CRYPTOGAMIE

Mycologie

ANCIENNE REVUE DE MYCOLOGIE

Fondée par R. Heim en 1936

Directeur scientifique: Mme J. Nicot

Secrétaire de Rédaction: M. Bruno Dennetière

Editeur: A.D.A.C. - 12 rue Buffon F-75005 Paris.

BUREAU DE RÉDACTION

Ecologie et Phytopathologie: G. Durrieu (Laboratoire Botanique et Forestier, 39 Allées

Jules Guesde, F-31062 Toulouse Cedex) - Systématique: P. Joly (Laboratoire de Cryptogamie,

Muséum National d'Histoire Naturelle, 12 rue Buffon, F-75005 Paris) - Physiologie: G.

Manachére (Laboratoire de Mycologie, Université de Lyon I, 43 bd du 11 Novembre 1918,

F-69622 Villeurbanne Cedex) - Cytologie: D. Zickler (Laboratoire de Génétique, Université Paris

Sud, Centre d'Orsay, Bat. 400, F-91405 Orsay) - Autres spécialités: M.F. Roquebert (Laboratoi-

те de Cryptogamie, Muséum National d'Histoire Naturelle, 12 rue Buffon, F-75005 Paris).

COMITE DE LECTURE

J. Boidin (Lyon), J. Chevaugeon (Orsay), J. Fayret (Toulouse), W. Gams (Baarn), G.L.

Hennebert (Louvain-la-Neuve), Ch. Montant (Toulouse), Cl. Moreau (Brest), D.N. Pegler (Kew),

B. Sutton (Kew), G. Turian (Genève).

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supplémentaires seront à leur charge.

TARIFS DES ABONNEMENTS Tome 15, 1994

CRYPTOGAMIE comprend trois sections: Algologie, Bryologie-Lichénologie, Mycologie.

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CRYPTOGAMIE, Mycologie est indexé par Biological Abstracts, Current Contents,

Publications bibliographiques du CNRS (Pascal).

Source : MNHN, Paris

CRYPTOGAMIE

MYCOLOGIE

TOME 14 FASCICULE 4 1993

CONTENTS

C. ILLANA, G. MORENO and A. CASTILLO - Spanish Е VIII.

Some nivicolous myxomycetes from central Spain

M.S.A. HARIDY - Occurrence of yeasts in yoghurt, cheese and whey ............

GI. NAUMOV, ES. NAUMOVA, E.D. SANCHO and M. KORHOLA -

Taxogenetics of the Saccharomyces sensu stricto yeasts from western

and South Africa ..... зана

A.H.M. EL-SAID - Keratinophilic and degno resistant fungi in soils of

Oman

C. WALKER, V. GIANINAZZI-PEARSON and H. MARION-ESPINASSE -

Seine c castanea, a С described arbuscular Da

fungus k ка poner:

M. MOSTAFA - Biological control of Drechslera teres: ability of antagonists

to reduce conidia formation, coleoptile infection and leaf infection in

barley (Hordeum vulgare)...

H.A.H. HASAN, M.M.K. BAGY and A.Y. ABDEL-MALLEK - The incidence

of fungi in human axillary hair and their toxigenic potentialities ...........

М. CONTU - The Agaricus genus. -I Agaricus bisporatus SPEC. nov. «eee

Bibliography sine

Table du volume 14 nn...

ІІІ

241

255

279

287

297

307

311

315

Bibliothèque Centrale Muséum

3 3001 00226854

рта

Source : MNHN. Paris

Cryptogamie, Mycol. 1993, 14 (4): 241-253 241

SPANISH MYXOMYCETES. VIII. SOME NIVICOLOUS

MYXOMYCETES FROM CENTRAL SPAIN

С. ILLANA, С. MORENO and A. CASTILLO

Departamento de Biologia Vegetal (Botanica). Universidad de Alcalá de

Henares. Alcalá de Henares, 28871. Madrid. Spain.

ABSTRACT - Six species of myxomycetes associated with Pinus sylvestris vegetation near melt-

ing snow were collected from the Sierra de Guadarrama (province of Segovia) and studied. Tric-

hia sordida var. sordidoides is proposed as a new variety and Comatricha chionophila as a new

combination. Trichia bicolor and Trichia contorta var. engadinensis are considered to represent

the same taxon as Trichia sordida. Comatricha alpina is new to Spain.

KEY WORDS : Taxonomy, chorology, myxomycetes, Spain.

INTRODUCTION

The nivicolous Myxomycetes have been poorly known in our country. As far

as we know only Gracia (1986, 1987) and Lado (1992) have published on them up to

now.

A microclimate favourable for the fruiting of some species myxomycetes is

created on dead and living vegetation covered by melting old snow. The fructifications

of nivicolous species can be numerous, but their period of fruiting is short, and they

are typically spring species. During the spring of 1991 and 1992, we visited the Sierra

de Guadarrama in the centre of Spain and collected some myxomycetes which were

growing on Pinus sylvestris vegetation covered for a time by melting snow. Four of

six observed taxa are thought to be typically nivicolous species: Comatricha alpina

Kowalski, C. chionophila (Lado) Moreno, Lamproderma sauteri Rostaf., and Trichia

sordida var. sordidoides Шапа & Moreno. Incidentally nivicolous (we think) are Di-

апета corticatum Lister and Enerthenema melanospermum Macbride & Martin; they

are rare species in Spain.

Microphotographs of spores and capillitium prepared with the critical point

method, were made under a Zeiss-DSM 950 SEM and under a Labophot Nikon photo-

microscope. Microscopic examinations were made of material mounted in water or

Hoyer's medium.

The material examinated has been deposited in the herbarium of the Depart-

ment of Plant Biology (Botany) of the University of Alcalá de Henares (AH).

Comatricha alpina Kowalski, Madroño 22: 152. 1973. (Figs. 1-2)

= С suksdorfii var. aggregata Meylan, Bull. Soc. Vaud. Sci. Nat. 53: 455.

1921. л

Source : MNHN. Paris

242 С. ILLANA et al.

Fig. 1. Comatricha alpina Kowalski, (AH 13453), a-b: sporocarps, с: spores, d-e: capillitium.

Collections examined. SEGOVIA: On fallen trunks and dead branches of Pi-

nus sylvestris, near melting snow, Sierra de Guadarrama, mountain pass of Los Cotos,

alt. 1900 m., 31.V. 1991, leg.: A. Castillo, G. Moreno, E. Valenzuela & C. Ochoa, AH

13453.

Our collection consists of about 20 sporocarps. These are densely aggregated,

3-4 mm tall, their sporophores are black, ovoid to cylindrical, about 1.6 diam, 2-3 mm

Source : MNHN. Paris

SPANISH MYXOMYCETES. VIII 243

Fig. 2. Comatricha alpina Kowalski, (AH 13453), а-Б: spores under the SEM

long, and their stalks are black and opaque and about 1 mm long. The peridium leaves

only a small violet-brown collar at the base of the collumella. The collumella is black

and opaque, and reaches about 2/3 the way up the sporophore and is rather sturdy

throughout, ending abruptly in a few capillitial branches. The capillitium is dark, dense

and strongly anastomosed. The spores are black in mass, dark brown in transmitted

light with a pale area, 10-12(13) rm in diam., densely and minutely warted. SEM,

however, shows that the "warts" are in fact baculae with irregular apices.

Comatricha alpina is a typically nivicolous species close to C. suksdorfii Ellis

& Ev., and it can further be segregated by its densely aggregated (touching) smaller

sporocarps on shorter stalks. Comatricha suksdorfii has loosely gregarious or scattered

sporocarps, 7-8 mm tall and stalks about 4 mm long (Kowalski, 1972, 1975; Meyer,

1986).

Comatricha suksdorfii has been cited previously from Spain by Gracia (1986),

but C. alpina is a new contribution to the national catalogue of myxomycetes.

Comatricha chionophila (Lado) Moreno, comb.nov. (Fig. 3)

= Collaria chionophila Lado, Anales Jard. Bot. Madrid 50: 9-11. 1992.

Collections examined. SEGOVIA: On fallen trunks and dead branches of Pi-

nus sylvestris, near melting snow, Sierra de Guadarrama, mountain pass of Los Cotos,

alt. 1900 m., 31.У.1991, leg: А. Castillo, б. Moreno, E. Valenzuela & C. Ochoa, АН

13441. Idem, 6.1.1991, leg.: A. Castillo, С. Moreno & С. Шапа, АН 14569.

Recently, Lado (1992) described a new species of Collaria, characteristically

found in nivicolous environments. The habitat and some locality and collection data

were the same as ours. We add M.O. photographs of sporocarps and capillitium to

complement the morphological studies made before.

We prefer following the classical concept of recognizing the genus Comatri-

cha, including within it those species assigned to the genus Collaria. Thus, we make

the appropiate new combination.

Source : MNHN, Paris

244 C. ILLANA et al.

Fig. 3. Comatricha chionophila (Lado) Moreno, (AH 13441), a-b: sporocarps, c-e: capillitium.

Dianema corticatum Lister, Monogr. Mycetozoa 205. 1984.

Collections examined. SEGOVIA: On dead branches of Pinus sylvestris, Sier-

ra de Guadarrama, mountain pass of Los Cotos, alt. 1900 m., 6.VI.1991, leg.: G.

Moreno, E. Valenzuela, A. Castillo & C. Шапа, AH 13467. Idem, 10.IV. 1992, leg.‘

A. Castillo & G. Moreno, AH 14545 y 16088.

This species has been collected in Spain only twice: in the province of Tarra-

gona (Gracia, 1977) on Populus alba and in Huesca (Carilla & Gracia, 1991) on Ficus

carica in a moist chamber culture. In both cases the spiral ornamentation of the capill-

itium is lacking.

SPANISH MYXOMYCETES. VIII 245

сосна melanospermun Macbride & Martin, J. Wash. Acad. Sci. 22: 91.

Collections examined. SEGOVIA: On dead branches of Pinus sylvestris, Sier-

ra de Guadarrama, mountain pass of Los Cotos, alt. 1900 m., 6.VI.1991, leg.: A. Cas-

tillo, G. Moreno, C. Шапа & E. Valenzuela, АН 13464. .

Enerthenema melanospermum is characterized by its columella ending in a

rather wide membranous brown funnel on the periphery and by its dark, with a pale

side, spinulose spores, 13-15 pm in diam. It was previously recorded in Spain from

the province of Lerida (Gracia, 1977).

Fig. 4. Trichia sordida var. sordidoides Wana & Moreno (typus), a-b: sporocarps, c-i: capillitium.

246 С. ILLANA et al.

Lamproderma sauteri Rostaf., Sluzowce Monogr. 205. 1874.

Collections examined. SEGOVIA: On dead branches and leaves of Pinus syl-

vestris and the stem of Cytisus oromediterraneus near melting snow, Sierra de Guadar-

гата, mountain pass of Los Cotos, alt. 1900 m., 31.V.1991, leg.: б. Moreno, В. Va-

lenzuela, A. Castillo & L.G. Montero, AH 13440, 13444, 13445, 13446, 13447, 13451

and 14501. Idem. 6.VI.1991, leg.: C. Ilana, A. Castillo, E. Valenzuela & G.Moreno,

AH 13464, 13465, 13468, 13469, 13591, 13592, 14502, 14503 and 14504.

Our collections are characterized by their 2-5 mm tall sporocarps, globose spo-

rophores, 2-2.5 mm diam, and short, about half the total height, black stalks on a well

developed hypothallus. The peridium is membranous, persistent, blue with silvery re-

flections, and which tears irregularly from apex to base, where pieces persist in the

form of a large collar. The columella is dark brown or opaque, long, almost reaching

the apex of the sporophore. The capillitium is dense, dark but paler outwards. The

spores are black in mass, dark brown in transmitted light with a small pale area,

(13)14-15 um in diam., densely warted.

This nivicolous species was recorded before in Spain from Lerida (Gracia,

1986). Most Lamproderma species are found at high altitudes in the mountains, where

they are often abundant and show great diversity (Kowalski, 1968). In Switzerland,

Meylan made many collections of L. sauteri and described six varieties; most of these

were not recognized by Kowalski (1975) who suggests that the differences are rela-

tively minor and thus not sufficient to recognize separate taxa; therefore L. sauteri is a

variable species.

Trichia sordida var. sordidoides Mana & Moreno var. nov. (Figs. 4-5)

Diagnosis: A typo differt capillitium "hemitrichioide" non "trichioide".

Etymology: From the latin "sordidoides" similar to Trichia sordida Johanne-

sen.

Habitat: on Pinus sylvestris, mountain pass Los Cotos (Segovia), 31.V.1991,

leg.: L.G. Montero, A. Castillo & G. Moreno, AH 13442.

Collections examined:

Trichia contorta var. sordidoides. SEGOVIA: On fallen trunks, leaves and

cones dead of Pinus sylvestris, near melting snow, Sierra de Guadarrama, mountain

pass of Los Cotos, alt.1900 m., leg.: L.G. Montero, A. Castillo & б. Moreno,

31.V.1991, AH 13442 Holotypus, 13443 and 13449. Nowotny 1714(1) -Austria-. Mey-

er 3280 -France-.

T. bicolor Stephenson & Farr, holotypus (BPI)

Trichia contorta var. engadinensis Meylan, lectotypus (LAU).

T. sordida Johannesen, holotypus (0). Nowotny no. 1714(2), 2225 and 4468

-Austria- Meyer no. 3611 -France-.

Sporocarps clustered and aggregated. Sporophores globose to subglobose,

0.4-1 mm diam., sessile, or on short thick stalks, Hypothallus dark brown to reddish,

continuous under a colony. Peridium single, membranous, the upper part showing dark

lines and rounded or irregularly star-shaped maculae consisting of particle deposits on

a dark ochraceous background, the base dark red, in transmitted light the peridium is

minutely irregular striated within and shows rounded depressions; dehiscence irregular.

Source : MNHN, Paris

SPANISH MYXOMYCETES. VIII 247

ЕЗ

Fig. 5. Trichia sordida var. sordidoides Шапа & Moreno (typus), a-b: peridium inner side, c-d:

spores under the SEM, e-g: capillitium.

Source : MNHN, Paris

248 C. ILLANA et al.

The capillitium is formed by elaters, profuse, long, tips tapering, yellow,

5-6 um in diam., ornamented with 4-5 spirals, with many short, thinner, tapering lat-

eral branchlets. The spores are yellow in mass, pale yellow in transmitted light, glo-

bose to subglobose, 12-15 um diam., very densely warted.

і Examination by SEM shows that the individual "warts" have a flat apex and a

narrowed base, so that they are in reality piles (Rammeloo, 1974; Saenz, 1978); these

Fig. 6. Trichia sordida Johannesen (typus), a: sporocarps, b-g: capillitium and spores.

SPANISH MYXOMYCETES. VIII 249

are very dense and many are confluent in rows and small clusters. The peridium

shows crateriform depressions, possibly spore impressions, and is irregularly reticulate

from minute wrinkles, like the peridium of Hemitrichia montana (Morgan) Macbride

(Rammeloo, 1984).

The proposed variety is like var. sordida (Figs. 6-7) in its habitat (nivicolous).

Moreover, the two varieties look alike and they are identical in peridial structure and

spore size. They differ only in the capillitium: in var. sordidoides the elaters are long

and show few free ends, and bear many, short, tapering secundary branchlets. The

i sordida has short elaters with many free ends, and scanty secundary branchlets

ig. Sa).

Fig. 7. Trichia sordida Johannesen (typus), a-b: spores under the SEM, c-d: capillitium.

250 С. ILLANA et al.

Fig. 8. Trichia bicolor Stephenson & Farr (typus), a: sporocarps, b-i: capillitium.

Source : MNHN. Paris

SPANISH MYXOMYCETES. VIII 251

Fig. 9: Trichia bicolor Stephenson & Farr (typus), a-b: inner peridium inner side, c-d: spores un-

der SEM, e-f: capillitium.

Source : MNHN, Paris

252 С. ILLANA et al.

We found that Trichia contorta var. engadinensis Meylan (1921) is very like

T. sordida: it is also nivicolous and was originally described as having globose to

ovoid, 1-2 mm in diam., sessile sporophores and a brown peridium. However, Kowal-

ski (1975) described the peridium of the type as speckled, and we were able to verify

this ourselves in the, mostly badly squashed, peridia; the maculae observed were very

like those of 7. sordida but less pronounced in the upper part. As were the microscop-

ical features: short elaters with many free ends and spores 12-14(-15) yum diám., simi-

larly decorated.

Trichia bicolor Stephenson & Farr (1990) is very much like 7. sordida; the

type (Figs. 8-9) consists of a dozen sessile sporocarps, in small scattered groups. Its

sporophores are subglobose to ellipsoidal. The peridium is yellow ochraceous with

ted lines and spots above and with a dark red base. The microscopical features are like

those of T. sordida, too except for the larger diameter (5.5-6.5 шт) of the elaters.

From Austria we were able to study nivicolous specimens kindly lent by Mr.

Nowotny, of which no. 1714(2) was particularly interesting, as it showed a gradation

in macroscopic features between T. bicolor and T. sordida. Because of these observa-

tions, we are agreed that the three taxa of which we have studied the types should be

included in only one species viz: Trichia sordida Johannesen, Mycotaxon 20: 81-82

1984; (= Т. contorta var. engadinensis Meylan, Bull. Soc. Vaud. Sc. Nat., 53: 460.1921;

= T. bicolor Stephenson & Farr, Mycologia 82:513.1990). The main macroscopic and

microscopic characters are summarized in Figs. 6-9.

For some authors (Kowalski, 1975; Meyer, 1986), Trichia contorta var. enga-

dinensis is a form "trichioide" of Hemitrichia montana (Morgan) Macbr. Hemitrichia

montana has brown-ochraceous sporocarps without the typical stains, it resembles ma-

croscopically H. clavata with the exception of the stalk. Rammeloo (1989) studied this

problem, adding data and SEM photographs.

Hemitrichia montana differs from Т. sordida by its capillitium not consisting

of free elaters and not bearing short lateral branchlets, and by its evenly brown ochra-

ceous peridium.

ACKNOWLEDGMENTS

We wish to express our gratitude to Mrs. Nannenga-Bremekamp for reviewing the mate-

tial of Comatricha alpina and for revising the manuscript; to Dr. Rossman and Dr. Stephenson

for revising the manuscript; to Mr. Nowotny and Mrs. Meyer for lending their collections of ni

colous Trichiales; to Mr. Gonzaga-García, curator of herbarium University Alcalá and curators

herbaria BPI, LAU, O; to Nikon (Rego & Cia, Madrid) for his photographic advice; to Mr. Pérez

from the Microscopy Electronics Service of the University of Alcala for the photographs and to

the "Comunidad de Madrid, Programa Regional de Investigación Ambiental", for the research

project C008/90 "Hongos de las zonas áridas y esteparias (calcoyesiferas) de la Comunidad de

Madrid".

BIBLIOGRAPHY

CARILLA J. y GRACIA Е., 1991 - Mixomicetes corticicolas de Aragón. I. Revista Iberoameri-

cana de Micologia 8: 3-7.

GRACIA E., 1977 - Contribución a la flora de mixomicetes de Catalufia. Mediterranea 2: 79-87.

GRACIA E., 1986 - Mixomicets quionófils. Collect. Bot. 16: 251-253.

Source : MNHN, Paris

SPANISH MYXOMYCETES. VIII 253

GRACIA E., 1987 - Mixomicetes quionófilos. П. Libro de resúmenes VII Simp. Nac. Bot. Crip-

tog., Madrid: 123.

JOHANNESEN E.W., 1984 - A new species of Trichia (Myxomycetes) from Norway. Mycotaxon

20: 81-84.

