

former situation they form an irregular layer in the periphery of

the medulla (Fig. 11), for not only do some dip down among the

testes, as mentioned above, but others extend outward to the main

longitudinal muscles; in the latter, however, they occupy almost the

whole of the medulla, as in C. laticeps. In the immature worm there

is, furthermore, some tendency for them to be arranged in two

lateral fields anteriorly, leaving a free strip in the median line dorsally

and ventrally. In the anterior region in particular they are very

numerous, irregularly ellipsoidal in shape, and vary greatly in size.

From 8 to 14 appear in transections, while their maximum diameter

is 0.20mm. Posteriorly they are slightly larger.

The process of the formation of the peculiarly clear yolk-cells

which are to be seen in the vitelline ducts (Fig. 14c) can be followed

with a considerable degree of satisfaction in the follicles. The cyto-

plasm of the small peripheral primordial cells from which they

develop is very compact, and consequently stains deeply as does the

nucleus (Fig. 14a). Numerous vacuoles appear in it and quickly

enlarge, so that in the intermediate stages the nucleus appears to be

suspended in the centre of the cells by protoplasmic strands radiating

from it to the cell-membrane, as shown in figure 14b. These strands

become modified into numerous, spherical deutoplasmic granules,

migrate outward and eventually come to lie just inside the cell-

membrane (Fig. 14c). In the proximal part of the uterus, where

from four to six vitelline cells are seen to be associated with each

fertilized ovum in the formation of the egg, the nucleus enlarges still

GLARIDACRIS CATOSTOMI GEN. NOV., SP. NOV. 19

more and becomes more transparent, while the cell-wall gradually

breaks down, thus liberating the vitelline granules. The enlarged

nuclei remain intact, however, during the passage of the egg thru

almost the whole length of the uterus.

The common vitelline duct varies in diameter from 30 to 75y, and

is lined by an epithelium similar to that of the oviduct. It is largest

immediately dorsal to the posterior end of the ovarian isthmus

where it forms a vitelline reservoir, as in C. /aticeps, as much as 220yu

in width by 45y in depth when filled with yolk. A little farther

forward it receives two main tributaries, varying considerably in

calibre according to the amount of vitelline material they contain.

Whereas these two ducts collect chiefly from the follicles ahead of the

uterus, at least one small tributary on each side drains the follicles

situated in the posterior end of the worm, and unites with the main

ducts near their point of union with each other.

Shortly after being joined by the common vitelline duct and as it

courses a little farther back on one side or the other, the oviduct

becomes surrounded by a poorly developed shell-gland. The ootype

is consequently inconspicuous. Beyond the ootype the epithelium is

syncitial in its nature since no distict cell-boundaries appear. More

than its inner half is deeply cleft to form pseudocilia, yet its nuclei

_ are comparatively large. As the oviduct—now, more properly

called the beginning of the uterus—continues backward in a dorsal

position in the medulla, it gradually enlarges, according as it becomes

filled with eggs, its wall becomes thinner and thinner, and the nuclei

diminish in number, flatten out and eventually disappear. The

latter takes place particularly after the organ turns in its course—

just ahead of the posterior group of vitelline follicles—and starts

forward towards the female genital atrium.

From a point just behind the level of the posterior border of the

Ovarian isthmus to its opening the uterus is surrounded by a volumi-

nous mass of club-shaped, unicellular glands (Fig. 13), similar to

those described for the species of Caryophyllaeus and closely resem-

bling those described by the writer (1919) for Cyathocephalus ameri-

canus and Bothrimonus intermedius. As to the function of these

cells no definite statements can be made as yet. Monticelli likened

the similar cells in C. tuba to those to be seen along the uteri of many

trematodes as well as of Gyrocotyle urna (Wagener), and called them

20 A. R. COOPER

glutin-producing glands. Will described them in C. laticeps, and

said that they were “‘fully identical” with those in Diphyllobothrium

latum. He also incidentally mentioned that Saint-Remy (1890)

looked upon them as a shell-gland. Schneider (1902) called them

glandular cells in C. fennicus, while Skrjabin considered them to be

shell-glands in C. syrdarjensis. In view of the fact that, as in the

species of the subfamily Cyathocephalinae just mentioned, the shell-

gland surrounding the ootype is poorly developed—altho it was

clearly seen in this species to initiate the formation of the egg-shell—

they may act as an accessory shell-gland. Even tho this whole region

of the uterus is lined with a deeply cleft cuticula, numerous droplets

of material were seen in the sections studied adhering to or lying

among the pseudocilia as if they were secreted from the cells in

question; and it is only in this portion of the uterus, not in the thin-

walled proximal region, that the shells of the eggs are thickest. At

any rate, since the uterus is provided with only a very few scattered

circular muscles, excepting just before its opening, they cannot be

myoblastic in their nature. Distally they diminish considerably in

number, yet they are directly continuous with the myoblastic nuclei

of the more numerous muscular fibres surrounding the terminal

portion of the duct and the female atrium, which in turn are continuous

with the subcuticular cells around the atrial opening. As stated

above, the uterus opens into the female genital atrium ahead of and

slightly to one side of the vagina. The atrium itself is from 0.20 to

0.30mm. in length by about 0.10mm. in diameter and lined with a

very irregular and deeply cleft cuticula.

The mature fresh eggs, when examined in normal saline solution,

were found to be ovoid in shape and from 54 to 66y in length by 38 to

484in width. The shell is from 2 to 3yu in thickness, and is provided

at its larger end with a small button-like boss and at its smaller end

with an operculum from 12 to 16y in diameter.

LIFE HISTORY

As regards the development and life-history of this species only

a few statements can be made at present. Larvae as small as that

shown in figure 15 were found in the stomach of the host, but, altho

a thoro dissection of the food-contents, which consisted of larvae of

Chironomus and Simulium, Ostracoda, Cladocera, “‘caddice-worms,”’

GLARIDACRIS CATOSTOMI GEN. NOV., SP. NOV. 21

dragon-fly nymphs and Mollusca, was made, their mode of entrance

was not discovered. Possibly further search will show that some

member of these groups of animals, if not a tubificid worm as in

Europe, is the intermediate host. Finally, from the standpoint of

the systematic position of the species it should be emphasized that

the smallest larvae found had nothing whatsoever in the nature of

appendages.

SYSTEMATIC POSITION

From the above description it is clear that this species, altho a

member of the family Caryophyllaeidae Liihe 1910, does not belong

either to Archigetes or to Caryophyllaeus. As pointed out above, the

scolex resembles that of at least one species of Archigetes, namely,

A. brachyurus Mrazek, but is quite different from the simple, leaf-like

anterior ends of the species of Caryophyllaeus. The reproductive

organs, it is true, are much more comparable to those of the latter,

but certain features of the muscular, excretory and nervous systems

do not permit of its being placed in either genus. Consequently a

new genus is erected to accommodate this form, and is given the

following characters:

Glaridacris gen. nov.

With the characters of the family. Medium sized caryophyllaeids with the

anterior end modified to form a scolex, provided on each surface with three suckers, of

which the median one is the deepest and most efficacious. Main longitudinal paren-

chymatous muscles in eight large fasciculi in the anterior part of the neck and the base

of the scolex. Only two main nerve strands in the medulla, connected in the scolex

by two more or less diffuse commissural loops. Excretory vessels form a single cortical

plexus with eight principal longitudinal channels; no true flame-cells present, terminal

renal organs, peculiar, highly vacuolated, simple cells. Expansion of the vas deferens

before entering the cirrus-sac to form a vesicula seminalis. 6dAapis, chisel; axpvs,

summit.

Type, and as yet only, species: G. catostomi sp. nov.

The principal specific characters may be set down as follows:

Glaridacris catostomi sp. nov.

With the characters of the genus. Small cestodarians, up to 25mm. in length by

1.0mm. in breadth. Scolex, short and broad, chisel-shaped in older specimens, hexa-

gonally pyramidal with prominent terminal disc in younger, base large in both; length,

0.30 to 0.45mm., width (posteriorly), 0.45 to 1.10mm., depth (posteriorly), 0.50 to

BZ A. R. COOPER

0.75mm. Neck only slightly narrower than body, 1.5 to 2.5mm. in length; whole

worm, apart from scolex, cylindrical, with somewhat conical posterior end.

Cuticula, 7 to 11 in thickness; subcuticula, 90 to 100u. No “fibrous strands’

nor calcareous bodies in parenchyma.

Female genital atrium, 0.5 to 1.0mm. behind opening of cirrus, 0.20 to 0.30mm.

in depth by 0.10mm. in diameter, opening crescentic, in same depression with male

opening.

Testes not completely surrounded by vitelline follicles; extend to cirrus-sac pos

teriorly; irregularly ellipsoidal in shape, from 0.10 to 0.18mm. in different diameters;

150 to 160 in number. Vas deferens, a loose somewhat triangular mass ahead of cirrus-

sac, 0.28 to 0.36mm. in diameter. Vesicula seminalis, 0.30 by 0.06 to 0.09mm.

Cirrus-sac large, almost spherical, occupying almost whole of medulla of region, 0.40

to 0.60mm. in diameter. Cirrus, 60 to 65 in diameter.

Vagina median, ventral, 30 to 55y in diameter. Ovary irregularly lobular, 0.8 to

0.9mm. in length, with nearly spherical isthmus, 0.4mm. in diameter. Oocapt, 20

by 25. Vitelline follicles not completely surrounding the testes, 8 to 14 in transec-

tions, 0.20mm. in maximum diameter. Vitelline reservoir, the expanded common

vitelline duct, 220 by 454. Ootype inconspicuous. Uterus in two portions, a proximal,

thin-walled, and a distal, extending from the posterior vitelline follicles to the opening

and surrounded by a large number of unicellular glands; empties into female atrium

slightly ahead of and to one side of vagina.

Eggs, ovoid, with small boss at larger end, 54 to 66y in length by 38 to 48y in

width.

Habitat: In stomach and intestine of Catostomus commersonii (Lacépéde).

Finally, Lithe’s (1910) characterization of the family will have to

be slightly emended to include this new species:

CARYOPHYLLAEIDAE Liihe 1910, e.p.

Monozootic pseudophyllidea with scolex unarmed; may or may not bear more or

less well expressed sucking organs which are set off from the rest of the body by a neck-

like constriction or are fused with the same without such. A caudal appendage bearing

on its hinder end the hooks of the oncosphere may also be present in the sexually mature

animal. Genital organs present only singly. Reproductive openings surficial, ventral,

medial and near the posterior end. Testes, numerous, exclusively anterior to the ovary

and the female genital ducts. Cirrus unarmed, ahead of the female sexual apertures;

vagina and uterus open at the bottom of a common vestibule which resembles in its

histological structure the shallow genital atrium and opens into it close behind the

cirrus. Ovary two-winged, directly behind the genital opening. Vitelline follicles

in the medulla, but peripheral to the testes and more or less completely surrounding

them like a mantle; mostly ahead of the ovary, but a group also in the hinder end of

the body, separated from the main mass by the ovary and the female genital ducts.

Uterus a winding canal, without sack-like expansions. Eggs, operculate.

College of Medicine,

University of Illinois.

GLARIDACRIS CATOSTOMI GEN. NOV., SP. NOV. 23

WORKS CITED

Cooper, A. R.

1919. North American Pseudophyllidean Cestodes From Fishes. Ill. Biol.

Monogrs., 4:289-541, 13 pls.

FRAIPONT, J.

1880. Recherches sur l’appareil excréteur des trematodes et des cestodes. Arch.

Biol., 1:415-36, 2 pls.

Ltue, M.

1910. Parasitische Plattwiirmer. II Cestodes. Die Siisswasserfauna Deutsch-

lands, Dr. Brauer, Berlin, Heft 18:1-153.

MonrvIceEttl, F. S.

1892. Appunti sui Cestodaria. Atti d. r. accad. sc. fis. mat. di Napoli, 5, ser.

2(6), 11 pp., 4 figs.

MrAzek, A.

1908. Ueber eine neue Art der Gattung Archigetes. Vorliufige Mittheilung.

Centrbl. Bakt., Orig., 46:719-23.

SAINT-Remy, G.

1890. Recherches sur la structure des organes genitaux du Caryophyllaeus muta-

bilis Rud. Rev. biol. du nord de la France, Lille, 2:249-60, 1 fig.

SCHNEIDER, G.

1902. Caryophyllaeus fennicus n. sp. Arch. Naturgesch., 68J, 1:65-71, 82-98,

3 figs.

SKRJABIN, K.

1913. Fischparasiten aus Turkestan. I. Hirudinea et Cestodaria. Arch.

Naturgesch., 79J, Abt. A, 2:2-10, 2 pls.

STEUDENER, F.

1877. Untersuchungen iiber den feineren Bau der Cestoden. Abhandl. naturf.

Gesellsch., Halle, 13:277-316, pls. 28-31.

Warp, H. B.

1911. The Discovery of Archigetes in America, with a Discussion of its Structure

and Affinities. Science, N.S., 33:272.

Warp, H. B. and G. C. WurepLe,

1918. Fresh-Water Biology. New York.

WI, H.

1893. Anatomie von Caryophyllaeus mutabilis Rud. Ein Beitrag zur Kenntnis

der Cestoden. Zeitschr. wiss. Zool., 56:1-39, 2 figs., 2 pls.

Wricat, R. R. and A. B. Macatium,

1887. Sphyranura osleri: a contribution to American helminthology. Journ.

Morph., 1:1-48, 1 pl.

EXPLANATIONS OF FIGURES

co cirrus opening g glands

cs Cirrus-sac ~ isthmus of ovary

ev excretory vessel Im longitudinal muscles

fa female atrium , n nerve(s)

ns nerve strand

Fig.

Tig.

Fig.

A. R. COOPER

ovary t testis

outer longitudinal muscles u uterus

renal cell v vagina

shell-gland vf _ vitelline folliscles

vs vesicula seminalis

Unless otherwise stated, the lines indicating the magnifications of the figures are

0.5mm. in length.

Fe eh ee os oa

PLATE I

Surficial view of scolex of specimen 3.5mm. in length.

Lateral view of same.

Surficial view of scolex of specimen 21mm. in length.

Lateral view of same.

Scolex of Archigetes brachyurus, surficial view. After Mrazek.

Genital organs in posterior end of worm, toto preparation, surficial view.

Pits in the mucosa of the host’s intestine, each showing only two of the several

larvae found in them.

A terminal renal cell and its connections, from a frontal section. The line

at the side represents 0.05mm.

10.

12e

13.

14.

PLATE IT

Transection thru the middle of the scolex.

Transection thru the anterior part of the neck.

Transection thru about the middle of the whole worm.

Transection thru the cirrus-sac.

Transection thru the ovarian isthmus.

Three stages in the development of the vitelline cells: a, the primordial cell

from the periphery of the follicle; b, an intermediate stage from the centre of the

follicle; c, the mature cell from the vitelline reservoir. The line repersents 0.02mm.

Fig. 15. The smallest larvae procured.

TRANSACTIONS OF THE AMERICAN MICROSCOPICAL SOCIETY

VOL. XXXIX

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PLATE I COOPER

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TRANSACTIONS OF THE AMERICAN MICROSCOPICAL SOCIETY

VOL. XX XIX

PLATE II COOPER

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THE GENERA OF THE EN CHYTRAEIDAE (OLIGOCHAETA)!

By Paut S. WELCH

INTRODUCTION

Michaelsen’s monograph (1900) on the Oligochaeta contains the

last general revision of the genera of the Enchytraeidae for the whole

world. Eisen (1905) modified, to some extent, the genera then

known to occur in North America. Since the publication of the

above-mentioned works numerous contributions to the knowledge of

these annelids have been made, so that the family has grown from a

relatively small group containing 13 genera and less than 100 species

to the present status of 16 genera and approximately 325 species.

With this marked increase has come the necessity for certain changes

and modifications in the limits of most of the groups.

The revision herein presented is the direct result of the discovery

of certain enchytraeids which failed to agree exactly with any of the

older generic descriptions and in order to properly assign them a

careful survey of several genera was necessitated. It was then

decided to extend the study to include all of the known genera of

Enchytraeidae and thus not only make available a considerable

amount of inaccessible material but also present something which

will serve as a basis for further revision as soon as more data are

secured. The writer wishes to acknowledge indebtedness to one

of his graduate students, Miss Helen M. Scott, who gave considerable

assistance in testing and rechecking the revised generic descriptions.

This revision can at best be regarded only as an attempt to

indicate progress to date. Certain unsurmountable difficulties

make it impossible at the present time to do more than work over

critically the published records as they now stand and to determine

the present status of each group as nearly as possible. The descrip-

tions of all of the species now assigned to the Enchytraeidae (approxi-

mately 325) have been re-examined in this connection and the work

of the writer on this group of annelids, covering a period of ten years,

has been brought to bear upon the task wherever possible. The

‘ Contribution from the Zoological Laboratory of the University of Michigan.

26 PAUL S. WELCH

principal difficulties are indicated below in order to point out some of

the features which should receive attention in future investigations

and revisions:

1. Descriptions based upon sexually immature specimens or upon

material not so stated but apparently immature.

2. Species placed tentatively in certain genera but data insuff-

cient to make final disposal of them.

3. Lack of information on morphological features now known to

have important systematic value, especially in the older descriptions

4, Deviations which strongly have the appearance of being errors

of observation or of printing.

5. Structures recorded as “‘not seen” or “not found” sometimes

the result of faulty methods of study, such as external examination

only or dissection only, or the use of poorly preserved specimens.

6. Difficulty in correct interpretation of certain descriptive terms,

as for example, does “lobulated testes’? mean divided testes as repre-

sented in Lumbricillus or something much less significant. In the

absence of illustrations which supplement the descriptions, such

expressions are very puzzling.

7. Interpretation of indefinite terms indicating differences of

degree, as for example, “‘setae slightly curved,” an expression which

when unaccompanied by further explanation or by figures is practically

unusable.

8. Difficulty arising from descriptions which fail to mention

important organs. Does lack of mention always or ever mean posi-

tive absence of the structure? Apparently, many investigators have

not realized the importance of stating positively that certain charac-

ters are absent. To leave these matters without mention is a distinct

detriment.

In this revision, the generic descriptions have been so modified as

to include what seems to be well founded changes demanded by in-

creased data as well as new features now regarded as having generic

value. In making these modifications the procedure has been as

follows:

1. The writer concurs with those who hold that the multiplication

of genera should be avoided except where the case is perfectly clear.

2. Genera and species founded upon sexually immature material

have been disregarded. It is well established that complete, depend-

THE GENERA OF THE ENCHYTRAEIDAE 27

able data cannot be secured from immature specimens and it is to be

hoped that future investigation will frown upon any disposition to

write descriptions from such imperfect material. It can only lead

to confusion and hinderance.

3. Data easily derivable from illustrations not supplemented by

description were regarded as valid.

4. Lymphocytes and the brain have been omitted purposely from

generic consideration since the writer doubts their value in generic

distinction.

5. In certain cases, statements known to be true for some of the

species of a genus have been incorporated in the generic description

since they represent all that is known at present about the features

mentioned. Re-examination of the other species will decide whether

such features will remain valid.

6. When the usual generic characters are not mentioned in the

descriptions, such omission is taken to mean that no information is

available, rather than that they are absent.

7. There have been incorporated into the genus descriptions

certain features which may prove later to be specific characters, as

for example, when structures are indicated as “‘present and absent.”

This is done largely for the sake of record and to indicate divergences

from the former descriptions. It remains for future investigation to

make the final disposal.

8. The term chylus cell is used to indicate the large, intestinal cells

each of which is characterized by a longitudinal, intracellular canal.

In the region involved, chylus cells usually alternate with the ordinary

epithelial cells which line the lumen of the intestine.

9. Eisen’s system of classifying the various forms of penial

bulb has been followed to considerable extent. There is increasing

evidence that the features of the penial bulb have distinct generic

value.

10. No attempt has been made to list more than the most impor-

tant literature involved in making this review. A complete set of

references and a statement of the synonymy up to 1900 is given in

Michaelsen’s monograph.

SUBFAMILIES

Some attempts have been made in the past to establish sub-

families in the Enchytraeidae. Eisen (1905, pp. 11-13) proposed

28 PAUL S. WELCH

four subfamilies, Mesenchytraeinae, Enchytraeinae, Achaetinae, and

Lumbricillinae, the major basis of distinction being the character

of the penial bulb. However, since the structure of the penial bulb

was not known for certain genera, the distribution of the genera

among these subfamilies was to some extent inferential. Cejka

(1910, p. 25) made use of three subfamilies under the names Frideri-

citinae, Mesenchytraeinae and Henleinae.

There seem to be some good grounds for considering the structure

of the penial bulb as a basis for the erection of subfamilies, but since

its structure is unknown in such genera as Achaeta, Distichopus,

Chirodrilus, and Stercutus, it does not seem profitable just now to

attempt to discuss this problem.

THE PENIAL BULB

The first attempt to use the characters of the penial bulb in the

classification of the Enchytraeidae was made by Eisen (1905, pp.

6-10) who, after an extensive study of a large number of North

American species, thought it possible to recognize three distinct

“types” which were definitely related to certain taxonomic groups.

The writer (Welch, 1914, pp. 173-180) presented a critical discussion

of this matter, pointing out that it seemed necessary to make some

modifications in Eisen’s original system. Since that time many more

of the North American forms have been studied and while it is still

probable that certain changes may ultimately be necessary, all of the

evidence at hand indicates that the characters of the penial bulb

are valuable in generic and possibly in specific diagnoses. For this

reason, statements as to the penial bulb have been incorporated into

revised definitions of the various genera, retaining Eisen’s terms for

the different types. If many of the species from the Old World can

be re-examined and the structure of the penial bulb described and

figured, the taxonomic status of this organ will be made more certain.

For sake of ready reference, Eisen’s summary of the three types

of penial bulbs will be quoted here.

“The Meseéenchytreid bulb is a single muscular structure, con-

taining circular muscles as well as fan-shaped muscular bands connec-

ting the body wall with the periphery of the bulb. Between the

muscular bands are generally found numerous penial glands which

THE GENERA OF THE ENCHYTRAEIDAE 29

open on the surface of the bulb around the penial pore. The sperma-

duct penetrates the bulb, opening on the center of its outer surface.

The Enchytreid bulb is multiple, consisting of several separate

cushions grouped around the penial pore. In these cushions we find

several sets or fascicles of glands, each fascicle opening by itself on

the surface of the body. There are no muscular bands connecting

the base of the cushions with its periphery. The sperm-duct never

penetrates the bulbs or cushions but opens close to and independently

of them. Exterior to the cushions there are numerous muscles

connecting the body wall immediately surrounding the pore with

other parts of the same somite.

The Lumbricillid bulb is always single and covered with a strong

muscular layer, which however never penetrates down between the

cells of the bulb. There are generally two or three distinct sets of

glandular cells in the bulb. Some of these open in the lower part of

the sperm-duct, or rather in a narrow groove in the elongation of the

sperm duct. Others open on the free surface of the bulb, either

irregularly or in narrow circular fields, bunched into fascicles. The

sperm-duct penetrates one side of the bulb. In Bryodrilus the gland

which opens in the extension of the sperm-duct is covered with a thin

cushion of muscular strands, forming a bulb within a bulb.”’

RELATIONSHIPS

It is not intended that any particular significance be attached to

the order in which the different genera are treated in this paper.

Certain genera are too poorly known at present to justify any attempt

to establish relationships, while others are little enough known to

make it a difficult and an uncertain task. It thus seems best in this

paper to omit efforts to determine phylogeny.

PROPAPPUS MICHAELSEN

Setae sigmoid; distal extremity cleft; those of a bundle equal;

four bundles per somite, two lateral and two ventral. Dorsal pores

absent. Oesophagus passing abruptly into intestine in 8; intestinal

diverticula absent; chylus cells absent; peptonephridia absent.

Origin of dorsal blood-vessel anteclitellar or intraclitellar. Nephricia

with small, slender, funnel-shaped anteseptal part and with loose,

scantily lobed, irregularly folded postseptal body, the folds being but

30 PAUL S. WELCH

little more than in close contact. Testes undivided; moderately

compact. Spermiducal funnel extremely short; shallow bowl-shaped.

Sperm duct very short; confined to 12. Penial bulb absent; small

atrial chamber at ectal end of sperm duct; atrial glands absent.

Spermathecae simple; no diverticula; no connection with digestive

tract; long, extending into 6-12.

DISCUSSION

Formerly, the genus Henlea (Michaelsen, 1903, p. 51) was re-

garded as the most primitive group of the Enchytraeidae because of

the diverse character of the setae manifested by the various species

although it presented no distinct transition features leading into the

near-standing, more primitive families of Oligochaeta (Phreodrilidae,

Tubificidae, Naididae). In 1905, however, Michaelsen (pp. 24-28)

described under the name Propappus a genus based upon specimens

found abundant in Lake Baikal, Southern Siberia, at depths of 2-8

meters. These specimens presented a complex of characters of par-

ticular interest. Most of the fundamental features are enchytraeid

leaving little doubt as to its membership in that group. However,

certain affinities with other families are manifested in the presence of

the following structures:

1. Cleft setae are recorded for the first time among the Enchy-

traeidae, all other known species having the simple-pointed type.

Cleft setae are common in Naididae, Tubificidae, and Lumbriculidae.

2. The spermiducal funnel is a very short, shallow, bowl-shaped

organ, resembling the funnel in certain other oligochaetes (Tubéfex,

et al) and showing little resemblance to the elongate, cylindrical,

glandular, thick-walled funnel found in practically all enchytraeids.

Apparently, only two other enchytraeids have spermiducal funnels

which at all resemble those of Propappus, namely, Mesenchytraeus

bunget Mchlisn. and Mesenchytraeus grebnizkyi Mchlsn. (Michaelsen,

1901, pp. 193, 199), in which they are very short, and ‘“‘pantoffel-

formig.”’

3. Each nephridium consists of a small, slender, funnel-shaped

nephrostome which constitutes the entire anteseptal part. The

postseptal part, however, departs strikingly in form and structure

from the typical enchytraeid condition in being very loosely con-

structed, having the appearance of an irregular knot of adherent loops

THE GENERA OF THE ENCHYTRAEIDAE 31

or folds, the free end of which composes the efferent duct. This

type of nephridium recalls the postseptal coils in the same organ in

Tubificidae, et al. Of all the other enchytraeids, Mesenchytraeus

alone shows any approach to such nephridial structure, although

its irregular, lobed, postseptal part in which the wide ducts are close

together is definitely coalesced into one mass.

Only two species are known at present to belong to this annectent

genus, namely, glandulosus and volki. The former, found in Lake

Baikal, in the one on which the genus was established. Recently,

Michaelsen (in a paper dated 1915 but which must have been pub-

lished in 1916 since papers dated 1916 are referred to in it) described

a second species, volki. It appears that this same writer first reported

it in the “Hamburger Nachrichten, Jahrg. 1916, Nr. 53, vom 30,

Januar, 3. Beilage, p. 1,” as Palpenchytraeus volki, n. gen., n. sp.

but later placed it in Propappus—a decision which certainly seems

more nearly correct. It is worthy of mention that in this species the

elongated spermathecae recall the condition in many of the North

American mesenchytraeids.

HENLEA MICHAELSEN

Setae straight and unequal in size, or straight and equal in size,

or slightly sigmoid and approximately equal in size; distal extremities

simple-pointed; four bundles per somite, two lateral and two ventral.

Head pore at 0/1. Dorsal pores absent. Oesophagus (with possible

rare exceptions) expanding abruptly into intestine. Peptonephridia

present or absent. Intestinal diverticula usually present. Origin of

dorsal blood-vessel anteclitellar; rarely intraclitellar; cardiac body

absent. Blood colorless. Nephridia with either large or small ante-

septal part; nephridial canal loosely wound and surrounded by con-

siderable amount of cell mass. Ventral glands absent. Testes com-

pact; not divided. Spermiducal funnel cylindrical; sperm duct short,

confined to 12, rarely longer. Sperm sacs and ovisacs absent. Penial

bulb of lumbricillid type. Spermathecae connecting with digestive

tract; diverticula present or absent.

DISCUSSION

Perhaps no genus of Enchytraeidae needs a thorough going revi-

sion as badly as does Henlea. Its heterogeneous nature has been

a2 PAUL S. WELCH

recognized by investigators for some time but certain conditions

surrounding the problem have thus far made such a revision almost

impossible. It therefore presents many difficulties in connection

with the present attempt to redefine the genus. Friend (1914b,

pp. 150-153; 1915, pp. 197-198) has pointed out the existence of

certain “groups”? within this genus. The writer [Welch] suspects

strongly that Hepatogaster Cejka should be regarded as a part of

the genus Henlea—possibly as a subgenus. It seems likely that these

“groups” will form the basis for the establishment of several sub-

genera when the genus is thoroughly worked over, particularly

when many of the foreign species have been re-examined and more

thoroughly described.

Certain deviations, apparent or otherwise, from the newly modi-

fied genus definition require some notice. Some ill-defined species

(lefroyi Beddard; scharfii Southern; et al) seem to offer exceptional

features, but the imperfect descriptions leave considerable doubt as

to whether they belong in this genus at all. Hence no significance

can be attached to them at present.

Eisen (1905, p. 98), in connection with his discussion of Henlea,

presents the following statement: ‘‘Chylus cells in the intestine in

the vicinity of clitellum.’’ However, in his subsequent descriptions,

no mention of them appears except in the case of H. guatemalae

(pp. 102-103) which is described as having no chylus cells at all.

In none of the American species of Henlea examined by the writer

have chylus cells been observed.

A number of species have been described in which the origin of the

dorsal blood-vessel is specified as intraclitellar and the definition has

been modified so as to include these forms. However, Friend (1913b,

pp. 460-461) has described a species under the name insulae which

has the dorsal blood-vessel arising in ‘117/18 or 19/20.” This form

is assigned to Henlea but taking the original description as it stands,

the writer is unable to place it with any more certainty in Henlea

than in one or two other genera, as for example, Enchytraeus. For

this reason, the apparent exception has not been given any particular

consideration. H. alba (Friend, 1913c, p. 83) and H. hillmani (Friend,

1914b, p. 135) are reported as having the origin of the dorsal blood-

vessel in'the region of 13-14.

THE GENERA OF THE ENCHYTRAEIDAE oo

Of the sixty or more species now assigned to this genus, there

are several which future investigations will certainly prove invalid.

HEPATOGASTER CEJKA

Setae straight and equal; distal extremities simple-pointed; four

bundles per somite, two lateral and two ventral. Head pore at 0/1.

Dorsal pores absent. Oesophagus merging gradually into intestine.

Peptonephridia present, dorsal and ventral. Intestinal diverticulum

present, surrounding digestive tract. Chylus cells absent. Origin

of dorsal blood-vessel anteclitellar; cardiac body absent. Nephridia

with smaJ] anteseptal part; nephridial duct loosely coiled and with

distinct cell mass. Testes not divided. Spermiducal funnel cylindri-

cal. Penial bulb of lumbricillid type. Spermathecae connecting

with digestive tract; diverticula absent. Characteristic, longitudinal

canals in epithelium of digestive tract in posterior part of body just

entad of perivisceral blood-sinus.

DISCUSSION

The genus Hepatogaster was established by Cejka (1910) for the

reception of two species which he considered as presenting characters

representing a new group. A careful examination of descriptions

reveals at least a close affinity with Henlea. In fact, it could be

included in Henlea with practically no change in the limitations of

the latter. Only one feature seems to offer any difference, namely,

that the oesophagus passes gradually into the intestine, but it seems

doubtful if a new genus could be established upon that character

alone. The presence of certain peculiar longitudinal canals in the

epithelium of the posterior part of the alimentary canal is stressed

in the original description and while these characters seem to be

unique, their value as a generic character remains to be demonstrated.

The structure of the penial bulb requires some notice. Cejka

thought that it resembled the enchytraeid type, interpreting certain

peculiar glands which open out through the body-wall in 12 and 13 in

the vicinity of the sperm duct termination as parts of the penial bulb

proper. Unfortunately, the penial apparatus is recorded in only one

of the two species. However, a careful study of the description and

Cejka’s plates leads the present writer to hold that the bulb is of the

lumbricillid type for the following reasons: 1. The sperm duct opens

34 PAUL S. WELCH

to the exterior through a compact, glandular bulb which is typically

lumbricillid. This duct actually opens out through it into a penial

invagination—a thing which does not occur in the typical enchytraeid

bulb. (2) Of the nearby groups of problematical glands, the one in

12 is single and median, thus apparently belonging to neither bulb.

(3) The other glands are in 13—another somite—a thing which has

not been observed in connection with the various parts of a typical

enchytraeid bulb. (4) In general appearance these peculiar glands

resemble the ‘‘ventral glands” found in certain enchytraeids although

they are unusual in being free from direct connections with the ven-

tral nerve cord.

Owing to the incompleteness of some of the data on this proposed

genus, it is allowed to stand for the present although there seem to

be good reasons for believing that it should be reduced at least to

the rank of a subgenus of Henlea.

~ BRYODRILUS UDE

Setae slightly or distinctly sigmoid; distal extremities simple-

pointed; those of a bundle equal in size; four bundles per somite, two

lateral and two ventral. Head pore at 0/1. Dorsal pores absent.

Oesophagus merging gradually into intestine. Peptonephridia pre-

sent. Four intestinal diverticula present. Origin of dorsal blood-

vessel intraclitellar; cardiac body present or absent. Nephridia with

small anteseptal part; nephridial canal loosely wound; cell mass large.

No ventral glands. Testes compact; not divided. Spermiducal

funnel cylindrical; sperm duct confined to 12. Sperm sacs and ovisacs

absent. Penial bulb of lumbricillid type. Spermathecae connecting

with digestive tract; diverticula absent.

DISCUSSION

While a few slight modifications have been introduced into the

description of this genus, no important comments are demanded

here. No mention of intestinal diverticula appears in the description

of B. sulphureus (Bretscher, 1904, p. 262) but since the material on

which the description was based was immature, this omission may

have no significance. The head pore in this same species is recorded

as appearing on the tip of the prostomium.

Four species are assigned to this genus.

THE GENERA OF THE ENCHYTRAEIDAE 35

BUCHHOLZIA MICHAELSEN

Setae sigmoid; distal extremities simple-pointed; those of a bundle

approximately equal in size; four bundles per somite, two lateral and

two ventral. Head pore at 0/1. Dorsal pores absent. Oesophagus

expanding abruptly into intestine. Peptonephridia present. Chylus

cells absent. Origin of dorsal blood-vessel anteclitellar or intraclitel-

lar; arising from summit of dorsal intestinal diverticulum; cardiac

body absent. Blood colorless. Nephridia with anteseptal part large

or small. Spermiducal funnel cylindrical; sperm duct confined to 12.

Structure of penial bulb unknown. Spermathecae connecting with

digestive tract; diverticula absent.

DISCUSSION

In Buchholzia focale (Friend, 1914a, pp. 118-119) no mention is

made of a dorsal intestinal diverticulum and the origin of the dorsal

blood-vessel is given as ‘‘Henlean.”’

But little is known concerning the penial bulb in representatives

of this genus. Eisen (1905, p. 12) places the genus under his sub-

family Lumbricillinae but explains (p. 6) that he does so on account

of its ‘undoubted relationship to the genus Henlea.”’

Buchholzia parva (Bretscher, 1900a, p. 24) is described as showing

no connection of the spermathecae with the digestive tract. However,

the sexual maturity of the material might be questioned since it is

stated that no trace of a clitellum was found.

Six species are now assigned to this genus.

MARIONINA MICHAELSEN

Setae sigmoid; distal extremities simple-pointed; those of bundle

approximately equal in size; four bundles per somite, two lateral and

two ventral. Head pore at 0/1. Dorsal pores absent. Oesophagus

merging gradually into intestine. Peptonephridia absent. Intestinal

diverticula absent. Chylus cells absent. Origin of dorsal blood-

vessel postclitellar; cardiac body absent. Blood red, yellow, or color-

less. Nephridia with anteseptal part large or small; nephridial canal

loosely wound; cell mass large. Ventral glands present or absent.

Testes undivided. Spermiducal funnel cylindrical; sperm duct con-

fined to 12. Sperm sacs present or absent. Penial bulb of the lumbri-

36 PAUL S. WELCH

cillid type. Spermathecae with or without connection with digestive

tract; never greatly elongate; diverticula present or absent.

DISCUSSION

In a few species, assuming that they are correctly referred to this

genus, there seems to be some deviation as to the origin of the dorsal

blood-vessel. Bretscher (1900b, p. 449; 1901, pp. 209-10) described

rivularis and guttulata as having this origin anteclitellar, and Eisen

(1905, p. 91) reported it in 12 in the single specimen of alaskae which

he described although he retained the general generic character of a

postclitellar origin (p. 90).

Friend (1912a, p. 224) has described a species, sialona, which he

assigns to Marionina, pointing out at the same time that it is striking-

ly like an Enchytraeus. ‘This species possesses peptonephridia—a

feature not represented in Marionina and sialona is unique in that

respect if it actually belongs in Marionina. However, the writer has

been unable, on the basis of the original description, to see why that

species should not be assigned to Enchytraeus, rather than to Marion-

tna. If this be the proper disposal of sialona, then the absence of

peptonephridia still stands as an invariable character of the genus.

Eisen (1905, p. 90) held that a generic character appears in the

presence of a small sperm sac in connection with each testis. Whether

this is true, remains to be determined by future investigations.

In antipodum (Benham 1904b, p. 294) the body of the penial

bulb appears to be of the lumbricillid type, but it is unique in possess-

ing a single, large accessory gland. Bretscher (1901, p. 210) recorded

guttulata as “‘ohne Prostata,’ but the whole description is so brief

that it is impossible to judge accurately as to the sexual maturity of

the material studied, or as to the exact meaning of the above quoted

statement.

M. werthi Mchlsn. (1908, p. 15) has a penial bulb which is de-

scribed as ‘‘einen winzigen, zwiebelférmigen, ganz in der Leibeswand

verborgenen Bulbus aus. An diesen Bulbus, der manchmal als

winzige diussere Papille etwas heraustritt, sitzt eine schwach gelappte,

in die Leibeshéhle hineinragende Prostata.”

About twenty-eight species are referred to this genus.

THE GENERA OF THE ENCHYTRAEIDAE 37

LUMBRICILLUS ORSTED

Setae sigmoid; distal extremities simple-pointed; those of a bundle

approximately equal in size; four bundles per somite, two lateral

and two ventral. Head pore at 0/1. Dorsal pores absent. Oeso-

phagus merging gradually into intestine. Peptonephridia absent.

Intestinal diverticula absent. Chylus cells absent. Origin of dorsal

blood-vessel postclitellar, rarely intraclitellar; cardiac body absent.

Color of blood yellow, red, or colorless. Nephridia with anteseptal

part either large or small; nephridial canal loosely wound, and con-

siderable cell mass between the folds. Ventral glands present or

absent. Testes divided deeply, forming a number of distinct lobes.

Spermiducal funnel cylindrical; sperm duct long but confined chiefly

to 12. Sperm sacs and ovisac absent. Penial bulb of lumbricillid

type. Spermathecae connecting with digestive tract; diverticula

absent.

DISCUSSION

While the limits of this genus have been but little changed,

certain variations may well be mentioned here. L. viridis (Stephen-

son, 1911, p. 48) has the dorsal blood-vessel arising in 13 and in ¢uba

(p. 43) it arises in 13, 14, 15. A variety (?) of minutus (Miill.)

described by Michaelsen (1911, pp. 1-4) has this vessel arising in 12.

Lineatus (Miill.) (=agilis Moore) has also been described by some

authors as having the dorsal vessel arising in 13.

Ventral glands do not appear in all representatives of Lumbricil-

Jus. Furthermore, they are said to occur in a few species of certain

other genera (Welch, 1914, p. 141).

Distinct sperm sacs and ovisacs appear to be absent in this genus.

A few references to very diminutive ovisacs restricted to the clitellar

region occur in the literature (Eisen, 1905, p. 77; Moore, 1905, p.

397) but these can scarcely be regarded as having any special signifi-

cance. Eisen (1905, pp. 75-76) stated that each division of the testes

is capped by a small sperm sac and evidently regarded this as a generic

character.

Stephenson (1911) has pointed out the close relation of Lumbricil-

lus to Enchytraeus on the basis of the discovery of certain species

which though assigned to the former, possess some characters

strongly suggestive of the latter.

38 PAUL S. WELCH

About thirty species are assigned to this genus at present, although

there is reason for doubting the validity of some of them.

FRIDERICIA MICHAELSEN

Setae straight or nearly so; unequal, those in bundle developed

in pairs, outer pair being largest, and enclosing smaller pairs; distal

extremities simple-pointed; four bundles per somite, two lateral and

two ventral. Head pore at 0/1. Dorsal pores present. Oesophagus

merging gradually into intestine. Peptonphridia present. Intestinal

diverticula absent. Chylus cells present. Origin of dorsal blood-

vessel postclitellar or intraclitellar, usually the former; cardiac body

absent. Blood colorless. Nephridia usually with large anteseptal

part, always consisting of more than nephrostome; cell mass well

developed. Ventral glands usually absent. Testes not divided.

Spermiducal funnel cylindrical; sperm duct short, usually confined

to 12. Sperm sacs and ovisacs absent. Penial bulb of lumbricillid

type. Spermathecae usually connecting with digestive tract; diver-

ticula present or absent.

DISCUSSION

While it seems advisable to make but little modification in the

description of Fridericia, a few variations as recorded in the literature

demand notice here.

F. tusca and F. valdarnensis are described by Dequal (1914,

pp. 15,17) as having setae which are sigmoid but those of a bundle

are of unequal length, the inner ones being shorter. The description

is very meager but it appears that even though they be sigmoid, they

still resemble the typical Fridericia arrangement and development.

Stephenson (1915, p. 47) describes the setae of F. carmichaeli as being

of the “Enchytraeus type’? but it may be that since in this species

there are usually only two setae per bundle and since the outer setae

of a Fridericia bundle are straight and approximately the same size,

this statement could be true, although if more were present per bun-

dle the inner ones might be shorter, smaller, and arranged in pairs.

Friend (1912b, p. 24) states that the head pore in F. anglica occurs on

the tip of the prostomium. In Fridericia peruviana, Friend (1911,

pp. 734-736) described the oesophagus as passing abruptly into the

intestine, but since the specimens on which the description was based

THE GENERA OF THE ENCHYTRAEIDAE 39

were immature, it seems best to attach no particular significance to

this case. According to Southern (1909, p. 165) F. magna Friend

has bright red blood. Friend (1899, p. 263) stated that in F. magna

an ovisac is present, extending caudad to 16.

At present about 90 species are assigned to Fridericia and while it

is very possible that some of them are not valid, it appears that this

is the largest of all the enchytraeid genera.

DISTICHOPUS LEIDY

Setae in two bundles per somite, representing the ventral rows

only; nearly straight; simple-pointed but blunt; very stout and

swollen in middle; hooked at proximal end. Head pore at 0/1.

Oesophagus merging gradually into intestine. Peptonephridia present.

Origin of dorsal blood-vessel postclitellar; small cardiac body present.

Blood colorless. Nephridia with small anteseptal part. Spermiducal

funnel cylindrical; sperm duct short, confined to 12. Penial bulb of

lumbricillid type. Spermathecae not described.

DISCUSSION

The genus Distichopus is known only from a single set of specimens

collected in Delaware and Pennsylvania by Leidy (1882, pp. 146-147).

Some of these specimens were later studied by Moore (1895, pp.

754-756) who extended the account, although even yet too little is

known concerning this unusual form. Certain important structural

features, such as the spermathecae, are yet undescribed and thus the

relationships of this genus are difficult to determine. Moore holds

that it is a close ally of Fridericia. The single known species bears

the name silvestris.

ACHAETA VEJDOVSKY

Setae entirely absent; dorsal and ventral rows preserved in some

species only as pear-shaped gland cells in body-wall, gland cells also

absent in other species. Head pore large; on tip of prostomium..

Dorsal pores absent. Oesophagus merging gradually into intestine.

Peptonephridia present or absent. Origin of dorsal blood-vessel

anteclitellar. Blood colorless. Nephridia with moderate or large

anteseptal part. Spermiducal funnel cylindrical; sperm duct short or

long but confined to region of clitellum. Penial bulb present; struc-

40 PAUL S. WELCH

ture practically unknown; probably of lumbricillid type. spermathe-

cae with or without connection with digestive tract; diverticula

absent.

DisSCUSSION

Achaeta is a small genus to which is assigned, at the present time,

eight species, none of which occur in the Western Hemisphere. Its

most striking characteristic is the total absence of setae. In most

of the species, specialized, pear-shaped seta-glands occur in the

positions where setae would be expected, although three species,

veydovskyi Bretscher (1902, p. 27), maorica Benham (1904a, pp. 221—

223) and camerani (Cognetti 1899, pp. 1-4) are described as being

completely devoid of seta-glands. Peptonephridia have not been

found in minima Southern (1907, p. 77) and incisa Friend (1914b, pp.

133-134) but occur in the other known species. The penial bulb is

practically unknown for this group since in no case is it described in

adequate detail. It was figured by Vejdovsky (1879, pl. I, fig. 11) for

eisenit but even there it is difficult to determine its exact composition.

It suggests the lumbricillid type of bulb. In maorica Benham (1904a,

p. 222) the spermathecae are greatly elongated, extending to 9 or

10, thus recalling the greatly elongated spermathecae of some of

American species of Mesenchytraeus.

ENCHYTRAEUS HENLE

Setae straight; those of a bundle equal; distal extremities simple-

pointed; four bundles per somite, two lateral and two ventral. Head

pore at 0/1. Dorsal pores absent; Oesophagus merging gradually

into the intestine. Peptonephridia present or absent. Intestinal

diverticula absent. Chylus cells absent. Origin of dorsal blood-

vessel postclitellar or intraclitellar; cardiac body absent. Blood

usually colorless. Nephridia with small anteseptal part; cell mass

well developed. Ventral glands sometimes present. Testes not

divided. Spermiducal funnel cylindrical; Sperm duct confined to

12, or quite long, extending caudad through several somites. Sperm

sacs present or absent. Penial bulb of enchytraeid type. Sperma-

thecae connecting with digestive tract; diverticula present or absent.

THE GENERA OF THE ENCHYTRAEIDAE 41

DISCUSSION

While it has been necessary to modify the older definition of the

genus, only a few points require mention here. E. dubius (Stephen-

son, 1911, p. 56) is unique in possessing testes which are divided very

much as is the case in representatives of the genus Lumbricillus.

However, Stephenson himself (1915, p. 43) indicates that there is some

doubt as to the generic position of this species. The same writer

(1915, pp. 43-44) gives a critical discussion of sperm sacs in the genus

Enchytraeus but makes no attempt to draw a general conclusion.

Since the matter of sperm sacs is still in doubt, it seems best, in this

paper, to use the data directly from the original descriptions and

consider the statements of absence of sperm sacs as valid until they

are definitely shown to be in error. The writer (Welch, 1914, pp.

177-178) previously discussed the penial bulb as a generic character

and pointed out that not all of the species included in Enchytraeus

conform to the enchytraeid type of bulb. However, it appears at this

time. that in the cases in which the penial bulb has been adequately

described the large majority have bulbs of the enchytraeid type as

proposed by Eisen and may so be incorporated into a revised state-

ment of the limits of the genus, at least until subsequent investigation

yields more complete data.

Eisen (1905, p. 61), in his generic description, states that the intes-

tine generally possesses chylus cells. However, no mention of these

cells is made later in his descriptions of species.

A genus, Litorea, described by Cejka (1913) for the reception of a

species which he called krumbachi, is certainly the same as Enchytraeus

and is so treated in this paper.

About thirty-five species are considered as belonging to this genus

at the present time.

MICHAELSENA UDE

Setae straight; distal extremity simple-pointed; one to two setae

per bundle, often but one; four bundles per somite, two lateral and

two ventral; present only on some of the somites (except in M. man-

gert Mchlsn.). Head pore at 0/1. Dorsal pores absent. Oesophagus

merging gradually into intestine. Peptonephridia present or absent.

Intestinal diverticula absent. Origin of dorsal blood-vessel post-

clitellar; cardiac body absent. Blood colorless. Nephridia with

42 PAUL S. WELCH

small anteseptal part, consisting of nephrostome only. Ventral glands

absent. Testes not divided. Spermiducal funnel cylindrical; sperm

duct confined to 12, or long and extending caudad to 14. Penial

bulb of the enchytraeid type. Spermathecae connecting with diges-

tive tract; diverticula absent.

DISCUSSION

Formerly the absence of setae from some or the majority of the

somites was regarded as one of the chief distinguishing features of

this genus but Michaelsen (1914, pp. 177-181) described a new species

under the name mangeri in which setae are present in four bundles

on all of the somites. This species may be regarded as a connecting

form between Michaelsena and Enchytraeus and at the same time,

according to Michaelsen, emphasizes the relation of Michaelsena

to Fridericia.

Southern (1913, pp. 8-12) described under the name Grania

what he regarded as a new genus, pointing out similarities with a form

then known under the name of Enchytraeus monochaetus Mchlsn.

The latter is now known to belong to Michaelsena and Michaelsen

(1914, p. 181) seems to be right in placing Grania there also.

Eisen (1905, p. 73) incorporated in his definition of this genus the

statement that there are ‘‘No penial bulbs,”’ but this seemed to be

based upon the condition which he found in a single specimen to which

he gave the name paucispina and which did not permit a full descrip-

ctiption. However, the descriptions of species now assigned to

Michaelsena indicate that it may be of the enchytraeid type. Eisen

(1905, p. 11) placed the genus in his subfamily Enchytraeinae.

It seems to be becoming increasingly difficult to separate Michael-

sena from Enchytraeus and it is possible that some future revision

based upon more intensive study of all of the species involved may

suggest the fusion of the two groups.

Eight species are assigned to this genus at present.

MESENCHYTRAEUS EISEN

Setae sigmoid; distal extremities simple-pointed; approximately

equal in size in bundle; four bundles per somite, two lateral and two

ventral. Head pore distinct; usually at or very near tip of prosto-

mium. Dorsal pores absent. Oesophagus merging gradually into

THE GENERA OF THE ENCHYTRAEIDAE 43

intestine. Peptonephridia absent. Intestinal diverticula absent.

Chylus cells absent. Origin of dorsal blood-vessel postclitellar; car-

diac body present. Blood either colorless or red. Nephridia with

small anteseptal portion, consisting merely of nephrostome; post-

septal part large, irregularly pluri-lobed, and with cell mass between

folds of closely wound nephridial canal greatly reduced. Ventral

glands absent. Testes compact; undivided. Spermiducal funnel

usually cylindrical; sperm duct short and confined to 12, or very long

extending caudad for many somites. Sperm sacs and an ovisac often

present. Penial bulb of mesenchytraeid type. Spermathecae con-

fined to 5, or elongated and extending caudad for varying distances,

sometimes to clitellum; diverticula present or absent; communica-

tion with digestive tract present or absent.

DISCUSSION

Eisen (1905, p. 14) stated that a ‘‘single median ovisac” and “‘one

pair of sperm-sacs generally of large size’ are present in Mesenchy-

traeus. However, Mes. altus Welch (1917, p. 71), which unquestion-

ably belongs to this genus, has a pair of ovisacs and it seems possible

that other cases of that sort will appear.

Several special cases which depart somewhat from the definiton

as proposed require mention here. In eastwoodi Eisen (1905, p. 50)

the head pore occurs on the upper side of the prostomium near 0/1.

Mes. mencli Vejd. (1905, p. 5) is described as having “‘Herz im 12,”

apparently referring to an intraclitellar origin of the dorsal blood-vessel.

A similar origin seems true of celticus Southern (1909, p. 155), although

the statement is not made positively. Bretscher (1902, p. 16)

claimed that specimens of setosus Mchlsn. (=megachaetus Bret.)

show the dorsal blood-vessel arising in 11,13, or 16 — which seems

an unusual variation. Mes. grandis Eisen (1905, p. 44) is described

as having nephridia with broad anteseptal parts. In orcae, mirabilis,

and kincaidi (Eisen, 1905, pp. 40-41), the testes are recorded as

composed of “lobes” but it is not clear whether they are deeply

divided as in the case of Lumbricillus or are merely lobulate at the

free extremity. Bretscher (1901, p. 212) states that in alpinus the

spermiducal funnel is in 8, but nothing is given concerning the sperm

duct. In nanus (Eisen, 1905, pp. 51-52), the penial bulb is described

44 PAUL S. WELCH

as absent, but there seems to be some possibility that immature speci-

mens were studied.

This genus contains at the present time about 50 species.

HYDRENCHYTRAEUS BRETSCHER

Setae sigmoid; distal extremities simple-pointed; four bundles

per somite, two lateral and two ventral. Dorsal pores absent.

Oesophagus merging gradually intointestine. Peptonephridia pres-

ent. Origin of dorsal blood-vessel postclitellar. Blood yellow or red.

Nephridia with large or small anteseptal part. Spermiducal funnel

cylindrical. Spermathecae without diverticula.

DISCUSSION

This genus was established by Bretscher (1901, pp. 208-209)

for two incompletely described species, stebleri and nematoides,

found in Switzerland. Its status is somewhat uncertain owing to the

fact that information on the head pore, relation of oesophagus to

intestine, chylus cells, cardiac body, ventral glands, testes, sperm

sacs, Ovisacs, penial bulb, and relation of spermathecae to the diges-

tive tract is entirely lacking. Likewise certain other features are

incompletely described. Since the original description is the only

record, nothing further can be done with these forms until specimens

are again found and studied critically. The fragmentary information

which is available seems to indicate that it is a valid genus.

STERCUTUS MICHAELSEN

Setae sigmoid; distal extremity simple-pointed; four bundles per

somite, two lateral and two ventral. Head pore absent (apparently)

or very small. Dorsal pores absent. Oesophagus merging gradually

into intestine. Peptonephridia absent. Intestinal diverticula

absent. Chylus cells absent. Origin of dorsal blood-vessel anteclitel-

lar; cardiac body present. Blood colorless. Nephridia with small

anteseptal part consisting of but little more than a mere nephrostome;

postseptal part large; nephridial canal loosely wound and surrounded

by considerable cell mass. Testes small; undivided. Spermiducal

funnel short and distinctly funnel-shaped. Penial bulb unknown.

Spermathecae not connected with digestive tract; diverticula absent.

THE GENERA OF THE ENCHYTRAEIDAE 45

DISCUSSION

The genus Stercutus was established by Michaelsen (1888) to

receive a species, zivews, which was found inhabiting fish excrement

in Germany. No other representatives of this genus have ever been

described. Lack of information as to the character of the penial

bulb, the sperm duct, and the presence or absence of ventral glands,

Ovisacs and sperm sacs gives some difficulty in determining the

affinities of this genus.

CHIRODRILUS VERRILL

Setae in six bundles per somite, two sub-dorsal, two lateral, and

two ventral; those in ventral and lateral bundles distinctly sigmoid,

those in sub-dorsal less curved; distal extremities simple-pointed.

Blood colorless.

DISCUSSION

The description of this interesting genus is extremely meager

and based entirely upon the original record by Smith and Verrill

(1871, pp. 450-451) in which but few of the important details are

described. Two species are assigned to this genus, lJarviformis and

abyssorum, both collected in Lake Superior. Both are apparently

deep water forms, larviformis being dredged from depths of 17 and

59 fathoms, and abyssorum from 47 and 159 fathoms. There is some

question as to the position of this genus, certain previous writers

having regarded it as a tubificid. Beddard (1895, p. 314) and

Michaelsen (1900, p. 88) have classed it among the Enchytraeidae,

although the latter (1903, p. 50) later placed it among the Tubificidae.

Eisen (1905, p. 13) retains it in the Enchytraeidae. It is unlikely

that the matter can receive any positive decision until material is

again secured and carefully studied. Since some of the characters as

described are apparently enchytraeid in nature, it is included in this

review, with the realization, however, that future investigation may

show it to have other affinities. If it be an enchytraeid, it is unique

for the entire family in possessing six sets of setae per somite. Eisen

(1905, p. 13) places it under the Lumbricillinae, but states (p. 6)

that it is “appended for convenience sake’’ and points out correctly

that nothing is known concerning the penial bulb and other internal

structures.

46 PAUL S. WELCH

GENERA DUBIA

Under the name Chamaedrilus, Friend (1913a, pp. 260-263)

described a new genus from material collected in England. Con-

sidering the generic characters as recognized at present for the

family Enchytraeidae, a careful comparison has made it impossible

for the writer to distinguish Chamaedrilus from Marionina and in this

paper they are regarded as the same.

Bretscher (1905) described what he regarded as a new genus of

Enchytraeidae under the name of Euenchytraeus but unfortunately

the record was made on sexually immature material and as a conse-

quence nothing could be determined as to the nature of the reproduc-

tive organs. Since no further record of this postulated genus occurs

in the literature and since the original description is unusable as it

stands, it is omitted from consideration in this paper.

KEY FOR THE IDENTIFICATION OF THE GENERA

OF ENCHYTRAEIDAE

1 ( 2) Setae entirely absent; represented in most species only by

four longitudinal rows of pear-shaped glands in body-wall

STN AER ee a ADEE ERLE CL. = 3 oes) sale Aenean Achaeta

2a): Setae present. 4).'54 8 2k Meee st... 3

3 ( 4) Setae arranged in two bundles per somite...... Distichopus

4 ( 3) Setae arranged in more than two bundles per somite....... 5

5 (6) Setae arranged in six bundles per somite, two subdorsal, two

lateral and two ventralereewerme yee... ee! Chirodrilus

6 (5) Setae arranged in four bundles per somite, two lateral and

two’ Ventral Ye ener saatieaa ls 6 arc Sten eae 7

7 (8) Setae cleft at distal extremities; spermiducal funnel wide,

open, shallow, and extremely short; postseptal part of

nephridia composed of a few coherent folds not intimately

fused, forming very loose organ...) . .'. ..4c eee Propappus

8 (7) Setae simple-pointed at distal extremities; spermiducal

funnel cylindrical or trumpet-shaped; postseptal part of

nephridia compact / 2.24.2). i). Pk ee ee 9

9 (10) Setae straight, arranged in pairs, inner pairs of bundle suc-

cessively smaller than outer; dorsal pores present; chylus

cells: present in* walls of intestine: ... 52/9. seu Fridericia

THE GENERA OF THE ENCHYTRAEIDAE 47

10 (9) Setae straight, sigmoid, or in pairs in bundle with smaller

ones within; dorsal pores absent; chylus cells absent from

WVAIS ONT LESCUME eR UL Ma aN UEN IC en anal ec A ae 11

11 (14) Oesophagus expanding abruptly into intestine........... 12

12 (13) Dorsal blood-vessel arising from anterior end of single, dorsal

Intestinal divertiGuUlun ye yy Ie Ne LL Buchholzia

13 (12) Dorsal blood-vessel arising directly from perivisceral blood-

BSRATTRS tee ee RUS Uh UN ei CIRO Oe LNRM SOLEMN Rte ty ALN Henlea*

14 (11) Oesophagus merging gradually into intestine............ 15

15 (16) Setae usually absent from several somites (except in Michael-

sena mangert Mchlsn.); usually one setae per bundle, never

TRAREN ClEAIE DW eel cists ma iaenevs is es eualaancr a yaa) Michaelsena

16 (15) Setae regularly present on all somites except first and last

and wossibly Che clicellar is iis Sk Neda NCL MA NI 4 17

MeO) intestinal diverticula present ona 18

18 (19) Setae sigmoid; four distinct intestinal diverticula; origin of

dorsal blood-vessel intraclitellar.............. Bryodrilus

19 (18) Setae straight; one intestinal diverticulum completely sur-

rounding digestive tract; origin of dorsal blood-vessel ante-

Fel] el BEV ROI SP OPS Re SUR tA os NU EA SO AE AN Hepatogaster

meen) intestinal diverticula absent...) 2c na le a 21

21 (22) Setae straight; those of a bundle equal........ Enchytraeus

POMS EAC GIOMIGIA oe Ua Ne ON uuC A MA lence 23

23°(24) Peptonephridia present..........6053..) Hydrenchytraeus

mame) peeptonephridia absent 04). oii.) e owen ae hae a ee 25

25 (26) Origin of dorsal blood-vessel anteclitellar; spermiducal fun-

Hempsnort, and trumpet-shaped. 2 )..).305).).)8 0 ue Stercutus

26 (25) Origin of dorsal blood-vessel postclitellar; spermiducal fun-

RNR NEIORN CAD G8 UO Uy ailhy eagle u) Wau ia OUi La BGreg 27

Di, ieee Meeecminarite is i MM Sey lr Ai Mls Lumbricillu

Za (an enteasonelimotidivideds yee ON in 8 29

29 (30) Cardiac body absent; penial bulb of lumbricillid type;

nephridial duct loosely coiled and cell mass of postseptal part

well developed; spermathecae never extending through

SEVEPAL | SOMMECS HMMs he eis4 aN ALIAS a te Marionina

* A few species, e.g., hillmani Fr., insulae Fr., marina Fr., alba Fr., and three or

four uncertain forms, have been assigned to Henlea, although they are described as

having the oesophagus pass gradually into the intestine.

48 PAUL S. WELCH

30 (29) Cardiac body present; penial bulb of mesenchytraeid type;

nephridial duct closely wound and cell mass reduced to mini-

PTAA Ss 92 US alo anes ee isccy. cs Sy RN Mesenchytraeus

LITERATURE CITED

BEDDARD, F. E.

1895. A Monograph of the Order of Oligochaeta. 769 pp., 5 pl. 52 fig. Oxford.

BENHAM, W. B.

1904a. Some new species of Aquatic Oligocheta from New Zealand. Proc. Zool.

Soc. London, 1903 (Vol. II), pp. 202-232. 3 pl. 1 fig.

1904b. On the Oligocheta from the Southern Islands of the New Zealand Region.

Trans. New Zeal. Inst., 37:285-297, 3 pl.

BRETSCHER, K.

1900a. Mitteilungen iiber die Oligochaetenfauna der Schweiz. Rev. Suisse Zool.,

8: 1-44. 3 pl.

1900b. Siidschweizerische Oligocheten. Rev. Suisse Zool., 8: 435-458. 1 pl.

1901. Beobachtungen iiber Oligocheten der Schweiz. Rev. Suisse Zool.,

9:189-223. 1 pl.

1902. Beobachtungen iiber die Oligochéten der Schwiez. Rev. Susse Zool.,

10:1-29. 4 fig.

1904. Beobachtungen iiber die Oligocheten der Schwiez. Rev. Suisse Zool.,

12:259-267.

_ _ 1905. Uber ein neues Enchytraeidengenus. Zool. Anz., 29:672-674.

CEJKA, B.

1910. Die Oligochaeten der Russischen in den Jahren 1900-1903 unternom-

menen Nordpolarexpedition. I. Ueber eine neue Gattung der Enchy-

traeiden, Hepatogaster. Mem. Acad. Imp. Sci. St.-Petersbourg, (8),

29: No. 2, 29 pp. 3 pl.

1913. Litorea krumbachi n. spec. n. gen.—Ein Beitrag zur Systematik der Enchy-

traeiden. Zool. Anz., 42:145-151. 10 fig.

Cocn_ettI, L.

1899. Descrizione dell’ Anachaeta camerani nuova specie della famigilia degli

Enchitreidi. Boll. Mus. Zool. Anat. Torino, 14, Nr. 354, pp. 1-4.

DEQUAL, L.

1914. Gli Enchitreidi della Toscana. Monit. Zool. Ital., 25:13-24. 7 fig.

EISEN, G.

1905. Enchytreide of the West Coast of North America. Harriman Alaska

Expedition, 12:1-166. 20 pl. New York.

FRIEND, H.

1899. New British Annelids. Zoologist, (4), 3:262-265.

1911. New British Enchytreids. Journ. R. Micr. Soc., pp. 730-736. 1 pl.

1912a. New British Oligochets. Zoologist, (4), 16:220-226.

1912b. British Enchytraeids. III. The Genus Fridericia. Journ. R. Micr.

Soc:, pp: 9-27.

THE GENERA OF THE ENCHYTRAEIDAE 49

1913a. British Enchytraeids. V. Species New to Science. Journ. R. Micr.

Sac:; pp: 295-271 135 fig:

1913b. Some Jersey Oligochaets. Zoologist, (4), 17:456-464.

1913c. A Key to British Henleas. Zoologist, (4), 17:81-91.

19142. Rare and Unique Sussex Oligochaets. Hastings and East Sussex Natural-

ist, 2:114-123.

1914b. British Enchytreids. VI. New Species and Revised List. Journ. R.

Micr. Soc., pp. 128-154. 5 fig.

1915. Studies in Enchytraeid Worms. Henlea fragilis Friend. Ann. Appl.

Biol., 2:195-208. 6 pl.

Lerpy, J.

1882. On Enchytraeus, Distichopus, and their Parasites. Proc. Acad. Nat. Sci.

Phil., pp. 145-148.

MICHAELSEN, W.

1888. Beitrige zur Kenntniss der deutschen Enchytraeiden-Fauna. Arch. f.

mikr. Anat., 31:483-498. 1 pl.

1900. Oligochaeta. Das Tierreich, 10 Lief. XXXIX-++575 pp. 13 fig. Berlin.

1901. Oligochaeten der Zoologischen Museen zu St. Petersburg und Kiew. Bull.

L’Acad. Imp. Sci. St. Petersbourg, (5), 15:137-215.

1903. Die geographische Verbreitung der Oligochaeten. 186 pp. 11 pl. Berlin.

1905. Die Oligochaeten des Baikal-Sees. Wissenschaftliche Ergebnisse einer

Zoologischen Expedition nach dem Baikal-See unter Leitung des

Professors Alexis Korotneff in den Jahren 1900-1902. Erste Lief.

68 pp. 9 fig.

1908. Die Oligochaeten der Deutschen Siidpolar-expedition 1901-1903.

Deutsche Siidpolar-expedition 1901-1903, 9:1-58. 1 pl.

1911. Litorale Oligochiten von der Nordkiiste Russlands. Travaux de la

Soc. Imp. Nat. St. Petersbourg, 42: 1-6. 2 fig.

1914. Beitrige zur Kenntnis der Land- und _ Siisswasserfauna Deutsch-

Siidwestafrikas. Ergeb. Hamb. deutsch-siidwestafrikanischen Stu-

dienreise 1911. Oligocheta, pp. 139-182. 1 pl.

1915. Ein eigentiimlicher neuer Enchytriide der Gattung Propappus aus der

Niederelbe. Verh. nat. Ver. Hamburg, (3), 23:51—-53.

Mookrg, J. P.

1895. The Characters of the Enchytraeid Genus Distichopus. Am. Nat.,

29 :754-756.

1905. Some Marine Oligochaeta of New England. Proc. Acad. Nat. Sci. Phil.,

pp. 373-399. 2 pl.

Situ, S. I. and Verritt, A. E.

1871. Notice of the Invertebrata dredged in Lake Superior in 1871, by the U. S.

Lake Survey, under the direction of Gen. C. B. Comstock, S. I. Smith,

naturalist. Am. Journ. Sci. Arts, (3), 2:448-454.

SOUTHERN, R.

1907. Oligocheta of Lambay. Irish Nat., 16:68-82. 2 pl.

1909. Contributions towards a Monograph of the British and Irish Oligocheta.

Proc. R. Irish Acad., 27:119-182. 5 pl.

50 PAUL S. WELCH

1913. Oligochaeta. Clare Island Survey, part 48. Proc. R. Irish Acad., Vol.

31,14 pp.1ipl. 1 fig.

STEPHENSON, J.

1911. On some Littoral Oligocheta of the Clyde. Trans. R. Soc. Edinburgh,

48:31-65. ipl. 14 fig.

1915. On some Indian Oligochaeta mainly from Southern India and Ceylon.

Mem. Ind. Mus., 6:35-108. 4 pl. 2 fig.

VEJDOVSEY, F.

1879. Beitrige sur Vergleichenden Morphologie der Anneliden. I. Mono-

graphie der Enchytraeiden. 14 pl. Prag.

1905. Ueber die Nephridien von Aeolosoma und Mesenchytraeus. Sitz. Gesell.

Wiss., Math.-Naturw. Classe, pp. 1-11. 1 pl.

WE cH, P. S.

1914. Studies on the Enchytreide of North America. Bull. Ill. State Lab. Nat.

Hist., 10:123-212. 5 pl.

1917. Enchytreide (Oligocheta) from the Rocky Mountain Region. Trans.

Am. Micr. Soc., 36:67-81.

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ANDERSEN AND WALKER

PLATE IV

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TRANSACTIONS OF THE AMERICAN MICROSCOPICAL SOCIETY

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PLATE V

ANDERSEN AND WALKER

TRANSACTIONS OF THE AMERICAN MICROSCOPICAL SOCIETY

VOL. XXXIX

PLATE VI ANDERSEN AND WALKER

AN ECOLOGICAL STUDY OF THE ALGAE OF SOME

SANDHILL LAKES

By Emma N. ANDERSEN AND ELDA R. WALKER

INTRODUCTION

Much of the western part of Nebraska consists of rolling sandhills

covered with bunchgrasses, yuccas, cacti, and other dry-land plants.

Cherry county which is situated in the north central part of the sand-

hills has in its valleys many bodies of water. In one area of 250

square miles there are about 75 lakes. This region is twenty-five

miles southwest from the village of Woodlake and about the same

distance south of Valentine. These lakes vary greatly in size. Some

are ponds a few hundred feet in diameter while the largest of the

group is about four and a half miles long and three-fourths of a mile

wide. All are comparatively shallow bodies of water, varying in

depth from a couple of feet to a maximum of fifteen feet. The

accompanying map! (Fig. 1) shows the relative size and arrangement

of the lakes of this region.

Surrounding the lakes are low meadows covered with grasses and

prairie flowers.» These meadows extend back from the lakes any-

where from a rod to, in some cases, a mile or more. Surrounding

the grasslands rise the “‘sandhills’”—dunes of yellowish sand extend-

ing to the next lake with its surrounding grasslands (Fig. 2). The

few native trees are stunted and produce no effect on the landscape.

The climate is that typical of the central plains, dry, windy, hot

in summer, and cold in winter. In the summer of 1912, the average

maximum daily temperature from June 24th to August 3rd was

91.2°, while the average minimum temperature was 59.8°. During

the same period the wind velocity reached a maximum of 21.9 miles

per hour, the average for a twelve hour period, while the lowest

average for a similar period was 1.3 miles per hour.

1 Furnished by Dr. G. E. Condra of the University of Nebraska.

* The vegetation of this region is well discussed by Pool (36).

52 ANDERSEN AND WALKER

The high daily temperature warms the water of the shallow lakes

and the prevailing high winds stir it, frequently mixing the warm

surface layers with those below causing thorough aération. The

climatic conditions together with the abundance of water plants, such

as Chara, Myriophyllum, Potomogeton, Scirpus, Nymphaea, and

Zizania, which give anchorage for attached forms, make an ideal

habitat for algae.

Although complete analyses of the water of the lakes are not

available, some idea of their alkalinity may be obtained from analyses

made by the department of chemistry of the University of Nebraska

of samples of water taken by Dr. R. H. Wolcott. These were made

in 1911 and showed the following parts per million of alkali:

Watts Lake, 111

Dewey Lake, 160

Hackberry Lake (no analysis made)

Big Alkali Lake, 622

Clear Lake, 1,129

As the analyses show some of the lakes are adapted to the algae of

fresher waters, while others are so alkaline that very few forms can

inhabit them. There is evidence that all of the series of lakes belonged

formerly to one general system. The larger lakes have well formed

shore lines except at their northwest ends where many of them are

swampy. This gives farther variations in the habitat for algae.

Surrounded by a large semiarid region, these lakes with their

large beds of wild rice and other seed producing plants prove a most

tempting resting and feeding place for migratory birds. They flock

here in large numbers and, no doubt, bring on their mud-laden feet

spores of algae from ponds both north and south—the only explana-

tion for some of the species present.

Earlier students working with the higher plants of the sandhills

reported a rich algal flora. This led the writers to spend the summer

of 1912 in studying the algae of the region and the conditions under

which they live. At first it was hoped to cover the entire group of

lakes, but after a few preliminary trips through the region, the work

was limited to a few localities so situated that they could be fre-

quently visited. These were chosen to represent so far as possible

the different types of habitat found in the region.

AN ECOLOGICAL STUDY OF ALGAE 53

From the various localities, specimens were collected by hand

either from a boat or by wading. A Birge net was also used to secure

free floating forms.

The identifications of species are based upon Tilden’s Myxophyc-

eae, Collins’ Green Algae of North America, and West’s British

Desmidiaceae. In groups such as the Oedogoniaceae, Characeae,

and Helminthocladiaceae covered by special publications, the identi-

fications were made with the works cited. Constant reference was

made to the other systematic works listed at the end of this paper.

The nomenclature of DeToni is followed except for the Desmids

where that of West is used. In the case of a few species not given

in the above general works, the terminology of the author describing

the form is used.

All diatoms were identified by Dr. C. J. Elmore. The one Volvox

found was identified by Dr. J. H. Powers. Acknowledgments are

also due Mr. F. H. Shoemaker for the photographs from which figures

2, 10, 11, and 12 were taken; to the Nebraska Conservation and Soil

Survey for help in prosecuting the work; to Prof. B. E. Moore for

suggestions as to physical problems; and to Prof. T. J. Fitzpatrick

for careful reading of manuscript and proof.

HACKBERRY LAKE

Hackberry lake (Fig. 1, 3) was chosen for a study of water condi-

tions, because it was representative and conveniently located. It is

a lake two and a half miles long and one-half mile wide. In depth it

varied during the summer of 1912 from three to seven feet. The

maximum depth, however, was found only in a few places. Usually

it did not exceed four feet.

The shore is sandy except at the northwest end where it is freshly

formed by the filling in of decomposing vegetable matter. This end is

swampy and passes gradually into dense beds of Zizania, Scirpus,

Myriophyllum, Potomogeton, and similar water plants. Here the

water is so filled with vegetation that it is almost impenetrable with

a boat or otherwise.

The southeast half of the lake was more open. Dense beds of

water plants were scattered through it but between them were

areas of open water, (Fig. 4). Here the algae were most abundant.

It was in this region that records of water conditions were made.

54 ANDERSEN AND WALKER

Algae were abundant all over the lake but more so in this part where

every rush stem, every lily pad, in fact, every submerged plant was

loaded with them (Figs. 5, 6). The number of species was not large,

as the list which follows shows, but the number of individuals was

very great. For example, a count was made of the thalli of Chaeto-

phora elegans. On one old Scirpus stem there were 592 thalli. Inan

area one meter square, there were 103 stems loaded in a similar way,

making over sixty thousand thalli of Chaetophora elegans in a square

meter. Chaetophora cornu-damae, Nostoc glomeratum, Gongrosira

debaryana, and Rivularia natans were equally abundant, while other

species were only slightly less so.

CLIMATIC CONDITIONS

The accompanying graphs (Figs. 15, 16, 17) are intended to illus-

trate weather conditions surrounding the lake. The station, (C on

Fig. 1) represented in Fig. 8, was in a blowout about 14 mile from

the lake. In the graph (Fig. 15) line A shows the daily variation in

temperature of this station as recorded by a Fries self-registering

thermograph. Line D shows the temperature on the north side of a

small house, a few rods from the lake (A on Fig. 1) as registered by a

maximum and minimum thermometer. Line C shows the tempera-

ture of the surface sand as recorded by a Fries self-registering soil

thermograph whose bulb was barely covered with sand. B gives the

temperature of the sand eight inches below the surface as recorded

by a similar instrument. The wind record was made by a standard

anemometer (Julien P. Fries), so only the averages of wind velocity

for twelve hour periods are available, but it gives some estimate as to

wind in the region. The high air temperature and its effect on the

surface temperature of the light sand when compared with the tem-

perature of the sand eight inches below the surface shows that

comparatively little heat penetrates deeply into the soil—a fact that

may have some bearing on the temperature of the water of the lakes

in some instances. Fig. 16 is a similar record for a much cooler week

taken in the grass of the meadow (Fig. 9) on the lake shore (B on

Fig. 1).

AN ECOLOGICAL STUDY OF ALGAE 55

WATER CONDITIONS

Temperature and Wind

For a month, July 8 to August 6, the same instruments were

stationed in a boat on this lake to determine, so far as possible, the

temperature conditions under which the algae were growing. During

this time the anemometer was placed on a post about six feet above

the surface of the water and a hundred feet from the water’s edge to

determine the velocity of the wind passing over the water. The

thermographs in the boat recorded the temperature (A) of the air

in the boat, the temperature (C) of the surface water, and the tem-

perature (2) of the water at the bottom of the lake. Due to drifting

of the boat to a slightly different position, the depth at which the

temperature was taken varied from three feet the first week to four

and one-half the second and fourth weeks and five feet the third week.

The temperatures were not taken in deeper parts of the lake because

there the algal growth was slight. As can be seen from the accom-

panying graph (Fig. 17) the temperatures of the surface and bottom

water approach each other very closely except on days of low wind,

when the temperature of the surface water varies considerably from

that at the bottom. This effect of air temperature and wind was

especially evident in the fourth week when great variations in air

temperature occurred.

It will be noticed that during a large part of this month, the water

temperatures were between 70° and 80° F., and that the temperature

of the water at all levels was remarkably uniform. It is evident from

the divergences of the temperature of surface and bottom water

during each period of lower wind that the uniformity of the water

temperature is due largely to stirring by wind. Also that a constant

stirring would produce good aération in all parts of the water is an

inevitable conclusion.

Hackberry lake is one of the less alkaline of the lakes in this group.

It is, however, more alkaline than some of its neighbors as was noted

before.

Light

A modification of the common solio paper photometer (text Fig. 1)

was devised to study the light conditions under which the algae were

56 ANDERSEN AND WALKER

growing.! This consisted of a water tight circular drum (A) at the

end of a metal tube (D). Inside of this drum was a disk (G) which

revolved by means of a rod (E) passing through the tube and con-

nected with a lever at the top. This disk (G) carried the paper

(F) on its under surface. It was perforated by eight holes (B) of the

same size as a Clear glass window (C) inthe drum. By revolving the

inner disk, the areas of solio paper under the perforation of the disk

could be exposed to the light through the window (C). The tube and

‘

Fig. 1

rod inside of it were made in sections so they could be extended to the

necessary length. The tube was marked in decimeters on the outside

so the depth could be ascertained at all times. With this instrument,

light readings were taken from a boat (Fig. 4) at various depths.

Care was taken to have the window in the drum exposed to direct

light at all times. In some instances, the period of exposure was

uniform and the depth was varied; in other cases both depth and

period of exposure were varied and in still others one depth was

maintained during the series of readings and the period of exposure

was varied.

Records taken by the last method are shown in (Fig. 7). The

top row shows exposures made in the air beginning at the following

hours, reading from left to right: 10:18 a.m., 10:30 a.m., and 2:10

p.m. In each record the exposures were 60, 45, 40, 30, 20, 10, 5, and

2 seconds.

Below are nine water records taken the same day and at intervals

immediately following or preceding the air exposures. These are

1 This instrument was made by the Spencer Lens Company.

AN ECOLOGICAL STUDY OF ALGAE a

arranged in order by depths, the first being one decimeter below the

surface and each succeeding one a decimeter deeper, making the last

nine decimeters deep. In all the periods of exposure were the same

as the air records, except the last two. Here no records could be

secured at the above exposure periods so the time was increased in

record 8 to 2, 3, 4, 5, 6, 7, and 8 minutes and in record 9 to 1, 2, 3, 4,

5, 6, 7, and 8 minutes. All water readings were compared with

exposures made in the air at the same hour. In no cases were the

readings satisfactory but some facts of interest were recorded.

1. Waves on the surface water varied the record of light intensity

greatly. On very windy days the intensity of light under the water

was so variable that one exposure showed almost no coloration while

the next at the same depth and period of exposure was deeply colored.

The only explanation evident was that the change in angle of surface

layer deflects the light almost entirely at times and not at other times.

On account of this variability readings on windy days were early

discontinued.

2. Before 9 a.m. or after 3 P.M. it was almost impossible to get a

usable tint on the solio paper, although to the eye it was entirely light

to the bottom of the lake. Evidently the angle between the sun’s

rays and the surface of the water was such that most of the rays were

deflected and the water light was simply reflected light and not of a

kind or quantity to affect solio paper.

3. Exposures made at a given depth but with different periods

of exposure gave very different records as to relative light intensity.

The shorter the exposure, the greater reduction of light, according to

the readings. When the exposure required more than three minutes

the error was so great that the reading was useless for comparison

with readings of shorter periods. Two series will illustrate the

point. The exposures were consecutive in both cases.

Time Relative intensity Time Relative intensity

All at 4 dm. deep All at 8 dm. deep

i sak AEA dhs 8 ce .0083

Le sZS Ever aga aE AE .0166

Lh .083 GMINA aye eta .0277

a .05 PUN SEMEN 584s, ghee avenz .0357

SPER inn sa sess .0416

58 ANDERSEN AND WALKER

Such a series can only be explained by the well known fact that

the reduction of silver in a photographic paper is not a uniform

process. Since these results proved a variable period impracticable,

exposures were made for a given period at various depths. These

readings showed clearly a constant reduction of light as the depth

was increased and agreed in general with the results obtained by

Needham and Lloyd (33), by Birge (11), and by Oltmanns (34).

4. Exposures made for constant periods at various depths showed

the light to be reduced by passing through the water. However,

this gave no accurate measure of the light at given depths, because

such exposures were compared with air tints of various periods of

exposure. These we have just shown to be unreliable because of the

inconstant reduction of the photographic film.

5. Solio paper is sensitive only to the blue-violet end of the spec-

trum. It is generally conceded that it is the red-orange end of the

spectrum that is most largely used by plants in photosynthesis

(Dangeard 22). It is, therefore, evident that tests made with solio

paper are of little value in determining the light conditions under

which algae grow. Such tests show in a most general way, as noted

above, that the direct rays of light are largely deflected except during

the middle of the day and that the light is reduced by passing through

water. Both facts are well known to physicists. This means that

water plants have a shorter day than air plants and that they grow

in less intense light at all times. In other words, as Oltmanns (34)

states, they are all shade plants.

6. Light readings taken in October show less light penetrating

the water than was found in July and August. The following table

shows this as well as could be determined from solio paper records.

Because of the reasons already given these records can only be

regarded as a crude estimate. The figures given are compiled from a

large number of readings so that the general diminution of blue light

per decimeter of depth and the relative intensity between the light

of midsummer and that of October are approximately correct.

Dm. August October

1 25 BD

2 25 al

3 2173 .066

AN ECOLOGICAL STUDY OF ALGAE 59

Dm. August October

4 083 .05

5 .05 .033

6 .033 .016

No doubt the reduction of light in October is due to the rays of the

sun striking the water more obliquely. Hence water plants have not

only a shorter day but also a shorter growing season than land plants.

It is evident that the main advantage in the present attempt to

measure light with reference to that used by plants under water is to

prove the absolute uselessness of the solio paper method.

Later the same photometer was modified slightly so that plates

sensitive to all kinds of light could be used in it (Figs. 10, 11, 12).

A color screen was added and in that way an attempt was made to

measure the red-orange light penetrating the water. Fig. 10 shows

the upper side of the drum which holds the photographic plate and

the detachable color screen over the exposure window. This is shown

again in Fig. 12 (below) with the tube removed. At the top of Fig.

12 is shown the under side of the upper half of the drum. The inner

disk shows two clips for holding the plate in place and eight perfora-

tions through which areas of the plate may be exposed to the light of

the window. This disk is revolved, exposing alternately a perforation

and a solid area, by a rod passing through the tube and connected

with the lever at the top (Fig. 11). This apparatus for making

exposures, while entirely rapid enough for solio paper, proved alto-

gether too slow for the highly sensitive plates. The only results

obtained indicated that red light penetrating many decimeters of

water was too strong for more than instantaneous exposures on these

plates. It is believed that a similar apparatus fitted with a shutter

that would make possible exposures of a fraction of a second would

come much nearer to giving an estimate of light available for the use

of algae. Even here the reduction of the photographic film would not

be uniform and some error would remain. Oltmanns (34) states that

physicists have shown that the light from each part of the spectrum

is deflected and absorbed differently by pure water. The water

inhabited by algae is always modified in color and transparency

both by its chemical content and by the presence of floating organ-

isms giving turbidity to the water. An instrument such as the above

would give an idea as to what effect such water conditions have upon

60 ANDERSEN AND WALKER

the quality of light used by algae even tho it did not give accurate

information as to the quantity of light available for them.

Murray and Hjort (32) investigating light conditions in the ocean,

by means of the Helland-Hansen Photometer and pan-chromatic

photographic plates found that red light penetrated the water less

than did blue and indigo. This agrees with the results cited by

Oltmanns (34) for pure water. This suggests, in some cases at least,

that actual light available for submerged plants is far below the

amount indicated by the solio paper photometer.

DISTRIBUTION

As is readily seen from the physical factors noted above, this

lake is characterized by fairly uniform temperature, aération, and

alkalinity. There remain but two factors that can influence the

distribution of algae in this lake—light and mechanical support.

The distribution may be entirely explained by these factors.

Attached forms as Rivularia pisum, Nostoc glomeratum, etc., grow

at a depth of about 3-4 dm. below the surface of the water. When

they are near the margin of the lake they grow on small submerged

sedges, chara, etc. In deeper water they grow on Potomogeton,

Myriophyllum, and such taller plants, but at remarkably uniform

distances below the surface. On the other hand, Nostoc pruniforme,

lying free at the bottom of the lake, was only found near the margin

where the water was two or at most three decimeters deep. At first

such forms led to the opinion that there was a zonal arrangement

about the shore such as is suggested by Comére (17) but the theory did

not stand the test. The distribution was entirely a vertical one caused

undoubtedly by the light intensity. There were several conspicuous

examples which demonstrate this. Chaetophora elegans extended on

Scirpus stems, almost its only support, over a distance of two deci-

meters but the growth was constantly most abundant in the upper

region, within 144-1 dm. of the surface of the water. Chaetophora

cornu-damae had almost the same distribution while Nostoc glomeratum

made its best growth at a depth of 2-4 dm. or even deeper. Evidently

it was crowded downward by the Chaetophora with which it grew.

Early in the season the Nostoc and Chaetophora grew together.

Later the Chaetophora occupied the upper zone while Nostoc grew

in equal abundance lower down.

TRANSACTIONS OF THE AMERICAN MICROSCOPICAL SOCIETY

VOL. XXXIX y -

PLATE VII ANDERSEN AND WALKER

TRANSACTIONS OF THE AMERICAN MICROSCOPICAL SOCIETY

VOL. XXXIX

Si ERIN nt iid na al Ran oa

WRN IEC act 5 B89 eB OS LANA EERE AEE A ies

PLATE VIII ANDERSEN AND WALKER

TRANSACTIONS OF THE AMERICAN MICROSCOPICAL SOCIETY

VOL. XXXIX

ANDERSEN AND WALKER

PLATE IX

AN ECOLOGICAL STUDY OF ALGAE 61

The following forms were found floating or attached almost

exclusively in the upper two decimeters of water or in water not more

than 2 dm. deep.

Clathrocystis aeruginosa Microspora amoena

Coelosphaerium Kuetzingianum Chaetophora cornu-damae

Nostoc muscorum Chaetophora elegans

Nostoc humifusum Bulbochaete sp.

Nostoc minutum Oedogonium fragile

Nostoc zetterstedtii Coleochaete orbicularis

Phormidium tenue Gongrosira debaryana

Chara contraria

Mixed with these were most of the unicellular forms of the

yellow-green algae and most of the desmids listed for this lake.

Such forms as the following predominated in the deeper water,

2—4 dm. from the surface: Rivularia natans, Rivularia pisum, Nostoc

glomeratum, and Nostoc austinii. With these were associated Meris-

mopedium aerugineum, Oscillatoria subtilissima, Dictyosphaertum

pulchellum, Gloeocystis gigas, Scenedesmus quadricauda, and S.

bijugatus. The distribution of unicellular free floating forms, how-

ever, was hard to determine because of the constant stirring of the

water. It will be noted that few yellow-green algae are characteris-

tically found in the lower zone.

Below a depth of 4 dm. many algae were found but not in any

charactertistic formation. Evidently they were there because

crowded out elsewhere and they were not growing as well as the same

species at higher levels.

As to grouping of the algae in these zones it was found that in any

given location one or two species were dominant and others grew

rather indiscriminately among them. The dominant species was

apparently determined by the support on which it grew. In a bed of

Scirpus, one species of Chaetophora and Nostoc glomeratum were

almost universally the dominating forms, the Chaetophora dominat-

ing the upper and Nostoc the lower zone. On submerged mosses,

Potomogeton, etc., Gongrosira debaryana, Nostoc glomeratum, or

Rivularia pisum dominated, while at the margin of the lake a free

floating form as Nostoc pruniforme or Rivularia natans might domi-

nate. Clathrocystis aeruginosa or Anabaena flos-aquae usually

62 ANDERSEN AND WALKER

dominated the groups floating at the surface of the water, tho the

constant winds kept these forms fairly well mixed.

SEASONAL DISTRIBUTION

Careful records of dates of collection were made to determine, so

far as possible, the grouping of algae by seasons. As the study ex-

tended only from the middle of June until the middle of October, the

observations included only summer and fall forms. Only the most

general results were obtained. Of the species found in Hackberry

lake the members of the Oscillatoriaceae were found only in the early

summer, Merismopedium and Coelosphaerium only in July. Other

members of the Myxophyceae were found in early stages in the first

part of the season but reached full maturity and maximum abundance

in October.

Most of the green algae were found throughout the period

observed but they reached their maximum in midsummer except in

a few cases. The species of Gloeocystis were found early in the

summer. Bulbochaete was found late in the season (August—October)

and the specimens were not fruiting. Nearly all the desmids were

found during July. Only a few were present earlier or later in this

lake, tho in places near by they occurred at other times in abundance.

The following table shows the forms found in the early, middle,

and later summer and suggests something of the seasonal occurrence

of the forms.

June 27-July 10 July 10-July 25 July 25-August

Nostoc humifusum Nostoc zetterstedtii Nostoc muscorum

Nostoc glomeratum Nostoc pruniforme Nostoc minutum

Nostoc austinii Nostoc humifusum Nostoc caeruleum

Nostoc glomeratum Nostoc pruniforme

Nostoc glomeratum

Anabaena flos-aquae Anabaena flos-aquae Anabaena flos-aquae

Oscillatoria subtilissima

Phormidium tenue

Rivularia pisum Rivularia pisum Rivularia pisum

Rivularia natans (young) Rivularia natans Rivularia natans

Merismopedium aerugi-

neum

Coelosphaerium kuetzing-

ianum

AN ECOLOGICAL STUDY OF ALGAE 63

June 27-July 10

Clathrocystis aeruginosa

Cosmarium vexatum

Cosmarium obtusatum

Cosmarium kjellmani

grande

Closterium lanceolatum

Chara contraria

Coleochaete orbicularis

Chaetophora elegans

Chaetophora cornu-damae

Gloeocystis vesciculosa

Gloeocystis gigas

Scenedesmus bijugatus

July 10-July 25

Clathrocystis aeruginosa

Staurastrum gracile

Spirotaenia obscura

Netrium digitus

Euastrum oblongum

Cosmarium subcrenatum

Cosmarium retusiforme

Cosmarium obtusatum

Cosmarium laeve

Cosmarium granatum

Cosmarium formosulum

nathorstii

Cosmarium elfingii

Closterium pritchard-

ianum

Closterium leibleinii

Chara contraria

Oedogonium fragile

Gongrosira debaryana

Chaetophora elegans

Chaetophora cornu-damae

Microspora amoena

Scenedesmus quadricauda

Scenedesmus obliquus

Dictyosphaerium pulchel-

lum

Pediastrium tetras

Pediastrum boryanum

Tetraedron trigonum

July 25-August

Clathrocystis aeruginosa

Closterium moniliferum

Chara contraria

Bulbochaete sp.

Chaetophora elegans

Chaetophora cornu-damae

A very interesting group of algae was found in a small ditch carry-

ing water from a spring into the northwest end of Hackberry lake.

Closterium lunula was so dominant here that at first it seemed to be

the only species present. Closer examination, however, showed the

following species associated with it.

Eremosphaera viridis

Cylindrospermum majus

Cosmarium angulosum concinnum

Cosmarium circulare

Cosmarium pachydermum

Cosmarium umbilicatum

Euastrum verrucosum

Pleurotaenium trabecula

Staurastrum meriani

Staurastrum punctulatum

Phormidium tenue, P. valderianum, and Spirulina major were

found on the lake shore just above the water’s edge, making the

sand green at many places.

64 ANDERSEN AND WALKER

The following algae were found in this lake:

Chroococcaceae

Clathrocystis aeruginosa

Coelosphaerium kuetzingianum

Merismopedium aerugineum

Oscillatoriaceae

Oscillatoria subtilissima

Phormidium tenue

Nostocaceae

Anabaena flos-aquae

Cylindrospermum majus

Nostoc austinii

Nostoc caeruleum

Nostoc glomeratum

Nostoc humifusum

Nostoc minutum

Nostoc muscorum

Nostoc pruniforme

Nostoc zetterstedtii

Rivulariaceae

Rivularia natans

Rivularia pisum

Bacillariaceae

Cocconeis placentula

Cymbella cymbiformis

Eunotia lunaris

Fragilaria capucina

Gomphonema acuminatum

Gomphonema parvulum

Navicula anglica

Navicula cuspidata

Navicula dicephala

Navicula sculpta

Navicula subcapitata

Navicula viridis

Nitzschia spectabilis

Nitzschia tryblionella

Surirella ovalis pinnata

Desmidiaceae

Closterium lanceolatum

Closterium leibleinii

Closterium moniliferum

Closterium pritchardianum

Cosmarium elfingii

Cosmarium formosulum nathrostii

Cosmarium granatum

Cosmarium kjellmani grande

Cosmarium laeve

Cosmarium obtusatum

Cosmarium retusiforme

Cosmarium subcrenatum

Cosmarium vexatum

Euastrum oblongum

Netrium digitus

Spirotaenia obscura

Staurastrum gracile

Tetras poraceae

Dictyosphaerium pulchellum

Pleurococcaceae

Scenedesmus bijugatus

Scenedesmus obliquus

Scenedesmus quadricauda

Tetraedron trigonum

Urococcus insignis

Protococcaceae

Gloeocystis gigas

Gloeocystis vesciculosa

Hydrodictyaceae

Pediastrum boryanum

Pediastrum tetras

Ulotrichaceae

Microspora amoena

Chaeto phoraceae

Chaetophora cornu-damae

Chaetophora elegans

Gongrosira debaryana

Oedogoniaceae

Bulbochaete sp.

Oedogonium fragile

Coleochaetaceae

Coleochaete orbicularis

Characeae

Chara contraria

AN ECOLOGICAL STUDY OF ALGAE 65

CLEAR LAKE

This lake (Fig. 1, 2, 13) is a little wider and deeper than Hack-

berry lake and is about two-thirds of a mile distant from it. The

water, however, is very alkaline (see discussion of alkalinity) and of a

characteristic yellowish color. About the edge in places were rushes

but there was little other vegetation in the lake. The algal flora was

scarce in species but rich in individuals. Throughout the season the

water was full of Closterium aciculare.'_ With this were found a

few specimens of Cosmarium obtusatum and Pediastrum boryanum,

and a few diatoms in fair abundance but that was all. Since in

Clear lake the algae were all free floating, and the species were

limited, no distribution studies were attempted. The same species

were found throughout the summer. Closterium aciculare was the

only characteristic form and was present throughout the season.

The few other forms present with it were characteristic of the springs

flowing into the lake and probably all came there by chance. It is

probable that they could not have continued to live in so alkaline

a water. Only one factor, alkalinity, could have influenced the

algal flora of this lake.

In some places on the shore of the lake were springs whose water

contained the richest algal flora found in the lake region. This

was especially true at the west end (x in Fig. 13). Here the springs

were on the sloping bank and the water seeped slowly down through

the boggy soil. The surface was covered with grass and ferns forming

sod enough to nearly support a hundred weight or more. Cattle

coming to the spring for water had tramped over this sod forming

holes from the size of a footprint to two or three feet in diameter.

In these holes (Fig. 14) the water stood undisturbed for long periods

with the thick grass sheltering the surface from the wind. The result

was a large number of small aquaria. The water in these was kept

uniformly fresh by seepage from the springs. While no chemical

analysis was made, it was noticeable that the alkalinity was low.

‘ This form in size fits the description of the species as given by West and West

(S1) but may be the form referred to by West (50) as Closteriopsis longissima tho some

of the specimens were over 600u long and 7p wide. He suggests that Closteriopsis

may be a degenerate of Closteriwm aciculare var. subpronum.

66 ANDERSEN AND WALKER

The temperature was fairly uniform throughout the summer period

varying from about 68° F. in the early morning to about 80° F. at

3P.M. So far as was observed the ecological conditions in the various

pockets were remarkably uniform in every way. Each was domina-

ted by some one species, the usual dominants being Anabaena toru-

losa, various species of Nostoc, Spirogyra, Scytonema, Oedogonium,

and Mougeotia. With these were mixed in various proportions the

other species found. Attempts were made to determine whether

there was any uniformity as to the species present with a given

dominant but there seemed to be none.

The only ecological groupings of forms observed were seasonal.

As will be seen from the accompanying list some forms were found

only early in the summer, others late in the summer, and still others

in the fal. The greatest variety of species was found in the middle

of the summer. In the case of unicellular forms this list can only be

looked upon as suggestive. Only a few specimens are found for some

of them and chance in collecting may have caused others to be over-

looked in one period or another. In this table early summer means

from the middle of June to the middle of July; midsummer from the

middle of July to the middle of August; fall is represented by collec-

tions made in October. Here fresh water, uniform temperature,

protection from wind, and very shallow water form a habitat as

uniform in every respect as is possible. Here the dominant forms

found in a given water pocket can be due only to one of two things,

seasonal periodicity which was very evident and the chance domi-

nance of certain forms. Often the soil beneath a mass of Spirogyra,

Mougeotia, or other such form was covered with desmids or diatoms.

It is apparent that smaller forms, especially unicellular ones, may

be shaded by the dominant ‘species and hence their presence would

be determined by their light relation. However, which of these

shade loving forms should be associated with a given dominant species

was chance so far as was determined. At other places on the lake

shore were springs where a few algae were found but they were of

so little consequence that the species were included only in the

general list of species.

AN ECOLOGICAL STUDY OF ALGAE 67

CLEAR LAKE PUDDLES

Early Mid- October

summer summer

Chroococcaceae

BepbanOTHeCe PASIAN eis ciple ce ts 2's | prahve ie lars'e sages My Ns Bl ee asiale ale wi eke

ISIn thro cyStis acruUpIBOSA de ie sole cies eletseeie cl ot clone eons AM) Aheue eaciuine ee

EROGGEADSAATCHATIA Santer cir eile sic cicisls's fora gece bmi letei eis > PE ACH SE Eee aA

Oscillatoriaceae

HOST ALOnIAMOLMOSA yes ws eiclne cies lols) sh ellisyels) seals ciate e > Sa ALT Sia etcin trie

MsaMlatonialimosa sec. e + sceiersk sed eic ss oA AUllohart ree ea tO a a

PATTIMICIMNLENIU Cases iettcs ale cies) [ee anne wgiclee 2 Nira WEI eae te

PHormidium valderianum’. 2). 2. es 6c. sl suse ee ws wae Bist) FINS can Maca meee

Nostocaceae

mabaena fOs-aquae.<. 2)... csc ease x RMN cam en a aaceaon

Pana tenarOscillanoidesis vise dalss 5 cine waliecalleanlonies ey eee XE HAN (tater Acie ab ee ©

Pima eM Ay COTUlOSA. sess era e alas « c/a)s, cai x x x

SMMLOsperna win COMALCM, .!)\0).5 5 fcc of vite vc dc tiele wallebioicie sists Be x

Nodularia harveyana .............. arstedel (peach aye eptek as KI hina EN eS ce

TumEMEMPBU IAG Kota oy St EN so loa boaikig Say aN tales a URE x

SNe MURIEERESR GE LY TV N25 15000272, 41 218 15/2 ke Xl cbse niflal pits #i a's eel > LUN Aone eee A

Pree MUSCOTUIM 2... ws oe cio vines ss x D-ce ee Mt reas kas ah une

Mgstoc'spongiaeforme.................. KEN Pel arogeigaph ticle Fanart pecais tae ene eee ait

Scytonemaceae

Seemerema Crispum:. 2.2.6 eck sce es x x x

Rivulariaceae

(LT TEES a CN ST AR Ue aM aad en VC x

Bacillariaceae* Cystopleura zebra

Achnanthes lanceolata Eunotia diodon

Cyclotella meneghiniana Eunotia lunaris

Cymbella amphicephala Eunotia major

Cymbella cuspidata Fragilaria construens

Cymbella naviculiformis Gomphonema acuminatum

Cymbella subaequalis Gomphonema constrictum

Cystopleura gibba Gomphonema gracile

Cystopleura gibba ventricosa Gomphonema montanum

* No seasonal studies were made of diatoms.

Melosira varians

Navicula ambigua

Navicula anglica

Navicula bacilliformis

Navicula brebissonii

Navicula cuspidata

Navicula dicephala

Navicula elliptica

Navicula gibba brevistriata

Navicula hilseana

Navicula major

Navicula mesolepta

Navicula iridis

ANDERSEN AND WALKER

Navicula pupula

Navicula sculpta

Navicula sphaerophora

Navicula stauroptera

Navicula viridis

Nitzschia brebissonii

Stauroneis anceps

Stauroneis anceps amphicephala

Stauroneis phoenicenteron

Surirella ovalis ovata

Synedra rumpens

Synedra ulna

Early Mid- October

summer summer

Desmidiaceae

Arthrodesmus convergens..............- x x x

Glostertumcynthias sn. sia: iets tases apiece x x x

Closterium didymotocum............... XK’ © Wades saci ee

@lostertumijennenlame.s seer ieee i HCl et.0) Soe oS | eee

Glosteniumrlanceolatuimep say. eee selene clsteieiis Chie cieeel|leeieisoe eee x

Clostenumblunulatee ere ceric x x. x

Closterium! monilifemim=..- =. oe a x x x

Glostentum parvulam acs 26512. felts eatets lio oratorereisin seni x x

Glosteritum pritchardianum cis) .5 cis aie leel||- stcteipiein sod sts [ses aes vs ote ee x

Glostersumpsiliqua sss semis ee raciie ce lsketesl| Ootsaes erciamieta tillorece eae eee x

G@losteriumistnolatumijen- eerie eet ae Kx )o..... 005082 eee

Clostertumiturciduntessereie ace ey ZX Vole aces) eee

Cosmariumabrptumeree rate seer ioe (teria ein it ae > REE FSS B80 0.c.c

Cosmaniumbangulostmbe amare aia ede| [eves creaeterses. til operas epee x

Cosmarium angulosum concinnum.......|............ x x

Cosmarium | boeckineesr cere ees sete Xo) eee eee x

Cosmariumibly bite anes bacon eet a|'civaens oats x x

Cosmarium botrytis tumidum........... > nn EE erE mcs |ocaccsocéolr

Gosmanium'circularesaseere eee e een Sh) AR x

Gosmanumiconnatomipe nent ery ceric eer x x x

Cosmanumicrenatum=ery eee ce > EE PAPA apa lob crate bclibc¢

Cosmarniumicychicuimiay rer cri ctr ee ie ene XW 0 wks oe eee

Cosmarium cymatopleurum............. > EE PREM) la,ccc 66 Oot 3+

Cosmarium formosulum nathrostii.......]............ x x

(Cosmaniumipalenttumepey eee ee eee o| cosine eee eee x; Verse eyepee

Cosmanumigranarumprmmen ees etic ee cic ares x x

AN ECOLOGICAL STUDY OF ALGAE 69

Early Mid- October

summer summer

Cosmarium*holmiensez). 25.4). 5 ccs seenle as x AG Heer x

Cosmarium)jkellmani grandeé< e240: nieces oe eres es x Seep é i

COSMALUMIIMENEL MUI leet eisi te sae dale <0 sillapare sin ci aoslene p clair aaa CNN Es

Cosmarium microsphinctum............. Xone MOT Lil ieveiettysccreee setae eeettegs Drahees ete

Cosmanum/ notabiley sos: cscs cies ys SVM ONT PES ace, cect anctra Panay bale AE EVEL MELA,

Cosmarium obtusatum........... oe x s aNslU po ohauel at aiabes| (sltentehoteee iota: !

Cosmarium ochthodes....... seit aee Pate te gS] [rite ae Me ela bee x asierne Bante

ORME PACDYGETMUAN: 6. o..s isies |e os cess wooo 8 x x

Cosmarium pachydermum aethiopicum. . . KDWP teers uvattee ae x

Cosmarium phaseolus elevatum...... SN HPN A Sa x al have te betelileyie

Cosmarium phaseolus forma minor....... > I RH | SPT eee cre bi Apesnaea SHooe

Rosmarium portianum.:...)......... 26525. SOUT esters at ie ek ate x

Cosmarium pygmaeum................. x alate tieictey sternal rere bln Bee

Brenaouen) PyTAMIGatwm so...) ss =n [esc eie dus cess x x

Cosmarium rectangulare hexagonum.....]........ Ee Xi x

Cosmarium sportella.......... Ay ain ab enact Meelis faye tes x Mi par Mak is toon 8

Cosmarium subcrenatum...... SY SANs x » AWE ele eeeee NP :

Cosmarium subtumidum...... Nonsesins al recon atid bests ba 2 Kalender oeue

Cosmarium subundulatum.............. KHIM ai eens vee emer sitions aye anus ae

Cosmarium taxichondrum...............].. ay Maa DN Mm a HES ete ee

Cosmarium tetraophthalmum........ Sal a Sey eee és Keg yl) es eee cca fete

Cosmarium trilobulatum................].. eben hepa elats x Beth as Soaionas

PUREE TUMETEN SE UATE HE Flos a Aira laine afte te kee seatac x Er eistoniotals

Cosmarium vexatum....... vats : NERVE POS Sry dl targetsoeer é

Euastrum attenuatum.............. Siete veil avetanetecese He.cluinl isomerate x

Semeersisri Dicken tatu: 1/542 6 4. hye els reine eile wincceds c retail >.< VES Caste bs oye ls

oS CETE SST RTT a OTE a ae feet li x x

Bwastrum dubium... 2.0.6... 4 <0: SAU Wo seven se x Sood ‘

Euastrum oblongum........ SIGE Roi tel | Tae Rea , x x

Euastrum verrucosum........ Ne eee et: x x x

Huastrum! verrucosum alatum. ...02.0%. 04 o<s0ces. 0: x x

Micrasterias pinnatifida............ pieeiebaiticictel aeanoeste teeta x x

PMEEASECTING FOCAEA (a2 C lia niet cise MIR ile enie a Lar ae me calvuey aetee, baci ats x

BRE ERINEL CIpAT US 5,5) yc) ches RA eM Mn aus ey x x

SMe MUMEANLSITUp CUI 3 <4 seks hess ui oleic teas hao e cs | (2 aa rea ese x

Penium libellula intermedium............)...... eal epee NE ate x

BEGIN NACPELL. s.... 6. sales: sal edtreieustleys rey SSA RD aT x Beil alahabache eave

emia SpirostriOlatim. .%) 4 s..c.isea rae ele yaa gens aaes x x

Pleurotaenium coronatum..... aaa ee ANS x x x

Pieurotaeniunl NOduUIOSUM, ..../.{2). 22 dood Posawele ce eae < x a caett ast sieetere

Pleurotaenium trabeculata........... ie 6 Sit R| ILE eM GHOULS aur tay Bae

70 ANDERSEN AND WALKER

Early Mid- October

summer summer

Spirotaenia CONGENSAtA Ls e:2))s cinta» ainfovcietsl| falter ein ioiua rs x x

Spirotaenia trabeculatay.).))s/ejsioe urs olen) steal ne sie ace osm iets i Ce oie ener

Staurastrumilaltemans sae tees cne ieee te] ouctevetea a eisioetel |e tare ree x

Staurastrumidickieltva qari eee tee ese tank x x

Staurastrumidilatatum seyioe ce iscisiirelericel| cette ie creer ell aiaiey oto slo a

Staurasteumi(dispars sy. 2. cis /2'eie). 6 sro stoves eta are alometerotet nse Xs eee eee

Staurastrumphirstitumssan sickis ents -iaroeya | erie erent x x

Staurastrumimargaritaceumlsccjs-s2 sis 1el|sevaelaine oe ccietc x xe

Staurastrumimentanlemery pene vercse eit erase cis eee alia teeters oie x

Staurastrumpmuticumys says cect etyeii ebe| secieeae epee X74 ee ees

Staurastrumforbiculare :2)2 fo oe etc iel-s.e vier x x x

Staurastrumporbiculare ralisliy.ciereicrits iiss cites eet aee Xt) | UGS eee

Staurdstrunajpaxd leer sic.) 5) hi abe) ostveje cts | sues ciel cial nels Miter t teins x

Staurastrum polytrichum............... Ko) OY [oa <syete ele oral areca eee

Staurastrum punctulatum............... x x x

Staurastrumisaxonicum Sere see crisis eisiee rept tiae Xi) 0) lease

Staurastrumiteliferum en ries iis elie salarel|icreiseel ete soot re pean WI EAS ets yeh. A

Zygnemaceae

Spirogyra arcta catenaeformis........... MY TL idiots elg ore! svoes Uo he Seen ee

SpMORyiACLASSAnc cs). ss\vlon cee ater an cle's > AMER: CORR PENI ESI Wrecks 5

Spirogyra Gublaaee cesta. Rive siecle x x x

Spiropyra lutetianar .2- cisco en <0) -eieivelsie Kh atthe st. oe ee rr

Splrageyaa Mer lecka mam cere piss steve iste alia Ste folaioxel eet i2ie (ate ol lovee lore ere halal x

Spirogyra WAniamS a yom re oe iostseyae siete axe | ele seye layers): os fal eheie te eiela ame jorels x

LY ENEMAS ace eee eters afore elelet oka: x x x

Mesocar paceae

Mougeotiarobuscaere rn carr ici c Gist ielll-ireys este, cie erie | slave e eee amet x

IMonpzeotia Sealarise ret nies ct leiayale sida iaeeiels ic avi viaiisl| s/.e oy aE x

Mougeotia viridiste.o4--ris fee seco > EE REARS alles ooo. coc

Volvocaceae

Volvox(aureuste mae see itocncioi eee ice |siaesiSeniens ct p a RIE ime

Tetras poraceae

Dictyosphaerium pulchellum............].........00. x x

Tetraspora Pelatinosa ye. c ses o ices sire = > ne Ere AN a aotMino koto o¢

AN ECOLOGICAL STUDY OF ALGAE pint

Early

summer

Pleurococcaceae

Mid- October

summer

Crucigeniamrectangularish ss sste tots sre ache teh cletate Rotini Gee Un Seeds x

PEMEMOSPUACTS VIFIGIS s siais/oiG (0,6 Sie ceheey oi erence ffele mveid ladle one ate

Nephrocytiummaegeliiyete ee lye oe see erate x

OGYSHSISOWMPa RIA ha tects chee evel elaraisisr ede Silt aeevers ature ees

Rhaphidium polymorphum falcatum.....|............

Scenedesmus antennatus...............

Scenedesmus bijugatus.................

ee sO ?

PPEErACORONETe LIC MAGLI Mee crepes seal ceat ale | aU spai cient anual abereia eis eee x

RU OGOECUSHIITSIOTIS Mean U Nis ae. yey e k toctiorclli aly USUI MOIA SMR RO PRN ULEE CN IR x

Protococcaceae

Gloeocystis vesciculosa................- x

Ps@cy Aa CAPYER TUT Veale Na AO IMMUN A) x

Hydrodictyaceae

BHSIALHOMISPNACHICUNA: (eis s/c) \ 5 waders Coarse) ates Malaoals cans

Pediastrumiang ulosmany yer narsra aia ans Mh ae | MINUS MCN) ee alia Cie | x

Pegiastriusy DOryanUHN |). 20.2 a), ie ake S x

IBEGIASEMIMLLELEAS Sainte MEL Nein hokey aca MAIN Ujanaaiah

Ulotrichaceae

iiterosporaypachy derma! cei Milk aw eC As

IMIGrOSpOTA SLAM ORM oho) e)dsrcitie se clase dais hess enol nis

Chaetophoraceae

Stigeoclonium glomeratum.............. x

Oedogoniaceae

| SUL OCS Sut o aed Aiea Uo ey Ache diate RU A a] WAS Aa x

Gedagoniam: spi:'4 +4 sageteisetak awe oes x

Oedogonium capilliforme australe........}............

Oedogonium cardiacum.. 2 .j)..0460.) 005). x

Oedogonium crispum uruguayense........ x

GCraoronnim: fragile yoy acy ec ntars eat uate eh eat es

economy Varian’). 0 igh ras hy aba sia oUt el Mia tis Cena x

Cladophoraceae

Rhizoclonium hieroglyphicum........... x

oija| eiejislelle|.eveol/al se} syeltm («reise .eueteleia

de ANDERSEN AND WALKER

V aucheriaceae

Early Mid- October

summer summer

Vaueheria Spices cian a oe hots cre ree © eet ete leliicte a ne arate e aera tate a x

BIG ALKALI LAKE

Big Alkali Lake (Fig. 1) is a little larger than Clear lake. While it

is more alkaline than Hackberry, Dewey, or Watts, it is far less so

than Clear lake. The water had the characteristic yellow color of

the alkaline lakes of this region.

Here again alkalinity must be

given as the factor governing the algal flora of the lake. One visit

only was made here. At first sight the water seemed absolutely barren

but further investigation showed the following forms to be present.

The Chara was quite abundant forming very much dwarfed patches

on the bottom near the shore.

Chroococcaceae

Clathrocystis aeruginosa

Merismopedium glaucum

Merismopedium tenuissimum

Oscillatoriaceae

Oscillatoria limosa

Bacillariaceae

Amphora ovalis

Campylodiscus clypeus

Cymbella cistula

Cystopleura gibba

Navicula cryptocephala veneta

Navicula gastrum

Navicula oblonga

Navicula sculpta

Desmidiaceae

Cosmarium angulosum

Cosmarium granatum

Cosmarium meneghinii

Cosmarium sexnotatum

Staurastrum gracile

Staurastrum paradoxum

Staurastrum polymorphum

Tetras poraceae

Dictyosphaerium pulchellum

Pleurococcaceae

Scenedesmus bijugatus

Scenedesmus obliquus

Scenedesmus quadricauda

Hydrodictyaceae

Pediastrum boryanum

Pediastrum duplex clathratum

Characeae

Chara foetida rabenhorstii

DEWEY LAKE

Dewey lake (Fig. 1) is situated about half a mile from Hack-

berry lake. It is less alkaline, nearly three times as large and propor-

tionally deeper. Otherwise the conditions were much the same.

Temperatures taken in the region of algal growth showed little varia-

tion from those in Hackberry lake.

AN ECOLOGICAL STUDY OF ALGAE 73

The algae were found along the margins of the lake and attached

to submerged plants near the surface. As no attempt was made to

study conditions in this lake no explanation of the conspicuous differ-

ence in the forms found can be given unless it was the larger amount

of water and difference in alkalinity. Collections were made here at

irregular intervals and the list must not be looked upon as complete.

Chroococcaceae

Clathrocystis aeruginosa

Merismopedium glaucum

Oscillatoriaceae

Beggiatoa alba

Lyngbya aerugineo-caerulea

Oscillatoria amphibia

Oscillatoria formosa

Oscillatoria subtilissima

Phormidium fragile

Phormidium tenue

Phormidium valderianum

Nostocaceae

Anabaena flos-aquae

Nostoc linckia

Nostoc pruniforme

Scytonemaceae

Tolypothrix distorta

Rivulariaceae

Rivularia echinulata

Rivularia natans

Bacillariaceae

Amorpha ovalis

Brebissonia vulgaris

Cocconeis placentula

Cymbella cistula

Cymbella cuspidata

Cymbella lanceolata

Encyonema turgidum

Eunotia lunaris

Fragilaria capucina

Fragilaria construens binodis

Gomphonema constrictum

Gomphonema gracile

Gomphonema montanum

Gomphonema parvulum

Navicula cuspidata

Navicula lanceolata

Navicula major

Navicula oblonga

Navicula pupula

Staureneis acuta

Stauroneis smithii

Stauroneis tenuissima

Synedra rumpens

Synedra ulna

Desmidiaceae

Closterium pritchardianum

Cosmarium blyttii

Cosmarium boeckii

Cosmarium formosulum nathorstii

Cosmarium impressulum

Cosmarium obtusatum

Cosmarium ochthodes var.

Cosmarium subcrenatum

Cosmarium turpinii podolicum

Cosmarium vexatum

Penium margaritaceum

Staurastrum orbiculare

Pleurococcaceae

Scenedesmus bijugatus

Scenedesmus obliquus

Tetraedron trigonum

Protococcaceae

Characium ambiguum

Characium subulatum

Gloeocystis vesciculosa

Hydrodictyaceae

Pediastrum boryanum

Ulotrichaceae

Hormiscia subtilis variabilis

Chaetophoraceae

Chaetophora elegans

Gongrosira debaryana

Stigeoclonium aestivale

74 ANDERSEN AND WALKER

Stigeoclonium glomeratum

Oedogontaceae

Oedogonium grande

Oedogonium vaucherii

Helminthocladiaceae

Batrachospermum vagum

WATTS LAKE

Watts lake (Fig. 1) is about half the size of Hackberry and only

about one-third of a mile from it. Conditions in the lake were very

similar to those in Hackberry in every respect. It was, however,

slightly less alkaline than Dewey lake.

Collections were made here about once a week but other data

were not taken. No boat was available on this lake and that may

account for some of the difference in the reports for Watts and

Hackberry lakes. No reason for a difference in species could be

given unless it were the slight difference in alkalinity. The following

species were found which it will be noted are nearly all included in

those found in Hackberry and Dewey lakes and the springs on the

shore of Clear lake.

Cosmarium geminatum

Cosmarium granatum

Cosmarium obtusatum

Cosmarium phaseolus

Cosmarium pseudopyramidatum

Cosmarium subcrenatum

Staurastrum gracile

Staurastrum margaritaceum

Chroococcaceae

Clathrocystis aeruginosa

Coelosphaerium kuetzingianum

Merismopedium tenuissimum

Microcystis marginata

Nostocaceae

Nostoc pruniforme

Bacillariaceae Tetras poraceae

Achnanthes lanceolata Dictyosphaerium pulchellum

Amphora ovalis Pleurococcaceae

Cocconeis placentula

Gomphonema gracile

Gomphonema montanum

Homoeocladia amphioxys

Navicula gastrum

Oocystis solitaria

Scenedesmus bijugatus

Scenedesmus obliquus

Scenedesmus quadricauda

Tetraedron minimum

Navicula lanceolata Hydrodictyaceae

Desmidiaceae Pediastrum boryanum

Cosmarium boeckii Characeae

Cosmarium formosulum nathorstii Chara contraria

OTHER LAKES

One visit was made to each of the following lakes in the early

summer. Trout and Dad’s lakes (Fig. 1) are among the larger of the

AN ECOLOGICAL STUDY OF ALGAE 75

group while Phalaris (Fig. 1) is one of the smaller. The Snake

Creek Falls, about 15 miles distant, were visited once. These are

falls in a creek which flows through the sandhill region. These

collections were made so superficially that the lists must stand only

as representing some species found in these localities.

The only form found which was sufficiently conspicuous to need

special mention was Nostoc verrucosum which was extremely abundant

on rocks in the cataract below the falls in Snake Creek.

The forms found in these localities are as follows:

PHALARIS LAKE

Chroococcaceae

Coelosphaerium kuetzingianum

Merismopedium tenuissimum

Dactylococcopsis rhaphidioides

Bacillariaceae

Amphora ovalis

Cocconeis placentula

Cystopleura gibba

Cystopleura turgida

Gomphonema montanum

Homoeocladia amphibia

Navicula cryptocephala veneta

Navicula cuspidata

Navicula elliptica

Navicula gastrum

Navicula oblonga

Navicula sculpta

Navicula sphaerophora

Sceptroneis fibula

Chroococcaceae

Clathrocystis aeruginosa

Nostocaceae

Anabaena flos-aquae

Nostoc zetterstedtii

Chroococcaceae

Clathrocystis aeruginosa

Desmidiaceae

Closterium acerosum

Desmidiaceae

Cosmarium angulosum

Cosmarium blyttii

Cosmarium formosulum nathorstii

Cosmarium granatum subgranatum

Cosmarium obtusatum

Cosmarium regnellii

Tetras poraceae

Dictyosphaerium pulchellum

Pleurococcaceae

Oocystis solitaria

Pediastrum boryanum

Scenedesmus bijugatus

Scenedesmus quadricauda

Tetraedron minimum

Characeae

Chara contraria

Chara evoluta

Chara fragilis

TROUT LAKE

DAD’S LAKE

Chara sp.

Hydrodictyaceae

Pediastrum angulosum

Characeae

Chara sp.

Tetras poraceae

Dictyosphaerium pulchellum

Hydrodictyaceae

Pediastrum boryanum

Pediastrum duplex clathratum

76 ANDERSEN AND WALKER

Chroococcaceae

Merismopedium glaucum

Oscillatoriaceae

Arthrospira jenneri

Oscillatoria brevis

Oscillatoria formosa

Phormidium retzii

Nostocaceae

Nostoe pruniforme

Nostoc verrucosum

Rivulariaceae

Calothrix parietina

Bacillariaceae

Achnanthes lanceolata

Amphora ovalis

Cocconeis placentula

Cymbella amphicephala

Cymbella cuspidata

Cymbella ehrenbergii

Cymbella gastroides

Cystopleura gibba

Cystopleura ocellata

Cystopleura turgida

Cystopleura zebra

Denticula elegans

Encyonema turgidum

Eunotia major

Fragilaria mutabilis

Gomphonema acuminatum

Gomphonema herculeanum

Lysigonium crenulatum

Lysigonium distans

Navicula ambigua

Navicula anglica

Navicula appendiculata

Navicula brebissonii

Navicula cuspidata craticula

Navicula dicephala

Navicula elliptica

SNAKE FALLS

Navicula humilis

Navicula iridis

Navicula lanceolata

Navicula limosa

Navicula gibba brevistriata

Navicula mesolepta

Navicula pupula

Navicula radiosa

Navicula sculpta

Navicula viridis

Homoeocladia amphibia

Homoeocladia brebissonii

Homoeocladia amphioxys

Homoeocladia palea

Rhoicosphenia curvata

Sceptroneis pacifica

Stauroneis anceps

Stauroneis phoenicenteron

Surirella robusta

Surirella spiralis

Synedra ulna

Tetracyclus lacustris

Desmidiaceae

Closterium striolatum

Cosmarium microsphinctum

Cosmarium portianum

Cosmarium sportella

Cosmarium undulatum wollei

Euastrum oblongum

Euastrum verrucosum

Penium margaritaceum

Staurastrum orbiculare hibernicum

Pleurococcaceae

Scenedesmus obliquus

Protococcaceae

Chlorococcum humicola

Cladophoraceae

Cladophora glomerata

CONCLUSION

It appears even from so brief a study as the one just described that

the occurrence of algae in a given body of water at a given time is

AN ECOLOGICAL STUDY OF ALGAE ih

due, to a certain extent, as Transeau (44), West (49), and others have

said, to seasonal periodicity.

It is also evident that West (48 and 50), Oltmanns (34), Brannon

(12), Wipple and Parker (53), Chambers (15), and many others are

correct in their decisions that the mineral and gas content of water

has much to do with its algal flora. Of these factors, alkalinity is

probably to a great extent, the explanation for the wide difference

in the algal flora of lakes so close together and so uniform in all other

factors.

In a given lake the distribution of species may be explained by the

one factor only that is variable, namely light intensity. Means for

measuring this factor were entirely inefficient and only the crudest

estimates can be made.

In small bodies of water where even the light is not variable to

any measurable degree the dominant species and its associates are

determined merely by chance except that forms lying beneath other

forms are more shaded. This exception does not affect the dominant

species but may affect the forms associated with it.

ALGAE FOUND IN CHERRY COUNTY

Chroccoccaceae. Oscillatoria limosa. (Roth) Ag.

Aphanothece prasina A. Braun

Clathrocystis aeruginosa (Kuetz.)

Henfrey

Coelosphaerium kuetzingianum Naeg.

Dactylococcopsis rhaphidioides Hansg.

Gloeocapsa arenaria (Hassall) Rabenh.

Merismopedium aerugineum Bréb.

Merismopedium glaucum (Ehrb.)

Naeg.

Merismopedium tenuissimum Lem-

merm.

Microcystis marginata (Menegh.)

Kuetz.

Oscillatoriaceae.

Arthrospira jenneri (Kuetz.) Stiz.

Lyngbya aerugineo-caerulea (Kuetz.)

Gom.

Oscillatoria amphibia Ag.

Oscillatoria brevis (Kuetz.) Gom.

Oscillatoria formosa Bory.

Oscillatoria princeps Vauch.

Oscillatoria sancta (Kuetz.) Gom.

Oscillatoria subtilissima Kuetz.

Oscillatoria tenuis Ag.

Phormidium fragile (Menegh.) Gom.

Phormidium retzii (Ag.) Gom.

Phormidium tenue (Menegh.) Gom.

Phormidium valderianum (Delp.)

Gom.

Spirulina major Kuetz.

Nostocaceae.

Anabaena flos-aquae (Lyngb.) Bréb.

Anabaena oscillarioides Bory.

Anabaena torulosa (Carmich.) Lager-

heim

Cylindrospermum comatum Wood

Cylindrospermum majus Kuetz.

Nodularia harveyana(Thwaites) Thuret

Nostoc austini Wood

Nostoc caeruleum Lyngbye

Nostoc commune Vaucher

Nostoc glomeratum Kuetz.

Nostoc humifusum Carmichael

Nostoc linckia (Roth) Bornet

Nostoc minutum Desm.

Nostoc muscorum Ag.

Nostoc pruniforme (L.) Ag.

Nostoc spongiaeforme Ag.

Nostoc verrucosum (L.) Vauch.

Nostoc zetterstedtii Areschoug

Scy tonemaceae.

Scytonema crispum (Ag.) Bornet

Tolypothrix distorta (Hofm. B.) Kuetz.

Rivulariaceae.

Calothrix parietina (Naeg.) Thur.

Rivularia echinulata (Smith) Born

Rivularia natans (Hedw.) Welw.

Rivularia pisum Ag.

Bacillariaceae.

Achnanthes lanceolata (Bréb.) Gr.

Amorpha ovalis (Bréb.) Kuetz.

Brebissonia vulgaris (Thwait) Kunze

Campylodiscus clypeus Ehr.

Cocconeis placentula Ehr.

Cyclotella meneghiniana Kuetz.

Cymbella amphicephala Naeg.

Cymbella cistula (Hempr.) Kirchn.

Cymbella cuspidata Kuetz.

Cymbella cymbiformis (Kuetz.) Bréb.

Cymbella ehrenbergii Kuetz.

Cymbella lanceolata (Ehr.) Kirch.

Cymbella naviculiformis Auersw.

Cymbella subaequalis Grun.

Cystopleura gibba (Ehr.) Kunze

Cystopleura zebra (Ehr.) Kunze

Encyonema turgidum (Greg.) Grun.

Eunotia diodon Ehr.

Eunotia lunaris Grun.

Eunotia major (W. Sm.) Rabenh.

Fragilaria capucina Desmaz.

Fragilaria construens (Ehr.) Grun.

Fragilaria construens binodis (Ehr.)

Grun.

Gomphonema acuminatum Ehr.

Gomphonema constrictum Ehr.

ANDERSEN AND WALKER

Gomphonema gracile Ehr.

Gomphonema herculeanum Ehr.

Gomphonema montanum Schum.

Gomphonema parvulum Kuetz.

Homoeocladia amphibia (Grun.) Kunze

Homoeocladia amphioxys (Ehr.) Kunze

Homoecladia brebissonii (H. Sm.)

Kunze

Homoeocladia palea (Kuetz.) Kunze

Lysigonium crenulatum (Kuetz.) Kunze

Lysigonium distans (Kuetz.) Kunze

Lysigonium varians (Ag.) D.T.

Navicula ambigua Ehr.

Navicula anglica Ralfs

Navicula appendiculata (Ag.) Kuetz.

Navicula bacilliformis Grun.

Navicula brebissonii Kuetz.

Navicula cryptocephala veneta

(Kuetz.) Rhabenh,

Navicula cuspidata Kuetz.

Navicula dicephala Ehr.

Navicula elliptica Kuetz.

Navicula gastrum Ehr.

Navicula gibba (Ehr.) Kuetz.

Navicula gibba brevistriata Grim.

Navicula hilseana Jan.

Navicula humilis Donk.

Navicula iridis Ehr.

Navicula lanceolata Kuetz.

Navicula limosa Kuetz.

Navicula major Kuetz.

Navicula mesolepta Ehr.

Navicula oblonga Kuetz.

Navicula pupula Kuetz.

Navicula radiosa Kuetz.

Navicula sculpta Ehr.

Navicula sphaerophora Kuetz.

Navicula stauroptera Grun.

Navicula subcapitata Greg.

Navicula viridis Kuetz.

Nitzschia brebissonii W. Sm.

Nitzschia spectabilis (Ehr.) Ralfs

Nitzschia tryblionella Hantzsch.

Rhoicosphenia curvata Grun.

Sceptroneis fibula (Bréb.) Schuett

AN ECOLOGICAL STUDY OF ALGAE 79

Sceptroneis pacifica (Grun.) Elmore (In

press)

Stauroneis anceps Ehr.

Stauroneis anceps amphicephala Kuetz.

Stauroneis acuta W. Sm.

Stauroneis phoenicenteron Ehr.

Stauroneis smithii Grun.

Surirella ovalis ovata (Bréb.) V.H.

Surirella ovalis pinnata (Bréb.) V.H.

Surirella robusta Ehr.

Surirella spiralis Kuetz.

Synedra rumpens Kuetz.

Synedra ulna (Nitzsch.) Ehr.

Tetracyclus lacustris Ralfs

Desmidiaceae.

Arthrodesmus convergens Ehrenb.

Closterium acerosum (Schrank)

Ehrenb.

Closterium aciculare Tuffen West

Closterium cynthia DeNot.

Closterium didymotocum Corda

Closterium jenneri Ralfs

Closterium lanceolatum Kuetz.

Closterium leibleinii Kuetz.

Closterium lunula (Muell.) Nitzsch.

Closterium moniliferum (Bory).

Ehrenb.

Closterium parvulum Naeg.

Closterium pritchardianum Arch.

Closterium siliqua West and G. S.

West

Closterium striolatum Ehrenb.

Closterium turgidum Ehrenb.

Cosmarium abruptum Lund.

Cosmarium angulosum Bréb.

Cosmarium angulosum concinnum

(Rabenh.) West and G. S. West

Cosmarium blyttii Wille

Cosmarium boeckii Wille

Cosmarium botrytis tumidum Wolle

Cosmarium circulare Reinsch.

Cosmarium connatum Bréb.

Cosmarium crenatum Ralfs

Cosmarium cyclicum Lund.

Cosmarium cymatopleurum Nordst,

Cosmarium elfingii Racib.

Cosmarium formosulum nathorstii

(Boldt) West and G. S. West

Cosmarium galeritum Nordst.

Cosmarium geminatum Lund.

Cosmarium granatum Bréb.

Cosmarium granatum subgranatum

Nordst.

Cosmarium holmiense Lund.

Cosmarium holmiense integrum Lund.

Cosmarium impressulum Elfv.

Cosmarium kjellmani grande Wille

Cosmarium laeve Rabenh.

Cosmarium meneghinii Bréb.

Cosmarium microsphinctum Nordst.

Cosmarium notabile Bréb.

Cosmarium obtusatum Schmidle

Cosmarium ochthodes Nordst. var.

Cosmarium pachydermum Lund.

Cosmarium pachydermum aethiopi-

cum West and G. S. West

Cosmarium phaseolus Bréb.

Cosmarium phaseolus elevatum Nordst.

Cosmarium phaseolus minor Boldt

Cosmarium portianum Arch.

Cosmarium protractum (Naeg.) De-

Bary

Cosmarium pseudopyramidatum Lund.

Cosmarium pygmaeum Arch.

Cosmarium pyramidatum Bréb.

Cosmarium rectangulare hexagonum

(Elf.) West and G. S. West

Cosmarium regnellii Wille

Cosmarium retusiforme (Wille) Gutw.

Cosmarium sexnotatum Gutw.

Cosmarium sportella Bréb.

Cosmarium subcrenatum Hantzsch

Cosmarium subtumidum Nordst.

Cosmarium subundulatum Wille

Cosmarium taxichondrum Lund.

Cosmarium tetraophthalmum Bréb.

Cosmarium trilobulatum Reinsch

Cosmarium tumidum Lund.

Cosmarium turpinii podolicum Gutw.

Cosmarium umbilicatum Luetkem.

Cosmarium undulatum wollei West

Cosmarium vexatum West

Euastrum attenuatum Wolle

Euastrum bidentatum Naeg.

Euastrum binale (Turp.) Ehrenb.

Euastrum dubium Naeg.

Euastrum oblongum (Grev.) Ralfs

Euastrum verrucosum Ehrenb.

Euastrum verrucosum alatum Wolle

Micrasterias pinnatifida (Kuetz.) Ralfs

Micrasterias rotata (Grev.) Ralfs

Netrium digitus (Ehrenb.) Itzigs and

Rothe

Netrium interruptum (Bréb.) Luetkem.

Pleurotaenium coronatum (Bréb.)

Rabenh.

Penium libellula (Focke) Nordst.

Penium margaritaceum (Ehrenb.)

Bréb.

Penium naegelii Bréb.

Penium spirostriolatum Barker

Pleurotaenium nodulosum (Bréb.) De-

Bary

Pleurotaenium trabecula (Ehrenb.)

Naeg.

Spirotaenia condensata Bréb.

Spirotaenia obscura Ralfs

Spirotaenia trabeculata A. Br.

Staurastrum alternans Bréb.

Staurastrum dickiei Ralfs

Staurastrum dilatatum Ehrenb.

Staurastrum dispar Bréb.

Staurastrum gracile Ralfs

Staurastrum hirsutum (Ehrenb.) Bréb.

Staurastrum margaritaceum Ehrenb.

Staurastrum meriani Reinsch

Staurastrum muticum Bréb.

Staurastrum orbiculare (Ehrenb.) Ralfs

Staurastrum orbiculare hibernicum

West and G. S. West

Staurastrum orbiculare ralfsii West

and G. S. West

Staurastrum paradoxum Meyen

Staurastrum paxilliferum G. S. West

Staurastrum polymorphum Bréb.

ANDERSEN AND WALKER

Staurastrum punctulatum Bréb.

Staurastrum saxonicum Bulnh.

Staurastrum teliferum Ralfs

Zygnemaceae.

Spirogyra arcta catenaeformis Kirchn.

Spirogyra crassa Kuetz.

Spirogyra dubia Kuetz.

Spirogyra lutetiana Petit

Spirogyra neglecta (Hass.) Kuetz.

Spirogyra varians (Hass.) Kuetz.

Zygnema Ag. sp.

Mesocarpaceae.

Mougeotia robusta (DeBary) Wittr.

Mougeotia scalaris Hass.

Mougeotia viridis (Kuetz.) Wittr.

Volvocaceae.

Volvox aureus Ehrenb.

Tetrasporaceae.

Tetraspora gelatinosa (Vauch.) Desv.

Dictyosphaerium pulchellum Wood

Pleurococcaceae.

Crucigenia rectangularis (A. Br.) Gay

Eremosphaera viridis DeBary

Nephrocytium naegelii Grun.

Oocystis solitaria Wittr.

Rhaphidium polymorphum falcatum

(Corda) Rabenh.

Scenedesmus antennatus Bréb.

Scenedesmus bijugatus (Turp.) Kuetz.

Scenedesmus obliquus (Turp.) Kuetz.

Scenedesmus quadricauda (Turp.)

Bréb.

Tetraedron minimum Reinsch

Tetraedron reticulatum (Reinsch)

Hansg.

Tetraedron trigonum (Naeg.) Hansg.

Urococcus insignis Hass.

Protococcaceae.

Characium ambiguum Hermann

Chlorococcum humicola (Naeg.)

Rabenh.

Characium subulatum A. Br.

Gloeocystis gigas (Kuetz.) Lagerh.

Gloeocystis vesciculosa Naeg.

Ophiocytium capitatum Wolle

AN ECOLOGICAL STUDY OF ALGAE 81

Hydrodictyaceae.

Coelastrum sphaericum Naeg.

Pediastzum angulosum (Ehrenb.)

Menegh.

Pediastrum boryanum

Menegh.

Pediastrum duplex clathratum A. Br.

Pediastrum tetras (Ehrenb.) Ralfs

(Turp.)

Ulotrichaceae

Microspora amoena (Kuetz.) Rabenh.

Microspora pachyderma (Wille)

Lagerh.

Microspora stagnorum (Kuetz.) Lagerh.

Hormiscia subtilis variabilis (Kuetz.)

Kirchn.

Chaetophoraceae.

Chaetophora cornu-damae (Roth) Ag.

Chaetophora elegans (Roth) Ag.

Gongrosira debaryana Rabenhorst

Stigeoclonium aestivale (Hazen) Col-

lins

Stigeoclonium glomeratum (Hazen)

Collins

Oedogoniaceae.

Bulbochaete Ag. sp.

Oedogonium Link (several species not

in fruit.)

Oedogonium capilliforme australe

Wittr.

Oedogonium cardiacum (Hass.) Kuetz.

Oedogonium crispum uruguayense

Magn. and Wille

Oedogonium fragile Wittr.

Oedogonium grande Kuetz.

Oedogonium varians Wittr. and Lund.

Oedogonium vaucherii (LeCl.) A. Br.

Coleochaetaceae.

Coleochaete orbicularis Pringsh.

Cladophoraceae.

Cladophora glomerata (L.) Kuetz.

Rhizoclonium hieroglyphicum (Ag.)

Kuetz.

Vaucheriaceae.

Vaucheria D.C. sp.

Characeae.

Chara Vaill. sp.

Chara contraria A. Br.

Chara evoluta Allen

Chara foetida rabenhorstii T. F. Allen

Chara fragilis Desv.

Helminthocladiaceae.

Batrachospermum vagum Ag.

PUBLICATIONS CONSULTED

1. Agardh, J. G. Species, genera, et ordines algarum. 3 vols.

1848-1880.

2. Allen, T. F. Characeae Americanae, illustrated and described. Parts I and II.

1879.

3. ——————. The Characeae of America, PartsI and II. 1880.

4, ———_———. Characeae. N. Y. State Museum Nat. Hist. 38th ann. report.

16, 15 Jan. 1885.

5. ——————.. The Characeae of America. Parts I and II, Fasc. 1, 2, and 3.

1888-1896.

6. Bessey, Charles E. Supplementary list of recently reported species. Webber’s

appendix to the catalogue of the flora of Nebraska, second edition.

tions from the Bot. Dept. of the Uni. of Nebr.

Contribu-

New series III:45-53, 1892.

7. Bessey, Charles E. and Webber, H. J. Report of the botanist on the grasses and

forage plants and the catalogue of plants.

Nebraska State Board of Agriculture.

Extracted from the report of the

1889-1890.

8. Botanical Survey of Nebraska, conducted by the Botanical Seminar, II, 1892, and

EIT, 1893;

16.

leis

18.

19.

20.

22:

23%

24.

25.

26.

27.

28.

29.

30.

31.

Rye

Sep

ANDERSEN AND WALKER

. Survey of Nebraska, conducted by the Botanical Seminar IV. Report on collec-

tions made in 1894-1895. 1896.

—————. Report on recent collections, Bot. Survey of Nebraska, pub. by

Bot. Sem. 1901.

. Birge, E. A. Absorption of the sun’s energy by lakes. Science, 38:702. 1913.

. Brannon, M. A. Factors influencing the flora of Devil’s Lake, North Dakota.

Int. Rev. d. ges. Hydr. u. Hydro. 4:291. 1911.

. Brown, H. E. Algal periodicity in ponds and streams. Bull. Tor. Bot. Club,

35:223-248. 1908.

. Brown, Wm. H. The plant life of Ellis, Great, Little, and Long’s Lakes in North

Carolina. Contr. from the U. S. Nat. Herb. 13:323-341. 1911.

. Chambers, C. O. The relation of algae to dissolved oxygen and carbon-dioxide,

with special reference to carbonates. 23rd ann. report Mo. Bot. Garden, pp.

171-207. 1912.

Collins, F. S. Green algae of North America. Tufts College Studies, vol. II,

No. 3, pp. 79-480. 18 pls. 1909.

Comére, Joseph. De l’action du milieu consedérée dans ses rapports avec la

distribution générale des algues d’eau douce. Mem. 25, Bull. Soc. Bot. France,

16:1-96. 1913.

Cook, M. C. British Desmids. 1887.

—————. British fresh-water algae exclusive of the Desmidiaceae and Dia-

tomaceae. 2 vols. 1882-1884.

Copeland, W. F. Periodicity in Spirogyra. Bot. Gaz. 47:9-25. 1909.

Danforth, C. H. Periodicity in Spirogyra with special reference to the work of

Benecke. Ann. Rep. Mo. Bot. Garden, 21:49-59. 1910.

Dangeard, P. A. Determination of the rays concerned in chlorophyll synthesis.

Bul. Soc. Bot. France 60:166-175. 1914.

DeToni, J. Bapt. Sylloge algarum I, 1889, V, 1907, IV, 1897-1905.

Engler, A. und Prantl, K. Die natiirlichen Pflanzenfamilien, Teil I. 1897-1900.

Fritsch, F. E. and Rich, Florence. Studies on the occurrence and reproduction

of British fresh-water algae in nature. Ann. Bot. 21:423-436. 1907.

Gomont, M. Maurice. Monographie des Oscillariees. (Nostocacees Homo-

cystees). 1893.

Halstedt, P. D. Classification and description of the American species of Chara-

ceae. Proc. Boston Soc. Nat. Hist. XX. 1879.

Hassall, Arthur Hill. A history of British fresh-water algae. 1845.

Hirn, K. Monographie und Iconographie der Oedogoniaceen. Acta Soc. Sci.

Fennicae 27. 1900.

Kiitzing, F. T. Phycologia generalis oder Anatomie, Physiologie, und System-

kunde der Tange. 1843.

Migula, W. Kryptogamen-Flora von Deutschland, Deutsch-Osterreich, und der

Schweiz’ im Anschluss an Thome’s Flora von Deutschland. Band II: 1 Teil,

1907, und 2 Teil, 1909.

Murray, Sir John and Hjort, Dr. Johan. The depths of the ocean. 1912.

Needham, James G. and Lloyd, J.T. Thelife of inland waters. 1916.

43.

33.

54.

SD:

AN ECOLOGICAL STUDY OF ALGAE 83

. Oltmanns, Dr. Friedrich. Morphologie und Biologie der Algen. 2 vols. 1905.

. Petit, Paul. Spirogyra des environs de Paris avec XII planches. 1880.

. Pool, R. J. A study of the vegetation of the sandhills of Nebraska. Minnesota

Botanical Studies, Vol. 4, Part III. 1914.

. Rabenhorst, Ludovico. Flora europaea algarum aquae dulcis et submarinae.

1864-1868.

. Robbins, W. W. A preliminary list of the algae of Colorado. Univ. of Colo.

Studies, 9:105. 1912.

. Robinson, Ch. Budd. The Chareae of North America. Bull. N. Y. Bot. Garden 4:

244. 1906.

. Saunders, D. and Woods, A. F. Flora of Nebraska, published by the Bot. Sem.

Parts land II. 1894.

. Schmidle, W. Einige Algen aus Denver, Colorado, U. S. Hedwigia 34:84-85.

Figs. 1, 2, 3a, and 3b. 1895.

. Smith, J. G. and Pound, R. Flora of the sandhill region of Sheridan and Cherry

counties and lists of plants collected on a journey through the sandhills in July and

August, 1892. Bot. Sur. Nebr. II. 1893.

Tilden, Josephine E. The Myxophyceae of North America and adjacent regions

including Central America, Greenland, Bermuda, the West Indies, and Hawaii.

Minnesota Algae TI. 1910.

. Transeau, Edgar N. The periodicity of algae in Illinois. Trans. Am. Mic. Soc.

erst. 1913:

. Webber, H. J. Fresh-water algae of the plains. Am. Nat. 23:1011-1013. 1889.

A second edition of Webber’s appendix to the catalogue of the Flora

of Nebraska. Contributions from the Bot. Dept. of the Univ. of Nebr. New

series III:1-44. 1892.

Appendix to the catalogue of the Flora of Nebraska. Contribution

from the Shaw School of Botany. No.9. Trans. Acad. of Sc. St. Louis, 6:1-47.

1892.

. West, G.S. A treatise on British fresh-water algae. 1904.

The algae of the Yan Yean reservoir, Victoria; a biological and

ecological study. Jour. Linn. Soc. 39:1-80. 1909.

Algae. Vol. I. 1916.

. West, W. and West, G. S. A monograph of the British Desmidiaceae. 4 vols.

1904-1912.

On the periodicity of the Phytoplankton of some British lakes.

Jour. Linn. Soc. 40:395-432. 1912.

Whipple, G. C. and Parker, H. N. On the amount of oxygen and carbonic acid

in natural waters and the effect of these gases upon the occurrence of micro-

scopic organisms. Trans. Am. Mic. Soc. 23:103-144. 1902.

Wolle, Rev. Francis. Desmids of the United States and list of American Pedia-

strums with 1,100 illustrations. 1884.

Fresh-water algae of the United States, (exclusive of the Diatoma-

ceae) complimental to desmids of the U. S., with 2,300 illustrations. 1887.

Fig.

ANDERSEN AND WALKER

EXPLANATION OF FIGURES

1. Map showing the lake region in Cherry County, Nebraska. Clear white areas

represent water, dotted areas wet meadows, and _|I|I II IL/I| swamps. (From

map by Dr. G. E. Condra).

2. A view taken from the top of a sandhill and showing at the front left a part of

Clear lake, at the upper left a part of Dewey lake and in the distance at the right

a narrow strip of White Water lake, also the “sandhills’’ and the meadows sur-

rounding the lakes. (Photo by F. H. Shoemaker).

3. Hackberry lake from the northeast shore.

4. Taking water photometer records among the rushes on Hackberry lake.

5. Submerged moss stems covered with Nostoc glomeratum—Hackberry lake.

(Photographed under water).

6. Section of Scirpus stem covered with Chaetophora cornu-damae and Nostoc

glomeratum. (Photographed under water).

7. Water photometer records on solio paper. Upper row exposures made in air.

Three lower lows exposures made under water.

8. Thermographs and anemometers in a blowout near the shore of Hackberry lake.

9. Thermographs and anemometers in the grassy meadow on the shore of Hack-

berry lake.

10. Lower end of water photometer showing water tight drum and window

covered with ray filter. (Photo by F. H. Shoemaker.)

11. Upper end of water photometer showing lever by means of which successive

areas of the photographic plate may be exposed to the window. (Photo by

F. H. Shoemaker.)

12. Above, under side of upper half of drum showing perforated, revolving disk

to which photographic plates are attached by means of two clips. (Photo by

F. H. Shoemaker.)

12. Below, upper half of water tight drum with lower half and tube removed.

(Photo by F. H. Shoemaker.)

13. Clear lake from a sandhill at its southwest end. At the right in the distance

a narrow strip of Willow lake. (X) the location of springs shown in fig. 14.

14. Pockets of spring water on the southwest shore of Clear lake.

15. Chart showing: A, temperature of air; B, temperature of soil 8 inches below

surface; and C, temperature of surface soil at a station in a blowout near the

shore of Hackberry lake. Also D, temperature of air in shade of a building on

the lake shore; E, wind velocity at the rim of the blowout; and F, wind velocity

at the bottom of the same blowout. Numbers at the left indicate, above tempera-

ture in Fahrenheit and below miles per hour of wind velocity.

16. Chart showing: A, temperature of air; B, temperature of soil eight inches

below the surface; and C, temperature of surface soil at a station in the grass near

the lake shore; also D, temperature of air in the shade of a building on the lake

shore; and E, wind velocity for the same period. Numbers at left indicate, above

temperature in Fahrenheit and below miles per hour of wind velocity at the

station.

TRANSACTIONS OF THE AMERICAN MICROSCOPICAL SOCIETY

PLATE X

June 24

June 25 | Tune 26

VOL. XXXIX

June 27

June 24 Tune 30

ANDERSEN AND WALKER

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TRANSACTIONS OF THE AMERICAN MICROSCOPICAL SOCIETY

VOL. XXXIX

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ANDERSEN AND WALKER

TRANSACTIONS OF THE AMERICAN MICROSCOPICAL SOCIETY, VOL. XXXIX

Jul 4 July 10 July s/t July 72 Jul 13 Jul 74 July IS July 76 Tuli 77 Jul 15 Jul 19 Jul 20 Juli 2/7 Jul 92 dul L Jul 24 Jul ag July 26 Tul 2

aes aa

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—

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VOL. XXXIX

cals Aug 4

July 37

Auda. 7

duly 29

July 2S | July26 July 2 Juty 23

WAG

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aUNUaUATE Neo T

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AN ECOLOGICAL STUDY OF ALGAE 85

Fig. 1/. Chart showing: A, air temperature; B, temperature of water at the bottom

of lake (3 feet below surface the first week, 414 feet below surface the second and

third week, and 5 feet below surface the fourth week); C, temperature of water at

surface of lake at a station located in a boat anchored in the lake; D, temperature

of air in shade of a building on the lake shore; E, wind velocity at margin of lake.

Numbers at left indicate above temperature in Fahrenheit and below miles per

hour of wind velocity. July 24-27 anemometer readings were not taken. It wasa

period of very low wind and is indicated approximately by the dotted line.

The University of Nebraska,

Lincoln, Nebr.

DEPARTMENT OF NOTES AND REVIEWS

It is the purpose, in this department, to present from time to time brief original

notes, both of methods of work and of results, by members of the Society. All mem-

bers are invited to submit such items. In addition to these there will be given a few

brief abstracts of recent work of more general interest to students and teachers. There

will be no attempt to make these abstracts exhaustive. They will illustrate progress

without attempting to define it, and will thus give to the teacher current illustrations,

and to the isolated student suggestions of suitable fields of investigation.—[Editor.]

LEECHES CONSIDERED AS OLIGOCHAETA MODIFIED FOR A

PREDATORY LIFE

Michaelsen (Mitt. Zool. Mus. XXXVI, Hamburg, 1919) was led

to a study of the relationships between these two groups of animals,

by noticing a figure in a recent paper on Sudanese Hirudinea. The

figure represented an organ that was interpreted by the author, as

a diverticulum of the alimentary tract of the leech, opening to the

exterior on the mid-dorsal surface of the 13th somite. Similar organs

have been described in certain leeches from Sumatra, in which they

are paired, and the external pores are ventrally situated. The

figured organ strongly resembles the spermathecae of certain oligo-

chaete species in the families of Enchytraeidae and Lumbriculidae,

in which the spermathecae communicate internally with the alimen-

tary tract. Similar relations have also been found in certain species

of other families of Oligochaeta.

As a result of his studies, Michaelsen has reached the conclusion

that the Hirudinea are, in reality, Lumbriculidae which have under-

gone special modifications in adaptation to a predatory mode of life.

He believes that such a conclusion receives much support from a

careful comparison of the structure of two intermediate types of

worms: the Branchiobdellidae, and Acanthobdella peledina Grube.

The former are parasitic in the gill chambers and on parts of the

surface of crawfishes, and, as their name indicates, were formerly

included with leeches; but recently their closer relationship with

the Oligochaeta is generally admitted. Acanthobdella peledina is a

peculiar leech-like parasite of certain fishes of the genus Salmo, in

northeastern Europe, and in western Siberia. On the ventral surface

NOTES AND REVIEWS 87

of several anterior somites, are paired bundles of setae, and the charac-

ters of the reproductive organs and of the body cavity are also nearer

to those of the Oligochaeta than to those of the leeches. Michaelsen

concludes that, although there is some justification for including

these two groups in the family Lumbriculidae, it is nevertheless

preferable to recognize them as two distinct families of Oligochaeta,

Branchiobdellidae and Acanthobdellidae closely related to the

Lumbriculidae. After making this disposition of these two groups,

the author makes a comparison of the various structural characters

of the Hirudinea and Oligochaeta.

Attention is called to the fact that there is a wide range of varia-

tion among different representatives of the Oligochaeta, and that

most of the characters which one is accustomed to think of as typical

of the Oligochaeta are not present in all members of the group,

though they may be in a majority of the better known ones. It is also

shown that many of the characters of Hirudinea which one is likely

to assume as distinguishing them from Oligochaeta, may be found

present in certain members of the latter group. Absence of setae

occurs in a genus of the oligochaete family Enchytraeidae, as well

as in Branchiobdellidae, and they are greatly reduced in numbers

and size in various other representatives. As previously mentioned,

four pairs of well developed setae are present on each of several anter-

ior somites in Acanthobdella peledina which has previously, without

question, been assumed to belong to the Hirudinea. The shortened

body and thickened body wall of the leeches, with a correlated

reduction of the body cavity, are already forecast in Chaetogaster

and in certain species of Lumbriculidae, to say nothing of the Branch-

iobdellidae and Acanthobdella. They are natural accompaniments of

a change of food, and assumption of a predatory mode of life.

There is great variation in the structure of the nephridia among

the Oligochaeta, and absence of ciliated nephrostomes and of cilia

in the excretory part of the ducts is found in species of diverse

groups. ‘The ventro-median position of the pores of the efferent

ducts of the reproductive organs of leeches has a counterpart in

certain species of Lumbriculidae and of the earthworm subfamily

Eudrilinae.

The most significant character which distinguishes the Hirudinea,

in general, from the Oligochaeta, is the position of the spermaries

88 AMERICAN MICROSCOPICAL SOCIETY

in somites posterior to the one which contains the ovaries. This

relative position of the two kinds of gonads is the opposite of that

normally found in Oligochaeta, and in the connecting forms, Branch-

iobdellidae and Acanthobdella. To account for this reversal of rela-

tions, the author refers to instances where Oligochaeta are found with

a considerable number of consecutive somites containing gonads;

and also to papers by different writers, in which gonads of certain

oligochaete species have been shown to produce one kind of germ

cells at one time, and at other times to produce those of the opposite

kind. From individuals with series of gonads of this type, he thinks

it not improbable that there may have been derived descendants in

which the relative position of the gonads of the two sexes is in the

reverse order from that of the ancestors. For details of structure

and references to the literature involved in these comparisons, the

original paper must be consulted.

The author thinks it desirable to modify the outlines of classifica-

tion, to fit these new views of relationship. He proposes a class

Clitellata which is co-ordinate with the class Chaetopoda, and with

three other classes which contain marine forms and are not involved.

The class Clitellata includes two orders, Oligochaeta and Hirudinea;

distinguished chiefly by the differences in the degree of development

of the body cavity, and the relative order of the gonads. The class

Chaetopoda includes two orders, Protochaeta and Polychaeta.

FRANK SMITH

Department of Zoology,

Univ. of Illinois

PROCEEDINGS OF THE AMERICAN MICROSCOP-

ICAL SOCIETY

MINUTES OF THE ST. Louis MEETING

The thirty-eighth annual meeting of the American Microscopical Society was held

in affiliation with the A.A.A.S. at St. Louis, Mo., Dec. 31, 1919.

In the absence of President Griffin, Vice-President Whelpley acted as chairman.

The report of the Treasurer for the years 1918 and 1919 was accepted and referred

to an auditing committee composed of Professors H. B. Ward and H. J. VanCleave.

The report of the Custodian was accepted, ordered printed, and referred to an

auditing committee composed of Messrs. Edw. Pennock and Edw. P. Dolbey.

A vote of appreciation was extended to Professor T. W. Galloway, the retiring

Secretary, for a most valuable service rendered to the Society during the past ten years.

The meeting voted approval of the action of the Executive Committee in appoint-

ing Mr. Wm. F. Henderson as Treasurer, and Professor Paul S. Welch as Secretary

at dates in advance of the regular annual business meeting.

The following officers were duly nominated and elected for the constitutional

periods: President, Professor T. W. Galloway, New York; First Vice-President,

Chancey Juday, University of Wisconsin; Second Vice-President, Professor A. D.

MacGillivray, University of Illinois; Secretary, Professor Paul S. Welch, University of

Michigan; Treasurer, Mr. Wm. F. Henderson, James Millikin University.

Professor Frank Smith of the University of Illinois, Professor J. E. Ackert of the

Kansas State Agricultural College, and Dr. B. H. Ransom of the Bureau of Animal

Industry were chosen as the elective members of the Executive Committee for 1920.

Minutes of the last annual meeting were approved as printed.

Adjourned.

Pau S. WELCH,

Secretary

CUSTODIAN’S REPORT FOR THE YEARS 1918 AND 1919

SPENCER—TOLLES FUND

Amount reported December 19075) 0) cc 60s os Fok aaa eaaee 533197

jmme.30; 1918 Dividendsreee ten. cerns Meee coer Gam eee 159.93

Dees Sl; 1918 Dividendseraree eres ee oa ticks cee eae 164.73

juners0: 1919) Divadendstemreaner se warts tee en ater heer eiays 226.24

Weerst 1919) MividendSeardyan eee ee ees ea isicheasciee 176.46) 12736

6058 .93

less Grant: Non Gas ce Mee eed enc cmaraorte 100 .00

Net amount invested ac cone ey a cee ea betes 8 5958 .93

Increase during last two years $627. 36.

90 AMERICAN MICROSCOPICAL SOCIETY

TOTALS

Alliicontributions 44°) Meee ee Ce eee eee 800.27

All Salesiofiransactionsenian aoe eee eee 878.38

AllcLigememberships sc0520 Sn ene cstaive mantels wants 300.00

AllinterestiaaDividendss sare eee eee eee eee 4270.28

LESS

(AU Granta ene ste nine ee one tn Merten) 7 EOL aan 250.00

ALND uestior Ibitesmembersitsn5 ee ee oe eee 40.00 290.00 5958.93

Life members: (Robert Brown, dec’d.); J. Stanford Brown: Seth Bunker Capp;

Harry B. Duncanson; A. H. Elliott; John Hatly.

Contributors of $50 and over; John Aspinwall; Iron City Microscopical Society,

Magnus Pflaum; Troy Scientific Society.

(Signed) M. Priaum,

Custodian.

Philadelphia, Pa., January 10, 1920.

The undersigned having examined the foregoing report certify that we find the

amount invested as shown therein $5958 .93—correct, as shown by the Pass-Book of

the Keystone State B. & L. Association, the same being brought down to the 2nd

instant inclusive.

(Signed) EDWARD PENNOCK,

Epw. P. DoLBey,

Auditing Committee.

ANNUAL REPORT OF THE TREASURER OF THE AMERICAN

MICROSCOPICAL SOCIETY

DECEMBER 22, 1917 TO DECEMBER 21, 1918

RECEIPTS

BalancetonyhandtromauditofA Olen ee eee ae eee $770.57

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FonwlO1gie ten fod tonnes ba vena Sob LoniNe At ae tia oe 226.00

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PROCEEDINGS OF AMERICAN MICROSCOPICAL SOCIETY 91

EXPENDITURES

Publishing sPransactions., 4.504 oo cicli hese iehase chccepeiete aloe eran dies ayaa ated ao see $877.82

Volume S6):num ber Aes 5s ticeieietss x dace od aes se bagels $293.82

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PL atesty se ee he ere ee ate eho seats clcratsaehe orotate 48.73

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TRL EASULOT pebehe ere so here lec eerste shemaene hae sicsree el meatah eters 12.10

Miscellaneous: itemise wastes Ascrsceiotetiticiarsier orcieruia a ai elaktye sia le laceiaceiatel etwas & 11.85

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Balance’on hand sane eotcrc Sern etre Hae ee MUNA te et Ors sare ovarian Vans eiahalie level 896.48

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Respectfully submitted,

H. J. VAN CLEAVE, Treasurer.

REPORT OF OUTGOING TREASURER FOR THE PERIOD OF DEC. 22, 1918

TO FEB. 28, 1919

RECEIPTS

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BACK AV ONITEE Sener ymmon tes eters lee Mie tua) 5 sat analasakar Spt $10.00

BGOrp LOL Ope cerry Meek ee tort cele SU al a Ee 60.00

FOr 1920 eas enh eure Solara e ay Val elcce ed ler ate shale lets 2.00

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92 AMERICAN MICROSCOPICAL SOCIETY

EXPENDITURES

Publication of Transactions Volume 37, number 4..................... $267.14

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TOTAL MSXPENDITURES ry teehee ie eet ee re $ 319.86

Balance transterreditoynewslneasurer-e pe) eee ee eee eee eee 730.02

$1,049.88

Respectfully submitted,

H. J. Van CLeave, Treasurer.

REPORT OF AUDITING COMMITTEE ON TREASURER’S

ACCOUNTS FROM DECEMBER 22, 1917 TO FEBRUARY 28, 1919

This is to certify that we have this day examined the accounts of H. J. Van Cleave,

Treasurer, and have checked the vouchers against payments; we find the accounts

correct and have verified the bank balance of $730.02 to be transmitted to the incom-

ing treasurer.

Henry B. Warp

FRANK SMITH

Audit Committee.

_ ANNUAL REPORT OF THE TREASURER

OF THE AMERICAN MICROSCOPICAL SOCIETY

March 1, 1919 to December 24, 1919

RECEIPTS

Balancemeceived:irom) former treasurer. 4.6 eee eens ee oe eee ae $ 730.02

Dues)received tromVolume 3/7 orbefore.. 0. cee os sane aeeeiee 36.00

Dues received trom Wolumesonee ne eee eo oe cee eee EERE 108.00

IDwesireceiveduronlavolume 39s sae os tei ee ic eae asian aaa ee 292.10

Duesireceivedurom Volume 40S.) nlc aoe cy Bee ee eee 2.00

NTE ALIONUEES hc AA le Tee era ey Cae oreo ct ce See ae AL ae oe 66.00

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pubscriptions dor Welume 38.42.0220 42> bobs ahs as eases oe Oe eR Oer 51.80

Subscriptionsitor Volume 39 7i4. cctod i oa) 8o.cs Sica cn here ere ee 34.00

Sales of Transactions, duplicates and back numbers ...........--..--+5 183.00

Donation from T. B. Magath for aid in publishing his paper..........-.. 50.00

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PSTN Ys bg (ac Gan cae ALG tenn hn) ee eae MI AL RMS eA Pe ae Oe tae 4.50

PROCEEDINGS OF THE AMERICAN MICROSCOPICAL SOCIETY 93

EXPENDITURES

Printing Transactions; Volume 38) NGIE yet ie eeoke o ak raters ttt $201.03

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Office. expensestoielreasurenkn este 5 cs acta leith eo cree ee 19.70

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Balancetonle bene ager ioe etal ra Cea ss. g atarer tise ti sacha aioraineaes wees loons 480.04

MNO TATH ORIN DITS Meee cn ier alree slayer edie vit Ns ean yt Cie pee $1,628.42

W. F. HENDERSON, Treasurer

This is to certify that we have examined the books and vouchers of the Treasurer

and find them to be in good condition and to give a correct record of moneys received

and expended as indicated.

Henry B. Warp

H. J. VAN CLEAVE

Auditing Committee

March 24, 1920.

TABLE OF CONTENTS

For VoLUME XXXIX, Number 2, April, 1920

Modern Dark-field Microscopy and the History of its Development, by Simon

IS ESTEIAE (CORZAS oie Geetehcene OS euch ONE oh SIE NC OCP OU ERCP R EOS Ue

A New Bladder Fluke from the Frog, with Plate XIII, by John E. Guberlet

Labeling Illustrations, with Plates XIV to XVII, by Z. P. Metcalf

Notes and Reviews: Position of Micropterygidae; Micropterygidae; Filariasis

in U. S.; Polyembryony and Sex; Origin and Significance of Metamorphosis

142

149

163

TRANSACTIONS

American Microscopical Society

(Published in Quarterly Instalments)

Vol. XXXIX APRIL, 1920 No. 2

MODERN DARK-FIELD MICROSCOPY

AND THE

HISTORY OF ITS DEVELOPMENT

Smmon Henry GAGE

Professor of Histology and Embryology, Emeritus Cornell University

INTRODUCTION

In most work with the microscope the entire field of view is

lighted and the objects to be studied appear as colored pictures or as

shadows—in extreme cases, as silouhettes—on a white ground. As

the field is always light, this has come to be known as Bright-Field

Microscopy (Fig. 1).

Fig. 1 Fig. 2

Bright- and dark-field photo-micrographs of the same objects (starch grains).

In contrast with this is Dark-Field Microscopy in which the field

is dark, and the objects appear as if they themselves emitted the

light by which they are seen (Fig. 2).

96 SIMON HENRY GAGE

The study of objects in a bright-field probably comprises 95%

of all microscopic work, and is almost universally applicable. On the

other hand dark-field microscopy has only limited applicability, and

yet from the increased visibility given to many objects it is coming to

be appreciated more and more.

Definition.—In its comprehensive sense, Dark-Field Microscopy

is the study of objects by the light which the objects themselves

turn into the microscope, and none of the light from any outside

source passes directly into the microscope as with bright-field

microscopy.

There are two principal cases: (A) The objects which are truly

self-luminous like phosphorescent animals and plants; burning or

incandescent objects, and fluorescent objects. (B) The objects

which emit no light themselves, but which deflect the light reaching

them from some outside source into the microscope.

These two groups are well represented in Astronomy. If one

looks into the sky on a cloudless night, the fixed stars show by the

light which they themselves emit, but the moon and the planets

appear by the light from the sun which they reflect to the earth,

the sun itself being wholly invisible at the time. As there is rela-

tively very little light coming from the intervening space between

the stars and planets, all appear to be self-luminous objects in a dark

field. This reference to the sky at night will serve to bring out two

other points with great clearness: (1) The enhanced visibility.

Everybody knows that there are as many stars in the sky in the day-

time as at night, but they are blotted out, so to speak, by the flood

of direct light from the sun in the daytime, while at night when these

direct rays are absent and no light comes from the back-ground the

stars and the planets show again by the relatively feeble light which

they send to the earth.

(2) The other point is that in dark-field microscopy the objects

must be scattered, not covering the whole field (Fig. 2). If there

were no intervening empty space the whole face of the sky would

look bright

It will b’e seen from this that ordinary sections or other objects

so large that they fill the whole field of the microscope cannot be

studied advantageously by the dark-field method, for they would

make the whole field bright. But for the liquids of the body, blood,

lymph, synovial, and serous fluids, fluid from the cavities of the

MODERN DARK-FIELD MICROSCOPY 97

nervous system, saliva, and all other mucous fluids, and isolated

tissue elements where the solid or semi-solid substances are distrib-

uted in a liquid, the appearances given by this method are a revela-

tion as was pointed out by Wenham and Edmunds and many others

over fifty years ago. No less is the revelation coming from the study

of bacteria, protozoa and other micro-organisms in the dark field.

DARK-FIELD AND ULTRA-MICROSCOPY

In both of these the objects seem to be self-luminous in a dark

field, and no light reaches the eye directly from an outside source,

but only as sent to the eye from the objects under observation.

The terms simply represent two steps, and merge into each other.

Dark-Field Microscopy deals with relatively large objects, 0.24

or more in diameter, that is, those which come within the resolving

power of the microscope.

Ultra-Microscopy deals with objects so small that they do not

show as objects with details, but one infers their presence by the

points of light which they turn into the microscope. This can be

made clear by an easily tried-naked-eye observation. Suppose one

is in a dark room, and a minute beam of brilliant light like sunlight

or arc light is directed into the room. Unless one is in the path of

this beam of light it will remain invisible, but if there are vapor or dust

particles present they will deflect some of the light toward the eye

and will appear as shining points. The character of the particles

cannot be made out, but the points of light they reflect indicate their

presence. As Tyndall used this method in determining whether a

room was free from dust in his experiments in spontaneous generation,

the appearance of the shining dust particles is sometimes called the

“Tyndall effect.”

The two forms are said to merge, because in studying objects like

saliva, etc., with the microscope designed especially for dark-field

work, some of the objects seen will show details, but some are so

small that they show simple as points of light usually in the form of

so-called diffraction discs. The larger objects in the saliva come in

the province of dark-field microscopy, and the smallest ones, of

ultra-microscopy, and in this case the instrument used might with

equal propriety be called a dark-field or an ultra-microscope.

The great purpose of the dark-field microscope is to render

minute objects or details of large objects plainer or actually visible

98 SIMON HENRY GAGE

from the advantages offered by the contrast given between the

brightly lighted objects and the dark background. For example,

with the homogeneous immersion objective the study of fresh blood

with the ordinary bright-field method enables one to see the red

corpuscles with satisfaction, but the leucocytes are not easily found

and the blood-dust (chylomicrons) and the fibrin filaments are not

seen at all or very faintly. With the same microscope using the

dark-field illumination the leucocytes are truly white cells, and the

blood-dust is one of the striking features of the preparation, and the

fibrin filaments seem like a delicate cobweb.

In this connection, perhaps a few words should be added on the

terms Resolution and Visibility. Both came over from the ancient

science of astronomy, and are properly used only when restricted as

in astronomy.

By resolution is meant the seeing of two things as two, not

blended. For example if two stars are close together they are re-

solved if they appear as two. When the telescope was invented it

was found that many stars that appeared single were really two stars

close together. Iftwo lines are placed close together they appear as two

to the naked eye when close up, but as one moves away the lines

seem to fuse and make one. Visibility refers only to the possibility

of seeing a thing. In the above examples the twin stars were visible

to the naked eye but not resolved into two, and likewise the lines

were long visible after they could be seen as two lines. Now the

purpose of the ultra-microscope is solely to increase the visibility

of small particles without reference to their details of structure.

Dark-field microscopy, on the other hand, while it gives greatly

increased visibility, also gives resolution of details.

As with bright-field microscopy the resolution of details of struc-

ture depends directly upon the numerical aperture (NA) of the

objective, and the brightness upon the square of the aperture (NA?).

METHOD OF DARK-FIELD MICROSCOPY

In this article the ultramicroscope and the study of self-luminous

objects will not be further considered, but the discussion will be

limited to objects which must be lighted by some outside source.

There are two principal cases: (1) objects which are lighted from

above the stage of the microscope or by so-called direct light (Fig. 3)

MODERN DARK-FIELD MICROSCOPY 99

and, (2) objects which are lighted from below the stage, or by trans-

mitted light (Fig. 4).

Fig. 3. Light from above the stage. (From The Microscope)

In both cases the light from the source is at such an angle that

none of it can enter the objective directly but only as it is deflected

or “radiated” by the objects in the microscopic field.

“| Objective

Stage

SAMARAS

Fig. 4. Light from below the stage.

When the light upon the object is from above the stage the back-

ground must be non-reflecting. If the background were white

there would be a kind of bright-field, not dark-field microscopy.

The black-background is secured either by placing the object

directly upon some black velvet or other non-reflecting surface, or

on a glass slide which in turn is placed upon black velvet, etc., or on

a dark well. The simplest way to produce a dark-well is to turn the

condenser aside and place a piece of black velvet over the foot of the

microscope. Or the condenser can be lowered well and the velvet

put over the top of the condenser.

100 SIMON HENRY GAGE

Diffuse daylight from a window, or more satisfactorily, artificial

light directed by a mirror or lens (bull’s eye),is directed obliquely

down upon the preparation (Fig. 3). Exactly the same preparation

will answer for light from below the stage. In this case the condenser

is turned out of the way, and some black-velvet put over the foot of

the microscope to cut out stray light.

For a good naked eye demonstration showing the increased

visibility due to the dark-field, some cotton may be placed on a piece

of black velvet, and a similar tuft of cotton on a white card.

For the special methods of lighting microscopic objects from

above the stage, see in the historical summary at the end of this

paper.

Dark-Field Microscopy by Transmitted Light—To make objects

appear self-luminous in a dark field when illuminated by beams of

light from below the stage, two things are necessary:

(1) The objects must be able to deflect in some way the light

impinging upon them into the microscope.

(2) None of the light from the source must be allowed to pass

directly into the microscope. These conditions are met when (a)

the objects to be studied are of different refractive index from the

medium in which they are mounted, and (b) when the transmitted

light thrown upon the object is at such an angle that it falls wholly

outside the aperture of the objective (Fig. 4-7).

The objects deflect the light into the microscope

(1) By Reflection

(2) By Refraction

(3) By Diffraction

Any one of these will suffice, but any two or all of the ways may be

combined in any given case.

For low powers where the aperture of the microscope objective

is relatively small it is comparatively easy to make the transmitted

beam of so great an angle that none of it can pass directly into the

microscope. A simple experiment will show this: A 16 mm. or lower

objective is used, the substage condenser is turned aside and on the

stage is placed a clean slide with a little starch, flour, or other white

powder dusted upon it. If now the mirror is turned to throw the

light directly up into the microscope the field will be bright and the

objects relatively dark, but if the mirror is turned at an angle suf-

MODERN DARK-FIELD MICROSCOPY 101

ficient to throw the whole beam at a greater angle than the aperture

of the objective will receive, the field wiil become dark and the

starch or flour grains will stand out as if shining by their own light.

If some black velvet is placed on the foot of the microscope so no

light can be reflected upward into the microscope from the foot or

the table, the field will be darker. This experiment succeeds by

either natural or artificial light. If some water containing para-

mecium and other micro-organisms is put on the slide and put under

the microscope, the organisms will appear bright and seem to be

swimming in black ink.

It is readily seen that with the method just discussed the light

is all from one side (Fig. 4). To light the objects from all sides,

that is, with a ring of light, the simplest method, and the method

utilized in all modern dark-field microscopy, is to use a hollow cone

of light, the rays in the shell of light all being at so great an angle

with the optic axis of the objective that none of them can enter the

microscope directly (Fig. 4-7).

With Refracting Condensers. With the condensers of the achro-

matic or chromatic type used for bright-field microscopy a solid

cone of rays is used. To get the dark-field effect the objects to be

studied must be lighted only by the rays at so great an angle that

they cannot enter the objective directly. This requires that the

condenser shall have a considerably greater aperture than the objec-

tive. The ordinary method of making the hollow cone is to insert a

dark stop—central stop—to block or shut off the central part of the

solid cone of light. The object is then illuminated with a ring of

light of an aperture greater than that of the objective (Fig. 6). Some

of this light is turned by the objects into the microscope. As only

a relatively small amount of the light is deflected by the objects

into the microscope, it is evident that there must be a great deal of

light to start with or there will not be enough passing from the

object to the microscope to make it properly visible. The question

also naturally arises how one is to determine the size of the central

stop to be used with any given condenser and objective.

This is easily determined as follows: The field is lighted well

as for ordinary bright-field observation and some object is got in

focus. Then the object is removed and the iris diaphragm of the

condenser opened to the fullest éxtent. If one then removes the

ocular and looks down the tube of the microscope and slowly closes

102 SIMON HENRY GAGE

the iris, when the full aperture of the objective is reached, that is,

when the back lens of the objective is just filled with light, the opening

in the iris represents the size of the central stop to use to cut out all

the light which would pass into the microscope from the condenser;

all the ring of light outside of this is of too great an angle for the

aperture of the objective. One can measure the size of the opening

in the iris with dividers and then prepare a central stop diaphragm.

Fig. 5. Ordinary condenser with sectional and face views of the central stop (D).

(From The Microscope)

A visiting card is good for this. It should be blackened with India

ink. To be on the safe side it is wise to make the central stop a little

greater in diameter than the iris opening (Fig. 5).

If now the microscope is lighted as brilliantly as possible, and

then the iris opened to its full extent and the blackened central stop

is put in the ring under the condenser, and a slide used with starch or

flour on it, the flour or starch particles will be lighted with the ring

of light, and they will deflect enough into the objective to make the

objects appear bright as if shining by their own light, the background

remaining dark. If the field looks gray or light instead of black it

is because the central stop is too small or not centered or the particles

used for objects are too numerous, not leaving enough blank space.

MODERN DARK-FIELD MICROSCOPY 103

One can determine what is at fault thus: The ocular is removed.

If the central stop is too small the back lens of the objective will show

a ring of light around the outside. If the central stop is not centered

there will be a meniscus of light on one side. If the objects are too

numerous the whole field will be bright. To verify these statements

one can use a specimen with flour or starch all over the slide. It will

look dazzlingly light, with the ocular in place and the back-lens will

be very bright when the ocular is removed.

For the meniscus of light when the central stop is decentered,

purposely pull the ring holding the stop slightly to one side and the

meniscus will appear in the back lens. To show the ring of light due

to a too small size of the stop, the easiest way is to use a higher

objective, say one of 3 or 4 mm. in place of the 16 mm. objective.

While it is necessary to eliminate all the light which could enter the

objective directly, the thicker the ring of light which remains to

illuminate the objects the more brilliantly self-luminous will they

appear, therefore one uses only the stop necessary for a given objec-

tive. If one makes central stops for the different objectives as

described above it will be greatly =mphasized that the objectives

differ in aperture, in general the higher the power the greater the

aperture, and consequently the larger must be the central stop, and

the thinner the ring of light left to illuminate the object. As one

needs more light for high powers instead of less than for low powers,

the deficiency of light caused by the large central stop must be made

good by using a more brilliant source of light for the high powers.

Reflecting Condensers. As was first pointed out by Wenham,

1850-1856, refracting condensers are not so well adapted for obtaining

the best ring of light for dark-field work as a reflecting condenser, on

account of the difficulty in getting rid of the spherical and chromatic

aberration in the refracted bundles of such great aperture. He first

(1850) used a silvered paraboloid and later (1856) one of solid glass

as is now used. Within the last 10-15 years there has also been

worked out reflecting condensers on the cardioid principle. The

purpose of all forms is to give a ring of light which shall be of great

aperture, and be as free as possible from chromatic and spherical

aberration, and hence will form a sharp focus of the hollow cone

upon the level where the objects are situated.

104 SIMON HENRY GAGE

“Glass. Slide

S AM

SSSSET TST

LLLEET TT LLL LLL LLL rd

Centrai Stop

Fig. 6. Bright-field condenser with central stop to give dark-field illumination.

This is a sectional view showing the hollow cone of light focusing on the object

and then continuing wholly outside the aperture of the objective.

The light deflected by the object into the objective is represented by broken lines.

The glass slide is in homogeneous contact with the top of the condenser, and the

medium beyond the object is represented as homogeneous with glass.

Centre! Stop

Fig. 7. Paraboloid condenser for dark-field illumination.

Axis—The principal optic axis of the microscope.

Central Stop—The opaque stop to cut out all light that would be at an aperture

less than 1.00 NA.

MODERN DARK-FIELD MICROSCOPY 105

Cover Glass—The cover for the object. For dry objectives it must conform to

the objective, and with homogeneous objectives it must be less than their working

distance in thickness.

C r-—Face view of the top of the paraboloid showing the centering ring, the

spot of white ink in the middle and the grains of starch for centering and focusing

high powers.

Glass Slide—The slip of glass on which the object is mounted. It is connected

with the top of the paraboloid by homogeneous liquid, and must be of a thickness

to permit the focusing of the hollow cone of light upon the object.

Hi, Hi—Homogeneous liquid between the cover-glass and the objective and

between the top of the condenser and the slide.

NA 1.00 to 1.40—The numerical aperture of the hollow cone of light focused on

the object by the paraboloid. As indicated on the left this is represented by a glass

angle of 41 to 67 degrees.

41° 67°—The limits of the angle of the rays in glass. Objective—The front lens

of the objective. The light rays deflected by the object are indicated by white lines

below and through the lens, then by broken, black lines above the front lens of the

objective. Water—The mounting medium for the objects.

In this diagram the course of the rays from the paraboloid are indicated as if the

objects were mounted in homogeneous liquid and that the rays passed beyond the

focus into a medium homogeneous with glass.

TABLE SHOWING THE Maxtuum ANGLE IN GLASS, AND THE CORRESPONDING

NUMERICAL APERTURE OF THE LIGHT WHICH CAN Pass INTO MEDIA OF

DIFFERENT REFRACTIVE INDEX ABOVE THE CONDENSER (Fic. 8-11)

Angle in Numerical Index of

Glass Aperture Refraction

HewAirover the condenser. 22.402 25...-4. 41° 1.00 1.00

DAV ALGI Nea fey. caiman snippet Gi Sen gtaeet 61° 133 1.33

4° Homogeneous liquid. :.).......0..2..- 90° 1.52 1.52

In the reflecting as in the refracting condensers the central part

of the light beam from the source is blocked out by a central stop

and only a ring of light enters the condenser.

Immersion connection of condenser and glass slide bearing the

specimen.—While the purpose of the reflecting condenser is to pro-

duce a very oblique beam of light for illuminating the objects, it is

seen at once that the laws of refraction will prevent the light from

passing from the condenser to the object unless the glass slide bearing

the object is in immersion contact with the top of the condenser.

106 SIMON HENRY GAGE

That is, for air (index 1.00) above the condenser, the rays in

glass at 41°, NA 1.00 and less can pass from the condenser into

the air and expand into a hemisphere of light in it (Fig. 8). Rays

above 41° are totally reflected back into the condenser.

For water (index 1.33) above the condenser, rays in the glass at

61°, NA 1.33, and less can pass into the overlying water and

make a complete hemisphere of light in it (Fig. 9). Rays above 61°

are totally reflected back into the condenser.

For glycerin (index 1.47) above the condenser, rays in the glass

at 75° 151, NA 1.47 and less can pass from the glass into the over-

lying glycerin and form a hemisphere of light in it (Fig. 10). All

rays at a greater angle are reflected back into the condenser.

For homogeneous liquid (index 1.52) over the condenser, there is

no limit to the angle of light that can pass from the condenser to it

(Fig. 11).

Immersion Liquid between Condenser and Glass Slide. While

water or glycerin answers fairly well it is recommended that homo-

geneous liquid be used in all cases. At first glance this would seem

unnecessary for, as just stated the aperture of the light is limited by

the medium of least refractive index between the condenser and the

object. Thus objects mounted in watery fluids, and especially those

mounted in air would seem to have the illuminating ray that could

reach them limited by an aperture of 1.33 in one case and of 1.00 in

the other (glass angles of 61° and 41°). This would be true if the

objects were suspended in the water or in the air, but many of the

particles are not suspended but rest on the glass slide, that is are in

so-called optical contact with the slide. This being true, the angle

of the light which can pass from the condenser to them depends upon

their own refractive index, and not upon that of the mounting

medium (air or water). This explains also why objects not in optical

contact with the slide are rendered more visible by the homogeneous

immersion contact of slide and condenser for the scattered light

from the particles in optical contact helps to light up particles not

in contact.

Another consideration also favors the use of the homogeneous

immersion contact of slide and condenser, even for objects mounted

in air. Physicists have found (see Wood) that beyond the critical

angle, while all light is turned back into the denser medium, it does

nevertheless pass one or more wave lengths into the rarer medium to

MODERN DARK-FIELD MICROSCOPY 107

find, so to speak, an easier place to turn around in. If now any

object is near enough the slide to fall into this turning distance of the

totally reflected light it may be said to be in optical contact, and the

light which meets it will pass into it instead of being totally reflected.

Fig. 8, 9, 10, 11. Diagrams showing the angle and numerical aperture of the

light in glass to fill the entire hemisphere above, with overlying media of air, water,

glycerin, or homogeneous immersion liquid.

As shown by the diagrams, the NA of the light in each case must equal the index

of refraction (Ir) of the overlying medium to fill the overlying hemisphere with light.

If the light is at a greater than the critical angle it is reflected back into the condenser.

Such light is represented by black in 8, 9, 10. With homogeneous liquid (Hom. imr)

above the condenser there is no critical angle.

It should be said in passing that the medium of least refractive index in the path

of the light beam from the condenser determines the critical angle at which the light

is wholly reflected, and hence determines the maximum angle of the illuminating

pencil that can light the object, but this does not apply if the object is in optical

contact with the glass (see below).

One can make a very convincing experiment to show the impor-

tance of remembering that some of the objects are in optical contact

with the glass slide and hence may utilize light which could not pass

108 SIMON HENRY GAGE

into the surrounding medium. If the upper face of the dark-field

condenser is cleaned as perfectly as possible, and then lighted well,

one can see no light emerging from the top except where the centering

ring is situated or where there are some accidental scratches. If one

dusts some starch, flour or other white powder on the clean surface,

the particles which make optical contact with the glass will glow as

if self-luminous. In case one wishes further evidence, the end of the

condenser should be carefully cleaned, and a glass slide of the proper

thickness connected with it by means of homogeneous liquid, then

some flour or starch can be dusted on the slide and it will glow as did

the particles on the top of the condenser. These demonstrations

show well with the naked eye and with objectives up to 8 mm. (Fig. 7, Cr.)

Aperture of the Ring of Light in the Condenser. As the angle of

the light illuminating the objects must be greater than can enter the

objective employed it follows that the central part of the illuminating

beam must be blocked out up to or beyond the aperture of the objec-

tive to be used. The greatest aperture rays possibly attainable

depends upon the opticians ability to so design and construct the

condenser that it will bring the remaining shell or ring of light to a

focus. For those designed to be used with all powers, the aperture

of this ring of light usually falls between 1.00 NA and 1.40 NA. As

water and homogeneous immersion objectives have a numerical

aperture greater than 1.00 NA. it follows that they could not be

used for dark-field observation with their full aperture, because much

of the light from the condenser could enter the objective, giving rise

to a bright or at least a gray field.

Reducing diaphragms for high apertured objectives. As the lower

limit in aperture of dark-field condensers is 1.00 NA, and sometimes

even lower, it follows that a condenser for use with all objectives

requires that none of them have an aperture over 1.00 NA. As all

modern immersion objectives have an aperture greater than 1.00

NA, this aperture must be reduced by inserting a diaphragm in the

objective.

The general law that the resolution varies directly with the aper-

ture, and the brilliancy as the square of the aperture, holds with

dark-field as with bright-field microscopy. In order to determine

by actual experiment with various dark-field condensers the best

aperture of the diaphragm to select, the writer requested, the Bausch

& Lomb Optical Company and the Spencer Lens Company to supply

MODERN DARK-FIELD MICROSCOPY 109

reducing diaphragms for their fluorite, homogeneous immersion

objectives ranging from 0.50 NA. to 0.95 NA. As measured by me

these diaphragms ranged from slightly above 0.50 NA, to 0.97 NA.

These varying apertures were tested on each condenser, using the

same light and as nearly as possible identical preparations (i.e., fresh

blood mounted on slides of the proper thickness). It seemed to the

BSS

Fig. 12. Large aperture objective with diaphragm to reduce the aperture to

less than 1.00 NA. (From Chamot)

D Funnel-shaped reducing diaphragm in the interior of the objective above the

back lens.

writer that the law of aperture as stated above held rigidly. The

question then is, which aperture shall be chosen if but one diaphragm

is available? It seemed to the writer that the one of 0.80 NA should

be chosen, at least for these fluorite objectives. If three are to be

had the range should be 0.70, 0.80 and 0.90. The reason why one

over 0.90 is not recommended is because some examples of the best

of the dark-field condensers tested, seemed to have their lower limit

somewhat below 1.00 NA, and hence the field could not be made

completely dark with the diaphragm of 0.97 NA. With others,

however, the field was as dark with this large aperture as with the

lower apertured diaphragms.

A considerable range of reducing diaphragms for the homogeneous

immersion objectives is recommended because all experience brings

home to the worker with the microscope the conviction that some

structures show better with the lower apertures and some with

higher ones, and it is believed from considerable experience that the

same fundamental principles hold in dark-field as in bright-field

microscopy.

110 SIMON HENRY GAGE

LIGHTING FOR DARK-FIELD MICROSCOPY

As is almost self-evident, only a very small amount of the light

passing through the condenser to the objects is deflected by the

objects into the microscope, consequently the source of light must be

of great brilliancy or there will not be enough to give sufficient light

to render the minute details of the objects visible, when high powers

are used. This visibility of minute details involves three things:

(1) The aperture of the objectives; (2) The aperture of the illuminat-

ing pencil; (3) The intensity of the light.

The most powerful light is full sunlight. Following this is the

direct current arc, the alternating current arc and then the glowing

filament of the gas-filled or Mazda lamps.

The reflecting condensers are designed for parallel beams con-

sequently the direct sunlight can be reflected into the condenser

with the plane mirror of the microscope. If the arc lamp, a Mazda

lamp, or any other artificial source is used a parallelizing system must

be employed. ‘The simplest and one of the most efficient is a plano-

convex lens of about 60 to 80 mm. focus with the plane side next the

light and the convex side toward the microscope mirror (Fig. 14)

i.e., In position of least aberration. This is placed at about its

principal focal distance from the source whether that be arc lamp,

Mazda lamp, or any other source and the issuing beam will be of

approximately parallel rays. These can then be reflected up into

the dark-field condenser with the plane mirror.

LAMPS FOR DARK-FIELD MICROSCOPY

Up to the present the small arc lamp (Fig. 13), using 4 to 6

amperes is practically the only one considered really satisfactory.

There is no question of the excellence of the direct current arc. The

alternating current arc has two equally bright craters which renders

its use somewhat more difficult.

For most of the work in biology the arc gives more light than is

comfortable to the eyes; but a still greater objection is that with the

burning away of the carbons the source of light is constantly shifting

its position, and hence the quality of the light varies from minute

to minute. A third difficulty for hand-feed lamps is that one must

stop observation frequently to adjust the carbons.

MODERN DARK-FIELD MICROSCOPY 111

In spite of all these difficulties, however, the arc lamp is indis-

pensable if one desires to attack all the problems for which the dark-

field microscope is available.

Fig. 13. Small arc lamp for dark-field illumination (From Optic Projection)

This figure is to show the wiring necessary and the arrangement of the arc and

lens to give a parallel beam.

A—Heavy base of the lamp support. By means of a clamp the lamp can be

fixed at any desired vertical height. HC and VC, the horizontal and vertical carbons.

The HC must be made positive. F, the wheels by which the carbons are fed.

TC—tThe tube containing the condenser. The condenser in the inner tube

can be moved back and forth to get a parallel beam. Sh, black shield, see E.

E—Black shield at the end of the lamp tube (Sh). It serves to screen the eyes

and to show when the spot of light is thrown back by the mirror into the parallelizing

lens.

W1, W2, W3, W4—The wires of the circuit passing from supply to the upper

carbon (HC) and from the lower carbon (VC) to the rheostat, and from the rheostat

back to the supply in W1. Never try to use an arc lamp without inserting a rheostat

in the circuit. As shown, it forms a part of one wire. It makes no difference whether

it is in the wire going to the upper or to the lower carbon, but it must be in one of them.

6-Volt Headlight Lamp.—Next to the arc lamp in excellence for

dark-field work is the 6-volt gas-filled headlight lamp (Fig. 14).

The reason of this excellence is that the filament giving the light is

in a very close and small spiral not much larger than the crater of the

small arc lamp, and hence approximates a point source of light.

112 SIMON HENRY GAGE

The brilliancy is also very great as the filament is at about 2800°

absolute. The two sizes that have been found most useful by the

writer are the bulbs of 72 watts and those of 108 watts. For the

bulb of 108 watts a mogul socket is essential; for the 72 watt bulb

the ordinary socket is used.

Ve HN

Cocty777770)

Fig. 14. Diagram of headlight lamp and transformer for dark-field illumination

(About one-sixth natural size).

Axis—Axis of the parallel beam from the lens (L).

Lamp—The 6-volt, 108 watt headlight lamp with its very small, close filament

centered to the axis of the lens. It is in a mogul socket (ms) and can be centered

vertically and horizontally by the inner and outer tubes and set screw (it, ot, s), and

the brass slide (sl).

Lamp House—The metal container for the lamp. (bb) Bafle plates near the

bottom to help avoid stray light. At the left over the lens (L) is the sloping eye shade.

L D G—Parallelizing lens cemented to polished daylight glass.

Lamp wires—The large wires from the transformer (Transf.) to the lamp (Double

heater wires are good).

m c—Mistakeless connection between the lamp wires and the transformer

(Transf). This is a Manhattan stage connector, and is different from anything else

in the laboratory and therefore the lamp can never be connected with a 110 volt cir-

cuit and burn out the lamp. Of course any other wholly different connection would

answer just as well.

Transf.—Diagram of a step-down transformer. As there are 18 coils around

the soft iron ring on the Primary (P) or 110 volt side, and but one coil around the

Secondary (S) side, the voltage is stepped down 18 times, or from 110 to 6 volts. In

an actual transformer the coils would be far more numerous, but in this proportion.

If the transformer were connected wrongly, i.e., with the lamp wires connected with

MODERN DARK-FIELD MICROSCOPY 113

the primary (P) side, and the 110 volt supply with the secondary (S) side, it would

then be a step-up transformer, and raise the 110 volts 18 times—with disastrous

results. C, separable connection for the 110 volt supply wires.

The only difficulty with these lamps is that as they are for a

6 volt circuit it is necessary to use a step-down transformer if one

has an alternating current with a voltage of 110 or of 220, as is usual.

If one has a direct current of 110 or 220 voltage, then it is neces-

sary to use a storage battery, in general like those used for the

lighting and ignition systems of automobiles. As a transformer

uses up but a very small amount of energy it will be readily seen that

in stepping down the voltage the amperage is correspondingly raised

from the general law that the wattage is the product of the voltage

into the amperage, and knowing any two the third may readily be

found.

For example with the 72 watt lamp, if the voltage is 6 the amper-

age must be 72/6 or 12 amperes. With the 108 watt bulb the

amperage must be 108/6=18 amperes.

The heating of the filament is determined by the amperage, and

also it must be remembered that the conductor of an electric current

must be increased in due proportion for an increased amperage, con-

sequently in the transformer the wires joining the 110 volt line is

small because a very small amperage is necessary to give a large

wattage; while from the transformer to the lamp the conducting

wires must be large, to carry without heating the amperage necessary

with the low voltage (6) to give the large wattage (108 or 72).

For the 18 amperes of the 108 watt bulb, the Fire Underwriters

specifications call for wire of No. 12 or No. 14 Brown and Sharp

Guage, i.e., wire 1.6 to 2 mm. in diameter or a cable composed of

smaller wire having the same conductivity. This specification is

for continuous service. In wiring the headlight lamp from the trans-

former, so called heater cable is good, provided one uses a double

cable, that is the entire cable for each wire. ‘This is easily done by

removing the insulation at the ends and twisting the two strands

together, then it can be treated as one wire and the two thus treated

used to join the lamp to the mistakeless connection (mc, Fig. 14, 15)

of the transformer. As the resistance is small in these large con-

ductors the full effect of the current remains to make especially

brilliant the glowing lamp filament, and brilliancy is what is needed

for this work.

114 SIMON HENRY GAGE

It should be stated that the transformer for this purpose should

be substantial and adapted to continuous service. It is known as a

“Bell Transformer”’ as it is connected to ordinary house light systems

for ringing door bells. The one used by the writer was obtained

from the General Electric Co. in 1920 and costs at present seven

dollars. It is marked: Transformer, type N D, Form P Volts 110

6. Capacity 108 KV-A, Cycles 60, Without taps in Primary.”’

(For making the connections, see the explanation of Fig. 14.)

In comparing the two 6 volt lamps for dark-field work, the 72

watt lamp answers well for most purposes, but the 108 watt one

approximates more nearly to the small arc lamp and is sufficient for

probably 99% of all dark-field observation in biology. For the

remaining 1% one could safely depend on sunlight.

Siereopticon and Mazda lamps for dark-field. In absence of the

head-light lamps described above, one can get good results by using

in the lamp-house (Fig. 14-15), a stereopticon lamp bulb of 100 to

25@ watts. These bulbs have the filament arranged in a kind of ball,

and hence fairly well concentrated. This filament must be centered

with the parallelizing lens as described for the headlight bulbs.

For the horizontal position, move the lamp back and forth by the

brass slide until the front of the ball filament is in focus on the 10-

meter screen. The microscope should then be placed from 15-25 cm.

from the lamp-house. The rest of the procedure is exactly as for

the headlight lamp.

If one has neither headlight lamp nor stereopticon lamp, still

good work can be done in biology by using the Mazda C bulbs where

the filament is in the form of a loop or C. This is centered and

focused as for the other lamps (Fig. 18). If one has only a lamp

similar to Fig. 18, the daylight glass can be removed and the micro-

scope placed close to the lamp. Fairly good results can be obtained

with a 100 watt mazda stereopticon or c bulb without a parallelizing

lens.

The Spencer Lens Company recommend in addition to the small

arc lamp, their small magic lantern (No. 394). This has either a

250 or a 400 watt stereopticon lamp bulb, and for parallelizing

system, the two plano-convex lenses common with simple magic

lanterns. The projection objective of the magic lantern is removed.

This yields good results especially when a piece of clear daylight glass

MODERN DARK-FIELD MICROSCOPY 115

is placed over the end of the cone left vacant by the removal of the

objective.

A real advantage possessed by these different lights is that the

lamps are connected directly with the 110 volt circuit, no transformer

being required, as with the headlight lamps. But if one is to do

much dark-field work the headlight lamps are much to be preferred.

Daylight effects with the headlight or Mazda lamps. For dark-field

work as for work with the bright field, daylight effects are of the

greatest advantage both for eye comfort and for the clearness with

which details can be made out. The daylight effect is readily ob-

tained by using a piece of daylight glass polished on both sides and

cemented to the flat face of the parallelizing lens by means of Canada

balsam (Fig. 14-15).

] \ Supply

||Tranef wires

Fig. 15. Headlight lamp in its metal house, and the step-down transformer.

(About one-eighth natural size)

D. Glass—The window of daylight glass on the side of the lamp-house to be used

for bright-field work. With the glass removed the centering of the lamp is facilitated.

P. lens—Parallelizing lens of about 75 mm. focus. It is cemented to a piece of

polished daylight glass.

m c—Mistakeless connection between the lamp wires and the transformer

(Transf.). Such a connection prevents joining the lamp with the 110 volt circuit,

and thus burning it out. This cannot be connected wrongly.

Transf.—Step-down transformer from 110 to 6 volts.

Lamp-House with centering arrangement. To avoid the non-

utilized light, and to place the source of light in the most favorable

position, there must be an opaque box to enclose and support the

head-light or Mazda lamp. As the filament giving the light must

be in the optic axis and practically in the focus of the parallelizing

116 SIMON HENRY GAGE

lens, the lamp or the lens must be sufficiently movable to attain the

end. In the lamp-house here figured (Fig. 14, 15) the lens is stationary

and the lamp is movable horizontally and vertically, that is, it can be

raised and lowered and moved toward and from the lens in the optic

axis. For the most perfect centering there should also be arrange-

ments for moving the lamp or the lens from side to side. In the one

here shown the parallelizing lens can be shifted slightly to take care

of the lateral centering.

Centering the Lamp-filament. As stated above the lamp-filament

must be centered, that is, put in the principal optic axis of the paral-

lelizing lens. This is most satisfactorily done by putting the paral-

lelizing lens in position in the lamp-house and measuring the distance

from the table to the middle point of the lens. The middle point of

lamp filament should be placed at the same height from the table.

This is easily accomplished by using the side window of the lamp-

house and raising and lowering the lamp by means of the vertical

adjustment (Fig. 14-15) until the filament is at the right height to

be on the level of the optic axis. Then the lamp is turned until the

spiral filament faces the lens. The two limbs of the fork holding

the filament then face sidewise. Of course, they would make a

shadow if they faced the lens.

To get a parallel beam. ‘The most satisfactory way of doing this

is to work at night or ina dark room. Having a white wall or white

screen at about 10 meters distant, light the lamp and move it back

and forth in the optic axis by means of the top slide (Fig. 14 sl) until

the filament of the lamp is in focus on the screen, the filament will

then be at about the principal focus of the parallelizing lens, that is,

in a position to give approximately parallel light to the microscope.

It is well to mark the position on the top of the lamp-house so that

if it gets accidentally displaced it can be returned without trouble.

It may be said in passing that the lamps are not all exactly alike so

that when a new lamp is installed it is necessary to center and focus

all over again.

Focusing the crater of the small arc lamp. The makers arrange

the carbons and the lens tube so that the crater will be approximately

in the optic axis (Fig. 13). Now to get the crater in the focus of the

parallelizing lens one can proceed in principle as with the headlight

lamp. In the arc lamp, the carbons are fixed and the lens movable.

Work at night or in a dark room and with the lighted arc move the

MODERN DARK-FIELD MICROSCOPY 117

lens back and forth until there is a sharp image of the crater on the

10-meter screen.

Lighting the Microscope. Assuming that the lamp filament

or the crater of the arc lamp is centered with the parallelizing lens,

one can find the best position for the microscope by holding some

thick white paper in the path of the beam and slowly moving out

along the beam. Where the spot of light is brightest and most

uniform is the best place for the microscope mirror. With the

headlight lamps and the arc light this is usually 20-30 cm. from the

parallelizing lens.

To get the spot of light to fall on the 45° mirror properly, the

center of the mirror must be at the level of the axis of the beam.

This can be brought about either by raising the microscope on a

block, by inclining the microscope, or by tipping the lamp-house over

toward the microscope. If some white paper is put over the mirror

one can tell easily when the cylinder of light falls upon it.

To get the light up through the condenser and into the objective

it is necessary to so tip the mirror that an image of the source of

light is directed back into the parallelizing lens. This image is

reflected back from the flat top of the condenser to the mirror.

With this arrangement of the mirror the microscope is almost always

well lighted, and the mirror will need but a slight adjustment to give

the best possible light. This will only be true however, when the

source of light is centered to the parallelizing lens and the condenser

to the axis of the microscope.

This method of lighting the microscope saves much time and

worry. It is effective with the microscope vertical or inclined, with

the lamp-house vertical or inclined, and finally it is unnecessary to

have the microscope in line with the beam of light. It may be at

right angles or at any angle provided the beam of light falls directly

on the mirror and the image of the source can be reflected back to

the parallelizing lens.

This method of lighting the microscope, so simple and generally

applicable, has the one draw-back that the reflected image is

rather faint and therefore not easily seen in a light room; at night or

in a dark room it is very easily applied. If one is using the head-

light lamp and the parallelizing lens is on the outside as shown in

Fig. 14-15, one can tell easily when the image is reflected back into

the lens from the bright image seemingly considerably nearer the

118 SIMON HENRY GAGE

lamp filament than the blue image of the filament shown in the lens.

To see these images one should look obliquely into the lens, that is,

along a secondary not along the principal axis. One can also gain

help in lighting by turning the mirror till a spot or ring of light

appears on the upper end of the condenser. If the slide isin place with

the oil for immersion, the spot of light will be bright. One must

usually change the mirror slightly after the preparation is in focus to

get the best light.

CENTERING AND FOCUSING THE DARK-FIELD CONDENSER

As can be seen by Fig. 6-7 the object must be in the focus of the

dark-field condenser and this focus must be in the optic axis of the

microscope.

The dark-field condenser must have a special mounting with

centering screws, which is the common method; or if the microscope

has a centering sub-stage arrangement the dark-field condenser

need not have a special centering arrangement, but be put in the

centering substage fitting. Ordinarily there is no centering arrange-

ment on a microscope and hence the dark-field condenser must have

a special centering arrangement of its own. The whole is then

placed in the usual bright-field substage condenser ring and raised

until it is at the level of the top of the stage. Asa guide to centering,

there is a circle scratched on the upper surface of the condenser

(Fig. 7 c-r). With a low power (16 mm. objective or lower, and

x5 ocular) one focuses down on the end of the condenser and if the

small circle is not concentric with the circle of the field the centering

screws are used with the two hands at the same time and adjusted

until the circles are exactly even all around. Unfortunately this is

not sufficient for the most satisfactory work, as it is rare that any

two objectives will be exactly centered even though screwed into

the same opening in the nose-piece, and much less likely to be

centered if in different openings. To get the best results the objective

to be used and the dark-field illuminator must be centered to each

other. To accomplish this the following procedure has been found

simple and certain: To start with the dark-field condenser is centered

by the low objective as described above, and then with a crow-quill

or other very fine pen one puts a very small point of Chinese white

or other white ink in the middle of the little centering circle. This is

MODERN DARK-FIELD MICROSCOPY 119

easily done if an objective of 20 to 40 mm. focus is used for centering

the circle on the condenser.

Now for centering the oil immersion or other high power objective

the field of which is less than the centering circle, the objective is

put in place, but no immersion liquid need be used for the centering.

The top of the condenser has dusted upon it some starch or flour or

other fine white powder so that in focusing down upon the top of the

condenser there will be some shining particles to focus on if the

white ink in the center of the circle should happen to be entirely

out of the field, which is often the case. When the objective is in

focus the centering screws are used to shift the condenser until the

minute spot of white ink in the center of the circle is exactly in the

middle of the field. In this way any objective may be centered with

the condenser, and so far as the centering is concerned, one can be

sure of getting the best results of which the condenser is capable.

When the condenser is centered to the high objective, the starch

particles and the white ink may be removed with a piece of moist

lens paper or a soft cloth.

Focusing the Condenser on the Object Level. This is one of the

most essential steps for good dark-field work. If the objects are not

in the focus of the condenser they will not be sufficiently lighted so

that they can radiate enough light into the microscope to show all

their details.

One can proceed as follows, it being assumed that the preparation

is mounted on a slide of the proper thickness for the given con-

denser:—Use a low power, 16 to 50 mm. objective and light the

microscope as described in the preceding section. Look into the

microscope and focus on a saliva preparation. Move the slide around

until there are plenty of epithelial cells in the field and then make

slight changes in the mirror until the most brilliant light is obtained.

With the screw device for raising and lowering the condenser shift

the position up and down slightly until the smallest and most bril-

liantly lighted point is found. When this is accomplished the

condenser is in the optimum focus for that slide and will give the

most brilliant light of which it is capable for the source of light used.

Any preparation for examination can have the condenser focused

upon it as just described.

For experimental purposes a very satisfactory preparation for

focusing the condenser is made as follows: A slide of the right thick-

120 SIMON HENRY GAGE

ness is selected and cleaned and on one face near the middle is painted,

with a fine brush, a very thin layer of Chinese white or other white

ink. When this is dry, a drop of Canada balsam is put upon it and

then a cover-glass. The white particles are very fine and serve

admirably to show the focal point of the condenser. Such a slide

can be kept as a standard and if the condenser is focused by its aid,

it will be in the right position for any preparation mounted upon a

slide of the same thickness as the standard. One must always

remember, however, that many preparations have an appreciable

thickness, and if the slide were of exactly the same thickness as the

standard the light might be made more brilliant in a given case by

focusing the condenser slightly upward for the higher levels of the

preparation. This shows also that the slides selected for prepara-

tions should be somewhat under the maximum thickness allowable

for the given condenser.

Fig. 16. Face and sectional views of the focus of the hollow cone of light from

dark-field condensers

A—Sectional view of an optically perfect dark-field condenser in which the sun

is represented as focused nearly to a point. No such condenser exists.

B—Sectional view of a possible condenser focus. It is drawn out somewhat and

spreads laterally. The variation in the thickness of slide which might properly be

used is shown by the two parallel lines enclosing the elongated focus.

C—Sectional view with a still more elongated focus. The parallel lines show that

the variation in thickness of slide permissable is correspondingly increased.

The apparent size of the sun’s image is shown on the axis above in each case.

It is least sharp in C.

The black line above the letters (A, B, C) represents the top of the condenser.

Thickness of glass-slide to use. Mention has been made of glass

slides of the proper thickness. What should this thickness be and

how can it be determined are pertinent questions for one who is

to get satisfactory results in dark-field work. The thickness of the

slide with any given condenser is that which will bring the focus of

the condenser—that is the image of the source of illumination—on

the upper face of the glass slide where the object is located. Either

MODERN DARK-FIELD MICROSCOPY 121

on the instrument or in the maker’s directions for its use the thickness

of slide which should be used with it is given. If such definite infor-

mation is not available or if a person wishes to determine for himself

the proper thickness of slide to use, it may be found out as follows:

An arc lamp and a dark room are necessary. The light should pref-

erably be parallelized as shown in Fig. 13. The tube of the micro-

scope is removed, and a piece of uranium glass with plane faces is

placed on the stage and connected with the top of the condenser by

homogeneous immersion liquid. The uranium glass is strongly

fluorescent and shows with great definiteness the exact path of the

beams of light from the condenser. One can see exactly where the light

comes to a focus above the condenser and then the diverging beams

above the condenser. If the condenser were perfect the rays would

focus very accurately at a point above the condenser face, Fig. 16 A.

This focal point is where the object should be placed and its distance

above the condenser face gives the thickness of the slide to use.

One can see that with an optically perfect condenser the thickness

should be very exact to get the most brilliant image. [Ii the optical

system is less perfect as shown in B Fig. 16 the rays do not all cross

at one point, but over an appreciable thickness and anywhere within

that elongated focus would give a brilliant illumination. In this

case the thickness of the slide used could vary the length of this focus.

In Fig. 16 C the focus is much elongated and the slide might

vary greatly in thickness and still give a brilliant image. Above the

sectional view of the focus in each case is given a face view of the

brightest point as described above in getting the focus of the con-

denser. One can readily see that the more perfect the focus at a

point the smaller will be the point of light, and as all the rays are at

that point it will be dazzlingly brilliant, while with B and C, where

only part of the rays focus at any given level the circle of light will

be less brilliant, but correspondingly greater in diameter. The

larger circle of light has the advantage of giving a larger illuminated

field, but the disadvantage of loss of brilliancy for the most exacting

work. It should be mentioned also that as the focus gives an image

of the source of light, the size of the source of light will also affect the

size of the bright spot seen in looking down on the image. This is

finely brought out by using the sun as a source and the arc light or the

incandescent light.

£22 SIMON HENRY GAGE

One can see also from these figures that if the slide is too thin the

objects will be partly in the dark space between the converging beams,

and if the slide is too thick a part of the objects will be in the dark

space between the diverging beams. If one sees the face view with

a low power in either case there will be a ring of light and a central

dark disc. and will look something like the central stop in Fig. 5 D

As the preparations (blood, saliva, etc.) usually studied by the

dark-field method have an appreciable thickness it is better to use a

slide somewhat thinner than the optimum where the object is almost

exactly at the level of the upper surface. If the slide is somewhat

thinner the various levels of the preparation can be focused on by

the condenser by slightly raising and lowering it as the case demands.

For example, if the optimum thickness is 1 mm. it is better to use

slides of 0.90 or 0.95 mm. and if the optimum thickness is 1.55 mm.

it is better to use one of 1.50 mm. for ordinary preparations.

Thickness of cover-glass and tube-length. These should be strictl

in accordance with the construction of the objective. In all mode n

objectives the makers state the tube-length and thickness of cover

glass for which unadjustable objectives are corrected. As the dark-

field illumination brings out very sharply any defects of correction

in the objective, one should select a cover of the thickness, and the

length of tube recommended by the maker of the objective. This

applies particularly to dry, inadjustable objectives. If the objec-

tives are dry and adjustable then corrections can be made for varia-

tions from the standard of cover thickness or tube-length.

If the objective being used is homogeneous immersion, the tube-

length must be carefully attended to, but the thickness of the cover-

glass is immaterial so long as it is thin enough to fall within the

working distance of the objective; of course if it were thicker than

that one would not be able to get the objective in focus (Bausch, ’90;

Gage, ’87, 1912).

PRACTICAL APPLICATION OF DARK-FIELD MICROSCOPY

In the practical application of dark-field microscopy it is self-

evident that it can be used successfully only with objects scattered,

leaving a certain amount of blank or empty space between the

objects. If the object being studied covered the whole field then it

would all appear self-luminous and give a continuous bright appear-

ance filling the whole field of the microscope.

MODERN DARK-FIELD MICROSCOPY 123

In Biology, used in the comprehensive sense applied to it by

Huxley, there come naturally the following groups of objects in which

it is applicable, and likely to yield much information :—

(A) Unicellular organisms in both the plant and the animal

kingdoms. This of course would include the Protozoan Animals,

the Bacteria, and other unicellular plants.

(B) In the multicellular animals and plants it includes the natural

fluid parts with their cellular and granular contents. In the verte-

brates, including man, this would, for example, comprise the blood,

and the lymph, with their cellular and granular contents; the tissue

fluids, and the fluids in the natural cavities like the pericardial, the

pleural and the peritoneal cavities, and the liquid found in the

cavities of the central nervous system, the joint cavities and tendon

sheaths. It is also of great service in the study of the liquids found

in mucous containers, as milk, urine, bile, the saliva, the mucous in

the nose, and other organs lined with mucous membrane.

Furthermore it is of help in the study of isolated elements of the

body like ciliated cells, etc. In a word it is applicable to the study

of all animal and vegetable structures—including the pathologic

ones—that are naturally isolated, or that can be artificially separated

so that there is sufficient blank space between the structural parts.

Dr. Chamot points out its help in the biological examination of

water, in the study of foods, fibers, crystallization phenomena, sub-

microscopic particles and colloids. He adds further (p. 40): ‘‘This

method is invaluable for demonstrating the presence of very minute

bodies or those whose index of refraction is so very nearly the same

as that of the medium in which they occur as to cause them to escape

detection when illuminated by transmitted light,” i.e., by bright-

field microscopy.

SUMMARY OF STEPS NECESSARY FOR SUCCESSFUL DARK-FIELD

OBSERVATION

1. A powerful source of light must be available.

2. The dark-field condenser is put in place in the substage, and

raised until the top is flush with the upper surface of the stage. The

condenser is then accurately centered. If there is an iris diaphragm

below the condenser it should be made wide open.

3. A homogeneous immersion objective with reducing diaphragm

of about 0.80 N.A. is screwed into one of the openings of the nose-

piece of the microscope.

124 SIMON HENRY GAGE

4. Slides and cover-glasses of the proper thickness are made very

clean, and put in position for rapid handling.

5. The preparation to be examined—blood, saliva, etc.—is

mounted on the slide and covered; the cover-glass is sealed with min-

eral or castor oil, or with shellac cement.

6. The mounted preparation is held in the hand and one or more

drops of homogeneous liquid put on the lower side of the slide oppo-

site the cover-glass. The slide is then put upon the stage so that the

homogeneous liquid makes immersion contact with the top of the

condenser. The condenser may need to be raised or lowered slightly

to make the contact perfect.

7. A drop of homogeneous liquid is put on the cover-glass.

8. The mirror is turned until there is a brilliant point of light in

the homogeneous liquid on the cover. The objective is then lowered

until it dips into the immersion liquid.

9. The microscope is then focused and the light made as brilliant

as desired by turning the mirror.

10. Dark-field microscopy requires more accuracy of manipula-

tion than does ordinary microscopy, but the increased visibility pays

for all the trouble. A dimly lighted room is desirable for then the

eyes are adjusted for twilight vision and can more easily make out

the finest details.

Method of Procedure. Asan example of the method to be followed

in dark-field work, blood may be used. As pointed out nearly 50

years ago, by Dr. Edmunds, blood with dark-field illumination seems

like a new structure, so many things are seen with the greatest dis-

tinctness that are wholly invisible or only glimpsed when seen by

the bright-field method.

(1) Slides of the correct thickness for the condenser are selected

and carefully cleaned.

Cover-glasses are also cleaned and placed where they can be

easily grasped.

(2) For obtaining the fresh blood the part to be punctured should

be cleaned well with 95% alcohol and then with a sterilized needle

or Dr. Morre’s Haemospast, the puncture is made. The drop of

blood exuding can be quickly touched by a cover-glass, and the cover

put on the center of one of the prepared slides. If a small amount

adheres to the cover, it will spread out in a very thin layer when

placed on the slide. At least one preparation should be made which

MODERN DARK-FIELD MICROSCOPY 125

appears quite red. In making the preparations one should work

rapidly so that the various corpuscles will be in their normal numbers,

and the fibrin will be formed only after the preparation is on the slide.

If all the preparations are quite red, after a few minutes, one can

be made thinner by pressing firmly on the cover by the ball of the

thumb covered with gauze or lens paper. The gauze or paper absorbs

the blood which runs out at the edge of the cover. In order to

prevent evaporation and to help anchor the cover-glass so that it

will not move by the pull of the viscid homogeneous immersion

fluid, it is advisable to seal the cover by painting a ring of liquid

vaseline (petroleum oil) or castor oil around the edge of the cover.

One of the thick preparations should not be sealed, but kept for

irrigation with normal salt to show especially the fibrin net-work.

When ready to study the blood, put a large drop, or two large drops,

of homogeneous liquid on the underside of the slide directly opposite

the specimen, and place the slide on the stage of the microscope so

that the immersion liquid will come over the face of the condenser.

Then a drop of immersion liquid is put on the cover-glass and the

objective run down into it. If the lighting is secured as explained

above one soon learns to focus on the specimen. In general, the

field all looks bright just before the objective gets down to the level

for seeing the specimen.

(a) The erythrocytes will appear like dark discs with bright rims

owing to the convex borders.

(b) The leucocytes appear as real white corpuscles owing to the

granules within them which turn the light into the microscope. If

the room is moderately warm—20 C or more—the leucocytes, some

of them, will undergo the amoeboid movement, and the picture they

present will be a revelation to those who never saw it or only with the

bright-field microscope. From the clearness with which everything

can be seen the minutest change can be followed, and also the most

delicate pseudopod detected. Another striking feature will be

noticed in the moving ones, that is, the vigorous Brownian movement

of the granules in the part of the leucocyte with the amoeboid move-

ment. In those showing no amoeboid movement there is usually no

sign of the Brownian movement of the granules; also if a part of the

leucocyte is not undergoing amoeboid movement the particles in it

are usually motionless.

126 SIMON HENRY GAGE

tween the corpuscles. In different parts of the specimen one can

find all the appearances of the fibrin shown in text-books on the blood.

(d) Chylomicrons appear everywhere like bright points in the

empty spaces. They are in very active Brownian movement. These

chylomicrons will probably be the most unusual part to those study-

ing blood with the dark-field for the first time.*

A very striking view of the fibrin net-work may be obtained by

irrigating the thick blood preparation. If a drop of normal salt

solution is placed on one edge of the cover-glass and a piece of

blotting paper on the other the liquid is drawn through washing out

many of the erythrocytes. If the washing out process is watched

under the microscope the erythrocytes will be seen gliding over or

through the fibrin net-work, or some of them will be anchored at

one end and if the current is rapid the corpuscles will be pulled out

into pear-shaped forms.

The leucocytes look like big white boulders in the stream, wholly

unmoved by the rushing torrent around them.

HISTORY

Almost always in human progress two steps must be taken (1)

The discovery of the fundamental principles involved, and (2) the

development of knowledge in other fields to make the application of

the principles possible. Often a long time, sometimes a very long

time, intervenes between the first steps and the final rendering of the

knowledge a part of the common knowledge of mankind. The

development of Dark-Field Microscopy is a good illustration of both

the statements made.

*The term chylomicron is from two Greek words; xbdés, juice or chyle, pxpév, any

small thing, technically the one-thousandth of a millimeter (u). I have introduced

this word to show the origin of these bodies from the chyle, and to indicate their

general average size. Gulliver in 1840-1842, called these minute granules the molecular

base of the chyle and showed that they were identical in the thoracic duct and in the

blood vessels of the same animal. He gave their average size as 1/36,000 to 1/24,000

of aninch. They have been called by others free granules or granulations, elementary

particles, etc. In 1896 H. F. Mueller described them as “A never-before observed

constituent of the blood” and gave the name of haemoconia, literally, blood-dust.

(See Gulliver, Lond. Edin. Phil. Mag. Jan. Feb. 1840; Appendix to Gerber’s Anatomy,

1842, and notes in the Works of Hewson, 1846; Mueller, Centralblatt f. allg. Path. u.

path. Anatomie, Bd. 7, 1896, pp. 529-539).