KOWALSKI D.T., 1968 - Observations on the genus Lamproderma. Mycologia 60: 756-768.

KOWALSKI D.T., 1973 - Notes on western Myxomycetes. Madroño 22: 151-152.

KOWALSKI D.T., 1975 - The Myxomycete taxa described by Charles Meylan. Mycologia 67:

448-494.

LADO C., 1992 - Collaria chionophila, а new myxomycete from Spain. Anales Jard. Bot. Madrid

50: 9-13.

MEYER M., 1986 - Les espéces nivales de Myxomycétes. Bull. Féd. Myc. Dauphiné-Savoie 100:

51-54.

MEYLAN Ch., 1921 - Contribution à la connaissance des Myxomycètes de la Suisse. Bull. Soc.

Vaud. Sc. Nat. 53: 451-463.

MEYLAN Ch., 1925 - Note sur divers Myxomycètes du Jura et des Alpes. Bull. Soc. Vaud. Sc.

Nat. 56: 65-74.

RAMMELOO J., 1984 - Icones Mycologicae 35-54. Jardin Botanique National de Belgique.

Meise.

STEPHENSON S.L. and FARR M.L., 1990 - A new species of Trichia from Montana. Mycologia

82: 513-514.

Source : MNHN, Paris

Source : MNHN. Paris

Cryptogamie, Mycol. 1993, 14 (4): 255-262 255

OCCURRENCE OF YEASTS IN YOGHURT, CHEESE

AND WHEY

Mamdouh S.A. HARIDY

Botany Department, Faculty of Science, El-Minia University, El-Minia,

Egypt.

ABSTRACT - 252 yeast strains were isolated from yoghurt (108 strains), soft cheese (50 strains),

whey (62 strains) and dairy laboratory (32 strains). On the basis of 25 physiological and morpho-

logical characteristics, the isolated yeast strains were assigned to 10 species belonging to 9 gen-

era. Kluyveromyces marxianus was the most common species followed by Debaromyces hansenii.

These species were of high occurrence in yoghurt, cheese and whey. Rhodotorula mucilaginosa,

Saccharomyces cerevisiae and Sporobolomyces roseus were represented by a considerable number

of strains, The total counts of yeast cells ranged between 9.8 x 10° and 1.8 x 10° in yoghurt, 1.1

x 104 and 6.4 x 104 in cheese and 4.0 x 10* and 1.1 x 105 in whey. Our results showed that the

composition of yeast flora as well as the total counts of yeast cells were dependent on manufac-

toring plants and contamination sources. Of particular note was the ability of yeast species to

grow in yoghurt at preservative temperature (5-6°C). The total counts of yeast cells were obvi-

ously higher at the surface of yoghurt packings than at the bottom, These counts increased three

fold at the bottom and ca. five fold at the surface of yoghurt packings after incubation of yog-

hurt packings at 5-6°C for 3 days from manufacturing date. It was of interest that Rhodotorula

mucilaginosa, Trichosporon beigelit and Candida rugosa, the human yeast pathogens, as well as

Candida steatolytica, the erreger of mastitic udder of cow, were isolated in this study.

RESUME - 252 souches de levures ont été isolées de yaourts (108 souches), de fromage mollet

(50 souches), de petit-lait (62 souches) et de de laboratoires laitiers (32 souches). Sur la base de

25 caractéristiques morphologiques et physiologiques, les souches isolées ont été réparties en 10

espàces appartenant à 9 genres. Kluyveromyces marxianus est l'espèce la plus commune, suivie

par Debaromyces hansenii. On trouve ces espèces en grande quantité dans le yaourt, le fromage

et le petit lait. Rhodotorula mucilaginosa, Saccharomyces cerevisiae et Sporobolomyces roseus

sont également représentées par un nombre considérable de souches. Le nombre total de cellules

de levures est compris entre 9,8.10? et 1,8.10° pour le yaourt, entre 1,1.10* et 6,4.10* pour le fro-

mage et entre 4,0.10* et 1,1.105 pour le petit lait. Nos résultats montrent que les espèces et le

nombre de cellules de levures isolées dépendent des techniques de fabrication et des sources de

contamination. On remarquera l'aptitude des espèces isolées à se développer dans le yaourt, aux

températures de conservation (5-6°C). On peut observer que le nombre global de cellules est plus

important à la surface qu'au fond du pot de yaourt. Durant les trois jours suivant la fabrication, à

la température de conservation, le nombre de cellules de levure est multiplié par 3 au fond du

pot, alors qu'il est multiplié par 5 à la surface du yaourt. Cette étude a également permis l'iso-

lement de levures pathogènes pour l'homme, Rhodotorula mucilaginosa, Trichosporon beigelii,

Candida rugosa, ou pour l'animal, Candida steatolytica, cause de la maladie du pis de la vache.

KEY WORDS : yeast, yoghurt, cheese, whey, human and animal pathogens.

Source : MNHN, Paris

256 M.S.A. HARIDY

Yeasts represent an obvious group of the microflora of milk and milk products

(Dombrowski, 1910). Depending upon type of yeast species as well as degree of prod-

uct contamination, they affect products positively or negatively. Their positive effects

were represented by deassimilation of acids and production of growth factors which

stimulate growth and activity of surface flora especially lactic acid bacteria as well as

aroma production by the formation of volatile acids and carbonyl compounds (Hosono

& Tokita, 1970; Kang et al., 1976; Engel, 1980; Schmidt, 1982; Winkelmann, 1983;

Lenoir, 1984). Their negative effects were a result of the fermentation of lactose prod-

ucing gas which in tum leads to swelling, flawing and bombage of product container

(Stocker, 1954; Fahmy & Youssef, 1974; Kroger, 1976; Spillmann & Geiges, 1983),

proteolytic and lipolytic activities (Schmidt, 1982) and production of disagreeable fla-

vour, bitter taste and loss of texture quality (Soulides, 1955; Davis, 1970, 1974; Kro-

ger, 1976). In Egypt, yoghurt and and soft cheese represent an important protein

source for farmers and the average class population which need to compensate for the

amino acids deficiencies of a vegetable protein diet. The study of sanitary condition

of these milk product is still under way, especially in villages. Except for the work

done on cheese by Ghoniem (1963, 1968), Fahmy & Youssef (1974), El-Bassiony et

al. (1980), Seham et al. (1982) and El-Essawy et al. (1984), little is known about yeast

flora of soft cheese especially at the El-Minia locality, where no study was performed

along that line. Moreover there is an obvious shortage in the knowledge about yeasts

of yoghurt as well as whey. Therefore, we report on the yeast flora of these milk pro-

ducts.

MATERIALS AND METHODS

Cheese and whey were collected in sterile conical flasks from supermarkets as

well as from farmer sellers in El-Minia markets. Yoghurt packings of three different

producers were bought from super markets. 5-10в samples from cheese or yoghurt

were mixed with 50 ml sterile Ringer solution (Haridy, 1987), blended and decimal di-

lutions were prepared for counting purposes. 0.5ml whey samples were mixed with 9.5

ml sterile Ringer solution and series of dilutions were also prepared. 0.511 portions

were spread on plates containing yeast malt agar medium (Lodder, 1970) adjusted to

pH 3.5. Plates were incubated at 28°C for 3-5 days. Developing yeast colonies were

counted and after microscopic examination, 252 strains were isolated, purified and pre-

served on yeast malt agar slants at 4°C. In order to survey yeast flora of the dairy la-

boratory, swabs were made from the floor and equipment and then streaked on the

above mentioned plates, whereas air of the dairy laboratory was surveyed by exposing

prepared plates to the air. In order to test the ability of yeasts to grow in yoghurt at

Preservative temperature (S-6°C), a set of yoghurt packings were bought from the

dairy laboratory after manufacturing and then incubated at 5-6°C. Following the above

mentioned procedures, the total count of yeasts cells were determined daily for a peri-

od of 6 days. Identification of the isolated yeast strains was performed on the basis of

25 physiological and morphological properties (table 3) as indicated by Lodder (1970),

Barnett et al. (1983) and Kreger van Rij (1984).

RESULTS AND DISCUSSION

I - Yoghurt:

Yoghurt represents a selective environment for the growth of yeasts because of

its low pH value. Spoilage of yoghurt was reported by several investigators (Ingram,

Source : MNHN, Paris

YEASTS IN YOGHURT, CHEESE AND WHEY 257

1958; Walker & Ayres, 1970; Davis, 1975; Peppler, 1976; Spillmann & Geiges, 1983;

Haridy, 1987). Results of our investigations of natural yoghurt products from 3 differ-

ent producers (A, B, and C) showed that natural yoghurt from producer C were free

from yeasts, whereas 40% of his fruit yoghurt products (D) contained yeasts (table 1).

The total count of yeast cells ranged between 4 and 55 yeast cells per g fruit yoghurt.

Introduction of fruits into his yoghurt products has amplified the risk of spoilage by

yeast by providing additional sources of contamination and fermentable substrates. De-

baromyces hansenii was the most common species. These observations were also re-

ported by other investigators (Davis, 1970, 1975; Tilbury et al., 1974; Spillmann &

Geiges, 1983; Haridy, 1987). Natural yoghurt of producer B showed higher counts of

yeast cells than those of the producer A (table 1). Moreover, only 2 species were iso-

lated from his yoghurt products. The isolation of only 2 species referred to internal

contamination of the manufacturing units. Natural yoghurt of producer A contained

lower counts of yeast cells and a wider spectrum of yeast species (table 1). Saccharo-

myces cerevisiae was only isolated from yoghurt of the producer A and represented

one of the dominant species. Generally, these results showed that the composition of

yeast flora of yoghurt and total counts of yeast cells were dependent upon the manu-

facturing plants and contamination sources. These observations were previously re-

ported by several investigators (Katrandshiev, 1965; Tilbury et al., 1974; Dubois,

1980; Suriyarachchi & Fleet, 1981; Comi et al., 1982; Spillmann & Geiges, 1983;

Haridy, 1987). Table 1 showed that not only higher counts of yeast cells but also wid-

er spectrum of yeast species was detected at the surface of yoghurt packings than at its

bottom. Kluyveromyces marxianus represented the dominant species at the surface as

well as at the bottom, whereas Debaromyces hansenii and Saccharomyces cerevisiae

were only isolated from the surface. These 3 species were the most common species

recovered from yoghurt and their occurrence in yoghurt was reported by several inves-

tigators (Katrandzhiev, 1965; Dubois et al., 1980; Suriyarachchi & Fleet, 1981; Spill-

mann & Geiges, 1983; Haridy, 1987). Table 2 showed the ability of yeasts to grow at

preservative temperature of yoghurt (5-6°C). It was clear from table 2 that yeasts

could grow at the surface as well as at the bottom of yoghurt packings. After the first

day of manufacturing, one gram of yoghurt contained 1.4 x 10“ and 5.4 x 10* yeast

ceils at the bottom and at the surface of yoghurt packing respectively (table 2), where-

as at the third day of manufacturing (consumption period allowed), the total counts of

yeast cells reached 2.4 x 10 per g at the surface and 4.1 x 10“ per gm. at the bottom.

These results showed that yoghurt was heavily contaminated with yeasts and was not

satisfactory for human consumption.

II - Soft cheese:

Table 1 showed that soft cheese of producer G was heavily contaminated by

yeasts. The total counts of yeast cells ranged between 2.1 x 10“ and 6.4 x 10“ рег в.

Its yeast cells counts was the highest count among the cheeses tested, where a limited

number of yeast species was recorded. These results indicate an internal contam-

ination from manufacturing units. The lowest number of yeast cell counts and the wid-

est spectrum of yeast species was recorded in soft cheese of producer F. Character-

istic for cheese of producer E was the dominance of the red yeast species,

Sporobolomyces roseus as compared to the common occurrence of Kluyveromyces

marzianus and Debaryomyces hansenii in cheese of the producers Е and б (table 1).

Isolation of Kluyveromyces marxianus as a dominant species in soft cheese (Damietta)

was reported by Fahmy & Youssef (1974). Dominance of Kluyveromyces marxianus

and Debaromyces hansenii in other types of cheese was reported by several investi-

gators (Stadhouder & Mulder, 1959; Lenoir & Auberger, 1966; Carini et al., 1977; Ro-

sini, 1976; Vergeade et al., 1976; Deiana et al., 1977; Nakase & Komagata, 1977;

Source - ММНМ Paris

M.S.A. HARIDY

258

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Source : ММНМ Paris

YEASTS IN YOGHURT, CHEESE AND WHEY 259.

Table 2: Growth of yeasts in yoghurt at 5-6°C.

Tableau 2: Croissance des levures dans le yaourt à 5-6°C.

Incubation \verage numbers of yeast cells/g yoghurt

period (days) at the surface at the bot

1 5.4 x 1.4 x 10*

2 7.6 x 10* 1.9 x 10*

3 х 107 4.1 x 104

4 3.3 x 10° 1510

5 4.7 10° 15 ce

6 6.1 x 10° 2.5 x 10°

Schmidt, 1978: Schmidt & Lenoir, 1978, 1980; Schmidt & Lambert, 1981; Haridy,

1987).

Ш - Whey:

Results in table 1 showed that higher counts of yeast cells and wider spectrum

of yeast species were recorded in whey than in cheese. It was clear from table 1 that

the most common species of yeasts recovered from cheese were generally dominant in

whey. Kluyveromyces marxianus, Debaromyces hansenii and Sporobolomyces roseus

were the dominant species. Rhodotorula mucilaginosa and Saccharomyces cerevisiae

were represented by considerable numbers of strains.

IV - Yeast flora of dairy laboratory:

In order to determine the effects of manufacturing practice on yeast flora of

dairy products such as yoghurt and cheese, we chose dairy laboratory of the producer

G. Air, floor and equipment were screened for their contents of yeast flora. Results in

table 1 showed that air, floor and equipment represented a contamination sources of

yoghurt and cheese by yeasts. Yeast flora of the dairy laboratory, especially yeasts of

the floor, was detected in yoghurt, cheese and whey. This observation was also re-

ported by Haridy (1987). Table 3 showed physiological and morphological properties

of the isolated yeast species. It was clear from table 3 that 252 isolated yeast strains

were assigned to 10 species belonging to 9 genera. Kluyveromyces marxianus was the

most common species followed by Debaromyces hansenii. Rhodotorula mucilaginosa,

Saccharomyces cerevisiae and Sporobolomyces roseus were represented by consider-

able numbers of strains. Of particular note was the isolation of Rhodotorula mucilagi-

nosa, Trichosporon beigelii and Candida rugosa, the human yeast pathogens, as well

as Candida steatolytica, the erreger of mastitic udder of cow.

M.S.A. HARIDY

260

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Moo 30 eppn 53314554 jo 2969223 = 6 змобоцзеў 4stoÁ мен = + SUNENIS Јо виојдовоа алізівсй Jo обезиволад = €

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загаду 15894 porejost oq Jo sonodoxd [eorsopoudrour pue [vordojorsKud :Є 21921.

Source : MNHN. Paris

YEASTS IN YOGHURT, CHEESE AND WHEY 261

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Source : MNHN, Paris

Cryptogamie, mycol. 1993, 14 (4): 263-270 263

TAXOGENETICS OF THE SACCHAROMYCES SENSU

STRICTO YEASTS FROM WESTERN AND SOUTH AFRICA

G.I. NAUMOV", E.S. МАОМОУА!, E.D. SANCHO? and M KORHOLA?

1, Scientific Research Institute for Genetics and Selection of Industrial

Microorganisms, I Dorozhnyi 1, Moscow 113545, Russia.

2. Departamento de Microbiologia, Escuela Tecnica Superior de

Ingenieros Agronomos, Universidad de Cordoba, Apartado 3048,

14080 Cordoba, España.

3. Research Laboratories of the Finnish State Alcohol Company

(ALKO Ltd), POB 350, SF-00101 Helsinki, Finland.

ABSTRACT - Using genetic hybridization analysis, we reidentified Saccharomyces sensu stricto

strains isolated from soil in South africa (J.P. van der Walt, 1970) as two biological sibling spe-

cies 5. cerevisiae Hansen and S. paradoxus Batschinskaia. The latter was found for the first time

in Africa. Three Saccharomyces strains isolated from different wines in Western Africa (A. Guil-

lermond, 1914) belong to S. cerevisiae and harbor each unique set of sucrose fermenting polym-

eric SUCI, SUC2 and SUC3 genes.

RÉSUMÉ - Des souches de Saccharomyces sensu stricto isolées du sol en Afrique du Sud (J.P.

van der Walt, 1970) ont été reidentifiées par analyse génétique comme deux espèces biologiques,

S. cerevisiae Hansen et S. paradoxus Batschinskaia. La dernière espèce a été trouvée pour la

première fois en Afrique. Trois souches de Saccharomyces isolées de vins différents en Afrique

Occidentale (A. Guilliermond, 1914) appartiennent à S. cerevisiae et ont chacune l'ensemble uni-

que des gènes polymériques SUCI, SUC2 ек SUC3 pour la fermentation du saccharose.

KEY WORDS : Saccharomyces paradoxus, S. cerevisiae, taxonomy, electrophoretic karyotyping,

SUC genes.

INTRODUCTION

Although natural and cultural yeasts of the genus Saccharomyces Meyen ex

Hansen from Europe and Asia have been intensively studied during a century, there is

a short information about Saccharomyces isolated from other continents (do Carmo-

Sousa, 1969; Phaff & Starmer, 1987; van der Walt, 1970). At the beginning of the

century a well-known French mycologist A. Guilliermond (1914) described Saccharo-

myces chevalieri, S. lindneri and S. mangini species isolated from different wines in

Western Africa. In 50th J.P. van der Walt isolated several soil strains of S. cerevisiae

Hansen, S. coreanus Saito and S. uvarum Beijerinck in South Africa. Besides, wine

strains of S. capensis v.d. Walt & Tscheuschner, 5. coreanus and 5. uvarum are known

* Present address: Departamento de Microbiologia, Escuela Tecnica Superior de Ingenieros

Agronomos, Universidad de Cordoba, Apartado 3048, 14080 Cordoba, España.

Source : MNHN Paris

264 G.I. NAUMOV et al.

from that region and S. cerevisiae strains were isolated from Bantu beer (van der Walt,

1970). Since that time many nomenclatural changes have been made in the genus Sac-

charomyces (Barnett, 1992). Without additional studies most of the species mentioned

above cannot be assigned to any of the currently accepted biological species of the

Saccharomyces sensu stricto group: S. cerevisiae, S. paradoxus Batschinskaia and S.

bayanus Sacc. (Naumov, 1987; Naumov et al., 1992a, b; Vaughan Martini, 1989; Vau-

ghan Martini & Kurtzman, 1985, Vaughan Martini & Martini, 1987). Only genetic hy-

bridization analysis or DNA/DNA reassociation are suitable for delimiting the sibling

species.

In the present study we reidentified by genetical methods some yeast strains

isolated by A. Guilliermond and J.P. van der Walt in Western and South Africa. The

Strains are maintained at the Centraalbureau voor Schimmelcultures in Delft (List of

cultures, 1990). Some of their genetic peculiarities have also been studied. Among Af-

rican natural Saccharomyces yeasts we found unique population of biological species

5. cerevisiae and yeasts of biological species 5. paradoxus.

MATERIAL AND METHODS

Strains

The list of Saccharomyces strains studied and their origin are presented in Ta-

ble 1. The reference strains for biological sibling species were as follows: S. cerevisiae

- CBS 5287, ATCC 48498, X2180-1A and S. paradoxus - CBS 5829. The reference

strains, methods for cultivation and hybridization of yeasts have been described else-

where (Naumov, 1987; Naumov et al., 1986). Hybrids of homothallic yeasts were ob-

tained by "spore to spore" mating method using a micromanipulator.