MODERN DARK-FIELD MICROSCOPY 127

The ancient opticians, thousands of years ago, knew well that

the principle of contrast. was of the highest importance in rendering

objects visible; but before this could be applied in microscopy, the

microscope itself must be devised. This we see in its simplest form

in the convex lenses of Roger Bacon (1266-1267) and in the now

rarely used compound form of the Dutch spectacle makers, Jansen

and Laprey (1590), composed of a convex objective and a concave

ocular (Fig. 17). As a result of the Dutch Compound Microscope,

Kepler was led to devise the modern form composed of a convex

objective and a convex ocular (1610). But this Keplerian com-

pound microscope has undergone many changes since its first concep-

tion and many modifications to render it suitable for giving

ability to show the delicate structures in nature with their true

appearance. Among these changes may be mentioned the prepara-

tion of achromatic lens combinations (Dolland 1757) for telescopes

and applied to microscopic objectives between 1820-1830, put on the

road to perfection by the introduction of the immersion principle

(Hooke 1678, Brewster 1813, Amici 1840-1855) and by the aperture

made available by the homogeneous immersion objectives of Tolles

1871-1874, and by the apochromatic objectives of Abbe. Condensers

for lighting the object have also played a prominent part from that

of Descarts (1637) to those recommended by Brewster (1831) and

the homogeneous immersion condensers of Wenham, Tolles (1856 to

1871) and those now regularly made for homogeneous contact with

the slide supporting the specimen.

Among the subsidiary discoveries were necessary the arc-light of

Davy (1800) and the right-angled arc lamp of Albert T. Thompson

(1894) (Fig. 13) and the electric generators now everywhere available.

In these last days also the gas filled or Mazda lamps with their

close filaments of Tungsten which approximate in brilliancy and

compactness of source to the arc lamp and greatly excel it in con-

venience; and lastly of the production of a glass filter to give the light

of the tungsten incandescent lamps true daylight quality, and make

microscopic work by this artificial light as comfortable as the light

from the northern sky (see Ives 1914, Gage 1915-1916).

The time also between the first appreciation of the dark-field for

the study of microscopic objects by Lister (1830), Reade (1838),

Wenham (1850), Edmunds (1877), and the appreciation of the micro-

scopical worker in general, came only after the invention of the ultra-

128 SIMON HENRY GAGE

microscope (1903) and the application of the dark-field method to the

study and detection of pathologic micro-organisms especially the

Spirochaeta pallida (1905). It now promises to give much help in

working out the activities and minute details of microscopic structure

in animals and plants from the lowest to the highest.

In the earliest stages of microscopic study the objects were seen

by the light which they directed toward the microscope, and if over

a dark background they appeared with varying degrees of brightness

as if self-luminous; but even as early as 1637 (Fig. 17) Descartes

microscope had provision for sending the light through the object.

In this case much of the light did not reach the object at all, but

passed on directly to the microscope. This mode of lighting showed

the object more or less as a dark body on a brilliant background.

Fig. 17. Descartes Dutch compound microscope with a parabolic reflector and

a condensing lens (From Descartes Dioptrique, 1637).

Ocular and Objective. The ocular is a plano-concave lens or amplifier, and the

objective (N O P R) is a double convex lens.

Reflector and Condenser. For objects to be lighted from above, there is a para-

bolic mirror (c c); for those to be lighted from below there is a condensing lens (i i).

MODERN DARK-FIELD MICROSCOPY 129

These two forms of lighting differed fundamentally in that with

the first no light from the source passed into the microscope; but only

that from the object, while with the second the light from the source

as well as from the object got into the microscope.

The significance of this fundamental difference for the aperture

of the objective and for dark-field microscopy were first appreciated

by Lister (1830), Wenham (1854), and Gordon (1906), and was

practically applied in the manufacture of dark-field apparatus by

Zeiss (1904) and Leitz (1905). In a word, it was the appreciation,

as stated by Lister (1830) that-if the direct light from the source

after it had reached the object, were prevented from entering the

objective, by blacking the central part of the objective, then only

the marginal part of the objective would be functional and that

would receive only those rays from the object that were directed to

it by the object itself, that is scattered light reflected, refracted, or

diffracted, from the object, none of the light from the source getting

directly into the microscope. As stated by Wright (p. 217) this is

the method of dark-field microscopy by lighting the object with a

solid cone of small aperture and, imaging it by hollow beams of large

aperture. In practice this method has been discarded for the one

by which the object is lighted by beams of light in such a direction

with reference to the axis of the objective that none of them can

enter the objective directly, and the light going to the microscope

comes only from the objects themselves; they will therefore appear

self-luminous on a dark background.

The two conditions are (a) where the light is directed upon the

object from above and, therefore away from the objective, and (b)

where the light is directed upon the object from below, and therefore

toward the objective (Fig. 3-4).

If the light is directed upon the object from above and the object

is over a non-reflecting background, the object will appear bright

in a dark field. Of course, if it is on a light background that will also

reflect light into the microscope and both object and background will

appear light. It is assumed here that the object or objects cover

only a part of the field, leaving plenty of empty space for background.

In striving after a truly non-reflecting background three distin-

guished men found the same thing, viz., that the only really black

thing in nature is a black hole, that is, a space with black walls into

which the light cannot enter directly. The dark walls absorb any

130 SIMON HENRY GAGE

stray light, and the empty space gives no reflection. The first of

these men devised for his microscopic purposes such a non-reflecting

background by means of a small cup or well with the walls painted

black. It is known as Lister’s black well (1826). The second dis-

coverer was Chevreul (1839), who found in his work on Contrasts

that a black space gave the only non-reflecting background. Sucha

background was used by Marey for making moving pictures to show

animal movements. Marey called it Chevreul’s black. The third

was J. H. Comstock (1901) who found in the study and photography

of spider webs that no pigment or fabric was black enough for a back-

ground He therefore devised a deep box with the inner walls

covered with black velvet and placed it so that the light could not

shine into it. Over the mouth of this box the web was placed and

lighted at right angles to the opening of the box. The feeble light

the webs reflected served well for photography.

These three men then absolutely independently found the same

solution to their problem and doubtless many others have found

also that Lister’s, Chevreul’s, and Comstock’s black space is the only

really black thing in nature.

From the time of Descartes (1637) the means for lighting objects

from above the stage have been many. Some of them, like the

bull’s eye condenser (Fig. 4, lens) and the side reflector send the

light only from one side, while with the circular mirror of Descartes

(Fig. 17) and the somewhat similar Lieberkuhn reflector (1740) the

light is reflected from all sides upon the object. If now the object

is on a dark background, it will appear as if self-luminous.

From 1850 to the present two additional means have been devised

for lighting from above. The first, following the suggestion of

Riddell (1852) aims to make the objective its own condenser, the

light being introduced into the side of the objective and reflected

down by a small mirror or a prism (H. L. Smith 1865, Tolles 1866).

(For a full discussion see W. A. Rogers, Journal of the Royal Micr.

Soc., 1880, p. 754-758.)

The other method referred to is that of Prof. Alexander Silverman

of the University of Pittsburgh. It consists of a circular electric

lamp and reflector which surrounds the objective and shines down

upon the object.

MODERN DARK-FIELD MICROSCOPY 131

Of course all objects lighted from above the stage will give true

dark-field effects only when there is a black background, and the

objects are scattered, leaving empty space between them.

Dark-Field Microscopy with Substage Illumination. The first

specific discussion of the possibility of dark-field microscopy with

light from beneath the stage is found in a paper by the Rev. J. B.

Reade of Cambridge University and is dated at Peckham, Nov. 1836,

and is published as appendix No. 2 in the Micrographia of Goring

and Pritchard, 1837. Reade says: p. 229: “To illustrate the two

methods (Bright-field and dark-field) by reference to the telescope

it may be observed that the discomfort of viewing spots on the sun

not unaptly corresponds with the view of microscopic objects on an

illuminated field; while the removal of all inconvenient and ineffective

light from the field of the microscope corresponds with the clear and

quiet view of stars on the dark blue vault of the firmament.” He

brings out very clearly in his paper that no light from the source

shall pass directly into the microscope, only that from the object,

and that the object appears “sparkling with exquisite lustre on a jet-

black ground.”

The first appearance of this method in the general literature of

microscopy which was found occurs in John Quekett’s Practical

Treatise on the Use of the Microscope, Ist ed. 1848, pp. 178-179. He

also furnishes a diagram to illustrate the method of lighting some-

thing like fig. 4 of the present article, and remarks: ‘““The method

consists in illuminating the object by a very powerful light, placed at

such an angle with the axis of the microscope that none of the rays

can enter it except those which fall directly upon the object, and are

so far bent as to pass through it into the compound body,” i.e., into

the tube of the microscope.

It is referred to in the first edition of W. B. Carpenter’s “The

Microscope and its Revelations” (1856) as follows:

“Whenever the rays are directed (from below the stage) with such

obliquity as not to be received into the object-glass at all, but are

sufficiently retained by the object to render it (so to speak) self-

luminous, we have what is known as the black ground illumination;

to which the attention of microscopists generally was first drawn by

the Rev. J. B. Reade in the year 1838 (1836-1837) although it had

been practised sometime before not only by the author (Dr. Carpen-

ter) but by several other observers.”’

132 SIMON HENRY GAGE

In addition to the condensing lens of Reade for throwing the very

oblique beam of light upon the object, the mirror was used for low

powers, and for higher powers, prisms were used especially by Nachet

and Shadboldt (1850). It was seen however, that light from only

one side might give rise to false appearances.

In the third volume of the Transactions of the Microscopical

Society of London, there appeared an epoch-making paper for

dark-field microscopy. It is entitled ‘On the Illumination of

Transparent Microscopic Objects on a New Principle.” It was

read by its author, F. H. Wenham, April 17, 1850. After discussing

the prisms of Nachet and pointing out the defect of oblique light from

one side only giving rise to false images, he proceeds to show how the

defect may be obviated by using two prisms giving light from oppo-

site sides, or, and this is the epoch making part of the paper for dark-

field work, by using a truncated parabolic reflector to give a circle of

light. A dark stop was present to cut out all but the rays which

exceed the aperture of the objective “‘So that the light which enters

the microscope shall be that which radiates only from the object,

as if it were seli-luminous.”’ The parabolic speculum was truncated

so that the light would focus on an object mounted upon the ordinary

glass slide.

From this fundamental beginning, illumination by a hollow cone

of light by the aid of the truncated parabola, all the advances in

dark-ground illumination have proceeded. In 1851, Mr. Shadboldt

says: “In order to obviate the objectional shadow (of lighting from

one side only) as well as to procure a more brilliant illumination

the parabolic condenser was projected by Mr. Wenham, to whom

alone belongs the credit of having suggested the use of oblique illumi-

nation in every azimuth, so as to produce a black field.” In this

paper Mr. Shadboldt commends the use of a condenser made wholly

of glass and depending upon internal reflections to take the place of

the metallic parabolic mirror of Wenham. ‘This he named a sphero-

annular condenser. In considering the obliquity required to have

all of the light going to the object of an angle to fall outside the

aperture of the objective, it seems to Shadboldt highly desirable that

each objective to be used in dark-field work should have its own

special condenser. That he understood as perfectly as we the

possibility of using a single condenser for all objectives is shown by

the following quotation, p. 157, “It is highly desirable that the

MODERN DARK-FIELD MICROSCOPY 133

condenser should be constructed specially with reference to the

aperture of the object-glass with which it is intended to operate;

and for a reason to be given immediately, it will be seen that cutting

off some of the rays, in order to make a condenser work with objec-

tives of very much larger aperture, although quite practicable and

even generally in use with the parabolic condenser, is not nearly so

advantageous as the use of a separate condenser for every object-

glass . . . of high power at least.”

In 1856 Mr. Wenham himself advocates the use of a truncated

paraboloid of solid glass with a central stop to cut out all the central

rays which would not be internally reflected from the upper surface

of the paraboloid. He brings out in the clearest manner possible

the need of using immersion contact with the paraboloid to permit

the very oblique rays to pass out of the paraboloid into the overlying

substance. If the object is in water, then water immersion and

when the object is mounted in balsam, he advocates the use of an

immersion liquid between the glass slide and the paraboloid of cam-

phine, turpentine or oil of cloves as their refractive index is nearly

the same as crown glass and permits the passage of the rays of great

aperture to pass on into the slide and the balsam containing the

objects. We now use cedar oil or other homogeneous liquid for the

same purpose.

In 1877 Dr. James Edmunds presented before the Quekett

Microscopical Club a paper on ‘‘A New Immersion Paraboloid

Illuminator.” It consisted of a paraboloid of glass cut off at an

exactly calculated distance below the focus, this distance varying

in the four lenses which constituted his set, and the plane top being

made optically continuous, and as nearly as possible optically homo-

geneous with the substance of the slide, by means of a cementing

fluid of high refractive index, such as anhydrous glycerine, castor

oil, copaiba-balsam, oil of cloves, etc. The paraboloid lenses acted

on the principle of total internal reflection, and each one was cal-

culated for the thickness of the slide beneath which it was to be used

(1/16th in 1/100 inch slides) so as to converge upon the object all

of the light entering the base of the paraboloid. Parallel light should

be thrown into the base of the paraboloid, and the most splendid

effects were obtained. by means of direct sunlight. Water immersion

objectives of 1/16th and 1/8th inch focus were used. After speaking

of some test objects he says, p. 19: “With bacterial fluids, the effect

134 SIMON HENRY GAGE

was equally remarkable. Saliva, blood, etc., viewed by a good dry

quarter of about 95° (NA 74), were seen almost as new objects when

lighted up by this paraboloid.”

As it was recognized from the time of Reade that to gain the

dark-field effect the light going to the object must be of an obliquity

so great that it could not enter the microscope directly; this in-

volved either a paraboloid or other dark-field illuminator of such

great range that it might be used with all objectives, or the suggestion

of Shadboldt must be followed that each objective have a paraboloid

especially constructed to give it the best possible effect. This ques-

tion naturally became very insistant when the water immersion

objectives of large aperture came into use, and especially when the

homogeneous immersion objectives came into common use (1880-

1890). It has finally been settled by adopting the first possibility,

viz., the use of dark-field illuminators adapted for all objectives, the

aperture of the objectives being reduced, where too great, to a point

somewhat below 1.00 NA. This makes it possible to utilize a ring

of light between 1.00 and 1.52 NA for the dark-field illumination, and

this ring of light produced by the sun or the electric light has been

found sufficient for practically all dark-field microscopy. It should

be stated in passing that the ring of light produced by the dark-field

illuminators usually falls between 1.00 NA, and 1.45 NA. Some fall

below 1.00 NA and some only go to 1.30 or 1.35. The reducing

diaphragms for homogeneous immersion objectives which have come

to the writer with objectives have ranged from 0.40 NA to 0.80 NA.

From 1907-1910 papers were written describing and figuring

reflecting condensers made on the cardioid principle to take the

place of the truncated paraboloid in dark-field work. The effort

was made to so figure the component segments of glass that the

spherical and chromatic errors would be largely eliminated, and that

the entire ring of light could be brought to a more perfect focus than

is possible with the truncated paraboloid: that is, to be optically

more like A than like B or C in Fig. 16. A simple plate form for

use on the top of the stage has also been devised. When this is

used the substage condenser is turned out so that the light can pass

directly up from the plane mirror to the condenser. This form is not

easy to keep accurately centered. From the writer’s experience

with quite a variety of these dark-field condensers in biological work

MODERN DARK-FIELD MICROSCOPY 135

the paraboloids have proved the easiest to work with and the most

generally satisfactory.

As a final word,—now that the means have been found for fuller

microscopic revelations, it behooves biologists to make the most of

them; and in the study of the finest details in living things by this

dark-field lighting, perhaps a truer conception of structure and action

can be gained than by a too exclusive dependence on dead material

treated with the endless variety of fixers and stains.

Fig. 18. Chalet microscope lamp for bright-field microscopy (Two-fifteenths

natural size).

The lamp has two daylight-glass windows under the overhanging roof. The

roof serves to shade the eyes. The source of light is a 100 watt Mazda C lamp bulb,

the filament of which is centered with the windows.

BIBLIOGRAPHY

AKEHUuRST, S. C.

1914. Substage illumination by hollow cones. Jour. Quekett Micr. Club,

Vol. XII, (1914) pp. 301-308. 3 pl.

Bauscu, Edward

1890. The full utilization of the capacity of the microscope and means for

obtaining the same. Proc. Amer. Soc. Microscopists, Vol. XII, 1890,

pp. 43-49.

Among other matters Mr. Bausch gives a very thoughtful discussion of

the effect of the cover-glass and of tube length.

BORELLUS, PETRUS

1655. De vero Telescopii inventore, cum brevi omnium conspiciliorum historia.

Ubi de eorum confectione, ac usu, seu de effectibus agitur, novaque

quaedam circa ca proponuntur. Accessit etiam centuria observationum

microcospicarum. Authore Petro Borello, regis christianissimi con-

ciliario, et medico ordinario. Hagae-Comitum, ex typographia Adriani

Vlacq, MDCLYV (1655).

136 SIMON HENRY GAGE

Evidence from those with personal knowledge that telescopes and micro-

scopes were made by the Dutch spectacle makers, Zacharias Jansen,

and Hans Laprey, 1590.

CARPENTER, WILLIAM B.

1856. The Microscope and its Revelations. First edition 1856.

An admirable statement of dark-field microscopy is given with the appara-

tus devised up to that time for effecting it. Showing how greatly dark-

field microscopy had been discarded in England one can compare the

first and the 6th (1856-1881) editions of this work with the 8th edition,

(1901).

Cxamot, EmMire Monnin

1915. Elementary Chemical Microscopy. New York, 1915.

This work is recommended not only for the account given of dark-field

microscopy and its application, but for the ultra-microscope, the polari-

scope, the micro-spectroscope and indeed all other chemico-physical

apparatus used with the microscope, and their application in chemical

and physical investigations.

CHEVREUL, M. E.

1838. De la Loi du Contraste simultané des ‘Couleurs et de l’assortiment des

objets colorés considéré d’apres cette loi. Paris, 1839. Work written

1835-1838. Third English edition, 1890. Part of Bohn’s Scientific

Library.

Comstock, J. H.

1912. The Spider Book. A manual for the study of spiders and their near

relatives, the scorpions, pseudoscorpions, whip-scorpions, harvestmen,

and other members of the class Arachnida, found in America north of

Mexico; with analytical keys for their classification and popular accounts

of their habits. New York.

In this book are given pictures of the spider webs photographed against

a black space, i.e., a deep box lined with black velvet. See p.181. The

first photographs made in this way were taken in 1901. They were

exhibited before the Entomological Society of America at its first

meeting, Dec. 28, 1906.

Conrapy, A. E.

1912. Resolution with dark-ground illumination. Jour. Quekett Micr. Club,

Vol. 11, (1912) pp. 475-480.

He says: “To get the utmost resolving power with dark-ground illumina-

tion, the condenser must have not less than three times the NA of the

objective. If the condenser has less than three times the aperture of

the objective then the limit of resolution is found by taking 144 the

sum of the apertures of objective and condenser: e.g., if cond. has

NA of 1.40, and of obj. 1.00 NA, their sum is 2.40, 14 of 2.40=0.60 NA;

‘limit in this case.”

Cox, Hon. Jacos D.

1884. Robert B. Tolles and the angular aperture question. Proc. Amer. Soc.

Microscopists, Vol. VI, (1884) pp. 5-39.

MODERN DARK-FIELD MICROSCOPY 137

This very able address, one of the ablest our society ever had the fortune

to hear from its president, brings out with absolute clearness and fair-

ness the steps in progress and the role played by Robert B. Tolles in

actually making possible the final step, and taking that step, in his

homogeneous immersion objectives. That is not all, he published the

formula by which the objectives were made. The reading of this

address is most strongly recommended to our younger members.

Descartes (LAT. CARTESIUS), RENE

1637. Ocuvres, Publiées par C. Adam et P. Tannery sous les auspices du ministére

de V’instruction publique, Vols. I-XII. Paris, 1902.

The Dioptrique is in Vol. 6 of this edition, and the French and the figures

are as in the original of 1637. In Cousin’s edition the figures are often

considerably modified and the French modernized.

Do.ionp, JoHN

An account of some experiments concerning the different refrangibility

of light. Read June 8, 1758. Philos. Trans. Roy Soc. Lond. 1758,

pp. 733-743. This is the original paper on achromatic telescopes, etc.

EDMUNDS, JAMES

1877. On a new immersion paraboloid illuminator. Jour. Quekett Micr. Club,

Vol. V, (1877) pp. 17-21. Monthly Micr. Jour., Vol. XVIII, 1877,

pp. 78-85.

The Paraboloid was made optically continuous and as nearly as possible,

optically homogeneous with the slide by the use of anhydrous glycerin,

castor oil, copaiba-balsam or oil of cloves. He says that saliva, blood,

and bacterial fluids gave remarkable effects, and were almost like new

objects when seen with this paraboloid.

GacE, S. H.

1917. The Microscope, an introduction to microscopic methods and to histology.

12th revised edition, Ithaca. 1917.

GacE, S. H. and H. P.

1914. Optic Projection. Principles, installation and use of the magic lantern,

the projection microscope, etc. Ithaca, 1914.

GacE, S. H.

1887. I. Microscopical tube-length and the parts included in it by the various

opticians of the world. II. The thickness of cover-glass for which

unadjustable objectives are corrected. Proc. Amer. Soc. Microscopists,

Vol. IX, 1887, pp. 168-172.

This paper gave the information that has led to greater uniformity.

GarpuKoy, N.

1910. Dunkelfeldbeleuchtung und Ultramikroskopie in der Biologie und in der

Medizin. 5 plates, 81 pages. Jena, 1910.

There is a bibliography of books and papers covering 9 pages (202 titles).

Gorpon, J. W.

1907. The top-stop for developing latent power of the microscope. Jour. Roy.

Micr. Soc., 1907, pp. 1-13. See also Wright, pp. 216-217.

The plan is to cut out all of the central beam by a stop at the eye point

instead of by opaqueing the central part of the objective.

138 SIMON HENRY GAGE

GorRING AND PRITCHARD

1837. Micrographia, containing practical essays on reflecting, solar, oxy-hydro-

gen gas microscopes, micrometers, eye-pieces, etc. 231 p. Many

figures in the text, one plate. Whittaker & Co., Ave-Maria-Lane,

London, England. 1837. Rev. J. B. Reade on dark-field, pp. 227-231.

HALL, JOHN CHARLES

1856. On an easy method of viewing certain of the Diatomaceae. Quart.

Jour. Micr. Sci., Vol. IV, (1856) pp. 205-208.

In this paper Dr. Hall figures natural size, the “spotted lens” of that time,

i.e., a very thick, more than hemisphere of glass with the central part

opaqued. (See Quekett, 3d. ed., p. 135 where it is said that it is the

invention of Thomas Ross.) Hall used this spot lens for oblique light

with the ordinary bright field microscopy. He expresses astonishment

that this instrument, designed to give dark-field effects, should give

bright ones. He did not consider the fact that the aperture of this

spot lens was insufficient to throw all the light outside of the aperture

of the objective. One would get the same effect if a wide-angled homo-

geneous immersion were used with a paraboloid, and no reducing dia-

phragm were put into the objective.

HEIMSTADT, OSKAR

1907. Neuerungen an Spiegelkondensoren (Aus der optischen Werkstatte von

C. Reichert in Wien). Zeit wiss. Mikr., Bd. XXIV, (1907) pp. 233-242.

HEIMSTADT, OSKAR

1908. Spiegelkondensor und Paraboloid. Zeit. wiss. Mikr., Bd. XXV, (1908)

pp. 188-195. Erwiderung an Herrn O. Heimstidt, by Siedentopf,

pp. 195-199.

Dr. Heimstidt objects to some of Dr. Siedentopf’s statements in his paper,

“Die Vorgeschichte der Spiegelkondensor.”’ Perhaps the spirit of the

polemic will best be brought out by a quotation from Heimstadt, p. 188.

“Vol allem beeintrachtigt es den Wert und auch die Neuheit dieser

Dunkelfeldbeleuchtung nicht im geringsten, dass dabei langst ver-

gessene Methoden ilterer englische Optiker wieder verwendt wurden.”

In a word, it is well brought out in these papers where the fundamental

ideas came from.

IGNATOWSKY, W. V.

1908. Ein neuer Spiegel-kondensor. Zeit. wiss. Mikr., Bd. XXV, (1908) pp.

64-67 with figures of the substage and the plateform. See also Jentzsch,

and Siedentopf. Jour. Roy. Micr. Soc., London, 1911, pp. 50-55.

Jentzscu, Dr. FELIx

1911. The reflecting concentric condenser. Physikalische Zeitschrift, Bd. XI,

pp. 993-1000. See also Ignatowsky and Siedentopf, Jour. Roy. Micro.

Soc., 1911, pp. 50-55.

KEPLER, JOHANNES

1604. Opera Omnia, Vol. II. Ad Vitellionem Paralipomena. (De modo visionis

et humorum oculi usu.) 1604, pp. 226-229. 11 figs.

Correct dioptrics of the eye here given, and also the explanation of the

effect of convex and concave spectacles.

MODERN DARK-FIELD MICROSCOPY 139

1611. Dioptrica.—Demonstratio eorum quae visui et visibilibus propter conspicilla

non ita pridem inventa accidunt, pp. 519-567. 35 figs., 1611.

The amplifier, real images, and erect images. The Keplerian microscope

(Modern microscope.)

LIsTER, JOSEPH JACKSON

1830. On some properties in achromatic object-glasses applicable to the improve-

ment of the microscope. Philos. Trans. Royal Society London, Vol.

120 (1830) pp. 187-200.

On p. 191 he discusses the effect of a ‘Stop behind the object-glass’’

(retro-objective stop) by which only the outer zone of the objective is

used, the central zone being stopped out. See Wenham, 1854.

Marey, ETIENNE JULES

1901. The history of Chronophotography. Annual Report of the Smithsonian

Institution for 1901, pp. 317-340.

On p. 320 Marey refers to Chevreul’s method of obtaining perfect black-

ness. .

MayALL, Joun, Jun.

1885. Cantor Lectures on the Microscope. Lectures delivered before the Royal

Society of Arts, Nov. Dec. 1885.

On pp. 95-96 are given the facts regarding the working out and production

of homogeneous immersion objectives. Tolles is given due credit.

Moorg, Dr. V. A.

1897. The Hemospast, a new and convenient instrument for drawing blood for

microscopic examination. Trans. Amer. Micr. Soc., Vol. XIX (1897)

pp. 186-188.

After using this “spring needle lancet” individually and with large classes

for many years I quite agree with Dr. Moore when he remarked to me

the other day, “It is the most humane instrument I have ever seen for

drawing blood.” I would like to add to this: And one of the most

efficient.

QUEKETT, JoHN

1848. A practical treatise on the use of the microscope including the different

methods of preparing and examining animal, vegetable and mineral

structures.

First edition, 1848. Reade’s method given and illustrated pp. 178-179;

Second edition, 1852, Reade’s method illustrated pp. 194-195. Third

edition, 1855, Reade’s method, the method of Wenham, Spot-Lens method

of Thomas Ross, the methods of Schadboldt and Nobert are all given.

READE, REV. J. B.

1837. On a new method of illuminating microscopic objects, pp. 227-231 of

Goring and Pritchard’s Micrographia, which see. (1837).

Rocers, Ww. A.

1880. On Tolles’ interior illuminator for opaque objects. (With note by R. B.

Tolles). Jour. Roy. Micr. Soc. London., Vol. IIT (1880) pp. 754-758.

In this paper Rogers gives the history of the devices for making the objec-

tive its own condenser by introducing light into its side and reflecting

the light down upon the object.

140 SIMON HENRY GAGE

SHADBOLT, GEORGE

1851. Observations upon oblique illumination; with a description of the author’s

Sphaero-annular condenser. Trans. of the Micr. Soc. of London.

Vol. III, pp. 132, 154.

This paper was read in 1851. As this condenser is like the glass para-

boloids now used for dark-field work, they are often called the Wen-

ham-Shadbolt paraboloids. Shadboldt discusses prisms in this volume.

SIEDENTOPF, H.

1907. Paraboloid-Kondensor, eine neue Methode fiir Dunkelfeldbeleuchtung

zur Sichtbarmachung und zur Moment-Mikrophotographie lebender

Bakterien, etc. Zeit. wiss. Mikr., Bd. XXIV, (1907) pp. 104-108.

1907. Die Vorgeschichte der Spiegelkondensoren. Zeit. wiss. Mikr., Vol. XXIV

(1907) pp. 382-395. 16 figures are given of early forms.

1908. Mikroskopische Beobachtungen bein Dunkelfeldbeleuchtung. (Mitteil-

ung aus der optischen Werkstitte von C. Zeiss, Jena) Zeit. wiss. Mikr.

Bd. XXV (1908), pp. 273-282. Two plates of photomicrographs of the

rays above the different condensers. See also under Heimstiadt.

1910. Cardioid-Condenser. Jour. Roy. Micr. Soc. Lond., 1910, pp. 515. See

also Ignatowsky, and Jentzsch, Jour. Roy. Micr. Soc. Lond., 1911, pp.

50-55, where will be found a statement concerning the historical relation

of these different condensers.

STEPHENSON, J. W.

1879. A catoptric, immersion illuminator. Jour. Roy. Micr. Soc. Lond., Vol.

II (1879) pp. 36-37.

This condenser does not depend on internal reflection, but by a silvered

surface around the central part. According to Siedentopf this is the

condenser copied by Reichert; and according to Heimstidt Wenham’s

truncated paraboloid was copied by Zeiss (See under Heimstadt).

WeEnzHAM, F. H.

1850. On the illumination of transparent microscopic objects on a new principle.

Trans. Micr. Soc. Lond., Vol. III. (1850) pp. 83-90.

This is the paper by Wenham in which dark-field illumination is produced

by a hollow silvered parabolic speculum.

1854. On the theory of the illumination of objects under the microscope with

relation to the aperture of the object-glass, and properties of light;

with practical methods for special differences of texture and colour.

Quart. Jour. Micr. Sci. Vol. IL (1854) pp. 145-158.

In this paper Wenham refers to the method of Lister (1830) for darkening

the central zone of the objective so that no light can enter the outer

zone, unless, as Wenham says, it is “radiated” from the object (See his

fig. 1, and pp. 149-150 of the article). On p. 153, in the reference to the

effect that his paper of 1850 had had in the microscopical world he says,

“As proof of the utility and correctness of my theory, I have only to

mention the many applications of it that have since that time (between

1850 and 1854) come into general use, in the way of adapting central

stops to the achromaatic condenser, single (ie., “spot lenses”) and

compound lenses, etc.”

MODERN DARK-FIELD MICROSCOPY 141

1856. On a method of illuminating opaque objects under the highest powers

of the microscope. Trans. Micr. Soc. Lond. in Quart. Jour. of Micr.

Sci., Vol. IV, (1856) pp. 55-60.

It is in this paper that Mr. Wenham insists on making homogeneous con-

tact with the slide and the top of the paraboloid. It will be noticed that

in this paper he speaks of Opaque Objects, while in the paper of 1850

he speaks of Transparent Objects. By reading the two papers it will

be seen that many of the objects mentioned in the two papers are

identical. This gains an explanation from the fact that he has ap-

parently given up the notion that the objects were visible by their

own “radiated” light, but by the light they reflect to the microscope.

Consequently he represents (Fig. 4) the light from the condenser going

to the cover-glass and being reflected from it down upon the object

and he says that it makes the most perfect kind of a Lieberkuhn reflector.

One can see instantly that when homogeneous immersion objectives are

used there can be no total reflection from the cover.

Woop, RoBert W.

1911. Physical Optics. New and Revised Edition, 1911.

On p. 373, he discusses, ‘‘Penetration of the disturbance into the second

medium,” and shows that going back to the time of Newton and Fresnel,

it was known that while there was total reflection, the light seemed to

pass for a minute distance into the rarer medium. This explains why

one may get a brighter dark-field picture than is expected if objects are

in optical contact with the slide.

WRIGHT, Sir A. E.

1907. Principles of Microscopy, being a handbook to the microscope. London

and New York, 1907.

The writer has found this book the best and most thought-provoking of

any that has been published on the microscope during the last 50 years.

A NEW BLADDER FLUKE FROM THE FROG*

Joun E. GUBERLET

Bladder flukes have been reported a number of times from North

American frogs but as yet very little work has been done on these

forms in thiscountry. The European species, however, have received

more attention and their complete life histories have been worked out.

In North America the studies on frog bladder flukes have been

carried on by only four authors, namely Leidy (1851), Bensley (1897),

Stafford (1902, 1905), and Cort (1912). The localities from which

these were reported are Toronto, Canada; Rice Lake, Ontario,

Canada; Urbana, Illinois; Bemidji, Minnesota; and North Judson,

Indiana. The writer has at hand another species of frog bladder

fluke from Rana catesbiana taken at Stillwater, Oklahoma.

In view of the fact that Cort (1912) has given a thorough review

of the literature as well as a discussion of the nomenclature of this

group, it is unnecessary to take up the history of the literature any

farther at this time. The frog bladder distomes have been grouped

into two genera by Loossand called Gorgodera (1899) and Gorgoderina

(1902). The basis for this classification is on the number of testes

which these animals have. The genus Gorgodera has nine testes

while Gorgoderina has two. Of the latter genus there are known from

North America three species, namely Gorgoderina simplex Looss,

G. translucida Stafford and G. attenuata Stafford. Of the former

genus there have been two species described, namely, Gorgodera

amplicava Looss and G. minima Cort. The writer adds another

species to the genus Gorgodera.

GORGODERA CIRCAVA NOV. SP.

In the summer of 1918 the writer found in the urinary bladder

of a large bull frog (Rana catesbiana) twenty trematodes (Figs. 1 and

2) which belong to a new species of the genus Gorgodera. In the

early part of the summer of 1919 another bull frog yielded two speci-

mens of the same species of trematode. These forms were so firmly

attached to the wall of the urinary bladder by means of the acetabu-

*Contribution from the Parasitology Laboratory of the Oklahoma Agricultural

Experiment Station, Stillwater, Oklahoma.

142

A NEW BLADDER FLUKE FROM THE FROG 143

lum that it was necessary to tear the bladder apart in order to make

the worms release their hold. The worms were killed by dashing

hot corrosive acetic over them when they were well extended. In

this way they were only very slightly contracted when killed.

It was thought at first that this form belonged to the species

Gorgodera amplicava Looss. Unfortunately, specimens of this

species could not be obtained for comparison. From a study of the

descriptions of G. amplicava in the literature on bladder flukes it

was concluded that the species were not the same. The only other

species of this genus known in North America is Gorgodera minima

Cort. That species is much smaller than the one to be described here.

The European forms are all much larger than any of the American

species of Gorgodera.

This species of distome is similar in activity and habit to the

others of this genus. The anterior portion of the body is very active

and moves about freely while the posterior region is less active but

not sluggish. The cuticle of the anterior part of the body is marked

with minute longitudinal striations. These markings extend to or

slightly beyond the acetabulum. The part of the body which is

anterior to the acetabulum is cylindrical but becomes flattened near

the acetabulum while the posterior portion is somewhat flattened

and rather opaque. The opacity extends from the posterior end

forward to the region of the ovary. That portion of the body occu-

pied by the ovary, vitellaria and acetabulum is fairly transparent.

The length of the animal varies from 2.5 to 3.75 mm. with a

width of .5 to .65 mm. in the region posterior to the ventral sucker.

This form appears to be considerably smaller than Gorgodera ampli-

cava which has a length of 3 to 5 mm., and larger than G. minima,

that form being 1to2mm.inlength. The individuals which measure

2.5 mm. in length have large numbers of eggs in the uterus while in

those of the larger size this organ is entirely filled throughout giving

it the appearance of being a mere egg sac. In individuals which are

less than 2.5 mm. in length no eggs are developed.

The ventral sucker ranges from .60 to .75 mm. in diameter and

is surrounded by a distinct circular sheath 0.05 to 0.135 mm. in

width (Figs. 1 and 2, vss). This circular sheath around the acetabu-

lum is very marked and is a rather distinct characteristic in this

form. Therefore, I wish to propose the name Gorgodera circava for

this species.

144 JOHN E. GUBERLET

The sheath around the sucker forms a distinct space or cavity

between the wall of the sucker and structures of the body (Fig. 7).

Small muscle bands (Fig. 7, mb) bind the tissues of the body to the

ventral edge of the sucker. There are also a few muscle bands and

connective tissue fibers extending across the cavity which connect

the sucker with the internal parts of the body. From the external

appearance of a normal animal the sheath is only slightly apparent

from a side view and appears only as a slight bulge around the sucker.

In an animal with both ends curved ventrally the sheath forms a

distinct fold around the acetabulum (Fig. 4). The ventral sucker

with the circular sheath produces a structure from .65 to .8 mm. in

diameter, which is somewhat broader than the greatest breadth

posterior to the sucker. The oral sucker has a diameter ranging

from .30 to .37 mm. with an average of .33 mm. for ten specimens.

The ratio of the oral sucker to the ventral sucker ranges from 1.8:1

to 2.3:1 with an average for ten specimens of 2.1:1. As stated by

Cort (1912:162) the acetabulum of G. amplicava is 2.5 to 3 times the

size of the oral sucker. Therefore, G. circava is different in this

respect.

The mouth is situated in the oral sucker and appears as a tri-

angular orifice in the posterior part of the sucker. The esophagus

(Fig. 1, e) is a short narrow tube 0.14 mm. in length and 0.03 mm. in

width. ‘The intestinal ceca (Fig. 1 and 2, i) are about 0.055 mm. in

width and are dorsal extending from the esophagus to within a short

distance of the posterior end of the body. They are widely separated

to give room for the reproductive organs which lie between as well

as ventral to them. ‘The ceca are dorsal and lateral to the testes.

Some folds of the uterus pass to the lateral margins of the body and

lie outside the ceca.

The reproductive system of Gorgodera circava is similar to that

of the other species of this genus. The principal differences lie in

the relative size and shape of parts, such as the number of vitellaria,

shape of ovary, seminal vesicle and ejaculatory duct. There are

nine testes, five on the same side with the ovary and four on the

other. They are irregular in shape and the anterior ones are some-

what larger than those posterior. The shapes and sizes of the

individual testes vary in different individuals but in general those

which are anterior are proportionately broader than those posterior.

With one exception the testes range about 0.23 mm. in length, 0.14

A NEW BLADDER FLUKE FROM THE FROG 145

to 0.17 mm. in breadth and 0.22 mm. in thickness. The testis which

is most posterior is usually much smaller than the others, measuring

about 0.17 mm. in length by 0.12 mm. in breadth and 0.21 mm.

thickness. The testes on either side are connected by minute

tubules. From the dorso-anterior edge of the anterior testis on each

side arises the vasa efferentia (Fig. 5, ve). These tubules extend

anteriorly and unite in the region of the vitellaria to form the vas

deferens which passes forward to the vesicula seminalis (Fig. 3, ves).

The vesicula seminalis is a large pyriform sac dorsal to the anterior

edge of the ventral sucker. It has a length of 0.15 to 0.2 mm.,

breadth of 0.14 mm. and thickness of about 0.15 mm. The shape

and size is somewhat modified according to the degree of expansion

or contraction of the worm. The vesicula seminalis is entirely filled

with sperm cells. From the dorso-anterior edge of the vesicula

seminalis the ejaculatory duct (Fig. 3, ed) arises and curves ventrad

for some distance and then extends forward to the common genital

pore (Fig. 3,g). This duct has a total length of 0.16 mm. and in the

proximal region has a diameter of 0.015 mm. Around the distal

portion of the duct are grouped the prostate glands (Fig. 3, p), a group

of unicellular gland cells. In this region the ejaculatory duct is

much enlarged forming a large pouch (Fig. 3, ep), or lumen in the

midst of the prostate gland. This pouch or enlargement of the

duct is 0.07 mm. in length and 0.05 mm. in diameter. The ejacula-

tory pouch as well as the duct is filled with sperms.

The vitellaria (Fig. 2, v) are immediately posterior to the ventral

sucker and anterior to the ovary. They are made up of two groups

of six to eight follicles each. One group lies toward each side of the

animal and they are connected by a transverse vitelline duct. This

duct becomes enlarged to form the vitelline reservoir in the median

line of the body (Fig. 6, vr). From the dorsal surface of the vitelline

reservoir arises a small median vitelline duct (Fig. 5, vd) which passes

dorsal into Mehlis’ gland where it unites with the ootype.

The ovary is a distinct three-lobed structure 0.27 mm. in length,

0.24 mm. in breadth, and 0.21 mm. in thickness. This organ lies

toward the ventral side of the body. It may occur on either the

right or left side as about half of the specimens studied showed it

on one side and the other half on the other. The oviduct arises

from the dorsal surface of the ovary as a funnel-shaped structure

with the broad part of the funnel attached to the ovary. It extends

146 JOHN E. GUBERLET

dorsad for some distance as it becomes narrow and then curves

laterally or anteriorly, after which it enlarges immediately into the

fertilization space (Figs. 5 and 6, f). It then becomes narrow again

and passes forward near the dorsal surface of the animal to Mehlis’

gland (Figs. 5 and 6, m) where it changes into the ootype. Mehlis’

gland is a small group of unicellular gland cells located between the

posterior edges of the vitellaria and dorsal to the transverse vitelline

duct. Laurer’s canal (Fig. 5 and 6, 1) passes from the proximal

region of the oviduct between the fertilization space and the ootype

and makes a slight lateral curve. It then goes anteriorly and dorsally

to the point where it opens on the dorsal surface of the body either

dorsal or lateral to the ovary.

In passing from the ootype the uterus curves ventrad and bends

back on itself (Fig. 5 and 6, u) in the median line of the body and

goes posteriorly between the testes and finally reaches the posterior

extremity of the body, where it fills with its numerous coils the

region of the body posterior to the ovary and testes. The coils of

the uterus become filled with eggs. Small masses of sperm cells

are scattered throughout the coils of the uterus. The uterus finally

emerges from the mass of coils in the region of the anterior testes

(Fig. 2) and extends forward ventral to the ovary and vitellaria,

passes dorsal to the ventral sucker and ventral, or slightly lateral to

the vesicula seminalis to the genital pore (Fig. 3).

The eggs of Gorgodera circava increase in size as they develop

and pass from the ootype to the genital pore as in other species of

the bladder flukes. In this case only the eggs in preserved specimens

have been studied and no doubt there has been some shrinkage

through the process of preservation. The eggs at the ootype measure

about 0.016 mm. in length by 0.013 mm. in breadth; at the posterior

end in the coils of the uterus 0.025 mm. in length by 0.019 mm. in

breadth; while at or near the genital pore where they contain fully

developed embryos, about 0.030 mm. in length by 0.023 mm. in

breadth.

The chief differences between the American species of Gorgodera

lies in the size and shape of the animals; the structure, size and ratio

in sizes of suckers; and the shape and relationship of the reproductive

organs. Gorgedera minima, described by Cort (1912) is the smallest

of the three species, it being 1 to 2 mm. in length and its acetabulum

is 1.6 to 2 times the size of the oral sucker. Gorgodera amplicava

A NEW BLADDER FLUKE FROM THE FROG 147

first described in this country by Bensley (1897), and reviewed by

Stafford (1902), and again compared with Gorgodera minima by Cort

(1912), is considerably larger being 3 to 5 mm. in length and its

acetabulum is 2.5 to 3 times the size of the oral sucker. Gorgodera

circava is 2.5 to 3.75 mm. in length and the acetabulum ranges from

1.8 to 2.3 times the size of the oral sucker. The acetabulum is also

surrounded by a distinct circular sheath which is a distinctive char-

acteristic of this species. In Gorgodera circava the vitellaria are

composed of six to eight follicles in each group while Gorgodera

amplicava has eight to ten in each and Gorgodera minima has nine to

eleven. The ovary of Gorgodera circava is a distinct three-lobed

structure while in G. minima it is only slightly lobed and in G. ampli-

cava it has three to five irregular lobes with smaller or secondary

lobes. The presence of the ejaculatory pouch in Gorgodera circava

is another structure not found in either of the other species. The

differences in the reproductive organs and the presence of the circular

sheath around the acetabulum clearly sets Gorgodera circava off from

the other species.

LITERATURE CITED

BENSLEY, R. R.

1897. Two forms of Distomum cygnoides. Centr. f. Bakt., u. Infekt., 21:326-331.

Cort, W. W.

1912. North American frog bladder flukes. Trans. Amer. Mic. Soc., 31:151-166.

Lery, J.

185i. Contributions to Helminthology. Proc. Acad. Nat. Sci. Phila., 5:205-209.

Looss, A.

1899. Weitere Beitrige zur Kenntniss der Trematoden-fauna Aegyptens.

Zool. Jahrb., Syst., 12:521-784.

1902. Ueber neue und bekannte Trematoden aus Seeschildkréten. Zool.

Jahrb., Syst., 16:411-794.

STAFFORD, J.

1902. The American Representatives of Distomum cygnoides. Zool. Jahrb.,

Syst., 17:411-424.

1905. Trematodes from Canadian vertebrates. Zool. Anz., 28:681-694.

EXPLANATION OF PLATE XIII

All drawings made with the aid of camera lucida.

Fig. 1. Dorsal view of Gorgodera circava, X35.

Fig. 2. Ventral view of Gorgodera circava, X35.

Fig. 3. Reconstruction from sagittal sections showing ends of reproductive organs

and genital pore, X130.

Fig. 4. Outline drawing of small specimen which is bent ventrally at both ends

causing the acetabular sheath to form fold around sucker, X35.

148

JOHN E. GUBERLET

Fig. 5. Reconstruction of female genital organs from sagittal sections, X120.

Fig. 6. Reconstruction of female genital organs from frontal sections as seen from

dorsal view, X120.

Fig. 7. Sagittal section through ventral sucker to show ventral sucker sheath, X35.

e esophagus

ed ejaculatory duct

ep ejaculatory pouch

ex excretory pore

f fertilization space

g genital pore

7 intestinal ceca

1 Laurer’s canal

m Meblis’ gland

b muscle bands

0 ovary

os oral sucker

zs.

Abbreviations

oviduct

prostate gland

uterus

vitellaria

vas deferens

vasa efferentia

vesicula seminalis

median vitelline duct

vitelline reservoir

ventral sucker

ventral sucker sheath

testes

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LABELING ILLUSTRATIONS

Z. P. METCALF

North Carolina State College and Experiment Station

Sometimes really good illustrations are spoiled by faulty or poorly

made labels and not infrequently biological illustrators do not do

sufficient labeling to make their illustrations clear. The thought is

frequently expressed by good draughtsmen that labeling is difficult

and is therefore to be avoided, or they say they wish that they had

been born with the ability to letter drawings properly thus expressing

clearly that in their own opinions their drawings are not labeled as

they should be. With these thoughts in mind it seems not amiss to

outline the following methods that may be used for labeling biological

illustrations.

It frequently becomes necessary to indicate separate parts on an

illustration, hence it becomes necessary to label the drawing. This

may be done in a variety of ways. If the person making the illustra-

tion has enough ability he may make the lettering free handed.

Another method is to select letters or figures from printed matter.

These may then be cut out and pasted on the illustration in the

proper place. Still another method is to buy the cut-out letters and

figures and paste these on the illustration. The sets desired may be

written on the typewriter and cut out and pasted on the illustration.

Labels may be printed directly on the illustration. And lastly guide

lines may be drawn to the margins of the illustrations where they

may be connected with type set up in the ordinary way when the

illustration is printed. Regardless of the method of labeling selected

care should be exercised to make the labels as neatly and accurately

as possible. Care should also be exercised to see that the lettering

is sufficiently large to stand the necessary reduction if the illustration

is reduced.

The labels of biological illustrations are generally indicated in

the following manner: (1) by Arabic numerals, (2) by the initial

letters of the name of the parts of the object labeled, (3) by abbrevia-

tion of the names of the parts of the object labeled, (4) by a sequence

of letters, and (5) by the full names of the parts of the object. In

the first four methods it is necessary to print an explanation of the

149

150 Z. P. METCALF

labels. The fifth method requires no explanation. The method

selected will depend upon the personal choice of the draughtsman.

There is much to be said in favor of the fifth method provided there

are not too many parts to label or the names are not too complicated.

By this method the attention is not distracted by having to search

through a long list of explanations. If it is preferred to abbreviate

the names, then there is much to be said in favor of the second and

third methods, for the initial letters and the abbreviation will indicate

the real name of the part. If either of these methods is selected the

explanations should be arranged alphabetically, so that it will not be

necessary to look through the entire list to find the explanation for

any abbreviation.

The labels may be placed at the margin of the illustration or

directly on the illustration. The method we select will depend to a

great extent on the nature of the illustration. If the parts are large

enough so that the labels may be placed directly on the part without

obscuring any details this is perhaps the best method. For many

illustrations, however, this is not possible. The labels should then

be placed at the margin. This latter method necessitates the use of

guide lines. Guide lines may be simple straight lines or they may

be brackets. Brackets are used to indicate areas of considerable

extent which could not be indicated by a single line. Instead of the

bracket, two lines drawn at an angle to each other may be used.

The straight lines may be solid, dotted or dashed lines. If a dashed

or dotted line is used care should be taken to get the dots of uniform

size or the dashes of uniform length. As a general proposition guide

lines should be straight and should run parallel to the main margin

of the drawing if possible. On very light drawings the guide lines

should be black. If the guide lines run over alternate dark and

light areas it is advisable to use a double line, one part being black

and the other part white. Care should be exercised to keep guide

lines of uniform thickness. This may readily be accomplished by

using a ruling pen. Ordinary liquid India ink may be used for

black lines and any good grade of Chinese white for white lines.

Care should be taken not to make the lines so thick that they will

look unsightly when the drawing is complete. On the other hand, the

lines should not be so delicate that they will not stand the necessary

reductions.

LABELING ILLUSTRATIONS 151

Cut out letters or figures which are gummed on the back may be

purchased in a wide variety of styles and sizes. These are very

desirable for labeling drawings especially if the correct size and a

suitable style are selected. Care should be taken to see that the

separate letters are pasted in a straight line. This may readily be

accomplished by drawing a faint pencil line to indicate the bottom

of the letters, and then bringing the letters to this line. Cut out

letters have the advantage that they may be pasted directly on the

illustration, and will obscure only a minimum amount of detail.

Labels may be written on the typewriter using a good black

record ribbon. In making labels on the typewriter only a new ribbon

should be used and the type should be thoroughly clean so that

good sharp impressions can be secured. These labels may then be

cut out and pasted on the drawing as recommended above for printed

labels. The chief difficulty with this method is that the labels are

too small unless the illustration is not to be much reduced in repro-

duction. Type written labels may be enlarged by making a negative

from them by any of the enlarging methods, and then printing a

positive from this negative on a smooth gaslight paper. This method

is advocated chiefly where we need a large number of labels of the

same kind.

Labels may be selected from printed matter and pasted on the

drawing. It is necessary to bear in mind the amount of reductions

that will take place in reproducing the drawing. The ordinary book

type is about 9 points, therefore if the drawing is reduced to one third

the original labels should be 28 point. If the drawing is reduced to

one fourth the original label will have to be 36 point. It is usually

difficult to find letters of sufficient variety in these sizes. Special

labels may be set up by the printer but this is usually a very expensive

method. Plate XIV shows the various sizes of printed letters

and may be used to determine the size of letters that it is necessary

to use. Thus if the drawing is reduced to one third labels the size

of 36 point will appear as 12 point or 18 point will appear as 6 point,

etc. The various sized letters on this plate may be traced on thin

tracing paper in India ink and pasted on the drawing.

HAND PRINTED LABELS FROM RUBBER TYPE

Labels may also be printed by hand from rubber type directly on

the illustrations. For printing labels from rubber type we will

152 Z. P. METCALF

need a set of rubber type of the proper size, holder to hold the type,

and a stamp pad filled with faint blue ink. The proper combinations

of letters are set up in the-holder, bearing in mind that the type are

inverted and reversed. The type in the holder are then stamped in

the pad and then on the drawing. The labels are then traced over

with India ink. It is necessary to trace over labels made with a

rubber stamp because the margins are not clear cut. The advantage

of using the pale blue ink is that if the illustration is reproduced by

ordinary photographic processes the blue will not show and need cause

no trouble if slight errors are made. Rubber type may be secured

in a variety of styles and sizes. A neat legible style should be selected

and a size selected so that it will stand the necessary reduction.

Sets of complete alphabets may be purchased or separate stamps

may be produced from dealers in office supplies. The former has

the advantage that any label may be readily set up. The latter is

preferable if many labels of the same kind are to be printed at one

time. The holders for rubber type are usually supplied with the

sets of type. These holders are convenient and since they are

adapted to the size type with which they are supplied they leave little

to be desired. Stamp pads for this purpose should be inked with a

faint blue ink as this color is not very active, photographically it

does not bother the engraver. After the labels have been stamped

with the faint blue ink they must be finished up with black India ink.

This requires a fine pen and a little attention to details, but can be

done with considerable rapidity after a little practice.

HAND PRINTED LABELS FROM METAL TYPE

Labels may also be printed from the regular metal type of the

printer very much as the rubber type is used. To print labels from

metal type we will need some black printers ink, a font of type of

proper size, a holder for the type and a compositor’s roll.

The ink used for hand printed labels of the metal type is the

regular black printers ink. This usually requires thinning to work

properly. Benzine, gasoline or xylol may be used for thinning the

ink. For this purpose the ink is placed on a piece of glass and the

solvent is added drop by drop while the ink is worked with a spatula,

until it is of the proper consistency. Experience soon teaches when

the ink is of the proper consistency. When it is thought that the ink

is properly mixed a small amount of the prepared ink is spread on the

LABELING ILLUSTRATIONS 153

compositor’s roll and the type in the holder is stamped on the roll

and printed on a piece of paper. If the ink is in the proper condition

and has been properly spread on the roll enough will adhere to the

face of the type to make a good label. If the ink is too thin it will

spread when we attempt to printa label. If it is too thick not enough

of it will adhere to the face of the type to print a good label.

Fig. 1. Type holder for printing labels.

b, bur of the stove bolt which is soldered q, quads which are used to fill out the

to the type box. line of type

h, heads of stove bolts used to clamp the _ 5, stove bolt used to clamp the type in

movable plate against the type the line

m, movable plate t, type

The type holder (Fig. 1) used for hand printed labels is a shallow

box made of brass. The height of the box is a little less than the

height of the type. One side of the box is movable and is fastened

to the opposite side by means of four set screws. This is used to

lock the type in rows. One end of the box has a set screw which is

used to lock the type. This holder is used very much as an ordinary

rubber stamp is used. If only one figure or letter is to be printed

in each label it is not necessary to use the type holder, but the individ-

ual types can be held in the hand very readily.

Type is set up in an inverted and reversed position. Each piece

of type has little grooves called the nicks which indicate when the

type is in a proper position. As soon as a line of type is set up a

glance will reveal whether all of the characters are in the proper

position. The characters to be used are picked out from the type

box one at a time and placed in the holder which is held in the left

hand in an inclined position so that the type will lay against the

fixed side of the holder. As soon as the label is completely set up

the rows of type are locked in place by tightening the set screws

154 Zz. P. METCALF

which fasten the movable side and the end. At first the set screws

are only set tight enough to hold the type firmly in place. A soft

wooden block is then placed on the face of the type and the type is

leveled up by pounding with a light hammer. The properly set up

label is firmly pressed on to the inked compositor’s roll and the inked

type is then stamped on the illustration. Care being taken to press

the type down firmly at all points without allowing it to move.

The advantages of hand printed labels are that they are very

neat and accurate and that they may be made by any one without

previous experience. The chief disadvantage is that it is somewhat

laborious to set up the type. But with the large sized type used in

printing labels for illustrations this is not a very large item. It must

be remembered that it requires several hours for the printers ink to

dry and care must be exercised in handling the illustrations or the

labels may be ruined.

FREE-HAND LETTERING

Occasionally it is not possible to letter an illustration by any of

the methods given above, in that case it is necessary to have recourse

to free-hand lettering. Free-hand lettering is a special kind of free-

hand drawing by means of which the draughtsman learns to draw

the design of letters neatly and rapidly. There are no special tricks

of the trade about lettering that cannot be learned by the biologist

who will conscientiously try to master the subject.

The first consideration in free-hand lettering as in other kinds

of free-hand drawing is to get the main proportions. After the main

proportions are secured the details are added. The more important

details are added first then the finer and finer details until the letter-

ing is complete. Just as the individual letters are found to vary one

from the other so in printing a line of lettering it will be found neces-

sary to space the letters carefully with reference to each other, other-

wise the lettering will not have a neat appearance when finished.

In lettering each letter is influenced by the letters on each side of

it so that no general rule can be laid down which will make it possible

to always place a letter the proper distance from its neighbors. The

only rule that can be given is that all letters should seem to have the

same amount of space allotted to them. Obviously this amount of

space will vary with the different letters and with the letters on each

side of it. Thus a capital I requires less space than a capital M.

LABELING ILLUSTRATIONS 155

Then, too, a capital I will require more space if placed between a

capital M and a capital N than if placed between a capital E and a

capital T because these letters have a great amount of free space

whereas the M and N have practically no free space. The letters

in each word must be studied, therefore, in order to determine the

proper spacing. Trials should be made in order to see just what

spacing looks the best. If this is done critically gradually a proper

conception of proper spacing of letters will be acquired.

The usual error made by beginners is to space the letters too far

apart. Letters look better when they are crowded well together.

The letters in any design that is to be treated in free-hand letter-

ing should be sketched in with a pencil complete before finishing up

any of the letters. This is done in order to insure a proper balance

of the words with each other and a proper spacing of the letters. In

sketching letters it is usually advisable to draw two faint lines one

for the top and the other for the bottom of the letters. Sometimes

it is advisable to divide this space by a third line so that certain

letters may be carefully drawn with rapidity. After the base lines

are drawn the letters are sketched in spacing each letter with refer-

ence to the other letters, and indicating at first only the general

outlines of the letters. Corrections should be made until the whole

has a well balanced appearance. Some letters may then be carried

forward and the principal minor details indicated, making any

necessary corrections in the other letters in the line to maintain the

balance. After experience has been gained letters may be sketched

in ink free-hand, especially such forms as the Gothic. And any one

having considerable lettering to do should practice this form of letter-

ing, but the art of lettering is soon lost unless it is used day by day.

After the pencil sketch of a line of lettering is finished the letters

may be finished with India ink on drawings or with black water

color or oil color on paintings. In doing this the borders of the main

letters are finished first and then the borders of the finer details; the

body of the letters being filled in last of all. Care must be taken in

finishing up the borders not to exceed the limits penciled and to keep

all straight lines straight and all curved lines a true curve. In filling

in the body of the letter care must be taken not to allow the color to

run over the borders that have been finished. In finishing large

letters a ruling pen and a straight edge may be used for the longer

156 Z. P. METCALF

straight lines on the borders of the letters but on the smaller letters

and for all fine details a pen must be used free hand.

The idea is prevalent that the forms of letters are fixed but

nothing is farther from the truth. There are certain broad general

styles of letters such as the Roman and Gothic but the variations in

these styles are as many as there are draughtsmen. A few of the

more important styles are discussed below and plates showing stan-

dard letters are given, not with the idea that these should be copied

slavishly but that these designs may be helpful in producing letters

for drawing and may serve to indicate the main styles. All letters

occur in two forms, capitals usually called caps and small letters

called lower case. If there are only a few letters in a group as in the

abbreviated signs used to label parts of a drawing either caps or lower

case or both kinds of letters may be used, but if there is a series of

words it is better to use lower case letter throughout as we are used

to reading words printed in lower case types. Words in lettering

should be well separated so that there is no doubt as to the limits of

the separate words. ‘The rule is that the words should be separated

at least by a space equal to that occupied by the widest letter and

slightly mere space would be better.

The Gothic letter is the simplest letter because it is formed of

lines of a uniform thickness throughout. Gothic letters exist in two

forms, a vertical Gothic and an inclined Gothic. In the vertical

Gothic alphabet the main axis of the letters is vertical and since the

lines are all of a uniform thickness it is a fairly easy alphabet to

letter in a free-hand manner. For convenience of discussion the

letters are divided into the following groups, (1) letters composed of

straight horizontal and vertical lines, (2) letters composed of horizon-

tal or vertical lines with diagonal lines, (3) letters composed of

straight and curved lines and (4) letters composed of curved lines

only. Furthermore it is convenient to define a full bodied letter as

a letter occupying as much horizontal as vertical space. For pur-

poses of analysis the letters are drawn on cross section paper each

letter occupying a vertical distance of five units. The thickness of

the stroke is only 2/3 of a unit.

In the first group of letters we have the capitals E, F, H, I, L and

T and the lower case letters i, ]. In the capital E it will be noted

that the letter is not a full-bodied letter as the foot occupies only

41¢ units and the cap only 4 units. The tongue of the letter occupies

LABELING ILLUSTRATIONS 157

only 21% units and is placed only slightly higher than the middle.

The capital F is identical with the E except that the foot is omitted.

Care should be taken not to extend the cap too much or the letter

will look top-heavy. ‘The capital H occupies about 4 units as other-

wise it looks too broad. The tongue is placed on a level with the

tongue in the E and F, that is slightly above the middle. The capital

I needs no comments as it is simply a straight line with a thickness

of 2/3 of a unit. The foot of the capital L is about 31% units in

length to prolong it makes it appear unwieldy. The full bodied

lower case letter occupies only three-fourths the space allotted to

the full bodied capitals and the width of stroke is only a half unit.

The small lower case letters occupy only two-thirds of the vertical

space occupied by the large lower case letters. Therefore the body

of a small lower case letter like i would occupy only one-half of the

vertical space allotted to a capital letter, hence 244 units the dot

being placed one full unit above the top of the letter. The lower

case 1 would occupy 34 of the vertical space allotted to a capital.

To the second group belong the capitals A, K, M, N, V, W, X,

Y and Z, and the lower case letters k, v, w, x, y and z. The capital

A occupies the full width of five spaces below and slopes to the top

line uniformly on both legs. The top does not end in a sharp point

but in a point that is about one-half unit wide. The tongue of the

A is placed about one and one-half units above the base line. The

capital K is somewhat difficult as it is composed of two diagonal lines

at different angles. The top diagonal is usually placed about three

and one-half spaces from the vertical stroke and at such an angle that

if it were projected the lower border of the diagonal would strike the

base line one full unit to the left of the vertical stroke. The lower

diagonal is placed four units from the vertical stroke and at such

an angle that its top border projected would strike the top of the

vertical stroke. The capital V is simply the capital A inverted and

the tongue omitted. The capital M is simply the V with two vertical

strokes added on each side. Note that these vertical strokes end

in their full width and not reduced as in the case of the top of the A

and the bottom of the V. The capital N consists of two vertical

strokes four units apart connected by a diagonal running from the

top of the left hand stroke to the bottom of the right hand stroke not

the reverse as is frequently seen in lettered signs. The diagonal is

placed at such an angle that the vertical strokes will end in full

158 Z. P. METCALF

width on both the base and top limiting lines. The capital W may

be considered as two V’s contracted to occupy only four spaces each

and united so that the apex of the jointed diagonals shall occupy only

half a unit each. The capital X is simply two diagonals which cross

each other in the center. This letter is therefore a full bodied letter.

The capital Y is composed of two arms which are six units apart

and run at such an angle as to unite two and one-half units from the

base line. The foot of the capital Z is four and one-half units long

and the cap only four units long. The cap and the foot are con-

nected by a diagonal placed at the proper angle. In the lower case

letters the stem of the K occupies the full vertical unit for lower case

letters and the diagonals of the letter bear the same relation to each

other that they do in the capital K but they are reduced to one-half.

The lower case v, w, x and z are the same as the caps except they are

reduced to one-half. The lower case y is the same as the lower case

v with the right diagonal extended below the base line, the full

length allotted to lower case letters.

In the third group we have the capitals B, D, J, P, R and U;

and the lower case letters a, b, d, e, f, g, h, m, n, p, q, r, t, and u.

The capital B may be considered as a capital E with the ends of the

cap and the foot connected to the tongue by arcs of circles. It will

be noted that this makes the top part of the letter somewhat smaller

than the bottom. The capital R may be considered as a capital F

with the cap and tongue connected as in the B and a tail added to

the lower part. The tail of the R should extend beyond the top part

of the letter at least a full unit otherwise the letter will look top-heavy.

The P is similar to the R without a tail but the top part of the P is

made longer by dropping the tongue about one-half unit below its

position in R. A capital D is produced by using a foot and cap

similar to the foot and cap in the capital B and connecting these two

horizontal lines by a regular curve. ‘The capital J and U are similar

to each other save that the J has a single vertical arm and the Ua

double arm. The J is somewhat narrower occupying only three

and one-half units whereas the U occupies about four and one-half

units. The vertical arms are in each case about three and one-half

units long. In the lower case letters b, d, p and g all have the same

form and the q is simply the g with the stem turned to the left to

distinguish these two forms; and the a is quite similar but with a

shorter stem. The lower case u may be taken as the type of another

LABELING ILLUSTRATIONS 2 nef 59

group of letters. It is essentially like the capital U with the right

arm extended to the base. The lower case n is simply the u turned

upside down and reversed and the m is simply two ns contracted

slightly and united. While the lower case h is simply the n with the

left side prolonged the full length of lower case letters. ‘The lower

case f, j, r and t are essentially vertical lines with short curved tails

added. The capitals O and Q are essentially complete circles

which extend slightly above the top line and slightly below the base

line. The capitals C and G are parts of circles and offer no special

difficulties. The capital S is composed of two curves joined by a third

curve and is one of the most difficult letters to handle. The upper

and lower limbs are arcs of ellipses whose major axes lie in horizontal

planes with the major axis of the upper ellipse slightly shorter than

the major axis of the lower ellipse and with their minor axes about

in the ratio of two to three. The lower case 0, a and s are essentially

the same as the corresponding capitals and lower case c is similar

with a horizontal line across the upper two-thirds of the circle.

The Gothic numerals may be taken as standard just as we took

the Gothic letters. It will be noted that the numerals 1 and 4 are

the only ones composed of straight lines only. The numeral 1 is a

straight line 144 unit wide, the numeral 4 has a total width of four

units and is drawn so that the horizontal tongue is one and one-half

units above the base line. The numerals 3 and 8 are essentially the

same being composed of two broad ellipses joined together the upper

ellipse having a shorter major axis than the lower ellipse. The minor

axes of the two ellipses bear a relation of about 2 to 3 to each other.

The numeral 0 is merely a flattened ellipse with a major axis of 5

units and a minor axis of 4 units. The numerals 6 and 9 are the same

being simply placed in different positions. It will be noted that

they are essentially the same as the numeral 0 except for the forma-

tion of the small ellipse at the bottom of the 6 and the top of the 9.

Note further that the tail of the 9 is somewhat expanded being near

the base line and that the tail of the 6 is somewhat contracted being

near the top line. This preserves the proper balance. The numeral

5 is essentially the same as the 6 except that the tail is composed of a

straight vertical and horizontal line. The top is somewhat contracted

to preserve the proper balance. The numeral 7 is four units wide

with the curved vertical stroke ending on the base line about one

unit to the right of the point where the horizontal line starts on the

160 Z. P. METCALF

top limiting line. The numeral 2 is the most difficult in the whole

series as it consists of acompound curve. ‘The top curve is somewhat

like the top curve of 3 but is flatter and the bottom curve is more

pronounced than the curve in the numeral 7.

The inclined Gothic is the vertical Gothic inclined at about 15°

from the perpendicular. We need not make any special analysis of

the separate letters as that has been done for the vertical Gothic.

This is a favorite alphabet for draughtsmen who do a great deal of

lettering as it can be done with great speed and if carefully done it

looks neat and is very legible. For these reasons it is especially

valuable for large amounts of labeling.

The Roman Gothic is in many respects a more pleasing alphabet

than the Gothic. The basis of the letters is the same as for the

Gothic but certain lines called body strokes are shaded by being made

thicker, while some lines are made thinner than in the Gothic letter.

The shaded or body strokes are usually made one unit wide and the

hair lines are usually made one-half unit wide. Roman Gothic

letters of widely different appearances may readily be secured by

varying the width of the hair line. It should be noted that the

curved body strokes are slightly thicker than the straight body

strokes. Where curved lines join straight lines the union is made

very gradually so that the eye cannot detect the point of union.

The Roman letter is a further modification of the Roman-Gothic

by the addition of serifs to the strokes so that no lines end with a line

of uniform thickness. This is the type of letter used in most printing

and is the most difficult letter for the draughtsman to handle. How-

ever, it is perhaps the neatest appearing letter and should be used

more extensively than it is at the present time. The separate letters

need not be analyzed separately because they have essentially the

same form as in the Gothic alphabet. The body stroke is usually

considered as one unit in width for the straight lines and slightly

wider for the curved lines. Sometimes variation in this standard

is made for some special purpose. The hair lines vary greatly in

different styles of this letter from lines as thin as they can be drawn

easily to lines at least half a unit in width. By varying the widths

of the hair lines, letters of quite different appearances may easily be

secured. ‘The serifs demand special attention and must be drawn

neatly and accurately or they will ruin the appearance of the lettering.

Horizontal serifs are usually about one unit in length and are con-

LABELING ILLUSTRATIONS 161

nected to the main stroke by a gentle curve which is made tangent

to the main stroke and to the serif. Vertical serifs are usually made

about one and one-half units long and are connected to the main

stroke by a gentle curve which is tangent to the serif but not tangent

to the main stroke. Exception, however, must be made in the case

of the double serifs found on the tongue of the E and F, which are

smaller than the other vertical serifs. In letters like E, s and Z in

which are two vertical serifs the upper one is made slightly shorter

than the lower one for the sake of appearance. In the capital J

and some of the lower case letters it will be noted that curved lines

instead of ending in straight serifs end in curved comma shaped

marks called kerns. Instead of filling in the body strokes of Roman

letters solidly they may be indicated by two hair lines. This makes

a neat appearing letter and is useful for display titles but is difficult

to execute and therefore seldom employed in labeling biological

illustrations.

162

Plate XIV.

Plate XV.

Plate XVI.

Plate XVII.

Z. P. METCALF

EXPLANATION OF PLATES

Letters and figures of various sized type.

Vertical Gothic letters analyzed.

Roman Gothic letters analyzed.

Roman letters analyzed.

TRANSACTIONS OF THE AMERICAN MICROSCOPICAL SOCIETY

VOL. XXXIX

30 point —————_________

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24 point

ABCDEFGHIJKLMNO

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TRANSACTIONS OF THE AMERICAN MICROSCOPICAL SOCIETY

VOL. X XX1IX

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TRANSACTIONS OF THE AMERICAN MICROSCOPICAL S¢ CIETY

VOL. XXXIX

PLATE XVI

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TRANSACTIONS OF THE AMERICAN MICROSCOPICAL SOCIETY

VOL. XXXIX

METCALF

XVII

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DEPARTMENT OF NOTES AND REVIEWS

It is the purpose, in this department, to present from time to time brief original

notes, both of methods of work and of results, by members of the Society. All mem-

bers are invited to submit such items. In addition to these there will be given a few

brief abstracts of recent work of more general interest to students and teachers. There

will be no attempt to make these abstracts exhaustive. They will illustrate progress

without attempting to define it, and will thus give to the teacher current illustrations,

and to the isolated student suggestions of suitable fields of investigation. —[Editor.]

ENTOMOLOGICAL ABSTRACTS

Position of Micropterygidae —Tillyard (1919, Proc. Linn. Soc.

N. S. Wales, 44:95-136) has made an extensive study of the remark-

able family of archaic moths, the Micropterygidae. Chapman (1917)

removed the genus Micropteryx from the remainder of the family

and proposed a new order, Zeugloptera, for its reception. Comstock

(1918), on the other hand, removed the whole family Micropterygidae

from the Lepidoptera and placed it as a new suborder of the Trichop-

tera. Tillyard finds no justification for either of these views. The

proposed “‘Zeugloptera”’ is found to lack a single character not found

in some other order. In all of the Micropterygidae M, does not occur

as a separate vein of the forewing; the characteristic trichopterous

wing-spot is lacking; the pupal wing-tracheation is complete; the

scales are broad and possess numerous striae; and functional frenula

are present. These characters definitely rule out the possibility of

these insects being Trichoptera, and necessitate the conclusion that

that they must be archaic Lepidoptera.

Micro pterygidae.—Braun (1919, Ann. Ent. Soc. Am., 12:349-367)

has also attacked the problem of the position of the Micropterygidae.

A study of wing structure in the primitive Lepidoptera shows, accord-

ing to this writer, that while the Micropterygidae stand close to the

common ancestor of Lepidoptera and Trichoptera they are true

Lepidoptera and have given rise to all of the remainder of that order

by several divergent lines, one represented by the Nepticulidae,

another by the Hepialidae, and a third ‘‘much branched line includes

the frenate Lepidoptera, of which some members such as the Prodoxi-

dae, Incurvariidae, etc., conserve some of the trichopterous characters

of their ancestry and must therefore be regarded as the most primitive

of the Frenatae.”’

163

164 PAUL S. WELCH

Filariasis in U. S.—F¥rancis (1919, U. S. Publ. Health Service,

Hygienic Lab. Bull. No. 117) reports on a study of filariasis in

Southern United States. Filaria bancrofti is the species concerned and

one endemic focus has been located in this country at Charleston,

S. C. Of 400 individuals examined in that city, 77 were infected with

microfilaria, whereas of 1,470 examinations in nine southern cities

outside of Charleston only 9 showed infection. The data indicated

that cases outside of Charleston have derived the infection either

from residence in Charleston or from residence at some place outside

of the United States, as in Cuba. Transmission occurs only through

the mosquito, but the certainty of the process is limited by the follow-

ing facts: (1) No multiplication of the filaria in the mosquito;

(2) The small number actually passing successfully through the

mosquito; and the still smaller number which reach the lymph glands

of man; (3) Male and female filaria must find lodgment in the same

lymph gland of man in order that reproduction occur; (4) Infection

of mosquitoes can occur only during a few hours before and after

midnight; (5) The biting act of the mosquito only drops the micro-

filaria on the free surface of the skin of man whence it must penetrate

the intact skin. The mosquito, Culex fatigans, was found to be the

transmitter. The anatomy of the mosquito proboscis in relation to

filaria transmission is discussed and the inward and outward courses

of the filariae pointed out. The former is through the stilette bundle

along with the ingested blood, while the latter is through the interior

of the labium. Eight well executed plates, mostly in color, add to the

value of the paper.

Polyembryony and Sex.—Patterson (1919, Journ. Heredity,

10:344-352) reports results of a study of the origin and development

of mixed broods in polyembryonic Hymenoptera and the ratio in

production of males and females. In 162 broods of Copidosoma

gelechiae, 90 were female, 62 male and 10 mixed. The sex ratio was

found to be approximately 3 females to 2 males. The great excess

of females in four of the mixed broods suggested the possibility that

both sexes might arise from a single fertilized egg. In Paracopidoso-

mopsis floridanus, 1.7% of the broods were pure female, 11.3% pure

male, and 87% mixed. The percentage of males varied from 0.06

to 72.07 and in over 58% of the broods less than 10% of the indivi-

duals in any brood were males. In Platygaster rubi not a single pure

male brood was found. This, however, might be explained by the

NOTES AND REVIEWS 165

prevailing conditions which make it unusual that an unfertilized

female might escape. Only 6 of the 105 broods were pure female.

In the 99 mixed broods, the number of females, in every brood,

exceeded the number of males. In 53 broods only one male per brood

appeared, 17 had 2 males each, and 13 had 3 each. The other broods

showed a varying number, but not exceeding 10. That some mixed

broods result from two parasitic eggs, one from a fertilized female

and one from a virgin female, seems very probable but two diffiulties

stand in the way of the exclusive application of this application,

namely, (1) simultaneous emergence of individuals of a mixed

brood, and (2) striking predominance of females over the males in

the great majority of broods. A Paracopidosomopsis female, in about

66% of the cases, deposits two eggs in the host egg at a single oviposi-

tion, and in the majority of cases both eggs were found to be fertilized.

A host egg mass of 28 eggs exposed to a mixed brood of parasites

yielded 14 with 1 parasitic egg, 11 with 2 each, and 3 with 3 each.

Eight of the 11 indicated two ovipositions, while 3 seemed to represent

one oviposition. In each of the 3 remaining eggs the three parasitic

eggs apparently represented different ovipositions. Therefore the

two-egg explanation seems inadequate for the mixed broods of

Platygaster. It is proposed that some of the mixed broods may result

by one fertilized egg giving rise to both sexes through abnormal

behavior of the two sex chromosomes during early cleavage, as for

example, somatic non-disjunction in which certain blastomeres

receiving but one x chromosome would produce male embryos.

Origin and Significance of Metamorphosis——Crampton (1919,

Bull. Brooklyn Ent. Soc., 14:33-40; 93-101) considers critically the

problems. of origin and significance of metamorphosis in insects.

Presence or absence of metamorphosis, although worthy of careful

consideration, cannot be regarded as an important factor in determin-

ing the relationships of insects, according to this writer. An ancestral

group, it is contended, may include some forms which have ‘‘devel-

oped the tendency towards a metamorphosis, to a marked degree,

while other representatives of the same ancestral group do not exhibit

any marked indications of such.a tendency.” Plecoptera, Embiidae,

Dermaptera, Coleoptera and their allies constitute the ‘“plecopteroid

superorder” and are regarded as the ancestral group from which the

higher insects were derived. This group contains forms exhibiting

well marked metamorphosis and some which do not. The higher

166 PAUL S. WELCH

forms are divided into two super orders: (1) the “‘psocoid superorder”’

containing the Psocodae, Mallophaga, Anopleura (Pediculidae, s.b.),

Hemiptera, Homoptera and their allies—a group in which few mem-

bers exhibit traces of metamorphosis; and (2) the “neuropteroid

superorder’’ comprising the Neuroptera, Hymenoptera, Mecoptera,

Diptora, Siphonaptera, Trichoptera, Lepidoptera and their allies, all

being predominantly holometabolous. Thus it is suggested that we

might expect the coleopterous representatives of the ancestral group

to be somewhat nearer the derived holometabolous group, while the

remaining representatives of the ancestral group would be nearer the

derived non-metabolous group. To account for the origin of meta-

morphosis among some of the ancestral forms, it is thought that there

arose a tendency (by mutation, etc.) of the immature stages to differ

from the adults, resulting eventually in stages which could enter an

environment untenable by the adult. Such forms, favored by natural

selection, would tend to persist and thus there would appear a

“propensity towards the production of complete metamorphosis.”

Against the claim of Handlirsch that cold produced metamorphosis,

Crampton argues that “insects in which the tendency toward meta-

morphosis was already well developed, were better equipped than their

less fortunate fellows, to penetrate the less favorable regions of

winter-frost, etc., and there establish themselves.’”? No support is

found in embryology or palaeontology for the view that larval stages

represent “free-living embryos.” Disagreement with any view that

environment causes metamorphosis is expressed. The pupal stage is

regarded as the ‘‘making over”’ period necessitated when immature

and adult stages come to differ so markedly that a great change must

be involved in the transition. Larvae stages are regarded by this

author as having some phylogenetic significance and may yield

valuable hints as to relationships. Whether primitive types of larvae

represent ancestral conditions more nearly than adults do seems

uncertain. In some cases it seems to be true but in other instances

the larvae have become far more specialized than the adult, thus

involving secondary characters.

PauL S. WELCH

Department of Zoology,

University of Michigan

TABLE OF CONTENTS

For VoLuUME XXXIX, Number 3, July, 1920

Protozoa of the Devil’s Lake Complex, with two plates, by C. H. Edmondson.... 167

Age, Growth, and Scale Characters of the Mullets, Mugil cephalus and Mugil

curema, with seven figures and seven plates, by A. P. Jacot................ 199

TRANSACTIONS

American Microscopical Society

(Published in Quarterly Instalments)

Vol. XXXTX JULY, 1920 No. 3

PROTOZOA OF THE DEVIL’S LAKE COMPLEX,

NORTH DAKOTA

CHARLES HowaArp Epwuonpson, PH.D.

University of Oregon!

CONTENTS

PAGE

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1. INTRODUCTION

Several reports previously issued? have described the physio-

graphic and chemical characteristics of Devil’s Lake situated in

Ramsey County, North Dakota. Ina recent paper Dr. R. T. Young,’

of the University of North Dakota, has indicated something of the

possibilities and limitations of the lake from a biological point of

view, as well as the general scope of the work already accomplished

in that direction. It will only be necessary, therefore, to set forth

a few of the specific features of this water area which may have some

bearing on the report to follow.

! The investigations included in this report were carried on at the State Biological

Station of North Dakota.

? Biennial Report of the State Biological Station of North Dakota, 1911-12.

Pope, T. E. B. Devil’s Lake, North Dakota, a study of physical and biological

conditions, with a view to the acclimatization of fish. U.S. Bureau of Fisheries Docu-

ment 634, 1908.

Simpson, H. E. The Physiography of the Devil’s-Stump Lake Region, North

Dakota. Sixth Biennial Report of the State Geological Survey of North Dakota, 1912.

Upham, W. The Glacial Lake Agassiz, Mon. 25, U. S. Geological Survey, 1895.

3 Young, R. T. The Work of the North Dakota Biological Station at Devil’s

Lake. The Scientific Monthly, December, 1917.

170 CHARLES HOWARD EDMONDSON, PH.D.

Biological studies of Devil’s Lake made by the United States

Bureau of Fisheries in 1908 indicate the presence of four vertebrate

inhabitants of the lake, namely: a stickleback, Eucalia inconstans;

a minnow, Pimephales promelas; the hellbender, Crytobranchus

alleghaniensis; and the leopard frog, Rana pipens. Among the

metazoan invertebrates reported are crustaceans, rotifers, nematodes,

a flat worm, an arachnid and a number of species of insects. One

may collect the shells of at least fifteen molluscs from the water line

on the shore, but no living forms have been taken from the lake.

Sponges, coelenterates, polyzoans and annelids are apparently en-

tirely absent.

Investigations of the protozoan fauna of the Devil’s Lake complex

were undertaken as a part of the general biological survey of that

water area. Although, in many respects, this fauna was found to be

such as one might expect in a fresh water lake of similar depth, yet

some very pronounced differences were disclosed. The almost total

absence of shell-bearing rhizopods may possibly find its explanation

in the chemical analysis of the water. Arcella vulgaris Ehrenberg,

a very constant and usually abundant form in fresh water, was rarely

observed and two species of Difflugia, which are among the most com-

mon protozoa in lakes where there is considerable ooze, were taken

only in situations where the salinity of the water must have been

materially reduced by the in-seepage of surface water. A species of

Euglypha was taken in the overflow of the lake water from the fish

tank. The only other shelled rhizopod observed was a single speci-

men of Cyhoderia ampulla Leidy, taken from the main lake.

The fact that the ooze at the bottom of the lake at times has been

found to be entirely free from oxygen might also be a contributing fac-

tor to the scarcity of these usually common bottom-dwelling rhizopods

of the shell-bearing type, although the presence of the larvae of a cer-

tain midge in this ooze as well as the work of Birge and Juday in

Wisconsin,’ where a considerable number of animals were found at

the bottom of lakes in the absence of oxygen, would hardly seem to

make this factor one of great importance.

Experiments of a preliminary character, recorded at the end of the

taxonomic part of this report, indicate that certain protozoa having

7 Birge and Juday, The Inland Waters of Wisconsin; Wisconsin Geological

and Natural History Survey, 1911.

PROTOZOA OF THE DEVIL’S LAKE COMPLEX den

adjusted themselves to fresh water conditions are not, in all cases at

least, readily adaptable to the waters of Devil’s Lake.

The writer wishes to acknowledge his indebtedness to Dr. R. T.

Young, Director of the State Biological Station of North Dakota,

whose co-operation made this report possible, and to Mr. E. G.

Moberg for his valuable assistance in collecting material.

2. TAXONOMY

SUBPHYLUM SARCODINA

CLASS RHIZOPODA

SUBCLASS AMOEBAE

ORDER GYMMAMOEBIDA

Famity AMOEBIDAE

Genus Amoeba Ehrenberg, 1831

Amoeba proteus (Résel).

Der kleine Proteus Résel, Insecten Belustigung, 1755, tab. 101.

Amoeba proteus Leidy, Pr. Ac. Nat. Sc., 1878.

Occurrence.—Associated with Ruppia in Whipple Bay, Creel

Bay, Minnewaukon Bay, Six-mile Bay, East Lake, and also taken

from the east side of the main lake and from the overflow of lake

water from the fish tank near the laboratory.

Amoeba radiosa Ehrenberg.

Amoeba radiosa Ehrenberg, Abh. Akad. Wiss., Berlin, 1830.

Occurrence.—Rarely observed. Taken with Ruppia from Minne-

waukon Bay, also from Big Mission Lake.

Amoeba limax Dujardin.

Amoeba limax Dujardin, Histoire Naturelle des Zoophytes In-

fusoires, Paris, 1841.

Occurrence.—Associated with Ruppia and algae at the head of

Creel Bay, Big Mission Lake (numerous), Little Mission Lake

(numerous), and the east side of the main lake (numerous).

Amoeba verrucosa Ehrenberg.

Amoeba verrucosa Ehrenberg, Die Infusionsthierchen als Volkom-

mene Organismen, 1838.

Occurrence.—Observed but once, from material taken along the

east side of Creel Bay.

172 CHARLES HOWARD EDMONDSON, PH.D.

Amoeba guttula Dujardin.

Amoeba guttula Dujardin, Histoire Naturelle des Zoophytes

Infusoires, Paris, 1841.

Occurrence.—Taken from algae near Brannon’s Island, from both

ooze and floating algae in Creel Bay, from the east side of the main

lake, and from sediment on rocks near the Station.

Amoeba striata Pénard.

Amoeba striata Pénard, Etudes sur les Rhizopodes d’eau douce.

Mem. Soc. Phys. et Hist. Nat. Geneve, 1890.

Occurrence.—One specimen only observed in plant infusion from

Stump Lake.

Amoeba vitraea (Hertwig and Lesser).

Dactylosphaerium vitraem Hertwig and Lesser, Ueber Rhizopoden

und denselben nahestehende Organismen. Arch. Mikr. Anat. Vol. 10,

Suppl., 1874.

Occurrence.—Taken from the east side of Creel Bay.

ORDER TESTACEA

FAMILY ARCELLIDAE

Genus Difflugia Leclerc, 1815

Difflugia pyriformis Perty.

Difflugia pyriformis Perty, Zur Kenntniss kleinster Lebensformen

in der Schweiz, 1852.

Occurrence.—Only observed from Big Mission Lake in a location

where fresh water seeps into the lake.

Difflugia constricta Ehrenberg.

Difflugia constricta Ehrenberg, Abh. Akad. Wiss. Berlin, 1841.

Occurrence.—Taken from Big Mission Lake in the same situation

as the preceding species, and also from the head of Creel Bay near

the entrance of a sewer.

Genus Arcella Ehrenberg, 1830

Arcella vulgaris Ehrenberg.

Arcella vulgaris Ehrenberg, Abh. Akad. Wiss. Berlin, 1830.

Occurrence.—Taken in ooze from the head of Creel Bay and from

near the station, also from Big Mission Lake near the in-seepage of

fresh water; abundant in the latter locality.

PROTOZOA OF THE DEVIL’S LAKE COMPLEX ilif/s)

FAMILY EUGLYPHIDAE

Genus Cyphoderia Schlumberger, 1845

Cyphoderia ampulla (Ehrenberg).

Diffiugia ampulla Ehrenberg, Bericht Preuss. Akad. Wiss., 1840.

Occurrence.—One specimen only has been observed. Taken from

Whipple Bay among Ruppia.

Genus Euglypha Dujardin, 1841

Euglypha alveolata Dujardin.

Euglypha alveolata Dujardin, Histoire Naturelle des Zoophytes

Infusoires, 1841.

Occurrence.—Taken from the overflow of lake water from the

fish-tank near the Station. Observed but once.

SUBCLASS HELIOZOA

ORDER APHROTHORACIDA

Genus Actinophrys Ehrenberg, 1830

Actinophrys sol Ehrenberg.

Actinophrys sol Ehrenberg, Abh. Akad. Wiss., Berlin, 1830.

Occurrence.—Rarely observed, taken from among Ruppia in

Minnewaukon Bay.

SUBPHYLUM MASTIGOPHORA

CLASS ZOOMASTIGOPHORA

SUBCLASS LISSOFLAGELLATA

ORDER MONADIDA

FaMILy RHIZOMASTIGIDAE

Genus Cercomonas Dujardin, 1841

Cercomonas sp. Figures 1-3, Plate XVIII.

Probably Cercomonas longicauda Dujardin. Very plastic with

caudal filament often developed. Diameter, when spherical, 10u

Occurrence.—Observed in infusions from Stump Lake only.

Famity HeTEROMONADIDAE

Genus Monas Miiller, 1786

Monas sp. Figures 4, 5, Plate XVIII.

Very plastic. Diameter, when spherical, 20u. May represent

Monas fluida Dujardin.

Occurrence.—In the ooze from Creel Bay.

174 CHARLES HOWARD EDMONDSON, PH.D.

Monas sp. Figure 8, Plate XVIII.

Length 9; body persistent in form, anterior region very granular.

Corresponds in some degree to Monas irregularis Perty.

Occurrence.—In the ooze from Creel Bay. From a stale culture

of Ruppia, Creel Bay.

Monas sp. Figure 7, Plate XVIII.

Body moderately plastic. Length, when extended, 15-18y.

Possibly same as figures 4 and 5.

Occurrence.—In the ooze from Creel Bay.

ORDER HETEROMASTIGIDA

FAMILY HETEROMITIDAE

Genus Heteromita Dujardin, 1841

Heteromita globosa (Stein).

Bodo globosus Stein, Der Organismus des Infusionthiere, Abth. 3,

1878.

Occurrence.—In dredged material from Creel Bay.

Heteromita sp. Figure 6, Plate XVIII.

But little of detail determined. Length 54. The form probably

represents Heteromita ovata Dujardin.

Occurrence.—Taken from ooze on rocks near the Station.

ORDER POLYMASTIGIDA

FamMILy POLYMASTIGIDAE

Genus Trepomonas Dujardin, 1841

Trepomonas agilis Dujardin.

Trepomonas agilis Dujardin, Histoire Naturelle des Zoophytes

Infusoires, 1841.

Occurrence.—Taken from Big Mission Lake, Whipple Bay, from

the ooze of the main lake and from the east side of the main lake.

Abundant in the latter locality.

ORDER EUGLENIDA

FamiILty EUGLENIDAE

Genus Euglena Ehrenberg, 1830

Euglena viridis Ehrenberg.

Euglena viridis Ehrenberg Abh. Akad. Wiss., Berlin, 1830.

PROTOZOA OF THE DEVIL’S LAKE COMPLEX WAS

Occurrence.—Observed from Minnewaukon Bay, Big Mission

Lake, in the ooze from Creel Bay and from the east side of the main

lake.

Euglena desus Ehrenberg.

Euglena desus Ehrenberg, Abh. Akad. Wiss., Berlin, 1830.

Occurrence.—Minnewaukon Bay, Six-mile Bay, near Brannon’s

Island, Big Mission Lake, Little Mission Lake, East Lake, and the

ooze from the main lake.

Genus Phacus Dujardin, 1841

Phacus pyrum (Ehrenberg).

Euglena pyrum Ehrenberg, Abh. Akad. Wiss., Berlin, 1830.

Occurrence.— Minnewaukon Bay, Creel Bay, Big Mission Lake

(numerous), and the east side of the main lake.

Genus Eutreptia Perty, 1852

Eutreptia viridis Perty.

Eutreptia viridis Perty, Zur Kenntniss kleinster Lebensformen in

der Schweiz, 1852.

Occurrence.—From the surface among Ruppia, Big Mission Lake.

Famity ASTASIIDAE

Genus Astasia Ehrenberg, 1830

Astasia tricophora (Ehrenberg).

Trachelius tricophorus Ehrenberg, Abh. Akad. Wiss., Berlin, 1830.

Occurrence.—Among Ruppia from Whipple Bay, from Creel Bay,

in the ooze from Big Mission Lake, and among algae near Brannon’s

Island.

FAMILY PERANEMIIDAE

Genus Petalomonas Stein, 1859

Petalomonas mediocanellata Stein.

Petalomonas mediocanellata Stein, Der Organismus der Infusions-

thiere, 1878.

Occurrence.—Taken from the surface of Big Mission Lake and

from the ooze of the main lake.

176 CHARLES HOWARD EDMONDSON, PH.D.

Petalomonas sp. Figure 10, Plate XVIII.

Has some resemblance to Petalomonas ervilia Stein. Conspicuous

groove entire length of the body. Length 36un.

Occurrence.—From the ooze of Creel Bay.

Genus Heteronema Dujardin, 1841

Heteronema acus (Ehrenberg).

Astasia acus Ehrenberg, Abh. Akad. Wiss., Berlin, 1830.

Occurrence.—From Six-mile Bay and from the ooze of Creel

Bay.

Genus Anisonema Dujardin, 1841

Anisonema grande (acinus) (Ehrenberg).

Bodo grandis Ehrenberg, Die Infusionsthierchen als Volkommene

Organismen, 1838.

Anisonema acinus Dujardin, Histoire Naturelle des Zoophytes

Infusoires, 1841.

Occurrence.—Among Ruppia and algae at the head of Creel Bay.

Genus Notosolenus Stokes, 1884

Notosolenus sp. Figure 9, Plate XVIII.

Length about 15y.

Occurrence.—From Whipple Bay, Stump Lake and from the

overflow of the fish-tank near the Station.

ORDER CHLOROFLAGELLIDA

FAMILY TETRAMITIDAE

Genus Tetraselmis Stein, 1878

Tetraselmis cordiformis (Carter).

Cryptoglena cordiformis Carter, Annals of Natural History 1858.

Occurrence.—Taken from Stump Lake only.

FAMILY POLYTOMIDAE

Genus Polytoma Ehrenberg, 1838

Polytoma uvella Ehrenberg.

Polytoma uvella Ehrenberg, Die Infusionsthierchen als Volkom-

mene Organismen, 1838.

Occurrence.—Found at the head and along the east side of Creel

Bay.

PROTOZOA OF THE DEVIL’S LAKE COMPLEX 177

FAMILY TRIMASTIGIDAE

Undetermined genus

Undetermined species. Figures 11, 12, Plate XVIII.

Description.—Body elongate, somewhat compressed, slightly

plastic, attenuated posteriorly; surface marked longitudinally by

several conspicuous ridges; flagella three in number arising from the

anterior extremity, equal and equalling the body in length; nucleus

and contractile vacuole unobserved. Length 20u.

Occurrence.—Numerous among Ruppia from Creel Bay.

FamMILy CHLAMYDOMONADIDAE

Genus Chlamydomonas Ehrenberg, 1833

Chlamydomonas pulvisculus Ehrenberg.

Chlamydomonas pulvisculus Ehrenberg, Abh. Akad. Wiss.,

Berlin, 1833.

Occurrence.—Taken from the head of Creel Bay.

SUBCLASS DINOFLAGELLIDA

ORDER DINIFERIDA

FAMILY PERIDINIIDAE

Genus Glenodinium Ehrenberg, 1832

Glenodinium pulvisculus Ehrenberg.

Glenodinium pulvisculus Ehrenberg, Die Infusionsthierchen als

Volkommene Organismen, 1838.

Occurrence.—Taken from the surface and from the ooze at the

bottom of Creel Bay.

SUBPHYLUM INFUSORIA

CLASS CILIATA

ORDER HOLOTRICHA

FAaMILy ENCHELINIDAE

Genus Holophrya Ehrenberg, 1831

Holophrya ovum Ehrenberg.

Holophrya ovum Ehrenberg, Die Infusionsthierchen als Volkom-

mene Organismen, 1838.

Occurrence.—Among algae from Creel Bay.

178 CHARLES HOWARD EDMONDSON, PH.D.

Holophrya sp. Figure 13, Plate XVIII.

Resembling Holophrya ovum Ehrenberg but much smaller.

Length 30-40y.

Occurrence.—In the ooze from Creel Bay.

Genus Urotricha Claparéde and Lachmann, 1858

Urotricha labiata, new species, Figure 14, Plate XVIII.

Description.—Body ovate, about twice as long as broad, equally

rounded at both extremities. Cilia covering the entire body, ar-

ranged in longitudinal rows and vibrating independently. A very

fine seta, nearly as long as the body, extending from the posterior

extremity. Mouth anterior, subterminal, beneath a prominent,

lobe-like lip. Nucleus central. Contractile vacuole posterior.

Reproduction by transverse fission. Length of body about 30g.

Occurrence.—Taken from numerous localities in Devil’s Lake.

Genus Prorodon Ehrenberg, 1833

Prorodon teres Ehrenberg.

Prorodon teres Ehrenberg, Die Infusionsthierchen als Volkom-

mene Organismen, 1838.

Occurrence.—Among Ruppia and algae of Big Mission Lake and

the main lake.

Prorodon edentatus Claparéde and Lachmann.

Prorodon edentatus Claparéde and Lachmann, Etudes sur les

Infusoires et les Rhizopodes, 1858.

Occurrence.—Infusions of Ruppia from Big Mission Lake and

Minnewaukon Bay.

Prorodon griseus Claparéde and Lachmann.

Prorodon griseus Claparéde and Lachmann, Etudes sur les Infu-

soires et les Rhizopodes, 1858.

Occurrence.—Taken from Stump Lake only.

Genus Enchelys Ehrenberg, 1838

Enchelys sp. Figure 15, Plate XVIII.

Length from 15-20u.

Occurrence.—Ooze from the main lake and from the overflow of

lake water from the fish-tank.

PROTOZOA OF THE DEVIL’S LAKE COMPLEX 179

Genus Spathidium Dujardin, 1841

Spathidium spatula Dujardin.

Spathidium spatula Dujardin, Histoire Naturelle des Zoophytes

Infusoires, 1841.

Occurrence.—Among algae from the head of Creel Bay.

Spathidium sp. Figure 16, Plate XVIII.

A very long, narrow and flattened form. Length 120y.

Occurrence.—Taken from infusions from the head of Creel Bay.

Spathidium sp. Figure 17, Plate XVIII.

A much shorter form than the preceding, with a conspicuous

collar about the oral extremity. Length 30y.

Occurrence.—From the ooze of the main lake.

Undetermined Genus®

Undetermined species. Figures 1, 2, Plate XIX.

Description.—Body elongate, plastic, slightly compressed dorso-

ventrally, inflated posteriorly, narrow anteriorly, rounded at both

extremities; cilia of uniform length arranged in longitudinal rows,

covering the entire surface; aperture a narrow slit diagonally placed,

sub-terminal; contractile vacuole posterior; nucleus concealed; endo-

plasm completely filled with green chloroplasts. Length 90y.

Occurrence.—From the surface of the main lake and from among

Ruppia and algae.

Genus Chaenia Dujardin, 1841

Chaenia teres Dujardin.

Chaenta teres Dujardin, Histoire Naturelle des Zoophytes Infu-

soires. 1841.

Occurrence.—Among algae from the head of Creel Bay.

Genus Mesodinium Stein, 1862

Mesodinium pulex (Claparéde and Lachmann).

Halteria pulex Claparéde and Lachmann, Etudes sur les Infu-

soires et les Rhizopodes, 1858.

Occurrence.—A common form on the surface and in the ooze of

the main lake.

8 The form is treated here with doubt as to its taxonomic position.

180 CHARLES HOWARD EDMONDSON, PH.D.

Genus Didinium Stein, 1859

Didinium nasutum (Miller).

Vorticella nasutum Miiller, Animalcula Infusoria Fluviatilia et

Marina, 1786.

Occurrence.—Among Ruppia from Minnewaukon Bay, Whipple

Bay, and from the east side of the main lake.

Genus Lacrymaria Ehrenberg, 1830

Lacrymaria olor Ehrenberg.

Lacrymaria olor Ehrenberg, Abh. Akad. Wiss., Berlin, 1830.

Occurrence—Among Ruppia in Creel Bay.

Lacrymaria truncata Stokes.

Lacrymaria truncata Stokes, Ann. and Mag. Nat. Hist., June,

1885.

Occurrence.—Among Ruppia from the north end of the main lake.

Lacrymaria cohnii Kent.

Lacrymaria cohnii Kent, A Manual of the Infusoria, 1881-1882.

Occurrence.—In an infusion from Stump Lake.

Lacrymaria lagenula Claparéde and Lachmann.

Lacrymaria lagenula Claparéde and Lachmann, Etudes sur les

Infusoires et les Rhizopodes, 1858.

Occurrence.—In ooze from the main lake.

Famity TRACHELINIDAE

Genus Lionotus Wrzesniowski, 1870

Lionotus fasciola (Ehrenberg).

Amphileptus fasciola Ehrenberg, Die Infusionsthierchen als Vol-

kommene Organismen, 1838.

Occurrence.—Abundant in many parts of the main lake, also

taken from Stump Lake and Big Mission Lake.

Lionotus sp. Figure 3, Plate XIX.

A very small species. Length about 40u. Often seen in conjuga-

tion.

Occurrence.—Among algae from Creel Bay.

PROTOZOA OF THE DEVIL’S LAKE COMPLEX 181

Genus Amphileptus Ehrenberg, 1830

Amphileptus meleagris (Ehrenberg).

Trachelius meleagris Ehrenberg, Die Infusionsthierchen als Vol-

kommene Organismen, 1838.

Amphileptus meleagris Claparéde and Lachmann, Etudes sur les

Infusoires et les Rhizopodes, 1858.

Occurrence.—Taken in Stump Lake and from algae at the head of

Creel Bay.

Famity CHLAMYDODONTIDAE

Genus Nassula Ehrenberg, 1838

Nassula rubens (Perty).

Cyclogramma rubens Perty, Zur Kenntniss kleinster Lebensformen

in der Schweiz, 1852.

Nassula rubens Claparéde and Lachmann, Etudes sur les Infu-

soires et les Rhizopodes, 1858.

Occurrence.—From the overflow of lake water from the fish-tank

near the Station.

Nasula ornata Ehrenberg.

Nasula ornata Ehrenberg, Die Infusionsthierchen als Volkom-

mene Organismen, 1838.

Occurrence.—Taken from Lake ‘‘N”’ only.

Genus Chilodon Ehrenberg, 1833

Chilodon cucullulus (Miiller).

Colpoda cucullulus Miller, Animalcula Infusoria Fluviatilia et

Marina, 1786.

Occurrence.—Infusions of algae from Creel Bay, Big Mission

Lake, and Whipple Bay.

Chilodon caudatus Stokes.

Chilodon caudatus Stokes, Am. Jour. Sci. 29, April, 1885.

Occurrence.—Among Ruppia from Minnewaukon Bay.

Genus Aegyria Claparéde and Lachmann, 1858

Aegyria pusilla (?) Claparéde and Lachmann.

Aegyria pusilla Claparéde and Lachmann, Etudes sur les Infu-

soires et les Rhizopodes, 1858.

Occurrence.—Among algae near the Station.

182 CHARLES HOWARD EDMONDSON, PH.D.

FAMILY CHILIFERIDAE

Genus Glaucoma Ehrenberg, 1830

Glaucoma scintillans Ehrenberg.

Glaucoma scintillans Ehrenberg, Die Infusionsthierchen als

Volkommene Organismen, 1838.

Occurrence.—In algae infusion from near Brannon’s Island.

Glaucoma margaritaceum (Ehrenberg).

Cyclidium margaritaceum Ehrenberg, Die Infusionsthierchen als

Volkommene Organismen, 1838.

Cinetochilum margaritaceum Perty, Zur Kenntniss_ kleinster

Lebensformen in der Schweiz, 1852.

Occurrence.—Very abundant. From the ooze of Creel Bay, the

surface of Creel Bay, Stump Lake, and near Brannon’s Island in the

main lake.

Genus Leucophrys Ehrenberg, 1830

Leucophrys patula (Miller).

Trichoda patula Miiller, Animalcula Infusoria Fluviatilia et

Marina, 1786.

Occurrence.—One specimen only observed, from the east side of

the main lake. A very typical specimen.

Genus Frontonia Ehrenberg, 1538

Frontonia leucas Ehrenberg.

Frontonia leucas Ehrenberg, Die Infusionsthierchen als Volkom-

mene Organismen, 1838.

Occurrence.—Taken from the east side of the main lake and from

East Lake. Abundant in Six-mile Bay and Minnewaukon Bay.

Genus Loxocephalus Eberhard, 1868

Loxocephalus granulosus Kent.

Loxocephalus granulosus Kent, A Manual of the Infusoria, 1881—

1882.

Occurrence.—Taken only in the ooze of Big Mission Lake near

the in-seepage of fresh water.

PROTOZOA OF THE DEVIL’S LAKE COMPLEX 183

Genus Uronema Dujardin, 1841

Uronema marinum Dujardin.

Uronema marinum Dujardin, Histoire Naturelle des Zoophytes

Infusoires, 1841.

Occurrence.—One of the most common species in the lake.

Abundant everywhere both at the surface and in the ooze.

Genus Colpidium Stein, 1868

Colpidium putrinum Stokes.

Colpidium putrinum Stokes, Ann. and Mag. Nat. Hist. Feb.,

1886.

Occurrence.—From algae at the east side of Creel Bay.

Genus Tillina Gruber, 1879

Tillina saprophila Stokes.

Tillina saprophila Stokes, Am. Nat., Feb., 1884.

Occurrence.—Taken only in the overflow of lake water from the

fish-tank near the station.

FAMILY PARAMAECIDAE

Genus Paramaecium Miiller, 1786

Paramaecium trichium Stokes.

Paramaecium trichium Stokes, Am. Naturalist, 19, May, 1885.

Occurrence.—From near the mouth of a sewer at the head of

Creel Bay, and from ooze near the rock pile in the main lake.

Paramaecium caudatum Ehrenberg.

Paramaecium caudatum Ehrenberg. Die Infusionsthierchen als

Volkommene Organismen, 1838.

Occurrence.—Taken from Big Mission Lake near the in-seepage

of fresh water.

FAMILY PLEURONEMIDAE

Genus Cyclidium Ehrenberg, 1838

Cyclidium glaucoma Ehrenberg.

Cyclidium glaucoma Ehrenberg, Die Infusionsthierchen als Vol-

kommene Organismen, 1838.

Occurrence.—Abundant everywhere, at the surface and in the

ooze in all parts of the lake.

184 CHARLES HOWARD EDMONDSON, PH.D.

Cyclidium litomesum Stokes.

Cyclidium litomesum Stokes, Am. Monthly Micro. Jour., 6, Dec.

1884.

Occurrence.—Numerous in infusions from the head of Creel

Bay and in the ooze from the main lake.

Genus Pleuronema Dujardin, 1841

Pleuronema chrysalis (Ehrenberg).

Paramaecium chrysalis Ehrenberg, Die Infusionsthierchen als

Volkommene Organismen, 1838.

Pleuronema crassa Dujardin, Histoire Naturelle des Zoophytes

Infusoires, 1841.

Occurrence.—Observed in infusions from Stump Lake only.

ORDER HETEROTRICHA

FAMILY PLAGIOTOMIIDAE

Genus Metopus Claparéde and Lachmann, 1858

Metopus sigmoides (Miiller).

Trichoda sigmoides Miller, Animalcula Infusoria Fluviatilia et

Marina, 1786.

Occurrence.—Common in dredged material from Minnewaukon

Bay, Creel Bay, and the main lake. Abundant in East Lake.

Genus Spirostomum Ehrenberg, 1835

Spirostomum ambiguum Ehrenberg.

Spirostomum ambiguum Ehrenberg, Abh. Akad. Wiss., Berlin,

1835.

Occurrence.—Observed in dredged material from Creel Bay.

FAmILy HALTERIDAE

Genus Halteria Dujardin, 1841

Halteria grandinella (Miiller).

Trichoda grandinella Miiller, Animalcula Infusoria Fluviatilia et

Marina, 1786.

Halteria grandinella Dujardin, Histoire Naturelle des Zoophytes

Infusoires, 1841.

Occurrence.—Common in infusions of Ruppia and algae from

Whipple Bay and Creel Bay and in the ooze of the main lake.

PROTOZOA OF THE DEVIL’S LAKE COMPLEX 185

ORDER HYPOTRICHA

FAMILY OxYTRICHIDAE

Genus Uroleptus® Ehrenberg, 1831

Uroleptus agilis Englemann.

Uroleptus agilis Englemann, Zeit. Wiss. Zool., Bd. 11, 1861.

Occurrence.—From the ooze of the main lake, also from Six-mile

Bay.

Uroleptus rattulus (?) Stein.

Uroleptus rattulus Stein, Der Organismus der Infusionsthiere,

1859.

Occurrence.—Among Ruppia from Whipple Bay.

Genus Oxytricha® Ekrenberg, 1830

Oxytricha fallax Stein.

Oxytricha fallax Stein, Der Organismus der Infusionsthiere, 1859.

Occurrence.—Among algae from Creel Bay.

Oxytricha pellionella (Miiller).

Trichoda pellionella Miller, Animalcula Infusoria Fluviatilia et

Marina, 1786.

Oxytricha pellionella Ehrenberg, Die Infusionsthierchen als Vol-

kommene Organismen, 1838.

Occurrence.—Taken from Ruppia near the Station, Big Mission

Lake, Whipple Bay, north end of Creel Bay, and the ooze from the

fish-tank after being flooded by lake water.

Oxytricha parvistyla Stein.

Oxytricha parvistyla Stein, Der Organismus der Infusionsthiere,

1859.

Occurrence.—Among Ruppia near the Station.

Oxytricha bifaria Stokes.

Oxytricha bifaria Stokes, Ann. and Mag. Nat. Hist., Aug., 1887.

Occurrence.—Abundant in Creel Bay, also taken from Whipple

Bay.

* Further study would, no doubt, result in the determination of other species

of the genus than those listed.

186 CHARLES HOWARD EDMONDSON, PH.D.

Genus Histrio Sterki, 1878

Histrio erethysticus Stokes.

Histrio erethysticus Stokes, Proc. Am. Philos. Soc, 24; 126, 1887.

Occurrence.—Among Ruppia from near the Station.

Genus Stylonychia Ehrenberg, 1830

Stylonychia notophora Stokes.

Stylonychia notophora Stokes, Ann. and Mag. Nat. Hist. June,

1885.

Occurrence.— With algae from Creel Bay.

Genus Holosticha Wrzesniowshi, 1877

Holosticha vernalis (?) Stokes.

Holosticha vernalis Stokes, Ann. and Mag. Nat. Hist., Aug., 1887.

A form bearing considerable resemblance to Stokes’ species

was occasionally observed. Length 140y.

Occurrence.—Among Ruppia from the main lake.

Genus Pleurotricha Stein, 1859

Pleurotricha lanceolata (Ehrenberg).

Stylonychia lanceolata Ehrenberg, Die Infusionsthierchen als

Volkommene Organismen, 1838.

Pleurotricha lanceolata Stein, Der Organismus der Infusionsthiere,

1859.

Occurrence.—Taken at the head of Creel Bay.

Genus Tachysoma Stokes, 1887

Tachysoma parvistyla Stokes.

Tachysoma parvistyla Stokes, Ann. and Mag. Nat. Hist. Aug.,

1887.

Occurrence.—Observed in infusions from Stump Lake only.

FAMILY EUPLOTIDAE

Genus Euplotes Ehrenberg, 1831

Euplotes charon (Miiller).

Trichoda charon Miller, Animalcula Infusoria Fluviatilia et

Marina, 1786.

Euplotes charon Ehrenberg, Die Infusionsthiere als Volkom-

mene Organismen, 1838.

PROTOZOA OF THE DEVIL’S LAKE COMPLEX 187

Occurrence.—Abundant among infusions of Ruppia and algae

from many parts of the main lake, and also from East Lake.

Euplotes patella (Miiller).

Kerona patella Miiller, Animalcula Infusoria Fluviatilia et

Marina, 1786.

Euplotes patella Ehrenberg, Die Infusionsthiere als Volkommene

Organismen, 1838.

Occurrence.—Found in Stump Lake, Big Mission Lake, East

Lake and in numerous localities in the main lake.

Genus Aspidisca Ehrenberg, 1830

Aspidisca costata (Dujardin).

Coccudina costata Dujardin, Histore Naturelle des Zoophytes

Infusoires, 1841.

Occurrence.—Taken in Whipple Bay; numerous among Ruppia

in Minnewaukon Bay and also on the east side of the main lake.

ORDER PERITRICHA

FAMILY VORTICELLIDAE

Genus Vorticella Linnaeus, 1767

Vorticella telescopica Kent.

Vorticella telescopica Kent, a Manual of the Infusoria, 1881-1882.

Occurrence.—Among Ruppia at the north end of the main lake.

Vorticella convallaria Linnaeus.

V orticella convallaria Linnaeus, Systema Naturae, Ed. 12, 1767.

Occurrence.—Attached to diatoms in the main lake, also among

Ruppia in Big Mission Lake.

Vorticella octavo Stokes.

Vorticella octavo Stokes, Ann. and Mag. Nat. Hist., June, 1885.

Occurrence.—Among Ruppia at the north end of the main lake.

Vorticella microstoma Ehrenberg.

Vorticella microstoma Ehrenberg, Die Infusionsthierchen als

Volkommene Organismen, 1838.

Occurrence.—Taken at the east side of the main lake.

188 CHARLES HOWARD EDMONDSON, PH.D.

Vorticella sp. Figure 4, Plate XIX.

A very common form, resembling Vorticella rabdostyloides Kellicott

but is considerably smaller and the body is transversely striated.

Length of stalk 124, with the diameter of the body nearly the same.

Occurrence.—Attached to floating diatoms.

Vorticella sp. Figure 5, Plate XIX.

A species with more elongate body than the preceding but also

transversely striate. Length of body 28y, stalk 68y.

Occurrence.—Attached to floating diatoms.

Genus Gerda Claparéde and Lachmann, 1858

Gerda annulata, new species. Figure 10, Plate XIX.

Description.—Body elongated, cylindrical, of nearly equal

diameter throughout, curved when extended; surface finely striate

transversely; a prominent annular ridge present usually about one-

fourth the distance from the posterior extremity; peristome border

revolute, disc slightly elevated; contractile vacuole conspicuous;

nucleus not observed. Length of body, extended, 80u.

Occurrence.—Among algae and Ruppia from the north end of

the main lake.

Genus Epistylis Ehrenberg, 1830

Epistylis plicatilis Ehrenberg.

Epistylis plicatilis Ehrenberg, Die Infusionsthierchen als Vol-

kommene Organismen, 1838.

Occurrence.—From the east side of Creel Bay.

Epistylis branchiophila Perty.

Epistylis branchiophila Perty, Zur Kenntniss kleinster Lebensfor-

men in der Schweiz, 1852.

Occurrence.—Among algae near the head of Creel Bay.

Genus Carchesium Ehrenberg, 1838

Carchesium epistylidis Claparéde and Lachmann.

Carchesium epistylidis Claparéde and Lachmann, Etudes sur les

Infusoires et les Rhizopodes, 1858.

Occurrence.—Among algae from Creel Bay.

PROTOZOA OF THE DEVIL’S LAKE COMPLEX 189

Genus Zoothamnium Ehrenberg, 1838

Zoothamnium alterans Claparéde and Lachmann.

Zoothamnium alterans Claparéde and Lachmann, Etudes sur les

Infusoires et les Rhizopodes, 1858.

Occurrence.—Among Ruppia and algae from Stump Lake.

Zoothamnium sp. Figure 6, Plate XIX.

Stalk very stout, zooids smooth, usually 2-8 in a colony. Length

of stalk 216y, of zooid 64y.

Occurrence.—From Stump Lake, East Lake, Creel Bay, Whipple

Bay, and from the main lake. Attached to algae or Ruppia. A

fairly common form.

Genus Vaginocola Lamarck, 1816

Vaginocola crystallina Ehrenberg.

Vaginocola crystallina Ehrenberg, Die Infusionsthierchen als

Volkommene Organismen, 1838.

Occurrence.—Numerous among algae from East Lake, also taken

from Stump Lake and from the north end of the main lake.

Genus Cothurnia Ehrenberg, 1831

Cothurnia imberbis Ehrenberg.

Cothurnia imberbis Ehrenberg, Die Infusionsthierchen als Vol-

kommene Organismen, 1838.

Occurrence-—Commonly attached to floating diatoms, from

dredged material and also among Ruppia in Creel Bay. Also taken

from Stump Lake.

Cothurnia curva Stein.

Cothurnia curva Stein, Der Organismus der Infusionsthiere, 1859.

Occurrence.—Among Ruppia at the north end of the main lake.

CLASS SUCTORIA

FAMILY PODOPHRYIDAE

Genus Podophrya Ehrenberg, 1838

Podophrya libera Perty.

Podophrya libera Perty, Zur Kenntniss kleinster Lebensformen in

der Schweiz, 1852.

Occurrence.—Numerous at east side of the main lake.

190 CHARLES HOWARD EDMONDSON, PH.D.

Podophrya sp. Figure 9, Plate XIX.

Bears some slight resemblance to Podophrya cyclopum Claparéde

and Lachmann. The lobulated border may have represented a

reproductive phase or possibly was abnormal. Total height 60y,

stalk 20u.

Occurrence.—Attached to algae from the main lake. Several

specimens were observed by Dr. R. T. Young.

Genus Sphaerophrya Claparéde and Lachmann, 1858

Sphaerophrya magna Maupas.

Sphaerophrya magna Maupas, Arch. de Zoologie Experimentale,

tom 9, Nov., 1881.

Occurrence.—From Stump Lake and the east side of the main

lake.

FAMILY ACINETIDAE

Genus Acineta Ehrenberg, 1838

Acineta sp. Figure 7, Plate XIX.

Body triangular in broad view, compre sed; endoplasm very

granular, nucleus concealed. Total height 50yu, stalk 20u. This

species resembles, in some degree, Acineta lemnarum Stein.

Occurrence.—From floating material in the main lake and also

among algae from Stump Lake.

Acineta sp. Figure 8, Plate XIX.

Body oval, slightly broader distally, greatly compressed; endo-

plasm granular concealing the nucleus and contractile vacuole.

Total height 60-72y, stalk about 15y.

Occurrence.—Attached to algae from Stump Lake. Commonly

feeding on Uronema.

3. EXPERIMENTS

Preliminary experiments in transferring protozoa from fresh water to the con-

centrated water of Devil’s Lake and vice versa.

In order to test the reactions of certain protozoa taken from other

sources to the more concentrated waters of Devil’s Lake a series of

simple experiments were carried out by which forms of protozoa

common to fresh water were transferred directly into the more saline

water of the lake.

PROTOZOA OF THE DEVIL’S LAKE COMPLEX 191

Infusions from a small body of fresh water near the southern

boundary of the main lake were prepared and certain protozoa which

readily appear in cultures were used in the tests.

By placing a drop of the fresh water culture on one end of a micro-

scopic slide and a drop of lake water near the middle of the slide and,

with a needle, drawing out from each drop toward the other a narrow

channel of water until the two met, the protozoa were conducted

from the fresh water drop into that of the lake water. To eliminate

possible influence of the fresh water a series of drops of lake water

were used and the organisms rapidly transferred from one to the

other until they reached a pure medium of lake water.

The waters from the two sources were kept at a uniform tempera-

ture and the effect of the change of environment thus brought about

was carefully noted by the activity of the organisms.

In similar manner the transference of certain protozoa from lake

water to fresh water was accomplished and the effect of such change

observed as hereinafter noted.

A. Transference of protozoa from fresh water to lake water.

1. Paramaecium sp. A specimen of a species, probably Paramae-

cium caudaium Ehrenberg, commonly occurring in the fresh water

was removed to the pure lake water with the following results: An

immediate change occurred in the organism. The body became

greatly compressed dorso-centrally with erratic movements at first

which soon gave way to a more steady, forward movement with slow

rotations on the long axis. A noticeable change also occurred in the

contractile vacuoles. The normal rhythmic collapse of the vacuoles

ceased after a few minutes and they became greatly dilated and dis-

torted. After ten minutes of rotary movements the organism became

quiet with the cilia of the periphera and the oral groove still active.

Many non-contractile vacuoles filled the endoplasm. Death occurred

at the end of twelve minutes.

A second specimen, after showing the same flattening of the body,

moved in circles for six minutes then assumed the forward movement

with rotations on the long axis. In eighteen minutes the organism

became quiet with a highly vacuolated endoplasm and the cilia of the

oral groove vibrating feebly. Death occurred in twenty-six minutes.

192 CHARLES HOWARD EDMONDSON, PH.D.

A third specimen after exhibiting similar physical and physiologi-

cal changes came to complete rest in twenty-two minutes. Death

resulted in twenty-five minutes.

A fourth specimen showed similar responses and died in fifteen

minutes.

Seven specimens were then transferred at the same time. Six of

these, after exhibiting similar responses as the preceding, were dead at

the end of ten minutes. One, after reacting in like manner, died at

the end of eighteen minutes.

2. Stylonychia sp. Several tests with a species of Stylonychia

were carried out. Unusual responses were less quickly manifested by

Stylonychia than Paramaecium when brought into contact with the

lake water. Commonly after five or six minutes of normal movements

a rapid whirling over and over of the body occurred gradually sub-

siding into complete rest. Death occurred in all specimens in from

sixteen to thirty-two minutes.

Reactions of similar character were obtained from Paramaecium

and Stylonychia by the introduction of small quantities of NaCL

into the fresh water in which they were normally living.

3. Metopus sp. A short type of Metopus, common in fresh water,

was transferred to the saline lake water. The most noticeable change

was an almost immediate flattening of the body. Normal rotary

movements continued for eight minutes when the organism came to

rest with the cilia of the surface still more or less active. Death

occurred at the end of fifteen minutes.

Numerous individuals of this species were used in successive

experiments with reactions similar in each case. Death resulted in all

specimens in from eleven to eighteen minutes.

B. Transference of protozoa from the concentrated lake water to

fresh water.

1. Uroleptus sp. The form used was one of the elongated types.

More than sixty specimens were used in the tests. With few excep-

tions but with considerable degree of variation, the following reactions

were very evident: After a period of from ten to fifteen minutes

contact with the fresh water, during which time more or less normal

activities were maintained, the organisms came to rest with the cilia

still in motion. The cell bodies became shortened and dilated, in

PROTOZOA OF THE DEVIL’S LAKE COMPLEX 193

many instances assuming a spherical form. After enduring this state

of depression for from ten to fifteen minutes the organisms showed

signs of recovery. The bodies gradually assumed an elongated form

and normal activities reappeared. Within a period of one hour and

twenty-five minutes from the time the organisms were first introduced

into the fresh water all, with the exception of a few which failed to

survive the state of depression, had fully recovered and were respond-

ing in a normal manner.

Considerable variation in the effect of the change was noted.

Of those surviving some were slightly affected and wholly recovered

in forty-five minutes, some in sixty minutes, while others required

the longer time noted above.

2. Euplotes patella (Miiller). Numerous individuals of this spe-

cies were transferred as in the preceding experiment. The effect in

this case was an immediate one. As soon as contact was made with

the fresh water the cell bodies became swollen and distorted, losing

the longitudinal striations and all resemblance to normal individuals.

During this state of depression the organisms were at rest with the

cirriin feeble motion. After a period of fifteen minutes the cells began

to resume movements although in a distorted condition. In fifteen

minutes more the longitudinal striations reappeared and soon after

normal responses were entirely restored.

3. Uronema marinum Dujardin. The transferrence of this spe-

cies from the lake water to fresh water resulted in no apparent state of

physical depression and no diminished or unusual responses to stimuli

could be detected. The species is commonly recognized as both a

marine and fresh water form.

4. SUMMARY AND CONCLUSIONS

SUMMARY OF THE Groups OF PROTOZOA RECORDED

SEVIS GIL REAM A aC dent NS 13 species

Mash OWNOnAt sage mwa wee ety (OS ks Ne Dew nr

PURELY yr, wh Sk ater eRe EE) toy

fo Fea Leese t ee A 0 111 species

Conclusions

1. The proportion of the number of species of the three groups of

protozoa recognized in Devil’s Lake corresponds favorably with the

same in a typical fresh water lake.

194 CHARLES HOWARD EDMONDSON, PH.D.

2. A most noticeable feature of the study of this fauna is the

apparent total absence of numerous forms universally found in fresh

water. The dearth of shell-bearing rhizopods was mentioned in the

introduction. Many common species of flagellates and ciliates were,

at no time during the survey, observed in the concentrated waters of

the lake.

3. The subdivisions of the classes of protozoa are fairly well

represented in Devil’s Lake. Two new species are described in the

report but with the exception of the facts mentioned in the preceding

paragraph, the protozoan fauna of Devil’s Lake cannot be considered

an unusual one.

4. Experiments of the interchange of protozoa between fresh

water and the lake water seem to indicate that the organisms of the

lake may adjust themselves to fresh water conditions with more

readiness than can the forms accustomed to a fresh water environ-

ment accommodate themselves to the concentrated water of the lake.

PROTOZOA OF THE DEVIL’S LAKE COMPLEX 195

5. INDEX

PAGE PAGE

PNGIAE UA tes coaipechehel mata GHEE LOOT Chaentarys syne ate errr ieee z caren 179

Acineta lemnarum..........0004: 190 i Chaentateresine jarnciceyee mice eee 179

PRENTIE CAS) «scene ais eiefeteiedn ae ie eel ove 1900 (eC@hiliferidaenasie iss sey eaieiscs: 182

UN STITT Eye se Ea 190) xiChilodontscuseeie aoe eee aes 181

EMEDTV Ss /sic als uite yee eo 17S ea Chilodonicaudatusss-5epeaeerieee 181

Mammophrys Sol... 4.65.6. 6 fe a6. 173% Chilodonjcuculluluss:oicceseo. sa 181

PRENSA ah srs s yo) 2b ovsseos't fencha Sisi dean 6 {Si Chiamydodontidaeya yy... 12a 181

PME MMUR SIN. eas sins a sce ce cise 181 Chlamydomonadidae............. 177

a STR IEL 3 pth eck Aah SIRE Ba a Pie Chlamydomonas.\s3, 5 ssa ae 177

OS PUL EELT ETE oe SL I a 171. Chlamydomonas pulvisculus...... 177

Amoeba guttula... 0.0... ee 12) -Chioroflagellidan ty) (oV. 08 alse) sare 176

Snare @layn lian lets eles ceva ae eee ena ae Ae/hattt Cilia taney ema pies eet everie to krnnyay Cay 177

MME AVATOLEUS 1.66 ooh es we as 171 Cinetochilum margaritaceum....... 182

PUINGEDATACHIOSA. coco c ick leone: ilg(al COCCUAINGIGOSIDIAs ak oe eee 187

ie na Stbiata..oo:<s2see hehe. Lj2ers Colpidimin vic ia ce sal sess atic ieee 183

Amoeba verrucosa............... Lit, Colpidium putrinum).(. 2. 3-< 55. 183

BeMmBeNa VIETAES.. 2. oe. ee es 172 Colpoda cucullulus............... 181

J) 5 0 EV Ais Cot huaniaree varia isis serves hare 189

BeBe PUS (015) 102 <i esge ls cleo ae 130) Cothurnuy cunva (44). sss. a 189

Amphileptus fasciola............. 180° Cothurnia imberbis......5.4..... 189

Amphileptus meleagris........... 181. Cryptobranchus alleghaniensis...... 170

/: SPERSLOTNE 30 Eo tid eae RO cRCaCRT OR 176 Cryptoglena cordiformis........... 176

Antsonema acinus..............- LG ¥ Cucliditmn yen tau eecreole ae Sane 183

Amisonema grande.............:. WOF \Cyclidium glancomanss 1). f2055.5. 183

Ppmrothoracida, .......:..255.:5 173 Cyclidium litomesum............ 184

| ULE CS Ie eee anaemia 172 Cyclidium margaritaceum......... 182

GoM VUNBATIS, ic. ... 2.5.2... 172 Cyclogramma rubens.............. 181

PREM MGAG Riera tect 2 SiS Goss Sluis: MADE AC yphioderiay. aya yas Soils sagt. 173

PAIN ery ative 425.2 e895, geese, he 187 Cyphoderia ampulla ARS TRIN Ss Beatie 17

Repuriised GOStAta s. . 6 6. a. esis ere vs 187

INSERT OS i a oe 175 Dactylosphaerium VItTAeCMs » Jeske 172

Astasta acus.......... 06.2... RAG apn dos clameratrni sy Y.92/5) WLV sav kia eae eae 180

Astasia tricopliora.... 620). 3.05 a. 4: 175 Didinium nasutum.............. 180

FaleS}IC ESI (6 EAC SIR gee Re i PR ee APSE) LIAR Lael sh 9s acihais fh dy oie 172

Difflugia ampulla......5-. 000050. 173

ERED PIGHO SUS 65 (0023205 e285 saic a ene Oe 74 Difflugia ‘constricta.. 00. ..). 2.564. 172

SER PUISEES YS dre. es sseiazais ob 0 Ses 176. “Dithugia pyriformis. ...).5 6.05.5 172

TLE RI ARE CI a8) si erdnie oases Seiad 177

MPP MGSPOIN 65. of). 2) 00s 34 8G sete 180) ePinohacellida ts 54: h.c6 Eels. wae 177

Carchesium epistylidis........... 188

MMMM 39205 o's, sis ahs) dk ee Wiss Wychelinndae nse s nie ON 177

Cercomonas longicauda........... PASM PEMGHEIVSRE DAN) sari 2 3. vtysaiwiate)s eats 178

Cereomonas spy... 2.6... 060.00.-2 P7St Peery S!SPy soo )3)s aye 23.3.4 sees 3 ote 178

196

PAGE

EIpistylis.d ducers pc etenomce once 188

Epistylis branchiophila........... 188

Eipistylis plicatilis.....2.-. 22.25. 188

Eucalia inconstans..........-.--- 170

Belen eee chetesejer alo alee Cush nor ohehae 174

Pugleradecess tte syste eeielaeee 175

Buslena Pyruin. 2002s eee cn ee 2 us 175

Buelenasvibldisen csi y5 carrer a 174

BuplermiGals eee. ple sis e see shee 174

Buclentdactase i ance nese 174

HCogly hae cent ees toe ae els 173

Euglypha alveolata.............. 173

Miuglyplidae. nse eset Aye ee 173

HGtES cme eA e cre tan es a ewiet 186

BplGtes CHALOM. isso we ee «oe 186

Euplotes patella, 3.) 0.2.2). 22 187

LDithol fo até Eos eins Aaicionin aos 186

Bbheptideres wate sacle 6 cre ve sees 175

PEMERE PLA) VATIGIS <<. cle sisi ee ares = 175

WTONtONIA ence eet ees 182

Prontonialeucass yess cele oe © 182

Glaucomasneten ce: oo eae ners 182

Glaucoma margaritaceum........ 182

Glaucoma scintillans............. 182

KGlenodinium’).. 22 ces ere oe seers 177

Glenodinium pulvisculus......... 177

Serdar meen cenieaeae neice cr 188

Gerdavannulata. 52h eee eee 188

Gymnamoebidas si). 6... 2 F<! 171

Hlalterianeesaee cine ac se eae 184

Halteria grandinella............. 184

MA GIT ULE. a5 p< 6 a ins0i s,s heb 2,02 179

Haltertdaenen.ce nas we eee oe 184

ATeOZOR TE eee ee ee eee 173

ieteromastigida. -s.)sc oe eects = ele 174

ieteromitasoo nh. so cseen eee 174

Heteromita globosa.............. 174

Heteromita ovata.............++.- 174

Heteromita Spyies ss sain. fea 174

Heteromitidae!< tia. 6. - 5 cs es 174

Heteromonadidae................ 173

Heteronema a-ceu o- cet teens 176

CHARLES HOWARD EDMONDSON, PH.D.

PAGE

Heteronema acus: oto...) eee 176

Heterotricha n-ne eee ene 184

LISHEIO Ss oe ih sian ee Ce 186

Histrio erethysticus.. 25). 4.2428 186

Holophrtya.oc. 2 oe eee 177

Holophrya ovum...... 5.2.2 a sae 177

Holophrya Sp. )v i.) Soo42. eee 178

Holosticha’..4)..2.3h4e saree eee 186

Holosticha vernalis.............. 186

Holotnchay.. 2chsie8 see ee eee 177

Hopotricha’ . s).:..'. boos Seaeeee 185

Thfusoria’.\. = 420 co oe eee 177

iKerona patella). 0. 9-2 eee 187

Teactymaria 4: Ja-cs. see eee 180

Lacrymaria cohnil.. 72.) seers 180

iacrymaria lagenula.-). +s aoe 180

acrymaria O10r. .-- see eee eee 180

Lacrymaria truncata: /2-e- eee 180

eucophrys.i2.)./)5.1.c ae cece 182

Leucophrys patula............... 182

Lionotus: «5.2 6. gan ls) ae see 180

Tionotus fasciola.-< 2.225... seeeee 180

Lionotus’sp. -.0\.!.:. 0). 2. eee eee 180

Lissoflagellata. . ) .'..: 2222 eee 173

(oxocephalus). : 12 => =. -ee see 182

Loxocephalus granulosus......... 182

Mastigophora ......:.:-.-=eeeeee 173

Mesodinium:. .. - 2): ... 0c) eee 179

Mesodinium pulex.............-. 179

Metopus:......-5.+:s5.5¢s eee 184

Metopus sigmoides.............. 184

Metopus Sp-1....2.554 4 -= eee 192

Monadidas o:.. oc... «ae 173

Monas. 2.00.65) +0 25 lense 173

Monas fluida. ..... +). v.72 eee 173

Monas tirregularis...........+.+-: 174

Monas sp.'......... 5-05 ++. 5-5

Wassula..iis. asc ns cst oh See 181

Nassula.omata:. -\nn4-4 eee 181

Nassula, rubens)...\-2.- tera 181

PROTOZOA OF THE DEVIL’S LAKE COMPLEX

PAGE

INGtosGlenluSs +) shktn te etaee sheen 176

INGEGSOIENUS SPy sc. s. desk see ok 176

PUBATUINGM BS 5 oi- y.os\ anc oes nhc eee 185

Oxytricha bifaria....... PS RN evel 185

Rerviricha fallax... ou kee 2 te 185

Oxytricha parvistyla............. 185

Oxytricha pellionella............. 185

DRPPSTICRIGAG. 2c. c.5.c'i ye ehieale et 185

IPAPAIMAECICAG. oa. o¢ Acre eciere Ss 183

PED GL TENGE Ue ree ee 183

Paramaecium caudatum.......... 183

Paramaccium chrysalis........... 184

PAGATAACCIUM SP), 2.4: 653 seit ec 191

Paramaecium trichium........... 183

Peranemildae:,. 2, 266, ca ee 175

BEICINUICAC ss walt ods Ged eitiva yes 177

[PEDAL LSE yagi ees CE oe eee 187

Rate eTMOMAS 4 cys c soe cheno sels = 175

Petalomonas ervilia.............5. 176

Petalomonas mediocanellata...... 175

IREtAlOMONASISP. fc w- feler ci ee sos 176

EAERGUISME PE Cars sie aia isiase < echals ie Da" 175

_ GTS Te) (hr ee 175

Pimephales promelas.............. 170

SM MIOLOMMNICAC. 2... cis eee ee 184

pleRTONC MIA. = Joos. 08k YeNlee vee 184

Pleuronema chrysalis............ 184

PIEUPONCMG GROSSE..5) 06. ae es 184

PISUTOHENNGCAC 4. 5\).0) << + os os ales 183

TSLiS( Lr ate) JE che Bore eRe eee 186

Pleurotricha lanceolata........... 186

PGMOP HEYA.) slisiases micas aie da weik 189

Podophrya cyclopum...........4. 190

Podophryaidibera.). 325 oes: 189

PodOphTVya/ SP isaac oN 190

Paaophnviddes ya) csNsk sae cones 189

PMSA: 2 os eee 174

Wemrmnstigidae, ........).02 4304.55 174

[ELS S STaE re aie Seat ee Pe ee ae 176

Polgtomauyella. ..2.)3 02022-46284 176

OMvUON Ae) <5 2 Fe cise leieees 176

212100 7514 Sl A eam tr AS Be 178

Prorodon edentatus.............. 178

PAGE

Prorotion griseus. 5 see ae ee 178

PHOFOdOM COLES iA fi.) rete ela lene Sie ore 5 178

Rand pipens ntl ee. Ue ss. . 170

Rhizomastigidaey noted. 3: 173

Rn ZOPOGane eee ia ee aie ere oe 171

Sarcadina® . ix Messen a aadioraes 171

Spathidium\2c ost jase eae oe aes 179

Spathidium spatula.............. 179

Spathidinga, Sp... nce Deen 179

Sphacrophrya. tinier seater ose 190

Sphaerophrya magna............ 190

SPUTOS LOMA fde aces avec chert tere ate 184

Spirostomum ambiguum.......... 184

Stylomyebia wai Ces aerate! aebaren 186

Stylonychia lanceolata...........- 186

Stylonychia notophora........... 186

Styonycelarspep les ierteiale eek < setae 192

Suctoriayeay vee mers scan arate te 189

PachySomay tosses winks cetots 186

Tachysoma parvistyla............ 186

Mestaccak eines Mes Fo os ete 172

Wetranmtidaen ss ove tae see ee 176

Metraselmist 4: eee crmynia canna 176

Tetraselmis cordiformis.......... 176

Sling ees fe oncee ese iai neers 183

Tillina sapropuila. 2 \'oa so. 183

ibrachelinidacwnn secure rte cert 180

Trachelius meleagris.............. 181

Trachelius tricophorus..........-+ 175

ARE PYOIMIONAS sche each delete 174

Trepomonas agilis............... 174

Dir tChedanGharowd sn lelte elena 186

Trichoda grandinella............. 184

DRUGhOMON PAULL Gs aoe ali sh ee 8 182

Trichoda pellionella.........-.... 185

Trichoda sigmoides..........-+.+:: 184

‘Atintncuahien ae pacnoue cosedeo we 177

Undetermined genus............ 177,179

Undetermined species.........-. 177,179

Wired erat ese cha)y d'=\Ggha ls te acess 2 = 185

Berle bs) AIMS io )4) iia) s/ ¢ slates cleo mie 185

198

PAGE

Uroleptus rattulus ), -2)..cis1 ip oe 185

Wroleptusispe:scscut > es Mecerp eee 192

Wronemaracwiat oy coccser eer eeee 183

Uronema marinum...........:.- 183

Wrotnchal tees ce eee 178

Urotnichaylabiatay oper -ee eee 178

WVarinocolane a.iescciic oieceeee 189

Vaginocola crystallina............ 189

Wonticella em. aac aceite eee 187

Vorticella convallaria............ 187

Vorticella microstoma............ 187

CHARLES HOWARD EDMONDSON, PH.D.

PAGE

Vorticella nasutum...........-.-. 180

Worticellajoctavo. =. -- eee 187

Vorticella rabdostyloides........... 188

Vorticella sp. 7. o< 31. i. eee 188

Vorticella,telescopica. i522. -ee8 187

Vorticellidaé:,..c.:... 2528. tt eee 187

Zoomastigophora. «7-255. 2 ees 173

Zoothamnitimeses sree ee eee 189

Zoothamnium alterans........... 189

Zoothamninmisp...).).2 coe 189

EXPLANATION OF PLATES

Prate XVIII

Figs. 1-3. Cercomonas sp. x 1090.

Figs. 4,5. Monassp. x 750.

Fig. 6. Heteromita sp. x 2000.

Fig. 7. Monas sp. «x 1200.

Fig. 8. Monas sp. x 1500.

Fig. 9. Notosolenus sp. x 900.

Fig. 10. Petalomonas sp. x 600.

Figs. 11, 12. Undetermined genus and sp. x 1250.

Fig. 13. Holophrya sp. x 550.

Fig. 14. Uronema labiata, new sp. x 750.

Fig. 15. Enchelys sp. x 1200.

Fig. 16. Spathidium sp. x 300.

Fig. 17. Spathidium sp. x 900.

(Posterior seta omitted in figure.’

PLATE XIX

Figs. 1, 2. Undetermined genus and sp. x 380.

Fig. 3. Lionotus sp. x 800.

Fig. 4. Vorticella sp., including stalk. x 800.

Fig. 5. Vorticella sp., including stalk. x 350.

Fig. 6. Zoothamnium sp., including stalk. x 250.

Fig. 7. Acineta sp., including stalk. x 400.

Fig. 8. Acineta sp., including stalk. x 280.

Fig. 9. Podophrya sp., including stalk. x 370.

Fig. 10. Gerda annulata, new species.

x 500

TRANSACTIONS OF THE AMERICAN MICROSCOPICAL SOCIETY

OILS FOODS

PLATE XVIII EDMONDSON

(9%

TRANSACTIONS OF THE AMERICAN MICROSCOPICAL SOCIETY

VOL. XX XIX

<a AEN TUDE TANT

Sores ere

PLATE XIX

EDMONDSON

AGE, GROWTH AND SCALE CHARACTERS OF THE

MULLETS, MUGIL CEPHALUS AND MUGIL

CUREMA

ARTHUR PAUL JACOT

CONTENTS

PAGt

Lol -SUNCUNTDS 5 Ss0.016 2c RIGA nN a ee PREIS. Aur Weer ee ROE cg 199

MR TENET OLEOU SPECIES re fo. 0.0) 88s Pe nce che Ie ee Se a Ce RL a eee 200

SU mA ceL TU SMIENINT Crete tetra rotor ee ot. Neila. ua lle eat eee a itne) oct. sled te Renan eyalnas 204

MPa PRGENMAIOLMOI My OUR Le oi). ee elar selec rec oeeiaeels es cad sulk Maat taevel eee 204

Mevelopment Of young. =... 2... y...5..... 6% PR Shh ea Tehst i sich eer 204

WIE EIDEITTOERY Sag l SUB Aa Ry cake ne TER At gO en lg ge RU EAD RD Ae 214

BENE EC EOS LOUP Am mit tie cont my ent wets <i" er Si cicarelen eg Haan ah a ee Ry a 220

SLUG. 5 STAGGERED ES PIES aC EPIC Tay ET SE Ge ED RR CRE RID 221

fle obull eparaesrébai: (CURSES ET) SEAR ry 2 SU a 22m ee Or pea Ce 223

“YABY ENTE. 8 oth ade sakt Raye atte OPN Ie og eee ese ea TEA op anaes Lao ie 223

SIT Tret TOTP eae os oy ee est ae earner S ely ee nee Ma tats Ee RU Se 226

ANSHUINIG., ois iS SE Peete NS Ope nD ed SRP AIC eae 2 A AAS a ee a 226

Sv TIERISEVEITEY? 0, Glo EMS Ne SENSED ER GRA MET lg CA care ees ARE oy ee NE Pe 226

MEARE IPO Ree ees) Lee Tie yes SIREN alg hed aay Sah wie Bhgsg aoe Ganga toe Sea 227

SeRMREIRCITIER PLATES oreo nis bests c MMe ean: ap ae Meares aRe esc) co eee cee rate 229

INTRODUCTION

During the summers of 1915 and 1916 the writer was given the

opportunity of studying the rate of growth and development of the

mullet of the Atlantic coast of the United States. The collecting was

done at and about Beaufort, N. C. Of the two species under consider-

ation, the striped mullet (Mugil cephalus) has far the more economic

importance. It ranges through all tropic and warm waters of the

globe and has long been used as food. On our south Atlantic and

Gulf coasts it has been sought so constantly and taken in such

quantities that its numbers have noticeably decreased so that the

supply continually falls short of the demand. For this reason the

artificial propagation of this species is very desirable and it is towards

this end that the present investigation has been made.

199

200 ARTHUR PAUL JACOT

Differentiation of Species

Along the Atlantic coast from New England to Florida, two spe-

cies of mullet may be encountered, Mugil curema and Mugil cephalus,

the latter being much the more common. Commercially, no distinc-

tion is made although the fishermen seem to be aware of two species,

calling the former the “‘silverside”’ or white mullet and the other the

“Jumping” or striped mullet. Other common names are locally used.

When asked wherein they differ, the fishermen give a variety of more

or less accurate answers, and generally end with some statement to

the effect that the ‘‘silverside”’ is very seldom caught. A review of the

fisheries literature on these species shows a lumping of the two, so that

no accurate information concerning their respective habits can be

secured.

Technically the silverside mullet (M. curema) differs from the

jumping mullet (M. cephalus) in having more heavily scaled second

dorsal and anal fins, nine rays in the anal fin in contrast to eight in

M. cephalus, and 38 versus 42 scales in the lateral series and 12 versus

14 scales in transverse (diagonal) series. The field marks are the

scalation of the anal and second dorsal fins and a lack of the longi-

tudinal stripes of M. cephalus.

Because of the unreliability of single characters in species deter-

mination, and because of the possible difference in coloration of

adult and young, a study of the variation of specific characters was

necessary. Relative measurements in these two species are imprac-

ticable as the slight difference in ratios is repeatedly exceeded by

individual variation. The amount of scalation on the second dor-

sal and anal fins is a fairly good character for both old and young fish,

but is relative only. The ¢otal number of rays and spines in the anal

fin is not constant. Specimens of M. curema with a total of 13 anal

fin supports (rays and spines), the last of which may be bi- or tri-

partite, are not rare, while specimens with 11 supports are rare.

M. cephalus has ten supports more often than twelve. Thus, though

quite constant, this character cannot be wholly relied upon. This

leaves the scalation of the two species to be considered.

The mullet is an unusually favorable subject for lepidology

because of the relatively large scales and the presence of the lateral

line groove (without the pore) which is found on nearly every scale,

and which materially aids in the alignment of the scale rows. The

tice

CHARACTERS OF THE MULLETS 201

general results are that the number of scales in the lateral series

varies considerably while the number in the transverse series is very

constant. For instance, although M. cephalus normally has 42

scales in the lateral series, it often has 41 though as many as 44 and

as few as 38 have been found. One specimen was found with 40

scales on the left side and 44 on the right side, two of the extra scale

rows were situated between the base of the pectoral fin and that

of the first dorsal, while the other two were below the second dorsal.

On the other hand a stunted individual 185mm. long whose ratio of

depth in length is 3 instead of 4, has 41 scales in lateral series and 15

in transverse series. Each scale, however, is much shorter along its

cephalo-caudal axis than scales of normal fish. M. curema varies

somewhat less. No variation was found in the number of lateral

rows (number of scales in transverse series). Thus it would seem that

the number of horizontal rows, as in the Ophidia (see Ruthven 1908),

is a reliable species character, or at least more so than the number of

transverse rows. Owing to the difficulty of counting the number of

scales in transverse series of small specimens under a binocular micro-

scope, due to the rotundity of the body and the consequent necessity

of rotating the specimen while counting, this character was found

impracticable for the determination of large numbers of young, and

the scalation was, therefore, further studied. The results are repre-

sented in figure 1, which is reproduced from a fairly typical individual.

In the absence of a lateral line row the median row on the caudal

peduncle was chosen for counting the number of scales in lateral

series. The last scale of this row is short and almost hidden by the

Figure 1. Scalation of the mullet on the side.

202 ARTHUR PAUL JACOT

forty-first scale. The other scales of the last row become more

evident dorsad! and ventrad. In the figure the numerals on the

lateral median line designate the number of the transverse row, those

on the body designate the number of scales in the transverse rows of

that area except where a numeral appears above or below the body, in

which case the numeral designates the total number, but as one of

them is situated on the dorsal or ventral median line, that scale

belongs as much to one side as to the other. All numerals include the

lateral median line (except those upon it). The numerals in parenthe-

ses are the corresponding figures for M. curema, (where omitted they

are not given). The reason for the diminution of the number of

scales in the transverse rows caudad or cephalad is shown in figure 2

Figure 2. Scalation of the mullet on the back.

which is a dorsal aspect of figure 1 with the lateral line grooves con-

nected by continuous lines. Thus when two lines run together, two

scale rows become one row, and where a single line ends, a scale row

becomes crowded out. A similar condition obtains on the venter.

It therefore seems that reduction of scale rows occurs on the dorsal

and ventral median lines—a condition very different from that in

the Ophidia (Ruthven 1908). The exact location of the termination

of a lateral row varies with the individual so that figure 2 is but indi-

vidual and the area between C and D varies in appearance with each

specimen. Likewise, there is variation in appearance between points

A and B and the corresponding section on the venter. The area

between points B and C down to, and including the venter, may be

definitely relied upon as constant. The transition band on the ven-

tral section (the area between 10(9) and 9(8)) is liable to shift caudad

or cephalad a scale or two, but this should cause no confusion.

1 The termination -ad as explained by Wilder and Gage (1882) signifies “towards,”

“in the direction of,’’ etc.

CHARACTERS OF THE MULLETS 203

From this analysis it should be evident that any variation in number

of scales in the horizontal row will shift the /imits of the various

areas caudad or cephalad, depending on the individual, and this in

turn means that only the middle area is unshifting. This middle area

is also so broad that the desired flexibility in counting is given and the

possibility of the complete loss of a row is greatly minimized.

A total of forty-four catches made between December 22 and

September 4, during several years were plotted on co-ordinate paper

using the abscissae for the length of the fish and the ordinates for the

date of capture. Simple lines are used for M. cephalus, and railroad

lines for M. curema and harengus (see fig. 3). The length of speci-

247 257

Sea

es aa

geurede

aceer

tre

Pears

ele

ian

‘a

Figure 3. Record of nan linead mullets;

*tugil cephalus.

te Savaule curems,

mens herein given is the greatest possible length and all measurements

unless otherwise stated are in millimeters.

204 ARTHUR PAUL JACOT

Mucit CepHatus LINNE

Determination of Young

Before anything can be done with the young of M. cephalus, it

will be necessary to go back to Myxus harengus of Giinther. In 1883

this species was established as the type of a new genus Querimana

(Jordan) which differs from the genus Mugil in having the following

characters—a serrated preorbital, thin lips, no adipose eyelid,

stronger teeth and two instead of three anal spines. Bean (1903) has

described the development of a third anal spine from the first ray.

Further investigation has brought out the fact that the adult also

has a serrated preorbital, as will later be described. The condition

of the lips, teeth and adipose eyelid will, in the proper place, be shown

to be but juvenile characteristics. Thus the genus Querimana,

consisting of juvenile mullet, becomes a synonym of Mugil.

The specimens of M. cephalus ranging from 23mm. up to 40 or

50mm. were very carefully examined and were found to be juvenile

M. cephalus, a heretofore undescribed Querimana (having the ‘‘Queri-

mana”’ formula of A. II, 9; scales 42-14).

The specimens running into the M. curema group answer perfectly

to the description given for Q. harengus. The description of Q. haren-

gus (Jordan, 1896), gives it thirty-eight scales in the lateral series,

twelve in transverse series and an anal fin formula of II, 10. Adulting

(changing to adult condition) this fin formula according to the evi-

dence given by Bean (1903) we have A. III, 9. This agrees with

M.curema. The development of Q. harengus further shows it to be

the young of M. curema, as will be shown below and not a distinct

species.

Development of Young

The juvenile stage of M. cephalus begins with individuals as small

as 23mm. Their first appearance is in the form of well developed fish

without the slightest larval appearance. As already described by

various writers, they form compact schools, swimming near the sur-

face of the water. They may be found in deep water, or more often

in water but a few inches in depth. The time of the year during

which they are to be found may best be seen by consulting figure 3.

They might easily be mistaken for the young of M. curema, because

CHARACTERS OF THE MULLETS 205

of the similar coloration. Collections made from December to

March consist of slim silvery individuals of small size. There seems

to be very little growth during this time. The sides are devoid of

pigment, being sharply defined from the dark brownish-green back.

In later March and early April this dark dorsal band is extended down

the sides by the gradual appearance of pigment cells. By mid-

April these pigment cells have so increased as to merge the dark back

into the silvery venter. With this advance in color, the fish rapidly

increases in length and the abdomen is bulged by the developing

intestine. Besides these external evidences of a turning point in the

life history of the species, the growing parts of the fish show this

change, though some more strongly than others. In the following the

juvenile characteristics of this species through the development of

the fish to its adult form, this turning point has been noted and the

reason sought. The lengths of the fish as given below are used for

the purpose of correlating the development of the various parts with

with the fish as a whole, and are of typical specimens. The silvery (or

juvenile) stage is found in specimens from 23 to 32mm. in length,

while those from 30 to 35mm. are somewhat difficult to distinguish

as silvery or dusky because of the merging of the two forms at this

size.

The preorbital in the juvenile stage has some 10 or 12 points, teeth

or serrations, of fair size. As the fish grows these points become more

and more numerous, less slender and less distinct. In older fish they

become blunt and stocky until in a large individual (502mm.) there

were 53 teeth on the margin, crowded so as to place about four to a

millimeter.

The adipose eyelid shows no marked acceleration in rate of growth

at the end of the silvery stage. It is entirely lacking in the smallest

specimens, but by the time the fish has reached a length of 28mm.,

with the aid of a high power binocular microscope, a slight translu-

cent growth can be detected just anterior to the eye. In describing

the state of transparency of the eyelid, it must be remembered that

only alcoholic specimens are being described, in life the adipose eyelid

being perfectly transparent at any age. For the sake of convenience

the eyelid has been divided into three parts: (a) the ring, which is

situated about the rim of the orbit, (b) the anterior lobe, and (c) the

posterior lobe. When the fish is about 30mm. long, the anterior lobe

206 ARTHUR PAUL JACOT

having slightly thickened, has become semi-translucent and has

stretched backward over the eye. At 32mm. length, the anterior lobe

has further thickened and become slightly more opaque, stretching

farther back over the eye and merging into the ring which has just

become visible. By the time the fish is 36mm. long, the anterior lobe

has become opaque, while the ring, which has very slightly stretched

posteriorly, has become semi-translucent. From this stage, the

gradual development of the growth can be easily followed without

the aid of the microscope. At 39 mm. the posterior lobe has become

quite definite while the anterior has thickened and become more

nearly opaque. On 42mm. specimens, the anterior lobe is visible to

the unaided eye. The ring has assumed an opaque cast at its inner

edge and stretched out over the rim of the nearest scales. The pos-

terior lobe has, by now, spread out over the preopercle but is still

translucent. The growth of the orbital ring is now very slow, its chief

expansion being inward over the eye. Ina specimen 47mm. in length

the posterior lobe has become so opaque as to become visible to the

unaided eye. Its lateral growth takes it not farther up or down than

the outer diameter of the ring, while its chief growth is posteriorly

over the preoperculum. The anterior lobe grows no farther forward,

having reached a point just anterior to the nostril, but it slowly grows

out over the eye. At 54mm., the more rapid growth of the lobes has

caused them to overrun the ring anteriorly and posteriorly, so that the

ring now assumes an elliptical shape, the long axis of the ellipse being

vertical. Sixty millimeter specimens show this ellipse more strongly

developed, the posterior lobe much lengthened posteriorly and the

whole eyelid in its typical form, so that it is now only a matter of slow

growth before the adipose eyelid has assumed its maximum develop-

ment in the full-grown adult. Thus it is seen that there is no break

between the juvenile stage and the adult.

The thin lips given as a characteristic of the genus Querimana do

not appear disproportionally thin for so small a specimen. If there is

any relative thickening of the lips beyond normal development, it is

so gradual as to be imperceptible.

That the teeth are slightly stronger in the juvenile young than in

the adult cannot be considered a generic difference in itself, as it may

well be due to a retrograde modification due to change in food habit;

‘as there seems to be grounds to suspect is the case.

CHARACTERS OF THE MULLETS 207

The development of the third anal spine from a ray was described

by Bean (1903) but deserves further comment. The ray is simple

and has about four articulations. At the close of the juvenile stage

Figure 4. Development and convolutions of the intestine of Mugil cephalus

when from 23 mm. to 40 mm. in total length. s=stomach; e=esophagus; a=anus;

c=line of cut of duodenal loop, see p. 208. At figure v the fish is 32 mm. long and

transforming into the dusky stage.

208 ARTHUR PAUL JACOT

this ray ceases growing with the same rapidity as the true rays and

becomes heavier basally, continuing to become relatively heavier

and stiffer, until it is about one-third of the space between the tips

of the second spine and the first ray, longer than the second spine

spine (Fig. 1). This relative length is maintained throughout life.

As this spine continues growing and thickening, the articulations

become obliterated until lost so that in adults the third spine is

basally as heavy as the second and quite equal to it asa spine. This

development should be of much interest to the morphologist and

systematist.

The reproductive organs are so rudimentary as to be invisible

throughout any part of the season or at any point in the juvenile

stage. This might be sufficient reason in itself for discarding the genus

Querimana.

The development of the intestine gives further evidence of the

relations of the two forms. Owing to the difficulty of describing this

development a series of outline sketches (Fig. 4) have been prepared,

illustrating, by a lateral aspect, each successive change.

The most simple form (fig. 4, I) consists of a duodenal loop and a

“straight-away”’ to the anus. When the figure V stage is reached the

fish is passing into the dark or dusky stage, the kink m having leng-

thened into a loop whose lower member has twisted upward and over

its upper one to form a loop. At the next figure (VI) the spleen

appears as a yellowish body, 25mm. in diameter, and from then on

becomes a factor in influencing the convolutions of, the intestine.

At the figure VIII stage the duodenal loop makes a kink which soon

becomes a loop and thus destroys the duodenal loop in its typical

form. Each odd figure from VII to XV shows the convolutions on

the further or inside by the cutting away of the duodenal loop or its

modification at line c. Beyond the stage shown in figures XIV-XV

the convolutions become so intricate that their study would surpass

the scope of this paper, the length of the fish at this time averaging

about 40mm. Thus, in the lengthening of the intestine, there is a

marked acceleration in the rate of growth at the time when the fish

is about 32mm. long, i.e., when the fish is passing into the young or

dusky stage.

Besides this development in length and complexity of the intestine

proper, the whole abdominal cavity is eloquent of the change exter-

CHARACTERS OF THE MULLETS 209

nally noticeable. To appreciate this change, it is necessary to begin

with the earliest individuals. All December specimens examined had

their entire viscera and the walls of the abdomen colored orange,

while the peritoneum in many cases was grayish with dark spots,

otherwise it was of a semi-translucent blackish color. The length of

the intestine was at times half that of the individual itself, though gen-

erally about three-quarters its length. In January, the coloring of the

intestine was the same as for the previous month with the exception

of a few individuals in which it was yellowish while the peritoneum

averaged darker, and the walls of the abdomen a little lighter. The

length of the intestine showed a slight increase over specimens of

corresponding lengths of the previous month. In February the vis-

cera were yellowish to pale, a very few individuals having traces of

vegetable matter in the intestine. The peritoneum showed no special

change, while the walls of the abdomen were pale. The intestine, on

an average, had increased in length to a slight extent, but in no cases

equaled the length of the individual. In late March, quite a few

specimens had entirely lost their internal orange or yellow color and

the intestine had traces of dark matter. The peritoneum was black

and the flesh about the viscera had assumed the more natural dark

coloration. These specimens showed marked increase in length of

intestine, it being considerably longer than the individual. These

fish were passing into the dusky stage. The majority of specimens,

however, were much like those of the previous month. The viscera of

April specimens are rarely orange or yellow tinged, the great majority

having the intestine more or less filled with dark matter. The length

of the intestine had also correspondingly increased. These fish were

well into the dusky stage, their intestine appearing as represented in

VII to XV of figure 4. It was from these slowly developing indivi-

duals that material for figure 4 was taken. From this time on the

growth of the fish is very rapid in comparison with that of the previous

month. Thus the increased length of the intestine can be directly

correlated with its own color.

An explanation for the change in visceral coloration described in

the preceding paragraph was sought by examination of stomach con-

tents. The silvery-sided individuals (juvenile fish) showed an almost

exclusive diet of crustacea, mainly copepods, and as alcohol almost

- invariably turns this form of life a salmon red, the coloration of the

210 ARTHUR PAUL JACOT

viscera is accounted for. The intestine as well as the stomach were

filled with this food, the latter not yet having reached the gizzard

development. In the most immature individuals the stomach’s form

was that of a simple sack. The stomach contents of the dusky stage

consisted, roughly, of 40% sand and mineral matter and 60% vege-

table and animal matter. This latter consisted of 50% diatoms,

35% algae and other soft vegetable matter and 15% animal miscel-

laneous. This seems to be the usual ration of the fish during the

remainder of its life, it being known as a mud feeder (See also Linton

1913).

From the above, two things should be apparent, namely (a) that

the first form (the juvenile or silvery stage) develops into the second

(the dusky stage), (b) that the juvenile stage is one of slow growth

and development which is more rapid after the fish has changed diet,

(made evident by the change in the color of the viscera). Because the

intestine, the stomach and the whole fish acquire an acceleration

in rate of growth and development at the time of change of diet, we

conclude that this period of change is due to change of diet.

A study of the development of the scale along with the development

of the individual is essential to the correct understanding and inter-

pretation of the adult scale. The simplest form procurable is found

in the juvenile or silvery-sided individuals (figs. 8-9). Text figure 5

represents one of these scales (in its natural position) divided into

areas using Masterman’s (1913) method. Rather than deal with the

axes, greater convenience is found in using the areas formed by these

axes as designated in figure 5. The terms “dorsal” and “ventral”

have not been used as no need was found for this differentiation of

sides. In the scale work here presented the scales were taken from

the lateral median line on the two rows originating at the latero-

anterior edge of the first dorsal fin (scales 10 and 11 of figure 1),

except in the very young, where this was done as nearly as possible.

For convenience, the appearance of the scale is described by means of

a formula in which a,/ and / stand for the anterior, lateral and poster-

ior areas respectively, while the number following each of these refers

to the number of circuli in that area. Thus, the formula for the scale

of figure 5 would be a. 11; 1.0; p. 22. The two lateral areas generally

differ in number of circuli when these are present; the average has then

been taken. In the more advanced stages of growth the number of

CHARACTERS OF THE MULLETS 210

av Pp

Figure 5. Advanced silvery stage scale, x45 divided into areas. a. v.=antero

ventral axis; p. v.=postero-ventral axis; a. d.=antero dorsal axis; p. d. =postero-dor-

sal axis; ].=lateral areas; a.=anterior area or basal end; p.=posterior area or apical

end. Formula=a. 11, 1. 0, p. 22.

circuli on the lateral area was computed by finding the average from

two or three scales (taken from the place above mentioned from one or

both sides of the fish). This does not affect the general result as

whatever variation is shown in these four scales is about as great as

the difference between the scales taken from corresponding places of

two different fish of the same size from the same school. In other

words, it was found that the average number of lateral circuli of scales

10 and 11 from fish number one was more constant than the average

number of lateral circuli of scales 10 and 10 from fish number one

and two (both being of the same size and from the same school).

With this, it must be remembered that the number of circuli does not

represent the number of “days” of growth, but that they testify to the

approximate development of the individual. The formula, then, is

useful in conveying a fairly good idea of the size and amount of

development of the fish from which it was taken.

The smallest fish have a scale already well developed, a 23mm.

specimen having a circuli formula a.7;1.0; p. 15 (fig. 8). This being

212 ARTHUR PAUL JACOT

the type of scale showing the least growth of any procurable, it is

inferred that the greater portion of this scale was formed at the

spawning grounds. Notice on this scale, (a) that the circuli on the

posterior area are much closer together than those of the anterior area,

(b) that the lateral areas are without circuli and (c) that the circuli

nearest the center are farther spaced than those further out. As the

scale enlarges, more circuli form on its anterior and posterior edges,

until the scale has reached a maximum development of a. 11; 1.0; p.

22 on a fish of 32mm. length. This type of scale may be found on any

silvery-sided individual, i.e., during the months of December, January,

February, many in March, anda few in April. The addition of circuli

during this season is very slow, so that the scale in three months’ time,

shows no more advance than illustrated in figure 9. Very often the

scale shows less development than this before the juvenile stage comes

toa close. From this point, the method of growth of the scale com-

pletely changes. Figure 10 shows a scale whose formula at the silvery-

side stage was a. 10;1.0 p. 20. A little later two closely spaced

anterior circuli were added, and, while this was going on, the tenth

anterior circulus stretched back along the outer rim of the scale, thus

forming a lateral circulus, so that the formula of the complete scale

has become a. 10+2;1.0+1; p. 20, and the posterior edge of the scale

shows a narrow border without circuli. In the next figure (11) three

anterior circuli have been added to the juvenile scale, two of which

have become lateral; the posterior border is quite wide but without

definite circuli. Note the shallow depression just posterior to the

center. This is the beginning of what is, in some fish, known as the

lateral line groove, and in this paper will be referred to by this term.

Figure 12 has two more anterior circuli, an additional lateral circulus,

a broken or fragmentary posterior circulus with suggestions of a

second, and a larger lateral line groove. Note the fine reticulations

at the anterior edge of the posterior circuli. The fish from which the

scale of figure 11 was taken had a length of 38mm. The circulation

is still further advanced giving the formula a. 9+10; 1.0+4 or 5;

p.19+1(or 2). The lateral line groove almost obliterates the first few

posterior circuli and the reticulation or veining is more extensive and

better developed. This series should clearly show the way the scale

changes its habit of sculpture (habit of growth of the configuration of

the surface). Figure 14 shows a scale from a 45mm. individual and

CHARACTERS OF THE MULLETS 213

gives the effect of this new development. The juvenile scale is seen

to be completely encircled by the later more rapid addition of circuli

which ¢end to be continuous. Thus, the scale is of the cycloid type.

Note, (a) that the anterior circuli are almost twice as numerous as

the lateral, their ends terminating near the anterior axes, (b) that

the anterior circuli of the outer series curve in the opposite direction

to those of the juvenile scale, thus making a definite demarkation

between the two scales, (c) that in the outer series the closely spaced

circuli are anterior while they are posterior in the juvenile scale, and

vice versa with the widely spaced circuli, so that in this respect the

habit of sculpture is reversed. This change, although taking place

at the same time as a change in diet, and occurring during the months

of March and April, is not due to seasonal or dietal change, for the

scale of M. curema passes through the same change at a different

season (during the summer) and unaccompanied by a change of food.

It is therefore inferred that this change is due to some previous change

in habit of growth of the scale, i.e., the change is phylogenic.

During this development the juvenile scale which is designated by

various authors as the nuclear area, nucleus,” centrum, initial field,

etc., occasionally passes through a process of deterioration of surface

face sculpture. This begins with the veining just anterior to the

posterior circuli (figs. 11-14) spreading farther and farther until the

lateral areas are covered (figs. 17-19). When the lateral areas are

fairly well filled in, the posterior circuli are gradually replaced by the

veining so that the veined area is linear or ovate in shape and not the

shape of the scale. A process giving a similar aspect has been des-

cribed and accounted for by Dahl (1911, p. 11-13).

The addition of circula continues more or less regularly for a longer

or shorter time according to the individual. Figure 15 illustrates a

scale taken from a 60-70mm. specimen, and serves to show the

nature of growth of the apical or posterior circuli. Note the way in

which the posterior circuli are bending out toward the apex of the

scale. With the bending out of these circuli the scale grows more

rapidly at the apex and on this posterior lobe narrow, pointed, pos-

teriorly directed cteni gradually rise from the surface. These cteni

are firm and strong, much longer than wide, slightly bent to give

2 It seems preferable to reserve the term “nucleus” for the structural center of the

scale as used by Cockerell (1913).

214 ARTHUR PAUL JACOT

more rigidity, and sharply pointed (figs. 20, 25, 27). These cteni

continue to form row after row, the scale taking on the appearance

of the one illustrated by figure 20. This scale (removed from a fish

taken on the 23rd of August) contains all the characteristics of the

species although the individual was but 145mm. long and not yet one

year old. The lateral line groove has extended backward to the post-

terior margin of the juvenile scale and forward as a narrower channel

to its anterior margin and to the posterior end of one of the basal

radii. This linear shape is that assumed by the lateral line in the

adult. In figure 22, although the scale shows nearly the same amount

of growth, the cteni have not as yet begun to form. Before the further

development of the scale is noted, it will be necessary to review what

is known of the migration of this fish.

Migration

The earliest reliable information we have concerning the migra-

tion of the mullet is a note left by Dr. Yarrow (Smith 1907) on the

fish in the Beaufort region in 1871. The substance of this note rela-

tive to migration is that small-sized individuals appear in May, and

that in later August fish commence to school preparatory to migra-

tion. He says:

The schools appear to come from the northward through Albermarle, Pamlico,

and Core sounds, gradually working their way southward. Their departure through the

various inlets seems to depend upon a favorable state of the wind, which should be

from the northward, for it has been noticed frequently that when the wind hauled, the

schools of mullet already without the harbor have suddenly turned, re-entering the

inlet, and pursued their course southward through Bogue Sound.

A few years later the U. S. Commission of Fish and Fisheries sent

out Mr. Ravenel (1887) to find out what he could about the mullet.

The method pursued was to visit various fishing centers and consult

the fishermen. The only reliable information we need note is that at

Beaufort three ‘‘runs’”’ were noted as follows:

small mullet 4-5 inches June—Aug. 30.

fat ve Sept.-Oct. 10.

roe y Oct. 10—Nov. 15.

The same year the Commission issued its comprehensive work

on the fishery industries in which there are two papers on the mullet.

The first one (Goode 1887) treating of the natural history will not

CHARACTERS OF THE MULLETS 215

be considered as it is based almost entirely on hearsay but on the

second (Earll 1887) which is much more comprehensive and reliable.

From under its caption ‘“‘movements” the following general notes

have been extracted:

. . . Small sized individuals are scattered about on the feeding grounds in the

grassy bays and marshes bordering the coast. Here they remain till late in July,

when they proceed to the deeper channels of the larger bays, where they gather in

schools of small size. Little is known of the whereabouts of the large mullet at this

season. Later the migrations begin, the fish of medium size moving southward. Their

places are soon filled by large fish. . . . These (roe mullet) remain until the first cold

storm occurs, when they start for the south, moving rapidly along the outer shore, or

through the inland passage. These fish are followed by smaller individuals known as

“frost mullet,” which remain through the greater part of the winter. The movement

seems to be general along the entire coast, all fish along the Atlantic seaboard being

reported as traveling southward, while those rounding Florida Keys continue their

coastwise migrations, gradually working northward and westward towards the Texas

line. No return movement is reported at any season along the Atlantic. . . .

In N. J. waters the mullet make their appearance in schools about the first of

September, gradually working southward and entirely disappearing by the last of

October. The same is true for the coast between Cape May and Cape Henry, including

the waters of Chesapeake Bay.

The small fish are seen in June on the N. C. coast, these gradually increasing in

numbers until the first of August, when the schools have attained considerable size,

but thus far no tendency to migration is noticeable. A little later a southern movement

begins, and school after school passes, the size of the individuals constantly increasing

till the first of September when the old or roe mullet arrive. . . If the weather con-

tinues pleasant they remain along the shores until the eggs have become well developed

before moving southward, but at the approach of the first cold storm they are off and

other smaller individuals follow in their wake, so that by the first of January the

greater part have disappeared. Comparatively few are seen from that date until the

following June, though scattering ones may be taken at any time.

At Wilmington [N. C.] small mullet are occasionally taken at any season, though

they are abundant from June to September only, and large ones are seen only in the

fall. As at Beaufort, the migration begins about the middle of August. The first

schools are composed of fish of medium size. . . . By the first of September these

have entirely disappeared, and their places have been taken by the ‘fat mullet.”

These are very abundant for several weeks, the roe mullet arriving about the middle of

October, before they have entirely disappeared. ‘Frost’ or “inch” [the distance

between the eyes] mullet, as they are sometimes called, follow in large, compact schools,

the last disappearing about the middle of December. Smaller fish, called ‘‘winter-

mullet,” are abundant till spring. . .

At Charleston the run is somewhat similar to that at Wilmington.

In East Florida, especially the St. John’s River, fish of all sizes may be seen at any

time. :°. .

216 ARTHUR PAUL JACOT

In the Gulf of Mexico it is claimed that the mullet are even more abundant than

along our Atlantic coast. . . . They are never entirely absent, though, as on the

Atlantic coast, they are much more abundant in the fall than at any otherseason. .. .

From the evidence at hand it is clear that the mullet fisheries for

different parts of West Florida continue from the middle of August

to the first of January, though the height of the season, for most

localities, is in October and November. Farther west the fish seem

less inclined to migrate, remaining more constantly in any given

locality, and on the Texas coast it is said that there is no special

time of abundance, but that mullet are equally plentiful at any

season.

Notes on Wood’s Hole (Smith 1897) state that M. cephalus is

“Found from September to end of October, going in large schools

about October 1.’’ For the same region Sumner (1911) reports M.

cephalus as ‘‘Present from July to December; most common in the

fall.’ In summary, Bean (1903) states that about New York the

earliest appearance of M. cephalus is in August when they are few,

that in September they are found in the New York markets and that

“the great schools were absent till October.”

The two most striking facts brought out by this literature are

those of a fall migration and the almost complete absence of large

mullet on our coast during the later winter, spring and early summer.

This migration seems to begin at the northern extremity of the range

of the species and extends southward with the migrating fish. The

migration seems to be orderly, deliberate, and in series, each series

being made up of a certain age group, almost the whole coast load of

mullet slipping around the peninsula of Florida and along the gulf

coast before all have scattered through the more torrid water which is

the real home of the mullet. Thus, there can be no question about a

definite fall migration down the Atlantic coast to warm water.

Another thing to be noticed and bornein mind is that the migration is

slow and leisurely, taking at least three months, so that it would

seem that each individual had time to live at its leisure on the way

south. Finally, notice should be taken of the lack of any noticeable

northward migration. Thus nothing is known of the fish from the

time it reaches the gulf until it reappears in late summer. There can

be no doubt that the fish does not return north during the winter,

but that it is living in southern waters where it can feed unrestrain-

CHARACTERS OF THE MULLETS 217

edly. After the winter therefore, in spring or early summer, this species

must return north. For a possible record of this period of the life

history of the fish, the scale may again be studied.

At the time that the young are from 40 to 60mm. long or about the

beginning of May, individuals of another age-group, as small as

120mm. in length and up, make their appearance. These individuals

keep increasing in size and numbers throughout the summer so that

by the end of August they are very common and range from 220-

370mm. in length. Their scales are all characterized by the single

“Tine” or break in the continuity of the circuli typically illustrated in

figure 25. The fish from which this scale was removed was taken

on July 2 (1915) and had a total length of 218mm. Notice (a) the

deterioration of the sculpture in the center, (b) the ctenoid area and

the position of the sharpest and the most worn teeth, (c) the unpored

lateral line groove, (d) the continuity of the “‘line’’ from the lateral

area posteriorly to and into the ctenoid area, and anteriorly across

the anterior area, (e) that the “‘line’”’ is formed (1) laterally by the

termination of the circuli in exactly the same way as they are ter-

minated at the outer edge of the scale, and (2) anteriorly by the

termination of the circuli in exactly the same way as they terminate

at the anterior edge of the scale, (f) that this “line” is the so-called

“‘winter-line”’ and (g) that the circuli within the “‘line” are all equally

spaced. With the last three points (d, e, f) in mind as well as the fact

that this is a south wintering fish, let us consult the scales of fish

which remain in cold northern waters during the winter. Good

illustrations of such scales have been published by Gilbert (1913),

Mastermann (1913), Lea (1913), Nilsson (1914) and Hjort (1914)

of the salmon, herring, mackerel and cod respectively. In the scales

of the salmon and cod, close examination will reveal that the winter

area is formed by the crowding together of the circuli (the circuli of

the cod are broken into dashes). The mullet scale is entirely lacking

in the crowding of circuli, testifying to undimished feeding during the

winter. The herring and mackerel scales, due to non-concentric

circuli on the older section of the scale show an entirely different type

of ‘‘winter line.” In this case it is formed by the pinching out of the

circuli. Thus they cannot be used to compare with the mullet.

Now, since the mullet is not affected by winter conditions and does

not show the typical winter crowding of the circuli, another cause

218 ARTHUR PAUL JACOT

must be sought for the break in the sequence of the circuli which does

occur. As already pointed out, this break is exactly similar to the

break caused by cessation of life. The break is sudden and complete.

We advance the hypothesis that this line is caused by a spring migra-

tion differing from the fall migration in being made (typically) by a

continuous run and not by a slow gradual shifting as in the fall.

Various types of these “‘lines’’ or linea may be encountered. Figure

25 illustrates its more typical and usual appearance, i.e., when the linea

is similar to the periphery. An occasional type of linea consists of a

straight but wide space between some two lateral circuli. In one

scale examined practically all the pre-migratory lateral circuli had

slightly shifted laterally and continued posteriorly as post-migratory

circuli. This may have been due to a migration of such a nature that

growth was retarded, not entirely stopped. Figure 24, if closely

examined, will show two closely spaced lineae, the outermost being

the most distinct. Such a form is occasional and may be due to a

second migration several weeks after the first, the fish going still

further north. Thus, the actual number of lineae cannot be absolutely

relied upon for the age of the individual Furthermore, one cannot

consider every linea a migration line as any cessation in feeding or

growth for any reason whatever, might cause the interruption and

renewed growth of the scale necessary to form a linea. Therefore,

though the actual number of linea is not always reliable for the

determination of the number of seasons which the individual has

passed through, the linea may be relied upon for age determination

when properly understood. Before this can be done, however, the

development of the scale of the species must be studied along with

the development and life history of that species.

The above mentioned hypothesis seems to be further substan-

tiated when one notices that specimens from 129 to 257mm. long

(clearly of a second age-component (fig. 3) having as many as 70

lateral circuli outside the juvenile scale) were taken in March. The

lateral circuli of the scales of these individuals were all evenly spaced

3 From the Latin linea, -ae, f; using the term in its more figurative application.

I introduce this new term to specifically label the definite feature of the scale typically

illustrated in figure 23 and explained above, restricting the terms peronidia, annuli,

winter band, annular ring, etc., to the area of circuli between the lineae or between the

first linea and the juvenile scale.

CHARACTERS OF THE MULLETS 219

and no linea of any kind could be detected, yet they were passing

through or had just passed through the winter, evidently at or in the

general vicinity of Beaufort. Thus, all mullet do not leave our coast,

those here in winter having probably come from much farther up the

coast. Furthermore, no fish with a single linea were taken before late

April which could at all be considered of this second age group (or

younger). The fish scale from which figure 23 was taken was removed

from an individual taken on the 28th of April. The linea is some

three circuli from the margin of the scale, thus setting the date of

migration during the earlier part of April. Other specimens taken

during the spring have the following number of post-linea circuli:

May 2—2 and 4

May 44

May 11—7

May 12—0, 3, 6, 6, 7, 7, 7, 10

May 25—3 and 10

Allowing an average accretion of five circuli per month, this data

gives early April as the norm of migration. That it is general and

fairly definite is brought out by figure 6, which is the record of the

Hara ees

ae ine ae |

ee a

Figure 6. Record of Postmigratory annuli of Jumping Mullet caught July 10,

1915.

number of post-lineal circuli of a catch made on the tenth of July

of one linead mullet. The fish being all taken on the same date, any

variation of the date of migration should be shown by the number of

circuli. The abscissae give the number of circuli and the ordinates

the length of the specimens; the points of greatest magnitude desig-

nate three specimens recorded at that point, etc. Although there is

a variation of 14 circuli, or nearly three months, it is not necessarily

all due to difference in date of migration, for individuals vary in rapid-

220 ARTHUR PAUL JACOT

ity of accretion of circuli, i.e., in a given time one individual may

acquire x circuli while another would acquire x+3 or 4. However

great or small a variation in time there may be, figure 6 clearly

shows that there is a definite time of migration which, as has already

been shown, takes place in earlier April and normally consists of a

single continuous run from southern feeding ground to more northern

waters. That the fish migrate in deep water off the coast seems

evident from the fact that the fishermen are unaware of such a move-

ment and that the fish is practically neglected by them until the fall

migration.

Second A ge-Group

The arrival of the jumping mullet in April marks the beginning of

its second season on our Atlantic coast; its age ranges from 14 to

17 months and its size from 120 to 200mm. (5 to 8 inches) (see figs. 3

120 140 260 280 320

Figure 7. Record of one linead mullet.

and 7). ‘The individuals are scattered about on the feeding grounds

in the grassy bays and marshes bordering the coast” (Earll 1887).

They can be found in small numbers over any mud bottom, mud

flats, etc., where vegetable plankton is abundant. Specimens may

be secured at any time during the spring and summer but they are so

scattered as to make fishing for mullet alone an expensive proposition.

Living thus they grow to a length of from 225 to 325mm. by mid-

August. Their flesh is very soft and oily, hence their name “‘fat

mullet.”” By this time they have begun to gather into schools of ever

increasing size, the social instinct becomes dominant as the reproduc-

tive organs rapidly develop. By late September the southward

CHARACTERS OF THE MULLETS 224

migration has begun and as the fish move down the coast and the roe

ripens, they spawn. Out of a batch of ten roe mullet purchased at

the Beaufort market on October 9, 20 and 25, four had but a single

linea as follows:

Length of fish 410, Scale formula 1.68-4-504

414 1.63-++-45

426 1.57--57

431 1.69-++57

The remaining six each had two migration lines giving the following

scale formulas:

Length of fish 426, Scale formula 1.42+60+28

440 1.51-+44+421

454 1.43-+-45-+ 24

473 1.40-+50+ 28

483 1.47-+55+424

493 1.60+46+ 23

The small number of circuli acquired during the third year indicates

that the rapid growth of the fish had been partially checked by having

attained sexual maturity—as is usually the case. If the series exam-

ined was typical, and every effort was made to make a very general

choice, this would mean that the jumping mullet generally attains

maturity and spawns for the first time in its second year. That this

is not always the case is evident from the scales of a male 392mm.

long with testes 34mm. long and 7mm. wide, taken on July 12 (1915).

The average number of lateral circuli for two or three scales reads

1.38-+-72+8, which means that it was spawned late in the season,

that it probably migrated north early the first year, late in the second

and in its third year was maturing early. No other fish was taken

with reproductive organs so far advanced, so early in the year; the

time of the year for organs of that size normally being in mid

August.

Adults

According to scale evidence, the majority of jumping mullet breed

for the first time during their second year. At this time they average

4 Asit is unnecessary to mention the number of circuli in the anterior area or in the

juvenile scale, the formula for older fish need include only the number of circuli in the

lateral area using a + sign for the linea. .

222 ARTHUR PAUL JACOT

less than a foot and a half, and constitute the great bulk of the mullet

fishery. The largest mullet that has come under our observation is

a 502mm. (20 inch) roe mullet whose scale (fig. 26), shows it to have

been in its fifth year. Some individuals are reputed to attain a

length of two and a half feet and a weight of ten pounds. What may

be said concerning the average adult mullet applies equally well to

the larger individuals. Those which return in the spring pass the

time in the marshes, mud flats, and mud bottoms of the wide shallow

estuaries, sounds, etc., so characteristic of our sunken and inundated

Atlantic slope, at least as far north as Cape Cod. The colder tem-

perature and rugged coast extending from Maine northward forms

a highly efficient barrier for such a highly specialized fish as the

mullet. In its spring and summer feeding grounds it can thrive

secure from man for it is so scattered as to make seining unprofitable

and it isina practically inaccessible locality due (a) to the soft muddy

bottom in which it seeks cover and in which man sinks so as to make

seining impossible, (b) to the reeds and grasses over which the lead

line will continually rise and allow the fish to run under—not to

mention those which clear the floats with three to eight feet to spare,

and (c) to the inaccessibility of the locality to power boats. Such is

the choice feeding ground of the mullet, and such is the locality from

which this fish returns to deeper water, fat and full, to enjoy a more

gregarious and social life. As the schools increase in size and the

temperature of the water lowers, their reproductive organs having

developed, they move slowly down the coast en masse both outside

and inside the Banks, spawning as necessity demands. At Beaufort

roe mullet are rare in September, common in October, abundant in

late October and early November, and rare in December; they are

caught both inside and outside the banks, though (according to the

fishermen) never with the eggs running (prime ripe); while spent

mullet are found wherever mullet are to be found. Some of the fisher-

men attribute this lack of ‘‘running” roe mullet to their going out to

sea to spawn while others claim that they spawn in fresh water

because the young are found there (although they are equally abun-

dant all the way out to well beyond the shore line). Thus nothing is

known of the spawning grounds of this species and therefore of its

eggs or larvae, the earliest stage known being the already well devel-

oped young described at the beginning of this paper.

CHARACTERS OF THE MULLETS 223

MuciL CurEMA Cuv. & VAL.

Young

The young of the white mullet, as above shown, is the so-called

Querimana harengus, and is undoubtedly found as far north as

Wood’s Hole. At Beaufort they have not been recorded earlier than

May 25th, but there is reason to believe that they could be found

even as early as late April. In habitat and habit they are similar to

M. cephalus.

The development of this species is, in general, like the former,

but without a definite silvery stage and with a constant rate of

development of the various parts and of the individual. The smallest

specimens normally procurable are 20 to 2imm. long and as much

developed as are 23mm. specimens of M. cephalus. At this least size

the alimentary canal contains no trace of the crustacean diet so

characteristic of the other species, their stomachs being filled with the

dark mud matter on which they continue to feed. Aside from this

difference the two species are similar in their juvenile characteristics,

i.e., they have cyclid scales, no adipose eyelid, and but two anal fin

spines.

The development of the scale though mainly similar to that of the

striped mullet is interestingly different. The juvenile scale differs

from that of M. cephalus in a tendency toward one pair less of basal

radii and in tending to have lateral circuli connecting anterior and

posterior circuli (figs. 16-19). The lowest formula found was a 10,

1.0, p. 14, thus being more advanced than corresponding M. cephalus.

As these juvenile fish acquire their adult characters the habit of

sculpture of the basal area of the scale changes in the same way as does

that of M. cephalus. The development of the apical portion of the

scale, on the other hand, is strikingly different. In M. cephalus

the lateral circuli generally extend backward following the contour

of the juvenile scale until they meet and thus form about it a close

fitting frame. This is so foreign to M. curema that it only rarely

occurs and then only with the first circulus. The second one in

extending backward tends to diverge from the first, the third from

the second though possibly less, and so on (figs. 16-19). This occa-

sionally occurs in M. cephalus scales (fig. 15) but only with a few

circuli. The typical lateral circulation for M. curema scales is this

224 ARTHUR PAUL JACOT

divergent type but without the close-fitting lateral circulus, the very

first one forming an acute angle and terminating very briefly at

the edge of the juvenile scale or continuing through it as an apical

circulus. Meanwhile each apical circulus has done one of two things,

it has entirely stopped growing or it has continued to grow. If all

apical circuli cease growing at the same time another apical circulus

may form about them as above described (figs. 16-19) and thus

very definitely mark off the juvenile scale as in the other species,

but, unlike it, this new circulus is close to the juvenile scale and

immediately followed by others so that the apical circuli are much

more closely spaced than in the jumping mullet. (Compare figs.

16-19 with figs. 12-15). If all the apical circuli of the juvenile

scale continue to grow in full strength and unchanged direction

(of very rare occurrence) the apical boundary of the juvenile scale

is undiscernible. Although these circuli will continue to extend

across the transition line between the juvenile and young scale,

until they meet lateral circuli or reach an equivalent distance, they

generally become thin at the transition line, or, in rare cases, become

obsolete at that point, (figs. 16, 17). Accompanying this weak-

ness of growth the circuli will often become curved or more widely

or irregularly spaced at the transition line, so that the boundar-

ies of the juvenile scale are plainly discernible. The first few apical

circuli of the juvenile scale never run beyond it, extending only

to the line of the posterior axes where they occasionally turn and

become lateral circuli. In the great majority of juvenile scales

all apical circuli do not pursue the same course, so that the scales

present an enormous amount of variation on the transition line (figs.

16-19). For this reason it is very rare when the juvenile scale is not

set off from the remainder of the scale posteriorly, while it is always

discernible anteriorly. When apical circuli meet lateral circuli they

do so at an acute angle thereby forming a type of circulation quite

characteristic of the scale of this species (figs. 17-19, 21). Figure 16

shows such an angle just formed, another about to form, and another

some distance from forming. Thus, although there is not so striking

a transition in the scale of M. curema as in M. cephalus, yet there is

a change so marked as to be unexplainable. It is certain that this

change in the scale sculpture is not due to migration for all stages of

the change, and scales some time before the change would take place,

CHARACTERS OF THE MULLETS 225

are procurable as long as juvenile fish are obtainable, and further, the

change is not merely a seeming cessation of growth for a short period,

but a complete change in sculpture habit; nor is it due to change in

diet for the intestine contents of the fish before and after the change,

in the scale, are alike. Thus again the change seems to be recapitula-

tory or phylogenic. A factor in the destruction of the central sculp-

ture, and more so than in the other species, is the spreading of the

lateral line groove (figs. 17, 18).

After a various number of apical circuli have been formed (general-

ly more than in M. cephalus) a break appears at the apical center in

which cteni are formed (figs. 18, 19, 21). These cteni are added and

develop much as in M. cephalus, but have an entirely different appear-

ance. Instead of being narrow, slightly curved, keeled, and sharply

pointed as in M. cephalus, the cteni of this species are wide and flat

with a very inconspicuous keel at the apical end (figs. 27, 28). More-

over, the cteni in M. curema practically all appear in a well defined

projecting band while in M. cephalus they gradually merge back into

the old worn stubs of former teeth called by Cockerell (1913) ‘‘apical

marginal elements” (herein, for brevity, called ctenobasii), and do

not project as a well-defined band beyond the normal outline of the

scale except in very advanced scales (fig. 26). In figure 27 notice

how the ctenobasii seem in places to be broken up circuli and in others

worn down cteni, as though the cteni were modifications of the cir-

culi. In M. curema (fig. 28) the transition is not so gradual, the fringe

of cteni seeming quite segregated from the remainder of the scale.

The ctenobasii, however, are present in even greater numbers than

in the other species and although they do not seem to be worn down

cteni they occupy an area once covered by them (figs. 18, 19, 21).

They must therefore be considered deteriorated cteni and noted as

another difference between the two species. The cteni are added

row after row along with the circuli throughout the summer until the

fish have reached a maximum size of 230mm. in September when they

migrate south. Figure 21 is from a scale of a 121mm. fish taken on

the 23rd of August, and shows all the characteristics of the scale of

this species. Compared with figure 20 (the corresponding scale of

the other species) the radii are seen to be fewer in number. This is

constantly the case. Both these scales having been taken from the

same position on the fish’s body; this difference is a real specific

226 ARTHUR PAUL JACOT

difference. The lateral circuli are also more closely spaced in M.

curema than in M. cephalus in scales of equal size. This does not

mean that one species accrues circuli at a greater rate than the other.

Migration

In the fall the scattered individuals and small schools gather

over the sandy bottoms in schools of ever increasing size, much as

do the other species, and each school in its turn migrates leisurely

south. During the winter this mullet is very rarely if ever found in the

Beaufort region but with the approach of summer an occasional

individual may be taken. It is, however, so uncommon in its second

season or older, that the fishermen consider it a matter of curiosity

or note when one is caught. Several specimens about eight inches

long were taken on the 27th of June. From the scale formula of an

individual 184mm. long (1.67+26) there seems to be little doubt

that this fish migrated in the early spring. Three other scales from

fish bearing no data show a similar linea, but situated farther from the

edge of the scale.

Adults

Due to the scarcity of this species at Beaufort no true adults were

procured so that practically nothing is known concerning their

habits. From figure 3 it is evident that the spawning period must be

rather protracted and, if an estimate of the time can be made from

the dates when the smallest fish are procurable the season would be

(conservatively) from mid-April to mid-August, the height of the

season probably being in May.

SUMMARY

Mugil cephalus Linné

1. To the synonymy of the genus Mygil should be added Queri-

mand.

2. To the synonymy of the species M. curema should be added

Q. harengus, its juvenile young.

3. M. cephalus spawns in October and November (September to

December).

4. The juvenile young pass the winter without much growth.

CHARACTERS OF THE MULLETS 227

5. In the spring the juvenile change diet and grow very rapidly

until fail when they school and migrate south not to return until

spring.

6. In the spring, the young, at that time from five to eight

inches long, return north by a (typically) continuous run.

7. By the second fall the fish have reached a length of a foot or

more and attained maturity.

8. In October and November these two-year-old fish migrate

south spawning as they go.

9. Jumping mullet may attain an age of five or six years, spawning

each year after maturity.

Mugil curema Cuv. & Val.

1. M. curema spawns in May and June (April to August).

2. The young are abundant in the bays and estuaries of our Atlan-

tic coast and develop rapidly.

3. In the fall the young school and migrate south.

4. After their first year, white mullet are but seldom caught north

of Florida.

BIBLIOGRAPHY

BEAN, T. H.

1913. Catalogue of the Fishes of N. Y., N. Y. State Museum Bulletin No. 60,

p. 366.

CocKERELL, T. D. A.

1913. Observations on Fish Scales. Bulletin U. S. Bureau of Fisheries, vol. 32,

1912, pp. 117-174.

Daur, Knut

1911. Age and Growth of Salmon and Trout in Norway as Shown by Their

Scales. Salmon Trout and Association. London. pp. 1-141.

Eartt, R. E.

1887. The Mullet Fishery. The Fishery Industries of the U. S., U. S. Commis-

sion of Fish and Fisheries, Section V, vol. I, pp. 555-582.

GILBERT, C. H.

1913. Age at Maturity of the Pacific Coast Salmon of the Genus Oncorhynchus.

Bulletin U. S. Bureau of Fisheries, vol. 32, pp. 1-22, pls. I-XVII.

Goopve G. B.

1887. Food Fishes of the U.S. The Fishery Industries of the U. S., U. S. Com-

mission of Fish and Fisheries, Section I, vol. I, pp. 449-456.

Hyjort, JoHAan

1914. Fluctuations in the Great Fisheries of N. Europe. Conseil Permanent

International pour |’Exploration de la Mer. Rapports et Proces-verbaux,

vol. XX, p. 122, pl. 3.

228 ARTHUR PAUL JACOT

Jorpan, D. S. & EvermMann, B. W.

1896. The Fishes of North & Middle America. Bulletin U.S. National Museum,

No. 47, pt. I, p. 809.

& GrBert, C. H. .

1883. Notes on a Collection of Fishes from Charleston, S. C. Proceedings

U.S. National Museum, vol. V, pp. 580-590 (588).

1884. Description of Ten New Species of Fishes from Key West, Florida. Pro-

ceedings U. S. National Museum, vol. VII, pp. 24-32 (26).

& Swain, J.

1884. A Review of the American Species of Marine Mugilidae. Proceedings

U. S. National Museum, vol. VII, p. 274.

KENDALL, W. C.

1892. See under Smith, H. M. 1892, but p. 192, footnote.

SmitH, H. M. & KENDALL, W. C.

1892. Extension of the Recorded Range of Certain Marine and Fresh-water

Fishes of the Atlantic Coast of the U. S. Bulletin U. S. Fish Commission,

vol. XIV, pp. 15-21 (21).

LEA, EINAR

1913. Further Studies Concerning the Method of Calculating the Growth of

Herrings. Conseil Permanent International pour I’ Exploration de la Mer.

Publications de Circonstance No. 66.

Linton, E.

1904. Parasites of Fishes of Beaufort. Bulletin U. S. Bureau of Fisheries, vol.

XXIV, pp. 321-428 (361).

MASTERMANN, A. T.

1913. Report on Investigations upon the Salmon with Special Reference to

Age Determination by Study of Scales. Board of Agriculture and Fisher-

ies of England, Fisheries Investigations, Series I, vol. I, p. 12.

Nitson, DAvip

1914. A Contribution to the Biology of the Mackerel. Conseil Permanent

International pour l’Exploration de la Mer. Publication de Circonstance,

No. 69.

RAVENEL, W. DEC.

1887. Information Bearing upon the Artificial Propagation of the Mullet.

Bulletin U. S. Fish Commission, vol. VII, pp. 197-202.

RUTHVEN, A. G.

1908. Variations and Genetic Relationships of the Garter-Snakes. U.S. Nation-

al Museum, Bulletin 61.

SmitH, H. M.

1892. Report on a Collection of Fishes from the Albemarle Region of N. C.

Bulletin U.S. Fish Commission, vol. XI, pp. 185-200 (192).

1897. Fishes Found in the Vicinity of Woods Hole. Bulletin U. S. Fish Com-

mission, vol. XVII, pp. 85-111 (94).

1907. Fishes of North Carolina. N.C. Geological and Economic Survey, p. 181.

CHARACTERS OF THE MULLETS 229

Starks, E. C.

1900. Osteological Characters of the Fishes of the Suborder Percesoces. Pro-

ceedings U. S. National Museum, vol. 22, pp. 1-10 (7), pl. I-III.

SUMNER, F. B., Osspurn, R. C. & Cote, L. J.

1911. A Biological Survey of the Waters of Woods Hole and Vicinity. A Cata-

logue of the Marine Fauna. Bulletin U.S. Bureau of Fisheries, vol. XX XI,

pt. II, p. 747.

Wiper, B. G. & Gace, S. H.

1882. Anatomical Technology. Barnes & Co., p. 27, & 47.

EXPLANATION OF PLATES

PraTte XX

Fig. 8. Juvenile scale from smallest fish normally obtainable (23 mm. fish), x 45.

Fig. 9. Juvenile scale with maximum amount of development (32 mm. fish),

x 45.

Figs. 10-14. Juvenile scale being enclosed by the more advanced type of scale

(29 mm.—45 mm. fish), x 45.

PLATE XXI

Fig. 15. Scale of a 60-70 mm. mullett with cteni first forming, x 45.

Figs. 16-17. Juvenile scale being enclosed by the more advanced type of scale,

x 45.

Figs. 18-19. Development of cteni on scale of the white mullett, x 45.

PLrate XXII

Fig. 20. Typical scale of advanced first season jumping mullett, x 25.

Fig. 21. Typical scale of advanced first season 121 mm. white mullet taken

August 23, x 21.

PEATE exer

Fig. 22. Scale of a 117 mm. mullet with unusual amount of circulation, x 30.

Fig. 23.. Scale of 120 mm. jumping mullet taken April 28 with linea very near

margin, x 25.

Fig. 24. Scale of jumping mullet, with a double linea, x 30.

PLATE XXIV

Fig. 25. Typical scale of second season jumping mullet. x 30.

PLATE XXV

Fig. 26. Scale of a five year jumping mullet 502 mm. (20 inches) long. x 13.

Development of scale of M. curema.

PLATE XXVI

Figs. 27-28. Ctenoid area of scales of adult M. cephalus and M. curema, highly

magnified. :

23D

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TRANSACTIONS

American Microscopical Society

(Published in Quarterly Instalments)

Vol. XXXTX OCTOBER, 1920 No. 4

MICRO-TECHNIQUE

SUGGESTIONS FOR METHODS AND APPARATUS

N. A. Coss

United States Department of Agriculture

SYSTEMATICALLY EXAMINING LARGE SERIES OF MICROSCOPICAL OBJECTS

There are various methods of recording the position and char-

acter of each member of a large series of objects mounted on a micro-

scope slide. One of the commonest methods involves the use of a

recording, mechanical stage. Each object on the slide receives a record-

number consisting of two separate readings from scales engraved

on the mechanical stage. The following method, however, is suc-

cessful without a mechanical stage or finder of any sort, and is charac-

terized by simplicity and expedition. It may be called the method of

charting.

The method consists in making a camera lucida drawing or chart,

at low magnification, of all the objects of which it is desired to make

record. Thechartisdiagrammatic; each objectis represented on the

chart by a simple, characteristic diagram, and the diagrams are then

numbered in series. The sheet that carries the chart may also carry

a series of printed numbers with corresponding spaces for records.

(See Figure 1.) Where the objects belong to a few great groups,

such as land-inhabiting, fresh-water, and marine, the printing of the

blank sheets in correspondingly assorted colors is an advantage.

The chart is made by using a camera lucida and an objective of

about five-inch focus.! In order to reduce the magnification, the

objective may be screwed into the end of the draw-tube of the micro-

scope barrel. A low power eye-piece is used with the objective, so

1 A very strongly magnifying spectacle lens will serve the purpose.

232 N. A. COBB

that all the objects on the slide can be seen at one time. A chart

having a magnification of five diameters is of convenient size. The

suitable illumination is secured by using a concave mirror without

sub-stage condenser. The light may be direct, in which case the

objects are seen as dark bodies on alight background, ora dark-ground

effect can be produced by inserting between the concave mirror and

the objects a small opaque disc. A suitable disc may be made by strip-

ping the barbules from a dark-colored six-inch wing or tail feather so

ylenchua gpiral. .. 26See-l11--....--

SrCepnalobue ties «eee

Soil - Imported roots of plants,"

Brazil - Diff. #510

No. 7 SDoryl, styracturus. 28 ».¥.... - cave

seabe 4Achromadora brazia ) 29) iN) ste ee

SDoryl. caudatus?_ . 30 Mononchus minor : -

6Elassonema - two. _ 31 " _ fragment minor -

p. Sie @Tylenchus_perfactua S32Rhabditie _. _ -_ -

ae rt 8Doryl. additicius _ 33Y.Doryl.protrudens-

Se MEE es 9 ."_ . protrudens. _ 34See.11 9. 2 J a8

---+--~-- teat aceite 10024. 20D SD BOBBYS Doryl ss eae

a G fo. ART ea lace Da te ee eS SG kchropad ores eee

a 16 +1 2Tropiconema tenuicolle 37See.11..._----

Teo De ¥ See.1] Egg... . S38Rhabditie - - . - -

POE ET Mer GaN ORNS Se fs BPN Hm) Achromadora~pepillae?39 Elassonema.. _ - -

ger (Se \ee RIDY eae pte pa oe OW aplos = 2 ss

eittacy? zt 16Mononchus __ _s: i. 41 See.1] .2 ts eee

sn? ee ey, 17Achromadora .... 42Pibre- - ------

< a nea en ae Bois ET! 18Rhabditis __-.- 43'Seesll) 2). aire eee

Ne j; + igirome __ ... _ 44Doryl. poor ._ Lu

2OElassonema._ _. _ _ 45 Rheabditia -.---.4

EP oat ae Ss 2 SPU ees ie lo =

Q2Rhabditis:..... 47 Doryl- sl. t1:-- =

Pylenchugy a 2 2) ES =)

94Mononchus minor _ _ a8 ner, Sl. Begs -.

Qoikhabditis ci Ll 602 2 2 eae

Fig. 1. Record chart used in tabulating large numbers of microscopic objects ar-

ranged on a series of slides. As printed the chart was 5x8 inches, and carried only the

two columns of figures 1 to 50 inclusive. At the left is seen the camera lucida drawing, or

chart, recording the form, size, and relative position of forty-nine microscopic objects,

—in this particular case, nemas. Immediately above the chart are seen the data relating

to the particular slide charted, which was No. 7 in a series of eleven slides (1-11), and

which carried a collection of forty-nine nemas gathered from soil attached to the roots

of plants imported from Brazil. Names and other notes with regard to the nemas were

typewritten opposite the appropriate numbers. Nos. 2, 7, 12, 13, 14, 23, 48, and 49

were encircled to indicate that these specimens were of especial interest. One-half

size.

as to leave only a small fan-shaped tip at the end, from one-half to

three-fourths of an inch across. With scissors, this is trimmed so as

to have a somewhat rounded contour. While the right hand is en-

gaged in making the chart, the left hand can flirt this little disc in and

out between the objects and the concave mirror and so produce a

MICRO-TECHNIQUE 235

is sawed from a sheet of German silver about one two-hundredth of an

inch thick. The edges of the central aperture are beveled so that the

mixture frozen on it becomes dove-tailed to the plate. In a similar

way, the small, washer-shaped piece of German silver fastened to the

top of the dome, as shown in Fig. 3,r, is

also beveled.

The German silver wheel is sol-

dered throughout to a round sheet

of exceedingly thin brass or German

silver. Then into six marginal per-

forations in the German silver wheel,

brass pins are soldered, giving to the

whole affair the appearance of a six-

legged table. The heads of the pins

are filed off so as to give clearance for

the microtome knife. The pins serve

to fasten the plate to a perforated

cork, being thrust into the cork eo Fig. 3. Perspective view and longi-

shown in the illustration. The rim tudinal section of a freezing-micro-

of the dome of thin sheet metal is tome object-holder mounted on a

somewhat similarly stiffened by sol- cork cylinder. The holder is made of

dering to it a ring of German silver ™¢tal only about 2/1000 of an inch

which is perforated and supplied with BEE Phe \edaes Orne ane)

i Repaie: : are beveled so that the imbedding

six brass pins in the manner just de- mixture when frozen is dovetailed to

scribed. the holder.

Though the dome-form is some-

what more difficult to construct than the flat, it is more efficient for

three reasons: It is more rigid, it gives a better clearance for the

microtome knife, and it contains less material.

In the case of small and moderate sized objects of which only a

few sections are required, the method is extraordinarily expeditious.

Objects of such a size that they can be imbedded in a few drops of the

freezing mixture placed on the control part of either of these metal

supports can be frozen in a few seconds by applying an ordinary ether

spray to the under side of one of these thin metal supports. The ex-

ceeding rapidity of the congelation gives rise to a consistency favor-

able to section cutting.

236 N. A. COBB

Ill

TO OBTAIN AN END VIEW OF A NEMA, ROTIFER, OR OTHER

SIMILAR SMALL OBJECT

Suppose the object is a nema of which an end view of the head is

required: decapitate the nema behind the pharynx with the aid of an

eye knife, or similar very small tool, having a very slender, thin blade.

The smallest and most slender-bladed knife used by oculists in oper-

ations on the eye is a very suitable tool, and it must have the degree

of sharpness characteristic of surgical instruments in good order.

Bring the nema by appropriate methods into glycerine; the decapita-

tion should be done in a drop of glycerine placed on the surface of a

transparent piece of celluloid. Push the nema to the bottom of the

glycerine and against the celluloid; decapitate by pressing the edge

of the knife against the nema as the latter rests on the celluloid. The

celluloid is sufficiently soft so that the edge of the knife will not be

dulled. If the knife is sharp, the cut will be clean, and the object

satisfactory. If the knife is dull, the nema will be more or less crushed

at the point of section and the preparation may prove unsatisfactory.

Mount the head in melted glycerine jelly, using sufficient jelly so

that the object may stand on end after being covered in. Place the

mount on the stage of a microscope, bring the object into focus, and

with a dissecting needle gently shove the cover-glass slightly back and

forth until the object is seen to be onend. Allow to remain on the

stage of the microscope until the jelly sets, watching from time to

time to see that the object maintains the desired position.

According to my experience, this is a better method of obtaining

end-on and sectional views of the heads of free-living nemas and other

similar small organisms than that of sectioning and imbedding. The

trouble with the method of sections is that the microtome knife very

seldom cuts the object to advantage. It is quite likely to cut in the

wrong place. If the ends of the setae or the surfaces of the lips are

removed in the first cut, it is a very troublesome matter to obtain a

good view or good sketch of the structures. Even if some of the parts

should not be lost or offer difficulty in mounting, there are so many

chances that the microtome blade will cut through at a disadvanta-

geous place that, as a rule, a very considerable number of nemas will

have to be sectioned before a good preparation is secured.

The method of sections has the further disadvantage that the

following of such small objects through the various dehydrating and

MICRO-TECHNIQUE 237

staining fluids, and the final orientation of them, is a tedious and diffi-

cult matter. Moreover in the case of nemas, there is considerable

difficulty in properly imbedding the object. The cuticle of nemas is so

impenetrable that unless special precautions are taken, the paraffine

will not thoroughly penetrate the tissues, and the results will be un-

satisfactory.

End views may be obtained by mounting the nemas in a micro-

scopic well made from a thin section of thermometer tubing. The

tubing should be like that used in the most delicate medical thermom-

eters, that is to say, with the smallest aperture procurable. This

tubing may be bought under the name thermometer, or barometer

tubing. It is well to have on hand ground sections of varying thick-

ness, from one-quarter of a millimeter thick to one millimeter or more.

The discs are cemented to a glass microscope slide at the time of using

by means of smoking hot wax or other suitable cement. Before

cementing the disc to the slide, fill the capillary aperture in the disc

with mounting fluid. This may be easily done by placing on the slide

a very tiny drop of the mounting fluid, and laying the disc onto the

small drop. The mounting fluid will enter the aperture by capillarity.

If it be desired to look at the head end of a nema, it is placed in the

microscopic well, tail down. If the nema is too long for the well,

it may be cut to fitit. The point is, to see that the object has about

the same length as the depth of the well, so that the end portion of

the object it is desired to view will come close to the under side

of the cover-glass when this latter is placed on the top of the well,

or rather on the disc of the glass containing the well. In placing

the nema in the well, a suitable tool is a small, curved hair cemented

to the end of a dissecting needle. Human eye-brow hairs are suitable

for this purpose. Using this method, the specimen can be examined

in clove oil, cedar oil, or any mixture of these or any other similar

thin mounting fluid. Cedar oil, having the same refractive index

as the glass composing the well, has advantages in connection with

illumination. The illumination in aqueous media is less satisfactory.

When the glass discs are not in use, it is best to keep them in

absolute alcohol in a glass-stoppered bottle. They should not be al-

lowed to become dry with mounting fluid in the capillary orifice,

otherwise they will be very troublesome to clean out.

238 N. A. COBB

DESTAINING OF NEMAS OR OTHER SMALL

OBJECTS IN THE DIFFERENTIATOR

In handling a mass of small organisms by the differentiator

method, there is sometimes considerable difficulty in securing satisfac-

tory destaining. There is little difficulty in getting a mass of organisms

thoroughly impregnated with the stain, no matter how varied they

may be in species and jn size; it is simply a matter of time. The

trouble comes in destaining. If the destaining process is carried on

until the largest of the objects, or the most impenetrable ones, are suf-

ficiently destained, it will generally happen that smaller specimens, or

those more easily penetrated, are deprived of too much of their color.

It is therefore a matter requiring considerable experience and judg-

ment to successfully destain such a miscellaneous collection. The

difficulty is considerably increased by the fact that when enclosed

in the differentiator tube, the specimens are not very easy to examine

critically by any ordinary method. If the differentiator be held

toward a strong light, the organisms may be examined by the aid

of an ordinary pocket lens, but not very critically. The most

satisfactory piece of apparatus for this work is what is sometimes

known as the chemical microscope, in which the objective is below

the stage and the light that passes through it from above is reflected

by a prism placed below so as to pass obliquely upward through

a barrel carrying an eye-piece. If the differentiator tube contain-

ing the destained nemas is laid on a glass stage over the objective

of such an inverted microscope, and a little water, or still better,

cedar oil, be placed between the differentiator tube and the glass stage,

it will be found that the nemas or other objects will sink to the bottom

of the fluid in the differentiator tube so as to come as near as possible

to the objective of the microscope. If the glass stage is thin, there is

no difficulty in using a one-half to two-thirds inch objective. In this

way, the nemas may be examined more critically with regard to the

extent of the destaining.

If it is desired to use a lens of higher power, it is sometimes possible

to do so by resorting to another method. Place a cover-glass on a

horizontal surface, and on the cover-glass a good-sized drop of cedar

oil. Lay the differentiator tube into this drop of cedar oil in such a

way that the nemas come opposite the cover-glass. It will now be

MICRO-TECHNIQUE 233

dark-ground effect as desired. To do this the feather ‘disc’? must be

materially smaller than the mirror.

The charts are nothing more than rude camera lucida drawings of

the objects, and with practice can be made with great rapidity. A

lot of fifty nemas mounted under a three-quarter inch round cover-

glass can be drawn in two to three minutes with sufficient accuracy to

make a very useful chart. (See Figure 1.) Each nema-diagram on the

chart has four very distinct properties, (1) Position, (2) Form, (3) Size,

(4) Orientation. For the most satisfactory work, it is desirable that a

certain optimum number of objects exist on the slide. This optimum

is determined by the number of them that will appear in a single field

of the lens afterward used in searching. Suppose a sixteen millimeter

objective is used as a searching objective, and a four millimeter for

the examination; then the optimum number of objects under the

cover-glass is that number which brings into each field of the sixteen

millimeter objective one to three objects.

After the chart is made, the short, crooked lines, representing the

nemas, say, are numbered in transversely arranged groups. Each

transverse group of the series constitutes a band of nemas running

across the mount and having such width as comes fairly well within

the scope of a single field of the 16 mm. objective. These imaginary

bands are illustrated in Figure 1. It will be seen that there are four

such bands. The nemas are numbered more or less consecutively.

Proceeding in this manner, on reaching the end of the first band, one

numbers the second band, also more or less consecutively, and so

on to the end.

In recording, begin with No. 1, placing it in the field of the 16mm.

objective. It is recognized by its size, form and orientation. Having

recorded No. 1 and examined it with the 4 mm. objective, a glance at

the chart will indicate at what distance, and in what direction, No.

2 lies from No. 1. Revolving to the 16 mm. objective and looking

through the microscope at Nema No. 1, the slide is moved in the in-

dicated direction until No. 2 is found and recognized. After record-

ing No. 2, No. 3 is found in the same way, and so throughout. The

novice will be surprised to find how easy it is, with a little practice, to

follow the series through without error.

The drawings should be so made and numbered that the chart and

the objects as seen under the microscope will resemble each other.

234 N. A. COBB

If no care be taken in this respect, the chart may be found to be “‘left-

handed.” Securing a “right-handed” chart is merely a matter of prop-

erly arranging the paper at the time the chart is drawn. Diagrams

should be so made with reference to the printed matter that when

it is right side up, the objects as viewed through the microscope

will have the same orientations as the diagrams.

This completes the description of this method, except to explain

that in the example illustrated, the numbers encircled are so marked

in order to indicate that those particular specimens present note-

worthy features.

The method may be elaborated in a variety of ways for the re-

cording of nemas, rotifers, protozoa, desmids and a vast array of other

microscopic objects. If the charts are of card-system size, say 5x8”,

they lend themselves to all sorts of convenient methods of filing. By

using thin paper, carbon copies can be made at the original draft.

The charts can be made and used by a grade of assistant that might

hardly be intrusted with the use of a recording mechanical stage, and

who may lack training in the accurate reading of scales and the record-

ing of numbers. Floating of the objects, of course, disarranges them.

Newly made slides are sometimes subject to this disadvantage. The

difficulty is avoided by keeping the slides always in a horizontal posi-

tion.

OBJECT SUPPORT FOR A FREEZING MICROTOME

In this freezing microtome attachment, the object is to reduce

the metal parts to a minimum and to concentrate the effects of the

freezing mixture as much as possible upon the object to be frozen.

To this end the object is placed on a thin

metal plate, only about one to three thou-

sandths of an inch thick, to which the nec-

essary rigidity is imparted either by solder-

ing it to a radiating framework in the form

of a flat wheel sawed from somewhat thicker

Fig. 2 metal, or, preferably, by giving to the metal

the form of a dome. These metals supports

are illustrated in Fig. 2 and Fig. 3 in which they are shown full size.

A six-spoked wheel, having a hub-hole one-eighth of an inch across,

MICRO-TECHNIQUE 239

found that the cover-glass will adhere to the differentiator by capil-

larity, so long as the differentiator is held in a horizontal position. If

the chemical microscope stage has a large aperture, it will be possible

to lay the differentiator across the stage, cover-glass downward. In

this way, if the differentiator tubing is thin, it will be possible to use

even quarter-inch objectives of long focus.

Where considerable work is done with differentiators, a chemical

microscope used in this way is a valuable accessory.

COMPRESSORIUM FOR CHROMOSOMES

When chromosomes or other similar minute bodies are so massed

together that one lies behind another and is thus liable to be missed in

counting, the compressorium described below may prove useful in

overcoming the difficulty, which none of the ordinary compressoria

will do.

When such a mass of chromosomes is flattened out by pressure,

the individual chromosomes behave somewhat as

would the seeds of a pulpy fruit under similar cir-

cumstances. They appear to be of a different con-

sistency from the material in which they lie,

and behave under pressure as if harder and

more compact than the surrounding matter.

Under moderate pressure they do not show much

tendency to break in pieces, but rather toaccommo-

date themselves to the narrower quarters by rear-

ranging themselves more nearly in one plane. So

far as enumeration of the chromosomes is concerned,

this new arrangement has two advantages: Ist,

they may all be more readily brought into a single

view, that is, all brought into focus at one time;

2nd, in the flattening-out process, they slip one over

another somewhat, and recede from each other—for

instance, as the seeds inside a grape will do, when

similarly pressed.

Fig. 4. Two curved,

perforated, steel

springs made from

thin, safety-razor

blades, as described

in the text. These

two forms, while of

the same length,

nearly one inch, are

of different degrees

of springiness; that

at the left being

the weaker.

The compressorium I have devised to secure this effect is con-

structed as follows: Take a safety-razor blade—one of the thinnest

kind, having perforations an eighth of an inch in diameter—and

240 N. A. COBB

soften it by heating it toa red heat. With shears, cut a somewhat

diamond-shaped piece from the softened blade, so that the ‘‘diamond”’

is about three to four times as long as wide, and has one of the round

apertures in its center; bend this elongated ‘“‘diamond” into asymmet-

rical bow whose depth is one-eighth of an inch or more. See Fig.

4. Heat the bow ina flame to a cherry-red and plunge it into cold oil

or water to hardenit. This will result in a springy piece of metal that

can be utilized to exert pressure on a small cover-glass under which

are mounted cells containing the chromosomes it is desired to scatter.

The length of the piece of springy steel may conveniently be made

to be about one inch, so that it will just reach across an ordinary

three-by-one glass microscope slide. Bind the slide in a piece of thin

metal having a three-quarter inch perforation at the back—that

is to say, so bend a piece of thin sheet metal that an ordinary slide

will slip into it through grooves along the two sides of the folded piece

of metal. See Fig. 5. This metal should simply pass around the edges

of the slide and lap over about a sixteenth of an inch at each edge leav-

ing one face of the slide uncovered. The grooves should be a little

wider than the thickness of the slide—at least enough wider so as

easily to admit the thin perforated metal spring. Place the cells, the

chromosomes of which are to be studied, on the slide opposite the

middle of the three-quarter inch aperture. Use very little mounting

medium; cover the cellular tissue to be treated with a small round

cover-glass. Tuck the ends of the bowed piece of springy perforated

steel under the edges of the

metal slide-case or holder,

holding the spring against the

small cover-slip in such a way

; that the cells to be compressed

/ lie opposite the center of the

x small perforation. Press and

lock the spring in the same

Fig. 5. Portion of a 3x1 inch glass microscope way aS in the case of the

slide enwrapped with thin metal as described in : ‘i

the text. a, thin metal wrapper; b, one of the 2 ee at the back of ee ae

springs shown in Fig. 4, cee in position onthe dinary photographic printing

slide so as to press the small round cover-glass, ¢ :

against the slide, e; d, aperture in the back of the frame. The cells will now be

metal wrapper, a. The ends of the spring,6, under pressure at or near the

enter through the notches on the edges of the + Jats

wrapper, a, so that in being applied the spring center of the perforation in the

does not need to be rotated more than a few steel spring. The entire con-

degrees. trivance will differ but very

=’

MICRO-TECHNIQUE 241

little in form and size from an ordinary microscope slide and can be

placed on any microscope stage in the same way as a slide. The

piece of springy steel is so thin that it in no way prevents the use of

a high-power immersion objective. Needless to say, it is for this

reason that it is made from such thin metal. The spring may be

manipulated with the aid of matches or wooden toothpicks.

Ordinary slides and cover-glasses are almost never perfectly flat.

Better results will be obtained by this method if the slide has its con-

vex surface up and the cover has its convex surface down, so that the

cellular tissues to be treated lie between two very slightly convex sur-

faces. It will be found that in this way very compact groups of chro-

mosomes and other similar objects can sometimes be scattered so as

to be counted, when otherwise they could not be counted.

There seems to be comparatively little danger of exerting too

much pressure. The beginner’s tendency at first is to exert, if any-

thing, too little pressure. The greatest difficulty arises from sliding

the glasses on each other, since much of this ruins the preparation. To

overcome this difficulty, a series of three or four notches, close to-

gether, may be filed in the edges of the metal holder before it is

folded about the slide,—or rather about the metal core on which it

is bent, or formed, and which naturally has a little greater width

and thickness than the slide. If now the bowed spring has a length

a little less than the distance between the bottoms of the notches

in the edges of the slide-holder, it will be found when it is pressed

down that the pointed ends can be tucked through the notches and

under the edges of the holder without materially sliding or rotating

the spring. The accompanying illustrations will assist in under-

standing this simple and effective device.

The particular cells to be compressed are prepared and searched

out in the usual way, then dissected out together with as little of the

surrounding tissue as possible, an operation performed with the aid of

an ordinary dissecting microscope. It may be advisable to look at

the group of chromosomes from both sides. To do this, the metal

holder, instead of having a three-quarter inch perforation, should

have a much smaller perforation, say about one-eighth of an inch.