CHEF gel electrophoresis

Chromosomal DNAs were prepared as described by Naumov et al. (1991).

Agarose slices were washed in 0.05 M EDTA, pH 8.0, prior to fractionation of chro-

mosomes by contour-clamed homogenous electric field (CHEF) gel electrophoresis. A

CHEF-DR™II apparatus (Bio-Rad, USA) was used to separate the chromosomal

DNAs. Agarose plugs containing chromosomal DNA were loaded into wells of a 1%

agarose gel in 0.5 x TBE (89 mM Tris, 89 mM borate, 20 mM EDTA, pH 8.2). Elec-

trophoresis was carried out at 200 V and 14°C for 15h with a switching time of 60 s

and then for 8h with a switching time of 90 s. After electrophoresis the gels were

stained with ethidium bromide for visualizing of the chromosomes. A standard set of

5. cerevisiae YNN 295 chromosomes was obtained commercially (Bio-Rad).

Southern blot analysis

After soaking the gels in 0.25 М НС! for 30 min, chromosomal DNA sepa-

rated by CHEF was denaturated, neutralized and transferred to nitrocellulose filters

which were then baked at 80°C for 2h. The SUC2 probe was a 0.9 kb BamHI-HindIII

fragment isolated from pRB117 (Carlson & Botstein, 1983). The probe was prepared

mainly according to Maniatis et al. (1982) and labeled with digoxigenin-11-dUTP us-

ing the Nonradioactive DNA Labelling Kit (Boehringer Manheim, FRG). Hybridiza-

tion was performed in 5 x SSC containing 0.1% N-laurol sarcosine, 0.02% SDS and

1% blocking reagent at 68°C overnight after which the filters were washed twice with

2 x SSC containing 0.1% SDS at room temperature for 5 min and with 0.1 x SSC

containing 0.1% SDS at 68°C for 15 min. Detection of hybridization was done with

Source : ММНМ Paris

SACCHAROMYCES FROM AFRICA 265

the Nonradioactive Kit. The filters were incubated in colour solution in the dark over-

night.

RESULTS AND DISCUSSION

Monosporic cloning

In genetic hybridization analysis only fertile monosporic parent strains should

be used. First, fertile homozygotic cultures of Saccharomyces sensu stricto were ob-

tained from single ascospores of collection strains listed in Table 1. All strains studied

showed high ascospore viability (89-100%) and were homothallic. For each strain 5-10

tetrads were dissected.

Genetic identification

Monosporic cultures of African Saccharomyces sensu stricto strains were

crossed with the reference strains of S. cerevisiae and S. paradoxus. The species de-

termination was judged on the basis of the viability of hybrid ascospores and the re-

combination of control markers (Table 2). Strains CBS 403, CBS 405 and CBS 2888

produced fertile hybrids with S. cerevisiae reference strain while their hybrids with 5.

paradoxus were sterile (Table 2). Thus, these strains belong to S. cerevisiae species.

On the contrary, strain CBS 2908 can be assigned to S. paradoxus species as it yielded

fertile hybrid only with S. paradoxus CBS 5829. In all intraspecies hybrids normal

meiotic segragation of control markers was observed. Strain CBS 400 was not in-

cluded in the crosses. Its belonging to the biological species 5- cerevisiae can be deter-

mined on the basis of hybridization analysis carried out by О. Winge and С. Robert

(1952).

Two strains CBS 400 and CBS 403 were previously studied by DNA/DNA re-

association. They showed high DNA homology with S. cerevisiae type culture (96%

and 87%, respectively) (Vaughan Martini & Kurtzman, 1985; Vaughan Martini &

Martini, 1987). Our genetic studies revealed in South Africa for the first time wild 5.

paradoxus (CBS 2908) and S. cerevisiae (CBS 2888) yeasts. Wild species 5. paradox-

us was previously isolated from a number of sites in Europe, Far East Asia and North

‘America (Naumov, 1987; Naumov et al., 1992а, 1993; Vaughan Martini, 1989). Wild

strains of S. cerevisiae occur very seldom іп nature and were found in Japan, Russian

Siberia and Finland (Naumov & Naumova, 1991; Naumov et al., 1992a; Naumov &

Nikonenko, 1988).

Identification of SUC genes

The fermentation of the sugars, viz. sucrose, maltose, o-methylglucoside, mel-

ibiose and starch is controlled in the yeast S. cerevisiae by the gene families (Barnett,

1981; Carlson et al., 1985; Naumov et al., 1991; Needleman, 1991; Pretorius & Mar-

mur, 1988; Winge & Roberts, 1958). Polymeric sugar genes are suitable as convenient

markers for strain identification but not for species delimination (Naumov, 1985). The

polymeric SUC gene family is known to contain 6 genes: SUCI (chromosome VID,

SUC2 (chromosome IX), SUC3 (chromosome ID, SUC4 (chromosome XIII), sucs

(chromosome IV) and SUC7 (chromosome VIII) (Carlson et al., 1985; Mortimer et al.,

1992). Each of the SUC genes encodes B-fructosidase (invertase) hydrolyzing sucrose

(Ottolenghi, 1971). ©. Winge and C. Roberts (1952) found that a strain of S. chevali-

eri (CBS 400) harbored three polymeric genes SUCI, SUC2 and SUC3. According to

our preliminary data strain CBS 405 had several SUC genes (Naumov, 1972). In this

Source : MNHN, Paris

266 G.I. NAUMOV et al.

Table 1 - Strains of Saccharomyces sensu stricto from which

monosporic cultures were used

Tableau 1 - Liste des souches de Saccharomyces sensu stricto

Species Strain Source Author

(original) designa-

designation tion

S. chevalieri CBS 400! palm wine, A. Guillermond

Ivory Coast

S. lindneri CBS 403! ginger A. Guillermond

wine,

West Africa

S. mangini CBS 4057 billi wine, A. Guillermond

West Africa

S. coreanus CBS 2888 soil, South J.R. van der Walt

Africa

S. cerevisiae CBS 2908 soil, South J.R. van der Walt

Africa

S. paradoxus CBS 5829 soil, V. Jensen

Denmark

S. cerevisiae ATCC wine, L.V. Turina

48498 Carpathians

Mountains,

Ukraine

S. cerevisiae CBS 5287 grape ber- I.A. Mazilkin

ries, Far

East of

Russia

5. cerevisiae YNN 295 genetic D. Vollrath &

line R.W. Davis

S. cerevisiae X2180-1A genetic R.K. Mortimer

line

Strain CBS 400 used was not monosporic. CBS 403 — VKM Y-407, CBS

405 = VKM Y-481, CBS 2888 = NRRL 12638, CBS 5287 = VKM Y-502

ATCC 48498 = M 437. ATCC = American Type Culture Collection

Rockville, U.S.A. CBS = Centraalbureau voor Schimmelcultures,

Delft, Holland. M = Magarach Institute of Viticulture and Wine

Making, Yalta, Ukraine. NRRL = Northern Regional Research

Laboratories Peoria, Ill., U.S.A. VKM = All-Russian Collection

of Microorganisms, Moscow, Russia. T = type culture.

connection, it was interesting to investigate the SUC genotypes of the S. cerevisiae

strains isolated from Africa.

Chromosomal DNAs of the strains studied were separated by pulsed field gel

electrophoresis (Fig. 1A). Strains CBS 400, 403 and 405 revealed karyotyping patterns

similar to one another and to references strain X2180-1A (Fig. 1A, lanes 3-5 and lane

2, respectively). Strain X2180-1A represents a “wild type" (in genetic terms) of 5. cer-

evisiae karyotype (Naumov et al., 1992b). Following electrophoresis the chromosomal

DNAs were transferred to nitrocellulose filter and hybridized with the SUC2 probe. In

Source : MNHN, Paris

SACCHAROMYCES FROM AFRICA 267

Table 2 - Genetic analysis of the hybrids of the biological

species S. cerevisiae (CBS 403, CBS 405, CBS 2888, ATCC 48498,

X2180-1A and CBS 5287) and S. paradoxus (CBS 2908 and CBS 5829).

Tableau 2 - Analyse génétique d'hybrides de S. cerevisiae (CBS

403, CBS 405, CBS 2888, ATCC 48498, X2180-1A et CBS 5287) et S.

paradoxus (CBS 2908 et CBS 5829).

origin of No. of No. of No. of Proportion Segrega-

hybrids spore zygotes tetrads of viable tion of

pairs obtai- isola- ascospores control

crossed ned ted of hybrids markers

a, (+:-)

5. cerevisiae x 5. cerevisiae

403 x 48498 30 3 21 90 2:2 (15)

405 x 48498 35 DI 22 83 2:2 (13)

2888 x X2180 55 3 29 42 24:21

S. paradoxus x S. paradoxus

2908 x 5829 69 6 29 66 39:38

5. cerevisiae x 5. paradoxus

403 x 5829 39 10 25 o =

405 x 5829 5 3 27 о -

5287 x 2908 37 5 23 о =

2888 x 5829 53 12 28 о =

Segregation of the control markers is in accordance with data of

random spore or tetrad analysis. Number of tetrads is indicated

in paranthesis. Reference strain no. 5829 was marked by а UV-

induced adenine (ade) auxotrophy (red colonies). Strains no. 403,

405, 48498, 2888 and X2180-1A have natural markers Mal, Mal’,

Mal’, баі” and Gal, respectively.

strains CBS 400, CBS 403 and CBS 405 isolated from different wines (Table 1), the

SUC2 probe hybridized to three different bands (Fig. 1B, lanes 3-5). Comparing with

standard strain YNN 295 having known order and sizes of chromosomes, these bands

correspond to chromosomes VII, П and IX to which the SUCI, SUC2 and SUC3 genes

respectively map. Additionally, the gene probes LYS2 (chromosome TI) (Eibel & Phi-

lippsen, 1983), LYS/ (chromosome IX) and TRPS (chromosome УП) (Balzi et al.,

1987) showed hybridization to the same bands as SUC2 probe did (data not shown).

Taking the data mentioned above into account, strains CBS 403 and CBS 405 are

more likely to have the same genotype as CBS 400: SUCI SUC2 SUC3. Strain CBS

2888 isolated from soil had only one SUC gene on chromosome IX (Fig. 1B, lane 6)

The cross with the reference strain X2180-1A (SUC2) confirmed that the only SUC

gene of strain CBS 2888 was allelic to SUC2, as no segregation of ability to ferment

sucrose was found.

Recently, we have studied by karyotyping and Southern analysis the SUC

genes of several dozens of natural S. cerevisiae strains isolated from different geo-

Source : MNHN, Paris

268 ОЈ. NAUMOV et al.

А 123456B123456

Figure 1 - Southern hybridization analysis of chromosomal DNAs from African S. cerevisiae

strains. Lane 1, YNN 295; lane 2, X2180-1A; lane 3, CBS 400; lane 4, CBS 403; lane

5, CBS 405; lane 6, CBS 2888. Ethidium bromide-stained gel (A) corresponding to hy-

bridization of chromosomal DNAs with the SUC2 probe (B). The linkage group number-

ing refers to the chromosomes of the strain YNN 295.

Figure 1 - Analyse Southern d'ADN chromosomique des souches de S. cerevisiae d'Afrique. Piste

1, YNN 295; piste 2, X2180-1A; piste 3, CBS 400; piste 4, CBS 403, piste 5, CBS 405;

piste 6, CBS 2888. Gel teint au bromure d'éthydium (A) correspondant à l'hybridation

d'ADN chromosomique avec la sonde SUC2 (B).

graphic regions (data not shown). Most of sucrose fermenting strains showed one

SUC2 gene strains non-fermenting sucrose possessed the silent sequences suc2°. It

seems that at least at the beginning of this centuary in Western Africa there was an

isolated population of yeast S. cerevisiae having an original set of SUC genes.

Source : MNHN. Paris

SACCHAROMYCES FROM AFRICA 269

This study showed that populations of Saccharomyces occurring far from Eu-

rope can have unique genetic constitution. Probably, the enlarged geographic screening

of natural Saccharomyces strains would allow revealing new genes in 5. cerevisiae.

ACKNOWLEDGEMENTS

This research was supported by Russian Fund for Basic Research N° 93 - 04-21836

(Russia) and by grant of Direccion General de Investigacion Cientifica у Técnica (Ministerio de

Educacion y Ciencia, España).

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Source : MNHN, Paris

Cryptogamie, Mycol. 1993, 14 (4): 271-277 271

KERATINOPHILIC AND CYCLOHEXIMIDE RESISTANT

FUNGI IN SOILS OF OMAN

A.H.M. EL-SAID

Botany Department, Faculty of Science, Qena, Egypt

ABSTRACT - Using goat hair fragments as baits at 28°C, 32 keratinophilic cycloheximide resis-

tant species and 1 variety belonging to 14 genera were collected from 50 soil samples gathered

from different places of Oman. Numerous keratinophilic fungi were isolated namely: Aphanoas-

cus sp. teleomorph of Chrysosporium tropicum, A. fulvescens teleomorph of C. keratinophilum, A.

terreus teleomorph of C. indicum, Arthroderma cuniculi, A. lenticulare teleomorph of Trichophy-

ton terrestre, A. tuberculata, A. curreyi, А. ciferii teleomorph of C. georgii, Chrysosporium carmi-

chaelii, C. lucknowense, С. pannicola, C. pruinosum, C. asperatum, С. xerophilum, Trichophyton

mentagrophytes, T. rubrum, Apinisia queenslandica teleomorph of С. queenslandicum, Mycelio-

phthora vellerea and Microsporum gypseum. Also several other saprophytic and cycloheximide

resistant fungi were isolated.

RÉSUMÉ - L'utilisation de fragments de poils de chèvre comme pidge à 28°C, a permis d'isoler,

а partir de 50 échantillons de sols récoltés à Oman, 32 espèces et une variété, appartenant à 14

genres, de champignons kéraünophiles résistant au cycloheximide. Les espèces les plus

fréquemment isolées sont: Aphanoascus sp. téléomorphe de Chrysosporium tropicum, A.

fulvescens téléomorphe de C. keratinophilum, A. terreus téléomorphe de C. indicum, Arthroderma

cuniculi, A. lenticulare téléomorphe de Trichophyton terrestre, A. tuberculata, A. curreyi, A. ciferii

téléomorphe de C. georgii, Chrysosporium carmichaelii, C. lucknowense, C. pannicola, С.

pruinosum, C. asperatum, C. xerophilum, Trichophyton mentagrophytes, T. rubrum, Apinisia

queenslandica téléomorphe de C. queenslandicum, Myceliophthora vellerea et Microsporum

gypseum. De nombreuses autres espèces de champignons saprophytes, résistant au cycloheximide

ont également été isolées.

KEY WORDS : Keratinophilic fungi, cycloheximide resistant fungi, soil bome fungi.

INTRODUCTION

Keratinophilic fungi are of importance and considerable significance and se-

veral investigations have been made on the contribution of these in soil of many coun-

tries all over the world (Ajello & Ziedberg, 1951; Ajello, 1952, 1954; Ajello et al.

1964; Randhawa & Sandhu, 1965; Ajello & Padhye, 1974; Caretta & Piontelli, 1975

Caretta et al., 1977; Crozier, 1980; Mc-Aleer, 1980; Sur & Gosh, 1980; Calvo et al.,

1984; Marsella & Mercantini, 1986; Sundaram, 1987; Chabasse, 1988).

In Arab countries, few surveys were carried out on keratinophilic fungi from

soil (Jana et al., 1979; Amer et al., 1975; Abdel Fattah et al., 1982; Abdel Mallek et

al., 1989; Youssef et al., 1989; Karam El-Din et al., 1990; Abdel Hafez et al., 1989a,

Source : MNHN. Paris

272 А.Н.М. EL-SAID

1991; EI Said, 1993). The present investigation aimed to study intensively composition

and frequency of occurrence of keratinophilic fungi in Omanian soil.

MATERIALS AND METHODS

Fifty soil samples were collected from different localities of Oman, according

to the method described by Johnson et al. (1959).

The soil samples were analysed chemically for the estimation of total soluble

salts, elements (Ca, Mg, K and Na) and organic matter. A pH-meter (WGPYE model

290) was used for the determination of soil pH. The soil type was determined by the

hydrometer method as described by Piper (1955) and most of samples are sandy.

Isolation of Keratinophilic fungi:

The hair baiting technique was employed as recommended by Vanbreuseghem

(1952), and as employed by Abdel-Fattah et al. (1982): 100g of soil were put in sterile

plate and a sufficient quantity of sterile distilled water (about 20-25% moisture con-

tent) was added and mixed throughly. Pieces of sterile goats hair were sprinkled on

the surface of the moistened soil. Two plates were used for each sample; the plates

were incubated at 28°C for 6-8 weeks, and the soil in plates were remoistened when-

ever necessary. The moulds which appeared on the baits were transferred to the sur-

face of Sabouraud's dextrose agar medium (Moss & Mc Quown, 1969) which was

supplemented with 20 units/ml of sodium penicillin, 40ug/ml of dihydrostreptomycin

and 0.05% cycloheximide (Actidione). Before adding to the agar, the first 2 antibiotics

were dissolved separately in sterile distilled water while the third was dissolved in

methanol. The plates were incubated at 28°C for 3-4 weeks and the developing colo-

nies were examined and identified.

RESULTS AND DISCUSSION

The soil samples tested were generally poor in organic matter content

(0.09-2.16% of dry soil) and their contents in total soluble salts widely ranged between

0.02-19.2%, in Ca: 0.02-2.17 mg, Mg: 0.02-0.66 mg, К: 0.02-0.51 mg, and Na:

0.02-2.2 mg/g dry soil. Abdel-Fattah (1973) found that the total soluble salts of Egyp-

tian desert soils varies between 0.4-6.6%. The PH values of the soils tested were all in

alkaline side (7.3-8.9).

Thirty-two keratoniphilic and cycloheximide resistant Species in addition to 1

variety which belong to 14 genera were collected from 50 soil samples baited with

goat hair fragments at 28°C (Table 1).

Aphanoascus teleomorph of Chrysosporium was the most common genus, oc-

curring in 52% of the samples. It was represented by 3 species of which Aphanoascus

sp. teleomorph of Chrysosporium tropicum was the most common species and was re-

presented in 28% of the soil samples. It was dominant species in Italy soils (24.5% of

the samples) as recorded by Todaro (1978). It was represented in 20.8 and 12.4% of

the soil samples in Marrakesh and Casablanca (ana et al., 1979). In India, С. tropi-

cum occurred in 18% of the soil samples tested (Sur & Ghosh, 1980); in Galapagos Is-

lands in 5.3% (Ajello & Padhye, 1974); in Chilean Andes in 3.9% (Piontelli & Caret-

ta, 1974); in soil of Volcano Etna, 20.5% (Caretta et al., 1977); in Spain in 24%

(Calvo et aL, 1984); and Yemen in 24% (El-Said, 1993). In Egypt, C. tropicum was

the most common fungal species recovered by baiting, comprising 36.6% of the total

Source : MNHN, Paris

KERATINOPHILIC FUNGI OF OMAN 273

fungal isolates (Mostafa, 1977). Abdel-Hafez et al. (1991) isolated C. tropicum in

25.3% of the soil samples collected from Eastern desert in Egypt. Abdel-Fattah et al.

(1982) isolated this species in 11.4% of the soil samples collected from Assiut Gover-

norate.

Table 1: Numbers of cases of isolations (NCI: out of 50), percentage frequency (%F) and occur-

rence remark (OK) of fungal genera and species recovered from 50 soil samples baited with goats

hair at 28°C.

Genera and species OR

Aphanoascus H

‘Aphanoascus sp. M

A. fulvescens (Cooke) Apinis L

A. terreus (Randhawa & Sandhu) Apinis в

Arthroderma M

A. cuniculi Dawson L

A. lenticulare Pore, Tsao & Plunkett R

A. tuberculata Kuehn R

A. сштеуі Berk R

A. ciferri Varsavsky & Ajello R

Chrysosporium 18 | 36 | M

C. carmichaelii Van Oorschot 6 12 it

С. lucknowense Gug 5 10 R

C. pannicola (Corda) Van Oorschot & Stalpers 5 10 R

C. pruinosum Gilman & Abbott 5 10 R

C. asperatum J.W. Carmichael 3 6 R

C. xerophilum Pitt 3 6 R

Aspergillus 17 34 M

А. flavus Link 8 16 E

A. fumigatus Fresenius 6 12 L

A. ustus (Bainier) Thom & Church 6 12 T

A. flavus var. columnaris Raper & Fennell а] 10 R

A. terreus Thom 4 8 R

A. niger Van Tieghem 3 6 R

Trichophyton Il 2 D

T. mentagrophytes (Robin) Blanchard 6 12 L

T. rubrum (Castellani) Sabouraud 5 10 R

Penicillium 7 14 È

P. chrysogenum Thom 6 12 L

P. funiculosum Thom 1 2 R

P. puberulum Bainier 1 2 R

Apinisia queenslandica Apinis & Rees 3 6 R

Cunninghamella echinulata Thaxter 3 6 R

Fusarium 3 6 R

F. oxysporum Shelecht 2 4 R

F. moniliforme Sheldon 1 2 R

Macrophomina phaseolina (Tassi) Goid 3 6 R

Mucor racemosus Fresenius 2 4 R

Verticillium lateritium Berkeley 2 4 R

Myceliophthora vellerea (асс. & Speg.) Van Oorschot 2 4 R

Microsporum gypseum (Bodin) Guiart & Grigorakis = 2 R

Occurrence remark: H = high occurrence; between 25 to 50 cases (out of 50 samples). M = mod-

erate occurrence; between 13 to 24 cases. L = low occurrence; between 6 to 12 cases. R = rare

occurrence; between 1 to 5 cases.

Source : MNHN, Paris

274 A.H.M. EL-SAID

A. fulvescens teleomorph of C. keratinophilum was the second most frequent

fungal species and was encountered in 20% of the soil samples tested. C. keratinophi-

lum emerged from 6% on children playgrounds sand samples (Bojanovsky et al.,

1979); from 13.2% of soil samples in W. Germany (Meissner & Qadripur, 1983); from

16.9% of soils of the Volcano Etna (Caretta et al., 1977); from 10% of the screened

soils of Spain (Calvo et al., 1984). In Yemen this species represented 14% of the soil

samples as recorded by El-Said (1993). In Egypt, Abdel-Fattah et al. (1982) found

this species emerged in 27.1% of soil samples collected from Assiut Governorate, but

Abdel-Hafez et al. (1991) recovered this species from Egyptian soils baited with ani-

mal hair (20% of the soil samples tested).

A. terreus teleomorph of C. indicum was the third most frequent fungal species

and was represented in 18% of the soil samples, whereas it was the most frequent spe-

cies in soil samples collected from Yemen (El-Said, 1993) and in cultivated soils col-

lected from Upper Egypt (Abdel Fattah et al., 1982), but it was less frequent in Egyp-

tian soils tested by Mostafa (1977), Abdel-Fattah et al. (1982) and Abdel-Hafez et al.

(1991). In India, it emerged from 31.3% of the soil samples (Sur & Ghosh, 1980); in

mountains localities in the Chilean Andes, from 19.8% (Piontelli & Caretta, 1974); in

Galapagos Islands from 2.6% (Ajello & Padhye, 1974); and Spain, from 4% (Calvo et

al., 1984) of the soil samples tested.

Arthroderma was the second most frequent genus and was encountered in 38%

of the samples tested. From the genus 5 species were collected of which A.cuniculi

was the most common species. The remaining Arthroderma species were scarcely re-

covered and these were A. lenticulare teleomorph of Trychophyton terrestre, A. tuber-

culata and A. ciferii teleomorph of С. georgii. The above species were also isolated

from soil samples collected from Yemen (El Said, 1993) and Egypt (Abdel-Hafez et

al., 1989a, 1991).

Chrysosporium occupied the third place with regard to the number of cases of

isolation of fungal genera and it recovered from 36% of the samples examined, Six

species of chrysosporium were isolated and these were C. carmichaelii (12%), С. luck-

nowense (10%), С. pannicola (10%), С. pruinosum (10%), С. asperatum (6%) and С.

xerophilum (6%). All the above species were isolated from the soil samples of Yemen

by El-Said (1993) and were emerged from 10, 10, 12, 12, 4 and 10%, respectively. In

Egypt, C. asperatum and С. pannicola were isolated from Egyptian soils by Abdel-Ha-

fez et al. (1989a, 1991).

Aspergillus (5 species + 1 variety) occupied the fourth place according to the

number of cases of isolation of fungal on genera and it encountered in 34% of the soil

samples. Among Aspergillus species, the most commonly collected were A. flavus, A.

fumigatus and A. ustus. The remaining Aspergillus species were isolated with rare fre-

quency of occurrence and these were A. flavus var. columnaris, A. terreus and A. niger.

Aspergillosis due to A. fumigatus and A. flavus has a world-wide distribution (Frey et

al. 1979). Most of the above species had been previously encountered, but with differ-

ent incidences from various types of soil from many parts of the world (Sundaram,

1987; Abdel-Hafez et al., 1989a; El-Said, 1993).

Trichophyton encountered from 22% of the samples tested. From the genus 2

species were collected of which Т. mentagrophytes was common and recovered from

12% of the samples. It is a human and animal dermatophyte (Nawok, 1970; Fleming,

1975; Frey et al., 1979). It emerged from 1% of sand samples from children's play-

grounds in Germany (Bojanovsky et al., 1979), from 68% of soil samples in Kuwait

(Amer et al., 1975) from 3% in England (Baxter, 1969) from 12% in Yemen (El-Said,

1993) and from 6% in Egypt (Abdel-Hafez et al., 1989a). Т. rubrum was less frequent.

Source : MNHN. Paris

KERATINOPHILIC FUNGI OF OMAN 275

Penicillium emerged in 14% of the soil examined. It was represented by 3

species: Р. chrysogenum, P. funiculosum and P. puberulum. Abdel-Hafez et al.

(1989b) isolated P. chrysogenum from the mud of Ibrahimia (Egypt). El-Said (1993)

isolated all the above species from the soil samples of Yemen.

The remaining isolated 8 genera and 9 species were recovered in rare frequen-

cies as present in Table 1.

Present results reveal that there is no correlation between the distribution and

occurrence of keratinophilic and cycloheximide resistant fungi and soil textures or site

of samples. But soil samples with low levels of total soluble salts coincided with a

wide range of genera and species and vice versa; this due to most of these fungi are

highly sensitive to high salinity. Abdel-Hafez et al. (1989a) found that soil samples

collected from salt marshes in Sinai Peninsula are free from keratinophilic fungi.

Aphanoascus teleomorph of Chrysosporium was the most frequent keratinophilic genus

in the soil samples from Oman as in case of other Egyptian soil collected from Delta

area, Upper Egypt, Sinai and Eastern Desert (Abdel-Fattah et al., 1982; Mostafa, 1977;

Abdel-Hafez et al., 1989a, 1991). This results agree with those which were recorded

by El-Said (1993) in the soil samples of Yemen.

Comparison between the present results and lists of keratinophilic and cyclo-

heximide resistant fungi recovered from soils collected from Egypt (Abdel-Fattah et

al., 1982; Mostafa, 1977; Abdel-Hafez et al., 1989, 1991) and Yemen (El-Said, 1993)

reveal that there is no keratinophilic or cycloheximide resistant fungi characteristic of

Omanian soils. But, these lists may differ in the order of frequency of occurrence of

some fungi.

ACKNOWLEDGEMENT

The authors wish to thank Mr. Y.A.M. Gherbawy for valuable technical assistance.

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Source : MNHN, Paris

Source : MNHN. Paris

Cryptogamie, Mycol. 1993, 14 (4): 279-286 279

SCUTELLOSPORA CASTANEA, A NEWLY DESCRIBED

ARBUSCULAR MYCORRHIZAL FUNGUS.

Christopher WALKER), Vivienne GIANINAZZI-PEARSON®)

and Huguette MARION-ESPINASSE®)

(1) The Forestry Authority, Northern Research Station, Roslin,

Midlothian EH25 9SY, UK.

(2) Station de Génétique et d'Amélioration des Plantes de Dijon, INRA,

BV1540 - 21034 Dijon Cedex, France.

(3) IBEAS, Department of Biology, Pau University, Faculty of Science,

64000 Pau, France.

ABSTRACT - Scutellospora castanea, a newly described fungus forming arbuscular mycorrhizas

was isolated from soil beneath Larhyrus sylvestris in France. It was established in pot culture with

Allium porrum and has been maintained for more than ten years on various host plants. The spe-

cies is characterised by its shiny, more or less globose, chestnut-coloured spores which are on av-

erage 300 jim in diameter and which possess a brittle outer wall group enclosing a flexible inner

wall group on which is formed an indistinct germination shield. It can be distinguished from 5.

arenicola mainly by the lack of an amorphous spore wall, and from S. erythropa on the basis of

spore size and wall structure.

RÉSUMÉ - Une nouvelle espàce de champignon, Scutellospora castanea, formant des mycorhizes

arbusculaires a été isolée d'un talus prés de Pau en France. La culture de ce champignon a été

établie sur Allium porrum en pot et ensuite maintenue pendant plus de dix ans sur différentes

plante-hótes. Cette espèce fongique forme des spores d'environ 300 um de diamètre qui se

caractérisent par une surface lisse et brillante, une forme plus ou moins globuleuse et une couleur

châtaigne. Elles possèdent une paroi externe cassante et une paroi interne flexible sur laquelle se

forme une plaque de germination mal définie. Scutellospora castanea se différencie de S.

arenicola principalement par l'absence de paroi amorphe, et de 5. erythropa par la taille et la

structure pariétale des spores.

KEY WORDS : Arbuscular mycorrhizas, Scutellospora castanea, Gigasporaceae, Glomales.

INTRODUCTION

A sample of soil (pH 7.7 in deionised water; Olsen-extractable P 11 ppm),

taken from beneath Lathyrus sylvestris L. on a roadside verge near Pau, Toulouse in

1983, was subjected to wet-sieving and decanting (Gerdemann & Nicolson, 1963).

Amongst the spores of members of the Glomales obtained in this way were those of

an apparently undescribed member of the arbuscular-mycorrhizal genus Scutellospora.

Some of these spores were used to inoculate seedlings of Trifolium pratense L. grow-

ing in irradiated clay loam (Epoisses) soil of pH 7.4 (in deionised water). The result-

ing pot cultures produced arbuscular mycorrhizas and abundant spores after a few

Source : ММНМ Paris!

280 С. WALKER et al.

months. The isolate thus obtained has since been repeatedly sub-cultured on several

hosts including Allium сера L., A. porrum L., Plantago major L. and Р. lanceolata І.

MATERIAL AND METHODS

Spores for the species description were extracted by a centrifugation-flotation

technique (Walker et al., 1982) or by shaking pot culture substrate in water and de-

canting the supernatant onto a 160 рт sieve. The spores were then suspended in a

dish of water and examined first under a dissecting microscope with reflected light and

later under a compound microscope with brightfield illumination or with Nomarski

Differential Interference Contrast. Roots were removed from pot cultures, subjected to

clearing and staining (Phillips & Hayman, 1970), and examined under a compound mi-

croscope for the establishment of mycorrhizas and associated extra-matrical mycelium.

In an attempt to standardise colour matching, spores suspended in water were

illuminated with light from a quartz-iodine fibre-optic source at a colour temperature

of 3200 K and examined under a dissecting microscope. Their colours were then com-

pared with those on a fungal colour chart (Anon, 1969) illuminated simultaneously by

a split fibre-optic from the same source. For more detailed examination, spores were

mounted on microscope slides in water (Spain, 1990), or polyvinyl alcohol lacto-gly-

cerol (PVLG) (Koske & Tessier, 1983). Colour matching of structures viewed with

transmitted light under a compound microscope is more difficult than with a dissecting

microscope, so for such observations colours (for example, of individual walls in the

description) were not precisely matched to a chart, and only generalised colour de-

scriptions are used.

Type material, consisting of a holotype collection on a microscope slide, and

isotype material on microscope slides, in dried soil, and preserved in both 5% formal-

dehyde solution and 0.025% NaN, has been lodged at the herbarium of the Royal Bo-

tanic Garden, Edinburgh (E). Isotype material has also been lodged at Oregon State

University (OSC). The species description is based on the style adopted by Gerdemann

& Trappe (1974), with modifications of spore wall terminology as suggested by Walk-

er (1983). Some of the terminology of Spain et al. (1989) has been used to describe

the hypha upon which the spore is borne.

SPECIES DESCRIPTION

Scutellospora castanea Walker sp. nov. Figs. 1-16

Sporae in solo singillatim efformatae, ad basis bulbosa terminales vel laterales,

globosae vel subglobosae, raro ovoideae vel obovoideae, 169-369 x 176-372 um; ju-

ventute candidae, opacae, maturitate ochraceae vel avellaneae, crescenter vacuola tae

e translucentes. Tunicae sporae stratis quatuor in turmis duabus. Turma externa stratis

duobus: stratum extimum laeve, nitidum, brunneum, 2-4 um crassum; stratum internum

lamellatum, hyalinum vel luteolum, 10-25 (-35) um crassum. Turma interna stratis duo-

bus: stratum extimum tenuissimum, hyalinum, flexile, minus quam 1 Wn crassum; stra-

tum internum flexile, 1-2 шт crassum. Turma interna scutello germinationis ovoideo,

complexo, hyalino vel luteolissimo, usque ad 208 x 181 ут. Basis bulbosa 38-51 um

crassa. Cellulae auxiliares pallide avellaneae vell brunneolae, singulares vel fascicula-

tae, prominentiis nodosis, obtusis.

Spores borne singly in the soil, terminally or laterally on a bulbous base; glo-

bose to subglobose, rarely ovoid or obovoid, 169-369 x 176-372 шт; pure white and

Source : MNHN. Paris

SCUTELLOSPORA CASTANEA sp.nov. 281

opaque when immature, shading through ochraceous (8G) to sienna (11) and becoming

increasingly vacuolate and translucent with maturity (Fig. 1). Spore wall of 4 walls

organised in 2 groups (Fig. 10). Outer group A of 2 walls (Fig. 3): wall 1 loosely ad-

herent, smooth, shiny, brown, unit, 2-4 um thick; wall 2 hyaline to pale yellow, lami-

nate, 10-25 (-35) um thick. Inner group В of two walls (Fig. 5 & 9): wall 3, very thin,

hyaline, flexible (membranous), less than 1 pm thick, closely adherent to wall 4; wall

4 flexible (membranous), 1(-2)mm thick. Germination shield on wall group B in ma-

ture spores hyaline to very pale yellow, with complex infolding of the edges, up to

208 x 181mm (Fig. 7). Bulbous base 38-51 mm wide (Fig. 2). Auxiliary cells pale

yellow brown to pale brown, single or clustered, with blunt, knobby projections.

Reaction to PVLG/Melzer's reagent (5:1 v/v) is variable, probably depending

on spore maturity. In wall group A, wall 1 becomes a deeper brown and wall 2 occa-

sionally becomes pink or red, but usually acquires a deeper yellow colour (Fig. 4). In

wall group B, neither wall reacts to Melzer's reagent, but the thin-walled, complex ger-

mination shield sometimes darkens to become a pale brownish yellow. This structure

is usually difficult to observe due to it being thin, a pale colour and the masking

caused by the brown pigmentation in wall group A, but it can be visualised by bleach-

ing spores (Fig. 7).

The bulbous base (= bulbous suspensor-like cell of Gerdemann & Trappe

(1974) or sporogenous cell of Spain et al. (1989)), which is concolorous with the outer

spore wall layers (Fig. 2) and can have up to five hyphal projections, is borne termi-

nally on a septate sporophore 14-20 шт wide formed from a broad, pale yellow-

brown, coenocytic hypha approximately 7-15 jim wide. The bulbous base detaches

particularly easily in this species, and is missing from a large proportion of spores ex-

tracted by processes involving sieving.

S. castanea forms endomycorthizas (Figs 11-16) with hyphae developing

appressoria (Fig. 12), H-connections (Abbott, 1982) (Figs 13 & 15), hyphal coils (Fig

14) and arbuscules (Fig. 16) but without vesicles.

DISCUSSION

There are only two other described species of Scutellospora possessing brown

spores with a smooth, shiny surface. These are S. erythropa Koske & Walker and S.

arenicola Koske & Halvorsen. Spores of the former have a greater size range with lar-

ger maxima (170-551 x 205-660 compared with 169-369 x 176-372 рт), are much

darker in colour, and have much more opaque outer walls than those of 5. castanea.

They also have a different wall structure and a much more distinct and robust germi-

nation shield (Koske & Walker, 1984). Scutellospora arenicola and S. castanea spores

are similar in colour, have similar size ranges (160-360 x 120-310 um for 5.

arenicola) and possess similarly indistinct germination shields. However, the wall

structures of 5. arenicola and 5. castanea differ in one important respect. The former

has one wall more than the latter - an amorphous innermost wall (Fig. 6) that reacts

distinctively to Melzer's reagent to give a purple reaction (Koske & Halvorsen, 1989).

Differences of this nature are considered to result from major evolutionary change at-

tributable to phylogenetic grouping such as species (Morton et al., 1992).

Unusually for a species of Scutellospora, wall 1 (the outer, unit wall) can be

detached with relative ease from wall 2 as the spore is crushed. The wall does not de-

tach completely, however, but cracks radially, and then separates in parts (Fig. 3). Oc-

casional spores can be seen to have lost patches of the outer wall even under the dis-

Source : MNHN, Paris

282 С. WALKER et al.

SCUTELLOSPORA CASTANEA sp.nov. 283

1 2 34

Fig. 1 - Freshly extracted spores suspended in water and illuminated by incident light. Immature

(lower arrow) and over-mature (upper arrow) spores are indicated. The vacuolate nature

of the spore contents in mature, healthy spores can be seen in the majority of the re-

maining spores. Scale bar 250 um.

Fig. 2 - Detail of spore showing the bulbous base. The point of connection of outer and inner

wall groups is arrowed, Scale bar 50 um.

Fig. 3 - Crushed spore mounted іп РУС, showing walls 1 and 2 in group A separating, and

walls 3 and 4 in group В remaining together. Scale bar 100 jum.

Fig. 4 - This preparation shows two heavily crushed spores (labelled a and b). Spore 'a' (right of

dividing line) was crushed in PVLG. Spore 'b' (left of dividing line) was crushed in

PVLG with Melzers reagent. The spores were then re-mounted in PVLG. Walls 1 & 2

have reacted in the Melzer's reagent (arrowed bl and b2), whereas walls 3 and 4 have

not. Scale bar 200 jum.

Fig. 5 - A spore crushed heavily іп PVLG. Walls 3 and 4 in the inner wall group are arrowed

and numbered. Scale bar 250 um.

Fig. 6 - A similar preparation to that in Fig 5, but from the isotype material of Scutellospora ar-

enicola, showing the additional wall (wall 5). Scale bar 250 pm

Fig. 7 - A spore bleached with undiluted domestic bleach to reveal the germination shield. The

lumen at the point of origin is arrowed. Scale bar 50 pm

Fig. 8 - Knobby auxiliary cells of Scutellospora castanea. Scale bar 25 pm

Fig. 9 - Detail of the inner wall group of S. castanea (walls 3 and 4 arrowed). Scale bar 100 pm.