Instead of using a three-by-one glass slide, cement to the inside of the

metal holder a thin cover-glass several sizes larger than that to be

242 N. A. COBB

Fig. 6. A metal holder for clamping a microscopic object between two thin cover-

glasses. a, metal holder; b, steel spring as illustrated in Figs. 4 and 5; c, small, round

cover-glass; d, rectangular cover-glass underneath the round cover-glass; e, notches

in the metal holder for the reception of the spring. This holder enables the micros-

copist to look at the object with an immersion lens from either direction.

placed over the object. As the metal holder, in order to be stiff enough,

has to be several times thicker than the bowed spring, it may be ad-

visable to bevel the edge of the round aperture in the holder, so that it

will interfere as little as possible with the use of an immersion objec-

tive. Onaslide constructed in this manner, the object is held between

two cover-glasses, and hence may be viewed from either side with

equalease. Such a slide furthermore permits the use of an immersion

lens as a condenser, a proceeding that has advantages.

LIST OF MEMBERS

HONORARY MEMBERS

Crisp, FRANK, LL.B., B.A., F.R.M.S.,

5 Landsdowne Road, Notting Hill, London, England

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LIFE MEMBERS

Brown, J. STANFORD, Ph.B., A.M........ P.O. Box 38, Far View, Black Hall, Conn.

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MEMBERS ADMITTED SINCE THE LAST PUBLISHED LIST

ARNOLD, L. P. Jackson, F. S.

ASHLEY, F. M. Jupp, H. D.

Brunoauist, E. H. Kenyan, W. A.

CASHEN, DorotTHY Kostir, W. J.

Ciovis, MABEL C. Kupo, R.

DERE UN Es la. Mancue_fF, E. D.

ELmorg, C. J. McCottoca, J. W.

Exits, C. McCuttoca, IRENE

Ensure, J. M. MottkowskI, R. A.

GRAVELLE, P. O. PIckarmae lr

Gross, F. O. Ryr, L. E;

Hayes, W. P. Scott, HELEN M.

Hupson, D. V. Smuon, C. L.

Husert, H. E. Tuers, R. V.

Hickman, J. R. TAvyior, W. R.

Hisaw, F. L. Watton, A. S.

Hortes, C. F. WARREN, SHIELDS

Hopkinson, D. YosHiDA, SADAO

Jacort, A. P.

243

244 LIST OF MEMBERS

ACKERT, JAMES EDWARD, Ph.D., ’11...... Kas. State Agr. Col., Manhattan, Kans.

ADAMS: sPREDERICK: © SB QWs oe cine pe pe obec ire ee aee cle Pee eet

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ATTEN Walt RA Vie MD ALSO SIS as a De 525 So. Park Ave., Bloomington, Ind.

ATEN: (WV YNERED fis. Al Mi O42 2 iene eke oreciaele arn Scripps Inst., La Jolla, Cal.

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BARKER, FRANKLIN D., Ph.D. ’03............ University of Nebraska, Lincoln, Neb.

BARRE SEW fBtOCl, Wel nD hee we reek Sy Bhs ty a) See cere Clemson College, S. C.

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IBROOKOVER*) CHAS: Ay Bee MES OSeeisenine ciacies Univ. of Louisville, Louisville, Ky.

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Brunoguist, E. H., A.M., 719..... Dept. Zoology, Northwestern Univ., Evanston, Ill.

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Fetiows, Cuas. S., F.R.M.S., ’83........ 107 Cham. of Comm., Minneapolis, Minn.

FERNANDEZ, Fr. MANUEL, B.S., 716........ San Juan de Latran College, Manila, P. I.

PINDUAM. (MEREINIC. ACM clone ees ccce nr viene e Park College, Parkville, Mo.

Pirz-RANDOLPH, RAYMOND B., FOROMIS 2 719 os Seis io ole ee leit ee

Beis ee Led ey Wy ena chy bieheae ake or Lace eee OMe State Laboratory of Hygiene, Trenton, N. J.

PVOTEAS UVP Ol. ciaeie come elena Creighton Medical College, Omaha, Nebr.

Peaniss, oH. Wo, MDa PD.) O52). oe 2 vei es cee 56 Brazos St., Hartford, Conn.

Ganeieer) spony Ge etn tea eae ne 2659 California St., San Francisco, Cal.

GAGEYPROF) SEION EL.) BiSij 202i er Selec mie. ee artes ait 4 South Ave., Ithaca, N. Y.

Gattoway, Pror. T. W., A. M., Ph.D., ’01....105 West 40th St., New York, N. Y.

Giprerui Me Ibn Di lose pictsrrte Biology Building, U. of Wis., Madison, Wis.

Garpsueren: iG Wi.,) Be eAey) Say cto sam mre hoe wise Bivsrte areteveteta let ete Lafayette, La.

Gowan, smAncrs EES 204 Fen) er ies eretepee sclera R. D. 1, Box 14, Exeter, N. H.

Grave; JouneH.; 19505 .0)). =. en: Research Dept., Brown Company, Berlin, N. H.

GeanAM,, CHARLES (Wey MERZ 11s one EN cat eeietn 447 W. 14th St., New York City

Gpanaw, JOHN YOUNG, PAs, 7142. diye stile Peietole dee aie = University, Alabama

CGWAVELED? BONO hele ctetie see clic tip sie 5 114 Prospect St., South Orange, N. J.

Grivein, LAWRENCE E., 7132.00 5002s ect eee eee ene Reed College, Portland, Ore.

GROSS WHO MID AO ae tinct aes coe inher ease 1816 Erie Ave., Philadelphia, Pa.

Guperter; Joan E,, Ph Dk, Mv... tees A. & M. College, Stillwater, Okla.

Guyer, MicHAkEt F., Ph.D., ’11............ University of Wisconsin, Madison, Wis.

HAGELSTEIN, ROBERT, ’16........... 165 Cleveland Ave., Mineola, Nassau Co., N. Y.

Hacur, Frorence, A.M. 7216... 0). icine see Nat. Hist. Bldg., Urbana, IIl.

Arr, Be Geecory, BAS do). oo. 2 33 Biology Building, Univ. of Wis., Madison, Wis.

Hance, Roperet T., B:A., 713.2... - 2.50. 22-s-- Zool. Lab., U. of Pa., Philadelphia, Pa.

HANKINSON, 1. L., 03. ..0..---.-..5%- New York College of Forestry, Syracuse, N. Y.

EDANSEN, 6 VAMESH US Cy.) nis s cielo ee eae 6 ie St. Johns Univ., Collegeville, Minn.

EEARD Ye UGENE EM ey eie oye cies 2 footed fever 1230 So. Keystone Ave., Indianapolis, Ind.

Harman, Mary T., 713........-.- Kansas State Agr. College, Manhattan, Kansas

eas, W. Ba MS, te eis eee eels sais meres 319 N. 18th St., Manhattan, Kans.

Heat, F. D., Ph:D.,7060.). 6)... 2.) ee Wash. State College, Pullman, Wash.

Heatu, Roy FRANKLIN, M.Sc., 718............---- P.O. Box 270, Billings, Montana

Hemeurcer, Harry V., A.B., ’714...........----.---: Fennville, Mich., R.F.D. 2.

HENDERSON, WILLIAM, ’11........... Mellon Inst., Univ. of Pittsburg, Pittsburg, Pa.

TATCRMPAN, | Jere, ACB rae ooo. o) Sin fore eleteialeyabelalninsoleiepeteiote, 3 Bristol, -West Virginia

Hatton, WirettaM A., PhD. 715..... 2.0.02 tee ee eee eee eee Claremont, Cal.

ERS Awe BE SBzS MUS 7 Staets 5-26 ce Kans. State Agr. College, Manhattan, Kans.

Hyorru, uopwue C5." 82722. ;- .-)- Meadowdale, Snohomish County, Washington

Hoty Cross CoLtteGE, PROFESSOR OF BIOLOGY...............--- Worcester, Mass.

Hopkins, FRANK’ B., B.S., *19.......-.--.----+- bys He aXe Oe North Salem, Ind.

FIOPKINSON, D.5 MEDS, ZO ree a itsinie) sees eR oie oa lee 1008 Third St., Milwaukee, Wis.

POSING) WAL TOMI E LAE emir 2 Sctine a vaciaeaeest eas 49 6th St., LaGrange, Ill.

AMERICAN MICROSCOPICAL SOCIETY 247

Horres, Cs E PRID S200 eis se hele Nat. Hist. Bldg., Univ. of Ill., Urbana, Tl.

HOWLAND, Henry R:, A.M), 7982. .0.255 055256055 217 Summer St., Buffalo, N. Y.

Eorer? Ee Ee aBESs Pa0ue was anda ade kao 3615 Melpomene St., New Orleans, La.

Hupson, 'D. V., BiS., 7208 6.05 052.0. Johns Hopkins Medical School, Baltimore, Md.

FLUGHES:) SABE WR U5 iy fe ce casiicaiers evalateitelebetadar sisi eyatets Grinnell College, Grinnell, Ia.

Wes; PREDERIGET O25 de eR ennai aletatelslcts 1201 Race St., Philadelphia, Pa.

IFACESON; ESS MED PAO tsa irs aa McGill University, Montreal, Canada

FACor; AY PEVAB ION Foe ee cs Sas No. China Language School, Peking, China

PEBES, | PROM ROCHE ers Le ENE oe ened catch e elena} Univ. of Okla., Norman, Okla.

INNER eA MA RDI ees A AN Oa te oy at Science Hall, Indianola, Ia.

SROEENS ON UB apeo ic BO Sevieea ters cpt eat Niahaes Bo ahaha chckis atl ahah Joplin, Mo., R.F.D. 4-147

JOHNSON, FRANKS; M.D), 793). 00. osc ceestelee ede Hotel Darby, Los Angeles, Cal.

JORDAN IE ROR ERED UTD sc iaee Nolet as) oohtckch abl. University Place, Charlottesville, Va.

jonas, Caancey, “OO. Hoy nbn ok Ss Biology Bldg., U. of Wis., Madison, Wis.

PDDH Ee DA Ope S219 oOo Nas Ae 460 W. Philadelphia Ave., Detroit, Mich.

ISNA SMORBRIGHPS CACME ey lie Tee ee Sd sials oynlh aubvdinennstets Bismarck, No. Dak.

ESTING WADS WWE An ACRE Se ZQ IO FOU. 8) Ik Ws ee a AL 2 a dims yee dd Milton, Wis.

KINCAID; PREVOR, A;M., 712d .0c5050 064% University of Washington, Seattle, Wash.

TICE inte 28 WE 2 A OS Pe Be Phillips Univ., Enid, Okla.

Kirscu, Pror. ALEXANDER M., M.G., ’16............. Notre Dame (Univ.), Ind.

RemepiInsD, HerMan, 1900.02 Sl ke Peek k ks 836 So. George St., York, Pa.

VON KERINSMID. PRES. Re Biy.)icJ/602 ics ven Univ. of Arizona, Tucson, Arizona

MOMRGTUE OE Ey BA ET TS PS. CSI eG ah eR ag 1015 Blondeau St., Keokuk, Ia.

Koro, Cuartes A., Ph.D., ’99, University of California, 2616 Etna St., Berkeley, Cal.

Kosriny Wi J., MA 20: 0.2 2065 3. Dept. Zoology, Ohio State Univ., Columbus, Ohio

nae NA POR) UN he a eke teed ee 32 S. Fourth St., Easton, Pa.

KRECKER, FREDERIC H., Ph.D., ’15........ Ohio State University, Columbus, Ohio

Kapow beDy 22 (ee iui. Sh rueea rt ak Loh Dept. Zoology, Univ. of Ill., Urbana, Il.

LamBeERT, C. A., ’12...Bank of New South Wales, Warwick, Queensland, Australia

LaRvE, GreorcE R., Ph.D., 11.......... University of Michigan, Ann Arbor, Mich.

DATHAM, Miss VAL McD D:DiS8:, FUR MLS, 885 02 )dod2, Uacdiasies ole Gnie oe dledaas

SPALSNe cba cratre SONA Wave ah HAR thewiad | wae aate ce bede pad t 1644 Morse Ave., Rogers Park, Chicago, Il.

Latimer, Homer B., M.A.,711....0.00 0.000000. Univ. of Minn., Minneapolis, Minn.

ERWIS)\ EVRY, FOREMAN SPB 2182), 52). 34 oco.48 PSS aaa dae oe os University, Va.

Lewis, Mes. KatHerine B., ’89............2.0505 Bellwood Farms, Geneva, N. Y.

LETteeep. Wie AM, BEB. 2068 i 52 2) 2 a ee gee a AE Nashville, Tenn.

HOM KAMOEBE 292 UW ae es dT aha ele 289 Westminster Road, Rochester, N. Y.

LONGFELLOW, RoBert Capes, M.S., M.D.,’11........ 1611 22nd St., Toledo, Ohio

POwoeEn, pen Bo 716.02. 2a cas 1312 York St., Denver, Colo.

MowREy,. "EEEANOR ©.) 719) 008 Be eee ee 1826 D. St., Lincoln, Nebr.

Lyon, Howarp N., M.D.,’84..............022. 828 N. Wheaton Ave., Wheaton, Ill.

MacGiiiivray, ALEXANDER D., ’12......... 603 W. Michigan Avenue, Urbana, IIl.

Mack, Marcaret EvizaBetuH, A.M., ’13............ Univ. of Nevada, Reno, Nev.

MAG ATE BeiMiS. PhP). 4S. )0 emanate Mayo Clinic, Rochester, Minn.

IMVANC@HIG. BD 97 Re be ae ee Latoh Rene 200 Glen Cairn Ave., Toronto, Can.

248 LIST OF MEMBERS

Marrs Grorcr/ Henry, MBS tei onee cee a: 94 Silver St., Waterville, Maine

Marsnatr Comins, MsD:; “960 2) oo sishcn 2507 Penn. Ave., Washington, D. C,

Wiens. Jom RID (Wsadscaodsuadonoqsnd: Rockford College, Rockford, Ill.

IMARSHATE: We SePhoD 12 iy hah: ee ae ee 139 E. Gilman St., Madison, Wis.

MArTLAND, Harrison S., A.B., M.D.,’14............ 1138 Broad St., Newark, N. J.

MATHER ES Me stPhiD OZR se rection etn 228 Gratiot Ave., Mt. Clemens, Mich.

May, JHenry jGustav, (PhD 215). 25 bleu eee eee tee eae

TR Re ee aaa ao aE Agr. Exp. Sta., Rhode Island State College, Kingston, R. I.

IMAVEE We HRONGE: Sib S ss loner cei slaceenoe oe Wesleyan College, Warrentown, Mo.

MAY WV AEDY PREDERTCK =, 702 serum nee ccieiieieeissieriee 222 Grand Ave., Nutley, N. J.

IMC COLEKOCH, IE Wr Dise lO ee eet ecprrcies Kans. Agr. Exp. Sta., Manhattan, Kans.

MCGREERY. (GEO DLWIS 2A rian estes sen ae mete 110 Nevada St., Carson City, Nev.

MCCULLOCH. [RENE (PhUD 520 ai se at ab anc wis Cale las hae eee OL eee

...Dept. Biology, Sophie Newcomb Memorial College, Tulane Univ., New Orleans, La.

McEway,A.,715..... Fifth Ave. Guarantee Building, 522 Fifth Ave., New York, N.Y.

MCKAY, i OSERH: G4) saree rior cia laeiciaasten etree meyoaelaieee 259 Eighth St., Troy. N. Y.

McK rrver! (Freep 1...) FORMS; 706.36. oss. as P.O. Box 210, Penticton, B. C.

IMCICAUGHETN SAT VAT. UIVISAL id Siete eee ete iar nerd ee eee Pullman, Wash.

IMG WALETAMS) SOHN Vol Ai 8 eave rosie ects ateraie) Blo ee chelate Lock Box 62, Greenwich, Conn.

Mercer, A. CLirrorD, M.D., F.R.M.S., 82... .324 Montgomery St., Syracuse, N. Y.

Mace CER. AW’: We SPMD, OO ocd oo Nee saison ana aden gnalatens 200 E. State St., Athens, Ohio

METCATE PROF ZENO ME iB Aey IZ Se ee. se Col. A. & M. A., W. Raleigh, N. C.

Miner | CHarirs Hs, oll. Med. School, Johns Hopkins U., Baltimore, Md.

Minter joan As PhDs BORIMGSS 789s. oe 44 Lewis Block, Buffalo, N. Y.

MOGKETD. i cbs sORS OLE es re tetaticredciasieudletetteya 2302 Sumner St., Lincoln, Nebr.

MGEECER ES ANAS 207 1 bis te bc Seep tery 341 West 57th St., New York, N. Y.

MontTGomeEry, CHARLES S., 719.............. 420 Riverside Drive, New York, N. Y.

Moopy, Rosert P., M.D., ’07......... Hearst Anat. Lab., U. of Cal., Berkeley, Cal.

Morcan, ANNA HAVEN, Ph.D., ’16............ Mt. Holyoke Coll., So. Hadley, Mass.

Mirren OAWSET, pRepAty 0) sit Opec weer eb eicte aaa: Univ. of Idaho, Moscow, Idaho

MVERS SBIRANK flere es csciteite 15 S. Cornwall Place, Ventnor City, N. J.

INESBIT, ROBE As UG. oie o's enh tcie Biology Building, Univ. of Wis., Madison, Wis.

NORRIS WEROREARRY NVALDO Ulan nee eee veer e 816 East St., Grinnell, Iowa

INORTONT CHARTERS HD VID) M11) ee eile eyate pepe i-cpeicierere 118 Lisbon St., Lewiston, Me.

OGEEVEEACG SPE SOC ple tae e's cists tae, 5 ou wise ee 1006 N. Union St., Lincoln, Il.

Osporn, Pror. HERBERT, M.S.,’05............ Ohio State University, Columbus, O.

Orr FTAB VEIN eNOS 25 area nb 20S oles acl eins Spencer Lens Co., Buffalo, N. Y.

PAGE, SERVINE PELEINE NG UIA) 0.5) one «0 o tape cite cine peiaet- 810 University, Ithaca, N. Y.

PATRICKAURANKAW Ee) E Ole. sls cy. Neieyelat 1500 Linwood Blvd., Kansas City, Mo.

IPRASE} BRED AN Sips ats <s sic's s clsjelale meat gains P.O. Box 503, Altoona, Pa.

PENNOCK> EDWARD, io ee ae, (0's <0 35-4) eocicnei 3609 Woodland Ave., Philadelphia, Pa.

PETERSON; NIELS PREDERICK, 711... 5 .yjcicicn neice ae pee ee Plainview, Nebr.

Pickett, F. L., Ph.D. ’20.Dept. Botany, State College of Washington, Pullman, Wash.

PrAte, FS.) -Phi Ds 10 pepe cS <5. ee puters ae 561 W. 141st St., New York, N. Y.

PATE EDWARD) alle: pemcriceiciiie es cece Brandhock, Gerrard’s Cross, Bucks, England

AMERICAN MICROSCOPICAL SOCIETY 249

ProucH, Haron H., A.M., ’16...... Dept. Biology, Amherst Coll., Amherst, Mass.

POHL, JOHN CoapRei lide tien cece sets es caisusieieistess s'seyei 204 N. 10th St., Easton, Pa.

POOL RAYMOND JRE LSD flO lnctara eas slevevernsatey as iclchpehale Station A, Lincoln, Nebr.

Pounpb, Roscor, A.M., Ph.D., ’98....... Harvard Law School, Cambridge, Mass.

ROWERS) BBE AGBe DED iae 20a ap stntchehtarciaherohshetevel ats Univ. of Nebraska, Lincoln, Nebr.

PRAEGER, Wie Ej Mi is 14s Sse crs cclets sloctotelenste 421 Douglas Ave., Kalamazoo, Mich.

PROcTER, WiILLTAm. PhiB., On eos. case ne 149 Broadway, New York, N. Y.

PURDY.. WILLIAM C2) MESG 21 One sere rete eee: 3rd & Kilgour Sts., Cincinnati, Ohio

Qumran) Marvin C5 AGM 2 1Si56 3.2 cieseciasiscws cock Wesleyan Col., Macon, Ga.

RANKIN, WALTER: Mi S/? 1S sic ceechs betters lelveisc Princeton University, Princeton, N. J.

Ransom, Brayton H., ’99..... U. S. Bureau of Animal Industry, Washington, D. C.

RECTOR. PRANK LESLIE, MDS eiline ac. se saad ae 227 Fulton St., New York City

REESE, ProF. ALBERT M., Ph.D. (Hop.) ’05...W. Va. Univ., Morgantown, W. Va.

Ricwarps, AvTE, Ph.D., ’12..Dept. Zoology, Univ. of Oklahoma, Norman, Oklahoma

Rimm ve. BaCuRTIS. MiSs. 15.5. .c usu eek Univ. of Manitoba, Winnipeg, Can.

ROBERTS SL WIGLISe 11S . lo. eee hine 65 Rose St., Battle Creek, Mich.

ROBERTS] EICMuean lan est), ose! Jace State Normal School, Cape Girardeau, Mo.

ROBERTS: Jamin oc. ge es a emilee eid 460 E. Ohio St., Chicago, Ml.

INOBINSON; JE ee IED. LS. cu Someta ae nee seco es Box 405, Temple, Texas

ROE} Ge! Cop Atseeli nies. cce 5 ces 'ofe cece 113 R Street, N. W., Washington, D. C.

IROGERS; WAL TOR MBL eo. hahahaha dover Lawrence College, Appleton, Wis.

ROSS; LUTHER SHERMAN) G:M., 711.5. .0 cee eee see 1308 27th St., Des Moines, Ia.

RUSH, Rf CAMP ae cys 5. ta Ste arora San id Budiara ah aa aapciee le wdioht Hudson, Ohio

eptanhid Dad a Jalal 710) 0 oto ARIS coo cee 62 Laureston St., Brockton, Mass.

SCHEAR: SEis SWisstesiyLO Mgrs hulocs so stds ete cine Btves'e rele 107 West Park, Westerville, Ohio

SCOTT ELEN MEA ee ee TN cave lala tors ele 26 Whites Place, Bloomington, Ill.

COTES Oper VV inl o eee NP tel sans lay elasecouals ibe Univ. of Wyo., Laramie, Wyo.

SHANTZ. HSE PReD ey OS as ee. oa Bureau Plant Industry, Washington, D. C.

SUE BOPNIISTDAT [ard Bg “hss s decid a o SBIR BEIGE ECE aE 809 Adams St., Bay City, Mich.

SHEERAR LEONARD) Pere Qe ene rvereteisrerestate lo Gore 158 W. State St., Wellsville, N. Y.

SHELDON, |) OHNE WISsm ne ened Seu ecun ey Cah (eae Sali te Morgantown, W. Va.

SHUETZ IY CHAS Or OLR okie a hecisied cee hes Box 135, Hoboken, N. J.

SIMON Cs) La Me) oa 2 Orne ra ae ates isin ia 8 1734 Linden Ave., Baltimore, Md.

SISTER Maewna, O.S.B., M.A., 716.......... St. Benedict’s College, St. Joseph, Minn.

SITEER IDAl Bs. loser sae tes saya ast or Lake Erie College, Painesville, Ohio

Smrrn, Pror) FRANK, AGM.) 7120 tos sa: 1005 W. California Ave., Urbana, III.

SmitH, GILBERT MorGAn, Ph.D., 715................ 1606 Hoyt St., Madison, Wis.

Swrre, Jo G.5° 96 ee eee ae eee 131 Carondelet St., New Orleans, La.

Soar, C. D., F.R.MLS., ’07...37 Dryburgh Road, Putney, London, S. W., England

SPAULDING, Mi, Hi VAUMueMiSmaeescsa ace 720 W. Babcock St., Bozeman, Mont.

SPURGEON, CHARLES Hy, AUME Stara nemeemecen eens occ Sheridan, Ind.

STEVENS, Pror. H. E., M.S., ’12...Agricultural Experiment Station, Gainesville, Fla.

SrewarT, THomAs S., MoD °ll. 22. eee 18th and Spruce Sts., Philadelphia, Pa.

STONE Gre Bara. de midsole TOL NE Taare nse 1725 LeRoy Ave., Berkeley, Cal.

STONE, GRACE As. ALM, Ona me cae cee sete Suh ey Teacher’s College, New York City

250 LIST OF MEMBERS

STUNKARD, Horace W., Ph.D.,’13...New York Univ., Univ. Heights, New York City

SUMMERS: PROE WE! Hie lOOs,. 14 hae se Serae see eee en nee Ames, lowa

SWEHzy, Onivry eh Dy edore cence. East Hall, University of California, Berkeley, Cal.

SWINGLE, PROF) LEROY D:0GE 2.2 -6- eS - Univ. of Utah, Salt Lake City, Utah

IDAVTORFWathe PasD) LU DOM rans Matra iy a wen. 1340 N. 12th St., Philadelphia, Pa.

‘TERRELL, (DRUMAN G..0MED Ss VAG) Wisely. ac oe 1301 Eighth St., Fort Worth, Tex.

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INSEE RANDOLPH: WORD) Biol Lonel ens eee eine eee Georgetown, Texas

opp; JaMEs'@., BeAvy MaDe Mh eee seo. sea daes een ea ae Boulder, Colo.

Tucker, ON Es PhiGe "Ages. hinec as aeenneeeee 898 S. Clarkson St., Denver, Colo.

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WiATTE FREDERICK (Cs (PRD. 7didee tenner et Behe a easaciaes oc etee eee

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WALKER LEV AD BERLE pal Gi thrccn Bisee ols wevstyeris See ee Station A, Lincoln, Nebr.

WAT TONS ANSP IO Gem aiccci A corse tenet ee 2819 W. Girard Ave., Philadelphia, Pa.

WARBRICK wy an Goede as ie. eieupe ne tarts Rie eyo nye ee 306 E. 43rd St., Chicago, Ill.

Warp, Beney B:, AM., Ph:Di787 2... oe. os University of Illinois, Urbana, Ill.

WARNER. B sAG MEDD tPiniGe. afin Oey Lik Sey cha iis eee gee iets ane Nevada, Iowa

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WARREN SSS AEB 210 © caus dolce eerelos ele eae 28 Hawthorne Rd., Brookline, Mass.

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WIESE SAT OxoOU 23) SUE cher eet cls eisteers eee are Univ. of N. M., Albuquerque, N. M.

WELCH IPAUE Ss) wd anit. cee eects ce Univ. of Michigan, Ann Arbor, Mich.

WEESH MLIb Ur oO. A ch Lee teetetateacyans 24 Upper Mountain Ave., Montclair, N. J.

WESTON, WILLIAM H., Jr., Ph.D., ’16......... Fed. Hort. Board, Washington, D. C.

WHEELER phim eeue ness ZOO cir sa Reicpenae. ciate s aos 79 Chapel St., Albany, N. Y.

WirEPER Yee yMie dvi dD) PhiG. VOOM mass ccisiea alsin: 2342 Albion Pl., St. Louis, Mo.

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WiIsmMER NE Trin Vie IO a. Es fe ia a Kans. St. Agri. Coll., Manhattan, Kans.

IWGDSEDAGEKS JERRY EDWARD! Phi D) 71157 oi) ae ee eiar spice eit Moscow, Idaho

Wotcortt, RoBert Henry, A.M., M.D., ’98...... Univ. of Nebraska, Lincoln, Neb.

Woon) ARTHURIKGNG Hae 225 422 neck wean ees 61 E. 65th St., New York, N. Y.

Yosuipa, SADAO, ’20...Pathological Department, Osaka Medical College, Osaka, Japan

ZOOKMDAVADY BAS eOOM aati «so dae eto 965 Holliston, Ave., Pasadena, Cal.

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INDEX

Age, Growth and Scale Characters of

Mullets, 199.

Algae, An Ecological Study of, 51

Andersen, Emma N. and Walker, Elda

R., An Ecological Study of the Algae

of some Sandhill Lakes, 51

Bladder Fluke, from the Frog, 142

Braun, Micropterygidae, 163

Cestodarian Parasite, Glaridacris

catostomi gen. nov., sp. nov., 5

Cobb, N. A., Micro-technique, 231

Cooper, A. R., Glaridacris catostomi gen.

nov., sp. nov.: A Cestodarian Parasite,

Crampton, Origin and Significance of

Metamorphosis, 165

Custodian’s Report for the Years 1918

and 1919, 89

Dark-field Microscopy, Modern, 95

Devil’s Lake, Protozoa of, 167

Edmondson, C. H., Protozoa of the

Devil’s Lake Complex, North Dakota,

167

Enchytraeidae, The Genera of, 25

Entomological Abstracts, 163

Filariasis in U. S., 164

Fluke, from the Frog, 142

Francis, Filariasis in U. S., 164

Frog, New Bladder Fluke from, 142

Gage, S. H., Modern Dark-field Micro-

scopy, 95

Glaridacris catostomi gen. nov., sp. nov.:

A Cestodarian Parasite, 5

254

Growth of the Mullets, 199

Guberlet, J. E., A New Bladder Fluke

from the Frog, 142

Henderson, W. F., Report of the Treas-

urer, 92

Illustrations, Labeling, 149

Jacot, A. P., Age, Growth and Scale

Characters of the Mullets, 199.

Labeling Illustrations, 149

Leeches, Considered as Oligochaeta Modi-

fied for a Predatory Life, 86

Meeting, Minutes of the St. Louis, 89

Metamorphosis, Origin and Significance

of, 165

Metcalf, Z. P., Labeling Illustrations, 149

Micropterygidae, 163

Micro-technique, Methods and Appara-

tus, 231

Microscopy, Dark-field, 95

Minutes of the St. Louis Meeting, 89

Mugil cephalus, 199

Mugil curema, 199

Mullets, Age, Growth and Scale Char-

acters of, 199

Oligochaeta, Leeches considered as, 86

Oligochaeta, The Genera of the Enchy-

traeidae, 25

Patterson, Polyembryony and Sex, 164

Pflaum, M., Custodian’s Report, 89.

Polyembryony and Sex, 164

Protozoa of the Devil’s Lake Complex,

North Dakota, 167

AMERICAN MICROSCOPICAL SOCIETY

Report of Auditing Committee on Treas-

urer’s Accounts, 92

Report of the Treasurer, H. J. Van Cleave,

Report of the Treasurer, W. F. Hender-

son, 92

Sandhill Lakes, An Ecological Study of

the Algae of, 51

Scale Characters of the Mullets, 199

Sex, Polyembryony and, 164

Smith, F., Leeches Considered as Oligo-

chaeta, 86

Spencer-Tolles Fund, Custodian’s Report,

St. Louis, Minutes of the Meeting, 89

259

dig

Tillyard, Position of Micropterygidae,

163

Van Cleave, H. J., Report of the Treas-

urer, 90

Walker, Elda R., Andersen, Emma N.,

and, An Ecological Study of the Algae

of Some Sandhill Lakes, 51

Welch, P. S., Entomological Abstracts,

163

Welch, P.S., Genera of the Enchytraeidae

(Oligochaeta), 25

Welch, P. S., Minutes of the St. Louis

Meeting, 89

TRANSACTIONS

OF THE

American

Microscopical Society

ORGANIZED 1878 INCORPORATED 1891

PUBLISHED QUARTERLY

BY THE SOCIETY

EDITED BY THE SECRETARY

PAUL S; WELCH

ANN ARBOR, MICHIGAN

VOLUME XL

NUMBER ONE

Entered as Second-class Matter August 43, 1918, at the Post-office at Menasha.

Wisconsin, under Act of March 3, 1879. Acceptance for mailing at the

special rate of postage provided for in Section 1103, of the

Act of October 3, 1917, authorized Oct. 21, 1918

The Colleniate Press

GerorcE BanTA PUBLISHING CoMPANY

MENASBA, WISCONSIN

1921

TABLE OF CONTENTS

For VotuME XL, Number 1, January, 1921

Acanthocephala from the Eel, with one plate, by H. J. VanCleave................... 1

A Brief Study of the Range of Error in Micro-Enumeration, by W. E. Allen............. 14

Department of Methods, Reviews, Abstracts, and Briefer Articles

Remarks on the Life-history and the Scale Characters of American Mullets, by

COL SHUpDS 2 Bicsocte dais aoe a SOS Pecan he ys ibe eo See 26

Spring Migration in the Crayfish, Cambarus argillicola Faxon, by H. Cummins... 28

Preparing Collections of the Mollusca for Exhibition and Study, with five figures, by

1 yap Coa. 83 co) cite ener ne cio Bini seek eee eee Reni ck ances. oc c 31

Proceedings of the American Microscopical Society............-2.00-0e-eeeceeeeeeee 47

TRANSACTIONS

American Microscopical Society

(Published in Quarterly Instalments)

Vol. XL JANUARY, 1921 No. 1

ACANTHOCEPHALA FROM THE EEL!

H. J. VAN CLEAVE

Introduction

Materials and Acknowledgments

Records of Acanthocephala infesting Anguilla chrysypa

Descriptions of Species

Tanaorhamphus ambiguus n. sp.

Neoechinorhynchus cylindratus (Van C., 1913)

Echinorhynchus coregoni Linkins in Van C., 1919

Echinorhynchus thecatus Linton, 1891

Uncertain Identifications of Species

Neoechinorhynchus agilis (Rud.)

Koleops anguilla Lockwood

Echinorhynchus clavaeceps Zeder

Echinorhynchus globulosus Rud.

Conclusions

Acanthocephala from Anguilla vulgaris

Literature Cited

Explanation of Plate

INTRODUCTION

Because of their migratory habits, eels offer some fascinating problems

to investigators interested in the geographical distribution of parasitic

organisms. The migrations of most other animals are between places

of essentially similar surroundings and for that reason little evidence is

available regarding the source of infestations borne by the wanderers.

Very little is known of the definite limitations to the distribution of para-

sites of purely limnetic and terrestrial organisms. On the other hand most

of the larger groups of parasitic organisms include species or even genera

which are distinctively limited to either a marine or a fresh-water habitat.

One of the most interesting and perplexing features of the parasitism of

migratory organisms is the fact that they may carry throughout their

range mature parasites the larval stages of which have been acquired in a

very narrowly restricted locality. Because of the fairly sharp contrast

‘Contributions from the Zoological Laboratory of the University of Illinois, No. 175.

2 . H. J. VAN CLEAVE

between the marine and the fresh-water faunas, organisms like the eel

which wander from one to the other offer exceptional opportunity for the

study of the influence of migration upon the parasitic fauna of the host.

In such migrants it becomes possible to recognize the source of a parasitic

infestation with much greater certainty than is possible when the migra-

tory movements are between localities of approximately the same physical

environment.

The investigations of Zschokke upon the parasites of salmon stand as

probably the most comprehensive work in the literature upon the parasites

of migratory fishes. In his studies he included observations upon individ-

uals ascending the Rhine and upon others taken from the Baltic Sea.

_ From the former he recorded twenty species of parasitic worms all of which

were apparently acquired by the salmon before they left the ocean. The

lack of parasites of limnetic origin is obviously correlated with the fact

that all evidence seems to indicate that the Rhine salmon commonly

refuses to take food after entering the river. In the Baltic salmon, however,

Zschokke found some typically fresh-water species of parasites occurring

with the ostensibly marine species. Zschokke’s work has been summarized

by Ward (1910:1160) in the following manner: ‘‘The Rhine salmon shelters

a purely marine parasitic fauna, while the Baltic salmon reckons many

limnetic forms among its parasitic guests. This remarkable condition

finds explanation in the continued feeding of the latter type, even in ~

fresh water, and the resulting enrichment of its parasitic fauna with

limnetic forms when it returns to the sea.”

Unfortunately, the parasites of the eels have not been investigated as

thoroughly as have those of the salmon and little effort has been directed

toward interpretation of the isolated observations. In the present paper

attention will be confined to the Acanthocephala infesting the eel. A

number of investigators have compiled lists of the parasites of the European

eel, Anguilla vulgaris, but to the present time very little material has been

available regarding the Acanthocephala of the North American species,

Anguilla chrysypa.

Any attempt to determine the origin of the parasitic fauna of the eel

must take into account both the pelagic and the limnetic periods in the

life cycle of an individual. Regarding the source of infestation of the

adult eel found in fresh-water, at least four possibilities must be con-

sidered:

1. The parasites may have been acquired in the marine habitat and

are retained for a longer or shorter period of time after the host enters

fresh-water. If the host remains in fresh-water for a period longer than

the life of the individual parasites constituting the original infestation,

the intestine would ultimately be freed from its parasites;

SS - . <_ — .

ACANTHOCEPHALA FROM THE EEL 3

2. Eggs or larvae of parasites acquired while the host was in the

ocean when discharged into fresh-water may succeed in becoming estab-

lished in the new habitat through their adaptability to entirely new

primary: hosts; A

3. The parasites may represent an entirely new infestation of typically

fresh-water species acquired after the loss of any marine species that

might have been carried at the time of leaving the ocean;

4, Marine and fresh-water species may become commingled in the

body of the same host individual.

A number of the earlier records of Acanthocephala from Anguilla

chrysypa have contained apparent misidentifications of species. In some

of these instances the present writer has had the opportunity of examining

the materials and has found that in one instance of a reputed occurrence

of a marine European species from A. chrysypa the specimens really belong

to a typically fresh-water species probably restricted to North America.

In the present paper the writer hopes to analyse the earlier records of

Acanthocephala from the eel and to add a considerable bulk of original

data which has accrued from the study of several important collections.

MATERIALS AND ACKNOWLEDGMENTS

In the investigation of this problem I have had the privilege of examin-

ing specimens and of incorporating data from a number of important

parasite collections. Specimens from the U.S. National Museum collected

by E. Linton and by A. Hassall have been especially interesting in this

connection. Through the courtesy of C. C. Adams specimens secured by

H. S. Pratt and F. C. Baker in the course of investigations by the New

York State College of Forestry and the U. S. Bureau of Fisheries upon the

fauna of Oneida Lake, New York, have been available for study.

In all the writer has examined four species of Acanthocephala from

the intestine of Anguilla chrysypa, of which one represents a new species

described here for the first time. In attempting to make a correct dis-

position of the species mentioned in the earlier works the writer has been

extremely fortunate in being able to examine collections which have

verified surmised incorrect determinations and have made corrections

in the identification possible.

For data concerning the Acanthocephala of Anguilla vulgaris the writer

has found it necessary to utilize the published records of European investi-

gators. In a number of instances, where records are the result of compila-

tion from various sources, there are probably errors in the determination

of the species. Through the kindness of Professor K. M. Levander of

Finland, I have been permitted to study the Acanthocephala encountered

in the course of his investigations upon the food and parasites of the eel.

4 H. J. VAN CLEAVE

RECORDS OF ACANTHOCEPHALA INFESTING Anguilla chrysypa

The first reference to the occurrence of an acanthocephalan in the

American eel is that given by Samuel Lockwood in t872. Ina highly enter-

taining but superficial manner he described a new genus and new species

of acanthocephalan from a cyst in the intestine of an eel ascribing to this

new form the name Koleops anguilla. Unfortunately his description and

his figures, based upon the study of a single living specimen, are so general-

ized that they possess but little of scientific value. Apparently the acan-

thocephalan from the hog is the only other species of these worms that

had ever come to his attention. Both his specific description and his

generic diagnosis consist in simple enumeration of a few points of difference

between his specimen and “‘Echinorhynchus gigas.’ Allof these differences,

with the possible exception of the poorly described proboscis, might apply

equally to any one of numerous genera belonging to the family Echinorhyn-

chidae. Consequently both the genus and the species stand as unrecog-

nizable.

Joseph Leidy in his extensive pioneer researches on fish parasites

has made no mention of ever encountering Acanthocephala in the American

eel.

Edwin Linton, in various reports, has given notice of the occurrence

of Acanthocephala in this host. He identified (1889 and 1901) as Echino-

rhynchus agilis specimens which I have determined (Van Cleave, 1913) as

Neoechinorhynchus cylindratus. Other specimens (1901) he believed to

belong to the European species E. globulosus. According to his statement

these last named specimens were from the collection of the U. S. National

Museum. I have examined specimens from this same collection which

bear a label indicating that they were identified as EF. globulosus by Linton

and seem to be the same individuals referred to in his paper just cited.

A thorough study of these specimens has demonstrated that they represent

an undescribed species of the genus Tanaorhamphus which is described

later in this same paper.

Data concerning the parasites of fishes for A Biological Survey of the

Waters of Woods Hole and Vicinity (Sumner, Osborn, and Cole, 1913)

were furnished by Linton. Under Anguilla rostrata (=A. chrysypa) but two

species of Acanthocephala were mentioned, namely, “FE. clavaeceps and

E. globulosus.’’ The first of these is apparently a renaming of what Linton

had earlier identified as ‘FE. agilis’”’ and what I have more recently shown

to be Neoechinorhynchus cylindratus.

In the investigation of parasites of fishes from the Illinois River eight

eels were examined by the writer (Van Cleave, 1919) but no Acanthocephala

were discovered. In addition the writer encountered numerous negative

ACANTHOCEPHALA FROM THE EEL 2)

records as the result of the examination of large numbers of eels taken

from salt water at Woods Hole, Mass., during the month of August.

DESCRIPTIONS OF SPECIES

In the light of the present investigation four valid species of Acan-

thocephala are to be attributed to Anguilla chrysypa. Of these one species

is new and two of the remaining ones are reported for the first time from

this host.

Tanaorhamphus ambiguus n. sp.

Definition. With the characters of the genus: Type female 7.9 mm.

long; maximum diameter 0.67 mm.; diameter of posterior region of body

about 0.35 mm. Proboscis cylindrical, 0.77 mm. long and 0.19 mm. in

diameter, armed with twenty longitudinal rows of about sixteen hooks

each. Hooks near middle of proboscis 41 to 47u long, with the hook at

base of each row about 24yu long. Proboscis receptacle 0.96 mm. long with

wall composed of a single muscular layer. Central nervous ganglion at

base of proboscis receptacle. Lemnisci 2.3 mm. long, cylindrical. In the

type female each lemniscus contains three giant nuclei. Subcuticular

giant nuclei arranged five in mid-dorsal line of body and one in mid-ventral

line.

Males have not been studied. Embryos not fully formed in specimens

under observation.

Type host: Anguilla chrysypa.

Type female collected by Albert Hassall at Baltimore, Maryland,

May 30, 1891. This specimen with others of NV. cylindratus and E. coregont

was deposited in the U. S. National Museum under catalog number 6301

of the Hassall Collection. The writer has examined two additional indi-

viduals helonging to this same species from the National Museum. These

bore the catalog number 6471 and Linton’s identification as E. globulosus.

Neither of these was in as good condition for study as the one selected as

type. One specimen, apparently a male of this species, had at some time

become dried out and for that reason an accurate determination of internal

structure is impossible. Linton (1901:435) states that there are three

specimens in the National Museum collection, but it is entirely probable

that one specimen was entirely lost at the same time that the damage was

done to the male. In the reference just cited, Linton states that this

species was numerous in collections made August 7 and 28, 1899.

One of the marked peculiarities of the type of this species lies in the

number of nuclei within the lemnisci. All other species of Neoechino-

rhynchidae from North America examined by the writer agree in possessing

two gidnt nuclei in one lemniscus and one in the other. The lemnisci of

the only other specimen of this species are entirely obscured by the develop-

(Of H. J. VAN CLEAVE

ing egg masses within the body cavity. In figure 5 the three nuclei of

one lemniscus are shown while two of those in the other lemniscus occur

in the region where the two organs overlap. Until additional specimens

are secured for study it is impossible to determine whether this deviation

from the customary number of nuclei is an individual abnormality or a

character of this species.

But one other species has been described for the genus Tanaorhamphus

and it is apparently very sharply restricted in its occurrence, having been

found in but one host, the gizzard shad (Dorosoma cepedianum) from the

Illinois River.

Neoe hinorhynchus cylindratus (Van Cleave, 1913)

Echinorhynchus agilis of Linton 1889 and 1901

Echinorhynchus clavaeceps of Linton in Sumner, Osborn

and Cole, 1913.

Definition. With the characters of the genus. Body almost cylindrical

except in immature forms which have the posterior region gradually

attenuated. Proboscis approximately globular, slightly broader than

long (0.172 by 0.150 mm.), provided with three circles of six hooks each.

Hooks of terminal circle 79 to 97y long, each bearing a root 58y long and

29u wide inside the tissue of the proboscis wall. Hooks of middle circle

about 37 long, without reflexed root. Basal hooks 21 to 25 » long, simple,

* thorn-like. Embryos within body cavity of gravid female 49 to 51y long

and 15 to 2ip broad. |

Specimens collected by Professor Linton at Woods Hole, Mass., were

identified as belonging to this species by the writer in 1913 (p. 188).

Additional records of the occurrence of this species in the eel have been ~

discovered since that time. Six juvenile specimens were taken from the

intestine of an eel from Oneida Lake, New York, in the materials collected

by F. C. Baker and H. S. Pratt. Three other eels from the same locality

were found to harbor Acanthocephala of another species but none of

N. cylindratus. Two individuals of this species were encountered in an

eel examined by A. Hassall at Baltimore, Md., along with the type of

T. ambiguus and two individuals of E. coregont. :

N. cylindratus is distinctively a fresh-water species, the development

of which is unknown. Without much question this species gains entrance

into the eels after they enter fresh-water. The infrequence of its occurrence

in the eel and the lightness of individual cases of infestation would indicate

that the eel does not serve as an important definitive host for this species

of parasite.

Echinorhynchus coregoni Linkins in Van Cleave, 1919

Definition. With the characters of the genus. Proboscis cylindrical,

carrying twelve to fifteen longitudinal rows of ten or eleven hooks each.

} Ro Waiieetuset *

ACANTHOCEPHALA FROM THE EEL 7

Hooks not crowded on proboscis. Basal hooks 28 to 53u long; those on

middle of proboscis 65 to 80u long; those near anterior tip smaller and

weaker than those near middle of proboscis. Ventral hooks slightly

larger and stronger than dorsal hooks. Lemnisci not longer than proboscis

receptacle. Cement glands of male in a compact mass. Embryos within

body cavity of gravid female 51 to 91 long by 17 to 204 wide, with an

approximateiy globular prolongation of the middle membrane at each

pole.

This species was originally described from the Great Lakes but present

indications seem to suggest that its distribution is fairly broad. The

writer has recently (1920:6) recorded its occurrence in fresh-water hosts

from the arctic regions of this continent. Two individuals of this species

were taken from an eel at Baltimore, Md., by Hassall. As indicated

earlier in this paper this species was found associated with NV. cylzndratus

and T. ambiguus.

Echinorhynchus thecatus Linton, 1891

Definition. With the characters of the genus. Proboscis when fully

extended approximately perpendicular to main axis of the body or forming

an acute angle with the axis. Proboscis usually about 1 mm. long. Pro-

boscis receptacle long and slender, about 1.5 times the length of the

proboscis. Hooks alternate in arrangement; in twelve longitudinal rows

of twelve or thirteen hooks each; those at base of proboscis 41 to 53 long;

near middle of proboscis 71 long; each hook surrounded by a conspicuous

cuticular elevation which frequently completely ensheathes the basal hooks

of each row. Lemnisci long and slender, about 1.5 times the length of the

receptacle. Embryos within body cavity of gravid female 80 to 110u

long by 24 to 30u broad.

This species is distinctively a member of the fresh-water group. It

has been reported from fishes taken from the ocean but these hosts fre-

quent fresh-water habitats. The general outline of the life cycle of this

species has been worked out (Van Cleave, 1921) thereby offering additional

evidence of its close association with the fresh-water fauna where it occurs

as one of the most characteristic acanthocephalan parasites of fresh-water

fishes of North America. In the collections from Oneida Lake, referred to

above, this species was found in fairly large numbers in the intestine of all of

the eels carrying acanthocephalan infestation. This constitutes the first

record of the occurrence of E. thecatus from Anguilla chrysypa.

UNCERTAIN IDENTIFICATIONS

Among the earlier American workers in parasitology there was a

marked tendency to ascribe to specimens of Acanthocephala found in

American hosts the names of European species. Thus much of the older

8 H. J. VAN CLEAVE

literature is replete with instances of the recorded occurrence of European

species in American hosts, whereas in the cases that have been investigated

carefully it has been shown that with few exceptions the acanthocephalan

fauna is distinctively different on the two continents. From point of view

of biology and of distribution of the individual species it is extremely

desirable that these errors in determination be cited and corrected if

possible. In the following section the writer has attempted to analyse

some of these older records in the light of more recent investigations.

Neoechinorhynchus agilis (Rudolphi)

E. agilis Rud.

The writer has examined some of Rudolphi’s type specimens (Van

Cleave, 1919:246) of this species and is confident that all records of the

occurrence of this species on the American continent are based upon

erroneous identifications. It seems probable that N. agilis is rather

sharply restricted in distribution to the Mediterranean region.

As indicated previously, Linton (1889 and 1901) identified under ae

name specimens from the el which unquestionably belong to the species

N. cylindratus, the commonest representative of the genus in North

American fishes. Furthermore NV. agilis is a marine species while J.

cylindratus is definitely associated with the fresh-water habitat.

Koleops anguilla Lockwood

In the description of this genus and species not a single diagnostic

character is given. Characters which are discussed are all contrasted

with the conditions found in “Echinorhynchus gigas.” Apparently this

last named species is the only other species of the group with which the

-describer was acquainted. Supposed peculiarities of the new form are in

reality common to all acanthocephalans except members of the family

Gigantorhynchidae. Even the drawings are not available as a supplement

to the meagre description for no detail of structure, either external or in-

ternal, is shown.

“Echinorhynchus clavaeceps” Zeder

This species has been considered by modern writers as a synonym of

Neoechinorhynchus rutili (Miller) which infests fresh-water fishes of

central Europe. In his later works Linton apparently uses this name to

replace his earlier identifications of E. agilis. It is highly probable that

specimens identified as E. clavaeceps belong in reality to the species WV.

cylindratus.

Echinorhynchus globulosus Rudolphi

Linton (1901:435) identified acanthocephalans from the eel which

were deposited in the U. S. National Museum as belonging to this speciese

ACANTHOCEPHALA FROM THE EEL 9

These same specimens have been found to belong to a previously un-

described species of the genus Tanaorhamphus which is described elsewhere

in this paper as 7. ambiguus.

CONCLUSIONS

Four valid species of Acanthocephala are known to occur in the intestine

of Anguilla chrysypa. All four of these are distinctively American species,

of which three are known to occur only in fresh-water fishes. The fourth

belongs to a genus which includes but one other species and it is restricted

to a single species of fresh-water host. In all of the instances that have

been examined carefully there is no evidence of marine species of Acan-

thocephala inhabiting the intestine of Anguilla chrysypa. In this connec-

tion a comparison with data from records of infestation of the European

eel is interesting.

ACANTHOCEPHALA FROM Anguilla vulgaris

Lack of unanimity in determining the synonymy of many species of

Acanthocephala infesting fishes makes direct comparisons between records

of European investigators extremely difficult. Before the time of Lihe

many specific names, the authors of which recognized them as synonyms

of earlier workers, had come into very general usage. Thus, for instance,

Rudolphi (1802:53) gave the new name Echinorhynchus angustatus to Ech.

lucii O. F. Miller (1778). In spite of this obvious renaming most European

workers have, until very recently, failed to recognize the priority of the

name £. /ucii and have continued to use the synonym instead of the valid

name of the species. Numerous instances of similar nature are encountered

in the literature dealing with the Acanthocephala of fishes. In the follow-

ing paragraphs names of commonly recognized synonyms have been

replaced by the valid names of the species in question.

In his ‘Register der Acanthocephalen und parasitischen Plattwiirmer,”

_Liihe (1911:91) has compiled a list of eighteen species of parasitic worms

(exclusive of nematodes) which have been recorded from the eel in central

Europe. Only six of these are recognized as strictly marine, while all

seven species of the Acanthocephala included in his list are typically

fresh-water forms with the possible exception of the encysted larvae of an

undetermined species of the genus Corynosoma.

K. M. Levander (1909) in his admirable contribution to the knowledge

of the food and parasites of the fishes of Finland has utilized the names

current in the literature for the species of Acanthocephala encountered in

his investigations. His data upon the parasites of Anguilla vulgaris

include three species of Acanthocephala from this host. Since the time of

the publication of his work the names Echinorhynchus angustatus, E.

10 H, J. VAN CLEAVE

globulosus, and E. clavaeceps have become recognized as synonyms of

Acanthocephalus lucii (Miller), Ac. anguillae (Miiller), and Neoechino-

rhynchus rutili (Miiller) respectively. Professor Levander very kindly

sent me specimens from Anguilla vulgaris and I have been able to confirm

his determinations with changes to the valid forms of the names mentioned

above.

Porta (1905) listed nine species of Acanthocephala from the eel of which

two, Echinorhynchus gadi and Acanthocephalus propinquus, are supposedly

of mariné origin. However, Porta’s records in this work are the result

of promiscuous compilation and it is entirely possible that either or both

of these species have been misidentified.

The following table gives some of the more important contributions to

the knowledge of the acanthocephalan fauna of the European eel.

TABLE I

Acanthocephala Reported from Anguilla vulgaris

Speci Porta Liihe Levander Stossich

Boake | 1905 1911 1909 1885-1898

at =e

ais ot

sh at

Neoechinorhynchus rutili

Acanthocephalus lucii

Acanthocephalus anguillae

Acanthocephalus propinquus

Echinorhynchus gadi

Echinorhynchus clavula

Echinorhynchus lateralis

Echinorhynchus salmonts

Pomphorhynchus laevis

Echinorhynchus miliarius (encysted) |

Corynosoma sp? (encysted)

++ +4+4+4++4+44

+ ++ +

In the above table are listed eleven species of Acanthocephala from .

Anguilla vulgaris. Two of these are larval forms thus leaving nine species

as recorded from the intestine of this host. One of these nine species,

A. propinquus is restricted to marine fishes while another, E. gadt, is

considered by European investigators as a typically marine species which is

brought into fresh-water only through the agency of migratory fishes.

Thus of the species of Acanthocephala infesting Anguilla vulgaris two are |

apparently acquired during the marine phase of its existence and the

remaining seven are introduced into the intestine of the eel while it is an

inhabitant of fresh-water. From the foregoing it will be seen that the two

species of eel differ not only in regard to the species of Acanthocephala

harbored but likewise differ in the origin of their infestations. The Acan-_

thocephala of Anguilla chrysypa seem to be entirely of fresh-water origin

ACANTHOCEPHALA FROM THE EEL Thal

while those of A. vulgaris comprise a mixture of marine and fresh-water

species.

LITERATURE CITED

LEVANDER, K. M.

1909. Beobachtungen iiber die Nahrung und die Parasiten der Fische des Finnischen

Meerbusens. Finnlindische Hydrographisch-Biologische Untersuchungen.

No. 5.

Linton, E.

1889. Notes on Entozoa of Marine fishes of New England, with Descriptions of several

New Species. Rep. Comm’r. U.S. Comm. Fish and Fisheries for 1886; 453-511.

1901. Parasites of fishes of the Woods Hole region. Bul. U.S. Fish Comm. 19:405—-492.

1905. Parasites of fishes of Beaufort, North Carolina. Bul. U.S. Bur. Fish. 24: 321-428.

Locxwoop, S.

1872. A new entozoon from the eel. Amer. Nat. 6:449-454.

Line, M.

1911. Acanthocephalen. Die Siisswasserfauna Deutschlands, Heft 16. Jena.

Porta, A.

1905. Gli Echinorinchi dei Pesci. Arch. Zoologico 2:149-214.

Stossicu, M.

1885. Brani di elmintologia Tergestina, II serie. Bollett. Soc. Adriat. di scienze

naturali in Trieste, IX.

1886. Brani di elmintologia Tergestina, Serie Terza. Ibid. 9:1-7

1898. Saggio di una fauna elmintologica di Trieste e provincie contermini. Programma

della Civica Scuola Reale Superiore, 1898:1-162.

SuMNER, F.B., Osporn, R. C., and Cots, L. J.

1913. A Bielceteal Survey of the Waters of Woods Hole and VACEEYs Part II, Section

Ill. Bul. U. S. Bureau of Fish. 31.

VAN CLEAVE, H. J.

1913. The genus Neorhynchus in North America. Zool. Anz. 43:177-190.

1919. Acanthocephala from the Illinois River, with descriptions of species and

synopsis of the family Neoechinorhynchidae. Bul. Ill. Nat. Hist. Survey

Vol. 13, No. 8.

1920. Acanthocephala of the Canadian Arctic Expedition, 1913-1918. Rep. Can. Arct.

Expedit. 1913-1918 Vol. 9, Part E.

1921. Notes on the Life Cycle of Two Species of Acanthocephala from Freshwater

Fishes. Jour. Parasitol. 6:167-172.

Warp, H. B.

1910. Internal Parasites of the Sebago Salmon. Bul. Bureau of Fish. 28:1151-1194.

ZSCHOKKE, F.

1889. Erster Beitrag zur Parasitenfauna von Trutta salar. Verhandl. d. Naturforsch.

Gesellsch. i. Basel, 8:761-795.

1891. Die Parasitenfauna von Truite salar. Centralbl. f. Bakt. u. Parasitenk. 10:

694-699; 738-745; 792-801; 829-838.

1902. Marine Schmarotzer in Siisswasserfischen. Verhandl. d. Naturforsch. Gesellsch.

i. Basel, 16:118-157.

12 H. J. VAN CLEAVE

EXPLANATION OF PLATE

Each figure is accompanied by a scale which indicates the magnification. The scales

accompanying figures 3 and 5 have the value of 1 mm., all others on this plate represent

0.05 mm.

All figures were drawn from stained whole mounts in damar with the aid of a camera

lucida.

Fig. 1. Profile view, portion of a single row of hooks from proboscis of E. thecatus,

showing characteristic cuticular prominence around each hook.

Fig. 2. Proboscis of NV. cylindratus, surface view, showing characteristic arrangement of

hooks.

Fig. 3. Female of E. coregoni showing general arrangement of organs. Note especially

the lemniscus (/) which is shorter than the receptacle of the proboscis (r).

Fig. 4. Embryo from body cavity of female of E. coregoni from A. chrysypa.

Fig. 5. Tanaorhamphus ambiguus, type female, showing general arrangement of organs:

b-brain, e-egg mass, /-lemniscus, n—subcuticular giant nucleus, r-receptacle of the proboscis.

Fig. 6. Proboscis of type female of T. ambiguus.

ACANTHOCEPHALA FROM THE EEL

aw &2 SS —

eer oc. = Ss

— ae & SS ee RSS y <s ees

RSs — nn Lox i SK ‘ < Gs point

A BRIEF STUDY OF THE RANGE OF. ERROR IN

MICRO-ENUMERATION

W. E. ALLEN

Like many other-people who have undertaken statistical study of

microscopic organisms, I worked for a long time without any very great.

effort to determine the accuracy of enumeration. This was probably due

to five reasons: first, various authorities state that by counting into so

many hundreds or thousands, the limits of accuracy are reached, second,

because two or three trial counts indicated substantial agreement, third,

because the series of counts seemed to follow a normal sequence, fourth,

because the insistent demands of routine work made it difficult to under-

take a study of this sort, fifth, because it seemed that if great care were

observed in handling and counting there was no great probability of

improving matters by making such a study.

But it so happened that my co-worker in the Scripps Institution, the

late Mr. E. L. Michael, when looking over the manuscript of my paper on

the plankton of the San Joaquin River, raised the question as to the

accuracy of my counts. We discussed the matter at various times and

he always remained sceptical as to my guess that my counts were not

in error more than plus or minus ten per cent. So, finally, when I got

settled down to my regular program of work on marine phytoplankton,

after adopting the measured water method of collecting, it became neces-

sary to get more definite information concerning the accuracy of the

counts.

I have made no thorough search of plankton literature for a record of ~

such studies, but I have had access to the most important European and

American papers, which I have scanned rather hastily without finding

any indication of sucha record. Hence, it seemed to me that my experience

might be of some value to other workers in this or similar lines. I also

thought it might lead some one to make a more thorough study of this

interesting problem.

To one who has not given any serious thought to the matter, it may

appear that the counting of microscopic organisms is quite similar to the

counting of any common objects such as beans or apples. In the case of

the plankton organisms, this is not true for several reasons. In the first

place, there is usually a certain amount of dirt or débris likely to hide some

individuals. Then there is the fact that if one wishes to be sure of getting

a required number of the organisms, he must (because he cannot see them)

filter a sufficient amount of water to give an actual excess over what he is

able to count. He must then (except in the use of one or two highly

RANGE OF ERROR IN MICRO-ENUMERATION 15

specialized methods) take a fractional part of his catch and estimate

the total from the number found in this fractional part. The extraction

of the fractional part from the whole and the even spreading of this under

the microscope for counting is an important phase in the routine of plank-

ton counting. One can take a pint of beans and after counting the number

contained, compute fairly accurately the number in a bushel or a car,

but he cannot take the individual organisms one by one from his fractional

measure and make such an exact estimate. Furthermore, the microscopic

things are necessarily handled in fluid through which they should be nearly

uniformly spread for count. If one had to take beans mixed in four or

- five or one thousand times their volume of water and make the count while

they were in the mixture, he might have a little better idea of the difficulty

of microscopic counting. Furthermore, there is the matter of eye fatigue

and the difficulty of recording the count as it progresses.

A few days after beginning the work of collecting by the measured

water method on September 1, 1919, I made a beginning at a study of

accuracy of enumeration which I was’ obliged to discontinue. I did,

however, make eight counts of a catch (7728) taken in the forenoon of

September 6. The results of these counts are partly summarized in

table I.

TABLE I

Eight counts by non standard method, of Catch 7728

No. of Count . 1 | ye 3 4 5 6 7 8

S =| ee S| S| | al Boar ee S| S|

fa) a a fa a a fa a

S S S 3S Ss S S S

oft] 0) Se ees Sse = (8 | | em k= ale Ree We v= a

Se) ie Sel SUN Be Sah Eh Sy eet est feel tt Ue se a | Ve Me 3

o oO o i) o Oo ° Cy) (=) a) lo) i) ° o o o >

: S/S Se) aye ale aly ep al el al a] a ee] al

Diatom cells....} 1) 91]! 17} 40) 8] 33}/ 5) 41/) 11) 8!) 3} 251] 24/100}) 27/150)| 12

Dinoflagellate

CCl ae ae sf 350} 6)|334) 1/228) 14)/292) 10/|275| 17||398} 23)|406| 24|/294) 11329

Diatom and Di- :

noflagellate cells|351| 3}/351] 3//296| 13]|297] 12]|/286| 16||401 15||/430| 21||321] 6)|341

The series was not very good because the conditions of counting were

not nearly enough alike, first because the first four counts were made

on the same day and the other four at intervals of one to four days, second,

because the material was kept in the mixing tube throughout the series,

merely being shaken up after return of the fractional amount from the

slide after each count. Furthermore, the number of diatoms was so small

16 W. E. ALLEN

as to make that part of the count unreliable and none of the counts were

carried quite so far as necessary to give sufficiently dependable results.

In spite of these deficiencies the table shows that the series was sufficiently

good to be considered statistically significant. Thus it appears that five

out of the eight counts of dinoflagellates showed deviations from the mean

of less than fifteen per cent and the highest per cent of such deviation

was twenty-four. The showing for total numbers of cells is even better,

a fact which calls attention to the general probability that a deviation of

count in one group may be largely obscured by the count of another group

when the two are combined for a total.

Although the table gives great emphasis to the point that the count of

such a few individuals as those of the diatoms is valueless as a basis of

generalization, it should not be forgotten that such a count may be worth

recording because of its positive indication of presence of organisms. Fur-

thermore, the system of random sampling to which we are usually forced,

may sometimes lead to just as great differences in estimating the plankton

population as is represented here. The significance of both errors becomes

rapidly less with increase in numbers of samples.

Constantly harassed by the feeling that I ought to still further improve

my basis of judgment as to the values of individual counts, I finally

returned to a study of the problem on January 21, 1920, and gave it a

large part of my time for the next two months. First I took some care

in the selection of a catch for study and finally decided on the one (8102)

for 8 p. M. on January 11, 1920, because it showed fairly good representa-

tion of both diatoms and dinoflagellates and also because it was relatively

free from dirt. First I made ten counts of this catch at intervals of one

day or more, the slide being emptied into the mixing tube each time, but

the whole being left there instead of being returned to the bottle. The

summary of results for this series is shown in table IT.

TABLE II

Ten counts of Catch 8102

Percentage of Ceviation from the mean

ee of Gon yr carne acti ‘a 2 3 4 nee 6 i 8 9 aon

Motal;Diatom'Cellsa tee ee a ZOOM TS a Ome eis 28 S10) 220 me

Total Dinoflagellate Cells........ ~aiis {2} 3] 91| 2/33 |45 [20 | 18

Above totals combined.......... 23 ce tint eae 24 So) lil i} ati 6 Bo

In this series a few of the more abundant organisms were only counted

on one tenth or one twentieth of the slide although most species were

RANGE OF ERROR IN MICRO-ENUMERATION 17

counted on one fourth. At any rate, there is some probability that the

counts of those forms which were most abundant were not carried quite

far enough to yield really satisfactory results. Even so, the table shows

that in only three counts out of the ten was there more than fifteen per cent

deviation from the mean in total numbers of organisms and of diatoms and

that there was similar deviation in only four out of the ten totals of dino-

flagellates. Stated in another way, the showing is that sixty to seventy

per cent of the counts deviated from the mean by not more than plus or

minus fifteen per cent. .

A momentary inspection of table II shows that the fourth count was

the only one in which the deviation exceeded thirty-three per cent and

that the enormous deviation in that case was due to some difference in

the count of diatoms. Three possible causes of this great deviation

TaBLeE III

Two counts each of ten successive catches

Catch Régular Recount Average Per cent of

Number Count Deviation

Total Diatoms........ atete, 1814 1463 24

8102 /Total Dinoflagellates... 468 576 522 10

Combined totals....... 1580 2390 z 1985 20

Total Diatoms........ 1572 2460 2016 22

8104 |Total Dinoflagellates... Zion © 440 358 pre. MES

Combined totals....... 1848 2900 2374 22

Total Diatoms........ 2098 2011 2054 2

8105 |Total Dinoflagellates. .. 358 404 381 S 6

Combined totals....... 2456 2415 2435 5 1

Total Diatoms........ 2898 2132 2515 “15 ia

8107 |Total Dinoflagellates... 256 568 412 38

Combined totals....... 3154 2700 2927 8

Total Diatoms........ 3298 2336 2817 a)

8108 |Total Dinoflagellates... 372 304. 383 3

_ a fe fe

Combined totals....... 3670 2730 3200 15

W. E. ALLEN

TABLE III (Continued)

Two counts each of ten successive catches

Catch Regular Recount Average Per cent of

Number Count Deviation

otal Diatoms: se. -e-e | 1602 912 1257 28

8110 |Total Dinoflagellates... 324 426 375 14

Combined totals....... 1926 1338 1632 12

MotaluDiatomse.. -.0- = _ 682 728 705 3

8111 |Total Dinoflagellates. .. 164 4 178 171 4

Combined totals....... 846 906 876 3

Total Diatoms........ 1516 1386 i 1451 4

8113 |Total Dinoflagellates... 316 2 294 7

Combined totals....... 1832 1658 1745 5

Total Diatoms........ 988 1652 | 1320 25

8114 |Total Dinoflagellates... 424 460 | 442 4

Combined totals....... 1412 2112 1762 20

Potal Diatoms. >= 2" 2760 3348 | 3054 9

8116 |Total Dinoflagellates. .. 158 110 134 18

Combined totals....... 2918 3458 3188 8 -

require particular mention, first, it is extremely difficult to secure even

distribution of the diatoms in the counting cell, second, there was an

insufficient count of the more abundant diatoms, third, there may have

been a personal error in keeping the tally of the count. My own opinion

was that this particular deviation was mainly due to difference in the

evenness of spread of the diatoms through the suspending fluid and to.

insufficient count.

For further test of the matter before making any very definite change

in method, I then took ten consecutive catches and made two counts of

each. The results are partly shown in table ITI. ;

In this table it may be noted that there was only one deviation from

the mean of as much as thirty per cent. The fact that this one deviation

of thirty-eight per cent was in the count of dinoflagellates might lead

RANGE OF ERROR IN MICRO-ENUMERATION 19

one to think that dinoflagellates could not be any more readily mixed

through the fluid than diatoms. My detailed record of the count shows,

however, that this deviation was mainly due to differences in the count

of extremely minute forms which I have been including under the name

Gymnodinium sp. The difficulty of seeing these forms is quite sufficient

to account for this error under the circumstances. It appears, then, from

this particular series that the deviation in the count of the fairly visible

forms is usually well inside of thirty per cent.

In order to have some basis of judgment as to what increase in accuracy

might be expected if counts were made covering the whole slide instead

of a fractional part, I then made eight counts of a single catch using four

different mounts. For each mount I made one count over the whole

slide and one count over one fourth of the slide. The most important

results are summarized in table IV. "

TABLE IV

Counts of four mounts of Catch 8102

Percentage of deviation from mean

Number of mount ......4......- 1 2 3 4

Percentage of deviation in full slide counts

Diatomscolonies... =o... on. es 18 5 3 D5

Disttommcellsy keke yh soaks 11 8 3 0)

Dinoflagellate: cells.............. 5 3 3 5

MROCAINGELIGE 5 vase otk. gaceo ee ae 10 7 2 2

Percentage of deviation in fourth of slide counts

Diatomrcoloniess..... 5a. pa: 47 30 | 12 16

Diatom cells...... PRE IE ahai cir eR 30 53 25 25

Ae chandiae. Cells fe eT ae 8 12 12 | 8

Petaleele ct 38. ee ou 25 ae ae) | 3

While the four counts of each kind are not enough for definite con-

clusions, they are quite suggestive. It was not practicable to carry the

series further because of the great amount of time required. As it is there

is Strong indication that under usual conditions the count covering the full

slide is much more likely to approach the mean than is the count made

over some part only.

20 W. E. ALLEN

After giving the matter a good deal of thought, I came to the conclusion

that by standardizing mixing processes, much could be done toward

reducing the errors of the fractional counts. I, therefore, adopted the

practice of shaking the storage bottle for one minute before pouring the

contents into a mixing tube, and of reversing twenty times each mixing

tube used. All other manipulations had already been made as nearly

uniform as possible.

I then selected for study catch number 8104 of 8 A. m., January 12,

because of its close resemblance to 8102 which had become somewhat

unreliable from repeated handling. Twenty counts were made of samples

from this catch. At least twenty-four hours intervened between each two

counts and the total catch was returned to the storage bottle after each

count,so that the sampling might be done in approximately the same

way each time. With the first ten counts a test was made of the method

of selecting fractional areas in the cell. In one case the areas were selected

at intervals around the margin and in the other.a median zone lengthwise

of the cell and covering one fourth of its area was selected. The second

ten counts were made by the median zone-method but record was kept

of the numbers at areas of one fifth as well as of one fourth of the slide.

The results are summarized in tables V and VI.

Table V shows the percentage of deviation from the mean by marginal

(twentieth to fourth of slide) and median (fourth of slide) counts in

the first ten counts, calculated from the mean for this ten, by fifth and

fourth of slide counts in the second ten calculated from the mean for

that ten and by fourth of slide counts in the twenty counts calculated

from the mean for the whole twenty. Without attempting extended

analysis of the tables, I may call attention to the fact that the deviations

shown by ten counts do not indicate very much difference in most cases

between the marginal count (which varied from 1/20 to 4 of the slide)

and the fourth of slide count, nor between the fifth of slide and fourth of

slide counts, but that there is a much greater range of deviation in the mar-

ginal counts. I also note the fact that there is a better approximation

to the mean in the fourth of slide counts in the case of Gonyaulax polyedra,

which is a dinoflagellate of sub-globular form. Such a difference in count

of this organism might be expected because its shape would favor fairly

even distribution in mixing and handling while most other organisms are

sufficiently irregular in form to lead one to expect them to be more erratic

in any distribution undertaken by shaking or stirring of the surrounding

fluid. In the twenty count series it may be noted that the difference

between Gonyaulax and total dinoflagellates tends to disappear but that

the difference between both and diatoms is accentuated. The close

resemblance of Gonyaulax to total dinoflagellates is attributable largely

to the fact that Gonyaulax contributed about two thirds of the total.

eee

= ACN -

RANGE OF ERROR IN MICRO-ENUMERATION

The increased difference in range of deviation between Gonyaulax and the

total diatoms is explicable on the basis of what has just been said as to

differences in distribution due to form.

» | = ||

See oie etal.

TABLE V—Catch 8104—Percentages of deviation from the mean

Diatom colonies Diatom cells Dinoflagellate Total cells Gonyaulax polyedra’

cells

Median 14 Median 44 Median 14 Median 14 Median 14

No. of counts No. of counts No. of counts No. of counts No. of counts

a 3S Ss a 3

a= a= a a= AS] <-

ae He AE Be Be

Se} 10 | 20-)SS)| a7 | 20 Ise 10 | 20 ssh ios) 20 || S38) 40° | 220

6 9 | 19 1 8 | 16 6 6 | 10 Os 16 || 29 i 2

12 12 22 17 10 18 21 3 1 12 8 16 21 3 1

30 13 1 15 5 4 15 10 6 49 6 3 11 1 5

7 0 dl 9 1 10 12 7 11 10 2 il 23 14 18

6 1 11 12 8 1 34 12 15 16 6 3 27 3 7

4 3 8 0 11 1 23 3 7 3 10 0 5 16 12

7 12 21 0 16 24 0 8 3 0 14 21 13 7 ll

24 14 23 37 29 36 0 10 13 33 27 33 5 12 16

7 5 16 ll 15 23 4 ul 2 10 12 20 5 5 0

31 38 23 35 56 42 15 9 5 28 50 37 19 5 0

eens een eek ||P ra per eminem [ee larlpee ( aac he oa

~— = ee =~ —

5 ol 5 onl - be) mm

33 |" 52 Ee £3 23

po | oro pole] Oo Oo

Ss | 10 ss | 10 S3| 10 ss | 10 sg | 10

0 1 10 14 15 26 4 5 10 13 14 24 13 12 17

2 1 10 4 1 8 6 0 3 4 1 6 18 7 3

13 12 3 6 7 0 10 2 6 4 6 1 10 2 6

7 5 17 9 1 10 5 0 3 u 1 9 1 2 2

8 4 15 3 2 7 5 6 P43 3 iD, 6 4 4 0

ll 10 21 12 5 14 12 13 9 10 3 12 1 df 3

8 13 4 1 6 3 5 3. 1 2 5 2 16 13 9

5 3 8 0 4 4 “4 0 4 0 4 4 8 2 6

13 6 18 19 8 17 4 10 14 18 8 17 1 7 12

5 5 17 4 8 0 LOS enG) 9 2 6 2 13 10 13

W. E. ALLEN

Table VI covers some of the same ground as table V but in a different

way. In this table enumeration totals are shown instead of percentages,

with the addition of a list of numbers of Gonyaulax polyedra in each of

the twenty counts and a list of numbers of both cells and colonies of

Nitzschia seriata in each of the twenty counts. It also includes a statistical

summary which the late Mr. E. L. Michael very kindly prepared for me.

The series is too short for statistical treatment but the summary has some

interest in a suggestive way.

This summary indicates that the extreme deviation is not only more

than twice as great in the-case of diatom cells as it is in the case of Gonyau-

lax and total dinoflagellates but that the same thing is true of both cells

and colonies of Nitzschia seriata, the most abundant diatom in the catch.

Nitzschia seriata is a slender spindle-shaped diatom occurring very largely

in colonies of two to six individuals. Its form would lead me to expect it

to be quite erratic in distribution by any possible method of mixing. This

is also to be expected of the other numerous diatoms, which belong mainly

to the Chaetoceras group. It is also interesting to note that the wide

range of deviation in diatoms is due to the tenth and eleventh. counts

and that in count ten the numbers of both dinoflagellates and Gonyaulax

are very close to the mean, though Gonyaulax approaches the extreme

deviation in the eleventh count.

This last point is important because of its indication that the error lies

in the mixing and distributing of the organisms rather than in the method

of counting. The normal count of the less erratic Gonyaulax indicates

that there was no serious mistake in counting, computing or recording,

while the known erratic distribution of the diatoms does indicate con-

siderable variability in results of mixing. In spite of the large extreme

deviation due to diatoms, the mean variability for total cells is only 12.2%,

a fact which gives ground for thinking that totals of most counts are

within a range of error of less than ten per cent.

A point which can be verified by the reader in table VI, but not in the

others (though true-of all), is that the deviations are fairly evenly dis-

tributed on both sides of the mean. This is an indication in this type of

study that the fluctuations are normal and that they appear approximately

according to expectation.

Although this study as a whole is distinctly brief and fragmentary

it seems to give a good practical basis for the following provisional con-

clusions: First, that by very great care the extreme deviation (in total

numbers of diatoms and dinoflagellates) could probably be kept within

twenty-five per cent; second, that the mean deviation can be easily kept

within ten per cent; third, that diatoms are more variable in the counts

than dinoflagellates; fourth, that the causes of variability are to be found

in the processes of mixing, sampling and spreading on the slide, rather

€

¥ toes

RANGE OF ERROR IN MICRO-ENUMERATION

TaBLe VI

Catch $104

Enumeration totals, deviations, etc.

Total Total Total Total Gonyaulax | Nitzschia

Count Diatom Diatom __|Dinoflagel- Cells polyedra serjata

Colonies Cells late cells

Col. 280

Ist 696 Phils? 304 2456 220 Cells 660

188

2nd 676 2100 332 2432 212 372

292

3rd 872 2464 356 2820 204 560

232

4th 708 2300 300 2600 176 436

252

Sth 764, 2532 284 2816 200 604

220

6th 792 2604 312 2916 240 552

160

7th 680 1948 348 2296 192 372

240

8th 664 1652 292 1944 180 536

204

9th 728 1984 344 2328 216 424

308

10th 1060 3644 352 3996 216 648

304

11th 948 3232 368 3600 252 796

264

12th 952 2770 348 3118 208 532

BY)

13th 840 2596 356 2952 228 508

E p

224

14th 1008 2828 348 3176 220 384

24 W. E. ALLEN

TABLE VI (Continued)

Catch 8104

Enumeration totals, deviations, etc.

Total Total Total Total Gonyaulax | Nitzschia

Count Diatom Diatom __|Dinoflagel- Cells polyedra | seriata

Colonies Cells | late cells

336

15th 992 2760 328 3088 216 608

316

16th 1048 2940 | 304 3244 208 632

252

17th 832 2640 . 340 2980 196 564

284

18th 932 2676 348 | 3024 228 556

304

19th 1016 3016 384 3400 240 738

| 304

20th 1008 2592 368 2960 244 616

Extreme : 103= 39.1%

deviation |246= 30.2%|1059=41% |52=15.5%|1089= 37 .4%|39= 18 .2%|242= 43.7%

263

Average 814 2585 336 > 2907 215 554

Standard 44

deviation 142 463 27 410 20 112

Average ; 37= 14%

deviation |124= 15.2%] 358=13.9%|23=6.9% | 354=12.2%|16=7.5% | 87=15.7%

than in the counting; and fifth, that the range of error in counting is at

worst far less for microplankton material than is the range of error in

locating, catching and preserving material.

It seems fair to regard these results as suggestive for microscopic

material in general, e. g., enumeration of blood corpuscles might be ex-

pected to show a range of error somewhat similar to that of Gonyaulax

and direct enumeration of bacilli to give results, resembling those from

diatoms.

As regards my own use of the study, I may say that it has led me to

decide on the mixing procedure already mentioned, and in counting to

are

RANGE OF ERROR IN MICRO-ENUMERATION

carry all enumerations to fifty individuals (or fifty colonies) or to a very

close approach to fifty at a convenient computing point, except that all

enumerations are stopped when one eighth of the slide has been covered.

I may say frankly that for a single count or for a very short series

of counts, this number limit and area limit are too small. But in handling

large numbers of catches in large series and working through long periods

of time, one must give close attention to the law of diminishing returns.

Would the counting of a larger number of abundant forms or the counting

of all over a larger area give enough greater approach to accuracy to

compensate for the greater effort and use of time? It has not seemed

to me that it would for present purposes. With the lens combination on a

monocular microscope which was used in making this study, it was con-

venient to work over the area of one fourth of the slide. Later when using

a different lens combination on a binocular microscope, it was found that

- an area of one eighth of the slide was more convenient. In fact some counts

are so fatiguing and so time consuming at one fourth slide as to be im-

practicable in a long series. With my present standardized procedure I

should expect the one eighth slide counts to show about the same range

of error as indicated for the one fifth slide counts in table V. I have not

yet had time to verify this assumption. At worst the range of error in

careful work will certainly not be as great as that due to other factors as

far as microplankton is concerned.

Finally, I may say that although the results which I have obtained

are inadequate for definite conclusions, they do indicate that with standard-

ized procedure the microscope phase of plankton study is much more

nearly accurate than some of the other phases.

DEPARTMENT OF METHODS, REVIEWS, ABSTRACTS,

AND BRIEFER ARTICLES

REMARKS ON THE LIFE-HISTORY AND THE SCALE CHARAC-

TERS OF AMERICAN MULLETS

Cart L. HuBss

Museum of Zoology, University of Michigan

Mr. Arthur Paul Jacot, in the July issue of these TRANSACTIONS for

1920 (pp. 199-229), has presented the results of his investigations on

scales of two American mullets: Mugil cephalus and M. curema. He has

discovered a number of facts bearing significantly upon problems in several

of the zoological sub-sciences. These facts, and, more particularly the

author’s interpretations of them, are discussed in this brief note.

Mr. Jacot’s discovery that the scales of these species of Mugil are

ctenoid proves an unexpected confirmation of the view recently held by

Jordan and Hubbs! that the group Percesoces, comprising the Mugilidae

and related families, is derived from the typical Acanthopterygii (which

‘is characterized in part by ctenoid scales), and hence is not transitional

between the cycloid-scaled malacopterygian fishes and the more specialized

spiny-rayed types. The wide differences found in the character of the

ctenii on the scales of the species of Mugil studied are also of considerable

taxonomic interest.

The detailed account given of the development of scale structure, and

the final proof of the transformation of the first soft-ray of the anal fin of

the juvenile or Querimana stage into the third anal spine of the adult

Mugil, are valuable contributions from the standpoint of the comparative

anatomy of these structures. This juvenile metamorphosis of Mugil has

long been in need of the detailed study which Mr. Jacot has accorded it.

The sharply defined mark on the scales of Mugil cephalus, which the

author appears to have interpreted as a metamorphic annulus, is obviously

the first winter annulus; apparently intensified, it is true, by the fact the

adult characters, appear first in the spring, synchronously with the resump-

tion of the growth of the scale and of the fish, following the cessation of

growth significantly demonstrated to occur during the winter. The portion

of the scale within this first winter annulus therefore corresponds with the

1 A monographic review of the family of Atherinidae or Silversides (Stanford Univ. Publ.),

1919.

DEPARTMENT OF METHODS 7

brief period of initial growth? between hatching and winter. The similarity

existing between the first annulus developed on the scales of Mugil curema

to that of M. cephalus indicates that this species likewise breeds in the

fall, rather than during the summer as Jacot supposed.

This altered conception of the nature of the first annulus involves a

different interpretation of the age at maturity of Mugil cephalus: the

first spawning fish appear to be just two years old (rather than in their

second year); similarly, the oldest individual examined was six years old.

The second and succeeding line-like annuli developed on the scales of

these species of Mugil being typical of the winter marks developed on the

scales of marine fishes of temperate waters, and of the coregonine fishes of

the Great Lakes, it is, to say the least, unnecessary to follow Jacot in

interpreting these marks as migratory rather than as winter annuli. The

fact that an annulus was not evident near the margin of scales of mullets

taken at Beaufort in early spring indicates merely that the spring growth

of these fishes had not yet commenced, and not that these fishes were

exceptional non-migratory individuals. Indeed, it is not at all certain

that the mullets actually do migrate southward during the winter, for a

growing body of evidence is indicating that in this season many shore-

fishes of the Temperate Zone merely retreat into deeper water and

become less active, and hence appear absent because not caught.

These altered interpretations bring Mr. Jacot’s facts into much better

agreement with the results of studies made on the life-history of other

fishes, and in the opinion of the writer, enhance the value of his contribu-

tions.

Mr. Jacot has introduced some new terms, none of which will probably

be adopted. Of these ‘“‘adulting (changing to the adult condition)” and

“circulation” (referring to the course of the circuli on the scales), require

no further comment. The term ‘“‘linea (from the Latin /inea, ae, f.; using

the term in its more figurative expression),’’ is unnecessarily substituted

for annulus or “winter band’’; a similar statement might be applied to

‘““ctenobasil.”’

2 It is probable that the juvenile mullets pass during this period through a pelagic stage,

for which the Querimana characters are well adapted. Labidesthes sicculus, a fresh-water

fish related to Mugil, passes through such an initial pelagic stage.

28 HAROLD CUMMINS _

SPRING MIGRATION IN THE CRAYFISH, CAMBARUS ARGIL- |

LICOLA BAXON

Harotp CUMMINS

Tulane University

Incidental to a study of the migration of frogs into their breeding ponds,

carried out near Ann Arbor, Michigan, in the spring of 1914, some interest-

ing observations were made upon migratory activities of this burrowing

crayfish. So little is published regarding the habits of crayfishes that even

these few notes seem to be worthy of publication.

The location and character of the pond and the method of obtaining

migration data by trapping are described in another paper.' Briefly it

may be said-that the pond is at the edge of a cultivated field, bounded

partially by a wood which adjoins the field. About this pond a trap was

constructed, extending approximately two-thirds of its circumierence. The

trap consisted of a cloth fence, provided with leaders of similar construction

extending radially outward from the main fence. At the junction of each

leader and the main fence a pail half-filled with water was sunk in the earth

with its top at ground level. Crayfishes migrating toward the pond came

in contact with this cloth barrier, and as they edged along it in an attempt

to enter the pond were entrapped in the pails. Since the pond was not

completely enclosed by the trap the number of crayfishes taken does

not necessarily represent the total number of migrants; some may have

gained entrance where the fence was incomplete.

Whité’s Wood, at the edge of which the pond is located, fulfills the

habitat requirements of this species, and so far as the writer’s collections

indicate, C. argillicola is the only crayfish that occurs there. In addition

to the observation pond there are four small ponds in the wood, two of

which like the observation pond are usually not dried in the summer.

The remaining two always dry during the summer. All of them are

frequented by crayfishes. The burrows, usually with chimneys, form a

characteristic feature of the habitat; sometimes they are found at some

distance from the ponds, but usually near them, and when the ponds dry

numbers of chimneys are thrown up on the exposed mud.

After their winter torpidity was dissipated by warm weather, crayfishes

reappeared in the pond, not only from the bottom of the pond itself but

also from outside sources. The first individuals to appear were those

which had spent the winter in burrows in the pond bottom. On March 23

the burrows were first opened to the surface, numbers of them being

1 Cummins, Harold, The rdle of voice and coloration in spring migration and sex recogni-

tion in frogs. Jour. Exp. Zool., v. 30, no. 3, April 1920.

Sever

DEPARTMENT OF METHODS 29

observed for the first time on that date. Each opening was circular,

averaging 1.5 cm. in diameter. Bordering the opening of each burrow

was an approximately circular area, averaging 9 cm. in diameter, of light-

colored sand, apparently brought up from a lower level in the process of

opening the burrow. The sand was not thrown into a high convex mass,

but rather was so small in quantity as to be not appreciably elevated from

the level of the pond bottom. Unfortunately no attempt was made to

collect crayfishes from the pond before March 23, therefore it is impossible

to state whether or not all individuals spent the winter in burrows. Data

on the reappearance of crayfishes which were outside of the pond during

the winter, presumably in their burrows, were obtained from:the trap.

The results of the trap are presented in the accompanying chart.

Sh a ies IMIG Eno Seno [salsa aa

mature, ©. as tree | ig iy CI

Se el A cI

}

finmatwe [1 Co

immature | | S| | |!

Sex?, immature

March ~ April Noy

Chart showing the trap catch from March 26 to April 30, inclusive, and temperature

and humidity records from March 20 to May 7, inclusive. Temperatures in the Fahrenheit

scale are indicated at the left, and the average temperature for the 24 hours ending 7 A. M. of

each day is plotted in a continuous line on the graph. Degrees of relative humidity are

indicated at the right, and the average for the 24 hours ending 7 A. m. of each day is plotted

in a broken line on the graph. In several instances the number of trapped crayfishes was

recorded in the field notes as ‘“‘several’”’; such records are here shown by the asterisk (*).

It is evident from the chart that migration occurs at irregular intervals.

There is a broken migration wave from March 26 to April 2, inclusive, and

an unbroken wave from April 26 to 30, inclusive, while in the period from

April 3 through April 25 but one crayfish was trapped. The migration

waves occurred during periods of relatively high temperature and humidity.

The lowest temperature with which a migration is coincident is 33.6 degrees

(April 21), but with only one individual. A temperature more favorable

to migration, if judgment can be based on the trap catch, is 42 degrees

or over. All the catches are coincident with high humidities, the lowest

being 87 (March 29), the others ranging between 90 and 100.

With the exception of the female carrying young, captured on April 21,

all the collections were made in the morning, representing crayfishes

30 HAROLD CUMMINS

migrating during the preceding night. The single exception is noteworthy

in demonstrating that spring migratory activity may occur in the daytime.

This migrant was not actually trapped, but was noted at 10 A. m. walking

in the grass near the edge of the pond. In view of the remaining migrations

having been nocturnal, probably this record represents a crayfish which

migrated during the preceding night, and, coming in contact with the

fence, walked away instead of alongside as did others.

The small number of immature individuals is suggestive. One trapped

April 30 was only an inch in length, and therefore unquestionably not

sexually mature. The remaining six were about two-thirds the size of

average adults. Whether there is some stimulus controlling the migration .

of adults, which is usually lacking in the young, or whether the young

will migrate later are questions which cannot be answered with the data

at hand. The facts of an early beginning of migration among adults and

the retardation of migration of all but one of the immature crayfishes

leads to the inference that migratory impulses occur both in young and

adults, but begin to function earlier in the season in the latter.

A total of ten plus “‘several’’ females with eggs were captured on March

29 and April 2, and none appeared thereafter. During the first migration

wave several females without eggs (two plus two lots of “‘several’’ each)

were trapped, and in the second wave five without eggs appeared. Three

females carrying young appeared in the second wave. The bearing of

these data upon the time and place of egg-laying and hatching is important,

but difficult of interpretation. If those females bearing eggs and young

furnish a standard of comparison, we must assume either that there is a

prolonged period for the egg-laying and hatching of the last five females

or that their eggs already had hatched. The same question does not arise

in connection with the females without eggs which were captured early.

It seems that a migratory movement of adult females in the spring would

prove advantageous to the young, for they would hatch in water which

presumably provides a more favorable environment for them than the

burrows.

DEPARTMENT OF METHODS 31

PREPARING-COLLECTIONS OF THE MOLLUSCA

FOR EXHIBITION AND STUDY!

~ FRANK COLLINS BAKER

Curator, Museum of Natural History, University of Illinois

The Mollusca form a large group of the Animal Kingdom and members

of this phylum are used for economic or biologic study by many biologists,

zoologists, geologists, ecologists, and others interested in the study of

animal life. Collections are also made for their beauty or interest by

amateur students. Whatever the cause of interest it is important that

the collections made should be properly prepared and preserved for

future consultation. The good appearance and permanence of a collection

of mollusks depend very largely upon the care taken in cleaning and

preparing the individual specimens. The modus operandi varies with

the size and the kind of mollusk.

CLEANING THE SPECIMENS

Mussels or River Clams. The river mussels, when only the shells are

to be preserved, should be placed in boiling water which will cause the

valves to open slightly. The adductor muscles may be cut with a thin-

bladed knife and the animal matter removed. Care should be taken to

remove all of the animal matter from the region of the muscles where it

is strongly fastened. During this process the collector must avoid break-

ing or injuring the edge of the shell where the substance is very thin, the

new shelly matter as well as the epidermis or periostracum being newly

formed at this part of the shell. This is especially true of the thin-shelled

mussels like Anodonta. After removing the animal parts the shells should

be washed carefully to remove the mucus and any parts of the animal

remaining. Care must be exercised to avoid breaking the igament which

holds the two valves of the shell together. When thoroughly cleaned the

two valves may be tied together with white string (never use colored string

for it will mark the shells) and the shells laid on boards or other objects

to dry in a warm place. Never allow the sun to shine on specimens of

this kind for they will then dry too quickly and the epidermis will peel off.

A few shells of each lot should be broken apart so that the interior, espe-

cially the hinge structure, may be studied.

Many shells will be marred by incrustations of lime or other matter.

This may be removed with muriatic or oxalic acid, which may be applied

with a small camel’s hair brush. As these acids, especially muriatic acid,

readily attack that part of the shell not protected by the horny epidermis,

‘Contribution from the Museum of Natural History, University of Illinois, No. £73

32 FRANK COLLINS BAKER

the specimens should be washed carefully and quickly after using the acid.

Many shells may need to be scrubbed with a small scrubbing brush or a nail

brush to remove the extraneous matter. In some cases, however, it may

be desirable to preserve the shells in their natural state, with all the

incrustations and other foreign matter attached, to indicate the character

of the water or bottom in which the animals lived. This may be necessary

in some ecological studies. After the shells ‘are thoroughly dry the strings

may be removed and the surface of the shells rubbed with vaseline. This

will usually prevent the epidermis from peeling or cracking and will give

the’ shell the appearance it had when living in the water. Great care

should be used to see that all of the surplus vaseline is removed or the

surface will become sticky and unsightly. A soft rag may be used to

rub the shells perfectly dry and clean.

Finger-nail Clams-Sphaeriidae. The smaller -bivalves—Sphaerium,

Musculium, Pisidium—are usually too small for the animal to be removed

from the shell and they may be killed in 70 per cent alcohol from which

they may be removed and dried in a few days. In the case of the larger

Sphaerium the animal may be removed, after having been killed by boiling

or by preservation in alcohol for a few days. As the valves of the shell.

are liable to open after being cleaned, and as they are usually too small

to be tied together, they may be wrapped tightly in a plain piece of tissue

paper until dry, when the paper may be removed. No oil or other pre-

servative should be used for these shells.

Fresh Water Univalves or Snails. The larger fresh water snails may be

killed by boiling or by preservation for a few days in 70 or 80 per cent

alcohol. The animals are then easily extracted with a dissecting needle.

A needle with a curved or twisted point is more effective in removing the

animal from the inner whorls than one with a straight point. In the

large Lymnaea, Planorbis, and Physa, the animals are easily removable,

but in the Pleurocera, Campeloma, and related genera, the animals must

be removed with great care as the upper part of the animal, containing the

liver and part of the sexual organs, is liable to break off and remain in the

shell. When removing these animals, get a firm hold of the body with the

dissecting needle and then by a slow, careful, twisting motion remove the

animal. All animal matter should be removed from the large shells.

If there are incrustations or other foreign material on the surface of

the shells this may be taken off with a brush, scraped off with a knife, or

removed with the acids mentioned for mussel shells, oxalic acid being the

best. The acids must be used with care that the fine texture of the shells

may not be injured. In those species having an operculum, like Campeloma

and Pleurocera, the opercula of a few individuals of each lot should be

removed from the foot of the snail, dried, and placed inside the aperture

of the shell, which may then be closed with a piece of fine cotton. It is

DEPARTMENT OF METHODS 33

not a good idea to glue the operculum to the cotton because the inner side

which bears the muscle scars for its attachment to the operculigerous lobe

of the animal may be needed for study. All shells should be thoroughly

dried before placing them in the cabinet and before placing the operculum

in the aperture. It is well in the larger Campeloma and Vivipara, to

wipe the surface gently with the rag used for vaselining the mussel shells,

using the same care as recommended for that group in this particular.

The smaller snails, Amnicola, Valvata, small Lymnaeas (Galba), Ancylus,

etc., may be killed in 70 per cent alcohol, from which they may be removed

in a few days and dried. The little fresh water limpets (Ancylus) should

have the animal carefully removed with the point of the dissecting needle.

As these small limpets are usually coated with foreign matter they may be

effectively cleaned by being allowed to float, upside down, on the surface

of a small quantity of oxalic acid, after which they may be washed and

carefully wiped with a camel’s nair brush. The shell is thus easily cleaned

if held, aperture downward, on the tip of the index finger.

Land Shells. The larger land snails or Helices should be placed in

warm water which should be quickly brought to the boiling point to kill

the animals. It is of importance to be certain that the water is boiling

for hot. water will not kill the animal at once and it will then be difficult

to remove from the shell. Land shells cannot be left too long in the

boiling water because the fore part of the body is liable to break away

from the part containing the liver, which will then remain in the upper

whorls of the shell and be very difficult to remove. If not killed quickly

by boiling, the columella muscle will not be loosened from the pillar lip

and the animal cannot be pulled out without breaking in pieces. The

larger species must be boiled for fully a minute but the smaller species,

the size of Polygyra hirsuta, will be ready to have the animal removed in

10 or 15 seconds. To prevent loss in a large tin or pot it is well to place

the snails to be boiled in a wire dipper which may be obtained in any

10 cent store.

To insure successful extraction of the animals it is necessary to use

great care and plenty of time. The same curved dissecting needle men-

tioned previously is well suited for removing the animals of land snails,

and the same twisting motion is necessary as described under fresh water

snails. If the animal breaks during the operation, leaving a portion in

the upper whorls of the shell, the remaining part may be removed with

jets of water from a small syringe, preferably a fine-pointed dental syringe.

It may be well sometimes to place the shell in alcohol for a day or two

in order that the part of the animal leit in the shell may be loosened,

after which the syringe will usually remove the matter. Sometimes a

vigorous shaking, or, with the hand holding the shell, striking the other

hand or the thigh, will aid in loosening the refractory matter. Much

34 FRANK COLLINS BAKER

patience and some ingenuity is necessary in removing the animals from

their shells in which the aperture is restricted or contracted by teeth or

folds, and in these cases the fine syringe will be found useful to start the

body from the shell. All shells should be washed out inside with the

syringe and scrubbed on the outside with a tooth brush, or other small

brush, to remove all traces of mucus, dirt, or other foreign matter. A

gentle flow of water from a tap or faucet is very effectual in removing

mucus and dirt from the interior of large shells. If the mucus is unusually

adhesive, as is sometimes the case, it may be necessary to use a small

piece of sponge or cotton attached to the curved dissecting needle, or

held with a pair of curved forceps, to remove the unsightly material. Land

shells do not require vaseline for the preservation of the epidermis as

suggested for fresh water mussels and large water snails. When perfectly

clean the shells may be laid on boards or other objects and laid in a con-

venient place to dry. Never allow shells to dry in the sun for they will

crack and be spoiled for cabinet purposes. Too strong emphasis cannot

be laid on the injunction to remove al/ animal matter from the larger land

shells, which have a peculiarly offensive odor all their own if placed in a

cabinet only partly cleaned.

The small land snails, especially the members of the Pupillidae and

those snails having teeth or folds in the aperture, cannot well have the

animals removed. If these are kept for a few days in a dry place the

animal will retract well within the shell and they may then be placed in

30 or 40 per cent alcohol for twenty-four hours, after which they may be

dried and no offensive odor will be retained. Vermin will not usually

attack a shell that is thus well soaked in alcohol. When dirt of any kind

remains attached to these small shells they may be effectually cleaned

by being put in a bottle with fine, clean sand, and a vigorous shaking will

remove the dirt. This process should not be used for fragile shells. It is

especially effective with the Pupillidae.

Marine Shells. The directions given above for land and fresh water

shells apply equally well for marine mollusks. The snails from the sea,

however, are more difficult to prepare because of the more powerful

columellar muscle by which the animal is attached to the shell. For the

larger species of sea snails the curved dissecting needle will hardly be

adequate to extract the animal. For this purpose nothing is better than

a stout fish hook which has been heated and then bent in the form of a

partial spiral. Plunging in cold water after shaping will return the temper

of the steel sufficiently for the purpose for which it is made. The shank

may be firmly fastened in a wooden handle made in convenient shape to

fit the hand, and the result is a very useful implement. In extracting the

larger animals from their shells, it is important that the hook be deeply

and firmly buried in the large, tough muscle attached to the columella

DEPARTMENT OF METHODS 35

pillar or axis of the shell. A strong, steady pull will usually bring the

animal.

Bivalve shells, clams, may be treated in a similar manner to Unionidae

mentioned on a previous page. Boring clams, like Pholas, Teredo, and

others, will require special attention to preserve the extra pieces of shelly

matter connected with shell. Small clams may be treated in the same

manner as mentioned under finger-nail shells. The same may be said

of the small snails which should be treated as the small fresh water or

land snails. Marine shells may be killed in boiling water or by preserva-

tion in alcohol. As in the case of land and fresh water mollusks, formalin

is not a good preservative on account of its action on the shells.

Many marine snails are encrusted with limy matter, the tubes of

worms, the hard shelly bases of corallines, and the dried remains of sponges.

These may be removed with an old file the end of which has been ground

toa point. Little chisels and punches like engraver’s tools are also excellent

for this purpose. With care and experience the collector will be able to

scale off the greater part of this extraneous matter without harming the

shell beneath. The judicious use of muriatic acid will also help in the

final cleaning process, but this reagent must be used with great discretion

in order not to mar the surface of the shell.

PREPARATION FOR ANATOMICAL STUDY

It is frequently desirable that some of the material collected should

be preserved for the study of the animal. Fresh water pulmonates, such

as Lymnaea, Planorbis, Physa, may be placed directly in 30 per cent alcohol,

where they may remain for twenty-four hours. They should then be

placed in 50 per cent alcohol for another twenty-four hours, and finally

preserved in 75 or 80 per cent alcohol. The fresh water operculate snails

may be preserved in the same manner, as may also most of the marine

snail shells.

Land shells, however, must be killed in osmic acid or by drowning, the

latter being the best, causing the animal to die in a fully expanded con-

dition. For drowning, the writer has obtained the best results by placing

the snails in a large, wide-mouthed bottle, filling the bottle level full with

water and placing a heavy piece of glass over the water to exclude all air

bubbles. In twelve to twenty-four hours the animals will be fully expanded

and quite dead and may then be removed to 30, 50, and 80 per cent alcohol

as recommended above for fresh water snails. Care must be exercised

that the snails are not taken from the drowning water too soon, for. in

this case they will contract badly when placed in alcohol.

Final preservation may be made ina 2 per cent solution of formalde-

hyde, but alcohol is better for the flexibility of the animal, which has a

36 FRANK COLLINS BAKER

tendency to harden and become brittle in formaldehyde. Even in a weak

solution of formaldehyde the shells gradually soften and easily break

when handled. A recent examination of some molluscan material in a

research collection of a well-known laboratory was found to be time wasted

because the material had been preserved in formaldehyde, and the shells

had softened and curled up, almost entirely losing their original character.

Valuable material upon which scientific conclusions are based is thus liable

to be ruined for future study and examination.

Slugs (Limax, etc.) and snails with small or very thin shells may

be preserved as mentioned for the animals of land shells. The eggs ofall

mollusks, fresh water as well as land, should be preserved in alcohol,

after passing through the different grades of the preservative. Some eggs,

as those of Pyramidula and Polygyra, have a more or less hard shell and

may be dried and preserved in bottles. In the case of large eggs of the

Bulimi and other large land shells, they must be treated in the same

manner as birds’ eggs and the contents removed by means of an egg blow

pipe. They may then be dried and placed in the collection.

For bringing out details of the surface structure of snails a 1 per cent

solution of chromic acid has been found to be a good reagent. Miiller’s

fluid is also an excellent fixing reagent. These reagents, however, harden

the body to such an extent that it is often difficult to make gross anatomical

examinations and the alcohol method described above is the best for all

purposes. When using the fixing reagents mentioned it is highly impor-

tant that the animals be washed thoroughly in running water before being

transferred to the different grades of alcohol. Twelve to forty-eight-hours

will be necessary for this purpose, depending upon the size of the specimen

treated. No specimens should be placed in strong alcohol at once as this

reagent extracts the water so rapidly that the internal organs are shrunken

and distorted. For sectioning and some histological purposes the harden-

ing methods mentioned are excellent.

PRESERVATION FOR STUDY OR EXHIBITION

The method of preserving and arranging a collection of mollusks

will depend wholly upon the purpose for which it was made. All collections

may be roughly divided into two types, those for display and those for

study. Each of these types requires a different treatment.

Collections for Display. Collections of this kind will probably be

confined almost exclusively to museums of one kind or another. An

exhibition collection of the Mollusca, even in a public museum, should

be more or less synoptic in character, and arranged to show the principal

features of classification, as well as facts relative to different kinds of

habitats—ponds, rivers, swamps, shallow water, deep water, rocky shores,

DEPARTMENT OF METHODS Si

sandy shores, forests, plains, and valleys—in short, the ecology of this

type of animals. The geographic distribution—Arctic, temperate, tropic,

island, continental, etc.——should be indicated by charts; the variation of

individuals and the economic use made of certain species should also be

clearly indicated. For some of this display, models may be used to illus-

trate ecology, geographic variation, and methods of life. Features of this

kind add much to the value of a collection and are always interesting to

those persons visiting a museum that are not particularly interested in the

generai subject of mollusks. Such economic displays as pearl buttons

and the clams from which they are made, both fresh water and marine,

shell money as used by the native tribes of this and other countries,

mollusks used for food, injurious snails, pearls, and other topics of like

nature, are very interesting, useful, and highly educational.

For exhibiting mollusks a strong, durable, attractive tablet is essential.

Such an one can be made of heavy binder’s board (no. 20) cut into con-

venient sizes and covered with such material as will give the best effect

to the collection. Many shells will look well on a black background and

these may be mounted on tablets that have been covered with a dull black

paper. Dark shells look better on a light background, and for these the

writer has used an ivory-colored cardboard known as Royal Worcester

Bristol Board, a material that withstands the fading power of light better

than any other paper used. For these light backgrounds the cardboard is

cut just a trifle smaller than the tablet, the edges of which have previously

been passépartouted with a dead black paper used for binding together

lantern slides, and the light cardboard is glued to the tablet (glue being

used only about the margin of the card), leaving a border of black. This

method produces a handsome tablet that is both durable and attractive.

When the label is attached (which should be made of the same cardboard’

used for the center of the tablet) the whole has a pleasing appearance...

The sizes of these tablets, as used by the writer in his museum work,

and found to be the most useful, may be 3x 2, 3x3, 3x4, 3x 6s S09",

6x6, 9x9, and 12x12 inches. All of these are multiples of the small

unit, 3 x 2 inches.

To make an exhibit collection of the greatest value from a teaching

standpoint, many drawings of structure and development, maps of dis-

tribution, and labels describing the function of organs, as well as notes

of interest concerning the animals or shells, should be freely used. A

famous museum man, Dr. G. Brown Goode, once said that a museum was

a collection of labels illustrated by specimens, and while this axiom is

pretty strong and the matter may be somewhat overdone, the fact never-

theless remains true that a collection for public exhibition must be largely

explained or interpreted by means of illustrations, models, and descriptive

Jabels. Perhaps the statement of the great British museum administrator,

38 FRANK COLLINS BAKER

Sir William H. Flower, more nearly describes the use and function of a

museum, who says: “It is not the objects placed in a museum that consti-

tute its value, so much as the method in which they are displayed and

the use made of them for the purpose of education.”

Printed labels are the best for permanent display, but as these are

expensive the next best are typewritten labels which may be printed on a

typewriter having a platen such as is used by the librarians for card ~

catalog work. The ribbon should be black carbon. Where large shells

or series of shells illustrating some feature of structure of variation are

to be exhibited a uniform black or ivory-colored background may be

employed, using a large sheet of dull black paper or a sheet of the bristol

board mentioned above for tablets.

" Cases. Nearly all molluscan shells are best displayed in horizontal

or flat cases. Shelving in an upright case can be used, but this method

of installation is not as attractive nor as easy to install as in a flat case.

Very large specimens or material preserved in alcohol or other fluid (these

should be flat-sided glass jars) are best shown in upright cases or in the

A-cases that are now used in many museums. In some museums the

space beneath the flat cases is utilized for the purpose of storing the study

series in drawers. In some of the older museums these drawers have (or

had) glass tops and the contents could be seen by the visitor by simply

pulling out the drawer. Excepting where the matter of space is vital

this should not be done. The open museum halls are poor places for the

proper storage of a research series which must be consulted in the presence

of curious visitors who greatly bother the student. These collections

should be stored in drawer cabinets kept in rooms especially reserved for

research collections and made convenient for their study. This subject

is more fully treated on a later page.

For holding cards and labels in an upright position the writer has

found the pins and ticket holders sold by stationers to serve the purpose

admirably. For attaching specimens to tablets it is better to use wax

than glue, the former being easy to remove if the shell is needed for exami-

nation, while glue is difficult to remove without injury to the shell. Bright,

polished shells, like Cypreaea and Oliva, are difficult to attach to the -

tablets on account of their smoothness. The prepared clay known as

‘plastene,’ ‘modelit,’ and ‘permodello’ has been used to some extent by

the writer and has been found excellent for this purpose, if the mixture

does not have too large a percentage of oil, which discolors ‘the tablet.

By drying the clay a trifle the amount of oil may be reduced. This clay

usually provides a mold in which the shell may be held in any desired

position. The clay is made in several colors among which gray-green,

terra cotta, or dark brown are the best. If the shells are of the common

kind and are not likely to be needed for study the liquid glues will pee

DEPARTMENT OF METHODS 39

the best medium for fastening the specimens to the tablets. The shells

may be propped in any desired position until the glue hardens.

COLLECTIONS FOR STUDY

While there are several ways in which a collection of mollusks can be

installed for exhibition, there is but one good method of caring for a

study or research series of these animals. The dry specimens should

be kept in drawer-cabinets. In considering the size of the cabinet the

dimensions of the primary unit, the individual tray containing the speci-

mens must first be decided upon.

Pasteboard Trays. These should be made as multiples of the smallest

unit. This unit may be 1x2 inches for the smallest species and 3 x 2

inches for the larger series, the unit of width here being three inches.