Fig. 10 - Murograph of S. castanea.

Source : MNHN, Paris

284 С. WALKER et al.

Figs 11-16 - Mycorthiza of Allium porrum and Scutellospora castanea.

Fig. 11 - Cleared and stained root squash showing the hyphal elements stained with

trypan blue. The stele is at the bottom of the photograph. A small cluster of auxiliary

cells is seen on the root surface (ac). Fig. 12 - Lateral view of an appressorium-like

structure (ap) with resultant coarse, intra- and inter-cellular hyphal elements colonising

the root. Fig. 13 - Plan view of an appressorium-like structure on the root surface. Fig.

14 - Lower focus of Fig 13, showing the hyphal coil occupying a cell beneath the.

appressorium-like structure. Fig. 15 - Inter-cortical hyphae, showing H-connections.

Fig. 16 - Arbuscule in a cortical cell

Source : MNHN, Paris

SCUTELLOSPORA CASTANEA sp.nov. 285

secting microscope at magnifications as low as 30x. This phenomenon was also

reported for spores of S. arenicola (Koske & Halvorsen, 1989).

Stimulated by the work of Spain (1990), fresh specimens were examined in

water. No great differences were noted in apparent wall structure when compared with

PVLG-mounted specimens, except that in the latter, wall 3 was a little easier to see.

On some specimens, both in water and PVLG, walls 3 and 4 were so tightly adherent,

that they were only distinguishable under magnifications of greater than 500 X. This

was particularly the case for spores that had been preserved in 0.025% sodium azide

solution. The laminae in wall 2 can be so fine that it appears superficially to be a unit

wall, though splits forming along a lamina in some spores gave a clue to its true na-

ture, and when heavily crushed, the laminae can be seen as steps where they have bro-

ken differentially. The bulbous structure at the base of the spore is not a cell, but is

continuous with the outer walls of the spore. We have therefore not used the terms

‘bulbous suspensor-like cell! (Gerdemann & Trappe, 1974) or 'sporogenous cell' (Spain

et al., 1989). The walls can best be understood through their development series. In

white spores, the earliest development is the production of an apparently 2-layered

wall (walls 1 & 2 in group A), formed directly by further development of the walls of

the suspensor-like cell. In some specimens, there seems to be a hyaline layer between

walls 1 and 2, but this is not always observable, and has not been considered to be a

separate wall. A similar wall or wall layer was described as being present in S. grega-

ria Koske & Walker, though in that species it was considered to be a separate lami-

nated wall (Koske & Walker, 1985). As the spores darken with age, both the outer-

most wall and the contents change colour. After the spores have started to change

colour, the flexible walls develop, first by formation of the extremely thin flexible

membranous wall (wall 3), and then by the production of the innermost flexible wall

(wall 4). The thin, membranous wall (wall 3) forms an endosporangium-like structure

enclosing the second flexible wall that forms a single entity similar to a sporangios-

pore.

Unlike most Scutellospora spp., the flexible walls of 5. castanea do not readily

separate from the outer wall group. Consequently, on casual observation, even mature

spores of this species can appear to belong to the genus Gigaspora. This misappre-

hension is reinforced by the difficulties of seeing the germination shield because of the

lack of either colour or thickening at its edges, and by the presence in some specimens

of a layer that appears to be similar to the germinal wall of Spain et al. (1989). Be-

cause this is detectable only on some spores, we have assumed it merely to be an in-

nermost lamina of wall 2, but ultrastructural study may prove this interpretation to be

erroneous.

Scutellospora castanea possesses the hyphal bridging or wound healing de-

scribed by Gerdemann (1955). However, it should be noted that this phenomenon does

not always result from wounding, but can occur on lengths of apparently healthy hy-

phae as a result of branching and later anastomosis. The hyphae may also form myce-

lial strands by repeated branching and anastomosis both within a hypha and between

adjacent hypha.

ACKNOWLEDGEMENTS

We wish to thank C. McEvoy, Forestry Authority, for help in preparation of the illus-

trations, F. Contour (INRA) and K. Clifford (Forestry Authority) are also acknowledged for their

skill in maintaining pot cultures. Thanks are also due to Drs M. Giovannetti, University of Pisa,

and D. Redfern, Forestry Authority, for reviewing the manuscript, to Dr J.M. Trappe, Oregon

State University, for preparing the Latin diagnosis and to Dr R.E. Koske, University of Rhode Is-

Source : MNHN, Paris}

286 С. WALKER et al.

land, for providing specimens of 5. arenicola. This work is the result of European cooperation

within the COST Action 8.10 (VA Mycorrhizas) group оп a Bank of European Glomales, and 5:

castanea is accession n°1 of the fungal germplasm database.

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Source : MNHN, Paris

Cryptogamie, Mycol. 1993, 14 (4): 287-295 287

BIOLOGICAL CONTROL OF DRECHSLERA TERES:

ABILITY OF ANTAGONISTS TO REDUCE CONIDIA

FORMATION, COLEOPTILE INFECTION AND LEAF

INFECTION IN BARLEY (HORDEUM VULGARE)

Mostafa-Mahmoud MOSTAFA.

Laboratoire de Biologie Appliquée,

Institut Universitaire de Technologie Louis Pasteur

3 rue de l'Argonne, F-67000 Strasbourg, France.

ABSTRACT - Preventive seed application of an Actinomycete, Trichoderma viride, Myrothecium

verrucaria or Trichoderma pseudokoningii reduced seed-borne Drechslera teres infection of bar-

ley coleoptiles by 90%, 87%, 79% and 76%, respectively. Moreover, leaf infection after artificial

inoculation was decreased by 87%, 84%, 79%, 77% and 71% when the Thibaut cultivar had been

previously treated at the three-leaf stage with Myrothecium verrucaria, Actinomycete, Trichoder-

ma sp., Trichoderma viride or Trichoderma pseudokoningii, respectively. These antagonists also

reduced conidia formation on straw colonized by D. teres.

RÉSUMÉ - L'étude sur orge а confirmé l'efficacité des germes antagonistes (Actinomycete,

Trichoderma viride, Myrothecium verrucaria, Trichoderma sp. et Trichoderma pseudokoningit)

sur les trois séquences épidémiologiques majeures de l'helminthosporiose causée par Drechslera

teres. L'inoculum séminicole (réduction du niveau d'attaque coléoptilaire par l'Actinomycète

90%, Trichoderma viride 87%, Myrothecium verrucaria 79% et Trichoderma pseudokoningit

76%). La contamination aérienne (diminution de l'importance et du nombre de lésions foliaires:

Myrothecium verrucaria 87%, Actinomycete 84%, Trichoderma sp. 79%, Trichoderma viride

77% et Trichoderma pseudokoningii 71%). Pour la conidiogénèse sur pailles, on note des actions

antagonistes significatives de Myrothecium verrucaria et de l'Actinomycète appliqués sur les

résidus de récolte.

KEY WORDS : biological control, barley, antagonism, barley net blotch, Hordeum vulgare,

Drechslera teres.

INTRODUCTION

Drechslera teres (Sacc.) Shoem. (anamorph of Pyrenophora teres f. teres

Drechs.) is the causal agent of barley net blotch. This cereal disease can cause consid-

erable damage in a number of barley-growing areas in various parts of the world

(Smedegard-Petersen, 1974; Sutton & Steele, 1983; Martin, 1985). The yield reduction

in barley plants infected with D. teres results mainly from decreased weight of grain,

whereas the number of grains per plant is usually less affected (Smith et al., 1988).

Source : MNHN, Paris

288 M. MOSTAFA

This pathogen predominantly attacks the leaf and overwinters on infected de-

bris and grain. Inoculum destruction and barriers to prevent colonization of the plant

by the pathogen are of primary importance for efficient control of the disease (Shipton

et al., 1973).

Biological control, in its broadest sense, which includes rotations and other ag-

ricultural practices, has been used since prehistory (Campbell, 1990). The direct use of

micro-organisms isolated from the soil to inoculate plants and reduce the disease was

first reported in a series of papers around 1920-1930 (Campbell, 1990), but these ef-

fects remained largely a scientific curiosity until comprehensive books (Baker & Cook,

1974; Cook & Baker, 1983 and Mukerji & Garg, 1988) collected and analysed the

available knowledge and stimulated further research, which resulted in many laborato-

ту investigations but few effective field trials (Campbell, 1990).

Treatment of wheat and barley seeds with Streptomyces griseoviridis decreased

damage caused by Fusarium spp. and Bipolaris sorokiniana on both inoculated and

uninoculated seeds (Tahvonen & Avikainen, 1990). Antagonists isolated from the resi-

dent microflora would seem to be good candidates for controlling foliar diseases, espe-

cially if a strategy of early introduction and establishment as residents is to be fol-

lowed (Spurr, 1981).

Certain groups of bacteria, including fluorescent Pseudomonas, Xanthomonas

and the Erwinia spp., are in relatively high densities on leaf surfaces and non patho-

genic members of these groups have been recognized as potential biocontrol agents

(Mukerji & Garg, 1988).

Apart from bacterization, inoculation of seeds with fast growing fungi can pre-

vent seed decay and seedling blight. Inoculation of corn seeds with Chaetomium glo-

bosum has resulted in field control of seedling blight caused by Fusarium graminea-

rum. Similarly oat seeds coated with Chaetomium spp. have some provided control of

Helminthosporium victoriae (Purkayastha, 1989).

The aim of the present study was to assess the ability of some antagonists

against D. teres on barley to suppress conidia formation and to reduce both coleoptile

and leaf infection.

MATERIAL AND METHODS

Antagonists

Isolates of Trichoderma pseudokoningii/N69, Trichoderma viride/NRG-1, Tri-

choderma sp/BRL 124, Myrothecium verrucaria/N76-1 and actinomycete/N51 were

selected from 50 strains of fungi, actinomycetes and bacteria isolated from soil and

straw obtained from the Department of Phytopathology, ENSAT, Toulouse University,

France and tested in vitro for their ability to control net blotch caused by D. teres

(Mostafa et al., 1992). These fungal isolates were grown on PCA (potato 20 g, carrot

20 g and agar 20 g in 1000 ml distilled water) in Petri dishes for 15 days using agar

disks from stock culture under laboratory conditions.

Actinomycete/N51 was grown in a liquid culture medium (meat extract 2 g,

peptone 2 р and NaCl 5 g in 1000 ml distilled water) for 8 days under laboratory con-

ditions as shake cultures. Aqueous suspensions of all the antagonists were prepared

and adjusted to 1 x 10° propagules/ml with a haemocytometer.

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BIOLOGICAL CONTROL OF DRECHSLERA TERES 289

Pathogen

A 5 mm disk of mycelium of D. teres (isolate R3) was taken from the edge of

8-day-old fungal colony on PCA in a Petri dish and cultured in 50 ml of 10% V8 liq-

uid medium. After incubation at 23°C for 10 days, the mycelium was ground in water

with a blender for 1 min to produce approximatively 1 x 10% mycelial fragments per

milliliter. Gelatin (0.25%) and Triton X100 (one sticker drop per 100 ml) were added.

Treatment of barley seeds

Seeds of barley cultivar Thibaut were surface sterilized using 95% ethanol (5

min) then 1% HgCl, (3 min) and carefully rinsed in sterilized distilled water (5 x 5

min). The seeds were dipped into the propagule suspensions of the antagonists and al-

lowed to dry for 24 h. After dipping the seeds into mycelium suspensions of D. teres

they were placed in test tubes containing cotton soaked in a nutrient solution (NaNO;

1 в, KNO; 0.25 в, MgSO,. 7 H,O 0.25 в, KH;PO, 0.25 g and FeCl, 0.001 g, in dis-

tilled water 1000 ml), and the tubes were incubated at 18°C with 12h daylength. When

the 2nd leaf was almost fully expanded, the necrotic areas on the coleoptiles were ex-

amined. A control set of seeds was inoculated with D. teres alone and another set with

sterilized distilled water. Each treatment included ten replications (10 x 5 seeds)

Treatment of barley leaves

Seeds of barley cultivar Thibaut were sown in pots (10 seeds/pot, with 6 repli-

cates per treatment), and when the 3rd leaf was almost fully expanded spore suspen-

sions of the antagonists were sprayed over the upper surface of all the leaves. Then

after incubation under polyethylene bags for 24 h at 18°C, the leaves were inoculated

with mycelial suspensions of D. teres. The pots were again covered with polyethylene

bags, for 48 h, and then transferred to a glasshouse for disease development. Through-

out the experiments, the temperature was in the range 18-20°C, and the relative hu-

midity was 85-95% with a 12 h daylength (light intensity = 60 Wm?).

After 15 days, the incidence% (number of leaves infected per treatment) and

severity (leaf area infected using a scale (Table 1) from 0 (no disease) to 9 (maximum

disease) were evaluated and the coefficient of infection (CI) was calculated by multi-

plying the incidence by the severity (Loegering, 1959).

Treatment of barley straw

Samples of barley cultivar Thibaut stubble were collected from the fields and.

the straw naturally covered with conidiophores of D. teres was cut into 8 cm lengths.

These were soaked in conidial suspensions of the antagonists (1 x 10* conidia/ml), and

a control sample was soaked in sterilized distilled water. Three replicated Petri dishes

were set up with 10 fragments of straw per dish (100% RH; 12 h daylength; 18°C),

and covered so as to promote conidia production. After 4 and 6 days, conidiophores

and conidia were counted microscopically after washing them with 25 ml sterilized

distilled water.

A statistical analysis was carried out on data using arc sin transformation

(Snedecor & Cochran, 1971).

Source : MNHN, Paris

290 M. MOSTAFA

SYMPTOM DESCRIPTION

INFECTED

È no visually observable infection

: : of the leaf

0 2:5. га : minute to small lesions : minute

2.5 E в тт necrotic areas that may or may not!

9% = 16 : 3. ) : be accompanied with chlorosis of

- : surrounding tissues

10 - 20 47) : small to medium sized lesions

20 - 30 5 )__ : necrotic areas medium in size

30 - 40 6 ) : and surrounded by chlorotic zones

40 - 50 Fed а

: medium to large lesions : heavily

: infected leaves with large necro-

50 - 75 87). : tic areas surrounded by large

75 - 100 9.) chlorotic areas ; blighted and

: dying leaves

Table 1 - Grading and description of net-blotch infection symptoms (determined by visual obser-

vation; Barrault, 1989).

Tableau 1 - Classes et description de symptómes foliaires induits par l'helminthosporiose de

l'orge; Barrault, 1989).

RESULTS

Seed treatment

Table 2 shows that the most severe symptoms (as defined by the extent of

necrotic areas) on the coleoptile were observed on the seeds inoculated with D. teres

alone (97.8%). All the antagonists used reduced the D. teres coleoptile infection signif-

icantly (Р < 0.05). Seed applications of actinomycete/N51 and Trichoderma

virideINRG1 gave the best control: the severity of the attack by D. teres was reduced

to 9.8% and 13%, respectively.

The mycelial fragments sampled at various locations on the necrotic coleop-

tiles and cultured on V8 agar belonged to express the pathogen D. teres in every case.

Leaf treatment

Application of antagonists 1 day before inoculation of the leaves by a mycelial

suspension of D. teres significantly reduced the area of the net blotch lesions which

subsequently developed on the leaves. Table 3 shows that the coefficient of infection

(СП) in the control plants (untreated with the antagonists) was much higher than in the

plants treated with the antagonists. The most significant reduction of leaf symptoms

was obtained with Myrothecium verrucaria/N76-1 and actinomycete/N51 which re-

duced the necrotic leaf area by 95%.

Source : ММНМ Paris

BIOLOGICAL CONTROL OF DRECHSLERA TERES 291

Mean coleoptile: Mean length of

Parameter Attack 8

length (cm) : necrotic areas :

: : per coleoptile :

Treatment 5 : per tube (cm) :

lactinomycete/N51 H 5.1 Е 0.5 Н efe

Trichoderma viride/NRGl: 4.6 : 0.6 : 13.0 са

Муговһесішт verrucaria : 4.7 : 0.9 19.6 be

N76-1 : :

Trichoderma pseudo- : 4.6 : 1.1 123.9 be

koningii/N 69 : :

Trichoderma sp/BRL 124 : 4.6 : is $32.6 Б

Control with D.teres : 4.6 : 4.5 :

alone : : :97.8а

control with sterilized: 4.6 : 2 0

[distilled water alone :

Table 2 - Effect of seed treatment on reduction of coleoptile infection by D. teres.

The values (averages of 10 replicates with 5 plants/tube) which are followed by the

same letter are not significantly different at the 5% level using the test of Newman-

Keuls.

Tableau 2 - Efficacité des germes antagonistes sur la réduction du niveau d'attaque coléoptilaire

par D. teres.

Straw treatment

The ability of the antagonists to suppress conidia formation in D. teres was

tested in the laboratory using infected straw treated with antagonists. The search for

the more effective antagonists (Table 4) showed that Myrothecium verrucarialN 16-1,

Trichoderma virideINRG-1 and actinomycete/NS1 could significantly (P < 0.05) re-

duce the number of conidia as compared with the untreated control.

Approximatively 79%, 75% and 70% of the conidia of D. teres were sup-

pressed in straw after 4 days and 76%, 81% and 85% after 6 days in the presence of

Myrotheciwn verrucarialN 16-1, Trichoderma viride/NRG-1 and Actinomycete/NS1, re-

spectively.

DISCUSSION

The in vivo experiments presented above have shown that three saprophytic

microorganisms (actinomycete/NS1, Trichoderma viride/NRG-1 and Trichoderma pseu-

dokoningii/N69) are efficient at three different stages in the life cycle of D. teres.

Source : MNHN, Paris

292 M. MOSTAFA

Parameter :% Number :Leaf area:Coefficent: Disease

infected of :reduction 9]

infected: рег :infection :

per :treatment: d

(Treatment treatment

lactinomycete/N51 S s 2085 15 : 95.3 al

IMyrothecium verrucaria : 10 : 1.6 : 16 : 95.3 a

N76-1

Trichoderma viride/NRG1: 18 : 2.7 : 48.6 i 85.7 ab

Trichoderma sp/BRL 124: 20 : 2.9 58 : 82.9 ab

Trichoderma pseudo- : :

koningii/N69 uus 2 50 48.1 : 85.9 ab

control with D. teres : :

alone 8 52 571 69 340.0 ^: 0 с

Table 3 - Effect of antagonists on reduction of leaf lesion by D. teres.

Leaf areas were determined visually and scored on a 0-9 scale (Table 1)

Disease reduction - percent reduction in coefficient of infection with respect to control.

The values (means of 3 replicates with 10 plants/pot) which are followed by the same

letter are not significantly different at the 5% level using the test of Newman-Keuls.

Tableau 3 - Efficacité des germes antagonistes sur la diminution de l'importance de lésion fol-

aires de D. teres.

These antagonists markedly reduced the production of conidial inoculum on

the straw. They could also decrease significantly the intensity of the coleoptile attack

by the seed-borne inoculum. Downes (1977) had already shown that the application of

a bacterium, Erwinia herbicola, on oat seeds prevented the appearance of coleoptile

necroses induced Бу Pyrenophora avenae. More recently, Уаппасі & Pecchia (1986)

reported that Drechslera sorokiniana on barley seeds could be partially inhibited by

Trichoderma harzianum and Chaetomium globosum, whereas Al-Hasmini & Perry

(1986) observed a reduction of the coleoptile attack by Gerlachia nivalis in barley af-

ter the application of a spore suspension of Trichoderma viride.

The antagonists selected also decreased the intensity of foliar symptoms

(through a reduction of the coefficient of infection). Scharen & Bryan (1981) had

shown the efficiency of a preventive foliar treatment with a bacterial suspension (Ba-

cillus licheniformis) against Drechslera teres on barley. Slessman & Leben (1976) had

also observed that a bacterium (AN77) could reduce dramatically the foliar symptoms

caused by Helminthosporium maydis on maize under controlled conditions but not in

the field.

Source : MNHN, Paris

BIOLOGICAL CONTROL OF DRECHSLERA TERES 293

Time after treatment : Sporulation after : Sporulation after

3 4 days 6 days

Parameter : Conidia % ,: Conidia %

per 100 :reduction: рег 100 :reduction

Treatment :conidioph- :conidioph-:

ores : ores

control = ory 2 1 зага

м. verrucaria/N76-1 JEUNE № = ва toss

Trichoderma viride/NRG1: 33.1 с 75 : 64.3 d : 81.2

lactinomycete/N : 39.9 be : 70 : 49.6 а : 85.5

Trichoderma sp/BRL 124 : 39-5 be : 70 | 88.0 = 74.3

Trichoderma pseudo- : 58.5 b : 28 : 183.0 b 5 46

koningii/N69

Table 4 - Reduction of D. teres conidia formation on the straw by antagonists,

Mean of ten replicates: each replication consisted of 10 fragments of straw for which

100 conidiophores were scored.

Reduction (%): in number of conidia with respect to control.

The percentages followed by the same letter are not significantly different at the 5% lev-

el using the test of Newman-Keuls

Tableau 4 - Efficacité des germes antagonistes sur la conidiogénèse de D. teres sur les pailles.

Among the three antagonists selected, actinomycete/NS1 displayed the broad-

est spectrum of action since its efficiency in the three epidemiological sequences in-

vestigated was good in every case. An increase in efficiency might be obtained

through the association of two or three different antagonists as a result of the synergis-

tic microorganisms (Mostafa, 1982).

The antagonistic activity of these microorganisms in other sequences of the di-

sease, particularly survival, still need to be tested. The necrotrophic character of the

pathogen and the importance of the saprophytic phase require the control of the multi-

ple survival forms which generate the primary inoculum. The survival forms on straw

are mostly responsible for the development of the disease on farm plots since the aeri-

al exogenous inoculum, as a result of its dispersion over short distances only, can be

considered as epidemiologically non significant (Piening, 1968; Barrault, 1989). With-

in this context, the efficiency of the antagonists selected might be tested on the follow-

ing survival forms: і the resting mycelium (inhibition of the asexual morphogenesis,

i.e. conidiophores and/or conidia); ii the sclerotia (inhibition of the myceliogenic acti-

Source - MNHN, Paris

294 M. MOSTAFA

vity); and iii the perithecia (inhibition of the perithecium ascus and ascospores mor-

phogenesis).

As to the action of the antagonists on the teleomorph, Pfender (1988) showed

that a basidiomycete (Limonomyces roseipellis), isolated from a microbial community

on straw originating from minimum tillage plots (where the disease was declining), in-

hibited the formation of perithecia of Pyrenophora tritici repentis on wheat.

The biological control of the pathogen in all its survival forms should be part

of an integrated control program aimed at reducing the inoculum pressure on the plot.

REFERENCES

AL-HASHIMI M.H. and PERRY D.A., 1986 - Fungal antagonists of Gerlachia nivalis. J. Phyto-

pathol. 116 (2): 106-112.

BAKER K.F. and COOK R.J., 1974 - Biological Control of Plant Pathogens. San Francisco,

W.H. Freer man & Co.

BARRAULT G., 1989 - L'Helminthosporiose de l'orge causée par Drechslera teres. Thèse Doc-

torat d'Etat. Institut National Polytechnique de Toulouse, France.

CAMPBELL R., 1990 - Current status of biological control of soil-borne diseases. Soil Use and

Management 6 (4): 173-178.

COOK R.J. and BAKER K.R., 1983 - The Nature and Practice of Biological Control of Plant Pa-

thogens. St Paul, The American Phytopathological Society.

DOWNES M.J., 1977 - Erwinia herbicola for control of Pyrenophora avenae on oat seedlings

and of other organisms. Royal Irish Academy; Irish National Committee for Biology:

Proceedings of Seminar on Biological Control (February 17 and 18, 1977 Dublin):

97-107.

LOEGERING W.D., 1959 - Method for recording cereals rust data. Washington, International

Spring Wheat Rust Nursery, 1959, USDA.

MARTIN R.A., 1985 - Disease progression and yield loss in barley associated with net blotch as

influenced by fungicide seed treatment. Can. J. Plant, Pathol. 7: 83-90.

MOSTAFA М., 1982 - Recherche sur la Lutte biologique contre l'Helminthosporium teres, para-

site de l'orge, par l'utilisation de microorganismes antagonistes et d'une souche hypoag-

ressive. Thèse Docteur-Ingénieur, n° 220, INP Toulouse.

MOSTAFA M., BARRAULT G. et ALBERTINI L., 1992 - Lutte biologique contre Drechslera

teres: Action in vitro de microorganismes antagonistes sur la croissance mycélienne et la

germination. Cryptog., Mycol. 13 (2): 125-133

MUKERII К.С. and GARG K.L., 1988 - Biocontrol of Plant Diseases (Volume 1). Florida, CRC

Press Inc, Boca Raton.

PFENDER W.F., 1988 - Suppression of азсосагр formation in Pyrenophora tritici repentis by Li-

monomyces roseipellis, a basidiomycete from reduced tillage wheat straw. Phytopatholo-

gy 78: 1254-1258.

PIENING L.J., 1968 - Development of barley net blotch from infected straw and seed. Can. J. Pl.

Sci. 4: 623-625.

PURKAYASTHA R.P., 1989 - Modern approaches to biocontrol of crop pathogens and their im-

plications. Indian J. Mycol. Res., 27 (1): 1-6.

SCHAREN A.L. and BRYAN M.D., 1981 - A possible biological control agent for net blotch of

barley. Phytopathology 71 (8): 902-903.

SHIPTON W.A., KHAN J.N. and BOYD W.J., 1973 - Net blotch of barley. Rev. Plant. Pathol.

52 (5): 269-290.

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SLESSMAN J.P. and LEBEN C., 1976 - Microbial antagonists of Bipolaris maydis. Phytopathol-

ogy 66: 1124-1218.

SMEDEGARD-PETERSEN V., 1974 - Reduction in yield and grain size of barley due to attack

by net blotch fungus Pyrenophora teres. Kgl. Vet. Landbohoe jsk Ars-skr 1974:

108-117,

SMITH 1.М., DUNEZ J., LELLIOT R.A., PHILLIPS D.H. and ARCHE S.A., 1988 - European

Handbook of Plant Diseases. London, Blackwell Scientific Publications: 377-383

SNEDECOR, G.W. et COCHRAN, W.G., 1971 - Méthodes Statistiques. Paris. Association de

Coordination Technique Agricole.

SPURR H.W., 1981. - Experiments of foliar disease control using bacterial antagonists. Pages

369-381. In: Microbial Ecology of the Phylloplane. New York, J.P. Blakeman ed., Ac-

ademic Press.

SUTTON J.C. and STEELE P., 1983 - Effects of seed and foliar fungicides on progress of net

blotch and yield in barley. Can. J. Pl. Sci. 63: 631-640.

ТАНУОМЕМ В. and AVIKAINEN H., 1990 - Effect of Streptomyces sp. on seed-borne foot rot

diseases of wheat and barley. I. Pot experiments. Ann. Agric. Fenniae 29: 187-197.

VANNACI С. and PECCHIA S., 1986 - Evaluation of biological seed treatment for controlling

seed-borne inoculum of Drechslera sorokiniana on barley. Med. Fac. Landbouww Ri-

jksuniv. Gent. 51 (2b): 741-780.

Source : MNHN, Paris

ea tee teh М іы

pies знак? ваты:

DA Fh vlone. аа,

Cryptogamie, Mycol. 1993, 14 (4): 297-306 297

THE INCIDENCE OF FUNGI IN HUMAN AXILLARY HAIR

AND THEIR TOXIGENIC POTENTIALITIES

Н.А.Н. HASAN, М.М.К. BAGY and А.У. ABDEL-MALLEK

Botany Department, Faculty of Science, Assiut University, Assiut,

Egypt.

ABSTRACT - The present study determined the incidence of fungi in 66 specimens of human

axillary hair. Eleven genera, 22 species and 2 varieties were isolated. Penicillium funiculosum

(32% of hair specimens) was the only fungus isolated in moderate incidence. Aspergillus fumiga-

tus (21%), A. flavus (17%), A. niger (17%) and Chrysporium tropicum (12%) showed low inci-

dence. However, Trichosporon catenulatum, Trichophyton mentagrophytes, Candida albicans,

Chrysosporium keratinophilum and Geotrichum candidum were rare. Twenty-eight isolates (out of

57 tested isolates) belonging to the most common fungal species (A. flavus, A. fumigatus and P.

funiculosum) recovered from axillary hair produced respective mycotoxins. The detected toxins

were: aflatoxins Bj, В», Gi, б», gliotoxin and ochratoxins А, B. The selective effects of Вас

(sweet deodorant) on the mycelial dry weight, protease activity and mycotoxins production of

some fungal species were also examined. At 1% concentration Bac inhibited the mycelial dry

weight of all tested fungi and protease activity of A. flavus and A. fumigatus. However, it pro-

moted protease activity of A. niger and Р. funiculosum. The specific production of aflatoxins by

A. flavus and ochratoxins by P. funiculosum showed their accumulation with Bac treatment.

RESUME - 22 espèces et 2 variétés de champignons, appartnant 4 11 genres, ont été isolées de

poils axillaires humains. Penicillium funiculosum (32% des échantillons de poils) est la seule

espèce présentant une fréquence notable d'isolement. Aspergillus fumigatus (21%), A. flavus

(17%), A. niger (17%) et Chrysosporium tropicum (12%) présentent des fréquences d'isolement

faibles. Trichosporon catenulatum, Trichophyton mentagrophytes, Candida albicans,

Chrysosporium keratinophilum et Geotrichum candidum sont rarement isolés. 38 isolements (sur

57 isolés) appartenant aux espèces les plus fréquentes (A. flavus, A. fumigatus, P. funiculosum)

produisent des toxines. Les toxines isolées sont: les aflatoxines B1, B2, ОТ et G2, la gliotoxine et

les ochratoxines A et B. L'action du "Bac" (déodorant) sur la croissance mycélienne, l'activité

protéasique et la production de mycotoxines a été étudiée pour un certain nombre d'espèces. A

une concentration de 1% le "Вас" inhibe la croissance mycélienne (poids secs) de toutes les

espèces étudiées et l'activité protéasique d'Aspergillus flavus et d'Aspergillus fumigatus. Au

contraire, А cette méme concentration, l'activité protéasique d'Aspergillus niger et de Penicillium

funiculosum est stimulée. La concentration mycélienne en aflatoxine et en ochratoxine respec-

tivement chez Aspergillus flavus et Penicillium funicusolum est augmentée par le traitement au

KEY WORDS : fungi, human axillary hair, mycotoxins, protease activity, sweat deodorant.

INTRODUCTION

The isolation of dermatophytes and other non dermatophytes from hair has

been investigated by several authors (English, 1976; Takotori & Ichijo, 1979; Lopez

Source : MNHN, Paris

298 НАН. HASAN et al.

Martinez & Rivera Lona, 1984 and Imwidthaya & Thianprasite, 1988). Dermatophytes

utilize protein as the main nutrient source and are capable of decomposing even a

resistant scleroprotein, such as keratin. During their growth on protein substrates they

secrete strongly active proteolytic enzymes. Mycotoxins are toxic substances produced

by fungi which cause diseases in animals or man. Acute diseases caused by

mycotoxins are called mycotoxicoses. No available literatures concerning with the

ability of fungi isolated from human axillary hair for producing mycotoxins. Therefore,

the present investigation deals with the incidence of fungi in human axillary hair and

their toxigenic potentialities to establish whether a potential hazard might exist due to

contamination of hair with toxigenic molds. Also, the selective effects of one of sweat

deodorants on the mycelial growth, protease activity and mycotoxin production by

some fungal species were examined.

MATERIALS AND METHODS

Isolation and identification of fungi:

Sixty-six specimens of human axillary hair were examined. These specimens

were randomly chosen from healthy males in Assiut city. The specimens were placed

in sterilized Petri-dishes and sent immediately to the laboratory. A portion of

specimens were examined in 10% KOH for the presence of fungal hyphae and

arthrospores. The specimens were also cultured on Sabouraud's dextrose agar

containing 0.05 mg/ml of chloramphenicol and 0.5 mg/ml cycloheximide.The plates

were incubated at 28°C (+ 2) and the fungal growth was observed once weekly for a

period of 4 weeks. The developing colonies were examined microscopically and

identified.

Toxigenic potentialities of common fungi:

Fifty-seven isolates belonging to Aspergilus flavus, A. fumigatus, A. niger and

Penicillium funiculosum which represent the most common fungal species associated

with hair were subjected to mycotoxin screening during this investigation. Twenty ml

of YES (yeast extract sucrose) medium in 100 ml Erlenmeyer flask were sterilized,

inoculated with 1 ml spore suspension of 1 week-old culture of each tested isolate and

incubated at 25°C (+ 2) for 7 days as stationary cultivation. Extraction and analysis of

the fungal toxins are carried out as mentioned below.

Antimycotic activity of sweat deodorant:

One type of sweat deodorant (Bac) was tested for its antimycotic activity.

Forty ml of sterilized Sabouraud's dextrose medium were poured into 250 ml

Erlenmeyer flask. Bac (manufactured by UTAC, Egypt under licence from Hans

Schwarzkopf GmbH Hamburg, Germany) was incorporated at concentrations 0.5, 1.0

and 5.0%. Two flasks were used for each concentration and control (free from Bac).

Then, the flasks were inoculated with 1 ml spore suspension (approx. 10° spores) and

incubated as described previously.

Estimation of protease activity:

Proteolytic enzyme activity in media was measured using casein powder as

substrate. The reaction mixture containing 2 ml of 1% (w/v) casein in 0.1 M sodium

phosphate buffer pH 7.6 was incubated with Iml of culture filtrate at 30°C (+ 2) for 2

h. The reaction was stopped and unhydrolyzed protein was removed by addition of

Source : MNHN, Paris

FUNGI IN HUMAN AXILLARY HAIR 299

1ml of 50% trichloroacetic acid. The precipitated undigested protein was removed by

centrifugation. The hydrolyzed protein in supernatant was determined by the method

of Lowry et al. (1951).

Extraction and analysis of the fungal toxins:

The content of each flask (mycelium + filtrate) was homogenized for 5 min in

high speed blender with 50 ml chloroform. Then the extract was evaporated till

dryness on a rotary evaporator. The chloroform extracts were analysed for the presence

of mycotoxins on Silica Gel-coated plates. The methods of Nesheim (1976), Moss &

Badii (1982) and Richard et al. (1989) were used for analyses of of ochratoxin,

aflatoxin and gliotoxin, respectively. Quantitative estimation of these toxins was made

according to Nabney & Nesbitt (1965) and Applegate & Chipley (1976).

RESULTS AND DISCUSSION

A total of 11 genera, 22 fungal species and 2 varieties were isolated from

healthy human axillary hair collected from 66 males in Assiut city Penicillium

funiculosum was moderately isolated, being present in 32% of the specimens. Four

fungal species showd low incidence viz: Aspergillus fumigatus, A. flavus, A. niger and

Chrysosporium tropicum. However, the remaining fungal species were rare in human

axillary hair (Table 1).

Dermatophytes were represented by five species belonging to Chrysosporium

(2 species), Trichosporon, Trichophyton and Candida (1 species for each). According

to the percentage inncidence (in relation to the total specimens) these species could be

arranged in the follwing order: C. tropicum (12%) > Trichosporon catenulatum (6) >

Trichophyton mentagrophytes (4.5) > C. keratinophilum and Candida albicans (1.5 for

each). Moharram et al. (1988) noticed that C. tropicum, C. keratinophilum, C. lobatum

and C. queenslandicum were associated with healthy human hair. In Iran, Moghadami

& Emami (1986) reported that dermatophytes isolated from tinea capitis included

Microsporum canis, Trichophyton violaceum, T. verrucosum, T. schoenleinii and 1

isolate of each of T. mentagrophytes, Candida albicans and Candida sp.

Seventeen species and 2 varieties belonging to 7 genera of fungi other than

dermatophytes were associated with axillary hair. Penicillium funiculosum, Aspergillus

fumigatus, A. flavus and A. niger were the dominant molds recovered (32, 21, 17 and

17% of the specimens, respectively).

Yeast (red colour), P. chrysogenum, A. flavus var. columnaris, Alternaria

alternata, Cladosporium cladosporioides, Emericella nidulans var. dentata and Mucor

racemosus were associated with 3-11% of the examined hair. El-Shanawani (1993)

reported that several saprophytic fungi were recovered from tinea capitis in Assiut and

New Valley Governorates; the most common species were A flavus, A. niger, A.

fumigatus, P. chrysogenum, Alternaria alternata and Cladosporium cladosporioides.

Also, El-Gendy (1988) and Mahmoud (1991) isolated A. flavus, A. niger, A. sydowii, P.

chrysogenum, Alt. alternata and C. cladosporioides from cases of tinea capitis. English

(1965) observed that A. fumigatus and A. terreus were able to grow on human hair.

Whereas, Botticher (1966) recorded that Alternaria alternata has been implicated as a

cause of dermitis in human. In this study, each of A. carneus, A. terreus, A. ustus,

Cladosporium carrionii, C. herbarum, Geotrichum candidum, P. canescens, P.

cyclopium and P. purpurogenum are encountered in 1.5% of human axillary hair (Ta-

ble 1). In this respect, Aho (1983) suggested that the presence of saprophytic fungi on

Source : MNHN, Paris

300 H.A.H. HASAN et al.

г нет oR

Genera and species

Alternaria alternata (Fries) Keissler 2

Aspergillus carneus (V.Tiegh. )Blochwitz 1

A. flavus Link n

A. flavus var. columnaris Raper & Fennell 3

A. fumigatus Fresenius 4

A. niger Van Tieghem 1

A. terreus Thom 1

EIS

A. ustus (Bain.) Thom & Church

Candida albicans (Robin) Berkhout

Chrysosporium keratinophilum (Frey) Carmichael

С. tropicum Carmichael

Cladosporium carrionii Trejos

cladosporioides (Fres.) de Vries

С. herbarum (Pers.) Link ex Fr 1

Emericella nidulans var. dentata Sandhu & Sandhu 2

Geotrichum candidum Link 1

Mucor racemosus Fresenius 2

Penicillium canescens Sopp. 1

Р. chrysogenum Thom $

CE Spee ы

Бапайоойоте

Р. cyclopium Westling

P. funiculosum Thom 2

Р. purpurogenum Stoll

Trichophyton mentagrophytes (Robin) Blanchard

Trichosphoron catenulatum (De Berum,Gougerot &

Vauchel) Ota

Yeast (red colour) $

ә-езешелешеше

DINED DDO RL]

Suuououououou

Total number of genera 11

Total number of species and varieties 2242

NCI = Number of cases of isolation (out of 66 specimens); OR-Occurrence

remarks; oderate occurrence (between 17-32 cases); L-Low occurrence

(between 8-16 cases); R-Rare occurrence (between 1-7 cases).

Table 1: Incidence of fungi in human axillary hair on Sabouraud's dextrose agar at 28°C.

Tableau 1: fréquence des espéces fongiques isolées de poils axillaires humains sur milieu de

Sabouraud à 28*C.

hair and skin creates an opportunity for them under special circumstances to become

invasive to the skin or hair and thus cause primary or secundary infection.