If one desires to carry out the 1x 2 unit for the entire series the larger

trays may be multiples of two inches. The various sizes that are the

most useful, as learned from experience, are as follows: a. 1 x 2, 2x2,

ieee A x 44 x 616 x6, 6x9), 9x9. 12 x12) bo 1 x 3,223,393,

3x4,3x6,3x9, 6x6,9x9,12x12. The depth of the trays should be

one-half inch, except in the largest size which should be three-fourths of

an inch in depth. The trays may be covered with black or white glazed

paper which gives them a pleasing appearance. These trays can be made

by any box manufacturer. Ingenious students may be able to make their

own trays if they have the time. To do this pieces of cardboard should be

cut as shown in figure 1 and the four pieces indicated by the dotted line,

folded together and attached by adhesive paper. Trays of this kind can

be quite economically made in a short time.

3 inches 2.

Fig. 1. Method of making a pasteboard tray for holding specimens.

Drawers. Having selected the size of the individual tray the next

step is the dimension of the individual drawer. This should be made of a

40 FRANK COLLINS BAKER

size to contain the trays in a manner that there may be no waste space.

A convenient size measurement, imside, is 1514 x 2114 -inches. This

allows five rows of the three inch unit trays which fit snugly in both

dimensions of the drawer. If the two inch is used the drawer should be

an inch wider or 1614 inches. This will hold eight rows of the two-inch

unit. The depth of the drawer will depend upon the character of the

specimens it will contain. The smallest specimens need a drawer not

over an inch in depth while the largest may require a depth of five or six

inches. For an all around depth the writer uses a drawer two mches in

depth and when larger specimens are installed the space of two drawers

is used for one. This method has been found quite satisfactory and does

not on the average take more room than when drawers are made of varying

depth. It is seldom that specimens of greatly different size will be placed

in the same cabinet, if room is wisely left for expansion, as should be done

in the larger collections. The drawers mentioned above, which are in use

in the University of Illinois Museum, are made of three-eighths inch

material for the sides and ends and compo board is used for the bottom.

These drawers require no handles and are very inexpensive.

Fig. 2. Storage cabinet of drawers, 32 in each cabinet. An open drawer at the bottom

shows the method of attaching labels to the back of the trays, as described in the text. Univer-

sity of Illinois Museum.

DEPARTMENT OF METHODS 4]

Cabinets. Having determined the sizes of the trays and the drawers

the next thing is the size and style of the cabinet to contain the drawers.

This should also be of the unit pattern so that several may fit together.

The cabinets that are in use in the University of Illinois Museum, and

which the writer has used in other museums he has had charge of, are shown

in figure 2. These have the following dimensions:

Height 46, width 3514, depth 25 inches, outside measurements.

Drawer space 161% inches wide, 2% inches between the drawer runners.

Drawer runners 3 inch pieces sunk in the sides of case 3/16 inch.

Each case holds 32 drawers, 8 in each of four sections.

If the drawers are to be one inch deep the space between the runners

should be 1-5/16 inches and the case would hold sixteen drawers in each

section or sixty-four in each cabinet. The drawers are really *¢ of an inch

deeper than the dimensions given, this extra space being occupied by the

runners. This space allows for extra large shells which may be included

with the smaller ones. It is usually essential that all cabinets be of the

same size and contain the same size of drawer so that additions and

rearrangements may be made without unduly changing the contents of

the drawers. This is very important when large additions are made neces-

sitating the rearrangement of a large part of the collection. The drawers

may be made of whitewood or basswood and simply shellaced or varnished.

The cabinets are best if made of oak and finished in some dark color.

The door of the cabinet should be made with a groove which extends

entirely around the inner margin. This should fit into a tongue in the

sides of the cabinet which also extends entirely around the cabinet. A

piece of plush or felt fitted into the groove in the door will keep out the

dust very effectually. Rubber has been used but this substance soon

loses its resiliency and becomes worthless. The door should be made so

that it may be entirely removed from the cabinet so that it will not be in

the way when the collection is being studied. The photograph, fig. 2,

indicates these points.

For smaller species, as the Pupillidae, Valloniidae, Sphaeriidae, Amni-

colidae, as well as many groups of minute marine shells, the writer has

used a case made to hold legal blanks which has proved very convenient

and satisfactory. The dimensions of the drawers are:

. Length 1414, width 9, depth 1 inch; height of case of ten drawers 14

inches.

Each drawer holds 56 of the 1 x 2 inch unit trays or 560 trays in the

cabinet. These cases are admirably adapted for holding these small

shells, the drawers not being large enough to be cumbersome as is the

case with a large drawer filled with these small trays. Several legal blank

cases may be installed in one of the larger cabinets if it is desired to keep

the cabinets perfectly uniform. The only possible criticism may be that

42 FRANK COLLINS BAKER

these legal blank cases are not perfectly dust proof and for permanent.

installation they should, perhaps, be enclosed in a cabinet as suggested

above. Each drawer should be labelled with the name of the contents

and each cabinet should have the name of the group it contains.

Bottles and Vials. For the safety of the collections glass bottles or

vials should be provided for all shells under 34 inch diameter. These

should be made in different diameters but only of two lengths to fit the

two unit widths of trays, two and three inches wide. Convenient sizes

are as follows:

134 x 3%, 134 x 4, 134 x &%, 134 x1 inch.

216 x %, 1, x Vé, 244 .x %, 2% x11 inch.

Occasionally a larger size will be needed and a vial of 114 inch diameter

will be found useful. Only a few of this size will usually be required.

The bottles known as shell vials, obtainable through almost any druggist,

are especially adapted for the preservation of molluscan material and can

be made of any of the sizes mentioned above. Short corks may be used

but these should be rolled or squeezed to soften them so that the fragile

tubes may not be broken when the cork is forced into the mouth of the

vial. Rolling with a hard piece of wood or metal or pressing between a

pair of large flat-nosed pliers will be found to accomplish this purpose

admirably. Homeopathic vials may be used but these are not as good for

dry specimens as the shell vials. Where expense is a serious item very

good containers may be made by rolling a piece of stiff paper over a lead

pencil or other round object the size of the required container and then

gluing or pasting the edges together and closing one end with cotton.

A cotton cork may also be used for the other end. The modus operandi

of this method is indicated in figure 3. The cylinders may be made in

lengths of legal blanks and then cut off in lengths to fit the trays.

glue glue

Fig. 3. Method of making paper shell tubes for holding small specimens.

STORAGE OF ALCOHOLIC MATERIAL

The proper storage of material preserved in alcohol is a matter requiring

considerable attention. For this purpose homeopathic vials are better

DEPARTMENT OF METHODS 43

than shell vials, because they are thicker and the strong, reinforced opening

makes it possible to press a cork in very tightly, which retards evaporation

of the liquid contents. Fairly large vials should be used even for small

specimens in order that the storage may be uniform and liquid enough

provided for the specimens to obviate frequent filling of the containers.

Several sizes of bottles, 2, 4, 6 ounce, will be required to preserve the

larger specimens. For mussels or series of specimens the clamp-top, all

glass fruit jars (Atlas for example) are excellent for storage purposes.

The old-fashioned Mason jar is not good because the liquid evaporates

quickly and the metal screw top cannot be made permanently tight,

besides becoming very unsightly in a short time. All of the jars should be

of glass, except the rubbers, and these will need to be changed at intervals.

The quart and pint jars have been found the most useful for purposes of

storage, although the two quart size may be necessary at times.

Professor Frank Smith, of the Department of Zoology, University

of Illinois, has made use of a method, first devised by the United States

National Museum, for the preservation and storage of small specimens in

vials which has much to commend it. The small vials, after being filled

with alcohol or other preservative and having a wad of cotton placed in

the mouth of the vial, are stored, bottom upward, in a large jar, of two

liter capacity or larger, which is then filled with alcohol or other fluid.

By this means the smaller vials may be kept without adding new liquid

for a long time. Also, the large jar will become empty before the small

vials and thus a warning is given before any damage can be done to the

specimen. It often happens when valuable material is stored in many

small vials that lack of proper attention permits the vials to become empty

of fluid and the specimens dry out and are thus ruined.

Many zoologists will prefer the single bottle method, however, on

account of the accessibility of the material, and for such the storage

should be in standard racks, which may be stored on compo board shelves

in the unit cabinets described previously. These racks will vary in width

but should be of the same length. Convenient dimensions are as follows:

Vials, length 22, width 1%, height side 214, height front 3 inches.

Bottles, length 22, width 24, height side 214, height front 3 inches.

The general form used is indicated in figure 4. The stock should be

LEZ LEE FFE z te A

ards GL vam FLEE IZE Ss, Z : a

= Baa

Fig. 4. Standard rack for holding alcoholic material, vial size.

44 FRANK COLLINS BAKER

3g inch for ends and 3/16 inch for bottom and sides. The large jars are

perhaps best stored on shelves, although a rack similar to the one sug-

gested for the vials and bottles, but made large enough to hold the jars,

may be used. These racks may be made by any good carpenter or the

lumber can be cut in a mill and the collector can put the racks together

himself.

REGISTRATION AND LABELING

Every set of specimens should have with it a label giving the name

of the species, its locality, the principal ecological conditions under which

it was found, the name of the collector, and the name of the authority who

determined the species, as well as the date of collection. For this purpose

cardboard labels just the width of the inside of the unit tray, 1 x 2 or1 x3

inches, may be used. The writer has found by experience that an excellent

method of attaching the label to the tray is to glue the upper edge of the

label] to the upper margin of the unit width of the tray, at the back. When

a whole drawer is arranged with the labels affixed in this manner the

different species and their localities may easily be read. The specimens

or vial of specimens lie in front of this label, as shown in figure 2. Genera

or group divisions may be indicated on labels fastened to the bottom of

the 1 x 2 or 1 x3 inch trays. A sample of label in use in the Museum of

the University of Illinois is shown below.

Museum OF NaTuraL History

ACTINONAIAS. MIGAMENTINA LLom]

SOAT Fork below. dam, Homer Park,

Gravel bolfom, water iS in.deep.

Mussel Survey BigVermilion River LA.

NoZI173 Det. by F.c.Baker Date 1X-30-1918

UNIVERSITY OF JLLINOIS

Fig. 5. Sample of label.

A catalog number should be given each set of specimens and this.

number should be placed in the vial containing the specimens or in the

case of large specimens, written on the shell. For mussels both valves

should be numbered. The best quality of indelible carbon ink should be

used for this purpose. The writer has found Higgin’s eternal ink (water-

proof) to be the best for all purposes. The alcoholic material should

have a label placed in each bottle written with the same kind of ink. Card-

board labels for this purpose are good. A permanent cloth known as

‘mapstock’ velum, sold by Jos. Bancroft & Sons Co., Rockford near

Wilmington, Delaware, has been found admirable for this purpose.

A serial catalog kept in a book and a card catalog are invaluable for

the proper recording and convenient classification of a collection. The

book should be arranged to contain the seria! numbers of the collection.

DEPARTMENT OF METHODS 4

This volume may be made as elaborate as the pocket book of the collector

will permit, varying from a simple note book to a large printed folio.

For museums and large collections of private individuals the large folio

is by all means the best. This may be arranged with the headings sug-

gested below.

No. of Received | Collected

specimens from | by Date Remarks

Other entries, such as original no., identified by, dry, alcoholic, etc.,

may be used if it is desired to elaborate further. For large institutions

an accession catalog is necessary, in which is recorded the material by

lots as received. In such cases an entry, accession number, is usually

made room for after catalog number in the species catalog.

The card catalog should contain the references to all of the lots of

one species, showing the different places from which they came, on one

card, or each lot of a species may have all of the information recorded on

one card. The first is more convenient for a small collection but the

latter is perhaps better for a large institution or collection, giving all of

the known data concerning each species lot on one card. The cards should

be arranged alphabetically under the genera, the names of which should

appear on guide cards, as is done for library card catalogs. Experience will

suggest many ways in which the cataloging may be so arranged as to make

the collection most useful, which is its legitimate function.

In closing let me say that a collection of mollusks is valuable principally

for the information which it may contain. It is of paramount importance,

therefore, that the data or information concerning each lot of specimens

be made as accurate and complete as possible. This should be done in the

field if possible and not left until later when memory may play one tricks

as to the exact habitat of some specimens in a large lot. There are many

questions still unsettled regarding the classification, geographic distribu-

tion, ecological habitat, and economic importance of this class of animals

and any conscientious collector may add real scientific knowledge con-

cerning some common species by exercising care and intelligence in making

collections.

Cat.

No. Name Locality

SOME PAPERS RELATING TO THE COLLECTING AND PREPARATION OF

MOo.t.usca FoR BotH EXHIBITION AND STUDY

BAKER, FRANK C.

1898. Mollusca of the Chicago Area. The Pelecypoda. Section VI, Eel aE BO2E for

Collecting Mollusks, pp. 25-32.

1900. A new Museum Tablet. Amer. Nat., XXXIV, pp. 283-284.

46 FRANK COLLINS BAKER

1902. The Descriptive Arrangement of Museum Collections. The Museums Journal

(English), II, pp. 106-110.

1904. The Arrangement of the Collection of Mollusca in the Chicago Academy of

Sciences. Museums Journal (English), IT, pp. 354-360.

1909. Suggestions for an Educational Exhibit of Mollusks. Proc. Amer. Assoc. Mu-

seums, III, pp. 56-59.

1910. Same title, Museums Journal, IX, pp. 394-397.

Dati, WIL11AM H. |

1892. Instructions for Collecting Mollusks and other Useful Hints for the Conchologist.

Bull. U. S. Nat. Museum, No. 39, Part G, pp. 1-56.

STERKI, VICTOR.

1916. Some Directions and Suggestions for Collecting the Sphaeriidae and Aquatic

Gastropods. Annals Carnegie Museum, X, pp. 478-480.

WALKER, BRYANT.

1902. Hints on Collecting Land and Fresh-water Mollusca. Journ. of Applied Micros-

copy and Laboratory Methods, V, No. 9, pp. 1954-1961.

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TABLE OF CONTENTS

For VOLUME XL, NuMBER 2, April, 1921

Larval Flukes from Georgia, with two plates, by E. C. Faust....................-- 49

On the Nature of Structures Characteristic of Cnidosporidian Spores, by R. Kudo.... 59

DEPARTMENT OF SUMMARIES

Recent Advances in=Parasitology, byab. GC. Faust... 2-9-0624. eee 75

DEPARTMENT OF MetuHops, REVIEWS, ABSTRACTS, AND BRIEFER ARTICLES

A Method for Orienting and Mounting Microscopical Objects in Glycerine, by

Charles:Bullard it sos ..c3 8 bc eee oe eee Cate os See eee 89

A Method of Demonstrating the Sheath Structure of a Desmid, with one

figure;by 'W.. R: Taylor: 22. ..c5e sat sets hot Be eon oe ee eee 94

™."

TRANSACTIONS

American Microscopical Society

(Published in Quarterly Instalments)

| Vol. XL APRIL, 1921 : No. 2

LARVAL FLUKES FROM GEORGIA!

(With two Plates)

ERNEST CARROLL FAUST

Parasitologist, Union Medical College, Peking

In a previous study (Faust 1919) I have reviewed the vercariae de-

scribed from the United States. I discussed the regions from which forms

had been reported and suggested that the Southeastern United States

offered an unexplored field where conditions were eminently favorable for

their existence.

From October 1918 to May 1919 I had the opportunity to examine

several hundred mollusks from the region of Rome, Georgia, and in two

of these species, Goniobasis carinifera Lamarck and Anculosa carinata

Brug, I have discovered the larval flukes described in this paper. I wish

to acknowledge my indebtedness to Mrs. Lola Swift Faust for the collection

of the material and to Mr. Bryant Walker for the determination of the

hosts.

The larvae represented in this investigation are all distomes and fall

into the following groups:

AMI Corelaseve Tire 4 Eee 1S lee a Pe a ed a ec 1

MUNOCORCAMIAC ae Wee eins Th. ae ay ou 3

MMEROCEMCOMS CeTeamiAn Grime is. |. kas. > ke ces 1

BchingslLombenla niacin ree a ee ek one 1

LUMECOCCHUCRIa eR ee ape ete G Wr ce iS Sal), 2

EYSUOCCEEOUSIEGhGariad. Sarena eee ics gh a ea, 2 1

All of these forms are new to this region. Two have been described in a

previous paper (Faust 1919a).

Cercaria thalia nov. spec.

(Figs, fT, 2)

Host: Goniobasis carinifera Lamarck.

This interesting larva for which I propose the name Cercaria thalia

was found in Goniobasis carinifera Lamarck, collected from Rotary Lake,

‘Contributions from the Department of Pathology, Union Medical College, Peking,

China.

50 ERNEST CARROLL FAUST

Rome, Georgia. The redia and cercaria stages were dissected out of the

digestive gland of the snail, while the agamodistomes were found in the

lung sac. Both the redia and the cercaria were characterized by a graceful

movement.

In the cercaria a good share of the organs are rendered opalescent by

the sub-integumentary cells which are muciferous rather than cystogenous

in structure. The larva is distinguished by a pair of eye-spots just lateral

to. the pharynx. The body measures 0.4 mm. in length by 0.15 mm. in

width. The tail is slightly longer and has a proximal diameter of 57yn.

The oral sucker measures 71 in section while the acetabulum when

extended has a diameter of 904. No spines have been observed on the body

or tail. There is a prepharynx and a small pharynx with a short esophagus

showing constrictions at intervals along its course. The ceca terminate

about one-third the body distance from the posterior end. There are three

pairs of mucin glands, with ducts opening on the dorsal aspect of the

orifice.

The excretory bladder has a long median shank and slender cornua

so that the entire appearance is that of a furculum with a supporting stem.

I have observed a triplet group of flame cells at the anterior extremity

of the body and a similar number at the posterior limit, but have been

unable to make out the number of groups intermediate. From the group

formula which I have previously shown to exist for this family of trema-

todes (Faust 1919a:334) four other flame cell units are to be expected. A

median collecting tubule runs almost the entire distance of the tail.

The agamodistome (Fig. 2) lacks the pigmented eyespots and, indeed,

shows few of the larval characters. The most readily recognizable common

feature is the excretory bladder. On the other hand this fluke shows

evidences of rapid maturity. The shape is that of a mature worm rather

than that of a larva. The ceca have enlarged and what is especially note-

worthy, the genital organs have reached a high degree of complexity. The

testes and ovary are in their relative positions, the vitelline follic’es are

well formed, the seminal vesicle and the seminal ieceptacle are each con-

spicuous and Laurer’s canal is distinguishable So well developed are all of

these organs that it is a simple matter to place the worm in the Allocreadii-

nae. Thus a considerable share of the life history of this animal is at hand,

even tho only the first larval host is known.

The cercaria is produced in a redia with a small orange-colored gut, a

short prepharyngeal region, a small pharynx, and a small but conspicuous

birth pore. Moreover, both daughter rediae and cercariae develop within

the same parent redia. It is noteworthy that the cercaria possesses eye-

spots, a common feature of the larval allocreadid form, but lacks a stylet.

LARVAL FLUKES FROM GEORGIA oe

p |

Cercaria camilla nov. spec.

(Fig. 3)

Host: Goniobasis carinifera Lamarck.

The stylet cercaria for which I propose the name Cercaria camilla is a

rapidly moving larva with an oblong-ovate body 0.16 mm. long by

0.066 mm. wide and a tail 0.11 mm. long and 18y in diameter at the base.

Both body and tail are spinose. The oral sucker is 26y in diameter while

the acetabulum measures only 13y in transection. The latter lies midway

between the anterior and posterior limits of the body.

Inserted into the roof of the anterior end is a typical quilled stylet.

The oral sucker leads into an enormous prepharyngeal pocket 53y in

diameter, with a thick, semi-muscular, semi-mucoid wall. This in turn

leads directly into a minute muscular pharynx. Beyond the pharynx a

short esophagus connects with a part of short blunt ceca. The mucin glands

consist of three pairs. Two of these pairs are granular, acidophilic, while

one pair is densely muciferous, basophilic in reaction. The contents of

each gland passes thru a long duct to empty at the side of the stylet.

The excretory bladder has the shape of an inverted truncated pyramid,

from which emerge delicate collecting tubules. These tubules when traced

to their sources reveal on each side of the body four pairs of flame cells

posteriorly disposed. A single collecting tubule runs down the middle

part of the tail. The flame-cell formula has an identical common denomina-

tor with that of Allocreadium isoporum Looss (Faust 1919a:327, 334),

namely

(aap aaa (2-2)

The parthenita of C. camilla is a very simple sac-shaped sporocyst

containing eight to twelve cercariae. Encystment of the cercariae has not

been observed.

Cercaria tabitha nov. spec.

(Fig. 4)

Host: Goniobasis carinifera Lamarck.

This cercaria for which I suggest the name Cercaria tabitha has an

ovoid body 0.15 mm. long by 0.088 mm. wide and a blunt tail 0.1 mm.

long by 0.017 mm. in diameter at the base. The body is covered with

heavy spines and the oral sucker is provided with a blunt stylet. The large

oral sucker, 32 in diameter, is provided internally with a thick mucoid

substance resembling that of the stylet. Behind this is a small pharynx.

There are four mucin glands on each side of the body, one of which contains

a basophilic substance. They all empty far laterad at the margin of the

oral sucker. The remainder of the digestive tract has not been traced.

The acetabulum is in the middle of the ventral side of the body. It meas-

ures 21y in diameter.

52 ERNEST CARROLL FAUST

The excretory bladder is cup-shaped. The collecting tubules emerge

from the anterolateral aspect of the bladder while the pore is posteriad.

The sporocyst is a simple, sacculate structure, containing up to twenty-

four larvae.

The tail of the cercaria is dropped readily under a cover glass, but

encystment has not been observed.

Cercaria pandora nov. spec.

(Fig. 8)

Host: Goniobasis carinifera Lamarck.

This little larva for which I suggest the name Cercaria pandora is

oblong-ovate with a body measurement of 0.145 mm. in length by 0.054

mm. in width and a tail not more than half as long by 174 in diameter at

the base. The anterior sucker is large, measuring 38 in diameter. The

acetabulum is mesad and very small (14). A small stylet with a sharp,

delicate point is inserted into the dorsal wall of the oral sucker. A small

pharynx is located just behind the orifice. It leads into a forked gut densely

surrounded with gland cells.. The mucin glands consist of four pairs, the

posterior one of which has a large nucleus and gives an acidophilic reaction.

The excretory bladder is a roughly truncated cone with conspicuous

lateral cornua. The caudal excretory-canal has several tributaries but no

observable flame cells.

The larva develops in large numbers in simple sacculate sporocysts.

Cercaria medea nov. spec.

(Fig. 7)

Host: Goniobasis carinifera Lamarck.

This larval fluke for which I suggest the name Cercaria medea has a

long, slender body and a short, stubby tail. The latter is so limited in

extent as to place the larva in the group of the microcercous cercariae.

The animal measures 0.22 mm. in Jength by 0.065 mm. in width and

has a tail only 21h long. The latter structure in semiglandular, the prod-

ucts of which are poured into a common atrium. The cells are chromo-

phobic and have irregular shaped nuclei. The anteriormost part of the

body together with the acetabulum bear small sharp spines. The oral

sucker has a diameter of 254 and the acetabulum of 27u. Inserted in the

dorsal wall of the oral sucker is a minute, simple-pointed stylet (See fig.

7a).

Behind the oral sucker is a small pharynx. A long, narrow esophagus

runs back from this to the anterior aspect of the acetabulum, whence the

ceca continue posteriad to the subcaudal region of the body. A group of

twelve to fifteen mucin glands is situated on each side of the body posterior

LARVAL FLUKES FROM GEORGIA 53

to the acetabulum. A very delicate bundle of ducts conveys the products

of these glands to the region of the stylet.

The excretory bladder is long and bag-shaped, giving rise to a pair of

cornua laterad just behind the acetabulum. The main collecting tubule on

each side of the body bifurcates just anteriad to the acetabulum. A

median tubule extends into the tail and opens into the mucin pocket.

Two strings of germ cells extend longitudinally across the acetabulum.

The sporocyst in which the cercariae develop varies greatly in size.

It has a muscular anterior end and at irregular intervals has constrictions.

The movement of both the cercaria and the sporocyst is slow.

Cercaria penthesilia nov. spec.

(Fig. 9)

Host: Goniobasis carinifera Lamarck.

This cercaria for which I propose the name Cercaria penthesilia is‘an

echinostome larva which is probably immature, in which the circlet of

collar spines has not yet developed. It measures 0.2 mm. in length by

0.084 mm. in width, and has a tail 0.135 mm. long by 14y in diameter at

the base. The body is covered with many short spines closely studded

together. The oral sucker has a diameter of 30u and the acetabulum of

32u. The entire body has a thick subintegumentary lining of long rhab-

ditiform cystogenous granules, which bear evidence of the animal’s future

encystment. A fluted keel is found in the distal third of the tail.

There is a short prepharyngeal region of the digestive tube followed by

a small pharynx. The esophagus forks almost immediately to form ceca

which extend far caudad. Paired groups of acidophilic mucin glands run

. mesad to the ceca.

The excretory bladder is spheroidal with a tage opening dorsocaudad.

The cornu on each side is quite inconspicuous. From it a dilated collecting

tube is traced which becomes narrow in the region of the pharynx. Several

flame cells have been found but the exact number has not been worked out.

Running posteriad from the bladder is a collecting tube for the tail. Half-

way down the tail it divides to form a pair of tubules opening laterad.

The genital system is represented by two groups of germ cells lying

longitudinally across the acetabulum. The nervous system is conspicuous,

with especially large ventral trunks.

The redia in which the cercaria develops is provided with the pharynx,

gut, birthpore and lateral appendages found in the echinostome group.

Cercaria quattuor-solenata Faust 1919

and

Cercaria furcicauda Faust 1919

(Figs. 5, 10)

These furcicaudous species, Cercaria quattuor-solenata, and C. furct-

cauda, were originally described in connection with a study of the excretory

54 ERNEST CARROLL FAUST

system of distome cercariae (Faust 1919a: 337, 338) and are included here

for the sake of completeness. The host of both species is Anculosa carinata

Brug.

Cercaria stephanocauda nov. spec.

(Fiz. 6)

This interesting larva which I have designated as Cercaria stephano-

cauda represents a group which has recently received considerable atten-

tion (Ward 1916, Pratt 1919, Faust 1918). Five species have been de-

scribed from North America. While in many respects the structure of

the body of the immature larva resembles that of the described species,

the tail is unique.

The worm has a body 2 mm. long and 1.2 mm. wide. The shank of the

tail is 4mm. long, while the lamellate furcae measure 1.1 mm. long and

0.5 mm. wide. The anterior fourth of the tail shank consists of a collared

region with about nine definite ringed constrictions, running around the

tail. At the posterior end of this collar there are numerous tubercules in a

single row which are the only traces of mammilations anywhere on the

body. Behind this collar the tail proceeds toward the distal portion with

gradual constriction. The tail is attached to the body proximally by a

number of powerful longitudinal muscles.

The oral sucker measures 430y in diameter and the smaller ventral

sucker, 3604. The pharynx just behind the oral sucker has a diameter of

200u. It connects with the ceca by a very short esophagus. The ceca

proceed directly laterad almost to the posterior margin of the body. They

are slightly convoluted.

The excretory system consists of a minute bladder and large collecting .

tubules. The latter reach to the region of the oral sucker, then are reflexed

and break up into capillaries. A large collecting tubule extends the length

of the tail, forking into the furcae and opening outward at the distal end of

each furca.

The genital glands are very immature as contrasted with the condition

in Cercaria macrostoma (Faust 1918). The germ glands are connected by

a chain of cells which lie in the median line, with one gland offside to the

left.

The sporocyst is large, being a simple sac with undifferentiated tissue.

SUMMARY

A study of cercariae taken from snails at Rome, Georgia, shows new

species with interesting relations to previously described forms.

LARVAL FLUKES FROM GEORGIA 55

REFERENCES CITED

Faust, E. C.

1918. Two new cystocercous cercariae from North America. Jour. Parasit., 4:148-153,

1 pl.

1919. A biological survey of described cercariae in the United States. Am. Nat.,

53:85-92.

1919a. The excretory system in Digenea. II. Observations on the excretory system in

distome cercariae. Biol. Bull., 36:322-339, 10 figs.

PRATT eH. 9:

1919. A new cystocercous cercaria. Jour. Parasit., 5:128-131, 2 figs.

Warp, H. B.

1916. Notes on two free-swimming larval trematodes from North America. Jour.

Parasit., 3:10-20, 1 pl.

56 ERNEST CARROLL FAUST

DESCRIPTION OF PLATES

Figs. 1, 2.—Cercaria thalia 1, cercaria, ventral view, X 126; 2, agamodistome, ventral view,

showing precocious development of genital organs, X 40.

Fig. 3.—Cercaria camilla, ventral view, X 400.

Fig. 4.—Cercaria tabitha, ventral view, X 285.

Fig. 5.—Cercaria quattuor-solenata, ventral view, X 200.

Fig. 6.—Cercaria stephanocauda, ventral view, X 28.

Fig. 7.—Cercaria medea, ventral view, X 90; 7a, detail of stylet.

Fig. 8.—Cercaria pandora, ventral view, X 285.

Fig. 9.—Cercaria penthesilia, ventral view, X 276.

Fig. 10.—Cercaria furcicauda, ventral view, X 200.

SS SE A A He inl ie

PLATE II.

LARVAL FLUKES FROM GEORGIA

58 ERNEST CARROLL FAUST

PLATE III.

ON THE NATURE OF STRUCTURES CHARACTERISTIC

OF CNIDOSPORIDIAN SPORES!

R. Kupo

CONTENTS

TET OUUG HOMINS Mee he Sheth SSE ey, Ri PRR «ee ee ok Phe MERE aL Senge Spec Rey 59

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INTRODUCTION

Although our knowledge of the morphology and development of

Cnidosporidia has recently been greatly increased, little is known about

the chemical nature of the different parts which compose the spores. In

the study of artificial cultivation of Cnidosporidia, it becomes necessary

to obtain a definite view regarding this point. Unfortunately the majority

of opinions advanced by several authors are not accompanied by any

definite experimental data. This is possibly due to the fact that because

the organisms have never been found in direct contact with higher verte-

brates as their parasites, they have not interested such a large number

of investigators as did other parasitic Protozoa which are directly respon-

sible for serious diseases among mammals, and that the number of organ-

isms found at one time is not generally large so that only little is left when

the study of their morphology and development is completed.

Erdmann (1917:317-318) has recently expressed a view that the polar

filaments of the spore of Chloromyxum leydigi were probably composed of

glycogen and plastin, and further suggested, in referring to my paper on

the polar filament of the spore of Nosema bombycis (Kudo, 1913), that her

method might be useful for the study of the nature of polar filaments of

the Microsporidian.

I have been working for some time on the subject by using several

species of Microsporidia and Myxosporidia. The results thus far obtained

in regard to the polar filaments are entirely different from those of Erd-

mann’s. Besides, as I believe the results of observations upon the nature

of spore membrane and the so-called iodinophilous vacuole seem to be

' Contribution from the Zoological Laboratory of the University of Illinois, No. 182.

*I am greatly indebted to Professor Henry B. Ward who kindly placed the material at

my disposal.

60 R. KUDO

more or less interesting, they are summarized and presented in the follow-

ing pages.

MATERIAL AND METHODS

For the study of spore membrane of Myxosporidia, Henneguya salmini-

cola Ward? was chosen. This species, as was described by Ward (1920),

forms numerous large cysts in the host tissue. Moreover, it was not

only obtainable in a very large number, but was also favorable for the

study due to the presence of the posterior process characteristic of the

genus. The Myxosporidian had been preserved in formol since 1914

together with the host tissue. The cysts were isolated, washed thoroughly

with distilled water, and the cyst wall was punctured. The emulsion thus

obtained was used to make numerous smears. For Microsporidia, spores of

Nosema bombycis Nageli and Nosema apis Zander were selected. Although

they are much smaller than such a form as Thelohania magna Kudo (Kudo,

1920), the enormous number that could be procured favored their selection

as representatives of Microsporidia. The infected silk-worm moths and

the infected ventriculus and intestine of honey bees were emulsified with

distilled water, and were used as stock emulsions.

The nature of the polar filaments was studied by using the fresh spores

of Myxobolus mesentericus Kudo, Mitraspora elongata Kudo, Leptotheca

ohlmacheri (Gurley) Labbé, Nosema bombycis, Nosema apis and Thelohania

magna.

The following four species of the family Myxobolidae were used for the

study of iodinophilous vacuole. They were fixed either in formol or subli-

mate alcohol as are indicated below: Myxobolus discrepans Kudo and

Myxobolus mesentericus Kudo fixed in sublimate alcohol, and preserved in

95% alcohol; Myxobolus aureatus Ward and Henneguya salminicola Ward

fixed and preserved in formol. Besides, the following five species from

other genera were also studied for the comparison: Wardia ovinocua Kudo,

Mitraspora elongata Kudo, Leptotheca ohimacheri (Gurley) Labbé, Chloro-

myxum wardi Kudo and M yxidium americanum Kudo.

The experiments were conducted both in smears and section prepara-

tions. In former case, the smears were first allowed to dry before the

application of chemicals or stains especially for the determination of spore

membrane. Detailed descriptions of methods used will be given in the

corresponding chapters.

THE SPORE MEMBRANE

The strong resistance of spores of Nosema bombycis against certain

chemicals has long been known by the studies of Frei and Lebert (1856)

and Haberlandt and Verson (1870), as the parasite is the cause of the well

known pebrine disease of silk worms. These authors, however, attacked

STRUCTURES CHARACTERISTIC OF CNIDOSPORIDIAN SPORES 61

the problem with the aim of destroying the spores rather than the deter-

mination of its nature.

As to the spore membrane of Myxosporidia, Biitschli seems to be the

first who studied the effect of concentrated sulphuric acid upon the spores

of Myxobolus miilleri Biitschli. He (Biitschli, 1881: 634) states as follows:

“Die Schalensubstanz besitzt eine recht ansehnliche Widerstands-fahigkeit

gegen Reagentien; dennoch ist die Angabe Balbiani’s, dass sie auch in

erhitzten Mineralsduren sich erhalte, nicht begriindet. Erstmaliges Er-

hitzen in koncentrirter Schwefelsdure liess zwar die Schalen nur in ihre

beiden Klappen zerfallen, zerstérte dagegen die sogleich zu erwiihnenden

Polkapseln vollig; nochmaliges Erhitzen bewirkt jedoch auch véll ge

Zerst6rung der Schalen.” Balbiani (1883:202) on the other hand states

that the boiling sulphuric acid does not affect the Myxosporidian spore

membrane. Gurley (1894:83) agreed with Biitschli, writing as follows:

“This (shell) substance is thin, very transparent, insoluble in the strongest

acids and alkalies in the cold, certainly in some, and probably in most

species destroyed by (soluble in ?) concentrated sulphuric acid at its boil-

ing temperature.”

It is my opinion that the controversy of the results of experiments

among these authors are probably due to the difference in the concentration

of the acid used. Although none of the investigators mentioned their exact

technique, it is almost certain that they added concentrated sulphuric acid

to the emulsion of the spores they had. Consequently they did not observe

the effect of the truly concentrated sulphuric acid, but that of more or

less diluted acid which varied from a stronger concentration (Biitschli and

Gurley) to a weaker one (Balbiani) according to the relative amounts of

water and the acid. In order to avoid this error in the present experiments,

the smears of spore emulsions were dried on the slides before they were

subjected to the action of reagents.

None of the above mentioned three authors, however, has expressed

an opinion in regard to the chemical nature of the membrane. It was

Thélohan who advanced his observations concerning this point. Yet in

his valuable work on Myxosporidia, Thélohan (1895:260) simply states as

follows: “Je n’ai pas déterminé la nature chimique de la substance qui

constitue l’enveloppe. Elle ne présente en tout cas aucun des charactéres

de la cellulose.”

Although a large number of papers on Cnidosporidia has appeared

lately, none touches this problem. Davis (1917:210) states that ‘‘sur-

rounding the spore is a thin, tough, transparent membrane, the sporocyst,

which is probably of a chitinoid nature.’ He, however, does not give any

experimental datum to support this statement. Auerbach (1910:17) in his

monograph wrote negatively as follows: ‘‘Die chemische Zusammensetz-

ung der Schale ist meines Wissens noch nicht sicher bekannt.”’

62 : R. KUDO

Thus the opinions of a few investigators regarding the chemical nature

of the Cnidosporidian spore membrane may be summarized as follows:

The spore membrane of Myxosporidia does not give a positive cellulose

reaction, and seems to be of chitinoid nature.

The results of my experiments will be reported here.

a) Tests for albuminoid substances:—Spores of Henneguya salmini-

cola, Nosema bombycis and Nosema spis are not affected by boiling potas-

sium hydrate solution (35 per cent), and do not give any recognizable

positive Millon’s reaction. It may therefore be said that the spore mem-

brane is not composed of albuminoid substances.

b) Tests for cellulose:—For the control of the cellulose tests, a filter

paper was used.

1) Ammoniacal solution of copper oxide. Six fibers of the filter paper

were taken out, and were mounted on two slides, each containing three

fibers. To one distilled water was added, while to the other ammoniacal

solution of copper oxide. Both were covered with cover glasses. The water

emulsions of Henneguya salminicola, Nosema bombycis and Nosema apis

were smeared on slides, were dried, and were treated in the same way as the

fibers of filter paper. The preparations were kept in a moist chamber.

The results of observations are as follows:

Soon after the treat-| 16 hours later 36 hours later

ment

Cellulose with water Outline of fibers sharp} No change No change

Cellulose with ammon.| Outline of fibers less

solution of copper oxide! sharp Outline faint More invisible

Henneguya salm., Nosema| Outline of spores sharp} No change No change

bombycis and N. apis

with water

Henn. salminicola, No-| Outline of spores sharp| No change No change

sema apis, and N. bom- \

bycis with amm. sol. of

copper oxide

From the above, it is clear that the ammoniacal solution of copper

oxide does not dissolve the spore membrane of Henneguya salminicola,

Nosema bombycis and Nosema apis.

2) Lugol solution and sulphuric acid. Small pieces of filter paper were

treated with Lugol solution, washed with distilled water, and were dried.

They were mounted on slides in distilled water and in 50 per cent sul-

phuric acid respectively. Dried smears of spore emulsion of Henneguya

STRUCTURES CHARACTERISTIC OF CNIDOSPORIDIAN SPORES 63

salminicola, Nosema apis and Nosema bombycis were treated in a similar

way. The results of observations are as follows:

Soon after preparation

Cellulose: water and H2SO, No coloration

Cellulose: Lugol Deep brown

Cellulose: Lugol and H2SO; Violet

Henn. salm.: water and H.SO, No coloration

Henn. salm.: Lugol Slightly yellowish

Henn. salm.: Lugol and H2SO, Slightly yellowish

Nosema apis and bombycis: No coloration

water and H,SO,

Nosema apis and bombycis: Almost unstained

Lugol

Nosema apis and bombycis: Almost unstained

Lugol and H2SO,;

3) Zinc chloride-iodine-potassium iodide mixture. Fibers of a filter

paper and dried smears of spores of Henneguya salminicola, Nosema bomby-

cis, and Nosema apis were treated with the following mixture: Zincum

chloratum pur. sicc. 20 gr., potassium iodide 6.5 gr., iodine 1.3 gr., and

distilled water 10.5 cc. The results of observations are as follows:

Soon after preparation 16 hours later

Cellulose with water No staining No staining

Cellulose with mixture Violet blue Violet blue

The spores with water No staining No staining

The spores with mixture Slightly yellowish; iodinophi-| No change

lous vacuole of Henneguya

brownish

As will be seen from the above experiments, none of the cellulose tests

gives positive reaction. It may therefore be stated as was remarked by

Thélohan (1895) that the spore membrane of Henneguya salminicola,

Nosema bombycis and Nosema apis is not of cellulose nature.

c) Tests for chitin:—For the control of chitin test, I have prepared

chitin from the wings of honey bees.

R. KUDO

1) Alkalies and acids. Dried smears of spores of Henneguya salminicola,

Nosema bombycis, and Nosema apis, were treated with potassium hydrate

solution and mineral acids.

The results are as follows:

Chitin Spore membrane of Henn.| Spore membrane of No-

salminicola sema apis and Nosema

bombycis

Boiling KOH (35%) Insoluble Insoluble Insoluble

Boiling dilute HNO; Soluble | Spore becomesmoreorless| Spore becomes - greatly

swollen; contents at-| swollen and hardly visi-

tacked ble

Boiling conc. HNO* Soluble | Spore becomes larger;) Spore becomes extremely

spore membrane of uni-| enlarged; invisible.

form thickness; less re-

Conc. HCl (room temp.)

Soluble

fractive

Slightly soluble; outline

Spore enlarged and

irregular invisble

Boiling conc. HCl Soluble | Attacked Disintegrates rapidly

Dilute H2SO, Insoluble Insoluble Insoluble

Conc. H2SO, (room temp)| Soluble | Membrane becomes thin-| Greatly enlarged and in-

ner; outline irregular;} visible

valves split; caudal fila-

ment broken

Boiling conc. H2,SO, Soluble | Completely dissolved | Completely dissolved

2) Zinc chloride and Lugol solution. The spore emulsions of Henne-

guya salminicola, Nosema bombycis and Nosema apis were mixed with

potassium hydrate solution (35 per cent), and washed thoroughly by centri-

fugation. After being partly dried, 1 cc. of a mixture of 3314 per cent

aqueous solution of zinc chloride (10 cc.) and of strong Lugol so ution (5

drops) was added, and observed. The results follow

| |

| Color reaction

Dark brown

| | =

Henneguya salminicola _ Spore membrane very slightly yel-

lowish -

Nosema bombycis and Nosema apis| Spore membrane unstained

3) Potassium iodide and sulphuric acid. Chitin and dried smears of

Hin: eguya salmin.cola, Nosema bomby- s and No ema apis were bo led at

STRUCTURES CHARACTERISTIC OF CNIDOSPORIDIAN SPORES 65

160°C with potassium hydrate solution (35 per cent) for thirty minutes,

washed thoroughly with 90 per cent alcohol, and then with distilled water.

They were then treated with a weak solution of potassium iodide which

had been acidified with sulphuric acid, and- were examined. The results

are as follows:

Color reaction

Chitin Bluish violet

Henneguya salminicola No visible staining of the mem-

brane

Nosema bombycis and Nosema apis\ No visible staining of the mem-

brane

As will be seen from the above experiments, the staining reaction

gives very ambiguous results. On the other hand, the effect of mineral

acids upon the spore membrane seems to be decisive. The spore mem-

brane of Nosema bombycis and Nosema apis behaves very much like chitin

under the influence of mineral acids, while that of Henneguya salminicola

is more or less different in this respect.

THE POLAR FILAMENT

Thélohan (1890:207) expressed an opinion that the substance compos-

ing the wall of the polar capsule was identical with that composing the

spore membrane, as both stained in the same way with safranin. This

view probably led Minchin (1912:399) to state that ‘‘a polar capsule is a

hollow, pearshaped body with a tough envelope, probably chitinoid in

nature. . . . Coiled up within the capsule is a delicate filament, often

of great length, probably of the same nature as the capsule, and continuous

with it.” Minchin does not give any evidence to support this view. Davis

(1917:210) possibly referred to Minchin, although he did not make it

clear, when he stated as follows: “‘Surrounding the capsule is a tough,

refractive envelope, probably chitinous. . .. Coiled up within the

capsule is a delicate filament, usually of comparatively great length,

which is probably of the same material as the capsule.” ;

Erdmann (1917:317), by studying the developing polar filament of

Chloromyxum leydigi, came to the conclusion that the polar filament is

composed of glycogen and plastin. She writes as follows: “Die vier

Polkapseln fiillen jetzt ihre Mutterzelle aus, die Plastinscheiben werden

zu Plastinringen, die durch Glykogen verbunden sind. Der Polfaden ist

66 R. KUDO

entstanden. . . . Dagegen kann ich die Umwandlung des Chromatins

and des Plasmas in eine stark farbbare Substanz, nach meinen Befunden,

Glykogen, bestitigen.’”’ To support this view, Erdmann further remarked

the difhculty in extruding the polar filament of spores o! the species of the

genus Chloromyxum as follows ‘‘Ausgestreckte Polfaéden von Chloromyx-

umsporen sind kaum an Praparaten beobachtet. Thélohan (Taf. IX,

Fig. 100c) bildet die Polfaden einer frischen Spore von Chloromyxum

quadratum ab, Erdmann Chloromyxum leydigi mit kurzem Polfaden.

Auerbach ist es nicht gelungen, bei Chloromyxum dubium sie zu zeigen.

Lebzelter erwihnt sie nicht bei Chloromyxum thymalli. Durch eine von

mir ausgeprobte Methode der Fixierung (100 Proz. Alkohol bis auf 40

Grad erhitzt) gelingt es leicht, die Polfiden zum Austreten zu bringen und

zu fixieren. Glykogenfairbung nach Fixierung zeigt, dass der Polfaden

aus Glykogen und einer plastinahnlichen Substanz zusammengesetzt ist

(Taf. 14, Fig. 27).’”’ I have, however, had no difficulty in causing the extru-

sion of the polar filament from the fresh spores of three out of five species

of Chloromyxum which I have studied up to date. In Chloromyxum mis-

gurni Kudo (Kudo 1916, Fig. 3e), Chloromyxum fujitai Kudo (drawings

were omitted due to the lack of space) and Chloromyxum trijugum Kudo

(Kudo 1919: Fig. 181), I have caused the filament extrusion. The other

two species, Chloromyxum catostomi Kudo (Kudo 1919) and Chloromyxum

wardi Kudo (Kudo 1919) were studied only in fixed specimens, and no

attempt was made to cause the filament extrusion. Erdmann has probably

studied a small number of spores.

Regarding the various methods which had been reported by several

investigators as successful in extruding the polar filaments of various

Cnidosporidian spores, I already summarized them in one of my papers

(Kudo 1918). Of many methods which I have tried since that date, the

following gave always the best results. For Myxosporidian spores, potas-

sium hydrate solution or perhydrol will always bring out satisfactory

results. This is especially true in the case of tissue infecting forms. When

the spores are found in the gall bladder or urinary bladder, the best results

are obtained by centrifuging the spore containing bile or urine followed

by repeated washing with distilled water before the spores are subjected

to the influence of the chemicals, although this is not of absolute necessity.

Yet when the number of spores present in the bile or urine is very small,

the treatment is favorable as the addition of potassium hydrate solution

to the bile produces a large amount of precipitation which hinders the

observation greatly. In most cases, no staining of the filament of Myxo-

sporidian spores seems to be necessary, due to the favorable thickness and

distinctness even in unstained state. For staining, Fontana’s method of

staining spirochoetes, or Giemsa’s solution gives beautiful results. The

STRUCTURES CHARACTERISTIC OF CNIDOSPORIDIAN SPORES 67

latter has its advantage over the former in bringing out the differentiation

of nucleus and sporoplasm beside the filament, although in some cases the

filament does not take the stain for unknown reasons. For Microsporidia,

mechanical pressure or perhydrol gives beautiful preparations of extruded

filaments. As I did not describe the detail of the method used (Kudo 1913),

I have recently described the exact technique for the application of mechan-

ical pressure elsewhere (Kudo 1920).

Although Erdmann was apparently unaware of it, the filament extrusion

under the effect of absolute alcohol had been described by Ohlmacher

(1893) in the case of Leptotheca ohlmacheri (Gurley) Labbé. In the section

preparations of kidneys of Bufo lentiginosus fixed with absolute alcohol,

Ohlmacher saw a number of spores with extruded polar filaments. Ohl-

macher was of the opinion that “‘it is, of course, evident that they (polar

filaments) must have been thrown out from spores before the organisms

were killed by the alcohol employed in fixing.” According to my own

observations on a large number of section preparations obtained from

Rana clamitans, it is clear that in Ohlmacher’s preparation, the fixation

with absolute alcohol which caused vigorous shrinkage of the spore mem-

brane and sporoplasm, was only responsible for the presence of spores with

extruded polar filaments. It is certain that the absolute alcohol method

of Erdmann is according to my comparative study on various methods

far inferior in having incompleteness and irregularity in its action.

The results of my observations are as follows:

1) The effect of water upon the polar filament. To determine nee

the extruded polar filaments of spores of Nosema bombycis, Nosema apis,

Myxobolus mesentericus and Leptotheca ohlmacheri are soluble in water or

not, fresh spores were subjected to mechanical pressure. After removing

the coverglasses, the smears were covered with distilled water, and were

kept in a moist chamber. The examinations were done under a dark field

microscope, and also in Fontana preparations. The results were similar

in four species, which are as follows:

Results of Observations

Control: soon after the application} Extruded polar filaments

of mechanical pressure

One day in water Polar filaments unchanged

Two days in water | Polar filaments unchanged

Four days in water Polar filaments unchanged

Eight days in water Polar filaments unchanged

68 R. KUDO

The experiments were repeated many times on other species than

mentioned above, but always giving the same results. From these experi-

ments it may be concluded that the polar filaments of the spores of Cnido-

sporidia mentioned above are insoluble in distilled water at room tempera-

ture.

2) The effect of filtered saliva upon the polar filaments. To determine

whether the extruded polar filaments of spores of the species mentioned

above are soluble in filtered saliva or not, fresh spores were pressed mechan-

ically. A drop of filtered saliva was added to each smear, and the smears

were kept in a moist chamber. The examinations were done as in the

preceding experiments, and revealed the following results which were

practically the same in the four species:

Observations

Control: soon after the extrusion | Extruded polar filaments

10 minutes in saliva Polar filaments unchanged

30 minutes in saliva Polar filaments unchanged

1 hour in saliva Polar filaments unchanged

3 hours in saliva Polar filaments unchanged

16 hours in saliva Polar filaments unchanged

32 hours in saliva Polar filaments unchanged

3 days in saliva Polar filaments unchanged

6 days in saliva Polar filaments unchanged

From the experiments, it may be said that the extruded polar filaments

of the spores of Leptotheca ohlmacheri, Myxobolus mesentericus, Nosema

bombycis and Nosema apis, are insoluble in filtered saliva at room temper-

ature.

3) Staining with Lugol solution. The extruded polar filaments of spores

of Nosema bombycis, Nosema apis, Thelohania magna, Leptotheca ohlmachert

and Myxobolus mesentericus stain uniformly as light yellowish as their

spore membrane by Lugol solution, and do not take any deeper color.

4) Staining after Best’s method. The extruded polar filaments of

spores of Nosema bombycis, Nosema apis, Thelohania magna, Leptotheca

Te ee ee

STRUCTURES CHARACTERISTIC OF CNIDOSPORIDIAN SPORES 69

ohlmacheri and M yxobolus mesentericus remain unstained by Best’s method.

5) Staining after Lubarsch’s method. The extruded polar filaments

of spores of Nosema bombycis, Nosema apis, Thelohania magna, Leptotheca

ohlmacheri and Myxobolus mesentericus stain uniformly slightly bluish-

violet by Lubarsch’s method. The polar capsules of the latter two species

and the spore membranes frequently stain deep violet.

6) Staining by Léffler’s method. The extruded polar filaments of spores

of Nosema bombycis (Kudo, 1913), and of Nosema apis, Thelohania magna,

Leptotheca ohlmacher: and Chloromyxum trijugum are stained deep violet

by Léffler’s method. The spore membrane is also stained in the same color.

7) Staining with Giemsa’s stain. The extruded polar filament of No-

sema bombycis (Kudo, 1916), and of Nosema apis and Chloromyxum triju-

gum (Kudo 1920, Fig. 181) have been stained in deep, dark red. The polar

capsules of the latter species and the spore membrane also frequently

stain the same color.

8) Staining with Fontana’s mixture for staining spirochoetes. The

extruded polar filaments of Nosema bombycis, Nosema apis, Thelohania

magna, Thelohania illinoisensis, Leptotheca ohlmacheri, Chloromyxum tri-

jugum stain in from yellowish to dark brown color by Fontana’s method.

The spore membrane takes the stain in the similar manner.

From these experiments, it is clear that the polar filaments of spores of

various species of Cnidosporidia, which have been listed in the above, are

not composed of glycogen as was thought by Erdmann in the case of

Chloromyxum leydigi. The only means which led Erdmann to the already

quoted conclusion regarding the chemical nature of the polar filament is

the results of Lubarsch’s staining. My experiments have shown clearly

that while this staining brings out more or less bluish stained filaments,

other tests for its glycogenous nature proved to be negative. The staining

effect of Loffler’s method is similar to that on the flagella of Bacillus

typhosus, and that of Fontana’s method is exactly the same as that on

various spirochoetes.

As to its true nature, I am, however, still unable to determine. It has

been noted by many investigators in numerous species of Myxosporidia,

and by myself in Leptotheca ohlmacheri and Thelohania magna that the

nucleus for the polar capsule becomes nebulous or diffused during the

formation of the polar filament. The chromatic substance of the nucleus

breaks up into numerous small granules and a large part of it unites with

a peculiar substance or substances which become differentiated in the

capsulogenous cell, first in a retort shape and then in rounded form. The

polar filament is apparently formed from this mixture.

70 R. KUDO

THE IODINOPHILOUS VACUOLE

In the mature spores of Myxosporidia belonging to the family Myxo-

bolidae, there exists regularly a more or less conspicuous rounded space

which is generally known as an iodinophilous vacuole because of its be-

havior toward iodine.

Miiller (1841) seems to be the first to notice this. peculiar structure

and figured vacuoles in his drawings. Biitschli (1881:636) observed the

vacuole in the spore of Myxobolus miilleri which had previously been seen |

by Miiller, and designated it as a nucleus. He described the structure as

follows: ‘“Von besonderem Interesse erscheint das unzweifelhafte Vor-

handensein eines Zellkerns in der plasmatischen Inhaltsmasse der Sporen.

Hiufig ist dieser Kern schon in frischem Zustand ohne Weiteres als kreis-

férmiger bis ovaler, heller Fleck recht deutlich sichtbar (Fig. 1n). Besser

tritt er jedoch nach Behandlung mit verdiinnter Essigséure oder Jodtink-

tur hervor und zeigt dann eine dunkle, etwas granulirt erscheinende

Hiille (Fig. 2n) und eine Anzahl ziemlich blasser Granula, welche durch

den Inhalt zerstreut sind. Leider setzten sich dem Versuch, den Kern zu

firben, sehr energische Hindernisse entgegen, da das Farbungsmittel nicht

in die Sporenschale eindringt; jedoch kann dieser Umstand nicht gegen

Kernnatur des fraglichen Gebildes angefiihrt werden, da auch das Plasma

der Farbung widerstand. Dennoch beobachtete ich einige Falle deutlicher

Kernfarbung bei Anwendung von Alaunkarmin.”’

That Biitschli’s view was not correct was shown by Thélohan (1899:

919-920) who studied the structure more closely with the following state-

ments: ‘‘Si, en effet, on traite ces spores par différentes réactifs, on ac-

quiert bienté6t la certitude que la tache claire observée par M. Biitschli, et

décrite et figurée par lui comme un noyau, est en réalité une formation

d’ordre tout différent. Peu visible a l'état frais, a-cause de la transparence

du protoplasma, elle apparait plus nettement par l’action de Valcool, des

acides acétique, azotique, osmique ou du nitrate d’argent a 2 pour 100.

On la voit alors entourée par le reste de la masse plasmique, qui, coagulée

sous l’influence de ces liquides, se distingue par son aspect finement

granuleux et sa moindre réfringence. Elle a tous les caractéres d’une vacu-

ole creusée au sein de cette mass et remplie d’une substance particuliére,

remarquable par sa résistance aux réactifs colorants caractéristiques de la

substance nucléaire. Seul Viode se-fixe sur elle, et, tandis que, sous son

influence, le reste de la spore prend une coloration d’un jaune pale, on

voit cette vésicule devenir d’un rouge brunatre qui rappelle absolument

la teinte que prend la matiére glycogéne par l’action de ce réactif. Comme

celle-ci, cette substance est insoluble dans l|’alcool et grade sa réaction vis-a-

vis de l’iode dans les spores conservées dans ce liquide. Comme elle encore,

elle est soluble dans les alcalis. Les acides la modifient, et aprés leur action

elle ne se colore plus. Toutefois, je n’ai pu, dans ces circonstances,

STRUCTURES CHARACTERISTIC OF CNIDOSPORIDIAN SPORES (fs

obtenir la réduction de la liqueur cupro-potassique.”’ Gurley (1894:209)

is in entire accord with the observations of Thélohan quoted above.

Keysselitz (1908:264) is the only other investigator who studied rather

closely the structure under consideration. He remarks as follows: ‘“‘Die

Vacuole hat eine rundliche Form; ihre Grosse ist nicht ganz konstant. An

der lebenfrischen Spore kann man sie nicht oder kaum bemerken. Nach

Behandlung mit Argentum nitricum, Alkohol, Osmiumsaure (Vgl. Thélo-

han) Aqua destillata, gew6hnlichem Wasser (bei einzelner Sporen) beim

Erhitzen sowie beim Antrocknen tritt sie deutlicher als heller Bezirk her-

vor. Sie ist gegen das umgebende Plasma nicht durch eine deutliche Mem-

bran abgesetzt. Beim Zusatz von wisseriger oder alkoholischer Jodlésung,

farbt sich ihr Inhalt mahoganibraun, eine Reaktion, die fiir die Sporen der

Myxobolen specifisch zu sein scheint. Er erscheint dann zuweilen fast

homogen, hiaufiger bemerkt man verschwommene dunklere und hellere

Flecke verschiedener Grdsse und Form. Der Inhalt scheint mir eine

zahfltissige Substanz zu sein, die in der Zelle gleichsam suspendiert ist.

In konservierten, mit Farbstoffen behandelten Sporen tingiert sich die

Vacuole nicht. Sie imponiert als heller Fleck in der Copula. Durch das

Jod wird in der Regel auch in den zwischen den Polkapseln befindlichen

Raume ein kleiner nicht scharf umgrenzter Bezirk mahoganibraun ge-

farbt.”” Auerbach (1910:16) simply states that ‘‘die Vacuole farbt sich

bei Zusatz von Jodtinktur braun.”

To summarize the views advanced by previous authors, the so-called

iodinophilous vacuole is stained with iodine mixtures, and therefore is of

glycogenous nature.

My observations gave the following results.

1) The effect of distilled water upon the vacuole. In order to deter-

mine whether the vacuole is affected by disti led water in partly exposed

conditions or not, fresh spores of Myxobolus mesentericus and Henneguya

mictospora were crushed under the cover glass, and were kept in distilled

water. After six hours, the smears were treated with Lugol solution, which

gave the following results on examination:

Observations

Control spores without being | Typical coloration of the vacuole

pressed

Spores crushed _ | No visible staining of vacuole-like

structure

The experiments show that the contents of the vacuole disappear when

placed in contact with distilled water.

72 R. KUDO

2) Treatment with Lugol solution. Smears and section preparations

of Henneguya salminicola, Myxobolus aureatus, Myxobolus discrepans and

Henneguya mictospora were treated with Lugol solution. The vacuoles

stained in brownish orange which on warming disappeared.

3) Staining by Lubarsch’s method. Section preparations of Henne-

guya salminicola were stained in Lubarsch’s mixture, the iodinophilous

vacuole, the polar capsules as well as spore membrane were stained in deep

bluish violet.

4) Staining by Best’s method. Section preparations of Henneguya

salminicola were stained by Best’s method. The vacuole took a faint

pink color.

5) Staining with Delafield’s haematoxylin and Lugol solution. Section

preparations and smears of Henneguya salminicola and Myxobolus mesen-

tericus were first stained with Delafield’s haematoxylin. After being

washed thoroughly, they were mounted in gum and Lugol mixture. The

iodinophilous vacuole stained in reddish brown.

From the experiments mentioned above, it is certain that the so-called

iodinophilous vacuole of spores of the family Myxobolidae contains a

substance similar to glycogen in characters.

In the section preparations of the cysts of Henneguya salminicola,

one sees the appearance of the vacuole as the spore formation proceeds.

The glycogenous substance in the sporoplasm remains inconspicuous while

the spore is in the pansporoblast, although one can trace the gradual con-

centration of the substance in it. When the spore matures and separates

itself from the other spore, the vacuole becomes sharply outlined. The

vacuole reaches its maximum size when the spore is completely formed.

No particular body that corresponds to the iodinophilous vacuole was

found in the spores of species belonging to families other than Myxobolidae,

although I have tested several species repeatedly.

It is generally understood without any experimental evidence that the

glycogen occuring in the spores of the family Myxobolidae is probably

used for the future development of the sporoplasm. Then it is strange to

notice the fact that the spores of other families which are essentially the

same in habitat and in many other respects, do not contain the glycogen in

such a conspicuous way as in this particular family. The majority of species

belonging to the family Myxobolidae attack the tissue of the host, yet

some species of the genera Chloromyxum, Myxidium and Sphaerospora

and all the species of the family Myxosomatidae which do not show any

iodinophilous vacuole in the spore at any stage of its development, inhabit

also the tissue of the host. Therefore the occurrence of the iodinophilous

vacuole does not seem to be correlated with the tissue infesting characters

of the Myxosporidia, as was suggested by Gurley (1894).

STRUCTURES CHARACTERISTIC OF CHIDOSPORIDIAN SPORES 73

SUMMARY

1) The spore membrane of Nosema apis and Nosema bombycis, taken

as representatives of Microsporidia, is proved to be composed of a sub-

stance similar to chitin in its chemical reaction.

2) The spore membrane of Henneguya salminicola, taken as a repre-

sentative of Myxosporidia, is proved to be composed of a substance, the

chemical reactions of which are less similar to those of chitin compared

with the microsporidian spore membrane.

3) The polar filaments of cnidosporidian spores are not composed of

glycogen as was suggested by Erdmann. They are formed by the mixture

of a part of the nucleus and a substance differentiated in the capsulogenous

cell.

4) A review of the methods which cause the filament extrusion in

Cnidosporidian spores is presented.

5) The so-called iodinophilous vacuole of the spores of the family

Myxobolidae contains a substance, the chemical reactions of which are

similar to those of glycogen.

BIBLIOGRAPHY

AUERBACH, M.

1910. Die Cnidosporidien. Lepzig. 255 pp.

BALBIANI, G.

1883. Myxosporidies ou psorospermies des poissons. Journ. micr., 7:197—204.

Burtscatt, O.

1881. Beitrige zur Kenntnis der Fischpsorospermien. Zeit. wiss. Zool., 35:627—651.

Davis, H. S.

1917. The Myxosporidia of the Beaufort region. Bull. Bur. Fish., 35:203-243.

ERDMANN, RH.

1917. Chloromyxum leydigi und seine Beziehungen zu anderen Myxosporidien. Teil II.

Arch. Protist., 37:276-326.

GuRLEY, R. R.

1894. The Myxosporidia, or psorosperms of fishes, and the epidemics produced by them.

Rep. U. S. Fish Comm., 5:65-304.

KEYSSELITZ, G.

1908. Die Entwicklung von Myxobolus pfeifferi Thélohan. Arch. Protist., 11:252-308.

Kupo, R.

1913. Eine neue Methode die Sporen von Nosema bombycis Nageli mit ihren ausge-

schnellten Polfaiden dauerhaft zu praparieren und deren Linge genauer zu

bestimmen. Zool. Anz., 41:368-372.

1916. On the structure and life history: of Nosema bombycis Niageli. Bull. Imer. Seric.

Exper. Stat., 1:31—51.

1916a. Notes on Myxosporidia found in some fresh water fishes of Japan, with the

descriptions of three new species. Jour. Parasit., 3:3-9. ;

1918. Experiments on the extrusion of polar filaments of cnidosporidian spores. Jour.

Parasit., 4:141-147.

74 R. KUDO

1920. On the structure of some microsporidian spores. Jour. Parasit., 6:178-182.

1920a. Studies on Myxosporidia. Ill. Biol. Monogr., 5:243-503, 25 pl. and 2 figs.

1921. Notes on Nosema apis Zander. Jour. Parasit., 7:85-90.

Mincam, E. A.

1912. An introduction to the study of the Protozoa. London. 517 pp.

MULreER, J.

1841. Ueber eine eigenthiimliche krankhafte parasitische Bildung mit specifisch organ-

isirten SamenkGrperchen. Arch. Anat. Phys. Med., 5:466-488.

OHLMACHER, A. P.

1893. Myxosporidia in the common toad with preliminary observations on two chromo-

phile substances in their spores. Jour. Amer. Med. Assoc., 20:561-567.

THELOHAN, P.

£889. Sur la constitution des spores des Myxosporidies. C. R. acad. sc., 109:919-922.

1890. Contributions a l’étude des Myxosporidies. Ann. microgr., 2:193-213.

1895. Recherches sur les Myxosporidies. Bull. sc. Fr. Belg., 26:100-394.

Warp, H. B. :

1920. Notes on North American Myxosporidia. Jour. Parasit., 6:49-64.

DEPARTMENT OF SUMMARIES

DEVOTED TO DIGESTS OF PROGRESS IN BIOLOGY

RECENT ADVANCES IN PARASITOLOGY!

By Ernest CARROLL FAUST

Parasitology has made extraordinary progress during the last decade.

In writing on this topic ten years ago Ward (1910) states increasing interest

in problems of medical zoology had made the mass of material at that time

so vast that a review of it was difficult. Since then stupendous progress has

been made.

These advances have been due in part to the more general recognition

of the importance of this science and the relation of its development to

human welfare and in part to the stimulus of the World War. While

individual investigators have contributed a great wealth of valuable data

to the science, the most outstanding discoveries have come as the result

of the work of commissions and bureaus, undertaking fundamental prob-

lems of parasitology on a comprehensive scale. Such accomplished facts

as the eradication of yellow fever and the minimization of malaria in the

Panama Canal Zone, the Hookworm and Tuberculosis campaigns of the

International Health Board, and results of the Bilharzia Mission in Egypt

are outstanding monuments of progress during this decade.

In attacking the problems in hand the life history of the parasite has

been frequently worked out. As in the solution of previous protozoan

and helminth diseases, a knowledge of the life history has not only been

valuable but in most cases the essential factor in the eradication of the

evil. Such a knowledge has shown the most practicable way of breaking

the vicious cycle.

As a result of the World War world problems have developed in para-

sitology from what were formerly matters of Oriental or tropical concern.

Troops coming from countries subject to tropical diseases, returning home,

have brought infections with them. Such is quite likely the case in such

protozoan diseases as amebiasis which require no intermediate host for

part of their life cycle. But, in addition, the added impetus which has

resulted from the study of such infections as World War problems, has

made it evident that the pre-war infection in England and America, for

example, was much higher than had previously been believed.

The period has been marked by the development of new laboratories

and intensive study of parasitic problems in new fields. In part this

‘Contributions from the Department of Pathology, Peking Union Medical College,

Peking, China.

76 ERNEST CARROLL FAUST

work has been done by investigators, who, native to the region, have gained

distinction in such problems. In part it has been accomplished by com-

missions which have been sent into the country to make these researches.

In part it has been brought about by the efforts of those, who, distant

from the field, have diligently sought out a solution to the problems,

brought to them by explorers.

The progress in parasitology has been stimulated and cooperation of

investigators secured by the appearance of several new Journals devoted

entirely or for a major part to parasitology. First must be mentioned

the Tropical Disease Bulletin, London, 1913, a review of all the important

literature on the subject of tropical parasitology and medicine. The estab-

lishment of the Journal of Parasitology (1914) in America affords oppor-

tunity for publications of investigations of a type midway between

Parasitology of Cambridge and the Annals of the Liverpool School. More

recently the Kitasato Archives of Experimental Medicine has entered the

field, affording opportunity for workers in the Orient to publish near at

home. Just recently a long-felt want has been filled by the appearance of

the American Journal of Hygiene and the American Journal of Tropical

Medicine.

Continuing the task of placing in the hands of investigators a depend-

able and indispensable index of Medical and Veterinary Zoology, Stiles and

Hassall (1912) have published their Index-Catalogue on Cestoda and

Cestodaria. Its value over the Trematode Catalogue of the series consists

in the more thoro analysis of specific and sub-specific citations with the

page reference for each and in the fewer number of errors which inevitably

creep into a work of such scope. The long-awaited companion volume

on Roundworms has recently been issued (1920) and meets the expecta-

tions of the most critical reviewer. Along this line one cannot commend too

highly the synopses of important papers relating to medical parasitology

appearing in the Tropical Disease Bulletin. It is to be regretted, however,

that the reviewers of this Bulletin have not seen fit to include certain other

reviews which, altho technically non-medical, are fundamentally related

to medical problems,

New species and new systematology in helminths are brought together

in a most workable digest in the chapters on Platyhelminthes and Nema-

thelminthes in Ward and Whipple’s Fresh-Water Biology (1918). Progress

in American helminthology is shown in the fact that many of these species

are described for the first time and in the introduction of a considerable

portion on cercariae to the subchapter on trematodes. The data are made

especially valuable by their relation to one another in the form of a key,.

and are made the more workable by ample illustrations.

The most comprehensive treatise on the subject of human parasitology

which has appeared within the decade is Fantham, Stephens, and Theo-

DEPARTMENT OF SUMMARIES 77

bald’s “Animal Parasites of Man’ (1916), a book which has no equal in

point of completeness and in up-to-the-minute information on human

entozoa. The writing of a brief review of progress in this field gives an

insight into the monumental character of this book. While strides have

been made in the science ever since the publication of ‘“‘Animal Parasites”’

it remains the reliable compendium and guide to the researcher or prac-

titioner encountering entozoic ailments. In their Manual of Tropical

Medicine (1919) Castellani and Chalmers -have not. only contributed

greatly to the knowledge of tropical protozoa, helminths and arthropods

from data largely drawn from their own wealth of experience in the Tropics,

but they have likewise secured the permanent cooperation of the practi-

tioner in problems of parasitology by presentng the clinical and patholog-

ical pictures of these parasitic infections. The manual stands as a lasting

memorial to the junior author, who gave his life for the work.

Perusal of the literature of parasitology of the period which is covered

in this review reveals a vast wealth of investigation, the major part of

which falls within the group of the protozoa. Workers on protozoa have

been many and a considerable share of their contributions significant.

Certain problems like amebiasis have been studied in new fields. In

other cases the life history has been elucidated. In many cases, however,

mere symptomatology and diagnosis have been set down, where the lack

of new data hardly warrants more than a statement of the case.

Foremost among workers in protozoology are those of the English

Schools, comprising Stephens, Fantham, Nuttall, Yorke, Macfie, Wenyon

and Porter. With these investigators life histories have played an impor-

tant réle. With them, too, detailed descriptions of morphological features

have not been neglected. One is most convinced of the thoroness of the

work of the Liverpool School in reviewing the elaborate and most detailed

methods which have been followed in the experimental treatment of

malaria.

Work on the Continent of Europe of a high character has been done

by Laveran, Leger, Frang¢a, Negri and Galli-Valerio. In the Americas

Craig, Kofoid, Darling, Hadley, Chagas and Magalhaes have made

noteworthy contributions, while Cleland’s solution of dengue in Australia

and Miyajima’s studies on the tsutsugamuchi deserve the highest praise.

In the words of Wenyon (1915) our knowledge of trypanosomiasis and

malaria has reached something like full fruition. Hardly so much can be

said of the majority of protozoon infections, partly because the circum-

stances have not been favorable, partly because the investigations have

been side tracked.

In 1911 Novy touched upon the progress that had been made in our

knowledge of protozoan infections and their treatment. The life history

of Trypanosoma brucei had just been demonstrated (1909) and remained

78 ERNEST CARROLL FAUST

one of the outstanding discoveries of the decade. Little was known of the

spirochaetes and their pathogenicity aside from the studies on treponema.

The life history of the malarial plasmodium had been well authenticated,

but other hematozoon forms were little known. Since then many groups

have been carefully studied. Nuttall (1913) has found the life cycle of

Babesia in dogs, horses and cattle to pass thru certain ticks as intermediate

hosts and has discovered curative salts for these infections. Stephens

(1914) describes a new tertian malarial parasite, Plasmodium tenue, from

the Central Provinces, India. Yakimoff (1917) contributes to the knowl-

edge of Piroplasma, Theileria, Nuttallia and Anaplasma infections of

domestic and semi-domesticated animals of Russian Turkestan. Fantham

(1910) and Hadley (1911) have given a clear morphological analysis of

Eimersa avium.

Again, the studies of Ross and Thomson (1916) on Egyptian sand

amebae show the necessity of preventing contamination of dry sand with

fecal matter.

Wenyon and O’Conner (1917) have helped to solve the practical treat-

ment of the protozoan infections of man in Egypt. They have been

able to standardize treatment of amoebiasis. Three new human Parasites,

Waskia intestinalis and Tricercomonas intestinalis and Entamoeba nana

have been found in these studies.

Craig (1917) has established a basis for classification of amebae parasitic

in man which allows one to profit from his numerous investigations in this

field. He recognizes as valid species, Craigia hominis and C. migrans,

Endamoeba coli, E. histolytica and E. gingivalis, and Vahlkampfia lobos pin-

osa. A more conservative standard is presented by Dobell in his mono-

graph on the amoebae (1919), a treatise which for its thoroness commands

the attention and admiration both of the theoretical and ‘the practical

parasitologist.

Von Prowazek (1913) has published an important paper on Balantidium

coli. He has carefully reviewed the geographical distribution of the

species, described its histology in minute detail and methods of propaga-

tion, and has worked out its pathogenicity.

Work of the character of Fantham and Porter’s (1914) contribution

to the life-history of Nosema bombi has been of increasing importance in

elucidating the general knowledge of protozoan life cycles and thus con-

tributing indirectly to a knowledge of related human forms where experi-

mental infections are obviously less possible. In reviewing the work on

protozoa one is struck by the mass of such work of an excellent character

of which lack of space unfortunately does not even permit mention.

Watson’s monograph (1916) on Gregarines constitutes a well organized

synopsis of new and described species of the group, many species of which

had previously been investigated only piecemeal

See SS an ke

DEPARTMENT OF SUMMARIES 79

Dobell (1918) has contributed a valuable memoir in his study of human

coccidia. Following up the work of Wenyon (1915) he has described three

definitely known species infecting man (Isospora hominis Riv. 1878,

Eimeria wenyoni n. sp. and E. oxyspora n. sp.,), in addition to throwing

doubt on the identity of a third Emeria species as that of the rabbit

(E. stiedae).

Moroff (1915), after a searching investigation, places the sarcosporidia

close to the gregarines and coccidian forms in the subclass Telosporidia,

along with the Haemosporidia.

Wolback’s work on the Rocky Mountain spotted fever (1918) has |

shown that the causal agent of the disease is a minute parasite, present in

the blood of infected mammals and in ticks which are capable of transmit-

ting the disease.

Kudo’s monograph on the Myxosporidia (1920) is memorable not

only as a collation of the work of earlier investigators, but as a survey of

the large number of myxosporidian forms studied by Kudo himself.

The work of Poche (1913) on the System of Protozoa is a comprehensive

treatment of nomenclature of the group. It is notable for the large number

of new orders, suborders and families proposed, many of which are readjust-

ments of rank justified by the increase in number of the group. With

the wealth of knowledge of morphology and life histories of the Protozoa

careful systematic readjustments of this type are increasingly necessary.

- Work on the helminths has been continued by many of the investigators

who have already established a name for themselves among parasitologists.

In Europe Odhner has contributed further studies to his work on phylogeny

and systematology, among the most interesting of which are those on Schis-

tosome and Holostome groups. Goldschmidt has extended his investiga-

tions on cytology most successfully. Kossack has monographed the

monostomes, while Monticelli and Liihe have contributed much to the

knowledge of trematodes. Fuhrmann, Leon and von Ratz have studied

the cestodes while Railliet and Henry and Seurat have made notable

contributions to the nematodes. The most brilliant work of the younger

helminthologists in Europe is undoubtedly that by Leiper.

In America such studies have been continued by Ward, Ransom and

Young on Cestodes, by Ward on Trematodes, and by Ward and Ransom on

nematodes. In addition there has arisen in the United States a considerable

group of younger investigators, of whom La Rue, Cort, Boeck and Van

Cleave deserve prominent mention.

In Japan Katsurada, Fujinami and Goto have produced work of high

merit. Yoshida, Okanama, Kobayashi and Miyairi have added much to

life-history problems.

The contributions on Australian helminths count among their number

the investigations of Nicoll, Cleland, S. J. Johnston, T. H. Johnston,

80 ERNEST CARROLL FAUST

Brein] and Sweet. This summary of important contributors to the science

would not be complete without mention of Ssinitzin, Skrjabin and Yaki-

moff for Russia and Southwell for India.

Ward (1917) has emphasized the necessity of rearranging forms “so

as to express better their correct relationships in the light of more perfect

knowledge of their structure.” But he adds the essential corollary that

it has been his fixed principle never to make any change until he was

personally familiar with the form discussed or had acquired such acquaint-

ance with its structure as to know that some change was inevitable and

that the proposed modification was defensible on morphological grounds.

On this basis he has made fundamental but conservative changes in

trematode and acanthocephalan groups and has established order in the

nematode group where previously taxonomy was confined most usually

to mere descriptions of new species.

The period has experienced an advance in helminthology from an

almost purely zoological science to one ministering to the needs of com-

parative bionomics and medicine.

The outstanding morphological and systematic contribution to our

knowiedge of the Cestoda during recent years is La Rue’s Monograph on

the Family Proteocephalidae (1914). Provided with a wealth of American

material, supplemented by more than on ordinary amount of types of the

group described by European and other workers, La Rue has been enabled

to mold the material into a comprehensive and practically exhaustive

treatise. His descriptions and drawings are detailed, yet clear, his types

are well defined and the amount of material collected, the amount used in

study and the location of each specimen in the collection are minutely set

down. Added to this are valuable synoptic tables and a workable natural

key to the group. The contribution as a whole is such as to place the author

immediately in the rank of the foremost helminthologists.

Recently Cooper (1919) has monographed another group of cestoda

from fishes which contributes to our knowledge in that group.

Among other contributions on cestode anatomy and phylogeny the

work of Douthitt (1915) on Anoplocephalidae is worthy of mention.

Because of the care which this investigator used in working over his material

and the gradual way in which he built up a natural classification of the

group the monograph will serve as a lasting memoir to his efforts.

Ransom (1913) has made possible the statement that Cysticercus ovis

is the intermediate stage of a dog tapeworm, Taenia ovis (Cobbold) Ransom

and in working out the life-history of this cestode experimentally has solved

a problem of long standing. This species in the bladder-worm stage

has thus been proved to be distinct from Cysticercus cellulosae and the

adult from Taenia tenella, T. solium, T. hydatigena, and T. marginata.

Treatment of dogs for the tapeworm is found not only to eradicate this per-

DEPARTMENT OF SUMMARIES 81

plexing economic problem of sheep measles but rids them of other worms

of equally serious pathogenicity.

Beddard (1911-1914) has contributed studies from time to time,

making known to science a large number of cestodes parasitic in animals

in the Zoologicai Society Gardens (London). Likewise Skrjabin (1914)

has contributed to our knowledge of the Cestoda of birds of Russian Turkes-

tan. Fuhrmann (1918) in a detailed survey of the avian cestodes from

New Caledonia and Loyalty Isle adds materially to the knowledge of the

families Tetrabothriidae, Anoplocephalidae, Davaineidae, Dilepididae,

Hymenolepidae, Acoleidae and Amabiliidae.

Thus the comparative work on cestodes has been greatly advanced.

The striking advances in our knowledge of the trematodes have come

as life-history problems. Members of the medica] profession have been

especially sympathetic to this work because it was concerned with flukes

most of which affected-man. It is particularly noteworthy that all of

these without exception have borne out the principle established for

Fasciola hepatica, that the miracidium penetrates a mollusk, and from

the mollusk the cercaria emerges which reaches its definitive host (immedi-

ately or intermediately according to the group to which the species belongs)

and there becomes mature. Faust (1918) following Ssinitzin’s work (1911)

has found the sporocyst and the redia stages to be true parthenitae.

A problem which Looss had repeatedly attempted to solve in Egypt

and on which Katsurada and Fujinami have contributed much in Japan

was the schistosome life history. Credit for the first solution of the life

cycle is due to Miyairi and Suzuki (1914) in Japan and later to Leiper and

Atkinson for Japanese species and Leiper (1915) for the two Egyptian

species. A clear understanding that the miracidium enters a gasteropod and

that by change of cycle the cercaria emerges from the snail and directly

infects man thru the skin or the mouth has made possible methods for pre-

venting the disease. It has also made possible a clear restatement of the

thesis that ‘‘The larval metamorphosis of all digenetic trematodes occurs

without known exception in the bodies of molluscs belonging to the classes

Gasteropods and Lamellibranchia.”’ Leiper (1918) has shown that when

once infected the patient is practically incurable. He has found from his

Egyptian Researches that

“(1) Transient collections of water are quite safe after recent contami-

nation.

‘“(2) All permanent coilections of water, such as the Nile, canals,

marshes and birkets, are potentially dangerous, depending upon the

presence of the essential intermediary host.

(3) The removal of infected persons from a given area would have

no effect, at least for some months, in reducing the liability to infection,

as the intermediate hosts discharge infective agents for a prolonged period.

82 ERNEST CARROLL FAUST

“‘(4) Infected troops can not reinfect themselves or spread the disease

directly to others. They could only carry the disease to other parts of

the world where a local mollusc could efficiently act as a carrier.

““(5) Infection actually takes place both by the mouth and through

the skin.

‘““(6) Infection in towns is acquired from unfiltered water which is

still supplied, ever in Cairo, in addition to filtered water, and is delivered

by a separate system of pipes.

“(7) Eradication can be effected without the cooperation of infected

individuals by destroying the molluscan intermediaries.”’

Nakagawa (1916) has unravelled the life cycle of Paragonimus Wes-

termani, showing that the cercaria is developed in Melania species and

the encysted larva in Potamon. These infected crabs when fed to pup-

pies gave rise to typical pulmonary paragonimiasis. Moreover the

route of infection has been found to be from the intestinal wall in the

vicinity of the jejunum, thru the abdominal cavity, thru the diaphragm

and pleural lining, where it bores thru the lung tissue and encysts. Yoshida,

working on the same problem entirely independently in Japan, was able to

substantiate Nakagawa’s results. Kobayashi’s work on this fluke in

Korea (1918) has hardly as convincing data as those of his colleagues.

On the other hand the latter investigator (1915) has clearly shown experi

mentally that the encysted larva found in several species of Japanese

fresh-water fish develops into the human fluke, Clonorchis sinensis. Thus

far, however, he has not worked out the cercarial phase of the life-history

of this worm. Nakagawa (1921) has just published his experimental

work on Fasciolopsis buski, which he finds to infect the hog as the encysted

post-cercarial distomule.

Recently interest in larval trematodes has been revived and the field

for study of this group in America has been studied by Cort and Faust,

who have shown that specific marks of discrimination in cercariae are

important even tho they are minute. These investigators have added

data on the larvae which should facilitate life history studies on flukes.

Among these studies are those on flame-cell constancy and homology,

including the use of this system as a basis for systematic relations. Of

importance both to pure science and to medical parasitology, Cort’s

monograph (1919) on the cercaria of the Japanese blood fluke sets a record

for careful study and detail in this group. Furthermore, Cort’s study on

the stages of development of the schistosome in the definitive host (1921)

makes a valuable addition to the ontogeny of the fluke.

One can not overlook the researches of Ssinitzin (1911) in this field.

This investigator has not only presented data on many interesting and

unique larval flukes, but has presented theories of their phylogenetic

relations which are at least extremely suggestive and stimulating.

DEPARTMENT OF SUMMARIES 83

A morphological paper which has done.much to show the necessity of

exactness in differentiation of closely related species is that of Ward and

Hirsch (1915) on the species of Paragonimus. These authors have found

the type, size and group relationships of the spines to be distinctly diagnos-

tic, and this fact, coupled with-the importance of one of these species to

medical science in the Orient makes the work especially significant.

Comparatively few investigators have made important studies during

the past decade on the morphology and systematology of parasitic nema-

todes. Railliet and Henry in France and Ward and Magath in America

have published researches which constitute marked exceptions to this

lack of such investigation in this group. The first significant analysis of

the parasitic Nematodes in America is embodied in Ward’s chapter on

these worms in Ward and Whipple’s Fresh-Water Biology (1918).

A most important contribution to the morphology of the nematode

is embodied in Magath’s monograph of Camallanus americanus (1919).

This thesis constitutes the most significant work on a single nematode

species since the researches of Looss on Ancylostoma duodenale. The

writer describes in detail the organs and systems of the worm and arrives

at the conclusion that formulae for measurement are not dependable but

that where doubt arises in systematalogy there remains only the accurate

morphological description of every organ and part of the form in question.

The most widespread campaign ever undertaken by governmental or

private interest for the eradication of a particular disease is that which

was undertaken by the Rockefeller Foundation for the banishment of

hookworm from the earth. In 1909 the Rockefeller Sanitary Commission

was created to combat the hookworm in the United States. The findings

of this Commission of the prevalence of the worm, the. “‘arrest of physical,

mental and moral growth, great loss of life, and noticeable decrease in

economic efficiency,” together with the success which attended treatment

of hookworm infection, led to the establishment in 1913 of the International

Health Commission (afterwards known as the International Health Board)

with the purpose in view of “extending to other countries the work of

eradicating hookworm disease as opportunity should offer” and, so far as

practicable, to follow up the treatment and cure of this disease with the

establishment of agencies for the promotion of public sanitation and the

spread of the knowledge of scientific medicine.

Forthwith this commission proceeded to determine 1) the geographic

distribution and the approximate degree of infection, 2) to examine

microscopically the cases and cure those infected, and 3) to establish

sanitary conditions which would prevent soil-pollution.

At the close of 1918 the Board had solely or cooperatively attacked

the problem in the Southern United States, Central Mexico, Cuba, Porto -

Rico, Jamaica, a considerable share of South America, Egypt, Ceylon,

84 ERNEST CARROLL FAUST

Siam, The Malay, South China, New Guinea, Papua, Java, Guam, and

Queensland, Australia, and new work was under way in the Madras

Presidency, India. ;

While the intensive method of microscopic examination and treatment

of patients within a limited area was utilized, the more fundamental

purpose of the campaign has been to develop an education propaganda

for better sanitary conditions so that the sources of infection will be

eliminated.

One of the fundamental life-history problems which has engaged

the attention of investigators in several geographically different centers

is that of Ascaris. Captain Stewart of the Indian Medical Service (1917,

1918) has shown that Ascaris lumbricoides, and A. mystax can be developed

to a certain larval stage in the mouse and rat.

Ransom and Foster (1917, 1919, 1920) have been able to produce indi-

viduals more nearly mature in the sheep and goat. The latter writers have

shown, however, that these stages of development in animals other than

the hog and man do not necessarily imply that the mouse, or rat, sheep

or goat serve as intermediate hosts for these parasites. Yoshida (1919),

working on guinea-pigs, was able to trace the life history as follows: ““The

ascarid larvae escape from the egg shell in the intestine of the host and

proceed to the abdominal cavity by boring through the wall of intestine.

Thence they pierce the diaphragm to enter the pleural cavity, finally

penetrating into the lungs from their surface. ... Furthermore, the

lungs are the only necessary and important organ to be passed by the

larvae in the course of their development. ... (They) continue their

development and migrate to the mouth cavity through the trachea, again

passing down the alimentary canal to the intestine of the host.

Work on the Acanthocephala has been relatively meager. Liihe’s

digest of the group (1911) has given a basis for Continental investigations

while Van Cleave’s numerous studies on American species constitute

marked progress in methods and thoroness of investigation.

Almost the entire amount of our knowledge of insects in the rdle of

catrier and intermediate hosts of parasitic disease has come within the

last few years. It is within this period that the life histories of the trypano-

some, the piroplasmas and the spirochaetes have been shown to develop in

specific flies, fleas, bugs or lice as the case may be. Moreover, certain

tapeworms, especially those of poultry, have been just recently shown to

develop as larvae within insect hosts. Finally there is further proof of the

importance in the spread of parasitic disease when the insect acts merely as

a vector. For these reasons important campaigns against these several

insects have been carried on by private forces and by government agencies,

foremost of which is the all but complete eradication from the Western

Hemisphere of yellow fever by controlling the mosquito transmitting the

AR lat,

DEPARTMENT OF SUMMARIES 85

disease, with plans under way for a campaign on this insect in the remaining

locus of infection.

Thus a considerable share of the problems which confronted parasitol-

ogy at the beginning of the decade have been carried to completion while

others are being gradually sifted out. In their place, however, have come

still others which require the greater skill and the wider point of view for

their full solution. All of these signs of progress.in parasitology indicate

that this science is rapidly coming to assume the place which it deserves as

the companion of bacteriology and gross pathology.

IMPORTANT LITERATURE ON PARASITOLOGY

General

CASTELLANI, A. and CHALMERS, A. J.

1919. Manual of Tropical Medicine. 2436 pp. London.

’ FANTHAM, H. B., STEPHENS, J. W. W., and THEOBALD, F. V.

1916. The Animal Parasites of Man. 900 pp. London and New York.

Stizes, C. W., and Hassatt, A.

1912. Index Catalogue of Medical and Veterinary Zoology. Subjects: Cestoda and

Cestodaria. Hyg. Lab. Bull., No. 85, 467 pp.

1920. Ibid. Subjects: Roundworms. Hyg. Lab. Bull., No. 114, 886 pp.

Warp, H. B.

1910. Recent Progress in Parasitology. Trans. Am. Micr. Soc., 29:119-158.

1917. On the Structure and Classification of North American Parasitic Worms. Jour.

Parasit., 4:1-12.

Warp, H. B. and Wurrte, G. C.

1918. Fresh-Water Biology. Chapters on ‘Parasitic Flatworms” and “Parasitic

Nematodes.” 1111 pp. New York.

Protozoa

Grave Cah:

1917. The Classification of the Parasitic Amoebae of Man. Jour. Med. Research

35:425-442.

DoBELL, C.

1918-1919. A Revision of the Coccidia Parasitic in Man. Parasit., 11:147-197, 1 pl.

1919. The Amoebae Parasiticin Man. 155 pp., 4 pl. London.

FanTuay, H. B.

1910. The Morphology and Life History of Eimeria (Coccidium) avium, a Sporozoon

Causing a Fatal Disease among Young Grouse. Proc. Zool. Soc. London, 1910:

672-691, 4 pl.

FanTuam, H. B. and Porter, ANNIE.

1914. The Morphology, Biology and Economic Importance of Nosema bombi n. sp.,

Parasitic in Various Humble Bees (Bombus spp.) Ann. Trop. Med. Parasit.,

8:623-638, 1 pl.

Hantey, P. B.

1911. Eimeria avium, a Morphological Study. Arch. Protistenkde., 23:7-50, 2 pl.

Kupo, R.

1920. Studies on Myxosporidia. Ill. Biol. Monogr., 5:244-503, 25 pl.

Mororr, Tu.

1915. Zur Kenntnis der Sarkosporidien. Arch. Protistenkde., 35: 256-315.

Novy, F. G. °

1911. Recent Achievements in Parasitology. 13th Rept. Mich. Acad. Sci., 18-32.

NUvuTratt, G. H. F.

1913. The Herter Lectures. III. Piroplasmosis. Parasit., 6:302-320, 14 figs.

PocueE, F.

1913. Das System der Protozoa. Arch. Protistenkde., 30:125-321, 1 fig.

VON PROWAZEK, S.

1913. Zur Kenntnis der Balantidiosis. Arch. Schiffs-Trop. Hyg., Beihefte 17:369-390,

2 pl.

Ross, R. and THomson, D.

1916. Studies on Egyptian Sand Amoebae. Proc. R. Soc. Med., Sec. Epiderm. and St.,

Med., 9:38-48.

DEPARTMENT OF SUMMARIES 87

STEPHENS, J. W. W.

1914. A New Malarial Parasite of Man. Ann. Trop. Med. Parasit., 8:119-128, 3 pl.

Watson, M. E.

1916. Studies on Gregarines. Ill. Biol. Monogr., 2:213—-468, 15 pl.

WEnyon, C. M.

1915. Leishmania Problems. Jour. Trop. Med., 18:241—247.

WEnvyovy, C. M. and O’Connor, F. W.

1917. An Inquiry into Some Problems affecting the Spread and Incidence of Intestinal

Protozoal Infections of British Troops and Natives in Egypt. Jour. R. Army

Med. Corps, 28:1-34, 151-187, 346-370; 4 pl.

WOLBACK, S. B.

1918. The Etiology and Pathology of Rocky Mountain Spotted Fever. Jour. Med.

Research, 32:499-508.

Yaxiorr, W. L.

1917. Parasites du sang des animaux en Transcaucasie. Bull. Soc. Path., exot., 10:

98-99.

Cestoda

BEDDARD, F. E.

1911-1914, Contributions to the Anatomy and Systematic Arrangement of the Ces-

toidea. I-XV. Proc. Zool. Soc. London.

Cooper, A. R. ;

1919. North American Pseudophyllidean Cestodes from Fishes. Ill. Biol. Monogr.,

4:295-541, 13 pl.

Dovutmrt, H.

1915. Studies on the Cestode Family Anoplocephalidae. Ill. Biol. Monogr., 1:353-446,

6 pl.

FUHRMANN, O.

1918. Cestodes d’oiseaux de la Nouvelle-Caledonie et des Iles Loyalty. In Sarasin

and Roux’s Nova Caledonia, Zoologie. 2: (Lief, 4) No. 14. 2 pl, 78 figs. Wies-

baden.

La Rug, G. R.

1914. A Revision of the Cestode Family Proteocephalidae. Ill. Biol. Monogr., 1:1-350,

12 pl.

Ransow, B. H.

1913. Cysticercus Ovis, the Cause of Tae Cysts in Mutton. Jour. Agr. Research,

i558. pl, 13 figs.

SKRJAKIN, K. J.

1914. Beitrag zur Kenntnis einige Vogelcestoden. Centralbl. Bakt. Parasit., (I) Orig.,

75:59-83.

Trematoda

Cort, W. W.

1919. The Cercaria of the Japanese Blood Fluke, Schistosoma japonicum Keareueede.

Univ. Calif. Pub., Zool., 18:485-507, 3 figs.

LOZ the Bevelaprien of the Japanese Blood Fluke, Schistosoma japonicum Katsur-

ada, in its Final Host. Am. Jour. Hyg., 1:1-38, 4 pl.

Faust, E. C.

1918. Life History Studies on Montana Trematodes. Ill. Biol. Monogr., 4:1-120, 9 a

Kopayasut, H.

1915. On the Life History and Morphology of Clonorchis sinensis. Centralbl. Bakt.

Parasit (I) Orig., 75:299-318, 4 pl.

1918. Studies on the Lung-Fluke in Korea. Mitt. Med. Fachschule, Keijo, 1918, pp.

97-115, 2 pl.

88 ERNEST CARROLL FAUST

LEIPeEr, R. T.

1915. Report on the Results of the Bilharzia Mission in Egypt, 1915. Jour. Roy. Med.

Corps, 25:253-267.

Mryarrt, K. and Suzukt, M.

1914. Der Zwischewirt des Schistosomum japonicum Katsurada. Mitt. Med. Fak.

Univ. Kyushu, Fukuoka, 1:187—197, 2 pl.

NakaAGawa, K.

1916. The mode of Infection in Pulmonary Distomiasis. Jour. Infect. Dis., 18:131-

142, 2 maps, 4 pl.

ODHNER, T.

1910. Results of the Swedish Zoological Expedition to Egypt and the White Nile

1901. No. 23A. Nordostafrikanische Trematoden. I. Fascioliden. 170 pp. 6

Tafs. 14 tfigs. Uppsala.

1911-1913. Zur natiirlichen System der digenen Trematoden. I-VI. Zool. Anz., 37:

181-191, 237-253; 38: 97-117, 513-531; 41: 54-71; 42: 289-318.

SSINITZIN, D. Tu.

1911. La Génération parthénogénetique des Trématodes et sa descendence dans les

mollusques de la Mer Noire. Mem. Acad. Sci. St. Petersbourg, (8) 30:1-127,

6 pl. (Russian).

Warp, H. B. and Hirscu, E. F.

1915. The Species of Paragonimus and their Differentiation. Ann. Trop. Med. Parasit.,

9:109-162, 5 pl.

Nematoda and Acanthocephala

Macatu, T. B.

1919. Camallanus americanus, nov. spec. Trans. Am. Micr. Soc., 38:43-107, 10 pl.

RAILLIET, A. ET HENRY, A.

1914. Essai de Classification des “Heterakidae.” IX Congr. Int. Zool., Monaco, pp.

674-682.

1916. La Famille des Thelziidae. Jour. Parasit., 2:99-105.

Ransom, H. B. and Foster, W. D.

1917. Life History of Ascaris lwmbricoides and Related Forms. Jour. Agr. Research,

9 :395-398.

1919. Recent Discoveries Concerning the Life History of Ascaris lumbricoides. Jour.

Parasit., 5:93-99.

1920. Observations on the Life History of Ascaris lumbricoides. U.S. Dept. Agr.

Bull. 817. 47 pp.

STEwart, F. H.

1917. On the Development of Ascaris lumbricoides Lin. and Arcaris suilla. Duj. in

the Rat and Mouse. Parasit. 9:213-227, 2 pl.

1918. On the Development of Ascaris lumbricoides and A. mystax in the mouse. Para-

sit., 19:189-196, 1 pl.

Yosuipa, S. O.

1919. On the Development of Ascaris lumbricoides L. Jour. Parasit., 5:105-115.

Ltue, M. ;

1911. Acanthocephalen. Siisswassfauna Deutschlands. 116 pp., 27 figs., Jena.

VAN CLEAVE, H. J.

1918. The Acanthocephala of North American Birds. Trans. Am. Micr. Soc. 37:19-48,

5 pl.

1919. Acanthocephala from the Illinois River, with Descriptions of Species and a Synop-

sis of the Family Neoechinorhynchidae. State Ill. Nat. Hist. Survey, 13:225-257,

7 pl.

DEPARTMENT OF METHODS, REVIEWS, ABSTRACTS,

AND BRIEFER ARTICLES

A METHOD FOR ORIENTING AND MOUNTING MICRO-

SCOPICAL OBJECTS IN GLYCERINE

CHARLES BULLARD

The object of this paper is to describe a method of mounting desmids

or similar microscopical objects in glycerine, so that they may not

only be drawn in different positions with the camera lucida, but may

form the basis of an herbarium of mounts which may be regarded as per-

manent, since the oldest preparations made by this method, now about

twenty-five years of age, show no signs of deterioration. The account

herewith presented is offered for the purpose of enabling others to utilize

it, as well as to answer certain inquiries that have been made in regard to

the subject. In its details the procedure presents no novel features. Its

principles are those used in the laboratory of Professor Thaxter for mount-

ing the lower fungi and other of the more delicate Thallophytes in glycerine,

or glycerine and eosin, and sealing with King’s Cement.! This method is

especially well adapted for mounting desmids for study from the point of

view of the systematist; since, in the vast majority of species only the well

developed empty cells and semi-cells are useful for this purpose. Those

species of Cosmarium or of Spirotaenia, for example, the cell-contents of

which have to be studied, require more exact fixation methods, with which

this paper is not concerned.

In mounting a sufficiently large object, no ‘‘finding ring”’ is needed.

But for marking the position of a small or large series of minute objects,

it is often essential. A ring of Brunswick Black has been found most

serviceable for this purpose, and should be prepared months in advance,

so as to become well seasoned. It may be placed centrally on the slide

by means of the turntable, and need not be larger than the field of the low

power.

THE MATERIALS AND THE METHOD OF THEIR USE

Since the procedure here described involves the use of a weak glycerine

jelly as a means of orientation, it is necessary, in order to avoid the diffi-

1 King’s Cement was an invention of the Rev. J. D. King of Cottage City, Massachusetts

who did not publish the formula or method of preparation. There is an antiquated recipe

published on page 235 in Rev. A. B. Hervey’s translation of Behren’s “‘Guide to the Micro-

scope in Botany” (S. E. Cassino Co., Boston 1885). Dr. Hervey assures me that it is genuine,

as he received it directly from King. If the more modern form of this cement cannot be

obtained from a dealer, it can be bought of its present maker, Professor R. E. Schuh, Howard

University, Washington, D. C.

90 CHARLES BULLARD

culties inevitably associated with the process in warm weather, to perform

the manipulations indicated in the cooler part of the year. After the

objects are oriented, the weak jelly may be satisfactorily set by placing

the slide-box containing the preparations outside a window where it will be

chilled. The drawing can be done in the warm part of the year, when the

light is also best. This weak jelly is made from any good clear glycerine

jelly, such as that prepared by Kaiser’s formula. A few drops of melted

jelly in a small vial is reduced by the addition of boiled or distilled water,

until the mixture will just set at the ordinary temperature of the room.

It should be perfectly limpid. The cork may be furnished with a dropper,

by pushing into its lower end a piece of platinum wire of such length that

a small loop at the lower end nearly touches the bottom of the vial. The

vial should be kept well corked.

Objects may be lifted, transferred and oriented by means of a fine

needle such as “No. 12 Sharps.”’ The needle is pushed eye first into the end

of a large match of straight grained wood, until a quarter of an inch, or

less, of the pointed end, which is thus as rigid as possible, remains project-

ting. A smaller instrument may be made by the addition of a proper

bristle, for which purpose a carefully selected whisker of a cat or dog

answers admirably; since it combines stiffness and elasticity with an

extremely delicate point. This bristle should be cemented to the mounted

needle, and bound in place by means of a long human hair, or a fine waxed

silk thread, in such a position that the point of the bristle projects slightly.

It is important in order to obtain the necessary rigidity that the free

portion of the bristle should be as short as possible consistently with con-

venient use.

The half inch circular cover glass is best adapted for general use, and

in mounts of this nature, it is necessary to employ a shallow cell. This may

be readily made by supporting the cover at one side by either a somewhat

compressible, or an entirely rigid support. The latter results n a better

mount mechanically, the former is easier to work with. The more flexible

support, consists in fibers of blue blotting paper completely picked and

teased out and then felted together again into a ball by means of the

forceps. This material has the advantage of compressibility, which permits

one to vary a little the amount of glycerine used for the mounting medium.

It has a certain disadvantage, however, from the fact that, as it is not a

rigid support, care must be taken that a mass of sufficient size is used, to

prevent the cover from touching the object, after the cement has dried

and contracted. A rigid cover glass support may be made by selecting a

very thin cover glass and placing it in an elongated folded paper. If this

is pressed against the edge of the tableand drawn back and forth, the glass

within will be finely comminuted. A minute fragment of this may then be

DEPARTMENT OF METHODS 91

used to support the cover. One must learn by experience the approximate

amount of glycerine needed to fill the space beneath the cover.

The cement may be kept in an ounce, or half-ounce, wide mouthed

bottle. Into the bottom of the cork a match, bearing the ringing brush is

inserted; the point of the brush nearly touching the bottom of the bottle.

The ringing brush itself should be the smallest obtainable with not many

but rather long hairs. A match may be pushed into the quill and firmly

bound with silk, and the whole fitted to the cork as mentioned.

For a mounting medium it is best to use only chemically pure glycerine,

filtered if necessary. It may be kept in a vial to the cork of which is fitted

a platinum loop, or a properly selected mourning pin, by means of which

the glycerine may be conveniently applied.

MANIPULATIONS

The details of the procedure for mounting large and small objects

are the same, but the smaller species are obviously the more difficult. A

slide is prepared for the reception of the objects by placing a minute smear,

or streak of the weak jelly, within the ring previously prepared; or if

desirable a series of minute drops may be used. It is convenient to employ

an ordinary dissecting stand, with a x12 aplanatic triplet, on which the

slide thus prepared is left, with the focus and light exactly regulated. The

material to be mounted having been spread out in glycerine on a slide,

the particular individual desired is selected under the compound micro-

scope, and pushed about with the needle beyond the edge of the glycerine

until freed from glycerine and all extraneous matter. In this condition it

will readily stick to the needle, and can thus be lifted from the slide, trans-

ferred to the surface of the weak jelly, and there left until all the individ-

uals desired for this mount have been transferred. The weak jelly should

then be liquefied by breathing gently upon it, and the slide at once placed

under the low power of the compound microscope. It will then be found

that, with a little practice, the objects can be easily and systematically

arranged with the needle. They may be set up in lines, or curves, in

whatever order may prove most convenient. Should the jelly harden too

rapidly, it may again be liquefied as above described. In a short time,

one acquires skill in setting up objects, such as desmid semi-cells, in

different positions under the compound microscope, without disturbing

those already in position. For drawing care should be taken so to set up

a symmetrical object that its vertical axis coincides with the optical axis,

or nearly so. The horizontal axes may be pointed in any direction, by

merely revolving the slide on the stage. After placing and orienting the

specimens, the slide may be put away upside down in a slide box, until two

or three more are brought to the same state of-preparation. In this con-

92 CHARLES BULLARD

dition they may be left outside a window all night in order to harden the

jelly.

When the preparation is ready to mount and seal, the support of

blotting paper, or cover chip, already mentioned, should be placed near

the edge where it will be just included by the cover as it is lowered. A

clean drop of glycerine should then be placed on the object, sufficient to

fill the cell as exactly as possible. If blotter shreds are used, and the

glycerine does not quite fill up the cell after the cover is placed in position,

cautious pressure over them with the point of the needle will spread the

glycerine, and fill the cavity completely. It may then be carefully sealed.

If more glycerine must be added to fill a cell, a small drop should be placed

at a short distance from the edge of the cover, and a narrow streak of it

drawn with the needle to the edge of the cover; so that a little will flow

beneath it. After repeating this process with the needle till the cell is

completely filled, the surplus glycerine must be carefully wiped off.

The complete removal, before sealing, of this surplus is absolutely essen-

tial for the preparation of a permanent mount; since this is the only way

to prevent subsequent leakage. A very little cement under the edge

within the mount serves to make it stronger. In order to remove all trace

of glycerine, it must be very carefully wiped off by means of an old, much

washed handkerchief, folded over the end of the forefinger in such a manner

as to form a point, which is moistened with alcohol. This can safely be

pushed up till it touches the edge of the cover, and then repeatedly re-

newed and worked around it unti. all glycerine is removed. Incase astill

larger surplus of glycerine must be removed, it is convenient to use small

strips of blotter folded A shape. One end is moistened with alcohol and

pushed up against the edge of the cover. Several pieces about the cover

edge absorbing simultaneously will gradually remove most of the excess,

after which, the slide must be very carefully cleaned with the handkerchief,

as before mentioned, and the mount at once sealed.

When there is only one specimen, and different aspects of it must be

drawn, the preparation is demounted after the first figure is made, and

before sealing the mount. The cover is lifted off, the glycerine drained

away, the jelly again liquefied as before, and the object transierred to an-

other prepared slide, remounted in a new position and drawn again. The

last remount, showing the object in its most characteristic position, may

be sealed.

To seal a perfectly cleaned mount, it may be held in the unsupported

left hand, while with the right, a light ring of cement is applied, after

resting the right hand on the left. With a full brush, a drop of cement is

then started on the edge of the cover, the hands being held as before, and

led around the circle by means of the brush in such a manner that the

DEPARTMENT OF METHODS 93

cement ring is partly on the cover and partly on the slide. The first ring

thus helps to keep the second under better control. Additional rings may

be applied on the turntable, when the sealing rings are hard. It has been

found convenient to finish with two coats of Brunswick Black. This is

soluble in turpentine, and offers more resistance to the solvent action of

alcohol used to clean immersion oil from the cover.

In glycerine mounts of most objects, collapsed specimens regain their

turgescence; and air bubbles, if present, disappear in a few days. Since

these mounts are delicate, they must be handled with care, and always

kept horizontal. Any necessary modifications of this method may be

made in order to mount other microscopical objects needing orientation

for camera drawing, and thus make it possible to obtain more accurate

figures. But the fact that mounts thus sealed have shown no signs of leak-

age for so many years, indicates that the correct principles have been

applied at this critical point, and should not be lightly changed.

Cambridge, Mass., April, 1921

A METHOD OF DEMONSTRATING THE SHEATH STRUCTURE

OF A DESMID

The structure of the cell wall in the Desmids is intimately concerned

with the method of formation of the mucilaginous sheath, which in many

members of the group is found to surround the cell. In the Saccodermae

the wall is believed to be continuous thruout, having no pores communicat-

ing with the exterior. On the other hand, the Placcodermae in addition

to other distinguishing characteristics frequently show pores connecting

the protoplast with the surrounding medium.’ In the Placcodermae it is

considered that the mucus exudes thru the pores, and may accumulate

outside the cell wall, so forming the sheath. It is not usually possible to

observe directly evidence of this extrusion, but in the filamentous desmid

Hyalotheca dissiliensis (Sm.) Bréb. the sheath shows under reduced illumi-

nation striae radiating from a zone around the ends of each cell.

HYALOTHECA DISSILIENSIS (Sm.) Bréb.

Showing sheath stained with Methylene Blue and Picric Acid. Magnification 415 diameters.

Photomicrograph with 100 watt condensed filament lamp, Wratten K; and B screens, yg"

Objective, X10 Ocular, field and sub-stage condensers.

The usual methods of staining algal cells, when applied in this case

with the hope of more clearly demonstrating the structure of this sheath,

caused much distortion. In the summer of 1919 at the Marine Biological

Laboratory, Woods Hole, Massachusetts, the writer worked out the

following method for the use of the students, and as it has been tried out

on subsequent occasions with uniformly satisfactory results, it is offered

1In this respect see Liitkemiiller, J., Die Zellmembran der Desmidiaceen. Beitrage zur

Biologie der Pflanzen, (Cohn), 8:347-414. 1902.

DEPARTMENT OF METHODS 25

as being suited for use with classes. The great abundance in which Hya-

lotheca dissiliensis (Sm.) Bréb. often occurs makes it peculiarly convenient,

but the method is no doubt adaptable for use with other forms.

Fresh living material is placed in a .05% aqueous solution of Methylene

Blue for 45 to 60 seconds. It is then removed, rinsed in distilled water and

placed in a zo saturated aqueous solution of Picric Acid. This serves to

fix the stain and brings out in a most striking manner the striations in the

sheath. The material may be examined in the Picric Acid solution, or

removed after a minute or two to water. Preparations are best used soon

after staining, as the sheath begins to disintegrate after a few hours.

Wn. RANDOLPH TAYLOR.