The results in table (2) show the toxigenic potentialities of fifty-seven isolates

belonging to the most common species. 61% of Aspergillus members tested produced

at least one toxin. All tested isolates of A. flavus produced aflatoxins from which two

isolates produced aflatoxins В), В», б; and б». The remaining 9 isolates produced B,

and В, only. These results are in accordance with the finding of Joffe (1969) who

reported that 8.4% of A. flavus members produced all four aflatoxins. However,

Youssef (1986) reported that 29 isolates of A. flavus (out of 30 tested isolates)

produced all four aflatoxins, and one isolate formed aflatoxins B, and В, only.

Aflatoxins are mutagenic, carcinogenic, teratogenic and acutely toxic to most

experimental and domesticated animals and man (El-Zawahri et al., 1977; Davis &

Diener, 1978).

Source : MNHN, Paris

FUNGI IN HUMAN AXILLARY HAIR 301

No.of positive No.of toxin Mycotoxin

Species

$ specimens producers produced

A flavus il 9

A. fumigatus 14 il Gliotoxin

A. niger u o -ve

P. funiculosum zi 6 Ochratoxin A ,b

Total isolates 57 28

Table 2: Mycotoxin production by common fungal species associated with axillary hair.

Tableau 2: Production de mycotoxine par les espèces fongiques associées aux poils axillaires hu-

mains.

Production of gliotoxin by 11 isolated of A. fumigatus out of fourteen tested

isolates agree with the finding of Moss (1977) and Richard et al. (1989). A. fumigatus

represents the predominant pathogen of aspergillosis in man. Eichner & Mullbacher

(1984) have hypothesized that gliotoxin may be produced during the pathogenic state

of A. fumigatus and contributes to the pathogenicity of the fungus. Six isolates (out of

21) of P. funiculosum produced ochratoxins A and B. Carlton & Krogh (1979)

classified P. funiculosum as ochratoxins producer. Ochratoxin A causes nephropathy in

pigs (Elling et al. 1985) and induces renal and hepatic carcinomas in mice (Kanisawa

& Suzuki, 1978; Bendele et al. 1983). Data obtained in this investigation, proved that

most of fungi tested produced seven mycotoxins. These data strengthen our initial

concern that potential hazard to human health may exist due to the presence of

toxigenic fungi on hair.

The selective effect of sweat deodorant (Bac) on mycelial growth and protease

activity of 5 isolates of fungal species associated with hair was illustrated in table (3).

The mycelial growth of three species species of Aspergillus namely: A. flavus, A.

fumigatus and A. niger were inhibited at 1% concentration whereas the mycelial dry

weight of Penicillium funiculosum and Chrysosporium tropicum were more sensitive at

0.5% concentration. The main constituents of Bac are water, SD alcohol 39-C, alumi-

nium chlorhydrate, ceteareth-11, Fragrance and hydroxy ethylcellulose. According to

Megalla et al. (1980), the essential oils as aliphatic alcohols exhibited a high

antifungal activity. Moharram et al. (1988) used creams, vaseline, shampoos and oils

which applied to human hair for treatment of dry skin and dandruff, as antimycotic

agent. They found that silk hair with lanolin cream (yellow). Relax bath (blue) and

Silk hair (herb-green) were highly effective against all tested fungi. Also,

Chrysosporium isolates were the most sensitive fungi to shampoos.

The ability of five fungal species to produce extracellular protease in broth

cultures was found to be greatly affected by the addition of sweat deodorant (Bac) as

shown in table (3). The response seemed to be fluctuated between inhibition and pro-

motion. Protease synthesis by A. flavus and A. fumigatus was greatly reduced at

Source : MNHN. Paris

302 H.A.H. HASAN et al.

i Prot

— еа Conc. Mycelial 222 rotease

% dry wt activity

(mg/40 ml) (ug/ h/ml)

A. flavus о 282 Ба 766.3

0.5 264 + 630.0

1.0 181 ^ 462.5

5.0 no growth

A.fumigatus о 235 + 283.8

0.5 223 + 230.0

1.0 198 a 202.5

5.0 no growth

A. niger о 394 + 126.3

0.5 349 + 200.0

1.0 125 = 165

5.0 no growth

Chrysosporium 0 249 + 223.8

tropicum 0.5 122 + 2325

no growth

P. funiculosum 0 310 153.8

о. 196 + 205.0

поб 162 з 212.5

5. no growth

Table 3: Effect of sweat deodorant (Bac) on mycelial growth, sporulation and protease activity of

five species.

Tableau 3: Effets d'un déodorant (Bac) sur la croissance mycélienne, la sporulation et l'activité

protéasique de 5 espèces fongiques.

concentration 1%. In this respect, Mahmoud (1991) found that citral, citronellol, nero!

and caproic acid reduced protease activity in Fusarium compactum and Trichophyton

violaceum.

Concentration of 0.5% of Bac has promotive effect on protease activity in A.

niger, P. funiculosum and C. tropicum. Carvone and Thymol were found to be

accelerate protease production by F. compactum and T. violaceum (Mahmoud, 1991).

Davidson & Branen (1980) found that the antioxidant such as butylated hydroxy

anisole and butylated hydroxy toluene caused leakage of intracellular protein from

Pseudomonas fluorescens. Cumming (1990) concluded that aluminium has inhibition

of nitrate reductase activity in roots of pitch pine (Pinus rigida) seedlings.

Source : ММНМ Paris

303

FUNGI IN HUMAN AXILLARY HAIR

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Source : MNHN, Paris

304 H.A.H. HASAN et al.

The mechanism of fungal inhibition might be due to the Bac constituents act

on the cytoplasmic membrane and might also be related to the destruction or

inactivation of essential enzymes and/or genetic material. These reasons are agreed

with the conclusion of Branan et al. (1980) on the mechanism of antioxidants on

microbial inhibition.

The selective effect of sweat deodorant (Bac) on the secondary metabolites

(aflatoxins and ochratoxins) of two toxigenic molds isolated from human axillary hair

(A. flavus and P. funiculosum) was illustrated in table (4). The inhibitory action of Bac

on mycelial growth of A. flavus has no effect on aflatoxin production. However, the

low specific production shows highly accumulation of toxins with treatments. In this

respect, Hasan & Mahmoud (1993) reported that the specific production of aflatoxins

increased at concentration 250 ppm of cumin and 500 ppm of clove oils.

The results of action of Bac on ochratoxins production by P. funiculosum (Ta-

ble 4) show that the active constituents of Bac maintained the ochratoxin B inhibitory

properties possibly by interfering with its biosynthesis pathway. On the other hand,

ochratoxin A was accumulated at 0.5 and 1% concentrations. Bac may be incorporated

into precursors of ochratoxins instead of the correct component in the biosynthetic

pathway and thus inhibit toxin B formation.

Our results indicate that, Bac has the unique capability of selectively altering

ochratoxin synthesis in P. funiculosum strain which produced both A and B type

toxins. Large increases in the amounts of ochratoxin A with corresponding decreases

in B toxin may be due to specific inhibition of the conversion of A to B by Bac.

REFERENCES

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suspected dermatophytosis. Mycopathologia, 83: 65-73.

APPLEGATE K.L. and CHIPLEY J.R., 1976 - Production of ochratoxin A by Aspergillus

ochraceus NRRL. 3174 before and after exposures to «Со Irradiation. Appl. Environm.

Microbiol. 31: 349-353

BENDELE S.A., CARLTON W.W., KROGH P. and LILLEHOJ E.B., 1983 - Ochratoxin A

carcinogenesis in mouse. Abstr. 3rd Mycological Congress, Tokyo.

BOTTICHER W.W., 1966 - Alternaria as a possible human pathogen. Sabouraudia 4: 256-285.

BRANAN AL., DAVIDSON Р.М. and KATZ B., 1980 - Antimicrobial properties of phenolic

antioxidants and lipids. Food Technol. 34: 42.

CARLTON W.W. and KROGH P. , 1979 - Ochratoxins, a review. Conference on mycotoxins in

animal feeds and grains related to animal health. Bureau of Vetenary Medicine. Food

and Drug Administration, Rockville Maryland U.S.A. June 8

CUMMING J.R., 1990 - Nitrogen source effects on aluminium toxicity in non-mycorrhizal and

mycorrhizal pitch pine (Pinus rigida) seedlings: И. Nitrate reduction and nitrate uptake

Canad. 1. Bot. 68: 2653-2659.

DAVIDSON Р.М. and BRANEN A.L., 1980 - Antimicrobial mechanisms of butylated hydroxy

anisole against two Pseudomonas species. J. Food Sci. 45: 1607.

DAVIS К.Р. and DIENER V.L., 1978 - Mycotoxins. p. 397-470. In: Beuchat L.R. (ed) Food

and Beverage Mycology. Westport, Connecticut. AVI Publishing Co

EICHNER R.D. and MULLBACHER A., 1984 - Hypothesis: Fungal toxins are involved in

aspergillosis and AIDS. Austral. J. Exp. Biol. Med. Sci. 62: 479-484.

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FUNGI IN HUMAN AXILLARY HAIR 305

EL-GENDY Z.K.A., 1988 - Studies in dermatophytes in Minia Governorate, М. Sc. Thesis. Bot.

Dept., Fac. Sci., Minia Univ., Egypt

ELLING F., NIELSEN J.P., LILLEHOJ E.B., THOMASSEN M.S. and STORMER F.C., 1985 -

Ochratoxin A induced porcine nephopathy: enzyme and ultrastructure changes after short

term exposure. Toxicon 23: 247-254.

EL-SHANAWANY A.A., 1993 - Human dermatophytes in Assiut and New Valley Governorates

Ph. D. Thesis, Bot. Dept., Fac. Sci. Assiut Univ., Egypt.

EL-ZAWAHRI M., MOUBASHER A.H., MORAD M. and EL-KADY LA., 1977 - Mutagenic

effect of aflatoxin Ву. Ann. Nutr. Aliment. 13: 859-866.

ENGLISH M.P., 1965 - The saprophytic growth of non-keratinophilic fungi on keratinized

substrate and a comparison with keratinophylic fungi. Trans. Br. Mycol. Soc. 48:

219-235.

ENGLISH МР. 1976 - Destruction of hair by two species of Chrysosporium. Trans. Br. Mycol.

Soc. 66 (2): 357-358.

HASAN Н.А. and MAHMOUD A.LE., 1993 - Inhibitory effect of spice oils on lipase and

mycotoxin production. Zentralbl. Mikrobiol. 148: In Press.

IMWIDTHAYA 5. and THIANPRASITE М., 1988 - A study of dermatophytes in Bangkok

(Thailand). Mycopathologia 102: 13-16.

JOFFE A.Z., 1969 - Nature, 221: 492. In: Detroy, R.W., Lillehoj ЕВ. and Ciegler A., Aflatoxin

and related compounds, р. 3-178. In: Ciegler A., Kadis 5. and АЛ S.J. (eds) Microbial

Toxins- Vol. 7- Fungal Toxins, (1971). Academic press. N.Y. and London

KANISAWA M. and SUZUKI S., 1978 - Introduction of renal and hepatic tumors in mice by

Ochratoxin A, a mycotoxin-Gann, 69: 599-600.

LOPEZ MARTINEZ В. and RIVERA LONA M., 1984 - Investigation of dermatophytes from

healthy skin from different parts of the human body. Revista Latinoamer. Microbiol.

26(4): 365-369

LOWRY OH, ROSEBROUGH МЈ. FARR AL. and RANDALL RJ, 1951 - Protein

measurement with the Folin phenol reagent. J. Biol. Chem. 193: 256-275

MAHMOUD А1.Е., 1991 - Some physiological and biochemical studies of fungi isolated from

skin of mammals. Ph. D. Thesis, Bot. Dept., Fac. Sci., Assiut Univ., Egypt.

MEGALLA S.E., EL-KELTAWI N.E.M. and ROSS S.A., 1980 - A study of antimicrobial action

of some essential oil constituents. Herba polon. 3: 181-185.

MOGHADAMI M. and EMAMI M., 1986 - Mycotic infection of tinea capitis in Teheran.

Iranian J. of Public Health 15 (1-1): 55-63.

MOHARRAM M., ABDEL-GAWAD K.M. and EL-MARAGHY 5.5.М., 1988 - Ecological and

physiological studies on fungi associated with human hair. Folia Microbiol. 33:

363-371.

MOSS M.O., 1977 - Aspergillus Mycotoxins. р. 449-524. In: Smith ТЕ. and Pateman ЈА

(Eds), Genetics and physiology of Aspergillus. London, New York, San Francisco,

Academic Press, Inc.

MOSS М.О. and BADII Е, 1982 - Increased production of aflatoxins by Aspergillus parasiticus

Speare in the presence of rubratoxin В. Appl. Environm. Microbiol. 43: 895-898.

NABNEY J. and NESBITT В.Е, 1965 - A spectrophotometric method for determining the

aflatoxins. Analyst 90: 155-160.

NESHEIM S., 1976 - The ochratoxins and other related compounds. In: Rodricks J.V. (Ed.):

Mycotoxins Adv. Chem. Ser., 149:276-295. Washington, D. С. Am. Chem. Soc.

RICHARD LL, LYON RL, FICHTNER RE. and ROSS Р.Е, 1989 - Use of thin layer

chromatography for detection and high performance liquid chromatography for

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306 H.A.H. HASAN et al.

quantitating gliotoxin from rice cultures of Aspergillus fumigatus Fresenius.

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TAKATORI К. and ICHIJO S., 1979 - Observation on human and animal hairs invaded by

Trichophyton equinum in vitro. Jap. J. Vet. Sci. 41: 655-663.

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Source : MNHN, Paris

Cryptogamie, Mycol. 1993, 14 (4): 307-310 307

STUDI SUL GENERE AGARICUS-I

AGARICUS BISPORATUS SPEC. NOV.

Marco CONTU

via A. Manzoni, 33.09128 Cagliari Italia.

ABSTRACT - A new species of Agaricus, viz. A. bisporatus spec. nov., is described from Sardi-

nia. The new species belongs to the subgen. Agaricus sect. Sanguinolenti and has been collected

under Eucalyptus and Dicksonia.

RESUME - Une nouvelle espèce du genre Agaricus, A. bisporatus, est décrite de la Sardaigne

méridionale. La nouvelle espèce, appartenant au sous-genre Agaricus sect. Sanginolenti, a été

observée sous Eucalyptus et Dicksonia.

MOTS CLÉS : Basidiomycotina, Agaricales, Agaricus, A. bisporatus, Sardinia

INTRODUZIONE

Da lungo tempo, e' divenuta evidente l'esigenza di condurre studi approfonditi

sull'ecologia e la diffusione delle specie del genere Agaricus in Sardegna. L'isola,

infatti, sembra molto ricca di tali entita', molte delle quali poco conosciute о

addirittura mancanti di una descrizione ufficiale. Particolare attenzione destano, in

particolare, le specie crescenti negli impianti artificiali frangivento ad Eucalyptus divv.

spp., nei quali Agaricus risulta essere di gran lunga il genere piu' diffuso. In questa

comunicazione viene descritta una entita’ ріш volte ritrovata sotto Eucalyptus

camaldulensis e, in una occasione sotto Dicksonia antarctica, caratterizzata in modo

assai netto soprattutto dal punto di vista micromorfologico. Tale entita va ritenuta

nuova per la scienza.

Agaricus bisporatus Contu, spec. nov.

Pileus 3-7.5 cm latus, carnosus, semiglobatus dein explanatus, haud

umbonatus, ad medium levis et brunneus vel pallide ochraceus, versus marginem

pallidior et radialiter fibrillosus vel subsquamulosus, vestigia veli saepe obtecto.

Lamellae tenues, latae, adnexae, confertae, roseae dein obscure brunneae, acie

pallidiore. Stipes 3-5.5 x 0.6.-1.6 cm, sat curtus, cylindraceus, haud bulbosus, saepe ad

basim radiculatis albidis praedito, albus vel cinereus, sericeus, levis. Annulus tenuis,

superus, albidus. Caro sat conspicua, alba, fracta leviter rubescens dein isabellina;

odor saporque debilis, gratis. Sporarum pulvis obscure brunnea. Sporae 6.7-7.5 x 4.5-6

um, subglobosae vel late ellipsoideae, mono vel pluriguttulatae, apiculatae. Basidia

12-22.5 x 7-8.5 um, mono vel bispora, clavata, Subhymenium cellularis. Cheilocystidia

15-25 x 7.5-I2 um, clavata, interdum sublageniformia vel appendiculata, hyalina vel

Source : MNHN, Paris

308 M. CONTU

brunnea. Pilei cutis ex hyphis cylindraceis intertextis, 3-7.5 шт latis, constituta,

pigmento intracellulari. Fibulae absentes.

Hab. - ad terram, sub Eucalypto camaldulense, nec non Dicksonia antarctica.

Autumno. Typus: M. Contu 92/66, Sardinia, prov. Cagliari, Serramanna, 23.x.1992, leg.

M. Contu & P. Dessi (CAG!).

Cappello 3-7.5 cm, semigloboso poi espanso, non umbonato, margine sovente

con evidenti resti di velo, secco; cuticola secca, al centro unita e bruna od ocracea

pallida, rotta, verso la periferia, in fibrille radiali concolori, piu' raramente in squamule

poco rilevate, nel complesso, comunque, la superficie pileica e' sempre liscia, senza

squame adnate. Lamelle piuttosto fitte, poco ventricose, larghe, annesse, rosa cariche

poi bruno-bistro, taglio bianco. Gambo 3-5.5 x 0.8-1.6 cm, proporzionato о corto

rispetto al diametro pileico, cilindrico о clavato, mai bulboso, alla base spesso con

radichette miceliari bianche; rivestimento liscio, sericeo, da bianco a cinerognolo.

Anello sottile, poco ampio, supero, cascante a gonnella, bianco, bordo brunastro. Carne

soda, bianca, al taglio legermente arrossante, con colorazione tendente al carota poi

isabella-madera nel gambo; odore e sapore leggeri, gradevoli. Sporata bruno-bistro.

Spore 6.7-7.5 x 4.5-6 um, brune cupe, da subglobose a largamente ellissoidi,

rare quelle sferiche, lisce, mono o pluriguttulate, lisce, con apicolo poco evidente.

Ваза! 12-22.5 x 7-8.5 um, mono o bisporici, clavati, а sterigmi robusti. Subimenio

cellulare abbondante. Cheilocistidi 15-25 x 7.5-12 шт, abbondanti, sovente in cespi,

clavati, piu' raramente ventricosi, sublageniformi o appendicolati, da ialini a brunastri,

non catenulati, talora settati. Rivestimento pileico formato da un intreccio di ife

cilindriche rastremate ai setti, larghe 3-7.5 um, pigmento intracellulare. Fibbie assenti.

Habitat: a gruppi sotto Eucalyptus camaldulensis, piu' raramente presso

Dicksonia antarctica. Autunno. Non raro in questo tipo di habitat.

Materiale studiato: Sardegna meridionale, prov. Cagliari, Serramanna,

23.x.1992, leg. М. Contu & Р. Dessi (typus, CAG) - ibidem, 15.x.1992, leg. М. Contu

= prov. Cagliari, Cagliari-citta’, Orto Botanico Comunale, in un vaso con Dicksonia

antarctica, 30.xi.1992, leg. M. Contu. Diverse altre collezioni dalla stazione typus

0006

у 000

3 4

Agaricus bisporatus, spec. nov. - 1) carposporo; 2) cheilocistidi; 3) spore; 4) basidi (typus!)

x 1000.