Botanical Laboratory,

University of Pennsylvania

TRANSACTIONS

OF THE

AMC ELCalt

Microscopical Society

ORGANIZED 1878 INCORPORATED 1891

PUBLISHED QUARTERLY

BY THE SOCIETY

EDITED BY THE SECRETARY

PAVE Ss Wit CH

ANN ARBOR, MICHIGAN

VOLUME XL

NUMBER THREE

Entered as Second-class Matter August 13, 1918, at the Post-offce at Menasha,

Wisconsin, under Act of March 3, 1879. Acceptance for mailing at the

special rate of postage provided for in Section 1103, of the

Act of October 3, 1917, authorized Oct. 21, 1918

Che Collegiate Press

GEORGE BANTA PUBLISHING COMPANY

MENASHA, WISCONSIN

1921

TABLE OF CONTENTS

For VoLtuME XL, Number 3, July, 1921

Observations on the Distribution and Life History of Cephalobium microbivorum Cobb

and of its Host, Gryllus assimilis Fabricius, with one plate and three figures, by J. E.

Ackert and Ph Men Wadley. mice oo See ts is ce wer mode ears oni eee 97

A Sarcophagid Parasite of the Common Field Cricket, by C. A. Herrick............. 116

Fresh Water and Marine Gymnostominan Infusoria, with four plates and three figures,

lon JE Aaledls bab iorynnlGee oe Grn oe minan RaSme Moan rr ociabid Sc imcaigMingacaol alc 22: 118

Copper: Its Occurrence and Role in Insects and Other Animals, by R. A. Muttkowski.. 144

DEPARTMENT OF MetuHops, REVIEWS, ABSTRACTS, AND BRIEFER ARTICLES

Microscope Illumination with Reference to Brownian Movement and Combina-

tonglaightings byaAe Silverman ers] oc ie eiari ele tie ile eee 158

B1,

| TRANSACTIONS

| OF

American Microscopical Society

(Published in Quarterly Instalments)

Vol. XL JULY, 1921 No. 3

OBSERVATIONS ON THE DISTRIBUTION AND LIFE HISTORY

OF CEPHALOBIUM MICROBIVORUM COBB AND OF ITS

HOST, GRY LLUS ASSIMTIEIS FABRICIUS

James E. AcKErRT AND F. M. WADLEY

CONTENTS

LPI RTGIEASLTOL, & scp aha oyster tote sche a So cae CRE Pee ae seh REST SER Eee sg Se eal eos SW Si. 98

ibevearasite, Cephalooiwm microuiorum Cobb....:.......+.s-.--sss 5-5-8) ene 98

DES ESET Set nrg, Sia ae th RS ere A a ce ena ee eT SIE 98

TEER DSUPANE .g Ssh Fh Sta OMI eg OOo OUST ea Ct Ne TR Tk rg 100

NVPEENO sO METOGEMUTE RM Meyeiess aa.ets tisrae SI ShaceEe Semmes Gans aT a ince 101

Removalrote Nematodestin.- seach cries ae ees a5. Site tats ciiohis. crete eae 101

CulimnarcrotNematodesa chance ai 2 serie rere rss ote ne a he cots on PAE RE 101

Misch aWOusTOnel) evelOpmety ry pe eee Aen oy as ects oe ew ea eae Ce et eh eee 102

HanlvaGleavaceto: Coiled Eimbryor «a..ace nea. 2 ss nek otis os oe eee ae 102

1 iE] TYE, 5 Basics MRE Soc SPST yp OP, PA PR eg ke eae eee 103

MUTE NC PILLS) COSSTIIUTLIEST Ais oe ops awe occ AeA ats, ee ee ae aS he eet hae teh Steers 104

IS EET OMUIOMEM es ance ato A reser aye cl nreene ees iat ute eve Mes ae ease eee 104

LALA DIIUUS) oh Gre rect hh pec Jere barl SRRSO SCTE ICR Sede BO ot a as ee 104

_ HIG BSI NTS, es ae te detali eloyer a aS SSE Us BE Umi te i a 105

Proportion or Male and) Female) @rickets invNature: 7.22.5. 20 06-2. 0-5--. eae. 105

Distribution of Cephalobiwm microbiworwm. 2... 6... oo oe ee ee ede es 105

Ceomtapiical WusbribWwelouiey.nt-..- cate oh eas Nok om Sees eg Sk he PY vised a Oats 105

Distibutionmnrthes CrickettHostsime sro yee ease 6a aioe faa clte soe eee 106

IPTOPOLMGn Gl Sexes ands barEneaOPenesismmm amr ite aslo Senn teats) eee 107

Batection NematodesonvblOSttrs.cs pte ene trtee te ie Seas itch sc dyofsd aichowe easvate eats ee 107

Beit eeLiStony. Ol Gephi onium microvluoriiminer | a-nation eae ool iss ea eee ee 108

1 LAU COS 10) paar enete ee ebaline ttet aaNet A fit, cops oR ah HEN Dac he oR cea eee ed 108

Beasonale Hn CUraM Ces shee eye arnt anaes tea A Seay iy, Ci tens ain Regret 109

Wrbermearasitesiol the Crickets ys fe cesn gets ee ec eee Pe hoe ad ween ee 109

SITEPLTTE 7 ant comers Mae, Fae Ghar Ponge fe sect hte DQ NA tc Bie ee dene aR meal ord tale 110

(LTS eee Cal OT cee ER on gE RNR es Ae a Bon a PAA ie ays he an olan ue tay ated 2 112

Bxpanation Ol Plate. > 2 .:/sisssth tas shee cee Wee Ted cL gaw TRS pales ClSee ake LS ree 114

1 Contribution No. 52 from the Zoology Department, Agricultural Experiment Station

of the Kansas State Agricultural College.

98 JAMES E. ACKERT AND F. M. WADLEY

INTRODUCTION

While securing gregarines from black field crickets for class use, in

October, 1918, the senior writer found heavy infestations of small nema-

todes, some of which were sent to Dr. N. A. Cobb for identification.

Determining that these nematodes represent a new genus, Doctor Cobb

suggested that studies be made on the distribution and life history of it.

Observations on its distribution have been made at Woods Hole, Mass.;

Falls Church, Va.; Douglas Lake, Mich.; Rockford, Ill.; and Manhattan,

Kan. ‘The studies on its life history and that of the crickets were made at

Manhattan. Further work on certain phases of these studies would be

very desirable, but as this cannot be done for some time, it seems best to

put the present findings on record. The writers wish to express their

indebtedness to Doctor Cobb for suggesting this nematode study, and to

Director Frank R. Lillie of the Marine Biological Laboratory, Woods Hole,

Mass., and Director George R. LaRue of the University of Michigan Bio-

logical Station for the privilege of using equipment at the respective stations.

THE PARASITE, CEPHALOBIUM MICROBIVORUM COBB

DESCRIPTION

These nematodes which are from 2 to 3 mm. in length were identified

as Cephalobium microbivorum n. g.,n. sp., by Dr. N. A. Cobb, who submits

this description.

The following characterizations and description, with figures, are taken from “‘Contribu-

tion to a Science of Nematology,” No. IX; “One Hundred New Nemas,” N. A. Cobb, 1920.

The characters other than specific are assembled from Cobb’s Keys.

Pxuyitum Nemates

SUBPHYLUM Laimia: Nemas having a more or less distinct pharynx.

Crass Anonchia: Nemas lacking onchia.

Supcriass Anodontia: Nemas lacking odontia.

OrDER Polylaimia: Nemas having an unarmed pharynx, composed of two or

more successive chambers more or less distinctly separated

from each other.

Genus Cephalobium

Cavity of the pharynx more or less prismoid or cylindroid (not conoid or very irregular),

and containing a glottoid organ at its base. Oesophagus with median bulb and posterior

swelling. Amphids none so far as known. Seta-like labial papillae 6. Single lateral wing

present; striae fine, plain. Spinneret absent.

Preanal and postanal papillae present on the male. Tail conoid or subconoid; terminus

acute, unarmed. Bursa none. Spicula two, equal, more or less arcuate; not jointed; their

width not uniform.’ Accessories (gubernaculum) present. Inner ends of spicula cephalated

by constriction. Length of the spicula 114 times as great as anal body diameter.

54. Cephalobium microbivorum, n. sp. The single wing begins near the head and ends

near the terminus. Its optical expression is either a pair of lines or a single line in the middle

of a field one-twelfth as wide as the body. The contour of the body may become crenate in

the anal region. There are about thirteen lateral organs on each side connected with pores

- CEPHALOBIUM MICROBIVORUM COBB 99

in the cuticle (see org. lat. fig. 54). Base of the pharynx containing a large, complicated and

peculiar dorsal glottoid organ (see fig. 54). No amphids. The rather thin-walled intestine

is set off by a collum one-eighth as wide

as the neck, and has a rather distinct

lumen. It becomes at once five-sixths

as wide as the body, and in cross section

presents two to four cells. From the

somewhat depressed anus, the narrow,

cutinized rectum extends inward and

forward a distance one and one-fourth

times as great as the anal body-diameter.

Scattered yellowish granules of variable

size occur in the cells of the intestine, the

wsth]

. Seg hl meh HGH (G)_ >a sm

msc org ylot — aldct dmoe seg ene murph al dt SCC past Ah x 750

Il pi)

ary yfat

largest being one twenty-fifth as wide as the body; in addition, there are

numerous very small granules. There isno tessellated effect. Subarcu-

ate, conoid tail tapers from in front of the anus to the acute, fine terminus.

There is no spinneret. From the elevated vulva, the rather small,

somewhat weak vagina extends inward nearly at right angles to the

ventral surface one-fourth the way across the body. Ajiong the middle

half of the body the two equal uteri contain thin-shelled, smooth, —

ellipsoidal eggs two-thirds as long as the body is wide, and appear

to be deposited after segmentation begins. No embryos were seen in

these eggs, only blastulas. For the most part®the ova are arranged

irregularly in the somewhat tapering ovaries. The two equal, rather

strong, slender, arcuate, tapering subacute spicula are one and one-

fourth times as long as the anal body-diameter, are more or less

cephalated by constriction and when seen in profile have proximal ends

nearly opposite the body-axis. Toward their distal ends four slender

stiffening pieces are apparent. There is a simple, strong, and rather

solid, straight accessory piece, one-third as long as the spicula,

bending back from them at an angle of about 90 degrees, so that its prox-

imal end lies opposite the body-diameter. There is no bursa. Near the

beginning of the second quarter of the tail there is a pair of lateral pores

similar to those on the female. Beginning just in front of the anus there

exists on the tail a series of six submedian pairs of flattish-conoid, rather

inconspicuous papillae. These occupy the anterior two-thirds of the tail,

and have a formula as follows: 1 ( ) 1; 111; 1. The members of the

posterior four pairs are located exactly opposite each other, the right

hand member of each pair being slightly behind the left hand member.

Papillae plainly enervated. Spicula conspicuous, rather close together;

at the widest part about one-eighth as wide as the corresponding portion

of the body, ending in minute ‘“‘buttons.’”’ The lateral pores on the tail

are the final members of the lateral

Ds 01023 02th ys AB IRB series of lateral organs.

Habitat: Intestine of the field

cricket, Gryllus assimilis Fabr. The

males are considerably smaller than

the adult gravid females.

100 JAMES E. ACKERT AND F. M. WADLEY

HABITAT

To ascertain the part of the cricket inhabited by C. microbivorum some

crickets were carefully dissected. In each case the thin-walled ileum

Fic. 1

Figure 1. Showing digestive tract of Gryllus

assimilis. c, colon; cr, crop; g.c., gastric caecum;

i, ileum; m.t., malpighian tubes; 0, oesophagus;

p, proventriculus; 7, rectum; s, stomach. X 4.

readily revealed the writhing

nematodes which appeared in

bold contrast to the dark fecal

contents. None of these parasites

was found in the coelom, nor in

any organ outside of the digestive

tract. In one case two dead adults

were taken from the colon of a

freshly dissected cricket, whose

ileum contained several live speci-

mens. To facilitate subsequent

discussion a brief description and

a diagram are given of the alimen-

tary canal of the cricket host.

The digestive tract (fig. 1) of

this black field cricket, Gryllus

assimilis, bears many resem-

blances to that of the large and

nearly wingless western cricket,

Anabrus, as shown by Packard

(1878-79, p. 175). The oesophagus

(o) connects with the mouth and

after making a sharp bend pro-

ceeds through the posterior part

of the head to open into the

spacious crop (cr) which occupies

the thoracic and anterior abdom-

inal portion of the coelom. After

greatly narrowing, the crop opens

into the strong proventriculus (p)

which is much larger than the

corresponding organ in Anabrus.

InG. assimilis the diameter of this

organ exceeds three times that of

the junction with the crop,

whereas in Anabrus the proven-

triculus is considerably reduced

in size.

CEPHALOBIUM MICROBIVORUM COBB 101

The proventriculus opens by a very narrow canal into the true stomach

(s) which immediately gives off anteriorly two large, flattened gastric

caeca (g.c. ), situated one above the other. The stomach is surprisingly

slender. After passing backward a short distance it makes an abrupt

turn upward, narrows slightly and terminates, giving off numerous Mal-

pighian tubes (m. t.) where it joins the intestine. Like Amabrus the re-

mainder of the digestive tract is distinctly divisible into three portions:

the ileum, colon, and rectum. Unlike the large western species, however,

the stomach of G. assimilis is considerably shorter and the ileum (i) longer

and much more capacious. The walls of the stomach are thick and

translucent, while those of the ileum are thin and transparent. So thin

are the walls of the latter that not only are the enclosed, motile nematodes

visible, but also the eggs in the females’ bodies. Before terminating, the

ileum narrows considerably, makes a pronounced twist and then opens

into the larger, thick-walled colon (c). Continuing posteriorly, the colon

with a slight constriction connects with the still larger rectum (r) with

which the anus communicates.

METHODS OF PROCEDURE

REMOVAL OF NEMATODES

The principal method employed for the removal of the nematodes is

here briefly described: After excising the head of the cricket an incision

was made in the posteroventral wall of the abdomen with fine scissors.

By cutting forward through the median ventral surface, being careful not

to cut deeply, nearly the whole lower body wall could be laid open without

disturbing the digestive tract. On inserting fine forceps into the meso-

thoracic region the crop could be seized, and with slow, sustained effort the .

entire digestive tract withdrawn. In this extended condition the food

tube was placed upon an ordinary glass slide, the rectum being excised at

the anus.. After covering with a few drops of normal salt solution, the

intestine was teased with needles. The optical examinations were made

with the aid of dissecting, binocular, and compound microscopes.

CULTURING OF NEMATODES

The culturing of the nematodes and their eggs was carried on for a time

with fair success, but this phase of the problem should be continued.

Having the nematodes on regular microscopic slides made it easy to remove

the substances not wanted in the culture, and to add materials desired.

With a little care the culture could be held to arestricted area on the slide

by the surface tension of the solution. To prevent drying, the culture

slides were placed in Petri dishes containing a few drops of distilled water.

Subsequent microscopic examinations of the cultures were made either in

or out of these small moist chambers.

102 JAMES E. ACKERT AND F. M. WADLEY

Culture fluids used included normal salt solution alone, with fecal

material, with peptone, and with both fecal material and peptone. Eggs

hatched in each fluid, but growth of young nematodes appeared to occur |

only when a few drops of 0.8% peptone (in distilled water) were added.

Dilute peptone was one of the successful solutions used by Welch and

Wehrle (1918, p. 151) in their extensive nematode cultures.

In normal saline solution adult nematodes lived from one to six days,

eggs developed and hatched, but even vigorous free embryos failed to

increase in length and died within three days. When a few drops of 0.8%

peptone were added to normal saline, embryos hatched and lived six days.

The best results were obtained with equal volumes of normal saline and

0.8% peptone solution and a trace of cricket feces. In this medium several

free embryos lived eight days, and a few thirteen days, the latter increasing

their body lengths 1624 per cent. However, before the young nematodes

had developed markedly, the culturing had to be abandoned on account

of failure to secure nematodes. The adult crickets which had been col-

lected prior to a cold wave succumbed in a few days, thus destroying

the source of supply.

OBSERVATIONS ON DEVELOPMENT

EARLY CLEAVAGE TO COILED EMBRYO

The observations on development were upon living material, no attempt

deing made to trace the formation of germ layers or organs. ‘The trans-

lucency of the dividing cells made it possible to follow the individual

blastomeres until they formed a more or less spherical mass, such as shown

in fig. 6. Whether such developing forms as represented in figs. 6 and 7

were hollow is uncertain, owing to the growing opacity of the embryos.

For convenience in description the terminology of Martin (1913) is partly

followed. For the uncertain stages shown in figs. 6 and 7, a morula

rather than the morula is used. Likewise, when the vermiform shape of

the embryo is first attained (figs. 8, 9) the term curved embryos is employed,

whereas, the fully attenuated enclosed embryo (fig. 11) is designated as a

coiled embryo.

These few observations on the development of the external form of C.

microbiverum are given in the hope that someone may find opportunity

to work out the embryology of this nematode. Such a study would be

valuable and would be greatly facilitated by the thin, elastic egg shells.

In the present studies fertilized eggs, which can be distinguished by

their clear nuclei, were mounted under cover slips in normal saline solution

and studied under low and high powers of the microscope, a few drops of

distilled water being added occasionally to compensate for evaporation.

In other cases the live females containing eggs were so mounted, and in

CEPHALOBIUM MIGROBIVORUM COBB 103

this way external development was traced from the fertilized egg through

hatching.

Under these conditions the early cleavage stages, as shown in figs. 2

to 5, develop somewhat rapidly, each cell division occurring in from ten

to sixteen minutes. As segmentation proceeds and the bulk of the embryo

increases, the thin, elastic shell expands accordingly. Eggs in an early

cleavage stage (fig. 5) at 5 p.m. were ina morula stage with large blasto-

meres (fig.6) at 8p.m. Eighteen hours afterward development had pro-

ceeded to a morula with small blastomeres (fig. 7), in twenty more hours

to a curved embryo (fig. 9), and six hours later to a coiled, motile embryo

(fig. 11).

HATCHING

The stages represented in figs. 2-11 inclusive were observed in the uteri

living worms mounted in normal salt solution under cover slips. Hatch-

ing occurred only after the eggs were ejected. Emergence from the egg

was accomplished by repeated thrusts of the anterior end of the embryo

against the thin shell which soon began to give way (fig. 12), finally ruptur-

ing (fig. 13) and liberating the writhing embryo.

FACTOR AFFECTING DEVELOPMENT

As noted above, eggs in nematodes which were in culture media devel-

oped somewhat rapidly, attaining the coiled, motile stage in approximately

two days. But in the body of the live host uterine eggs do not appear to

to develop beyond the four-cell stage, as numerous examinations have

shown. However, such eggs in a dead and somewhat macerated cricket

were in more advanced stages of development. Also in the voided feces

of an infested cricket, eggs in a morula stage were found. Thus it appears

that the failure of uterine eggs to develop beyond the four-cell stage in the

living cricket is due to an inhibiting factor. This factor the writers believe

to be lack of sufficient oxygen. In the higher animals it is well known

that the intestines contain enormous numbers of bacteria which must take

much of the free oxygen, especially in the colon and adjacent regions where

anaerobic bacteria thrive. In the ileums of these crickets there were

numerous bacteria. The cases of advanced development of uterine eggs

of the nematodes in the macerated cricket would favor this interpretation,

as the thin wall of the disintegrating ileum quickly ruptures, admitting

oxygen. Likewise, eggs which developed up to hatching in the uteri of

cultured nematodes would be in the presence of more oxygen than when

in the ileum of the cricket. An ample supply of oxygen to eggs in the

separate fecal pellets is obvious. That additional oxygen accelerates

development in nematode eggs has been determined by the senior writer

104 JAMES E. ACKERT AND F. M. WADLEY

who observed more rapid development in cultured (normal saline) eggs of

Ascaridia pers picillum on the addition of 10% hydrogen peroxide (unpub-

lished results).

Tue Host, GRYLLUS ASSIMILIS

DISTRIBUTION

In October, 1919, when the cricket examinations were at their height

some adults were sent to Mr. James A. G. Rehn, who identified them

as Gryllus assimilis Fabricius. Rehn and Hebard (1915, pp. 295, 296)

regard this species as the common black field cricket of the Americas,

which ranges from Canada to Argentina and from the Atlantic to the

Pacific. .

In the vicinity of Manhattan there are two races of this species, one

maturing in August and September, the other in Apriland May. The fall

adults lay their eggs in October, depositing them in the soil, under stones,

and in other protected places. These eggs hatch the following spring

according to Bruner (1886, p. 194), and the young mature in August and

September. Concerning the occurrence and behavior of the spring adults,

a few notes from the senior author’s records for another problem are given,

“About the last of May, 1915, adult crickets were found in nature mating.

Several pairs of these were placed in cages containing sterilized earth, some

carefully selected stems of alfalfa, and a small block of wood. Care was

taken not to introduce any other animals.” These records show that

every second day the caged crickets were given one of the following foods:

green alfalfa, fresh apple, algae, and small bits of fresh beef. Young

crickets hatched in three weeks. By October the nymphs were approxi-

mately half-grown, averaging one-half the length of the adults.

From these and other observations it appears that in the vicinity of

Manhattan, Kan., G. assimilis produces only one brood per year, but is

represented by two races, the fall adults, laying their eggs in the autumn,

passing the winter in this stage, hatching in the spring, and maturing in late

summer or autumn; and the spring adults, depositing their eggs in the

spring, hatching in early summer, passing the winter in a nymphal stage,

and maturing the following spring. These respective findings are in close

accord with the observations of McNeill (1891, p. 5) for G. abbreviatus in

Illinois and of Blatchley (1901, p. 439) for G. pennsylvanicus in Indiana.

HABITS

Concerning the habits of the common black cricket, Blatchley (1901, p.

436) states that it is nocturnal, omnivorous and cannibalistic. The present

studies indicate that these crickets in nature are largely nocturnal, but

that they may stridulate, move about, and feed to some extent in the day

time.

+ CEPHALOBIUM MICROBIVORUM COBB 105

That they are omnivorous in nature is amply confirmed by these obser-

vations. Plants on which they have been seen feeding include alfalfa

(Medicago sativa), bluegrass (Poa pratensis), bindweed (Convolvulus spp.),

crabgrass (Syntherisma sanguinale), and Bermuda grass (Capriola dactylon).

Decomposing plant and animal tissues appear not to be distasteful, as the

crickets have been seen feeding on both. Portions of dead crickets and

other arthropods have been taken in preference to wilted grass, and in a

few instances the animal tissues were in an advanced stage of decomposi-

tion.

Cannibalism is of frequent occurrence among the common black

crickets, but apparently they seldom attack each other in life. In ordi-

nary captivity mortality is high; some of the captives usually survive and

frequently feed upon their deceased mates. The senior writer in connec-

tion with another problem reared crickets from eggs in large life history

cages, making almost daily observations for months. Crickets were often

seen dying, and sooner or later others began to devour them. Inasingle

instance, a live cricket was observed to approach a dying one, lying on its

back, and begin feeding on the latter’s hindfemur. Atno other time, either

in 1915-16 or during the present studies, has one cricket been seen to feed

upon another living one.

HABITATS

The wide distribution of G. assimilis.is doubtless due in part to its

omnivorous feeding habits and to its varied habitats. Among the habi-

tats from which it has been taken are the following: at edges of side walks,

in holes in the ground and chinks in walls of buildings, under old hardened

ox feces, sticks, boards, logs, stones, and stone walls, and among various

kinds of vegetation.

Besides proximity to food and a reasonable amount of protection the

diurnal habitat of this cricket must afford an atmosphere of comparatively

high humidity. In artificial rearing the mortality was exceedingly high

until water was sprinkled into the cages, when the percentage of survivals

markedly increased.

PROPORTION OF MALE AND FEMALE CRICKETS IN NATURE

The writers found the number of male and female crickets to be approxi-

mately equal in nature, except during late October after the breeding season

is over. At this time the adult females, with their abdomens distended

with eggs, far outnumbered the surviving adult males.

DisTRIBUTION OF CEPHALOBIUM MICROBIVORUM

GEOGRAPHICAL DISTRIBUTION

Examinations of black field crickets for C. microbivorum have been

made in five states: Kansas, Massachusetts, Virginia, Michigan, and

106 * JAMES E. ACKERT AND F. M. WADLEY

Illinois, but to date these nematodes have been found only in Kansas and

Virginia. From April to June, 1919, Dr. N. A. Cobb examined a few black

field crickets at Falls Church, Va., and found C. microbivorum in nearly

every cricket. At Manhattan, Kan., these nematodes are known to have

been of common occurrence in the adult black crickets during the autumns

of the last three years (1918, 1919, 1920).

DISTRIBUTION IN THE CRICKET HOSTS

A study of the data collected during the search for C. microbivorum

in the local black field crickets reveals some interesting points in the

distribution of these nematodes in their hosts. Most of the examinations

of the crickets were made during the periods between September 19 and

October 31, in 1919 and 1920. From Table I it is seen that the number of

female crickets examined exceeds that of the males. This was due to

certain collections madé late in October after the breeding season and after

the consequent heavy mortality of male crickets. Collections made in

September included nearly equal numbers of males and females.

TasLe I. SHowING NEMATODE INFESTATION OF ADULT CRICKETS EXAMINED BETWEEN

SEPTEMBER 19 AND OcTOBER 31, 1919 anp 1920, ar MANHATTAN, KAN.

Pe erred ise Nes : : ____| Average In- | -

|No. C rickets No. Crickets Per cent Total No. \festation per | Range oO

Examined Infested Infested | Nematodes ee Infestation

Male

Crickets 14 10 71.4 217 | lei 2 tonsil

Female

Crickets 33 30 1; 9029) 822 27.4 3 to 91

All |

Crickets AT 40 85.1 1,029 Dyaih 2 to 91

1 | { |

From Table I it is seen that over eighty-five per cent of the crickets

examined by the writers between September 19 and October 31 were

infested with this nematode. Approximately seventy per cent of the males

and ninety per cent of the females contained these parasites. Table I

likewise shows that both the range and average infestation of the females

exceed that of the males. That the intestine of the male cricket furnishes a

suitable environment for these parasites is evident from infestations

amounting to as many as fifty-one C. microbivorum. Consequently the

explanation of these phenomena must be sought elsewhere. In October

the females’ bodies are usually gorged with eggs, and are larger than those

of the males. Obviously, to afford this greater development, more food

CEPHALOBIUM MICROBIVORUM COBB 107

would be required than for the males, thus increasing the chances of the

females ingesting a larger number of nematode eggs or larvae. The

large, distensible crop (fig. 1, cr) is adapted for receiving quantities of food,

and the numerous fecal pellets voided daily by these females are evidences

of large appetites. Thus, to the writers, the most plausible explanation of

the higher percentage, average, and range of nematode infestations in the

females is that the engorged females take more food, and thus, on the

average, swallow more eggs or larvae.

Occasional examinations of three or four specimens of G. assimilis were

made during June, August, and September, 1920. No specimens of C.

microbivorum were found in any of these crickets until August 21 when

one was taken from an adult female. Of six mature crickets—three males

and three females—examined on this date, two of the females contained

nematodes, the other infestation consisting of two immature specimens.

In September the recorded infestations ranged from seven to thirty-one

nematodes, and in October from seven to as many as ninety-one.

In addition to the examinations of adult crickets shown in Table I,

some half-grown black crickets were dissected on November 3, 1920.

In the ileum of one of these nymphs were two mature specimens of C.

microbivorum. The significance of this observation will be discussed later.

PROPORTION OF SEXES AND PARTHENOGENESIS

The nematodes observed included noticeably more females than males,

and in one case males were entirely lacking. This case will be discussed

presently, but concerning the presence of more female nematodes than

males, the writers have no explanation to offer. Merrill and Ford (1916,

p. 127) likewise found the females more numerous in the two species of

nematodes they studied.

In the case just mentioned in which males were lacking, the infestation

consisted of three females each containing fertilized eggs. These females

may have been fecundated by males which had already left the host, as two

dead nematodes were found in the colon of a cricket immediately after

killing. Or, the females in question may have been parthenogenetic as

Welch and Wehrle (1918, p. 159) and others have observed in small

nematodes. It is possible also that they may have been protandric

hermaphrodites, but the almost constant occurrence of males in the

infestations favors the view that the females were probably fecundated

by males which subsequently passed from the cricket.

EFFECT OF NEMATODE ON Host

The effect of C. microbivorum on the host does not appear to be serious,

as apparently normal crickets often harbored thirty or more of these

nematodes. On the other hand, it seems probable that so many compar-

108 JAMES E. ACKERT AND F. M. WADLEY

atively large entozoa must be detrimental to the host. Flury (1912) has

shown in the cases of nematodes parasitic in higher animals that they

cause injury not only by taking food material and by stoppage, but that

on account of their imperfect digestive system their excreta contain toxins

which are absorbed by the host. Some of these injuries would be likely

to occur in the infested crickets.

Lire History oF CEPHALOBIUM MICROBIVORUM

LIFE CYCLE

As stated elsewhere, C. microbivorum matures in the intestine of the field

cricket and eggs normally develop to the four-cell stage. When such eggs

are ejected and kept moist development proceeds to the coiled, motile

embryo stage in about two days. Cricket feces voided during the night

and examined the following morning contained eggs of C. microbivorum

in a morula stage. To ascertain the probable fate of such eggs in nature,

four adult female crickets were placed in a lantern globe cage over some

moist, sterilized earth. They lived from four to six days, the dead ones

being removed before they were attacked by their mates. Two of these

crickets were subsequently found to be infested with mature female

nematodes containing eggs. The earth in the cage was moistened nearly

every day, but as it was kept uncovered in the laboratory, evaporation was

rapid and the culture dried out several times. Ten days after the infested

crickets were placed over the sterilized earth examinations of portions of

the latter were made which revealed two dead larvae, slightly larger than

newly hatched embryos.

These observations indicate that in nature the eggs pass from the body

of the cricket in early cleavage stages, and since the diurnal habitat of the

cricket must have an atmosphere of comparatively high humidity, as

shown elsewhere, a fair percentage of the nematode eggs voided in the

daytime would be protected by the humidity of the cricket habitat. Since

in culturing, these nematode eggs hatched in each separate medium, it is

logical to infer that they would hatch in the moist débris of a cricket

habitat. The thin elastic shell which bursts after a few thrusts would

probably not long confine the embryos. Another reason which leads the

writers to think that these young nematodes pass a period free in nature

is that while immature stages of C. microbivorum are found in the ileum

of the cricket, these nematode larvae are always much larger than the

newly hatched embryos. If infection were caused by the cricket’s inges-

tion of nematode eggs, one would expect occasionally to find in the crickets

young nematodes the size of newly hatched ones, but this has failed to

occur in the removal of over one thousand of these nematodes. The

writers made no attempt to infest crickets by giving them hatched embryos

or free larvae, but Merrill and Ford (1916, p. 127) succeeded in infecting

termites with free larvae of the nematode, Diplogaster aerivora Cobb.

CEPHALOBIUM MICROBIVORUM COBB 109

The indiscriminate feeding habits of the crickets would give ample

opportunity for the ingestion of larvae of C. microbivorum, for as stated

elsewhere, they feed upon decomposing plant and animal tissues, and these

substances are commonly found in the moist diurnal habitats. Once in the

ileum of the cricket the young C. microbivorum evidently thrive, for they

occur in active stages ranging from one-third to normal lengths.

SEASONAL ENDURANCE

In the light of the information available, this nematode’s problem of

enduring the seasons seems relatively simple. It has been noted that in

the vicinity of Manhattan, Kan., two races of G. assimilis occur, one

spending the winter in the egg stage, and the other in the nymphal stage.

The finding in November of mature specimens of C. microbivorum in nym-

phal black field crickets which live over the winter here and elsewhere in

protected places indicates one way in which the cold season may be endured.

These nymphal crickets mature in May, and deposit their eggs early in

June, thus making it possible to shelter these adult parasites until summer.”

By this time the eggs of the other race are hatched, furnishing possible

hosts for the young C. microbivorum liberated as eggs from the winter-

enduring nymphal crickets. Protection of the larvae against sudden

desiccation and high temperatures would be afforded deeper in the habitat.

At any rate, the fact that the fall infestations are the heaviest indicates

that this nematode’s problem of enduring the summer is not a serious one.

Another possible means of C. microbivorum enduring the winter might be

afforded by the habit of the fall adult crickets crawling into the ground

and into other protected places when their life work is finished. If such

infested crickets were killed by freezing and remained congealed through-

out the winter, the spring season might be well advanced before maceration

of the crickets’ bodies proceeded far enough to admit sufficient oxygen

for the development of the enclosed nematode eggs. The plausibility of

this method is strengthened by the fact that many nematodes can with-

stand some freezing. It is possible, of course, that larvae of these nema-

todes pass the winter free in the soil. Further studies in connection with

the culturing of C. microbivorum will doubtless settle this and other points

in its life problems.

OTHER PARASITES OF THE CRICKET

While searching for C. microbivorum in the crickets certain other

parasites were encountered; viz., gregarines, gordiacea larvae and dip-

terous larvae. In July, 1919, gregarines were present in many of the

common black crickets examined at Woods Hole, Mass. At Douglas

° The writers made no examinations of the spring adults, but Doctor Cobb’s examinations

from April to June were of adults of this race in nearly all of which he found C. microbivorum.

110 JAMES E. ACKERT AND F. M. WADLEY

Lake, Mich., these protozoan parasites were of frequent occurrence in

crickets in August, 1920, and larval gordiacea were occasionally found.

Of the 106 mature or nearly mature crickets examined gregarines were

found in thirty-seven per cent, and gordiacea larvae in 9 per cent of them.

The crickets were taken from four localities; viz., Douglas Lake shore line

about seventy-five yards wide; Burt Lake shore line approximately ten

yards in width; Sedge Pool shore line about eight yards in width; and an

upland pasture one and one-half miles from water.

The only gordiacea infestations occurred in the Sedge Pool locality,

which was also the most favorable for gregarine infestations. This pool,

which is separated from Douglas Lake by a narrow ledge, is approximately

200 feet long by 120 feet wide. It is protected by two to four foot banks

and by a substantial growth of timber on three sides, leaving the east and

southeast sides open to the direct rays of the sun. Most of the crickets

were taken on the narrow ledge at the east side of the pool within ten

to twenty feet of the protected water’s edge. The infested crickets con-

tained gordiacea larvae in later stages of development, some of them having

attained the dark adult coloration. Two that escaped from a cricket in a

bottle of water were readily identified as Paragordius varius (Leidy).

This was the only species obtained by May (1919), who examined several

hundred crickets from the east shore of Douglas Lake in connection with

his studies on the life history of this species. The lowest percentage (30

per cent) of gregarine infestation was in the upland pasture one and one-

half miles from Douglas Lake, while the highest (45 per cent) occurred at

Sedge Pool. This indicates that these protozoan parasites thrive better

under moister conditions.

Records for gregarine infestation of the crickets examined at Man-

hattan, Kan., are not complete, but of twenty crickets taken from nature

between June 24 and October 21, 1920, eleven, or fifty-five per cent of

them, were infested. The infestations ranged from one to as high as 517

gregarines, the average being slightly over sixty-two. Not a specimen of

Paragordius was found here, but from two crickets examined by Herrick

(1921) a few sarcophagid larvae were taken, this apparently being a new

case of parasitism in G. assimilis.

SUMMARY

1. In the autumns of 1918, 1919, 1920 black field crickets, in the

vicinity of Manhattan, Kan., were infested with a new species of nematode

which has been identified as Cephalobium microbivorum Cobb.

2. In the body of the living cricket development of the eggs has not

been observed to exceed the four-cell stage. This is attributed to lack of

sufficient oxygen.

CEPHALOBIUM MICROBIVORUM COBB 111

3. In culturing, eggs hatched in all moist media used, but young nema-

todes grew only when 0.8% peptone was added, two specimens increasing

their lengths 1624 per cent in thirteen days. In early cleavage, each cell

division was accomplished in from ten to sixteen minutes; and in approxi-

mately two days the embryo was fully formed. Hatching is accomplished

by repeated thrusts of the anterior end of the embryo against the thin

elastic shell which soon ruptures and liberates the embryo.

4. The parasitic habitat of this nematode is the spacious ileum of

Gryllus assimilis Fabricius, which is the common black field cricket of the

Americas, ranging from Canada to Argentina and from the Atlantic to

the Pacific. In the vicinity of Manhattan, Kan., this species is represented

by two races, one maturing in August and September, the other in April

and May; each race produces one brood of crickets per year. The race

maturing in the spring winters here in the nymphal stage, while the adult

fall race spends the cold season in the egg stage. Eggs of G. assimilis

hatched (June) in three weeks after deposition. Late in October they were

half grown, and by the last of the following May they were mature and

mating.

5. These crickets are omnivorous, feeding on various kinds of plant

and animal tissues, both fresh and decomposed. Cannibalism is of common

occurrence among them. They are known to devour their dead and

dying, but not to attack each other in normal condition. Their diurnal

habitats, which may include a variety of situations. must furnish some

protection and sustain a certain amount of moisture.

6. The numbers of adult male and female crickets observed in nature

were about equal, except in the autumn after the breeding season, when

more females survived.

7. Of crickets examined in five states, only those from Kansas and

Virginia have been infested with C. microbivorum. At Manhattan, Kan.,

about 85 per cent of the fall adult crickets examined were infested; 70

per cent of the males and 90 per cent of the females. The females also

contained a larger number of these parasites. Both the higher percentage

of parasitism and the heavier infestations of the females are attributed to

their greater voracity.

8. Infestations of fall adults were first found in August, the parasites

being young andfew. By September adult nematodes were taken and the

size of infestations increased to 31. In October both young and adults

were numerous, a maximum infestation amounting to 91 of these nema-

todes.

9. Nymphal, black field crickets of the race which winters in this stage

were infested with mature nematodes in November.

02 JAMES E. ACKERT AND F. M. WADLEY

10. Female nematodes were more numerous than males; for. this

phenomenon no explanation is offered. One cricket contained 3 female

nematodes, each having fertilized eggs. Death of the males after fecun-

dation is deemed more probable than parthenogenesis or protandric

hermaphroditism.

11. No positive deleterious effect of the parasites on the host was

observed, but this does not preclude possible injury.

12. The life cycle of C. microbivorum appears to be as follows: The

nematode matures in the ileum of the common field cricket. Its eggs are

deposited inearly cleavage stages and passed from the body of the cricket.

Under moist conditions furnished by the diurnal habitat the eggs soon

hatch, and in the presence of nutritive substances the liberated embryos

grow. Sooner or later the larval nematodes are swallowed by the omniv-

orous cricket in whose ileum they mature.

13. The nematode’s problem of enduring the seasons is apparently

solved by the occurrence of the two races of G. assimilis, the winter nymphs

sheltering some of the mature nematodes through the colder months and

the young of the fall adults ingesting larval nematodes during the warmer

ones. ;

This method is probably supplemented by the infested bodies of certain

fall adult crickets which, though dead, pass the winter in a somewhat

congealed condition, the macerating bodies later liberating mature nema-

todes and eggs.

14. Other parasites encountered in the crickets examined included greg-

arines, gordiacea larvae and dipterous larvae. Gregarines were found

generally, Paragordius varius larvae only at Douglas Lake, Mich., and

sarcophagid larvae only at Manhattan, Kan.

LITERATURE CITED

BLATcHLey, W. S. .

1901. Orthoptera of Indiana. Rept. State Geol. Ind., 27:123-471.

BRUNER, L.

1886. Second Contribution to a Knowledge of the Orthoptera of Kansas. Bull. Wash-

burn College, Lab. Nat. Hist., 1:193-200.

Frory, F.

1912. Zur Chemie und Toxikologie der Ascariden. Arch. exper. Path., 62:273-390.

HERRICK, C. A.

1921. A.Sarcophagid Parasite of the Common Field Cricket. Trans. Amer. Micr. Soc.,

40:115-116.

Martin, A.

1913. Recherches sur les Conditions du Developpement Embryonaire des Nematodes

Parasites. Ann. Sci. Nat. (Zool.), Paris (9), 18:1-151.

May, H. G.

1919. Contributions to the life histories of Gordius robustus Leidy and Paragordius varius

(Leidy). Ill. Biol. Monographs, 5:1-118.

CEPHALOBIUM MICROBIVORUM COBB 113

McNE 1, J.

1891. A List of the Orthoptera of Illinois. I. Psyche, 6:3-9.

Merrit, J. H., AND Forp, A. L.

1916. Life History and Habits of Two New Nematodes Parasitic on Insects. Jour. Ag.

Res., 6:115-127.

PACKARD, A. S.

1878-79. The Western Cricket. Second Report U. S. Ent. Commission, Washington,

pp. 163-178.

Resn, J. A. G., AND HEBARD, M.

1915. The Genus Gryllus (Orthoptera) as found in America. Proc. Acad. Nat. Sci.

Phila., 67:293-322.

WEtcu, P. S., AnD WEHRLE, L. P.

1918. Observations on Reproduction in Certain Parthenogenetic and Bisexual Nema-

todes reared in artificial Media. Trans. Amer. Micr. Soc., 37:141-176.

114 JAMES E. ACKERT AND F. M. WADLEY

EXPLANATION OF PLATE

All drawings were made with the aid of a camera lucida and are of the same magnification,

X 400. The figures show stages in the embryological development of the external form of

Cephalobium microbivorum Cobb.

Figs. 2 to 5. Eggs in early cleavage. ‘

Fig. 6. Morula with large blastomeres.

Fig. 7. Morula with small blastomeres.

Figs. 8,9. Eggs containing curved embryos.

Figs. 10, 11. Eggs containing coiled embryos.

Figs. 12, 13. Eggs in process of hatching.

115

CEPHALOBIUM MICROBIVORUM COBB

PLATE IV

A SARCOPHAGID PARASITE OF THE COMMON FIELD CRICKET!

CHESTER A. HERRICK

The purpose of this article is to record what seems to be a new case of

parasitism in the cricket, Gryllus assimilis Fabricius, which, according to

Rehn and Hebard (1915, pp. 295, 296), is the common black field cricket,

not only of Kansas but of America. The cricket hosts of this parasite

were taken in an alfalfa field near a stone wall. This wall and the débris

along the south side of it afforded excellent protection for the congregated

crickets. Numerous dead crickets were in the wall and under the débris

near it and may have attracted the sarcophagids.

While examining these black field crickets for nematodes under the

direction of Dr. J. E. Ackert, two insect larvae were found on Sept. 29,

1920. From the method of examination it is apparent that these larvae

were in the body-cavity of the crickets. After excising the extreme

posterior end of the cricket the thorax and abdomen were gently separated,

and by sustained effort the whole intestine was removed from the abdomen.

While the intestine was being examined the active larval parasites escaped

through the large open end of the abdomen. One larva was 3 mm. in

length, while the other, which was further developed, was 14 mm. long.

The large larva was placed in a small covered tin box where, within 24

hours, it had pupated and become cemented to the floor of the box.

In this container the pupa was kept at laboratory temperatures, which

ranged from 55° to 96° F., with an average temperature of 72° F. On

the nineteenth day after pupation the adult fly emerged. It was sent

to the Bureau of Entomology of the United States Department of Agri-

culture, where Dr. J. M. Aldrich identified it as Sarcophaga kelly: Aldrich.

This sarcophagid was first seen by Kelly (1914), who discovered it asa

parasite of grasshoppers at Wellington, Kan., in mid-summer. His

attention was attracted to certain flies that struck flying grasshoppers and

caused them “to drop to the ground asif shot.’”’ On examining such

grasshoppers he found tiny larvae crawling toward the base of the unfolded

hind wing. Similar observations were subsequently made in New Mexico

by Smith (1915), who found that this fly chose healthy, freshly molted, or

inactive grasshoppers for the deposition of its larvae. He states (p. 8)

that, ‘“The female (Sarcophaga kellyi) upon locating a suitable victim was

observed to alight upon the dorsum of the thorax and quickly deposit sev-

1 Contribution No. 29 from the Zoology Department, Agricultural Experiment Station

of the Kansas State Agricultural College.

116

A PARASITE OF THE FIELD CRICKET 117

eral living maggots, which, encountering only the soft tender membrane,

speedily made their way into the body cavity of their host. The maggots

are capable, however, of entering a host which is fully dried out and

hardened.”’ This observer found as many as sixteen of these larvae in the

body-cavity of a single grasshopper. He found that the larvae usually

escaped through the wall of the thorax immediately behind the anterior

coxa, but that others either bored through the abdominal wall or escaped

through the anus.

Kelly (p. 438), who reared large numbers of S. kel/yi, found that the

larvae deposited in the fall, on escaping from the grasshopper host, pene-

trated the ground to a depth of from one-half an inch to 2 inches, where

they remained throughout most of the winter. Pupation in nature

occurred early in March and adults emerged from late March until the

last of May. In warm weather Kelly (p. 439) found that the life cycle was

completed in much shorter periods. By June or July a second generation

had matured, and from this time “until November no distinction could be

made between generations on account of overlapping. However, judging

from the rapidity of their development, there were probably three or four

additional! generations, making about five or six for the season.’”’ Accord-

ing to this author, S. kellyi has been reared from grasshoppers at Welling-

ton, Kan., Washington, D.C., and points in New Mexico, Arizona, and

Utah.

SUMMARY

1. Larvae of Sarcophaga kellyi Aldrich were found inhabiting the

body-cavity of black field crickets, Gryllys assimilis Fabricius at Manhat-

tan, Kan., in September, 1920.

2. This seems to be a new case of parasitism in the black field cricket.

_ 3. The larvae of Sarcophaga kellyi have been reported from grasshop-

pers in Kansas, Washington, D.C., New Mexico, Arizona, and Utah.

LITERATURE CITED

Knity, B. OG, |

1914. A New Sarcophagid Parasite of Grasshoppers. Jour. Agr. Research, 2:435-446.

Regn, J. A. G., AND HEBARD, M.

1915. The Genus Gryllus (Orthoptera) as found in America. Proc. Acad. Nat. Sci.

Phila., 67:293-322.

Smita, H. E.

1915. The Grasshopper Outbreak in New Mexico During the Summer of 1913. U. S.

Bur. Ent., Bul. 293:1-12.

FRESH WATER AND MARINE GYMNOSTOMINAN

INFUSORIA

Lron Aucustus Hausman, Px.D.

Biological Laboratory, Cornell University

INTRODUCTION

The present contribution to the survey of the protozoa deals with the

characteristic appearance, habits, and habitats of members of one of the

largest and most important groups of the protozoa, namely, the Gymnos-

tomina. Within this suborder are included many of the largest of these

unicellular forms of animal life; forms which constitute one of the most,

if not, indeed, the most important source of food supply for the smaller

aquatic organisms, which in their turn form the bulk of the food of fishes,

Their presence in ponds and streams is of great importance, for they

convert refuse matters which might pollute the water into an available

source of food for higher forms of life. A study of the protozoan faunas

of waterways should, it seems, go hand in hand with a study of the problems

of water purification, and of the preservation and utilization of our aquatic

resources.

The majority of the species of Gymnostomina treated in this paper are

fresh water. Several marine species are also included.

The water samples of which examination was made were secured from

various portions of New York, Connecticut, Massachusets, and Mississippi,

and over 1,000 were examined. They were taken from open lakes, ponds, -

roadside pools, rivers, brooks, rills, marshes, watering troughs, and the

like.| The marine samples were secured from the Connecticut shore of

Long Island Sound and from tidal estuaries and embayments, in the

vicinity of New Haven.

METHODS OF STUDY

Methods of collecting material containing protozoa, in the field, are too

well known to need much discussion here. The methods used in the

present investigation underwent no decided original modifications from

the methods commonly employed.?

Half a dozen pint fruit jars, fitted into a small, suit-case-like convey-

ance, together with a small silk plancton net, a large, long handled cooking

spoon, and several glass tubes of various lengths (with detachable compres-

sion bulbs for ‘‘sucking’’), comprised the entire field equipment. The

jars were labelled, and a record kept of the nature of the locations from

1 See Hausman, L. A. Observations on the Ecology of the Protozoa, Am. Nat., vol. 4,

1917, p. 157.

2See Hausman, L. A. A Contribution to the Life History of Amoeba proteus, Leidy,

Biol. Bull., No. 5, May, 1920, p. 340.

118

FRESH WATER AND MARINE INFUSORIA 119

which the samples were taken, for future reference. Likewise each precise

spot whence samples came was indicated on a topographic map. Possibly

this may be found useful at some later time.

Upon arrival at the laboratory the samples were transferred to wide,

open-mouthed jars. An examination was made of each sample immedi-

ately, and for a week or so, on each succeeding day, with the view of

keeping record of the new species which emerged from encystment with

the gradual stagnation and putrification of the water, for except in a very

few cases the samples contained algae or other vegetal matter. The

small bolting silk net shown in Fig. 3 was used for concentrating the

infusoria content of one or more pipettefuls of water from the middle or

bottoms of the samples where the water was usually more or less clear.

No concentration methods were needed in the examination of the surface

scum of the putrescing material.

All measurements were made with an ocular micrometer, or from a

ruled millimeter slide, from retarded living, or freshly killed specimens.

The characters which are, perhaps, the most satisfactory for use in the

identification of the living animals are: the contour of the body, the posi-

tions of the buccal cavity and of the largest contractile vacuole, and the

disposition of the cilia. Killing and staining, or intra vitam staining may

make apparent the structure of the pharynx and also of the nuclear ele-

ments. This treatment may sometimes be necessary for bringing out of

the cilia. Methods of post mortem and of intra vitam staining will be

discussed later.

For the first examination of samples a small drop of water was taken

from the top scum, or from concentrated material (the results of straining)

and mixed with an equal volume of very viscous gelatine solution, and the

whole thoroughly stirred together on the slide with a curved needle. Or

often several drops were mixed with an equal part of the gelatine in a watch

crystal and used on the slide when needed. The drop on the slide was now

carefully flattened out and examined without a cover glass under low

power (16 mm. objective and 4x eyepiece) to ascertain if the solution were

of a viscosity great enough to check sufficiently the movement of the

protozoa. If not, it was allowed to concentrate still more by evaporation,

until properly viscous, and a cover glass applied. Magnification with the

16 mm. objective and the 4x and 10x eyepieces, and with the 4 mm. objec-

tive, and the same two eyepieces was usually found of sufficient strength

for the determination of the species described in this paper. A word of

caution is to be given here concerning the clarity of the gelatine solution.

The gelatine used must be of the best grade and the solution must be

’See Hausman, L. A. The Manipulation and Identification of the Free-Swimming

Mastigophora of Fresh Waters, Am. Nat., vol. 44, 1920, p. 333.

120 LEON AUGUSTUS HAUSMAN, PH.D.

perfectly fresh. It was found that gelatine which had stood for some

time became cloudy in appearance and stringy in texture, due to the

growth of colonies of mould plants and bacteria.

Another method of quieting the movements of the protozoa, which

was developed, consisted in chilling the slide and its supported water drop

on a small block of ice. As the temperature decreased the motions of the

protozoa became slower and slower, though never so slow as those incar-

cerated within the gelatine mixture. This method was devised more in

the spirit of curiosity than in any hope that it would be as great an aid as

the gelatine method of quieting movement.

Permanent mounts of the infusoria are believed to be very unsatisfac-

tory, with the exception of those made of Difflugia, Arcella, Euglypha, the

Foraminifera, and others whose bodies secrete a protective shell or test.

And here it is the test and not the creature itself which is preserved in its

original form. During the process of killing, of staining, and of mounting,

the body form is more or less distorted, and the cilia deformed or lost.

The most convincing demonstration of the poverty of the mounted slide

can be had by examining together a living Paramoecium retarded in the

gelatine solution, or one freshly stained intra vitam, and a mounted slide,

of the same creature, of the best manufacture obtainable. For optimum

results in the study of gross anatomy, at least, or for the needs of the

systematist, nothing, I think, can equal the intra vitam staining, with

the creature hampered in its movements in the gelatine solution. The

movements of the cilia or of the contractile vacuoles are often of the greatest

aid in determining their position and form. In fact the presence and form

of the pharynx in its entire length can often be made out, in certain species,

only by means of the cilia vibrating within it.

The stains‘ most frequently used were methyl] blue, and gentian violet.

Safranin, methyl] green, and iodine were also used. Safranin, it was found,

stained the deepest, and methyl] blue the least. For certain forms, there-

fore, the one was used, and for others, the other. In the case of each stain.

a 95% alcoholic solution of the dry stain was made and kept in a small

bottle ready to be diluted before applying to the slide. The staining set

holder (Fig. 1) was designed to contain in a compact and convenient form

the requisite number of stains, and other reagents, together with solid

glass dipping rods for each. Thus any mixture of reagents was prevented.

The labels (shown underneath the holes for containing the dipping rods)

bore the names of the reagents. A great deal of comfort was derived from

this very simple piece of apparatus.

The killing and staining was accomplished in two ways; either by killing

first and staining afterwards, or by performing both operations simultane-

4 See formulary of reagents at end of paper.

FRESH WATER AND MARINE INFUSORIA Wil

ously. The killing fluids used were: a 10% aqueous solution of tannic

acid, a 1% aqueous solution of copper sulphate, a 2% aqueous solution of

osmic acid, a 4% aqueous solution of acetic acid, a 3% aqueous solution

of mercuric chloride, a 1% aqueous solution of formaldehyde. The osmic

acid and copper sulphate solutions seemed to be the best killers, killing the

animals at once, and without apparent distortion. Neither did disinte-

gration set in with such rapidity as was the case when some of the other

killing reagents were employed. These killing reagents can be used in

other strengths than those given here but these percentages seemed to

give the best results.

The killing was done either with a large amount of the material in a

watch crystal, or underneath the cover glass, and the staining was accom-

plished in the same way. Where the protozoa were extremely abundant,

as they were usually in surface scums or infusions of decaying marine

algae, the watch crystal ‘“‘mass”’ staining or killing was found to be the most

satisfactory, as well as the easiest and quickest. This method had also to

commend it the fact that both the killing reagent and the stain could be

most readily controlled. Several watch crystals full of material were

placed side by side and very delicate gradations of color secured.

As has been previously stated, the intra vitam staining gave by far

the best results. This was accomplished either under the cover glass,

or in the watch crystals, following the methods noted above, after the

gelatine had been added and the proper degree of viscosity secured.

PREPARATION OF CULTURES

In order that a large number of individuals of a given species may be

available for examination, it is necessary to depend upon cultures. For

convenience in designation, there have been recognized in this paper the fol-

lowing types: (1) natural cultures, that is, those in which large numbers of

a species appear, in natural conditions in the field and without any artificial

manipulation of the medium in which they occur, (2) indirect cultures, or

those which result from merely collecting the material and allowing it to

stand and to decompose in the laboratory, and (3) artificial cultures, or

those which are prepared with a definite nutritive medium (determined by

experimentation) and inoculated with the desired species.

There is little or no exercise of technique involved in securing either

natural or indirect cultures. One soons learns to recognize good natural

culture environments such as greenish duck ponds, for Euglenae of various

species; boggy water supporting. growths of Sphagnum, for Prorodon

niveus and armatus ; watering troughs with Spirogyra or other Chlorophyceae,

for species of Chilodon and Holophrya; clear, cold waters for Astasia, etc.

12>. * LEON AUGUSTUS HAUSMAN, PH.D.

For indirect cultures one has merely to allow the collected water and

vegetation to stand in the warmth and light of a south-exposed laboratory

window, and make regular examinations day by day.

Where, however, but few individuals of a desired species occur, it

becomes necessary to aid their propagation artificially. Results which

gave earnest of better ones with further experimentation, were obtained

by what is here termed artificial culturing. This was accomplished by

segregating desired individuals, and then introducing into the jar of water

in which they were placed some favorable nutritive substance. The

methodology of preparing such cultures has been well enough developed

at the present time, possibly, to make an account worth while, though

many problems of detail still await solution.

For capturing individual protozoa under the microscope, there was

devised what is here called an isolation pipette, shown in Fig. 4. A soft

glass tube is drawn out to a hair-like degree of fineness at one end, and

inserted into a thin walled rubber tube at the other. The opposite end of

the rubber tube is tightly closed by means of a sealed glass tube. The

hair-like point of the pipette is first dipped into clear water to allow

capillary attraction to draw as much as it will up into the bore. The

forefinger of the left hand is laid lightly upon the rubber tube near to its

closed extremity, compressing it slightly and thereby driving out a small

drop of the water from the tip of the glass pipette. The latter is now

inserted with the right hand underneath the objective and into the uncoy-

ered drop on the slide. Release of the pressure of the left forefinger results

in the withdrawal into the hairlike bore o the pipette a small quantity

of water, the amount of which can be delicately regulated.

After the desired animal has been thus captured it is forced out into the

water in the isolation jar, and there is added the proper nutritive substance.

Thereafter the whole is set in a warm, light place to “ripen.”” It was found

advisable, from the standpoint of ease of handling, to make the culture

in small 3 or 4 cm. stender dishes. To inoculate such small cultures it

sufficed, on several occasions, to introduce but a single individual. This,

however, it must be confessed, was because we could secure no more, and

the successes resultant from this meagre inoculation were regarded

merely as fortunate accidents.°

Several inoculations, aggregating some half dozen, or preferably

more, individuals are usually necessary. There was no certain way of deter-

mining, save after the anticipated development of the culture, whether the

° Single individuals can be removed by means of the isolation pipette, and introduced,

for long-continued observations, into a device termed the micro-aquarium (See Hausman,

L. A., The Vibratile Oral Membranes of Glaucoma scintillans, Ehr., Am. Nat., vol. 44, 1920,

p. 427.

FRESH WATER AND MARINE INFUSORIA h23

animals had actually been introduced into the isolation jar and inocula-

tion actually accomplished. Rather clumsy attempts, yet in several

instances not unsuccessful ones, were made to make as certain as possible

the incarceration of single individuals within the isolation jar by first eject-

ing the captured animal into a drop of clear water on a slide while under

the microscope, noting the presence of the creature, and then washing it

off carefully into the material in the isolation jar with a fine stream of clear

water.

Tubes of different tip diameters were used, and it was noted that the

most success attended the use of the finest of these which it was possible to

use for a given species. It is well to give the tip a slight turn when draw-

ing it out, as shown in the figure. This seemed to make it easier to man-

ipulate under the microscope.

It was found practicably impossible to manipulate the pipette and to

capture the protozoa under any power greater than that afforded by the use

of the 10x eyepiece and the 16 mm. objective. To insure the best results

the drop of water must be well flattened out, and first freed from annoying

débris.

For maturing the cultures rapidly, and under conditions which could

be regulated and tabulated for further referene, a culture oven, such as is

illustrated in Figure 2 was devised. A large aquarium jar was equipped

with perforated tin shelves hung by copper wires from the upper rim of the

jar; heated with a carbon filament lamp, placed on a copper wire platform

to prevent it coming into contact with the glass bottom of the jar, and

covered with a cardboard cover bearing a thermometer. This had the

advantage of furnishing to the stender dish cultures placed on the shelves,

at once the requisite amounts of heat and light. The temperature of the

interior of the oven was regulated by raising or lowering the cardboard

cover, propping it up with little wooden blocks.

Samples were dessicated in this oven by removing the cover and allow-

ing the cultures to remain until they had dried. Inthis way cultures of

Holophrya, Prorodon, and Loxophyllum were kept and resuscitated ‘at

pleasure. This method of keeping material by dessication might be

a useful one for class requirements. The dried material could be removed

from the dishes and placed in labelled envelopes, and filed away in a card

catalogue tray. More experimentation along this line might reveal the

fact of its being possible to have on file any quantity of protozoa. material

which could be revivified at will for class room use!

KEY TO THE FAMILIES AND SUBFAMILIES

I. Protozoa possessing, at some stage of the life cycle, locomotor appen-

dages in the form of cilia, either single or fused into membranes.

Macro and micro nucleus present. CLASS INFUSORIA

124 LEON AUGUSTUS HAUSMAN, PH.D.

II. Cilia present during the entire life cycle; buccal cavity and anal orifice

normally present; contractile vacuole often connected with an

excretory tube system.

SUBCLASS CILIATA

III. Cilia more or less alike in form and distribution over the entire body,

having a tendency to lengthen (or in some cases to be present only)

on the oral or aboral side. Buccal cilia usually a trifle longer than

the others. j

ORDER HOLOTRICHIDA

IV. Lacking undulating membranes about the buccal cavity, the latter

being closed except during the ingestion of food.

; SUBORDER GYMNOSTOMINA

A. Body outline usually oval or extended; neither oral nor aboral

sides flattened

Family Enchelinidae

AA. Body outline sometimes oval or extended, more often,

however, with oral side either flattened or concave.

B. Buccal cavity terminal or nearly so

Family Trachelinidae

BB. Buccal cavity not terminal

C. Gullet with pronounced curve

Family Enchelyidae

CC. Gullet without pronounced curve

D. Body entirely and evenly ciliated

Subfamily Nassulinae

DD. Cilia longer on, or confined to aboral side

Subfamily Chilodontidae

DDD. Cilia confined to oral side

Subfamily Erviliinae

KEY TO THE GENERA MENTIONED IN THIS PAPER :

A. Body ovoid, ellipsoidal, or almost spherical

B. Body distinctly ovoid or spherical

C; Posterior spinous process present: =). > .uL eee xe UROTRICHA

CC. Posterior spinous process not present

D. Possessing spiral band of longer cilia.......... PERISPIRA

DD. Not possessing spiral band of longer cilia

E. Cilia restricted to one or two circles about the body

F. One midway circle of cilia present.......... MESODINUM

FE) Two such.citcles present: a0 2 eee eee DIDINIUM

EE. Cilia not restricted to one or two circles about the body

EF. Cilia restricted: tovone'side of bodyasee- ee TROCHILIA

FF. Cilia not restricted to one side of the body

FRESH WATER AND MARINE INFUSORIA 125

G. Buccal tavity anteriorly terminal

Hea Nucleus long*and: curved) 20 0. ENCHELYODON

HH: Nucleus usually ovoidal.......... HOLOPHRYA

GG. Buccal cavity not anteriorly terminal

H. With a short neck like, or lip like projection from

EUTNCE TOT CMC Beg ick ee ae ste omnes TRACHELIUS

HA” Without? such “a. projection. 22.5). .1.-- NASSULA

BB. Body not ovoid or nearly spherical, but drawn out into the form of

an elongated ellipsoid

C. Body armored ‘with rectangular plates

D. Cilia arising from middle of rectangular plates...... COLEPS

DD. Cilia not arising from? middle of plates: ...-...:: TIARINA

CC. Body not armored with rectangular plates

D. Cilia restricted to ventral surface

E. Body about five times as long as, or longer than, greatest

(GIN ein Wg VERUTEY ek neh Dacia aN RE at ou ee 8 LIONOTUS

EE. Body less than five times as long as its greatest diameter

Bee aE AR ey en lei Tm a TG. wap Ne LIONOTOPSIS

DD. Cilia not restricted to the ventral surface

E. Body longitudinally furrowed.......... PLAGIOPOGON

EE. Body not longitudinally furrowed.......... PRORODON

AA. Body not ovoid, ellipsoid, or almost spherical

B. Body purse or flask shaped

C. With long flexible neck or proboscis

D. With circle of longer cilia about the anterior extremity of the

[ARO WOSCIGHE er, ere Meet Pan, eyes cet dar TRACHELOCERCA

DOP NWWithoOutosieh: a: /eircle con icililan we. a «ou LACRYMARIA

CC. With short neck

D. Neck obliquely truncated

E. Ciliation entire

Fo Body “capable -of” distortion at’. will. | 2%... ENCHELYS

pt Body not capable of distortion... ...<'. SPHATHIDIUM

PB Oilrtione WMOti embibers metres stake ets.) PHASCOLODON

DD. Neck not obliquely truncated

E. Tentacular process arising from the buccal cavity in the

ANPELIOEIECMU. OM Mechanar ee is Br ee aN eas ILEONEMA

EE. Without such a tentacular process TRACHELOPHYLLUM

BB. Body not purse or flask shaped

C. Body ribbon or leaflike

D. Body very elongate

Be Amtenon ‘end: TOUNCEC Maser. 4 dear seo! AMPHILEPTUS

Pin vAitbenor end wnop prounded.. <2... FLEXIPHYLLUM

126 LEON AUGUSTUS HAUSMAN, PH.D.

An ideal composite gymnostominan ciliate, to

show the various anatomical divisions and organs

of the body.

a. proboscis cilia

c. proboscis

d. buccal cilia

e. buccal cavity opening into pharynx

f. food vacuoles

g. nucleus

h. principal contractile vacuole

i. hyaline border of body

1. caudal cilia

m. anus

n. smaller contractile vacuole

. discharged trichocyst

undischarged trichocyst

pharynx

cila band

. pharyngeal rods

ay tre (e tale e)

FRESH WATER AND MARINE INFUSORIA LD

DD. Body not very elongate

Ke With antenor ‘border. crenulates 2202-0. 6s. LOXODES

EE. Without crenulations on the anterior border

F. Neck elongated and constricted............ DILEPTUS

FF. Neck not elongated and constricted.. LOXOPHYLLUM

CC. Body not leaflike; usually kidney or bean shaped, or nearly so

DP harynxe longa aiid CULVEUM tars gebaacs neta ket. a TILLINA

DD} -Pharynx ‘short rand: straight: oe... CHILODON

OBSERVATIONS ON THE RECORDED SPECIES

Genus Holophrya

This genus, a very common one, and widely distributed, possesses

either an enormous number of distinct species, or a much smaller number,

many of which are very variable in size and form, and often, indeed, in

coloration. From our limited observations we are inclined to take the

latter view. Little attempt has heretofore been made to accord all of

these diverse forms specific names.

The species here called Sp. 1 and 6 were commonly found in brackish

tidal estuary water about a mile from Long Island Sound among detached,

floating Fucus and green Sea Lettuce, the first day after having been

brought into the laboratory. All of the individuals of these species

observed (and there must have been hundreds seen) varied but little away

from an average. Later on, however, considerable variation occurred

as the numbers became greater in the slowly putrescing material. It is

difficult to say whether this was due, however, to an increasing variation

among the individual members of the species, or whether new species

Species 5, 6, and 7 were present in enormous numbers in a four days

old infusion of fresh cabbage leaves, and in an infusion of dried corn

leaves of the same age.

Species 2, 3, and 4 were met with occasionally in almost all samples,

both marine and fresh water, particularly in those from ponds.

The globularity of body, and the small anterior or anterior-lateral

buccal cavity, together with the uniform length and distribution of the

cilia, appear to be constant characteristics of the members of the genus.

Among such small, globular forms, mutilated individuals seemed not to be

very common. We are of the opinion that figures of very irregular forms

assigned to this genus were made from such mutilated individuals.

Holophrya sp. 1 to 7 shown in Figs. 5-11.

Genus Urotricha

Members of Urotricha may easily be recognized by the presence of the

caudal spine or seta, and by their curious habit of swimming slowly and

128 LEON AUGUSTUS HAUSMAN, PH.D.

evenly and then suddenly jerking ahead, or to the right or left, as though

shot by a spring, a motion resulting from a ghia lateral snap of the rigid

caudal seta.

Kent has observed that the walls of the pharynx are surprisingly elastic,

and that this enables the creatures often to take in food, the bulk of which

may equal their own bodies!

If the posterior spine is not easily seen, staining makes it easily visible.

Urotricha farcta and platystoma, Figs. 12 and 13.

Genus Perispira

The Perispira ovum (Fig. 14), of Stein, which we have recorded from

stagnant pond waters, may be the same as the Holophrya ovum, of Ehren-

‘berg. If this be so then it is anaberrant form of Holophrya, for it possesses

a spiral band of longer cilia characteristic of Perispira. Since even cilia-

tion is characteristic of Holophyra, this form had best be placed in Perispira.

Genus Enchelys

This genus is hardly distinguishable from Holophrya, the presence of the

laterally opening buccal cavity of Enchelys being apparently the sole point

of difference. And when the members of the latter assume a globular form,

which they do, with apparent volition, the buccal cavity becomes almost

exactly anteriorly terminal, much like that of many species of Holophrya.

This assumption of globularity occurs, often, when the animal is gorged

with food granules. The young, soon after division has been completed,

also take on a globular body.

The smaller species figured is regarded as the Enchelys farcimen (Rie

16) of Ehrenberg. EE. pupa (Fig. 15) was met with several times in p6nd

water.

Genus Enchelyodon

The ovate-elongate body and the terminal. buccal cavity, together with

the large size, should serve to distinguish Enchelyodon farctus (Fig. 17)

from forms in the genera Prorodon and Enchelys. Note that the cilia are

very short. We were unable to see them in the unstained animal. This

form was rarely found in the waters of bogs, ponds, slowly moving streams,

ele:

Genus Spathidium ;

The chief difference between this genus and Enchelys, from which it is

separated only with difficulty, appears to be the possession of a longer

pharynx, usually furnished with pharyngeal rods. The latter, however,

are not easily visible.

The species figured, which seems to be Spathidium spathula (Fig. 18),

was found in pond and slow stream waters.

FRESH WATER AND MARINE INFUSORIA 129

Genus Prorodon

Both Prorodon armatus (Fig. 19) and P. ovum (Fig. 20) were rarely

found in pond waters. The buccal cavities of both are distinct, and the

prominent pharyngeal rods of the latter were very good as an identification

characteristic.

Genus Lacrymaria

Lacrymaria olor (Fig. 21) is a common form in infusions of leaves both

of deciduous tress and of aquatic plants. Like Trachelocerca olor (Fig. 22)

it often lies with its lenticular body concealed among a mass of débris and

shoots forth its long serpentine neck in all directions. Whether this is a

deliberately willed concealment for the purpose of protection, or for the

advantage which it secures for the seizure of prey is uncertain. I have

not seen this habit mentioned elsewhere, and yet I found it to be a very

common one among the many individuals observed.

Its size is extremely variable, but the constant body form offers a ready

means for identification.

Genus Trachelocerca

To be distinguished from Lacrymaria chiefly by the smaller size of its

members. Trachelocerca olor (Fig. 22) and Lacrymaria olor are almost

identical in habits. The movements of the smaller form are, however,

the more rapid. Tracheolocerca olor is found commonly among the smaller

aquatic vegetation in small quiet pools and coves.

Trachelocerca phoenicopterus (Fig. 23) is a marine species occurring

among algae along the shore, as well as in putrifying infusions. Its length

seems to be very variable.

Genus [leonema

Ileonema dispar (Fig. 24) occurs among Spirogyra, Zygnemea, Oscilla-

toria, and can probably be found among any of the fresh water filamentous

algae. It is not a common form, and usually disappeared soon from fresh

material.

The cilia are sparse and apparently weak.

Genus Plagiopogon

Plagiopogon coleps (Fig. 25) which we figure from Kent, we believe

to have found in salt water among decaying Fucus and other algae. It

closely resembles Coleps hirtus (Fig. 26) though the longitudal furrows

of the body and the absence of armor plates are apparent under high

powers. It seems to be a species of fairly constant form and size.

Genus Coleps

Coleps hirtus (Fig. 26) is a very common form of ciliate, the commonest

of its genus, among decaying vegetation and in old infusions, and can be

130 LEON AUGUSTUS HAUSMAN, PH.D.

readily identified from its size and armored body. In swimming it twirls

rapidly on its longitudinal axis and pursues a rapid, wavering reckless

course. It is an exceedingly voracious species and appears to feed on both

animal and vegetable tissue, and the bacteria which are disintegrating them.

Genus Tiarina

Tiarina fusus (Fig. 27), a marine form from among decaying algae,

resembles Coleps in structure very closely. The form of the body is,

however, different. The form which we figure we take to be Tiarina fusus,

(Fig. 27) which is apparently the same as the Coleps fusus of Claparéde

and Lachmann.

Genus Didinium®

Didinium nasutum (Fig. 28), not an uncommon form in decaying

and fresh aquatic vegetation, is one of the largest of the ciliates. Its two

zones of cilia offer an easy character for identification. This form appears

freely where an adequate supply of smaller ciliates appear, for it is upon

these that it feeds. The habits of this species have been exhaustively

studied by S. O. Mast (22) and recorded in one of the most interesting

of the recent papers on protozoan habits.

The natatory movements of this species are much like those of Uro-

tricha, namely a slow gliding progression interrupted frequently by spas-

modic jerks.

Genus Mesodinium

Mesodinium cinctum (Fig. 30) is not an uncommon form in salt water

and when swimming rapidly looks very much like a minute replica of

Didinium nasutum. ‘The constricted median line and the single zone of

median cilia make it easy to identify when at rest.

The smaller Mesodinium (Fig. 29), which is found rather rarely asso-

ciated with the preceding species, I consider to be the Mesodinium pulex

of Claparéde and Lachmann.

Genus Tillina’

Tillina magna (Fig. 31) were found frequently in a ten day’s old infusion

of dried corn leaves associated with various species of Holophrya, Chilodon,

and Colpoda. Its distinguishing characteristics are the irregular, asym-

metrical body and the curved, ciliated pharynx.

Genus Amphileptus

Fig. 32 I have called provisionally, Amphileptus gutta. It seems to occur

in both marine and fresh water infusions. They bear either many smaller

contractile vacuoles distributed over the posterior two-thirds of the body,

6 See Mast, S. O., The Reactions of Didinium nasutum, etc., Biol. Bull., vol. 16., 1908,

p: ol:

7See Gregory, L. H. Observations on the Life History of Tillina magna, Jour. Exp.

Zool., vol. 6, 1909.

FRESH WATER AND MARINE INFUSORIA 131

or occasionally, yet not so frequently, one single large vacuole, situated

in the posterior half or posterior end, or slightly to one side. Because of a

lack of very definite characteristics forms like this are difficult to place

with certainty.

Genus Lionotus

Members of this genus are among the most graceful ciliates. Viewed

from above, the apparently slender neck is seen to be broad and leaflike.

Figures of these species should, therefore, indicate this and not lead to

the impression that the neck is of the same type as that possessed by

Lacrymaria or Trachelocerca.

Lionotus wrzesniowski (Fig. 36) is a large form, found in pond waters

amid living and dead aquatic vegetation, where often occurs, also, Lionotus

fasciola (Fig. 34). The latter species is also found in salt water with

Fucus or other marine algae. A similar form, entirely restricted to fresh

water, is Lionotus pleurosigma (Fig. 35). This species can be distin-

guished from fasciola only by its clear, deep, hyaline border.

The smallest of the species figured (Fig. 33) was found in brackish

water in a tidal estuary among detached, floating marine algae.

Genus Lionotopsis

Fig. 37 is, perhaps, the Lionotopsis anser of Conn, drawn from but a

few poorly defined individuals found in pond water. The position of the

buccal cavity could not be determined.

Genus Loxophyllum

Members of this genus can usually be recognized by the gracefully

flexible way in which they glide about over and through débris or wrap

their pliant and leaflike bodies about it. The ease with which the curved

anterior portion of the body is used for the examination of possible food

substances reminds one of the sensitive exploratory gropings of the tip

of an elephant’s trunk. The deep, clear, hyaline border possessed by all

the Loxophylla is constantly characteristic.

Loxophyllum setigerum (Fig. 39) and rostratum (Fig. 40) were found

quite abundantly in brackish water. The latter appeared in great abun-

dance in an eight days old infusion of green Sea Lettuce and Fucus in

salt water.

Loxophyllum sp. 1 occurred in fresh water among aquatic vegetation

(Fig. 38).

Genus Trachelophyllum

Fig. 41 has been called Trachelophyllum tachyblastum, from a single

specimen found in pond water.

Genus Flexiphyllum

Flexiphyllum elongatum (Fig. 42) is frequently met with in pond water

among growing vegetation. Its motion is a graceful and sinuous gliding

132 LEON AUGUSTUS HAUSMAN, PH.D.

and it makes rapid progress through the water. We have found that it

prefers to move concealed amid débris.

Genus Trachelius

Trachelius ovum (Fig. 43) possibly the most common species, can be

distinguished by its large size, its curious little neck, and its deliberate

motions. The buccal cavity and gullet are quite prominent in most

individuals. The size and shape of the neck is apparently subject to con-

siderable variation. Within the body the number of contractile vacuoles

is normally very large.

Genus Dileptus

Dilepius gigas (Fig. 44), fairly common form, is of unusual variability

in size and shape. It is entirely carnivorous and possessed of a voracious

appetite. The prey is stung and rendered helpless by the discharge of the

trichocysts located along the border of the long neck like process, and if too

large to be swept into the buccal cavity by the lashings of the buccal

cilia is forced in by the writhings of the neck. The body often rotates |

on its longitudinal axis during progression through the water.

Individuals have been reported which measured 800 uy.

Genus Loxodes

Loxodes rostratum (Fig. 45) was found only rarely in pond water among

fresh and decaying vegetation. It is reported to occur also commonly,

in infusions.

Genus Nassula

This is a beautiful genus, its members being symmetrically ovoid, and

many of them iridescent. Nassula microstoma (Fig. 48) is a very pretty

species. It is usually brownish or yellowish, the color depending upon 1S

contained food. Under strong light, as it revolves through the water, it

scintillates brightly, reminding one of a small, ovoid, minutely faceted

epidote. This was a very common species in brackish tidal estuary water.

Nasulla ornata (Fig. 46 and Sp. 7 (Fig. 47) were found in pond waters

among fresh and decaying vegetation.

Genus Chilodon

Chilodon, much like Holophrya, is a genus containing a great number

of species of considerable variability of form and size. Of all the species

which vary in this way among themselves, Chilodon cucullulus (Fig. 52),

the commonest, is the most flagrantly disregardful of maintaining its

proper dimensionsand contour! In the same infusion we have found no less

than a dozen differently shaped and sized specimens! Calkins says of this

species that it is “extremely variable. . . and has received so many

different names that it hardly pays to enumerate them all.”’ It is “one

of the most common and widely spread ciliates known.”

FRESH WATER AND MARINE INFUSORIA 133

This species appeared, in a remarkably pure and rich culture, ina five

days old infusion of dried flaky deposit from the sides of an old wooden

watering trough. It occurred also abundantly in fresh and putrid sea

water.

Chilodon megalotrocha (Fig. 49) and caudatus (Fig. 51) are sometimes

found associated with cucullulus. Chilodon vorax (Fig. 50) is much

less common. ;

Genus Phascolodon

Phascolodon vorticella (Fig. 53) is a freshwater form from swamps,

-rather rare. It is a beautiful form, graceful and deliberate in its move-

ments. The body plasm is clear and crystalline.

Genus Scaphidiodon

Scaphidiodon sp. 1 (Fig. 54) is perhaps the form shown in Fig. 54.

This occurs in sea water containing putrid animal and plant tissues.

Genus Trochilia

Trochilia sigmoides (Fig. 55) is another marine form, very striking and

beautiful. It is found rarely in clear salt water with living algae, and

can be easily recognized by the oblique banding on the body and the

meagre number of cilia restricted to one side of the body. Near the small,

anterior end of the body is located the buccal cavity, apparently not

followed by a definite pharynx, but surrounded by cilia a trifle longer

than the rest. It swims slowly, rotating.

FORMULARY OF REAGENTS FOUND USEFUL IN QUIETING,

KILLING AND STAINING

I. QUIETING SOLUTIONS

PeeeGela bine solition——VWatet- so tiers a: a. 5.02;

: Gelabintenm tes os ese Iso?

Heat slowly until gelatine is dissolved; then allow to cool and

congeal to the desired viscosity.

2. Chlorotone—a 1 per cent aqueous solution

II. Kuitiinc REAGENTS

1. 10 per cent aqueous solution tannic acid

2. 2 per cent aqueous solution osmic acid—Invert slide with

drop of water containing the protozoa over a bottle of the

solution, uncorked. The fumes kill almost instantly.

3. 3 per cent aqueous solution acetic acid

4. 1 per cent aqueous solution copper sulphate

5. 2 per cent aqueous solution chromic acid

III. Srarminc Reacents (These can be made up as saturated solutions

in either water or 95 per cent alcohol, and diluted to the desired

134 LEON AUGUSTUS HAUSMAN, PH.D.

depths of color. When used in aqueous solutions, very dilute, _

they make good intra vitam stains.)

1. methyl blue 6. gentian violet

2. methyl green 7. iodine, with potassium io-

3. Lichtgriin dide (a killing stain, either

4. Bismark brown with water or alcohol).

5. Safranin h

BIBLIOGRAPHY

BLOCHMANN, F.

1895. Die Mikroscopische Tierwelt des Siisswassers, Abt. I. Protozoa, Hamburg.

Butscatt, O. ‘

1880-1889. Protozoa, in Bronn’s Klassen und Ordnungen des Thierreiche, Bd. I, Th.

J-III, Leipzig u. Heidelberg.

CaLKins, G. N.

1901. Marine Protozoa of Woods Hole, Bull. U. S. Fish Comm. vol. 21.

CaLxkins, G. N.

1901. The Protozoa, N. Y.

Catxtns, G. N.

1909. Protozoology, N. Y.

Carter, H. J.

1856. Notes on the Freshwater Jnfusoria of the Island of Bombay, No. I Organization

Ann. and Mag. of Nat. Hist., 2nd Ser. vol. 18, p. 115.

CLAPAREDE ET LACHMANN

1868. Etudes sur les Infusoires et les Rhizopodes, Genéve et Bale.

COcCKERELL, T. D.

1911. The Fauna of Boulder County, Colo., Univ. of Colo. Studies, vol. 8.

Conn, H. W.

A Preliminary Report on the Protozoa of the Fresh Waters of Connecticut, Conn. State -

Geol. and Nat. Hist. Survey, Bull. 2.

DOFLEIN

1909. Lehrbuch der Protozoenkinde, Jena.

Epmonson, C. H.

1906. The Protozoa of Iowa, Pro. Davenport Acad. Sci. vol. II, p. 1.

Grecory, L. H. :

1909. Observations on the Life History of Tillina magna, Jour. Exp. Zool., vol. 6, no. 3.

GRUBER

1882. Neue Infusorien, Zeit. Wiss. Zool. vol. 33, p. 439.

GRUBER

1884. Die Protozoen des Hafens von Genua, Nov. Act. des K. Leop. Car. Deutsch.

Akad. der Naturfor., vol. 46, p. 475.

Hausman, L. A.

1917. Observations on the Ecology of the Protozoa, Am. Nat., vol. 4, p. 157.

HENDERSON, W. D. '

1905. Notes on the Infusoria of Freiburg im Bresgau, Zool. Anz., vol. 29, p. 1.

JENNINGS, H. S.

1899. A Report on the Protozoa of Lake Erie, etc., Bull. U. S. Fish Com., p. 105.

KEntT, W. S.

1880-1881. A Manual of the Infusoria, 3 vols., Lond.

FRESH WATER AND MARINE INFUSORIA 135

Koror, C. A.

1897-1901. Plancton Studies IT, Bull. Ill. State Lab. Nat. Hist., vol. 5, p. 273.

LANDACRE, F. L.

1908. Protozoa of Sandusky Bay and Vicinity, Ohio Acad. Sci., vol. 4, p. 421. (This

paper contains an excellent bibliography, chiefly of American writers, brought

up to the date, 1904)

LankKEstER, E. Ray (Editor)

1903. A Treatise on Zoology, Part I, Introduction and Protozoa, 2nd fascicle, Lond.

LIEBERKUBN, N.

1856. Contributions to the Anatomy of the Jnfusoria, Ann. and Mag. of Nat. Hist. 2nd

Ser., vol. 18, p: 319.

Mast, S. O.

1908. The Reactions of Didinium nasutum, etc., Biol. Bull., Woods Hole, Vol. 16, p. 91.

MERESCHKOWSKY, C.

1881. On Some New or Little Known Jnfusoria, Ann. and Mag. of Nat. Hist. 5th Ser.,

vol. 7, No. 39, p. 209.

MEUvNIER, A.

1910. Microplankton des Mers de Barents et de Kara, Duc d’Orleans Campagne

Arctique de 1907, Brussels.

Mincain, E. A.

1907. Protozoa, in Allbutt and Rolleston’s A System of Medicine, Lond.

MIncany, E. A.

1912. An Introduction to the Study of the Protozoa, Lond.

PRATT, H. S.

1916. A Manual of the Common Invertebrate Animals, etc., Chicago.

Rovx, J.

1901. Faune Infusorienne des Eaux Stagnantes des Environs de Genéve, Genéve.

Rovx, J.

1902. Note sur les Infusoires Ciliés du Lac Leman, Revue Suisse Zool., T. 8, fasc. 3, p.

459.

SuitH, I. F.

1914. A Preliminary Report on the Jnfusoria of Kansas, Kansas Univ. Sci. Bull.,

vol. 9, No. 13.

STEIN, F. ~

1854. Die Infusionsthiere, auf Ihre Entwicklungsgeschichte, Leipzig.

STEIN, F.

1859. Der Organismus der Infusionsthiere, 3 vols., Leipzig.

Stevens, N. M.

1901. Studies on Ciliate Infusoria, Proc. Calif. Acad. Sci., 3rd Ser., vol. 3.

Stokes, A. C.

1885. Some New Jnfusoria, Am. Nat., vol. 19, No. 5, p. 433.

STOKES, A. C.

1887. Notices of New Fresh Water Infusoria, Pro. Am. Phil. Soc., vol. 24, p. 244.

STOKES, A. C.

1888. A Preliminary Contribution Towards a History of the Infusoria of the Fresh

Waters of the United States, Jour. Trent. Nat. Hist. Soc., vol. 1, no. 3.

STOKES, A. C.

1918. Aquatic Microscopy for Beginners, 4th ed., N. Y.

WARD AND WHIPPLE ;

1918. Fresh Water Biology, N. Y.

136 LEON AUGUSTUS HAUSMAN, PH.D.

EXPLANATION OF PLATE

Fig. 1. Stain or reagent set holder.

Fig. 2. Culture oven a, switch, c, cardboard

cover, @, tin shelf, e, lamp rack, /, copper wire

for suspending shelves, g, thermometer.

Fig. 3. Strainer for concentrating samples.

Fig. 4. Isolation pipette

Fig. 5. Holophrya sp. 1, 15-25u

Fig.. 6. Holophrya sp. 2, 15-254

Fig. 7. Holophrya sp. 3, 15-25y

Fig. 8. Holophrya sp. 4, 15-25y

Fig. 9. Holophrya sp. 5, 45-55y

Fig. 10. Holophrya sp. 6, 30-35u

Fig. 11. Holophrya sp. 7, 40-45y.

Fig. 12. Urotricha farcta, 15—-25y

Fig. 13. Urotricha platystoma, 35-45

Fig. 14. Perispira ovum, 80-100y

Fig. 15. Enchelys pupa, 80-100z

Fig. 16. Enchelys farcimen, 25-50u

Fig. 17. Enchelyodon farctus, 175-225

Fig. 18. Spathidium spathula, 60-80u

Fig. 19. Prorodon armatus, 25—30u

Fig. 20. (See Plate I)

Fig. 21. (See Plate IT) :

Fig. 22. Trachelocerca olor, 320-380u

7 FRESH WATER AND MARINE INFUSORIA 137

*_ PLATE V

138

LEON AUGUSTUS HAUSMAN, PH.D.

Fig. 20.

Fig. 21.

Fig. 23.

1000.

Fig. 24.

Fig. 25.

Fig. 26.

Fig. 27.

EXPLANATION OF PLATE

Prorodon ovum, 100-130y.

Lacrymaria olor, 320—380y.

Trachelocerca phoenicopterus, 450—

Ileonema dispar, 115-125y.

Plagiopogon coleps, 75—90x.

Coleps hirtus, 45-55y.

Tiarina fusus, 75-80y.

Fig. 28. Didinium nasutum, 850-1000u

(the only species not drawn to scale. If repre-

sented in its relative proportions, it would be

more than twice and a half as large as Trachelius

ovum, Fig. 43, Plate IV).

Fig. 29.

Fig. 30.

Fig. 31.

Fig. 32.

Fig. 33. .

Fig. 34.

Fig. 35.

Fig. 36.

Fig. 37.

Fig. 38.

Mesodinium pulex, 10-20.

Mesodinium cinctum, 30-45.

(See Plate ITT).

Amphileptus gutta, 40-60y.

Lionotus sp. 1, 25-35.

Lionotus fasciola, 75-125.

Lionotus pleurosigma, 110-125y.

Lionotus wrzesniowski, 175—200y.

Lionotopsis anser, 75—100y.

Loxophyllum, sp. 1, 45-50x.

FRESH WATER AND MARINE INFUSORIA 139

willie

ey : O;

PLATE VI

140 LEON AUGUSTUS HAUSMAN, PH.D.-

Fig. 31.

Fig. 39.

Fig. 40.

Fig. 41.

120-150p

Fig. 42.

Fig. 43.

Fig. 44.

Fig. 45.

Fig. 46.

Fig. 47.

Fig. 48.

Fig. 49.

Fig. 50.

Fig. 51.

EXPLANATION OF PLATE

Tillina magna, 195-225y.

Loxophyllum setigerum, 100-125

Loxophyllum rostratum, 125-150

Trachelophyllum tachyblastum,

Flexiphyllum elongatum, 200-300u

(See Plate IV)

Loxodes rostratum, 250—350u

Nassula ornata, 200-400p

Nassula sp. 1, 30-35y

Nassula microstoma, 50—60u

Chilodon megalotrocha, 40-50u

Chilodon vorax, 50-70u

Chilodon caudatus, 35-50u

FRESH WATER AND MARINE INFUSORIA 141

PLATE Vil

142 LEON AUGUSTUS HAUSMAN, PH.D.

EXPLANATION OF PLATE

Fig. 43. Trachelius ovum, 280-350y

Fig. 44. Dileptus gigas, 450-800n

Fig. 52. Chilodon cucullulus, 125-225

Fig. 53. Phascolodon vorticella, 40-70u

Fig. 54. Scaphidiodon sp. 1, 25-35

Fig. 55. Trochilia sigmoides, 30-40.

FRESH WATER AND MARINE INFUSORIA 143

PLATE VIII

COPPER: ITS OCCURRENCE AND ROLE IN INSECTS AND

OTHER ANIMALS!

RicHarp A. MutTKowsk1, Px.D.

University of Idaho, Moscow, Idaho

. General.

. Respiratory Proteins in Insects; the Copper Nucleus.

. Copper in Other Animals.

The Sources of Copper: Soil, Water, Plants.

Discussion. :

Summary.

Bibliography.

Be wh Re

“ID On

I.. GENERAL

The present paper arose from certain experiments on the respiration

of insects, particularly on the gases of the blood, and its réle as a respir-

atory factor.

As at present understood, respiration in insects proceeds by the tracheal

method: Atmospheric air is led directly to the cells by the tracheae, while

the blood acts primarily in the transportation of food and metabolic

products. This is modified in aquatic stages, in some of which the oxygen

in solution in the water is absorbed by means of tracheal gills. In certain

secondarily aquatic insects,—that is, insects originally aquatic which

became terrestrial in habit, but which in some stages again have sought

the water,—there are found structures which fundamentally are of the gill

type. Here a thin membrane separates the blood from the water; the

blood takes up oxygen through this membrane and distributes it directly

to the tissues, or indirectly by yielding it to the gaseous supply in the

tracheae. Such structures are the gill filaments and gill pouches of

Trichoptera larvae and aquatic caterpillars, the caudal gill pouches of

Chironomid larvae, Culicid larvae, Simulium larvae, etc.

Considered from both the physiological and morphological standpoint,

these structures are meaningless unless a respiratory protein is postulated

in the insect blood to fix the transfusing oxygen. Without such a protein,

there could exist only an oxygen balance between the fluids divided by the

animal membrane,—between the oxygen in solution in the blood serum

and that in solution in the water. But since the available dissolved

oxygen decreases with the rising temperature of the water, and the tem-

perature of the insect rises with that of its environment, the oxygen

supply of the water becomes impoverished, and with it the amount in

solution in the blood,—that is, if the blood lacked a respiratory protein.

1 (Contribution from the Zoological Laboratory of the University of Idaho, Moscow,

Idaho.)

144

COPPER: ITS OCCURRENCE IN INSECTS 145

Furthermore, many insects are found in places where the available oxygen

is nearly entirely used up in the decomposition and oxidation of organic

waste. Indeed, a number of species are known which live, grow, and

transform under anaerobic conditions (Juday 1909, Muttkowski, 1918).

Now, a respiratory protein is known for a few insect species, among

them some of the anaerobes just referred to, especially the so-called “‘blood-

worms’ or Chironomid larvae. This pigment has been identified as hemo-

globin, dissolved in the blood plasma, and not included in the corpuscles.

As far as known to the writer, it is restricted among insects to the ‘“‘blood-

worm”’ type of Chironomid larvae. The important point in connection

with the hemoglobin of these larvae is this: It is confined to a few species,

but not.all of these species live under anaerobic conditions; nor do all

anaerobic Chironomids contain hemoglobin. One is forced to the con-

clusion that among these latter the hemoglobin is replaced by some other

respiratory pigment. Hemocyanin has been suggested by the writer

(1920), altho it had not been demonstrated for a single insect species.

It is known from the study of vertebrate blood that hemoglobin forms

oxyhemoglobin with oxygen and carbohemoglobin with carbon dioxide.

From its identity with vertebrate hemoglobin it can be supposed that the

activity of the hemoglobin as found in the Chironomid ‘‘blood-worms”’ is

similarly two-fold,—that it transports both oxygen and carbon dioxide.

For the rest of insects it was shown by the writer that both oxygen and

carbon dioxide are present in the blood (account published elsewhere).

Hence it is logical to assume that, similar to Chironomid ‘‘blood-worms,”’

there is a definite carrier which fixes both oxygen and carbon dioxide.

The following recounts a series of experiments undertaken in an attempt

to prove or disprove the foregoing assumption. The experiments were

performed during the spring, summer and fall of 1920, altho some earlier

observations made in the course of the past ten years are included.

II. RESPIRATORY PROTEINS IN INSECTS; THE CoppER NUCLEUS

In its development the problem presented several distinct phases:

(1) Aside from the few insects possessing hemoglobin, is a respiratory

protein available at least in those aquatic insects provided with blood

gills? (2) If such a protein can be demonstrated, what is its nature?

(3) lf available, is it confined to aquatic stages or is its distribution univer-

sal among insects?

The presence of hemoglobin in Chironomid larvae (blood-worm type)

is easily verified. For the blood responds to the various oxidation (Guiac,

O-tolidin, and Benzidine reactions) and crystallization tests (Hemin)

that have been elaborated for the recognition and study of hemoglobin in

vertebrate blood. Except for the hemin ‘test, none of these is conclusive

146 RICHARD A. MUTTKOWSKI, PH.D.

as far as general differentiation between Invertebrate and Vertebrate

blood is concerned. The hemin test alone indicates positively the presence

of hemoglobin as such in Chironomid “‘blood-worms,” or any other animal.

Yet in a number of tests made for hemoglobin in the colorless blood of

species like Anax, Aeshna, Dytiscus, and others, isolated crystals other

than Sodium chloride were found which resembled the familiar prisms of

hemin.

The oxidation tests are conclusive only in so far as they reveal the

presence of blood, specifically the respiratory protein. They do not

indicate the identity of this pigment. For it is noteworthy that the

blood of crayfish as well as that of all insects reacts with Guaiac, and

Benzidine, and produces coior changes identical with those produced by

vertebrate blood (See table I). Note that these oxidations are produced by

blood which shows little or no trace of hemoglobin. Furthermore, as with

vertebrate blood, boiling the test substance does not stop the reaction.

Tas LE I. REACTIONS OF INSECT BLOOD TO. HEMOGLOBIN TESTS

Name of Species Guiac Test Benzidine Test | Hemin Test-Crystals

(Oxidation) (Oxidation) (Nippe’s Reagent)

Number | Result | Number | Result | Number Result

Aeshna larva....... 8 pos. 3 pos. 5 4 neg., 1 trace

Anax, yg. larvae.... 2 x 1

Enallagma larvae... My 1

Mayfly nymphs.... 1

Belostoma...2:.... 3 1 7 5 neg., 2 trace

Chironomus larvae | 6 | 3 12 12 pos.

Dytiscus larvae... .| 2 6 ) neg., 1 trace

Dytiscus adults. ... Z 1 7 5 neg., 2 trace

Controls |

Cambarus blood... | 6 | 3 5 5 neg.

Blank, with FeCh. .| 5 | 3

Blank, with CuSO; | 5 oie ae “

In hemoglobin the iron is the active agent in the oxidation, in the

hemocyanin of the crayfish it is the copper. That such is really the case

was readily shown by the introduction of a crystal of ferric chloride or

copper sulphate into some of the blank control tests. Such ‘‘salted’’

controls reacted positively, before and after boiling.

The tests described proved two things: (1) The oxidation tests for

hemoglobin do not serve to differentiate between this and other respira-

tory proteins, or between the blood of Vertebrates and Invertebrates.

(2) They proved the presence of a respiratory protein in insects.

Two possibilities at once presented themselves,—that the carrier in’

question could be either hemoglobin, or hemocyanin; or both, as in some

COPPER: ITS OCCURRENCE IN INSECTS 147

mollusks. It was definitely ascertained, however, that in only a very few

insects could hemoglobin be the respiratory protein. In most insects, if

present at all, it was found only in infinitesimal quantities, and therefore

negligible as a respiratory factor. This left the alternative of hemocyanin.

This respiratory protein has been reported for a number of higher Crustacea

and some Arachnida (Scorpion, Limulus). It is also widely known among

the Mollusca, and there is no valid reason to assume that it might

not be present in other groups of animals, including the insects.

To ascertain if this is the case, both direct and indirect methods were

resorted to in this study. Unfortunately, no direct method for the recog-

nition of hemocyanin is known such as the hemin test for hemoglobin. A

large number of experiments were attempted to find such a reagent, but

all were unsuccessful. Hence an indirect method was adopted..

In hemocyanin copper forms the nucleus of the respiratory compound.

If the presence of copper could be shown in insect blood in amounts com-

parable to the copper content of equal quantities of crayfish blood,—

then it would be logical to assume that this copper forms the nucleus of a

respiratory protein similar to that of crayfish. Since, as already related,

various tests indicated the presence of extremely minute quantities of

hemoglobin, and since hemocyanin responded positively to the various

hemoglobin tests, the latter were useless for differentiation between an

iron and a copper compound. It therefore became necessary to separate

the two, and to test separately for copper. This, of course, could be

accomplished only after incineration of the tissues. ;

Among larger insects the blood and entire specimens, in small insects :

only whole specimens, were incinerated in the course of this study. The

incinerations were begun in June 1920 and continued thru the summer and

fall, as material became available. The usual analytical methods were

followed: the ash was dissolved in hot dilute hydrochloric acid, a portion

tested for iron, while the remainder was treated with excess of ammonia,

precipitating the iron and dissolving the copper. The solution was then

filtered, the filtrate concentrated by slow heating, acidulated with acetic

acid, and tested for copper. Where the amount of ash was very small,

the residue was redissolved and reprecipitated several times in order to

obtain all the copper present.

As expected, iron showed heavily in all the incinerations, as it is

universally present in animal cells. The thiocyanates were the chief

reagents used in testing for this substance. Only qualitative tests were

performed on copper, with notation of the approximate intensity of the

reaction as compared with the control substance, namely crayfish blood.

No exact quantitative estimates were possible, as the amounts dealt with

148 RICHARD A. MUTTKOWSKI, PH.D:

were microscopic. The reagents employed were Potassium ferrocyanide,

Ammonium mercuric sulphocyanate, especially after the test drop had

been inoculated with zinc salts (acetate or sulphate) or Caesium and

Rubidium chloride; and finally, the Lead acetate—Potassium nitrite

method for the formation of the triple nitrite Lead-Copper-Potassium.

The ammonia so generally employed as a test for copper was not sufficiently

delicate. It is sensitive only to about 1:2000, and the copper obtained in

the few milligrams of ash was insufficient to react with it. The other

reagents mentioned are sensitive to copper in dilutions up to 1:50000 and

over, sufficiently so to give definite reactions.

The incinerations covered practically every order of insects (see

Table-I1). The material incinerated was collected by the writer from

Paradise Creek, two or three ponds, and the fields in the immediate

vicinity of the university at Moscow, Idaho; except no. 45, Sialis larvae,

which were obtained from Lake Mendota, Madison, Wis., and kindly

sent me by Prof. Chancey Juday, of the Wisconsin Geological and Natural

History Survey.

TABLE II. Copper In INSECTS

: : : No. of Incin-| Result

Name Stage Tissue pe Ge Remarks

| tS) | |

Coleoptera

1. Dytiscus _ larva blood 2 pos Equal to Cambarus

Dee i ‘3 whole 2 pos. slightly less

oe oa adult blood 5 pos. nearly equal

4. dg a whole 4 pos slightly less

5. Gyrinus adult whole 1 pos. less

6. Harpalus adult whole 1 pos. less

7. Leptinotarsa - sf 1 pos. less than C.

Hymenoptera

8. Apis mellifica jadult whole 2 pos. less than C.

9. Bombus sp. = ng 1 pos. less

10. Polistes t Ped es 2 pos nearly equal

11. Formica i 1 pos. less

Lepidoptera

12. Pieris rapae larva blood 1 pos. about equal

gia Be - ey whole 1 pos. about equal

14, 3% 4 adult | whole 2 pos less

15. Noctuid moths |adult | whole 2 pos. less

Diptera

16. Musca domestica} larva whole 2) pos. slightly more

iS nor “jadult whole 2 pos about equal

18. Stomoxys adults E 44 pos. less

19. Tachinid flies Hg e 1 pos. less

Hemiptera

20. Belostoma

Bile yn" *

22s i

2a. ih

24. Ranatra

25. Gerris sp.

26. Notonecta sp.

27. Corixa sp.

28. Aphis sp.

Odonata

29. Anax & Aeshna

30. Anax & Aeshna

°31. Anax

33. Aeshna

34. 4

35. Sympetrum

36. ce

37. Libellula

38. .

39. Enallagma

40. igs

41. ec

Ephemeroidea

42. Several spp.

Trichoptera

43. Several spp.

Neuroptera

44. Myrmeleon

Megaloptera

45. Sialis infumata

Tsoptera

46. Termes sp.

Orthoptera

47. Gryllus

48. Ceuthophilus

49. Locusta

50. Melanoplus

oe bivittatus

51. Dissosteira

COPPER: ITS OCCURRENCE IN INSECTS

TABLE II.

yg. nymphs

adults

mixed

yg. larvae

larvae

“ce

yg. larvae

old larvae

yg. larvae

late larva

adult

nymphs

larvae

adults

larvae

mixed

adult

“ce

adult

Copper IN Insects (Continued)

Tissue

blood

whole

blood

whole

blood

whole

blood

whole

blood

whole

blood

whole

whole >

whole

149

No. of Incin-

erations

BEN WORF RUD WwW Ww

NWR WwWrRNN TORR bd DY

Se — —= WwW — wd

—- dO

Result

Cu.

Remarks

slightly less

about equal

slightly less

ce c

less

nearly equal

less

nearly equal

ce ce

less

“ce

slightly less

less

about equal

less

slightly less

nearly equal

equal or stronger

nearly equal

less

nearly equal

less

Total—34 species, 108 incinerations _

A glance at the results in Table II shows positive reactions for copper

in all insects incinerated, no matter what the stages chosen for ashing.

150 RICHARD A. MUTTKOWSKI, PH.D.

This universal presence of copper among the Insecta, not only in aquatic

forms, but also in terrestrial species, indicates that it has an important

function which hitherto has been overlooked. Its universal distribution is

certainly not adventitious. Such a contingency might be explained for

aquatic insects on the basis of the food supply (a large percentage of

Crustacea), but would hardly apply to terrestrial insects, especially those

among the latter which feed on plants only, or whose food is even more

restricted, as in the case of honey bees. Furthermore, the copper present

in the blood of many insects exists in practically the same proportions

as in the blood of Crustacea. In Belostoma, for instance, five cc. of

incinerated blood reacted to ammonia, showing a slightly fainter shade of

blue than an equivalent amount of incinerated Cambarus blood. Indeed,

copper was present to such extent, that an incineration of one cc. showed

decisive reactions with all the reagents listed except ammonia. Other

examples might be adduced, such as wasps and ant-lions. Here the ash of

a single individual gave positive response to tests for copper.

Based on the foregoing results, the writer offers the hypothesis that

the role of copper in insects is to form the nucleus of a blood protein,—

namely a hemocyanin, similar in constitution to the known hemocyanins of

Crustacea and mollusks; and that it serves a similar purpose, that of a

respiratory pigment.

Based on experiments, recorded elsewhere, on the presence of oxygen

and carbon dioxide in the blood of insects, the writer advances the further

suggestion, that the function of this hemocyanin is parallel to that of

hemoglobin,—namely, that the hemocyanin carries both oxygen and

carbon dioxide, that compounds are formed in the respiratory cycle

similar to the oxy- and carbohemoglobins. This second hypothesis has

not been proved directly, but it is logical to assume that analogy of function

in a respiratory protein, as hemocyanin is analogous to hemoglobin, should

result in analogous compounds during the respiratory cycle. In short,

it is reasonable to assume the formation of oxyhemocyanin with oxygen,

and of carbohemocyanin with carbon dioxide.

From this standpoint, the various respiratory structures of advanced

aquatic insects, such as the gill filaments and gill pouches of Trichoptera lar-

vae and aquatic caterpillars, and the others referred to in the opening

paragraph of this paper, acquire a real significance. If considered as of the

category of true gills, to which type they undoubtedly belong, it is easy to

understand how: e ective they would be with a respiratory protein.

Without such a pigment to fix the gases they would seem purposeless as

structures, and inefficient physiologically.

COPPER: ITS OCCURRENCE IN INSECTS 151

III. Copper IN OTHER ANIMALS

As already stated, hemocyanin has been reported for mollusks, several

species of higher Crustacea, scorpions and Limulus among Arachnida,

and more recently for Coelenterates and fish. To ascertain whether or not

it is found in the other classes of Arthropoda, the writer incinerated several

species of plankton Crustacea, spiders, daddy-long-legs, centipeds, and

millipeds (see Table III). In all of these copper was discovered in quanti-

ties equal to or exceeding the amount present in Cambarus blood.

As seen from this same table, examples of other phyla were also incin-

erated, including snails and slugs among mollusks, Lumbricus among

Annulata, Ascaris among Nemathelminthes, Volvox among Protozoa, and

human blcod and snake blood for Chordata.

This material also was collected locally, except nos. 13-16 inclusive,

which were collected from Wisconsin lakes, and kindly sent me by Prof.

Chancey Juday, of the Wisconsin Geological and Natural History Survey.

The determination of copper in the blood of Cambarus has been

noted repeatedly in this paper. The blood of this Crustacean was used

constantly as a control for other incinerations. Only a small number

of these controls are listed in the table. A second pigment has been

reported for marine Decapoda, called Tetronerythrin, which is found also

in our fresh water crayfish. The function of this pigment is unknown,

altho it has been stated definitely that it is not a respiratory pigment.

It is probable, however, that the pigment is used in the coloration and

markings of the exoskeleton, and that it is carried passively in the blood-

stream, similar to the pigments found in insect blood, and elaborated

during the ecdysis. It is readily perceived in crayfish blood, from which it

may be crystallized in orange-red crystals. Ordinarily it is not very

abundant, but previous to moulting it is present in quantities sufficient

to give a distinct pink or reddish color to the blood. Indeed, in larger

crayfish, exceeding four inches inlength, I have found the blood a bright red

or scarlet, so that it resembled the diluted and aerated blood of a verte-

brate. This blood clots in dark red masses, also resembling the clots of

vertebrate blood. In “‘soft,’’ freshly moulted crabs the blood appears

transparent and contains little or none of this pigment.

Among species other than Arthropod Volvox furnished perhaps the

greatest surprise by its show of copper, not merely as-a trace, but in

appreciable quantity. About 15 cc. of filtered Yolvox were used in this

incineration. That the reaction could not have been due to residual

water is indicated by the fact that 100 cc. of water from the same pond

showed not the slightest trace of this element. Its function in Volvox is

problematical.

Ascaris furnished an additional surprise. Surely no one would suspect

copper in an internal parasite. However, as barely a trace was found,

RICHARD A. MUTTKOWSKI, PH.D.

TaBLe III. Copper mn ANIMALS OTHER THAN INSECTS

Name

Crustacea

1. Cambarus sp.

2 “e

3 “c

4 cc

& “cc

6. ce

of “c

8 ce

9 “

10. ie

11. Hyalella

12. Cladocera &

Copepods

13. Daphnia pulex

14, Microcystis

15. Copepods

chiefly Limno-

calanus)

16. Daphina pulex

Arachnida

17. Argiope sp.

18. Phalaena sp.

19. Several spiders

Myriapoda

20. Millipeds spp.

21. Centipeds spp.

Annulata

22. Lumbricus

Mollusca

23. Physa sp.

24. Slugs

Nemathelminthes

25. Ascaris

Protozoa

26. Volvox

Chordata

27. Thamnophis

Sirtalises te

28. Homo sapiens

and this due probably to mechanical storage, it would hardly be justi-

fiable to attribute any physiological réle to copper in this parasite.

source is most probably the plant food taken in by the host.

No. of in-

Stage | Tissue eevee Result

cinerations

1 in. long) blood 3 pos.

1 in. long} whole 4 pos.

2 in. long} blood 4 pos.

2 in. long} - whole 3 pos.

3 in. long| blood 6 pos.

3 in. long} whole 3 pos.

4 in. long] blood 5 pos.

4 in. long) whole 3 pos.

5 in. long} blood 3 pos.

5 in. long} whole 2 pos.

adults whole 2 pos.

adult whole 1 pos.

adults whole 1 pos.

adults -whole 1 pos.

adults whole 1 pos.

adult whole 1 pos.

adults whole 1 pos.

whole 1 pos.

mixed whole 5 pos.

adult whole 2 pos.

of & Q whole 1 pos.

mixed whole 1 pos.

adult blood 1 pos.

adult blood 1 neg.

Remarks

More Cu. than in No. 1.

ce “ce ae ce

No. 3

a4 “ “ ce “e

“e a4 c ce

No. 5 {

ce “ce ce ce ce

ce “ce ce ce “ce

ce (74 “e “ce

No. 7

“ce cc ce (74 oe

less than No. 2 ;

cc “cc (a4

ce (a4 oc |

Equal to No. 9

More than No. 9:

Equal to No. 9

More than No. 9

trace

Equal to No. 3

Equal to No. 4

trace

Equal to No. 2

trace in 2.8 gr.

about 1.6 gr. used.

Its

COPPER: ITS OCCURRENCE IN INSECTS 153

In snails and slugs the copper undoubtedly occupies the same réle

that it has in squids, clams, and other mollusks. LLumbricus, like Ascaris,

showed only a trace. Aside from leeches, this is the first time copper has

been noted for an Annelid.

The fact that an abundance of copper was found in Myriapoda and in

several representatives of the Arthropoda, in some even more than in the

control substance, lends definite support to the assumption that for all

Arthropoda copper is an essential element, and functions in the réle of a

respiratory protein in all members of this largest of phyla.

The discovery of copper in snake blood was due to pure chance. Two

and eight tenths grams of snake blood besides a small quantity of human

blood had been incinerated for another purpose. While waiting to utilize

the ash at some later date it occurred to the writer to test for copper. (At

the time I did not know of Rose and Bodansky’s discovery of copper in ma-

rine fish.) The various tests were negative except two in which the test drop

had been placed under alcohol vapor for several hours. No reaction showed

in the first fifteen minutes, but after that indications of a positive reaction

were noticeable. Later, when examined after an interval of several hours,

the reactions showed definitely positive.

The ashed human blood referred to in the foregoing paragraph was also

tested, but gave negative results. However, since the quantity was even

less (1.83 gr.) than the snake blood used, the experiment is inconclusive.

IV. THE SOURCES OF COPPER

For aquatic animals the source of copper is the slight amount in solu-

tion in the water. It is thus that mollusks and Crustacea obtain the

copper necessary to their respiration. The soluble copper originates from

the soil. Since the distribution of mollusks and Crustacea is universal,

copper must likewise be available universally.

For terrestrial animals such as bees, wasps, caterpillars, moths, spiders,

centipeds, etc., the soil cannot be considered as a direct source of copper.

Their food consists largely of plants and animals, and perhaps minute

droplets of water from wayside pools. It is evident that their copper must

eventually come to them by way of their plant food. To determine this

positively, a number of plants were incinerated and tested for copper

(Table IV).

All plants reacted positively, but only to the more sensitive reagents, as

the copper is present only in traces, not at allin amounts comparable to that

of Arthropoda. All parts of the plant showed the presence of copper, with

this difference: the fruit generally contained a less amount than the stem,

leaves, or root. Because of the minimal amounts, its réle in the plant is

154 RICHARD A. MUTTKOWSKI, PH.D.

probably not an active one, and its presence due to mechanical storage.

As far as the relation of copper and plants is concerned, the copper ion is

known to be highly toxic to plants, especially to the lower forms of plant

life.

TaBLe IV. Copper In PLANTS

Name Part No. of Incinerations Result for Copper

1. Watermelon Rind J) Pos. trace

2 eebear Leaves 2 pos. trace

3p Pear fruit 2 pos. less than No. 2

4. Tomato leaves and stem 2 pos. trace

5. Tomato fruit 2 pos. less than No. 5

6. Potato leaves 1 pos.

7. Lettuce leaves y) pos.

8. Red Beet leaves 2 pos.

9. White Beet leaves 2 pos.

10. Apple leaves - 2 pos.

11. Apple fruit 2 pos. less than No. 11

12. Currant leaves 2 pos.

13 Celery, stem 2 pos.

14. Clover heads 1 pos.

15. Clover | leaves 2 pos. more than No. 14

Total—11 species, 30 incinerations.

Here in Moscow the copper content of the soil is very low. The surface

soil is of aeolian origin and what copper it contains is brought by dust-

storms from mountains 100 to 200 miles to the west in Washington and

Oregon. Ina quantitative estimate of copper in the soil 50 grams yielded

only sufficient copper to permit a qualitative test.

Samples of water, each 3500 cc., taken from Paradise Creek during

the summer and concentrated to 5 cc., likewise showed little more than

traces of copper. A sample of water taken more recently (November)

from under the ice showed a copper content of 0. 0187 gr. by the sulpho-

cyanide method or approximately 0.00534 gr. per liter. The mud of

Paradise Creek shows somewhat larger amounts, some of which may be

due to organic matter.

V. DIscUSSION

In a recent paper on the occurrence of copper in marine organisms, Rose

and Bodansky (1920) note the previous demonstration of copper in the

following groups::

1. Echinodermata—starfish, urchins, sea-squirts.

Annulata—leech.

Crustacea—various Decapoda.

Arachnida—Scorpion, Limulus.

SoS TS)

COPPER: ITS OCCURRENCE IN INSECTS 155

5. Mollusca—clams, oysters, snails, cuttle-fish, octopus.

6. Tunicata—Ciona.

7. Pisces—shark, 2 Teleosts.

To this list Rose and Bodansky add:

1. Coelenterata—jellyfish, Portuguese Man-of-War.

2. Mollusca—oysters, clams.

3. Curstacea—shrimps and crabs.

4, Pisces—Torpedo and sting ray, 12 Teleosts.

In all they add some 35 species, demonstrating for the first time that

the copper in fish is not due to pathological causes. A survey of the

groups studied in the present paper shows some interesting additions to the

foregoing lists:

1. Protozoa—Volvox.

2. Nemathelminthes—Ascaris. -

3. Mollusca—snails, slugs.

4. Annulata—Lumbricus.

5. Arthropoda.

a. Crustacea—plankton, Cambarus, Hyalella.

b. Arachnida—Phalaena, spiders.

c. Myriapoda—centipeds, millipeds.

d. Insecta—13 chief orders, over 35 species.

6. Chordata—snake.

To these must be added the occurrence of copper in higher plants.

Such a wide distribution of an element in a variety of living organisms,

representing eight of twelve phyla, must have some significance. Its

occurrence cannot be wholly adventitious, especially since it may be

present in considerable quantities in the organism. Where present only in

traces, it may well be ignored. In a number of forms its physiological réle

has been known for some time, altho physiologists believed that it was

restricted to a few scattered species, and really was more or less an abnor-

mality or rarity. Schulz, for instance (in Abderhalden 1910), states,

“‘Hemocyanin occurs in the blood of higher Crustacea. It is present only

in a few members (italics mine) of this class (Homarus, Maja, Portucuco,

etc.)”” Yet it is evident from the work of Rose and Bodansky and from

the experiments herein noted that copper is not at all restricted to a few

Decapoda among Crustacea, but that even the simplest and smallest

Crustacea contain it.

Indeed, the writer, once he found positive indications of copper in a

few species of insects, set out to test representatives of as many different

groups as he could obtain locally. These were taken wholly at random,

representing a variety of living conditions, from aquatic to parasitic, and

entirely without regard to possible favorable results. (As a matter of fact,

156 RICHARD A. MUTTKOWSKI, PH.D.

in the end I purposely selected some of the least likely animals, such as

Volvox, Ascaris, Lumbricus, and snakes.) This same attitude held for

the work on lower Crustacea, spiders, and so on. The results, I believe,

more than justified such a procedure, by indicating copper in the most

unexpected places.

Yet this very fact of random selection is all the more convincing in a

general application of the phenomena discovered. It signifies that these

random selections are representative of whole groups and that what

pertains to the few pertains also to the many. Since in these representa-

tive species copper has been found, and since in at least some of these few

the physiological réle of the copper has been definitely ascertained, it can

be concluded that all or most remaining members of these groups also

possess copper and that its physiological réle is also similar. In other

words, in at least the Mollusca and Arthropoda all species contain copper in

appreciable quantities, and this copper functions as the nucleus of a respira-

story protein, hemocyanin.

In thus extending particulate findings to entire groups of organisms I

do not think I am overstepping the bounds of proper scientific conser-

vatism. For copper was found in many groups, while in certain groups

where a greater variety of material was available every species studied

showed positive results. The uniformity of the results in these groups,

unexpected as they were, is convincing and, I believe, warrants the above

generalization.

VI. SumMMARY

i Both oxygen and Carbon dioxide are present in insect blood in

appreciable quantities. Insect blood aids in the transportation of gases,

and the respiratory function is not confined to the tracheae.

2. Over 100 incinerations were made of insect blood, or whole speci-

mens, and the ash tested for copper. Copper was found in all cases.

The 35 species studied represent 13 of the chief orders of insects, in both

larval and adult stages.

3. Copper is found in insect blood in quantities comparable to that

of crayfish blood. Its réle is therefore interpreted as being identical,—

namely that it serves as the nucleus of a respiratory protein,—hemocyanin.

Insects, therefore, have two sources of oxygen,—atmospheric air led

directly to the tissues by way of the tracheae, and fixed oxygen carried by

the respiratory protein of the blood.

4. Incinerations of plankton Crustacea, spiders, and centipeds gave

positive results for copper, showing that copper is distributed among all

classes of Arthropoda. It may therefore be regarded as an element

essential to the physiological activity of Arthropods, its réle being to act

in a respiratory pigment for all members of this phylum.

COPPER: ITS OCCURRENCE IN INSECTS Wey

_ 5. As representatives of other phyla Volvox, Ascaris, snails and slugs,

Lumbricus, human and snake blotd were incinerated. All of these, except

human blood, showed varying amounts of copper.

6. As sources of copper the water, soil, and plants were tested. All

plant ash showed traces of copper. The water samples of this region

showed only small quantities, while the soil showed varying amounts.

VII. BrBLIoGRAPHY

GuERITHAULT, B. Sur la présence du cuivre dans les plantes et particuliérement. dans les

matiéres alimentaires d’origine végetale. C. R. Acad. Sci., Paris, 171, pp. 196-198

(no. 3), 1920.

Jupay, C. Some Aquatic Invertebrates that Live Under Anaerobic Conditions. Trans. Wis.

Acad. Sci., 16, part I, pp. 10-16, 1909.

Motrxkowsk1, R. A. The Fauna of Lake Mendota. Trans. Wis. Acad. Sci., 19, part I,

pp. 374-482, 1918.

Muttkowski, R. A. The Respiration of Aquatic Insects: A Collective Review. Bull.

Brooklyn Ent. Soc., 15, pp. 89-96, 131-140, 1920. Bibliography.

_MourtrKxowskt, R. A. Copper in Animals and Plants. Science, N.S., 53, No. 1376, pp. 453-

454, May 13, 1921.

Mourrxkowsk1, R. A. Studies on the Respiration of Insects. I. The Gases and Respiratory

Proteins of Insect Blood. Ann. Ent. Soc. Am., 14, pp. 150-156, 1921.

Rose, W. C., and Bopansxy, M. Biochemical Studies on Marine Organisms.

I. Copper in Marine Organisms. JN. Biol. Chem., 44, pp. 99-112, 1920. Bibliography.

Scuutz, F. N., in Aberhalden, HB. d. Biochem. Arbeitsmethoden, Vol. II, pp. 335-346, 1910.

DEPARTMENT OF METHODS, REVIEWS, ABSTRACTS,

AND BRIEFER ARTICLES

MICROSCOPE ILLUMINATION WITH REFERENCE TO BROWN-

IAN MOVEMENT AND COMBINATION LIGHTING!

ALEXANDER SILVERMAN

BROWNIAN Movement.—For the study of this phenomenon china

clay was mixed with distilled water and passed through a double quanti-

tative filter paper. The opalescent filtrate was run into the cavity of

a hollow-ground slide, and this in turn placed over dull black paper on

the stage of the microscope. A 4mm. apochromat objective surrounded

by a ring-lamp” was lowered into the solution and the instrument focused.

Eve

resell

1 Published by courtesy of the American Chemical Society. Read before the Division

of Industrial and Engineering Chemistry at the Rochester, N. Y. meeting of the American

Chemical Society, April 25-29, 1921.

2 J. Ind. Eng. Chem., 9(1917), 971; 10(1918), 1013; 12(1920), 1200.—J. Soc. Chem. Ind.,.

38(1919), 126.—J. Royal Mic. Soc., No. 253 (1920), 98.

158

_ DEPARTMENT OF METHODS 159

A 10x compensating ocular gave a magnification of 440 diameters. Not

only was Brownian movement clearly evident, but the dot-like, rod and

lenticular shapes of particles were shown with great definition in white

against a black background.

CoMBINATION LicHtinc.—A comparative study was made of (1) the

effect of transmitted light, (2) of direct super-stage illumination from

a ring-lamp surrounding the objective, and (3) of combination lighting

from above by the same lamp, a part of whose light passed through a glass

slide and was reflected back by a sub-stage mirror placed parallel to the

stage.

For this study a fossil insect in amber was employed. The length of

the insect, 0.9 mm., will enable the reader to judge the magnification in the

illustrations. A combination of 32 mm. objective and 10x ocular together

with the length of bellows used in photographing gave a magnification of 60

diameters in the original photographic prints.

Figure 1 shows the effect of transmitted light from below, placing the

mirror at an angle and using a 25 watt frosted spherical lamp. While

sufficient contrast is ob-

tained details are lacking

because of the varying

thickness of the insect.

Figure 2 shows the effect

of cirect light from the

ring-lamp above the object

wit) dull black paper

placed. under the slide.

Some details’ are visible,

but there is no contrast.

Beautiful detail may be

obtained by substituting

white paper for black.

Figure 3 is the result of

a combination of light from

the ring-lamp above, with

its own light reflected up- Fic. 2

wards by the sub-stage mirror placed parallel to the microscope stage. A

black paper was held over the sub-stage mirror after 15 seconds. This

160 ALEXANDER SILVERMAN

Hess

, Department of Chemistry,

University of Pittsburgh.

affords contrast and detail,

and the method may prove

desirable for photographing

an object which has trans-

parent, translucent and

opaque parts.

Seeds Grafiex plates,

exposed 60 seconds, with a

Davies shutter closed to

the smallest stop, gave the

results obtained in the in-

sect photographs.

Conclusions.—This

paper emphasizes the im-

portance of a study of

background colors and of

combination lighting in-

microscopy and cites specific

examples. .

TRANSACTIONS

OF THE

American

Microscopical Society

ORGANIZED 1878 INCORPORATED 1891

PUBLISHED QUARTERLY

BY THE SOCIETY

EDITED BY THE SECRETARY

PAUL, S:. WELCH

ANN ARBOR, MICHIGAN

VOLUME XL

NuMBER [’ouRr

Entered as Second-class Matter August 13, 1918, at the Post-office at Menasha

Wisconsin, under Act of March 3, 1879. Acceptance for mailing at the

special rate of postage provided for in Section 1103, of the

Act of October 3, 1917, authorized Oct. 21, 1918

The Collegiate Press

GEORGE BANTA PUBLISHING COMPANY

MENASHA, WISCONSIN

1921

TABLE OF CONTENTS

For Volume XL, Number 4, October, 1921

On the Effect of Some Fixatives upon Myxosporidian Spores, with four figures, by

New Species and Collections of Arrhenuri: 1921, with three plates by Ruth Marshall.... 168

Some Work on Marine Phytoplankton in 1919, by W. E. Allen....................... fie

The Accessory Chromosome of Anasa tristis again, by A. M. Chickering .............. 182

DEPARTMENT OF MetHops, REVIEWS, ABSTRACTS, AND BRIEFER ARTICLES

Dhesiterature of Diatoms = by, Hred\BssPaylor. ss ace =e ee eee 187

A Compendium of the Hosts of Animal Parasites contained in Ward and Whipple’s

Fresh-water Biology, compiled by HZ J- Van:Cleave.. .- 5-2 24.0222 ae -eee 195

The Endocrines, by S. W. Bandler, reviewed by T. W. Galloway................-- 200

East of:members:. 30a 2e «caches BG hes 6 7 eo eee 206

TRANSACTIONS

American Microscopical Society

(Published in Quarterly Instalments)

Vol. XL OCTOBER, 1921 ; No. 4

ON THE EFFECT OF SOME FIXATIVES UPON MYXOSPORIDIAN

SPORES!

R. Kupo

Although it has generally been recognized that when myxosporidian

spores are fixed, stained and mounted as either smears or as section pre-

parations, they appear smaller than in the fresh state, it was Cépéde who

first called attention to the matter. He recognized differences between

fresh and stained spores of the species he studied, and concluded as follows

(Cépéde, 1906:63): “en présence de telles différences de taille et de

Vimportance donnée actuellement aux dimensions des spores des Myxos-

poridies dans la systématique comme caractére distinctif des espéces, je

crois utile de faire remarquer qu’il serait bon d’indiquer si les mensurations

des spores ont été faites in vivo ou sur des préparations fixées et colorées,

et montées au baume.”’

My observations upon the species which I have studied up to date

agree with Cépéde’s results and have suggested that the dimensions of

spores of a species should be accompanied by the statement of conditions

under which the measurements were made (Kudo, 1920:49).

Most authors agree that at the present state of our knowledge regard-

ing this group of Protozoa, a satisfactory classification of genera and species

of Myxosporidia, must have as its basis the study of the spore (Kudo,

1920:52-59). The size, dimensions and structure of spores show a certain

amount of variation even in one species, yet they are far more typical of

the species than are the vegetative forms. In every case, the identification

of a species has been successfully done only when the spores were present.

Since the characters of the spore vary to a more or less recognizable

extent, according to the difference of conditions under which the spores

are observed, it naturally follows that the characters of spores of two

different species can only be correctly compared when they are observed

1 Contributions from the Zoological Laboratory of the University of Illinois, No. 191.

161

162 R. KUDO

under exactly the same conditions. In other words, if the spores were origi-

nally observed and measured in the fresh state, those of the species to be

compared with the former, should also be studied in the same condition.

This kind of comparison, however, has happened only in a few cases, and

can not always be carried out even in future. Many species of Myxo-

sporidia have accidentally been found and described from stained smears’

or section preparations only by several authors, who were engaged with

studies on other topics. Furthermore, even if one deals exclusively

with Myxosporidia, one is frequently compelled to omit the study of

fresh spores and to confine oneself to that of stained preparations under

various circumstances.

The characters of spores observed only in section preparations can, of

course, not only be compared properly with those obtained from fresh

spores, but also should not be used for the data of establishing a new species.

Special precaution must be exercised in cases where two forms are similar

in habitat and locality and whose vegetative stages are not known.

Recently Schuurmans Stekhoven (1920) described three new species

of Myxosporidia, Sphaerospora gasterostei, Myxidium rhomboideum and

Henneguya renicola, found in the-uriniferous tubules of the kidney of Gaster-

osteus pungitius (misprinted as pungiticus) from Holland. The author

studied section preparations of the host kidneys which were fixed with 60

per cent. alcohol and stained with Delafield’s hematoxylin, and compared

the characters of the spores observed therein with those of already known

three species, Sphaerospora elegans, Henneguya media and Henneguya

brevis (Kudo, 1920:30). Thélohan’ (1895) who described these latter

forms, seems to have studied them in both fresh and fixed conditions,

although his description is unfortunately brief. When the forms are so

similar in every respect except the size of the spores, one finds it extremely

difficult to decide whether the newly observed species studied only in

sections, are identical with the former or not. If, however, we can calcu-

late the dimensions of spores in the fresh state from those obtained from the

stained ones, we can undertake a more satisfactory comparison between

the species observed under diverse conditions.

In order to see exactly how the fixation, staining and mounting would

affect the shape, dimensions and structure of Myxosporidia, a few ex-

periments were performed on the spores of Leptotheca ohlmacheri. This

' Myxosporidian was first found by Ohlmacher and Whinery (1893) in the

kidney ot Bufo lentiginosus. I have recently observed it in the kidneys of

Rana clamitans and Rana pipiens. Although I formerly placed it pro-

visionally in the genus Wardia (Kudo, 1920:83-84), my recent study on its

morphology and life history which will be published elsewhere, has shown

that it should be placed in the genus Leptotheca as Labbé suggested.

EFFECT OF FIXATIVES UPON MYXOSPORIDIAN SPORES 163

The fresh spore of Leptotheca ohlmacheri (Figs. 1 to 3) is oblong with its

longest diameter at right angles to the sutural plane. The anterior end is

slightly attenuated due to the thickening of the spore membrane at that

point, while the posterior end is rounded. In profile, it is nearly circular

with the slightly attenuated anterior extremity. In the anterior end

view, it is regularly oblong. The shell is moderately thick. The sutural

Spores of Leptotheca ohlmarcheri (Gurley) Labbé. Figs. 1, 2 and 3. Three optical

sections in front, side and anterior end views respectively of fresh spores which were kept in a

hanging drop preparation with physiological solution and which are typical of the species in

form, structure and dimensions. Fig. 4. A spore from a smear fixed with absolute alcohol,

stained with Giemsa stain and mounted in cedar oil, showing the shrinkage of the entire body.

All about X 2100.

ridge is well marked and protrudes conspicuously at the ends. The

spore membrane is somewhat irregularly striated. Three to seven strie

run parallel to the sutural line on each valve, the remaining ones make

somewhat similar angles with the former. The strize in lateral view are

mostly placed horizontally. The number of strie on each valve varies

from 25 to 35. Two spherical polar capsules usually equal in sizé in one

spore, occupy the anterior portion of the spore. The polar filament, coiled

4 to 6 times, is distinctly visible. Two independent sporoplasms occupy

the extracapsular cavity of the spore. They are extremely homogeneous

and each to be the karyosome of the nucleus). The size of the spore varies

to some extent. There are some larger and some smaller than the average

spores as is the case with every species ‘ound, doubtlessly due to the mal-

formation. The average dimensions are as follows:

Sutural diameter ; 9.5 to12u; average 10.8u

Breadth 13 to 14.54; average , 13.75p

Thickness 9.5 to12u; average 10.8u

Diameter of polar capsule 3.5 to 4.5 uw; average 4.0

Length of polar filament (KOH) 42 to 62u; average 52.0u

164 R. KUDO

A drop of the emulsion of fresh spores in physiological salt solution was

smeared on a slide. The amount of the emulsion and the area over which

it was smeared were made approximately the same in every smear so as to

obtain the similarity in the number of spores and the conditions under

which the spores existed until they were fixed. Soon after the smear was

made, it was fixed in one of the following fixatives before it dried up one

smear being purposely made to dry up: 50 per cent., 70 per cent. and

absolute alcohols, 4 per cent. formol, Schaudinn’s (warmed), Bouin’s and

Flemming’s (weak) mixtures. The fixatives were allowed to act upon

the smears for 16 hours, after which they were removed from the latter by

proper washings. At the same time, small pieces of the infected host

organ were fixed in Schaudinn’s fluid and 4 per cent. formol respectively,

sectioned in paraffin and stained with Heidenhain’s iron hematoxylin.

The fixed smears were stained with Heidenhain’s iron hematoxylin, Dela-

field’s hematoxylin or Giemsa stain, and mounted in Canada balsam or

cedar oil (for Giemsa stained smears only).

From each of the preparations, one hundred mature spores which

could be distinguished distinctly from those that are in the course of devel-

opment, were drawn at a scale of 2,100 magnification, and measured. The

results are as follows:

| |

; Sutural Diameter | 3readth Diameter of |

| Polar Capsules

Range |Average| Loss | Range |Average| Loss | Range |Average} Loss

in in in inp in ingM| inp in w in

Fresh spores (control). .......|9.5-12.0 10.8| 0 \13-14.5 13.75) 0 3.5-4.5 4.0] 0

Air-dried, unstained, alcohols, |

and balsame . senceiaeee|t0— OF oO 8.6 | 2.2 \10.5-11.4 10.7 | 3.05 |2.6-3.2 ZEON tea:

' 150% alcohol; Giemsa......... 19 .2-10.0 OS Wate |10.5-10.9 11.0 | 2.75 |3.0-4.0 375| OES.

70% alcohol; Delafield........ 9.0-9.8| 9.4] 1.4 | 9.8-10.9] 10.4] 3.35 j2.8-3.9] 3.4! 0.6

Absolute alcohol; Giemsa.... . 9.0—- 9.5 9.25] 1.55 | 9.5-10.9} 10.2 | 3.55 |2.8-4.0 3.4) 0.6

n | | |

& 4% formol; Giemsa.......... I8.9- 9.3 9.1 Wt 9.5-11.4] 10.5 | 3.25 |2.9-3.8 3.4!) 0.6

‘SS |Schaudinn; Giemsa or Heiden-|

n Hains. seen cee oriser oe i9.2- 9.8] 9.6) 1:2 |10.9-11 9) 11.4) 2.35°|3.0-3.8 3.4] 0.6

Wi Ninouin SHedeaiaan, em ls.5s-9.5] 9.0] 1.8 | 9.5-10.9] 10.2 | 3.55 |2.64.0] 3.3] 0.7

Flemming; Heidenhain....... j8.8- 9.6] 9.2] 1.6 |10.0-11.2} 10.6 | 3.15 |2.4-3.2| 3.3) 0.7

Averages ce. 2 12 rye seer iis = |Bramterarers 0) 245 (ale Sieg | eee ects 1026) eSa098| fees eee 3.4| 0.6

Per cent. of loss, calculated

from the dimensions of | 8

Sporesi{romismears series ayer eee |e ia ROA Peete SH) [aie - otter 290%) 5 reer | eee 17.09

— !

Schaudinn; Heidenhain....... 8.6-10.0 923))) 257 1086=11,28)) 11126575132 0—-36 SES Ons

A 4% formol; Heidenhain....... \8.4— 9.9 OPIS OST ROE S13!) MOM ise Son2E8—oe8 eed! (U5 7/

Gt | Average. sthanweccee sat otek 3 a 1) Na bast | Sa ee 10)75)/23200) heer 3.3] 0.7

oO EE

Hal Per cent. of loss, calculated| |

from the dimensions of A

spores in sections. ....... | i ee ees US Alacigopocsalehooans 2830 Sb ier: cu ee 21.0%

EFFECT OF FIXATIVES UPON MYXOSPORIDIAN SPORES 165

From the above table, the following may be noted:

1) The amount of loss in sutural diameter of the spore of Leptotheca

ohlmacheri is greatest when the spore is airdried and smallest when it is

fixed with either Schaudinn’s fluid or 50 per cent. alcohol. The average

loss amounts to about 14 per cent. of the sutural diameter of the fresh

spores.

2) The amount of loss in breadth of the spore is greatest when the spore

is fixed with Bouin’s fluid or absolute alcohol and smallest when it is fixed

with Schaudinn’s fluid. The average loss amounts to about 22 per cent. of,

the breadth of the fresh spores.

3) The amount of loss in the diameter of polar capsule is greatest when

the spore is air-dried and smallest when it is fixed with 50 per cent. alcohol.

The average loss amounts to about 15 per cent. of the diameter of polar

capsules in fresh spores.

4) The losses in smear and section preparations are almost similar.

In the case of Myxobolus cycloides, Cépéde gave the following dimen-

sions: Fresh spores: sutural diameter, 13.5-16u, breadth 11-13u4. Schau-

dinn-Heidenhain’s spores: sutural diameter 10.5-12u, breadth 7.5-8y.

Thus in this case, the fixation caused loss of 3.54 and 4.25 respectively in

sutural diameter and breadth of the spore. These losses amount to 25

and 35 per cent. compared with the fresh spores.

The loss in the latter species is greater than the former species. It,

however, shows distinctly from these two different types of Myxosporidia,

that the sutural diameter undergoes a smaller amount of shrinkage than

the breadth.

Concerning the change in the dimensions of spores by fixation, Cépéde

states that the reason why the spores appear smaller in fixed and stained

conditions than in fresh condition, is simply because the refractive power

of Canada. balsam makes the unstained spore membrane invisible, and not

because the shrinkage of the body caused by fixation takes place.

In the case of Leptotheca ohlmacheri, this is not the case. Very fre-

quently spores such as shown in Fig. 4, are seen in the smears. The spore

undoubtedly occupied the entire area which appear as blank zone before

it was fixed. When fixed, the contents underwent a strong shrinkage, thus

leaving a clear unstained zone between it and the smear. In such a

spore, one can distinctly see the spore membrane in an irregular outline.

I have noticed similar change of spores in smears of many species from

various genera suggesting shrinkage as the main cause for the loss in

dimensions. I, therefore, consider the decrease in the dimensions of myxo-

sporidian spores, in general, is caused by the shrinkage of the entire spore

body under the influence of the fixative and subsequent treatments.

The amount of shrinkage caused by fixation will apparently be different

in different genera and species. Unless a large number of measurements

166 R. KUDO

on different species be made, we do not know exactly the data on which the

dimensions of fresh spores of the species observed by Schuurmans Stek-

hoven may correctly be calculated. Assuming that the spores of Sphaero-

spora gasterostei and Henneguya renicola underwent shrinkage similar

to that of Leptotheca ohlmachert, we obtain the following comparison:

Sphaerospora gasterostet Sphaerospora elegans

Schuurmans Stekhoven Calculated Thélohan

Length 6.7 pw 7.8 pw 10 uw

Breadth 7.0 p 90 u lp

| Length of

polar caps. 3.5 pu 4.1

If the calculation is correct, it seems probable that Sphaerospora gas-

terostei is independent from S. elegans.

Henneguya renicola Henneguya media Henneguya brevis

’

Stekhoven

Length of :

spore | 8 u 9.28 wu 20-24 wu (?)

Breadth Sy 45u 5-6 p

Schuurmans Calculated Thélohan Thélohan

Polar caps- |

sule (length)| 4.5 u 5.4 u 4-5 u

Tail Persie 17.4 he

The calculated value of Henneguya renicola resembles closely the dimen-

sions given by Thélohan for Henneguya brevis. The form of spore becomes

so highly modified in section preparations that it is hard to make out the

form in the fresh state. Therefore, it is highly doubtful whether Schuur-

mans Stekhoven saw a new species or not.

The form of spores changes to a variable extent according to the dif-

ference of the fixatives used. The more shrinkage the spore undergoes,

the more irregular outlines it assumes. Careful fixation in Schaudinn’s

fluid often preserves the form of spores very nicely. .

When a spore is fixed with any one of the fixatives, the coiled polar

filament becomes entirely invisible. This is probably caused by the

coagulation of the wall of polar capsule which becomes opaque by the

EFFECT OF FIXATIVES UPON MYXOSPORIDIAN SPORES 167

fixation. The distinction between the two sporoplasms is harder to deter-

mine in fixed and stained spores than in fresh spores. The sporoplasms

become coarsely reticulated, losing the homogeneous condition seen in the

fresh state.

In conclusion, I may again suggest that a new species should be de-

_ scribed after studying the spores in fresh as well as fixed and stained con-

ditions and if possible the fixed vegetative forms.

SUMMARY

1) The ordinary fixation causes about 14 and 22 per cent. decrease

respectively of the sutural diameter and breadth of fresh mature spores of

Teptotheca ohlmachert.

2) The possibility of calculating the dimensions of fresh spores from

those of fixed and stained spores is discussed.

3) The decrease in the dimensions of spores is due to the shrinkage of the

entire spore body.

4) Fixation makes the coiled polar filament invisible.

Works CITED

Kuno, R., 1920. Studies on Myxosporidia. A synopsis of genera and species of Myxospor-

idia. Ill. Biol. Monogr., 5:245-503, 25 pl. and 2 textfig.

SCHUURMANS STEKHOVEN, JR., J. H., 1920. Ueber einige Myxosporidien des Stichlings.

Arch. Protist., 41:321-329, 1 pl.

NEW SPECIES AND COLLECTIONS OF ARRHENURI: 192%

RutH MarsHALL

Rockford College

The genus Arrhenurus, the largest group of the water mites, continues

to yield new material from collections in lake regions. The new species

described in this paper came from regions as far apart as Canada and China;

while additional notes on already described species are based on material

secured in several states of northeastern United States, some of them from

new localities. Through the kindness of Professor N. Gist Gee, of Soochow

University, material was secured for the description of new species from

China. Professor Frank Smith and Dr. H. R. VanCleave, of the Univer-

sity of Illinois, were good enough to contribute some material from Michi-

gan, New York and Massachusetts. Through the interest of Dr. R. A.

Muttkowski an opportunity was given for the examination of some col-

lections of the Biological Station of the United States Bureau of Fisheries

at Fairport, lowa; and more recently the author was privileged to see some

collections of Dr. F. A. Stromsten, of the University of Iowa. The

author’s own collections from the Muskoka Lake region of Ontario,

together with other material from various sources, form the basis of

a preliminary account of the genus Arrhenurus as it has been found in

Canada. This topic will be discussed first.

Practically the only account of the water mites of Canada so far is

that contained in a paper by Dr. F. Koenike, ‘‘Nordamerikanische Hy-

drachniden,”’ and a revision of this paper, ““A Revision of my ‘Nordameri-

kanische Hydrachniden.’’’ The descriptions were based upon material

sent to him by Dr. J. B. Tyrell, of Toronto, and were collected in Alberta

and British Columbia, near the international line. Of the thirty species

listed by Dr. Koenike, four were Arrhenuri and new species. These were

A. lautus, A. inter positus (a young male), A setiger and A. krameri. The

last named species has since been found by the author and further notes

are given in this paper. In addition to this, one new species is now added

for Canada (A. uniformis nov. spec.), and four more are recorded for the

first time, as follows. A. americanus Mar. and A. manubriator Mar. were

found by the author at Parry Sound and A. americanus var. major Mar. in

a small lake near Bala, Ontario. <A. marshalli Piers., the most widely

distributed species, had previously been found in material from Long

Point, Canada, in some collections of the United States Fish Commission.

In the descriptions of species which follow the Canadian material will

be discussed first.

168

NEW SPECIES AND COLLECTIONS OF ARRHENURI: 1921 169

Arrhenurus uniformis nov. spec.

Be PX tie: 123)

This species resembles A. scutiliformis Mar. and belongs in the group of

“long-tailed”’ Arrhenuri in which the very long and rather simple appendix

is decidedly narrower at the end than it is at the base. The outline of the

body is approximately circular; the enclosed dorsal area and the appendix

are moderately high and rounded. Details of structure given in the

figures show this to be a new species. The last joint of the fourth leg is

long and slim; the spur on the fourth joint is moderately developed.

The single male on which this description is based is 1.33 mm. long

and 0.73 mm. wide. The color is dull olive green. It was found in a small

lake near Long Lake, at Bala, Ontario, August 25, 1920.

Arrhenurus kramert Koenike.

Pl. IX, fig. 7-9.

The author has already recorded (1908) the finding 0° a mite from

Oregon which appeared to be A. krameri. Drawings of this specimen are

now given for the first time and its identity with the single male from

British Columbia on which Dr. Koeinke’s description was based seems to

be established. It is slightly smaller, however, measuring only 1.29 mm.

A dorsal view is shown, which did not appear in the original paper, together

with the lateral view and a drawing of the palpus.

Arrhenurus simulans nov. spec.

Pl Xho t/=2 te

Material sent to the author by Dr. H. J. Van Cleave from collections in

Dump Lake, Woods Hole, Massachusetts, contained sixty individuals of

this species. It was at first thought to be A. krameri, altho a larger form,

the length being 1.45 mm. and the extreme width, 0.83 mm. The males

bear some resemblance also to A. rectangularis Mar., especially when a com-

parison is made of the side views of the long appendix, the end of which in

the three named species shows a double scallop, one part above the other.

The body of the new species is conspicuously elevated where it joins the

appendix. The wing-shaped genital areas are rather small and the ends

of the line enclosing the dorsal area are far behind them on the appendix.

Twenty-seven males were present. The color in the preservation is dull

brown green.

Over half of the individuals in the collection were females; the examina-

tion of the palpi shows that they belong to this species. The body of

A. simulans fem. is broadly ovate. The epimeral plates are relatively

small; the third and fourth have about the same width throughout, the

two posterior groups being well separated from each other and from the

genital area. The fourth epimera are narrow, scarcely wider than the

third. The genital plates are of nearly uniform width and extend straight

out from the aperature, which is large. The body is 1.32 mm. in length.

170 RUTH MARSHALL

Arhenurus pseudosetiger nov. spec.

Pl. IX, fig. 4-6.

In a former paper (1910) the author identified as A. setiger Koen. an

individual from Madison, Wisconsin. But a more careful study of this

specimen shows that it belongs to another, though closely related species,

which will be designated as A. pseudosetiger. The body proper is stouter

than it is shown in Dr. Koenike’s figures of the Canadian species, being

nearly circular in outline, not oblong, and the appendix is smaller. The

dorsal enclosed area runs over on the appendix and is depressed. The

entire length of the body is 0.8 mm., the greatest width, in the region of the

fourth leg,0.7 mm. The color is deep brick red.

Arrhenurus trifoliatus Marshall

PIATX, fig. 10-12:

It is not often that the collector succeeds in securing a large number of

Arrhenuri at any one time; it is still more unusual to find any one species in

numbers large enough to make possible a thorough examination of all

structures and to identify with certainty the females of the species. Col-

lections made by the author in the marshy sloughs at Burlington, Wiscon-

sin, July 5, 1919, consisted largely of individuals of the rather uncommon

species, A. trifoliatus, twenty-five males and fifteen females being secured.

It is now seen that the earlier description of the species (1908) did not show

completely the details of the appendix of the male. As seen in Fig. 10,

a young male, a delicate bladder-like structure, A, is attached to either

side of the stout petiole, a structure which is easily in ured in preservation.

The female, which is now described for the first time, is broadly oval

inform. The posterior groups of epimera are close together and the geni-

tal area lies immediately behind them. The genital wing-shaped areas

are unusual in form, the outer ends curving strongly upward. The

length of the body is 1.15 mm., the extreme width, 1.05 mm.

A. major was represented in this Burlington collection by one male.

The Lake Beulah region which was visited at the same time yielded four

species of Arrhenuri: A. marshalli, A. megalurus, A. americanus and A.

reflexus, the latter having the unusual color of orange red.

Arrhenurus compactilis Marshall

Pl. X, fig. 13-16.

This somewhat rare species was found in collections from Fairport,

Iowa, and again in the collections of Professor Frank Smith from Douglas

Lake, Michigan. This new material adds two states to the range of the

species and makes possible a more complete study of its structure. Draw-

ings of the palpus and the last leg are now given. The latter is seen to

be very characteristic of this group of stout bodied petiolated Arrhenuri

in having a long fourth joint with a conspicuous spur ending in a tuft of

curved hairs, while the fifth and sixth segments of this appendage are short.

NEW SPECIES AND COLLECTIONS OF ARRHENURI: 1921 7a

The female of the species is now known from the study of the palpi

which agree in all details with those of the male except that they are some-

what larger, as usual. The body is stout and oval and measures 1.3 mm.

in length. Details of the dorsal and ventral suriaces are shown in the

figures.

The collections of water mites from the Biological Station at Fairport,

summer of 1917, have already been referred to. They were secured from

small lakes in the vicinity of the Station and were rich in Arrhenuri. This

material is especially interesting since there have been no previous records

of Arrhenuri from Iowa that the author is aware of. Nine species were

found, as follows:

A. marshalli Piers. A. fissicornis Mar.

A. americanus Mar. A. compactilis Mar.

A. americanus var. major Mar. A. laticaudatus Mar.

A. apetiolatus Piers. A. dentipetiolatus Mar.

A. birget Mar.

The last named species is rare, only two other specimens, from Colorado,

having been recorded.

The collections of Dr. F. A. Stromsten, mentioned in the introduction,

add A. lyriger Mar. to the list.

The following descriptions of two new species of Arrhenuri from

China continue the study of the water mites of the region of Soochow which

was begun in a former paper (1919), and are made possible through the

continued interest of Professor N. Gist Gee who furnished the material.

Arrhenurus soochowensis nov. spec.

Pl. XI, fig. 22-25

The new species belongs to a type of Arrhenuri which is seen in A.

kraepelini Koen. described from Java, a type apparently common in the

Asiatic members of the genus. This is the type of A. forpicatus Neum.

of Europe, represented in America by A. lyriger Mar. It is characterized

by a deep incision in the end of the appendix, the hyaline appendix lying

on the dorsal side of this.

The single individual upon which this description is based has these

characteristics well marked. The appendix is relatively long; the median

incision runs into a round opening over which lies the large and highly

developed hyaline structures, closely resembling the same parts in A.

limbatus Koen. of Madagascar. The dorsal enclosed area of the body is

rather small and the line which encloses it is not quite closed. The fourth

joint of the fourth leg has a short spur. The fourth joint of the palpus is

unusually broad at the distal end. This is a small mite, 0.8 mm. long and

0.5 mm. wide. The color is dull green. The specific name refers to the

locality where it was found.

1D RUTH MARSHALL

Arrhenurus geet nov. spec.

Pl. XI, fig. 26-29

This species resembles A. madaraszi Daday found in Ceylon and

belongs to the forpicatus group mentioned under the last species. The

appendix is small and narrow but well developed; its median incision is

large at the upper end. The hyaline appendix is an oblong structure;

on either side of it is developed a delicate claw-like piece. The body is

broad; the enclosed dorsal area is large, the line enclosing it not quite

closed posteriorly. The fourth leg lacks the spur on the fourth joint.

The palpus, as in the related species, has a broad fourth joint. The single

male on which this description is based is 0.73 mm. long and 0.56 mm.

broad. The species is dedicated to Professor Gee.

LITERATURE CITED

KoENIKE, F. 3

1895. Nordamerikanische Hydrachniden. Abh. natur. Vereins Bremen, 13:167-226.

1912. A Revision of My ‘“‘Nordamerikanische Hydrachniden.” Trans. Canadian

Institute, Toronto, p. 281-296

MarsHALL, R.

1908. The Arrhenuri of the United States. Trans. Am. Mic. Soc. 28: 85-134.

1910. New Studies of the Arrhenrui. Trans. Am. Mic. Soc., 29:97-110.

1919. New Species of Water Mites of the Genus Arrhenurus. Trans. Am. Mic. Soc.,

38 :225-281.

EXPLANATION OF THE PLATES

Plate IX

1. Arrhenurus uniformis, dorsal view.

2. Arrhenurus uniformis, genital area.

3. Arrhenurus uniformis, lateral view.

4. Arrhenurus pseudosetiger, dorsal view.

5. Arrhenurus pseudosetiger, lateral view.

6. Arrhenurus pseudosetiger, left palpus

7. Arrhenurus krameri, dorsal view.

8. Arrhenurus krameri, lateral view.

9. Arrhenurus krameri, left palpus.

10. Arrhenurus trifoliatus, appendix of young male.

11. Arrhenurus trifoliatus, female, epimera.

12. Arrhenurus trifoliatus, right palpus.

Plate X.

13. Arrhenurus compactilis, female, dorsal view.

14. Arrhenurus compactilis, female, ventral view.

15. Arrhenurus compactilis, male, part of left fourth leg, the last three joints rotated.

16. Arrhenurus compactilis, right palpus.

17. Arrhenurus simulans, female, epimera.

18. Arrhenurus simulans, male, dorsal view.

19. Arrhenurus simulans, appendix, ventral view.

20.

PANE

bdo hm & bh bo

Ww hm

NEW SPECIES AND COLLECTIONS OF ARRHENURIT: 1921

Arrhenurus simulans, lateral view.

Arrhenurus simulans, left palpus.

Plate XI.

Arrhenurus soochowensis, dorsal view.

Arrhenurus soochowensis, lateral view.

. Arrhenurus soochowensis, ventral view.

Arrhenurus soochowensis, left palpus.

Arrhenurus geel, lateral view.

Arrhenurus geei, dorsal view.

. Arrhenurus geei, appendix, ventral view.

. Arrhenurus geei, left palpus.

173

RUTH MARSHALL

174

PLATE IX

NEW SPECIES AND*COLLECTIONS OF ARRHENURI: 1921 175

PRATEEX

176 RUTH MARSHALL

PEATE Xd

SOME WORK ON MARINE PHYTOPLANKTON IN 1919

W. E. ALLEN

Scripps Institution for Biological Research of the University of California.

On September 1, 1919, efforts to use tow nets for quantitative work

with phytoplankton in the La Jolla area were abandoned and the resources

of the Scripps Institution available for such work were temporarily con-

centrated upon collection and study of a series of catches made from our

pier by the measured water method at intervals of twelve hours. These

catches were taken by the simple procedure of dipping water from the

surface of the sea at a point about one thousand feet from shore and imme-

diately pouring it through a filtering net made of number 25 bolting silk.

This net was made in the form of a funnel into the small end of which a

bottle or other receptacle could be tied. After filtration catches were

preserved in formalin for quantitative study at convenience.

Quantitative studies consisted in roughly approximate identifications

of the species present and in an enumeration of representatives of each

found in a certain fractional part of a catch mounted in a Sedgwick-Rafter

counting cell. Necessary aids to this work were a Whipple eyepiece

micrometer and a mechanical stage. Records were later assembled in

the form of tables which were studied with reference to occurrence and

prominence of species in the locality at the time and through the period

of observation.

It is interesting to note that the locality in which this work was done

was much farther south than points at which extensive studies of phyto-

plankton have been made in Atlantic waters, i. e., it is in about the lati-

tude of Northern Egypt. Furthermore the fact that catches were made

regularly and continuously for months at intervals of twelve hours (8 a. m.

and 8 p.m.) marks the series as somewhat different from most other

groups of catches of marine plankton material.

Surface water temperatures taken at the time of making the catches

showed a range from 23° C. in August to 13° C. in December, but the

range within the limits of discussion of this paper was from 208° C. in

September to 13° C. in December.

Although there are other plants which take some part in synthetic

activities in the open sea and although dinoflagellates of many kinds are

generally analytic rather than synthetic in character, it has appeared from

preliminary studies of marine plankton in the Southern California region

that the groups most promising for quantitative study as synthesizing

organisms are the diatoms and dinoflagellates, (or at least the armored

dinoflagellates). There seems to be no reasonable doubt that these two

Las

178 W. E. ALLEN

groups of organisms are on account of their small size, large numbers,

rapid growth, facile reproduction, wide distribution and cosmopolitan char-

acter peculiarly favorable objects for quantitative study especially since

they seem to be the most easily and continuously accessible of all marine

organisms.

Both groups show rhythms and pulses of production which are more or

less evident in each month of the year. Such rhythms and pulses are,

however, characterized by changes in prominence of particular species

according to season and according to certain other variable conditions.

For convenience in the present discussion a pulse may be defined as a

marked increase in numbers of organisms which extends over a period of

three or more days before decreasing to or near the numbers found at its

beginning. In this four month period there were five such pulses of

diatoms and four of dinoflagellates. In both groups they were unevenly

distributed in the period. The records show that out of four times possible

for coincidence of pulses of the two groups there were two close approxi-

mations to coincidence. This, of course, indicates that both groups of

organisms are sometimes favored by the same stimuli to production.

But it is true that some other evidence indicates different possi-

bilities, e. g., there were fourteen cases in which a catch of diatoms was

more than three times as large as either the catch preceding or the one

succeeding it and there were fifteen such cases of dinoflagellate catches.

Out of fourteen chances for coincidence of such catches in the two groups

only four occurred, a fact which leads one to think they may be to some

extent mutually deterrent. This view gains support from the fact that

catches distinctly low in numbers as contrasted with those catches nearest

them show no coincidence in five chances although we might expect that

there would be coincident absence in both groups if both were similarly

responsive to changes in local conditions. In view of such considerations

one seems to be driven to the provisional assumption that plants in the

open sea like those on land may sometimes find such generally favorable

conditions that widely diferent types may live and thrive together without

prejudice but that usually some factor has given one form a better oppor-

tunity than another which may be used to the detriment or to the complete

exclusion of that other.

The above mentioned exhibits of presence and absence are still more

suggestive in regard to the perennial assumption that marine organisms

are uniformly distributed through considerable areas of marine waters.

A catch markedly larger than both of those at twelve hours from it or a

catch markedly smaller than both surely indicates that distribution is not

uniform in the given area.

Since catches were taken rather early in the day and early in the night

the records were examined for evidence of greater productivity in light

SOME WORK ON MARINE PHYTOPLANKTON IN 1919 179

or darkness. In October and November, two months out of the four,

about four-fifths of the larger catches were made in the morning and in the

other two months there was not much difference. So far as this limited

evidence goes it favors the view that growth and reproduction occurs

most vigorously at night, as might be expected from our general knowledge

of distribution of plant activities in the twenty-four hours.

Although many species of diatoms and dinoflagellates may be found in

the Southern California region there are not many which are ever very

prominent or numerically important and there are very few which are

frequently and continuously thus important. Since most of these can be

identified fairly well under ordinary conditions of examination, statistical

study is not seriously hampered by the requirements of taxonomy.

For most purposes it is best to study the distribution of diatoms and

dinoflagellates as separate groups. ‘Thus considered the following points

may be noted concerning diatoms: Some representatives of the group were

to be found throughout the period although distribution was very irregu-

lar. Large numbers appeared in the last three months of September thus

producing an autumnal maximum similar to those noted in European

waters.

In connection with this maximum I was interested in noting that for

two or three days previous to its inception there had been rather strong

and constant currents from the north. I also noticed that large numbers

of mackerel came to the vicinity of our pier in the latter part of August

and left about the time that the great increase in diatom prduction began.

Whether these points were mere coincidences or whether they had signift-

cant relationship to increased diatom production, I have no means of

knowing

Forty six species of diatoms were recorded in the four months but only

twelve of these were readily identified although fourteen were usually

approximated, i.e., confusion limited to only one or two other forms.

These included most of those of numerical importance. Eleven forms

were found to have been represented in the most abundant five in one or

more months. Five of these belonged to the genus Chaetoceras. They

were mostly rather small species and difficult to identify.

Detailed study of the records has clearly shown the important fact

that when there is increased production of the most prominent forms there

is also increased production of the less prominent forms and an increase

in the number of different forms. Such facts naturally lead to the assump-

tion that conditions favorable to high productivity of diatoms in the sea

affect a large number of forms in the same way. They also lead to the

inference that determination of the species which shall lead in production

is largely due to biological factors such as rapid multiplication and vigorous

development. }

180 W. E. ALLEN

As to the dinoflagellates I may say that they are usually much fewer

in numbers than are the diatoms. Otherwise the general features of their

‘distribution are not greatly different except in the periods of maximum

production. In the last four months of 1919 the greatest numbers were

produced in November but there had also been some heavy production

in August several weeks before the maximum production of diatoms.

Thirty-seven forms of dinoflagellates were recorded eight of which

were fairly easy to identify. Usually satisfactory identification of twelve

forms could be made and these included most of those showing numerical

importance. Six easily identified species were found amongst the five

‘most numerous in one or more of the four months. Two of them belonged

to the genus Ceratium.

Two of these most prominent species deserve special mention because

of their connection with the phenomenon called “‘red water.” Gonyaulax

polyedra Stein has at various times been mentioned as responsible for

extensive areas of ‘‘red water’ in Southern California which have attracted

especial attention because of the bad odor where it was washed upon the

beaches and because of the large number of littoral animals killed by it and

then stranded upon the beaches. The brownish or reddish color of the

‘water is due merely to the vast numbers of these small organisms present

in it. The destruction of littoral animals is usually said to be due to

products of decay after death of such quantities of the microscopic organ-

ism. But it is possible that the living Gonyaulax is also poisonous to ani-

mals. In the last three or four years Prorocentrum micans Ehr. has been

more often detected as a cause of ‘‘red water’ than has Gonyaulax but no

cases have been reported in which littoral animals died as a result of its

presence. It is noticeable that in “red water’’ areas (some of which extend

for miles in open water) very few other organisms, large or small, are

found amongst those which cause the discoloration.

Several different kinds of dinoflagellates cause the appearance of

water called “phosphorescence.’’ More or less glow of this sort may be

‘observed in waters of our section at almost any time of year although not

continuously present. At times there are present in the water sufficient

numbers of individuals of this sort to cause at night a glowing pathway

where fishes stimulate them by swimming through.

A more detailed report of this work in 1919 is awaiting publication in

another place. Its conclusions may be briefly stated as follows:

First, the measured water method seems to be by far the best to use

‘for a standard method and the surface level to be the best for a standard

level of collecting for quantitative study. Other methods and other

‘levels should be regarded as special methods more or less supplementary to

‘the standard.

SOME WORK ON MARINE PHYTOPLANETON IN 1919 181

Second, there is evidence that drift currents have pronounced influence

on phytoplankton production at our pier.

Third, large numbers of phytoplankton organisms respond to condi-

tions of production favorable for any one.

Fourth, it seems probable that some of the more prominent forms

may be useful as indicators of certain conditions in the ecologic complex.

Lastly, it is evident that the problems of the ecologic complex of the

sea are fascinating as well as intricate and baffling and that in many ways

good returns are sooner or later to be expected as the results of time and

energy expended in study upon them.

THE ACCESSORY CHROMOSOME OF ANASA TRISTIS AGAIN

A. M. CHICKERING

Albion College, Albion, Mich.

During the examination of literature in connection with cytological

studies on other Hemiptera I became much interested in the case of Anasa

tristis. I was astonished to find a very marked variation in the results of

the cytologists who have studied the spermatogenesis of this form. There-

fore, when in the summer of 1919 there occurred an excellent opportunity

to procure an abundance of material, I decided to make some obser-

vations of an independent nature. It would seem not out of place occa-

sionally to examine some of the commonly accepted cases and particularly

where, as in this instance there has been a decided disagreement.

Altogether ten investigators have worked on the male germ-cells of

Anasa tristis. At one time or another four of this number have been

opposed to the now generally accepted view, first stated by E. B. Wilson.

Two of these have corrected their former statements and now agree with

the latter in his conclusions.

Paulmier (’99), who was the next after Henking (91) to study the

history of the accessory chromosome, decided that the spermatogonial

number of chromosomes was twenty-two. He discovered the pair of

m-chromosomes in the spermatogonia and described their behavior. He

believed these united in synapsis to form a single condensed bivalent

chromosome-nucleolus which persisted throughout the growth period and

became the small central tetrad of the first maturation division. Further-

more he stated that this tetrad divided equally in the first division but

that the products of this division passed undivided to but one of the poles

of the second spindle giving ten and eleven chromosomes respectively to the

spermatids. He therefore identified the chromosome-nucleolus of the

growth period as the microchromosome bivalent and thought this to be

identical with the accessory.

Montgomery (’01, ’04) followed Paulmier in giving the spermatogonial

number of chromosomes as twenty-two. He also regarded the accessory

chromosome as being derived by a fusion of the m-chromosomes. A

re-examination of his material after the publication of Wilson’s second

paper on chromosomes led to a change in his statements so that they

were then in agreement.

After a very careful study of Paulmier’s material as well as his own Wil-

son came to the following conclusions in a remarkable series of papers

(05, ’06, ’07, 711); (1) that there are twenty-one and not twenty-two

182

THE ACCESSORY CHROMOSOME OF ANASA TRISTIS 183

chromosomes in the spermatogonia; (2) that these chromosomes exist in

pairs and this leaves one without a mate; (3) that this odd one is one of the

three largest and is the so-called accessory chromosome; (4) that this

chromosome exists as a chromosome-nucleolus throughout the growth

period; (5) that the m-chromosomes previously identified by other obser-

vers as the accessory have an entirely independent history and divide in

both of the maturation divisions; (6) but that the real accessory divides

in the first and then passes undivided to one only of the two spermatids

derived by division of the secondary spermatocytes, thus giving rise to two

kinds of spermatozoa.

Foot and Strobell (07), using smear methods to the entire exclusion

of sections and illustrating only with photo-micrographs, took sharp issue

with Wilson. ‘These investigators asserted that Paulmier was right in his

spermatogonial count; that the so-called chromosome-nucleolus of the

growth period is but ‘‘morphologically the equivalent of a nucleolus”

or in other words the plasmosome; that there is no odd or accessory chrom-

osome; that what has been called such is but a lagging chromosome which

divides in each division as do all the others; that therefore all spermatids

receive eleven chromosomes.

This disagreement among cytologists, of course, became a serious

matter. Many fundamental facts came into direct question and conse-

quently several people were interested enough to make independent

investigation of the conditions.

Closely following the papers of Foot and Strobell there appeared a brief

treatment of the question by Lefevre and McGill (08). Their observa-

tions confirmed those of Wilson.

In connection with work on the chromosomes of some of the coreid

Hemiptera Morrill (10) confirmed Wilson’s spermatogonial count.

The climax of the researches on the accessory chromosome of Anasa

came in 1910 when McClung and Pinney went over the whole matter with

great care. Miss Pinney made an entirely independent study and in order

to avoid bias or prejudice in the matter refrained from reading any of the

accounts published by other investigators until her own conclusions had

been reached. McClung studied the original material of Paulmier, Wilson

and Lefevre and McGill. Both McClung and Pinney agreed that the

spermatogonial number is twenty-one. They further agreed with Wilson

that there are ten bivalent chromosome and one, the accessory, which ts

univalent in the metaphase of the first division. This univalent body

exists as a short, heavy thread, a compact mass or finally as a straight

longitudinally split rod all through the prophase stages of the first division.

It divides as do the others in the first but does not divide in the second

mitosis.

184 A. M. CHICKERING

In making this brief and confirmatory study of Anasa tristis! Ihave

used the ordinary cytological methods now somewhat standardized. As

usual I have found Bouin’s and Flemming’s fluids very valuable. Perhaps

the best preparations have been made with Bouin’s. The iron-hematoxy-

lin method of staining has again proven the best general stain although I

have had difficulty with the domestic preparations

My own conclusions are not enough different from those of Wilson to

warrant a lengthy treatment. In fact as regards the important stages the

matter might be dismissed by a statement that the facts as I see them are

as stated by Wilson, Lefevre and McGill, and McClung and Pinney. In

order, however, that my results may be on record and that the constancy

of the chromosome relationship within the species may be further evidenced

I will state the main facts as I see them.

There are without question twenty-one chromosomes in the normal

spermatogonia. I have examined dozens of these cells in the metaphase

when the chromosomes are well spread out but I have never found one

which clearly showed more than the expected number. The three large

bean-shaped chromosomes and two small m-chromosomes are always

present together with sixteen others of about equal size.

In the late prophase stages of the first division at least nine typical

tetrads can be seen accompanied by what I think are three dyad bodies.

I am sure one of these is the accessory. Probably the others are the m-

chromosomes but the study of these has not made me entirely sure of this.

When the chromosomes become placed in the metaphase plate preparatory

to division the accessory usually occupies a position outside of the ring

formed by the position of the nine ordinary tetrads. The m-chromo-

somes, now unquestionably formed into a tetrad, occupy a central position

within the ring. All the bodies divide equally in this division. As a

result then all secondary spermatocytes possess eleven chromosomes each.

When these become arranged in the flat metaphase plate again the ring-

like arrangement is succeeded by an irregular placing. In spite of this

the accessory can usually be identified. When the second division occurs

this accessory does not divide like the rest but goes to one pole undivided

thus giving rise to two kinds of spermatids. Lateral and polar views of

these stages show without the shadow of doubt that half the spermatids

receive ten while the other half get eleven chromosomes.

At this point I will mention a condition observed several times in Anasa

and other Hemiptera. Follicle cells in division frequently show double the

normal spermatogonial number of chromosomes. How this is brought

about and what the fate of the cells involved is I am not able to state.

1 For the identification of my material I am indebted to Dr. Paul S. Welch of the Univ.

of Michigan, Ann Arbor, Mich.

THE ACCESSORY CHROMOSOME OF ANASA TRISTIS 185

It is amazing that there should have been so much disagreement in the

results of investigations up to this point. As is often the case in the Hemip-

tera when the preparations are good most of the stages so far outlined stand

out with diagrammatic clearness. There should be no further difference

of opinion in regard to these matters. Anyone can demonstrate the

truth of the statements by using reasonable care in preparation and obser-

vation.

When we come to the question of the behavior of the accessory chromo-

some from the time of the last spermatogonial division down to the late

prophases of the first maturation division the facts are more difficult to

determine. Those who have worked upon the question of spermatogenesis

will agree that it is often difficult to discover just what is going on in the

nucleus at this time. However, I believe the main contentions of Wilson

can be proven true

In many cells before and during synezesis a compact body can be

found just outside of the more or less tangled mass of chromatin threads.

Because of its size and appearance I think it reasonable to conclude that it

is the accessory chromosome. In every stage following this the same body

can be identified and need not be confused with the plasmosome which

occurs with it for a large part of the growth period. Thus the accessory

maintains its individuality throughout the maturation process.

In conclusion I would say then that I believe the current explanation

of the “‘case of Anasa tristis’”’ as given by text-books 0 zoology and genetics

based upon the work of Wilson is correct. This classical example may be

regarded as a permanent addition to our stock of knowledge. Our inter-

pretations may change, of course, but the facts will stand as Wilson stated

them.

The different stages of the spermatogenesis of Anasa are on the whole so

clear and beautiful that I can recommend them for class use where it is

desired to demonstrate the main facts connected with the behavior of the

sex chromosome.

LITERATURE CITED

Foor, KATHERINE and STROBELL, E. C.

1907. The ‘Accessory Chromosome” of Anasa tristis. Biol. Bull., Vol. XII.

A Study of the Chromosomes in the Spermatogenesis of Anasa tristis. Amer.

Jour. Anat., Vol. VII. :

HENKING, H.

1891. Ueber Spermatogenese und deren Bezeihung zur Entwicklung bei Pyrrochoris

apterus. Zeitschrift. wiss. Zool. Band LI.

LEFEVRE, GEORGE and McGIr1, CAROLINE. -

1908. Chromosomes of Anasa tristis and Anax junius. Amer. Jour. Anat. Vol. VII.

McCune, C. E. and Pinney, Ebru.

1910. An Examination of the Chromosomes of Anasa tristis. Kan. Univ. Sci. Bull.,

Vol. V.

186 A. M. CHICKERING

MontcomMery, T. H.

1901. A Study of the Germ-cells of the Metazoa. Trans. Amer. Phil. Soc., Vol. XX.

1904. Some Observations and Considerations upon the Maturation Phenomena of the

Germ-cells. Biol. Bull., Vol. VI.

1906. Chromosomes in the Spermatogenesis of the Hemiptera heteroptera. Trans.

Amer. Phil. Soc., Vol. XXI.

Morrit, C. V.

1910. The Chromosomes in the Odgenesis, Fertilization and Cleavage of the Coreid

Hemiptera. Biol. Bull., Vol. XTX.

PAULMIER, F. C.

1899. The Spermatogenesis of Anasa tristis. Jour. Morph. Supplement, Vol. XV.

Witson, E. B.

1905. Studies on Chromosomes, I. Jour. Exp. Zool., Vol. II.

1905. Studies on Chromosomes, II. Jour. Exp. Zool., Vol. II.

1906. Studies on Chromosomes, III. Jour. Exp. Zool., Vol. IIL.

1907. The Case of Anasa tristis. Science, N. S., Vol. XXV.

1911. Studies on Chromosomes, VII. Jour. of Morph., Vol. XXII.

DEPARTMENT OF METHODS, REVIEWS, ABSTRACTS,

AND BRIEFER ARTICLES

THE LITERATURE OF DIATOMS

Frep B. TAYLOR

Bournemouth, England

The literature on the subject of diatoms is both extensive and expen-

sive; many of the books are long out of print; much is scattered through

periodicals English and foreign; and there is no recent general hand-book

at a moderate price. The nearest approach to such a volume is in German,

it is the part on Peridiniales and Bacillariales in Engler and Prantl’s

Pflanzenfamilien, published at Leipzig in 1896. It is written by F.

Schuett, and gives an account of the morphology and biology of diatoms,

with a scheme of the genera divided into Centricae and Pennatae, a

description of each genus then established, and a drawing of one or more

species to illustrate the genus.

The Diatomaceae of the Hull District by Mills and Philip contains

several plates covering most of the species commonly found in- England.

There is also a most instructive paper by Philip in the Transactions of the

Hull Scientific and Field Naturalists’ Club vol. IV. part iv (1912) p. 205,

on Diatoms of the Humber.

For fresh water diatoms Die Siisswasser Flora Deutschlands, Oester-

reichs, und der Schweiz, part 10, by H. von Schénfeldt deals with diatoms,

it is published by Fischer of Jena, and contains many figures of English

diatoms.

The early history of our knowledge of the Diatomaceae is given by

Ehrenberg in the introduction to the Bacillaria in his first great work,

Die Infusionsthierchen, published in 1838. Kitton in Science Gossip,

1880, pp. 78 and 133, gives a résumé of this introduction, which describes

the work of Ehrenberg himself and other diatomists. Kitton also gives

an interesting account of the views of. Corda (1835) in Science Gossip,

1882, pp. 6, 22. The imperfection of the instruments then available, and

the then commonly received belief in the animal nature of diatoms often

led to the enunciation of opinions, which now appear ridiculous.

Kuetzing’s Kieselschaligen Bacillarien, (1844), also contains an histori-

cal preface. This was translated by Professor H. L. Smith oi Geneva,

U.S. A. in nos. 2 and 4 vol. II of The Lens. A résumé of this translation

with comments of his own was given by Kitton in Science Gossip, 1874,

pp. 2, 25, 149.

187

188 FRED B. TAYLOR

Ehrenberg may be regarded as the father of diatomology, he was

able to command a vast quantity of material; and although many of

his views are now known to be erroneous, and many of his figures are incor-

rect or insufficient owing to the imperfect objectives he used, and to the

want of sufficient magnification, his labors will always be of great value

to the diatomist. About one third of the genera now recognized were

founded by him. After him Greville and Grunow are the most prolific

creators of genera, accounting for about another third between them.

Diatom literature may be taken as starting with Agardh’s Systema

Algarum in 1824, and Ehrenberg’s early papers on the Infusoria, in which

he included diatoms, published in 1829-1832. Kuetzing’s Synopsis

Diatomearum followed in 1834, and in 1838 Ehrenberg brought out his

great work on Infusorial Animalcules, Die Infusionsthierchen, nine of the

64 plates being devoted to diatoms.

Kuetzing’s second work, Die Kieselschaligen Bacillarien, 30 pl. was

published in 1844; the following year saw the first edition of Pritchard’s

Infusoria, further editions were issued in 1852, 1861, and 1864. The

part on the Diatomaceae was written by Ralfs, and plates iv to xvii give

figures of diatoms. This is, perhaps, the book on diatoms most readily

accessible to English students, and though portions of it are out of date or

incorrect, it forms a very useful introduction to the subject.

Rabenhorst’s Siisswasser Diatomaceen, 10 pl., appeared in 1853.

Owing to the imperfection of optical apparatus many of the figures in these

earlier works are wanting in detail. It is therefore often difficult to

determine the identity of the species named and described; yet it is won-

derful how much was seen and accurately recorded with instruments that

would now be despised and rejected.

The last half of the nineteenth century was a period of great activity

in this branch of research. From the year 1853 to 1866 the Transactions

and Journal of the Microscopical Society, and the Transactions of the

Royal Society of Edinburgh contain a number of papers by Brightwell,

Gregory, Greville, Lauder, Roper, Walker-Arnott, Wallich and others,

many of which are beautifully illustrated by Tuffen West. William

Smith’s Synopsis of British Diatoms, containing 69 plates by the same

artist was published in 1853-1856; Gregory’s Diatoms of the Clyde ap-

peared in 1857 (Proc. Royal Soc. Edinburgh, vol. xxi, p 473). The

figures were drawn by Greville.

Ehrenberg’s: colossal Mikrogeologie appeared in 1856. Besides his

two large books he wrote numerous papers on diatoms in the Transactions

of the Berlin Academy of Science. Naturally in the course of the progress

of knowledge, and with the improvement of instruments, and the increase

of material, the early conceptions of many genera and species have been

modified, and relations have been acknowledged between forms at first

DEPARTMENT OF METHODS 189

sight widely differing. The Micrographic Dictionary first appeared in

1854, with enlarged editions in 1859, 1872, and 1882. It contains several

plates of diatoms.

Greville’s fine series of 20 papers on New and Rare Diatoms, mostly

from Trinidad, Barbadoes, Moron, Monterey, and Ceylon, appeared in

the T. M. S. and Q. J. M. S. from 1861 to 1866, with monographs on

Asterolampra, Campylodiscus, and Auliscus. He also wrote on Diatoms

from Hongkong (A. M.N.H. 1865), the South Pacific (Edin. N. P. J.

1863) and The Tropics and Southern Hemisphere, (T. B.S. Edin. and

Edin. N. P. J. 1865, 1866). A collection of poor photographic reproduc-

tions of 81 plates mostly Greville’s from T. M. S., etc., compiled by Moe-

bius was published in New York. It is sometimes on the market.

In America J. W. Bailey wrote on American Bacillaria, and published

several papers between 1841 and 1860. L. W. Bailey also wrote on the

subject, and numerous papers by Dr. A. M. Edwards appeared in various

periodicals between 1859 and 1877; his sketch of the Natural History of

the Diatomacea is dated 18/4. (cis Bull=Mforr.-Bot. Club, 1374 p21 34):

Lewis wrote in 1861 on New and Rare Diatoms, and in 1865 on White

Mountain Diatoms.

Cleve, a Swede, began to write in 1864, and published various papers

and books, many of them in English, on Diatoms from Spitzbergen, the

Sea of Java, West Indian Archipelago, Greenland and Argentina, and

Finland; also on Arctic Diatoms, and New and Little Known Diatoms, and

on the diatoms found by the Vega Expedition, and later, several papers on

Plankton Diatoms. His great work is his Synopsis of the Naviculoid

Diatoms published in 1904, 1905. In this last book he proposes a re-

arrangement of the Naviculae and the related genera setting up a number

of new genera, which have not been universally accepted; although many

of them are recognized as useful subdivisions of the older genera; some

writers, however, accept his proposals en masse.

Grunow, an Austrian, wrote from 1860 to 1890; his papers on New

Diatoms, and on Austrian Diatoms (1860, 1862, 1863, and 1882 and

1883) are important contributions to diatom classification; and much of

his work is embodied in Van Heurck’s Synopsis of the Diatoms of Belgium,

and in Schmidt’s Atlas. His account of the diatoms of the Novara

Expedition is dated 1867; the same year saw his paper on the diatoms of the

Sargasso Sea of Honduras, an abstract of this is givenin M. M. J. 1877,

p- 165. He also wrote on Caspian diatoms, and conjointly with Cleve a

book on Arctic diatoms, followed by his Diatoms of Franz Josef Land.

Kitton wrote from 1868 to 1884 a number of articles, many of them

in Science Gossip, a magazine which during the years 1867 to 1877 printed

several useful papers on the subject of diatoms, including some by Kitton

on North American deposits. Kitton translated some of Grunow’s

190 FRED B. TAYLOR

papers for various English periodicals; among these were the Novara

diatoms, which appeared in Grevillea in 1872.

Harting’s Banda See, Janisch on Guano diatoms, Janisch and Raben-

horst on the Marine Diatoms of Honduras, Heiberg’s Danish Diatoms,

and Schumann’s Prussian Diatoms belong to the early sixties.

In the seventies followed De Brébisson on Diatoms contained in Cor-

sican Moss. He had previously written on diatoms from the Cherbourg

littoral, and wrote other papers.

Donkin published only three parts, (12 plates), of his Natural History

of the British Diatomacee. About the same time appeared O’Meara on

Irish Diatomacee (poor plates) and on diatoms from Kerguelen’s Land;

and Petit on Table Bay, Campbell Island, Cape Horn, etc. In 1872

Pfitzer wrote on the structure and development of diatoms; an abstract

of this important paper is given by O’Meara in Q. J. M.S. 1872, p. 240.

The Lens, published in Chicago, only lived two years, 1872, 1873. It

contains valuable articles by Professor H. L. Smith and other American

diatomists; among these is the professor’s Classification of diatoms into

Rhaphidiee, Pseudo-Rhaphidiee, and Crypto-Rhaphidiee, which was

followed by Van Heurck and De Toni, and by most diatomists until the

present division into Centrice and Pennate, (a modification of it) ap-

peared. Many French diatomists follow Pfitzer’s division into Placo-

chromatice and Coccochromatice, which is based upon the nature of the

endochrome.

In 1874, Adolf Schmidt, a canon of Aschersleben, commenced his splen-

did work, Atlas der Diatomaceenkunde. This is published in parts of four

plates issued at irregular intervals. An index to the first four series, 240

plates, has been published. Up to August 1914 parts 1 to 79, containing

plates 1 to 316 had appeared: parts 80-83 have been published since the

war. Adolf Schmidt died in 1901, but the Atlas has been continued by his

son and by Fricke, Heiden, and Hustedt, with the assistance of Cleve,

Grunow, and other diatomists.

J. Brun of Geneva entered the field in 1880 with his Diatomées des

Alpes et du Jura; following in 1889 with Diatomées Fossilies du Japon

written in collaboration with Tempére; and in 1891 with Diatomées,

Espéces Nouvelles. The plates in the two last are excellent: the first

commences with an important sketch of the natural history of diatoms.

In 1886 Count Castracane’s report on the diatoms found by H. M.S.

Challenger was issued by the British Government. This contains 30

plates and valuable notes. Peragallo’s Villefranche appeared in 1888

and Wolle’s Diatomacee of North America (112 plates) was published

in 1890 The plates are mostly copied from other authors, and are far

behind the originals in execution; but the book is useful for reference, and

is generally procurable at a moderate price, considering its size.

DEPARTMENT OF METHODS 191

Le Diatomiste edited by Brun, a periodical devoted to the study

of diatoms, only lived from 1890 to 1896. It contains many notable

contributions including monographs on Pleurosigma and Rhizosolenia

by Peragallo, on Entogonia by Bergon, and on the miocene diatoms of

Barbadoes by Brun and Barbo.

Le Micrographe Préparateur under the editorship of J. Tempére ran

from 1893 to 1906. It contains valuable papers on the structure and

reproduction of diatoms, on the movement of diatoms, and on mounting

and cleaning. But the most important is Les Diatomées Marines de

France, of which some 115 p!ates were published in this periodical; the

complete work, 137 plates, was separately published in 1908.

Leuduger-Fortmorel between 1879 and 1898 published books on the

diatoms of the north coast of France, Ceylon, Malaysia, and West Africa.

In 1886-1887 in the Journal of the Quekett Microscopical Club appeared

the well known paper on the Oamaru deposit in New Zealand by Grove

and Sturt; also in 1886 Pantocsek published the first volume of his great

work on the fossil-diatoms of Hungary. The second volume appeared in

1889, and the third in 1892. The whole contains 102 plates. In the third

volume are many new forms from Kusnetzk in Russia, and from fossil

deposits in Japan. A second edition was issued in 1903-1905. Pantocsek

has also written on the diatoms of Lake Balaton (Platten See), and of

Kertsch, and of Szliacs in Hungary, (1902,3). About this time Walker and

Chase, and Kain and Schultze wrote on diatoms in America, the latter pair

bringing to notice the interesting deposit of Atlantic City.

Van Heurck’s Synopsis des Diatomées de Belgique, (1880-1885) con-

tains 141 plates, and is a work of the greatest value. His Treatise on

Diatoms translated into English by Dr. Wynne Baxter, was published

in 1896 before the original; it contains 35 plates illustrating all the species

found in the North Sea and the neighboring countries; and in the text are

descriptions with typical figures of all the genera known at the time of

publication, some 193 in number. Otto Witt in 1885 gave an account of

the diatoms from the marine deposits of Ananino and Archangelsk in the

province of Simbirsk in the interior of Russia.

Rattray’s monographs on Aulacodiscus (J. R. M.S. 1888); Auliscus

(J. R. M.S. 1888); Actinocyclus (J. Q. M. C. 1890); and Coscinodiscus

(Proc. Royal Soc. Edin. 1890) are standard authorities.

About the same time appeared Pelletan’s Les Diatomées, he was

assisted by Deby, Petit and Peragallo. The account of the natural history

of diatoms is good, and there are numerous illustrations of the most

common species and genera. Truan and Witt’s book on the fossil deposit

of Jérémie in Haiti is dated 1888. Another work of great utility, though

it is not illustrated, is Les Diatomées du Mond Entier, issued by Tempére

and Peragallo as a companion to the series of 625 slides from various locali-

192 FRED B. TAYLOR

ties distributed by them. It gives lists of the diatoms found by them on

the slides, and is a valuable aid to identification. A second series of

1000 slides is accompanied by a volume bearing the same title, the second

edition of this is dated 1915; it is of course more comprehensive.

Deby’s monograph on Campylodiscus appeared in 1891; with three

or four exceptions the figures are copied from other works. About the ~

same time a portion of Janisch’s report on the diatoms of the Gazelle

Expedition was privately distributed to certain favoured diatomists

without text or list of contents. This work has never been completed;

out of 22 plates numbers 7, 8, 10, 12, 13, 14, 17, and 18 are wanting.

Many of the specimens figured are reproduced in Schmidt’s Atlas.

In 1891 to 1894 De Toni, a professor of the University of Padua,

published his monumental work, Sylloge Bacillariearum as part of his Sylloge

Algarum. This book has no illustrations, but it contains in Latin de-

scriptions of all the genera and species described or named at the date of

publication, with their synonyms and varieties, and references to all

published figures and drawings. Nearly 6000 species are admitted. The

book also contains a bibliography to date of the literature of diatoms by

Deby.

Frére Héribaud of the college of Clermont-Ferrand wrote in 1893 on

the diatoms of Auvergne. The volume commences with a short, but very

clear, instructive, and succinct account of the subject, forming an admir-

able introduction to the study of diatoms. Between 1902 and 1908 he

published four memoirs on the fossil diatoms of Auvergne.

In 1896 Schuett wrote the part on diatoms in Engler and Prantl’s

Pflanzenfamilien, (lieferung 143-145). This book, with its one or more

typical figures of each genus, and its account of the morphology and

biology of diatoms, forms a most useful handbook at a moderate price.

It gives a description of all known genera classified as Centric and Penna-

te, the division now generally adopted; the Pennate are since 1902

subdivided into Mobiles and Immobiles, the Centrice are all Immobiles.

This last distinction is due to Mereschkowsky, a Russian diatomist:

(Script. Bot. Hort. Imp. Petrop. 1902) and A. M. N. H. 1902, p. 65).

Karsten’s book, Die Diatomeen der Kieler Bucht, 1899, is praised by

Cleve and Mereschkowsky as a vade mecum for students of living diatoms.

In the present century we have Dippel’s book on the Rhine and Maine

districts, 1905; and a most useful and instructive report by Mann on the

diatoms found in the Pacific by the U. S. ship Albatross, published by the

Smithsonian Institute in 1907. Von Schonfeldt’s Diatomace Germaniz

was published in the same year. ;

Peragallo’s magnificent work Les Diatomées Marines de France, (13

plates, 1897-1908), and Meister’s Kieselalgen der Schweiz, (48 plates, 1912),

contain splendid figures, and include most of the English forms.

DEPARTMENT OF METHODS 193

La Diatomologia Espanola by Azpeitia, (1911, 12 plates), treats of vari-

ous Spanish deposits, including Moron. This establishes two quite new

genera, Dossetia and Secallia.

I must not omit mention of three important papers by Laubeg, on

the Paleobotany of France, (Soc. Bot. de France. Mém. XV. Jan. 1910):

the Study of Sedimentary Deposits of Diatoms (Bull. des Services de la

Cante Géologique de France, Mém. 125, 1910): and on Diatoms, their

Deposits and Uses, (Revue Générale des Sciences, 1911).

In Nuova Notarisia professor Achilli Forti has written several papers,

with photographic illustrations; on Bergonzano and Marmorito deposits in

1908 and 1914; and a monograph on the genus Pyxilla in 1909. His

paper on the classification of diatoms as Mobiles and Immobiles in 1912

was anticipated by Mereschkowsky in 1902, (Script. Bot. Hort. Imp.

Petrop. Fasc. xvii. p. 96.), as I have already pointed out. Mereschkowsky

has also written other papers on diatoms, and has suggested the forma-

tion of various new genera, some for new forms, and some for species

already described.

West’s Algee (Camb. Univ. Press, 1916) devotes 43 pages to Diatoms,

and compresses much information into that space. Boyer’s Diatomacee

of Philadelphia, 40 plates, bears date 1916: it contains a useful intro-

duction and fine illustrations.

Considerable attention has been given during the last thirty years to

the Plankton diatoms. Cleve in 1889 wrote on Pelagic diatoms from the

Kattegat, and followed this by papers on diatoms from Baffin’s Bay and

Davis’ Straits (1894), Phytoplankton of the North Atlantic (1897), Diatoms

of the Jackson-Harmsworth Expedition (1898), North Sea and English

Channel (1900), Swedish Expedition to Greenland (1900), and Plankton

of the South Atlantic (1900). And Oestrup wrote on the Marine diatoms

of East Greenland (1895, 6 plates).

Van Heurck in 1909 wrote on the diatoms found during the voyage

of S. Y. Belgica in 1897-1899 in the Antarctic Ocean, (13 plates).

Karsten wrote the account of the diatoms found during the German

Deep Sea Expedition in 1898, 1899. This is a costly and magnificent work

in three parts, on the Antarctic, Atlantic, and Indian Phytoplankton, with

splendid plates. Other writers on the subject are Ostenfeldt, Aurivillius,

Hensen, Jorgensen and Lemmermann.

The Belgian Museum of Natural History has published Microplankton

de la Mer Flamande, by Meunier, (Tome VII. Fasc. 2, 3. 14 plates. 1913,

1915), Nordisches Plankton, Botanischer Teil, by Gran, 1905, contains

about 180 figures of interesting and. new forms. Gran also wrote several

other papers on Arctic Diatoms and Plankton.

I must also note Mangin’s paper on the Study of Plankton in Annales

des Sciences Naturelles, 1908, pp. 177-219; and Bachmann on the Phyto-

194 FRED B. TAYLOR

plankton on Fresh Water, with special reference to the Lake of Lucerne,

(1911, pub. Jena). The report of the Imperial Fisheries Institute of

Japan for 1911 contains 6 plates of littoral diatoms of Japan.

It is impossible in the space of a few pages to note all the contribu-

tions to the knowledge of the structure and history of diatoms, even to

name more than some of the best known writers on the subject such as

Nelson, Morland, Lauby, Cox, Brightwell, Roper, Wallich, the two

Miillers, Butcher, O’ Donohoe, and Murray.

THURGARTON, BOURNEMOUTH.

A COMPENDIUM OF THE HOSTS OF ANIMAL PARASITES CON-

TAINED IN WARD AND WHIPPLE’S FRESH-WATER BIOLOGY

COMPILED BY

H. J. VAN CLEAVE

University of Illinois

Ward and Whipple’s Fresh-water Biology contains by far the most

comprehensive treatment of the animal parasites of the North American

fauna that has ever been published. The chapters dealing with the

parasitic worms, represent contributions, on the part of Professor Henry B.

Ward, not only in a compilation of results of almost innumerable researches

of varying magnitude but also in the inclusion of extensive data based upon

previously unpublished records. Much of this information of especial

interest to parasitologists and to field zoologists is not available for ready

reference because the names of hosts mentioned in the text are not in-

cluded in the general index of the book.

For personal use the present writer prepared a compendium of the

hosts mentioned in the Fresh-water Biology. This proved so valuable

an aid and received such favorable comment from workers to whom the

manuscript was shown that it was considered desirable to put it into

a form in which it could be generally available.

The authors of the Fresh-water Biology have made no attempt to

include complete check lists of the hosts in the chapters dealing with

parasitic forms, yet in many instances references are inclusive enough to

be of great value as a point of departure in determining the recorded

parasitic fauna of any given host animal. In using this compendium

it should be recalled that in many instances only one or a few typical

species are listed for each genus and even the hosts of such species as are

cited do not constitute full check lists. Doubtless there have been nu-

merous erroneous determinations of hosts in the works from which the host

lists have been assembled but host names have been quoted directly as they

stand in the original citations without attempt at verification. Asa result,

some of the names of hosts current in the older literature appear along

with the valid names of the same species in this compendium.

In many instances where there seems to be no fixed specificity of hosts,

as well as in the discussion of families and genera of parasites, group names

such as ‘fish,’ ‘birds,’ or ‘water birds,’ are used frequently.

In the chapters dealing with Protozoa the parasitic forms have not

received the attention of the writers, consequently the great group of

Sporozoa and all other parasitic protozoans have received no treatment

in this compendium.

195

196 H. J. VAN CLEAVE

To facilitate locations of words on the page in referring back to the

text, specific, generic, vernacular, or group names have been used in the

compendium as they stand in the text reference. In a few instances cross

references have been.inserted between vernacular and scientific names, but

in such instances the page references to the two names have not been

assembled. This is due to the belief that the inconvenience of cross

citation is less than the confusion resulting from the necessity of visualizing

both vernacular and scientific names while scanning the printed page in

search for a given reference.

A host name cited under a given page reference may appear more than

once on that page. .

A direct means of determining the groups of parasites listed for any

given host without necessity of referring back to the text is afforded by

reference to the following list of page inclusions for the various groups

containing parasitic forms:

Trematoda 374-424

Cestoda 429-451

Nematoda 520-535

Gordiacea 537-542

Acanthocephala 545-551

Rotatoria 553-620

Discodrilide 644

Hirudinea 646-660

Copepoda 782-788

Malacostraca 841-850

Hydracarina 851-874

MAMMALIA skunk 522

‘mammals’ 390, 404, 440, 441, 442,444,447, wapati 389

522, 549 weasel 522

beaver 386 whales 433

cat 390, 393, 447 wolf 523

cattle 389, 409 AVES

Didelphis virginiana 410 ‘bird’ 402, 404, 409

dog 390, 523 ‘birds’ 440, 441, 442, 443, 444, 446, 447, 448,

Lepus 409 526, 549, 550

man 389, 409, 432, 433, 434, 521, 522, 523, Anas platyrhynchos 388

534, 656 Anseriformes 449, 548

mink 522, 523, 530 Ardea herodias 410, 524, 527, 532

muskrat 383, 386, 391, 404, 447, 451, 522, Ardea minor 391

534, 535 ; bittern 532

Mustelidae 522 Botaurus lentigenosus 550, 551

otter 522, 523 Bolaurus minor 526

pig 390 canvas back 442

seals 433, 523, 549 chicken 391, 402, 446

sheep 389, 408 coot 443

DEPARTMENT OF METHODS 197

cormorant 533 Bascanion constrictor 405

crane 444 Chameleon 400

crow (‘fish’) 446 Chelonura ser pentina 524

duck 411, 441, 442, 447 Chelydra serpentina 376, 384, 387, 396, 529

egret (see Herodias) Chrysemys marginata 377, 385, 394

eider duck 549 Chrysemys picla 394

Gallinago wilsont 382 Cinosternum pennsylvanicum 3706

Gavia immer 391 (see loon) Cistudo carolina 522, 524, 533

geese 442, 447 ‘Emys’ 546

grebe 440, 448 Emys guttata 529

grebe (horned) 448, 449 Emys scripta 529

gull 440, 442, 446 (see Larus) Emys serrata 524, 533

Herodias egretta 549 garter snake 407

‘heron’ 408, 440, 444, 445, 532 Heterodon platyrhinus 405, 406, 407

heron (green) 444 Malacoclemmys geographicus 546

heron (little blue) 444 Malacoclemmys lesueurti 378, 380

herring gull 408 Nanemys guttata 528

‘ibis’ 448 Natrix rhombifer 407, 439

kingfisher 525 Pseudemys 387

Larus philadelphia 391 Pseudemys elegans 546

loon 440 (see Gavia) Pseudemys scripta 377

pelican (brown) 533 snakes 405, 438

pelican (white) 433, 533 terrapin (common, food-) 376, 380, 531

pintail-duck 442 Trionyx ferox 378

Plotus anhinga 524 (see water-turkey) Tropidonotus rhombifer 406

Porzana carolina 549 Tropidonotus sipedon 406, 526

scoter 442 turtles 385, 433, 526, 652

scoter (American) 449

scoter (black) 391 AMPHIBIA

eee tees amphibians 398, 399, 404, 408, 434, 438, 449,

snipe (grey) 382 522 547 :

Somateria dressert 549 :

sparrow (English) 384

spoonbill 444

spoonbill (roseate) 445

stilt (blacknecked ) 447. 448

waterbirds 401, 431, 432, 442, 443, 440, 447,

548

water turkey 523, 533 (see Plotus)

wood-ibis 531

Amblystoma 399

Amblystoma mexicanum 528

Amblystoma ligrinum 438

Anura 400, 403

Bufo 399

Bufo americana 522, 533

Bufo lentigenosus 521

Cryptobranchus alleghaniensis 525

Diemyctylus viridescens 547

frog 382. 399. 400, 402, 404, 387, 408, 411,

REPTILIA Te : cor

652, 654

alligator 382, 408 Necturus lateralis 379

Alligator lucius 410 Necturus maculosus 439

Alligator mississippiensis 391, 530,531,532 . Rana catesbiana 408

Amyda 402 Rana halecina 531

Ancistrodon piscivorus 439 . Rana pipiens 410, 524, 530

Aromochelys carinatus 376 salamander 382, 399

Aromochelys odoratus 376, 377, 397 Salamandra rubra 533

Aspidonectes 402 Siredon mexicanus 528, 533

198

PISCES

fishes 382, 398, 401, 408, 411, 433, 434, 523,

524, 528, 529, 547, 551, 652, 653, 654,

655

Acipenser (European) 392

Acipenser oxyrhynchus 433

Actpenser rubicundus 378, 392, 396

Acipenser sturio 374

Ambloplites rupestris 401, 436, 548, 785

Ameiurus nebulosus 399

Amza calva 392, 401, 432, 435, 436, 526, 548,

787

Anguilla vulgaris 435

Anguilla chrysypa 401, 435, 524, 546

A plidonotus grunniens 381, 395

black bass 375, 379, 392, 395, 408, 524, 534

white bass 528, 529

blue-gill 408, 411

Boleosoma nigrum 379

bull-head 382, 408, 439

carp (German) 529

catfishes 786

cat (channel) 439

Catostomus commersonii 787

Catostomus teres 375

chub 431

Coregonus nigripinnis 437

Coregonus prognathus 437

Coregonus artedi 437

crappie (black) 528, 529

Cristivomer namayeush 437

Cyprinidae 408, 430

dace 395

dace (horned) 411

darter 395

Dorosoma cepedianum 545, 547

Eromyzon sucetta oblongus 787

Esox lucius 392, 399, 401, 437, 546

Esox reticulatus 392,437, 524,529 (see Lucius)

Fundulus 655

Fundulus ocellaris 784

Gasterosteus 432

herring (lake) 527

Ictalurus punctatus 395, 401

Lepomis pallidus 785

Leptsosteus osseus 788

Lepisosteus platostomus 435, 436

Lepisosteus tristoechus 788

Lota lota 392

Lucioperca 392

Lucius masquinongy 787

H. J. VAN CLEAVE

Lucius reticulatus 788 (see Esox)

Micropterus dolomieu 374, 392, 398, 401, 436,

548, 785

Micropterus salmoides 392, 400, 436, 546

minnow 379, 395, 398, 408, 411

minnow (red-finned) 395

Moxostoma macrolepidotum 394

Perca flavescens 401, 524

perch 379, 395, 396, 399, 408, 411

pickerel 788

pike 408, 411, 521

pike (wall-eyed) 530

Plychochelius oregonensis 424

pumpkinseed 395, 411

rays 434

Roccus lineatus 375, 548, 785

rockbass 375, 379, 392, 395, 398, 408, 411

Salmo sebago 434, 437

salmonid fish 527

Salvelinus namaycush 392

shad 533

sharks 434

sheepshead 528

Siluride 439

Stizostedion canadense 788

Stizostedion vitreum 401

sturgeon 395

sturgeon (lake) 530

sucker 395, 431, 787

sunfish 375, 395, 411

teleosts 431, 524

trout 408, 431, 433, 550

trout (lake) 374

trout (Great Lake) 431, 527, 547, 548

whitefish 432, 521, 527, 547

INSECTA

insect 534, 851

insect larve 528

Achaeta abbreviata 539

Acridide 538, 540

Blasturus cupidus 395

Blattide 538

Gryllus (see Acheta)

Hexagenia 395

Locustide 540

mayfly 395

Neobius fasciatus 539

CRUSTACEA

crustacea 528, 534

Apus 411

DEPARTMENT OF METHODS 199

Copepoda 451, 523

crawfish 395, 401, 644

Cyclopidee 442

Decapoda 842

Diaptomus 442

Isopoda 547

Ostracoda 451, 523

Palaemonetes 842

Palaemon 842

GASTROPODA

snails 409, 415, 451

Campeloma decisum 416, 419, 420

Gasteropoda 417

Goniobasis virginiana 416

Helix albolabris 423

Helix alternata 423, 531

Helix arborea 411

Lymnaea 419

Lymnaea catascopium 411

Limnaea humilis 389

Lymnaea proxima 412, 417, 418, 419

Lymnaea refiexa 415, 422

Lymnaea stagnalis 411

Physa anatina 417

Physa gyrina 412, 417, 419, 420, 423, 424

Physa heterostropha 411, 413

Planorbis parvus 413, 415

Planorbis trivolvis 413, 417, 420, 421

Pleurocera elevatum 416

slugs 451

Succinea 423

Succinea ovalis 409

LAMELLIBRANCHIA

Anodonta 379, 380

Fresh-water mussels 423, 851, 872

Unionidae 421

ANNELIDA

annelids 451, 521

earthworm 430

Lumbriculus 451

Tubificidee 430

ROTATORIA

rotifers 554

Albertia 589

Pleurotrocha 589

COELENTERATA

Hydra 287, 291

PORIFERA

freshwater sponges 856

PROTOZOA

colonial protozoa 856

THE ENDOCRINES, By SamuEL Wy tis BANDLER, M.D., W. B.

Saunders Co., Philadelphia and London, 1920. 486 pp.; price, cloth,

$7.00 net.

REVIEWED BY

T. W. GALLowAy

In the sudden enthusiasm generated by new discoveries that are clearly

important, we human beings are reasonably sure to be swept into fads and

over-emphasis. Both our generalizations in scientific philosophy and

the more practical applications of them to the treatment of human ailments

are full of illustrations of the momentum of credulity. The student of the

history of science will recognize instances under the terms,—‘‘natural

selection,” ‘“‘adaptation,” “eugenics,” “eye strain,” “‘uric acid,”’ and the

like, in which truth has beep over-pressed.

It would be a miracle under the circumstances if the thoroughly admitted

omnipresence of endocrine secretions in the blood, produced by all sorts

of normal and abnormal groups of cells, should not congest human traffic

over the same course, and lead the willing student of the subject into

statements which are not now established. For example, the present

author asserts that they (the endocrines) are the underlying factors in

heredity;”’ and, more specifically, ‘“‘The differences between animals

of various species (and among individuals of the same species) are due to

the ductless glands,” and more of like tenor. While it may well be true

that the hereditary structural elements in the germ cells are influenced

by the internal secretions of the soma in which the germ cells lie, and both

soma and germ cells are profoundly influenced in their later development

by their mutual secretions, such over-sweeping statements of causation

are not altogether impressive. It is a bit like saying that the environment

is responsible for heredity! which is doubtless true. In spite of this over-

enthusiasm, the body of the book contains a mass of most interesting

statements supported by many cases,—with history, symptoms and

experimental therapy given,—as well as by inference. The author is a

gynecologist and naturally stresses the hormones linked most closely with

the functions of sex development, reproduction, pregnancy, parturition,

and with the emotions conne_ ted with these.

The author with commendable boldness even undertakes to redeem

some territory from the deluge of Freudian interpretation, by way of endo-

crine action. He makes a strong case for believing that many of the com-

plexes, hysterias, bad orientations, and phobias which the psychoanalysts

accredit to mental conflicts are in reality due to a poor balance in the

endocrine system.

> 66

200

DEPARTMENT OF METHODS 201

Rather unfortunately, because the author does not seem to have a

large first-hand knowledge of investigation on inheritance, the book is

introduced by a somewhat rambling and inconsequential jumble of illus-

trations and exhortations which is rather euphemistically labeled ‘‘Envi-

ronment and Heredity,’”—presumably because these terms combined make

room for about all that can be said on any subject. An analogous loose-

ness and lack of system in arrangement mars the treatment at many points

and leads to much unnecessary repetition, and to some seeming conflicts.

This is particularly illustrated in comparing chapter 3, The Introduction

to the Story of the Endocrines; chapter 4, Internal Secretions, and chapter

18, The Balance between the Endocrines. The total is something like

what one might use in a series of lectures on the subject to a class in which

inattention or lack of preparation would make much restatement seem

necessary, rather than what one expects in a scientific book.

Aside from those mentioned, the following are the principal chapter

headings: Environment and Heredity; The Endocrines in Gynecology;

Hypergenitalism and Hypogenitalism; Skin Affections and Internal

Secretions; Puberty and Climacterium; The ‘‘Higher up” Theory of Steril-

ity in Women; Pregnancy, Labor and Placental Gland; Constitutional

Dysmenorrhea; Instincts and Emotions; Mental and Nervous Defects;

Mental Deficiency and Criminality; Neuroses and Psychoses; Phobias;

The Autonomic Nervous System; Therapeutic Suggestions concerning

Endocrines; and several chapters dealing with histories, symptoms, clinics

and cases.

Possibly an illustration will aid in making vivid the author’s repeatedly

emphasized point of the interdependence of the internal secretions. In

respect to lactation after labor, the following facts are significant:—The

mammary glands are developed at puberty by the interaction of secretions

from the ripening ovaries, the posterior hypophysis, the thyroid and the

adrenals. A secretion from the ovaries or from the endometrium causes

them to swell at menstruation. Placental secretions during pregnancy

produce hypertrophy and differentiation of tissue at that time, and seem

to inhibit the action of the above mentioned secretions. Milk itself

will not form however until this placental secretion is removed or inhibited.

After birth the ovaries, posterior hypophysis and other endocrines resume

sway and stimulate degenerative process by which milk is formed.

Assuming that feminine metabolism and emotional states are more

variable and instable than the masculine, the explanation by the endo-

crinologist would include such facts as these:—men and women alike possess

these numerous glands and with‘all degrees of original potency; all these

glands are normally pouring their secretions into the blood constantly; the

secretion of each gland modifies those of certain, if not all the other glands,

as well as general metabolic conditions thruout the body; the glands, directly

202 T. W. GALLOWAY

or indirectly are also modified in their action by the metabolic conditions

and by sudden changes of state (as of the emotions) which may be initiated

by the environment; gradually under normal conditions and in average

individuals these various forces come into an adjustment that represents a

person’s constitutional norm; in women 13 times a year there is a cyclical

interruption of this balance by the introduction of new factors which

influence all the endocrines involved and thus upset the balance; in the

case of pregnancy, new secretions (and consequent modification of the

endocrine balance and of the mental states dependent upon this) are

introduced about each of these points: pregnancy, fetal development,

parturition, lactation, and the cessation of lactation.

Probably it will be of most service to the readers of this review, in

trying to give an idea of the comprehensiveness of the book, to outline the

sources, effects and interrelations of the various internal secretions as the

author conceives them, especially the more intimate interrelations between

the thymus, thyroid, pituitary and the sex glands. The experiments and

deductions pointing to, and partially explaining, the cyclical character

of these sex-linked endocrine activities seem to the reviewer the most

effective and valuable part of the book. The treatment of the sexual and

reproductive secretions of the male is not nearly so adequate, in spite of

their greater simplicity, as is that of the female.

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LIST OF MEMBERS

In order to reduce the cost of printing, the complete

list of members will be omitted.

MEMBERS ADMITTED SINCE THE LAST PUBLISHED LIST

Bicknell Anna Bao ere are ote a eho SE eee: 4411 39th St., Washington, D. C.

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Herrick, Chester A., B. S. ... Kansas State Agricultural College, Manhattan, Kansas.

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Sister Monica Mie S*e a eee sess Notre Dame Training College, Glasgow, Scotiand.

Sperry, Arthur, Bt S) ase Kansas State Agricultural College, Manhattan, Kansas.

206

INDEX

Abstracts, 14; 89; 158; 190. _

Acanthocephala from the Eel, 1.

Ackert, J. E., and Wadley, F. M., Observa-

tions on the Distribution and Life History

of Cephalobium microbivorum Cobb and

of its Host Gryllus assimilis Fabricius, 97.

Allen, W. E., Some Work on Marine Phyto-

plankton, 177.

Allen, W. E., A Brief Study of the Range of

Error in Micro-enumeration, 14.

American Mullets, Life History and Scale

Characters, 26.

Animals, Occurrence and Réle of Copper in,

144.

Animal Parasites, Compendium of Hosts of,

195.

Annual Report of Treasurer, 48.

Arrhenuri, New Species and Collections,

168.

Baker, F. C., Preparing Collections of the

Mollusca for Exhibition and Study, 31.

Bandler, S. W., The Endocrines, a Review,

200.

Briefer Articles, 14: 89; 158; 187.

Brownian Movement, Microscopical Ilumi-

nation with Reference to, 158.

Bullard, Chas., A Method for Orienting and

Mounting Microscopical Objects in Gly-

cerine, 89.

Cambarus agrillicola Faxon, Spring Migra-

tion in, 28.

Cephalobium microbivorum Cobb, Distribu-

tion and Life History, 97.

Cnidosporidian Spores, Structures Charac-

teristic of, 59.

Combination Lighting, Microscope Ilumi-

nation with Reference to, 158.

Common Field Cricket, A Sarcophagid Par-

asite of, 116.

Compendium of Hosts of Animal Parasites,

195.

Copper, Its Occurrence and Rdéle in Insects

and Other Animals, 144.

Crayfish, Spring Migration in, 28.

Cummins, H., Spring Migration in the Cray-

fish, Cambarus agrillicola Faxon, 28.

Custodian’s Report for the Year 1920, 47.

Desmid, Method of Demonstrating Sheath

Structure, 94.

Diatoms, Literature of, 187.

Distribution and Life History of Cephalo-

bium microbivorum Cobb, 97.

Eel, Acanthocephala from, 1.

Endocrines, Review, 200.

Faust, E. C., Larval Flukes from Georgia, 49.

Faust, E. C., Recent Advances in Parasit-

ology, 75.

Fixatives, Effect upon Myxosporidian Spores,

i Lee

Fresh-water Biology, Ward and Whipple,

Compendium of Hosts of Animal Parasites

contained in, 195.

Fresh-water and Marine Gymnostominan

Infusoria, 118.

Galloway, T. W., Review of Endocrines, 200.

Georgia, Larval Flukes from, 49.

Glycerine, Mounting Microscopical Objects

in, 89.

Gryllus assimilis Fabricius, Distribution and

Life History of, 97.

Hausman, L. A., Fresh-water and Marine

Gymnostominan Infusaria, 118.

Henderson, W. F., Treasurer, Annual Re-

port, 48.

Herrick, C. A., A Sarcophagid Parasite of

the Common Field Cricket, 116.

Hosts of Animal Parasites, A Compendium,

195.

Hubbs, C. L., Remarks on the Life History

and Scale Characters of American Mullets,

26.

207

208 INDEX

Infusoria, Gymnostominan, 118.

Insects, Occurrence and Réle of Copper in,

144.

Kudo, R., On the Effect of Some Fixatives

upon Myxosporidian Spores, 161.

Kudo, R., On the Nature of Structures Char-

acteristic of Cnidosporidian Spores, 59.

IU,

Larval Flukes from Georgia, 49.

Life History and Distribution of Cephalo-

bium microbivorum Cobb, 97.

Literature of the Diatoms, 187.

Marine and Fresh-water Gymnostominan

Infusoria, 118.

Marine Phytoplankton, 177.

Marshall, Ruth, New Species and Collections

of Arrhenuri, 168.

Methods, Department of, 14; 89; 158; 187.

Method of Demonstrating Sheath Structure

of a Desmid, 94.

Method for Orienting and Mounting Micros-

copical Objects in Glycerine, 89.

Micro-enumeration, Range of Error in, 14.

Minutes of Chicago Meeting, 47.

Microscope Illumination, 158.

Mollusca, Preparing Collections for Exhibi-

tion and Study, 31.

Mounting in Glycerine, 89.

Muttkowski, R. A., Copper; Its Occurrence

and Réle in Insects and Other Animals,

144.

Myxosporidian Spores, Effect of Fixatives

upon, 161.

Orienting and Mounting in Glycerine, 89.

Pflaum, M., Custodian, Report for the year

1920, 47.

Phytoplankton, Marine, 177.

Preparing Collections of Mollusca, 31. °

Proceedings of the Society, 47.

Range of Error in Micro-enumeration, 14.

Recent Advances in Parasitology, 75.

Reports of Auditing Committee on Treas-

urer’s and Custodian’s Reports, 48.

Reviews, 14; 89;

Sarcophagid Parasite, 116.

Scale Characters in Mullets, 25.

Sheath Structure of Desmid, 94.

Silverman, A., Microscope Illinination with

reference to Brownian Movement and

Combination Lighting, 158.

Spencer-Tolles Fund, Report on, 47.

Summaries, Department of, 75.

4p ~

Taylor, F. B., The Literature of the Diatoms,

187.

Taylor, W. R., A Method of Demonstrating

the Sheath Structure of a Desmid, 94.

Treasurer, Annual Report, of, 48.

Van Cleave, H. J., Acanthocephala from the

Hel) 1.

Van Cleave, H. J., 4 Compendium of the

Hosts of Animal Parasites contained in

Ward and Whipple’s Fresh-water Biology,

195.

Wadley, F. M., and Ackert, J. E., Distribu-

tion and Life History of Cephalobium mi-

crobivorum, 97.

Ward and Whipple, Compendium of Hosts of

Animal Parasites, 195.

Welch, P. S., Secretary, Minutes of Chicago

Meeting, 47.

et oe

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Adapter for FFS Microscope, $50.00. licf from eve fatigue.

Allowing the natural use of both eyes, this apparatus will be

particularly appreciated by all those who are obliged to do frequent

or extended work with the microscope.

Write for illustrated, descriptive circular

502 St. Paul St., Rochester, N. Y.

New York Chicago Washington San Francisco London

Makers of Photographic Lenses, Microscopes, Projection Apparatus

(Balopticons), Ophthalmic Lenses and Instruments, Photomicro-

graphic Apparatus, Range Finders and Gun Sights for Army

and Navy, Searchlight Reflectors. Stereo-Prism Binocu-

lars. Magnifiers and Other High-Grade Optical

Products.

Members and Friends Will Find Our Advertisers Reliable

THE AMERICAN MICROSCOPICAL SOCIETY

Leitz Microscopes are the Standard of the World

LEITZ “MON-OBJECTIVE

BINOCULAR” MICROSCOPE

The Modern Research Type

This Binocular Microscope can be used with any of

the standard objectives from lowest to highest power.

This model originates with Leitz and was success-

fully introduced in 1913. Other firms have copied this

model but the individual design, superior workmanship

and efficiency of the Leitz pattern will fully protect the

prestige for the original type.

Points of Merit

1. Binocular vision.

2. Perfect accommodation to any

interpupillary distance.

Adjustment for any difference

in refraction between the

EVes.

Complete elimination of eye

strain.

>. Improved quality of image.

> Parallelseyepieces:

7. The possibility of using any

favored objective from, the

lowest power to the highest

oil immersion.

8. Reduction in numerical aper-

ture.

—

ELEITZ WerZba*

Write for Pamphlet No. 1003

A OPTICAL ano” MECHANICAL NEw : ORK \ SURPASSED “oa EQUALLED

Nea WORKMANSHIP RY. iris “SUPREME

60 East JOUStr

Members and Friends Will Find Our Advertisers Reliable

THE AMERICAN MICROSCOPICAL SOCIETY Vil

THE SILVERMAN ILLUMINATOR

offers important advantages for practically every application of the

Microscope:

a—It shows more detail.

b—A clearer and better defined picture is presented to the eye

and the camera.

c—Several novel methods of illumination can be provided.

d—It saves much valuable time.

e—It prevents eye strain, eye fatigue and brain fag.

f—It can be lowered into deep hollow objects.

g—It gives excellent results for very low power work as well as

higher magnifications, also in oil immersion work.

h—It can be used with any microscope, ordinary or binocular.

A small circular tube lamp surrounds the objective and fur-

nishes a diffused and uniform illumination directly where it is

needed.

The Silverman Iliuminator marks A GREAT ADVANCE in Microscope

Illumination

WRITE FOR BULLETIN 45-C

LUDWIG HOMMEL & CO.

530-534 Fernando St.

ELECTRIC

DARK FIELD

ILLUMINATOR

(U. S. Army Medical School Type)

A Combined

Dark Field liluminator

and Microscope

Lamp

It Fits the Substage Ring of All Standard Makes of Microscopes

It Is New, Original, Unique, Compact, More Efficient

ANOTHER FORWARD STEP IN MICROSCOPE CON-

STRUCTION. ANOTHER SPENCER TRIUMPH

Pittsburgh, Pa.

Send for Booklet

SPENCER LENS COMPANY

Manufacturers

MICROSCOPES, MICROTOMES, DELINEASCOPES

BUFFALO, N. Y.

SPENCER

BUFFALO

[eurcac]

SPENCER

BUFFALO

Members and Friends Will Find Our Advertisers Reliable

VIII THE AMERICAN MICROSCOPICAL SOCIETY

Watson's Apparatus for Microscopes

Watson’s manufacture a series of Achromatic Objectives of high

and low aperture, and of varying powers, enabling the maximum effect

to be obtained with every description of Object Glass.

They correspond in correction and workmanship with Objectives of

similar powers and apertures, and the high power Condensers can be

made suitable for lower power Objectives by the removal of the top lens.

THE HOLOSCOPIC OIL IMMERSION CON-

DENSER. Power .22”, numerical aperture 1.35, the

whole of which is aplanatic if used on the thickness of

slip for which it is corrected. The finest Condenser

obtainable for high power work.

Optical part only, to fit the Royal Microscopical

Society's Screw Thread, £9/12/6.

Condenser Full ; ‘2 aero = | Diameter

ser Anrtiipe Aplanatic Aperture Equivalent Focus of Back

{AP Lens

| Complete op Lens Complete lop Lens

| Removed Removed

Mac ro. Inch Ineh =

Illuminator = ae = | 2 1225

Aplanatic = ; E d or; means

ees ae -90 | 48 ily -66 = 5

I'he 1.0 95 40 4 1.0 77

Universal 3

Parachromatic 1.0 90 40 29 4 .62

WATSON’S HOLOS. IMMERSION PARABOLOID for the

examination of Spirochetae, and the exhibition of ultra-microscopic

particles, is the best high power Dark Ground Illuminator.

Optical! Pantvonlw Priceres/11/6:

Particulars of all the above, and of Watson’s Microscopes and

Accessories of every description, are contained in their catalogue of

Microscopes, which is published in 4 parts, as follows:

Part 1: Students’ Instruments.

Part 2: Research and Other Microscopes, and appliances of every

description.

Part 4: Instruments for Metallurgy, Petrology and Mineralogy.

Part 5: Photo-Micrographic Apparatus and Accessories. Sent post

free on request to:

W. Watson ¢& Sons, Limited

Established 1837 313, High Holborn, LONDON, ENGLAND

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