Source : MNHN, Paris

AGARICUS BISPORATUS SPEC. NOV. 309

DISCUSSIONE

Le raccolte piu' tipiche provengono dagli impianti artificiali ad Eucalyptus

camaldulensis е presentano il rivestimento pileico radialmente fibrilloso. La collezione

dell'Orto Botanico di Cagliari differiva per l'assenza di fibrille radiali e per la tendenza

di alcuni cheilocistidi ad assumere un profilo ventricoso o sublageniforme, piu'

raremente appendicolato: poiche' i caratteri micromorfologici, in particolare le spore

subglobose ed i basidi bisporici, erano molto simili ho ritenuto di doverla considerare

solo un fenotipo. A. bisporatus appartiene, a causa della carne arrossante e dell'anello

semplice e supero, al sottogenere Agaricus sezione Sanguinolenti (Moell. & J. Schaef.)

Sing. ed e' simile soprattutto ad A. fuscofibrillosus (Moel.) Pilat, dal quale differisce

per la carne molto meno arrossante, le spore piu' larghe, i basidi costantemente

bispirici ed i cleilocistidi piu' stretti. A. silvaticus Schaeff. differisce per i basidi

tetrasporici, la carne con arrossamento piu' netto е vivo ed il gambo bulboso. Non e'

stato possibile rinvenire altre entita' similari nella letteratura europea (Bon, 1985;

Cappelli, 1984), пе in quella esotica (cfr. per tutti Heinemann, 1977). Се da

osservare, conclusivamente, che la specie e'stata osservata, almeno fino ad ora, presso

essenze, come Eucalyptus e Dicksonia, tipicamente australi: cio' induce a ritenere che

non possa del tutto essere scartata l'ipotesi che A. bisporatus possa essere rinvenuto

anche nel continente australe, sempre che non si tratti proprio di un'entita' originaria

dello stesso e presente in Sardegna unicamente perche' introdotta con dette essenze.

LETTERATURA

BON М. 1985 - Clé monographique du genre Agaricus L.: Fr. (sous-genre Agaricus). Doc.

Mycol, 60: 1-35.

CAPPELLI A., 1984 - Fungi Europaei. І. Agaricus. Saronno.

HEINEMANN P., 1977 - Essai d'une clé de détermination des genres Agaricus et Micropsalliota.

Sydowia 3

Source : MNHN, Paris

Source : MNHN. Paris

Cryptogamie, Mycol. 1993, 14 (4): 311-313 311

ANALYSES BIBLIOGRAPHIQUES

G.C. CARROLL and D.T. WICKLOW (eds.), 1992 - The fungal community. Its

Organization and Role in the Ecosystem. Second edition. 976 pages. Marcel

Dekker, М.У.

Selon les auteurs, le but de la première édition (1981) était de développer les

échanges entre les mycologues écologistes et les théoriciens de l'écologie générale. Le

but fut sans doute atteint puisque voici, onze ans après, un nouveau volume de 976 pa-

ges, 44 articles groupés en 9 parties: Ecologie fongique et théorie de l'écologie; Popu-

lations fongiques, paramètres environnementaux et concept de niche; Organisation des

communautés et des populations fongiques; Interactions entre espèces dans les

communautés fongiques; Développement des communautés; Influence des champi-

gnons sur la dynamique des communautés de plantes; Biomasse fongique et

productivité dans les écosystèmes; Rôle des champignons dans le recyclage des

nutriments; Application des théories écologiques dans les systemes mycologiques. La

moitié des contributions sont nouvelles par rapport à la premiere édition et 80% des

auteurs sont aussi nouveaux venus. C'est dire combien les chercheurs mycologues ont

participé à ce besoin d'échange entre la spécificité de la mycologie et les théories et

méthodes contemporaines des sciences de l'écologie.

On note, dans le présent volume, un important développement de l'étude de la

génétique des populations et de leur biologie évolutive. Plusieurs chapitres portent sur

le ròle des microarthropodes dans la structuration des communautés de champignons.

Une partie tout à fait nouvelle traite de l'importance des champignons dans le

développement des communautés de plantes supérieures. Nouveaux aussi sont les cha-

pitres sur l'écologie des Chytridiales dans le rumen, sur la résistance des champignons

génétiquement modifiés, l'utilisation des techniques de biologie moléculaire pour

étudier l'écologie, les statistiques multivariables, le mutualisme et l'influence des mo-

difications naturelles ou artificielles sur la structure des communautés de champignons

épigés. Dans la huitième partie (rôle des champignons dans le recyclage des

nutriements) un chapitre nouveau aborde la théorie de la modélisation de la contribu-

tion des champignons au recyclage dans les écosystèmes, la comparaison des

méthodologies d'estimation de la biomasse fongique dans les litières et la production

des basidiocarpes et des lichens épiphytes. Enfin la dernière partie est constituée d'une

série d'exemples montrant l'interprétation écologique de données d'observation

mycologique (champignons coprophiles, Hyphomycètes aquatiques, moisissures de sto-

ckage, etc.).

Dans l'ensemble ce volume est très dense, riche en notions théoriques, infor-

mations et mises à jours rédigées par des spécialistes chez lesquels on perçoit bien,

dans la plupart des cas, l'osmose réalisée entre les deux spécialités de mycologie ana-

Iytique et d'interprétation écologique. L'hétérogénéité du contenu et de la forme des

différents chapitres ne rend pas la recherche facile mais on peut y trouver beaucoup

d'informations actuelles et originales et bénéficier d'une bibliographie abondante

groupée par thème.

МЕ. Roquebert

Source : MNHN, Paris

312 ANALYSES BIBLIOGRAPHIQUES

SOCIETAT CATALANA DE MICOLOGIA, Ed. - Bolets de Catalunya, XII

col.leccio, 50 pl. col. (n° 551 à 600), sous encart. Facultat de Farmacia,

Barcelona, 1993. I.S.S.N. 0212-3460.

Comme les précédentes séries de la documentation iconographique éditée par

la Société Catalane de Mycologie, la douziéme livraison réunit cinquante fiches des-

criptives de champignons. Les espèces ont été choisies parmi les Basidiomycètes

(Agrocybe, Amanita, Boletus, Geastrum, Meruliopsis ou Stropharia...) et les

Ascomycétes (Heyderia, Otidea, Poronia, Tarzetta ou Tuber...).

Chaque fiche, plastifiée, propose au recto une photographie de format 19,5 x

13 cm, reproduisant différents aspects des sporocarpes de l'espèce étudiée. Au verso se

trouvent noms, auteurs, synonymie, énumération des caractéres macro- et microsco-

piques ainsi que des références et divers renseignements sur l'écologie, la comestibilité

ou l'étymologie des appellations scientifiques. Signalons а ce propos que le genre

Mycocalia a été établi par J.T. Palmer sur mykes (gr.), fungus, champignon et kalia

(gr.), bird's nest, nid d'oiseau (pl. 584). D'autre part, l'épithète spécifique du Nidularia

duriaeana Tulasne (maintenant Mycocalia duriaeana) rappelle que ce champignon est

dédié au Capitaine Ch. Durieu de Maisonneuve, né vers 1796, décédé en 1878 et di-

recteur du Jardin des Plantes de Bordeaux de 1853 à 1876. Ce botaniste entretint une

active collaboration avec C. Montagne, les frères Tulasne et J.H. Léveillé (pl. 588:

Durieu & Léveillé).

Plusieurs index et, en particulier, une récapitulation par ordre alphabétique des

genres, selon familles, ordres et classes, des espèces présentées depuis le commen-

cement de la collection sont joints aux fiches dont l'illustration photographique est tou-

jours bonne et - souvent méme - excellente.

J. Perreau

FRAITURE A. - Les amanitopsis d'Europe. (Genre Amanita, Agaricales, Fungi).

Synthése critique de la littérature. Meise, Jardin Botanique National de Belgi-

que, Opera Botanica Belgica, vol. 5, 1993, 128 p., ill. LS.S.N. 0775-9592.

LS.B.N. 90-72619-09-9.

Le sous-titre de l'ouvrage reflète exactment la teneur de cette étude consacrée

aux amanites exannulées européennes. Il s'agit en effet, présentée de façon claire et lo-

gique, d'une compilation - dans le sens didactique du terme - de nos connaissances sur

ces champignons qui, comme le souligne l'Auteur, ne sont pas toujours faciles а identi-

fier exactement.

Les données générales concernant les amanitopsis sont exposées dans un pre-

mier chapitre: caractères macro- et micromorphologiques, organoleptiques et

toxicologiques pour les basidiomes, paramétres écologiques et chorologiques pour les

mycéliums. L'accent est mis tout particulièrement sur les différents types de structures

du voile général dont la consistance détermine en grande partie la forme de la volve.

La deuxieme partie traite des problémes de taxinomie et de nomenclature.

L'Auteur y commente d'abord les classifications établies depuis une trentaine d'années

pour le sous-genre Amanita auquel appartiennent les champignons considérés. A ce su-

jet, il propose ensuite dans un tableau synoptique sa propre conception en délimitant

huit stirpes selon deux sous-sections au sein de la section Vaginatae sensu Bas. Des

clés permettent de reconnaître les vrais amanitopsis ainsi que plusieurs espèces rele-

vant de la section Amanita mais dont les basidiomes sont souvent dépourvus d'anneau.

Trois planches délicatement dessinées illustrent l'habitus des champignons étudiés.

Source : MNHN, Paris

ANALYSES BIBLIOGRAPHIQUES 313

Accompagnée de nombreuses références sur l'iconographie, l'écologie ou la

distribution géographique, avec également de multiples notes, l'analyse détaillée des 25

espéces et variétés retenues fait l'objet du chapitre 3. Celui-ci est suivi d'une fort abon-

dante bibliographie - quasi exhaustive sans doute - et de plusieurs index. Par la docu-

mentation solidement argumentée qu'elle apporte, cette étude monographique ne peut

manquer d'étre une aide de grande valeur pour tous les travaux ultérieurs sur les

Amanites.

J. Perreau

Source : MNHN, Paris

Cryptogamie, Mycol. 1993, 14 (4): 315-317

TABLE DU TOME 14

ABDEL-MALLEK A.Y. - voir HASAN Н.А.Н.

ALBERTINI L. - voir JUGNET M.P.

ALBERTINI L. - voir MOSTAFA M.

ALTES A. - voir WRIGHT J.E.

AZBUKINA Z.M. - voir NAUMOV СІ.

BAGY М.МК. - voir HASAN H.A.H.

BAGY М.МК. - voir HEMIDA S.K.

BARRAULT С. - voir JUGNET M.P.

BARRAULT G. - voir MOSTAFA M.

BIOCCA E. - voir DUPRE C.

1. BOIDIN, P. LANQUETIN et б. GILLES - Contribution à la connaissance des

Phanerochaetoidea de France (Basidiomycotina)

BRUN A.M. - voir DAILLANT 0.

BRUXELLES G. - voir JANEX-FAVRE М.С.

CARON D. - voir JUGNET M.P.

CASTILLO A. - voir ILLANA C.

CHEVALIER G. - voir DUPRE C.

CONTU M. - Studi sul genere Agaricus-I Agaricus bisporatus spec. NOV. scure

CUVELIER J.J. - voir DAILLANT О

DAILLANT O., CUVELIER J.J. and BRUN A.M. - Radium and radium decay os

in some macromycetes: first results ..

DIAZ G. y HONRUBIA M. - Respuestas de crecimiento del albardin Som spartum

1.) а la inoculacion con hongos micorricicos y la fertilizacion fosforada ……

DUPRE C., CHEVALIER G., PALENZONA M. et BIOCCA E. - Caractérisation des

mycorhizes de différents Tuber par l'étude du du polymorphisme enzymatique

EL-SAID A.H.M. - Keratinophilic and cycloheximide resistant fungi in soils of Oman

GAILLARDIN C. - voir NAUMOV G.I

GIANINAZZI-PEARSON V. - voir WALKER C.

GILLES G. - voir BOIDIN J.

GLORIES Y. - voir VIVAS N.

HARIDY M.S.A. - Occurrence of yeasts in yoghurt, cheese and whey .....

HASAN H.AH. - Differential action of Cercoran and mn M on sensitive and tolerant

strains of toxigenic fungi... "

HASAN H.A.H. and OMAR S.A. - Selective effect of = insecticides on

metabolic activities and aflatoxins biosynthesis by two Aspergillus spp.

315

195

307

163

271

255

185

Source - MNHN, Paris

316 TABLE DU TOME 14

HASAN H.A.H., BAGY ММК. and ABDEL-MALLEK А.У. - The incidence of fungi

in human axillary hair and their toxigenic potentialities ............

HAWKSWORTH DLL. - Protection des noms dee, courant: un avantage ou une mena-

ce pour le taxinomiste

HEMIDA S.K., BAGY ММК. and KHALLIL А.М. - Utilization of icons »

fungi

HEREDIA G. - Mycoflora associated with green leaves and leaf litter of Quercus

germana, Quercus sartorii and Liquidambar styraciflua in a Mexican cloud

forest ...

HONRUBIA M. - voir DIAZ G.

ILLANA C, MORENO G. & CASTILLO A. - Spanish myxomycetes. VIII. Some

nivicolous myxomycetes from central Spain ....

JANEX-FAVRE МС. PARGUEY-LEDUC A. et BRUXELLES G. - Etude

ultrastructurale de ун de Morchella deliciosa Fr. н,

Discomycètes) ..

JUGNET M.P., BARRAULT G., CARON D. et ALBERTINI L. - Epidémiologie de

Fusarium culmorum (W.G. Smith) Sacc. sur blé dans le sud-ouest de la France.

Mécanismes et agents de dissémination des conidies . ss

KHALLIL A.M. - voir HEMIDA S.K.

KORHOLA M. - voir NAUMOV G.I.

KHOROLA M. - idem

KOTLABA Е. and POUZAR Z. - Pilatoporus maroccanus sp. nov. a new ses of

the Polyporus palustris group ns

LANQUETIN P. - voir BOIDIN J.

MARION-ESPINASSE H. - voir WALKER C.

MATZER M. - Beitrag zur Kenntnis der Ascomycetengatungen Casas

Roselliniopsis und Synaptospora ... Я Si

MORELET M. - voir PRZYBYL K.

MORENO G. - voir ILLANA C.

MORENO С. - voir WRIGHT J.E.

MOSTAFA M., BARRAULT G. et ALBERTINI L. - Essai de lutte biologique contre

Vhelminthosporiose de l'orge par l'utilisation d'une souche peu agressive de

Drechslera teres

MOSTAFA M. - Biological control of Drechslera teres: ability of antagonists to reduce

conidia formation, peso infection and leaf infection in barley (Hordeum

vulgare) ... di NR г

NAUMOV GI, NAUMOVA ES. AZBUKINA ZM., KORHOLA М. and

GAILLARDIN С. - Genetic and Li identification of Saccharomyces

yeasts from far east Asia .

NAUMOV G.I, NAUMOVA E.S., SANCHO E.D. and KORHOLA М. - Taxogenetics

of the Saccharomyces sensu stricto yeasts from western and South Africa .........

NAUMOVA E.S. - voir NAUMOV

NAUMOVA ES. - idem

NUNEZ M. Full compatibility and течи of i arcularis from Spain and Costa

Rica TRE

OMAR S.A. - voir HASAN H.A.H

Source

297

149

207

171

215

229

287

263

MNHN, Paris

TABLE DU TOME 14

PALENZONA M. - voir DUPRE G.

PARGUEY-LEDUC A. - voir JANEX-FAVRE M.C.

POUZAR Z. - voir KOTLABA F.

PRZYBYL K. and MORELET M. - Morphological differences between ru

317

piceae and O. querci, and among O. querci isolates 219

SANCHO E.D. _ voir NAUMOV G.I.

SUBRAMANIAN C.V. - Blastophragma gen. nov. for two interesting Sonya

from southeast Asia .... 39

SUBRAMANIAN C.V. - Phialicorona pleomorpha gen. et sp. nov. and its one È 45

SUBRAMANIAN C.V. - Nusia gen. nov. for two interesting Hyphomycetes from

southeast Asia 109

VIVAS N. et GLORIES Y. - Etude de la flore fongique du chéne (Quercus DI

Caractéristiques du séchage naturel des bois destinés à la tonnellerie 127

WALKER С. GIANINAZZI-PEARSON У. and MARION-ESPINASSE H. -

Scutellospora castanea, a newly described arbuscular mycorrhizal fungus ..... 279

WRIGHT JE, MORENO G. and ALTES A. - PA ыя attenuatus

(Gasteromycetes, Basidiomycetes) new for Europe . wor cen

Analyses bibliographiques 69, 159, 239, 311

Instructions aux auteurs .......

Source : MNHN, Paris

Commission paritaire 16-1-1986 - N° 58611 - Dépôt légal 4 trimestre 1993 - Imprimerie Е. Paillart

Sortie des presses le 31 décembre 1993 - Imprimé en France

iteur : A.D.A.C. (Association des Amis des Cryptogames)

Président : D. Lamy; Secrétaire : B. de Reviers

Trésorier : E. Bury; Directeur de la publication : Н. Causse

Source : MNHN Paris

CRYPTOGAMIE

Diffusion de CRYPTOGAMIE

LE PERIODIQUE FRANCAIS

CONSACRE A LA CRYPTOGAMIE

CRYPTOGAMIE est un périodique édite

par ГА.О.А.С. (Association des Amis des

Cryptogames), dont le siége est au Labo-

ratoire de Cryptogamie du Muséum Na-

tional d'Histoire Naturelle. Les cher-

cheurs de tous pays y publient leurs

travaux en francais, allemand, anglais, es-

pagnol et italien, aprés accord des

Comités de Lecture constitués de

spécialistes de réputation internationale.

CRYPTOGAMIE propose trois sections:

Cryptogamie, Algologie

Cryptogamie, Mycologie

Cryptogamie, Bryologie-Lichénologie

Chaque section publie 4 numéros par an

(tirage: 450 exemplaires).

THE FRENCH JOURNAL

DEVOTED TO CRYPTOGAMY

CRYPTOGAMIE is a periodical

published by A.D.A.C. (Association des

Amis des Cryptogames), settled at Labo-

ratoire de Cryptogamie, Muséum National

d'Histoire Naturelle. Research workers

from the whole world publish their papers

in French, German, English, Spanish and

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commettee that comprises experts of inter-

national renown.

CRYPTOGAMIE offers to its subscribers

three sections:

Cryptogamie, Algologie

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Cryptogamie, Bryologie-Lichénologie

Each section publishes 4 numbers a year

(printing: 450 ех.).

E europe | NMiAfrine C Australie

Amérique Ш Aste

Origine des 453 articles publiés de 1986 à 1991

E Europe Ш Afrique

Amérique [Asie

Répartition des articles publiés de 1986 à 1991 selon la langue

anglais О Allemand

ЕЕ E Espagno! ШШ italien

Source : MNHN, Paris

SOMMAIRE

С. ILLANA, б. MORENO and A. CASTILLO - Spanish myxomycetes. УШ.

Some nivicolous myxomycetes from central Spain...

M.S.A, HARIDY - Occurrence of yeasts in yoghurt, cheese and whey

ОЈ. МАЏМОУ, ES. NAUMOVA, ED. SANCHO and М. KORHOLA -

Taxogenetics of the PERDI sensu stricto yeasts from western

and South Africa È

AHM. ERE Keni n and sche resistant fungi in soils of

АХ F V. GIANINAZZI-PEARSON and H. MARION-ESPINASSE -

Scutellospora castanea, a newly described arbuscular poet

fungus . ze

M. MOSTAFA - EE. control of Drechslera teres: ability of antagonists

to reduce conidia formation, coleoptile infection and leaf infection in

barley (Hordeum vulgare)...

Н.А.Н. HASAN, M.M.K. BAGY and А.У. ABDEL-MALLEK - The incidence

of fungi in human axillary hair and their toxigenic potentialities

М. CONTU - Studi sul genere Agaricus-I Agaricus bisporatus spec. nov.

Table du tome 14 .....

Cryptogamie, Mycol. 1993, 14 (4): 241-317,

241

255

263

271

279