IV ... MYCOTAXON 135(1)

MYCOTAXON

VOLUME ONE HUNDRED THIRTY-FIVE (1) — TABLE OF CONTENTS

Nomenclatural novelties & typifications ....... 0... eee eee eens vii

DRONA OWES, 6 ote an 8 oy wie nt aterg YS cig asl ata gene ert Meth ot ae iy rh le ogee Ray ix

Coriivenda.... Samy. <6. bx MP ee Mc on 6 Rpt CRS AO SU ein aR wR %

EP OE TTI ABOU 9 cect ante Bigs Rigas POL Rl Baca te lies ete et IE age cae ky xi

DOLOSADUIISSION PROCEG HTC. § «AW Dons Sant Perret hea cab ine he apenas ahd Hote hhen sey xiii

TAXONOMY & NOMENCLATURE

Two new genera of gymnopoid/marasmioid euagarics

RONALD H. PETERSEN & KAREN W. HuGHeEs_ 1

Caliciopsis sambaibae sp. nov. from the Brazilian Cerrado

JosE Luiz BEZERRA, MARUZANETE PEREIRA MELO,

JosE EVANDO AGUIAR BESERRA JR, ELLIOT WATANABE KITAJIMA,

SAMARA RAQUEL SOUSA, CRISTIANE DUARTE SANTOS 97

Morphological and molecular identification of

Phlebia wuliangshanensis sp. nov. in China

Ruo-X1A HUANG, KAI-YUE Luo, RuI-XIN Ma, CHANG-LIN ZHAO 103

Pseudocercospora seropedicensis & P. solani-cernui spp. nov.

on Solanum K.M. ANDRADE, P.S. MEDEIROS,

JESSICA REMBINSKI, JUCIMAR M. OLIVEIRA, C.A. INAcIo 119

The nomenclatural history of Umbilicaria spodochroa

and nomenclatural corrections in Umbilicariaceae

Evceny A. Davypov, TEUvo AHTI, ALEXANDER N. SENNIKOV 131

Ochroconis terricola sp. nov. from China

XIN ZHANG, KUN-YING WANG, PENG-PENG REN, YU-LAN JIANG 143

Neomyrmecridium asymmetricum sp. nov. from Ecuador

LIZETTE SERRANO, DAYNET SOSA, FREDDY MAGDAMA,

FERNANDO ESPINOZA, ADELA QUEVEDO, MARCOS VERA,

MIRIAM VILLAVICENCIO, GABRIELA MARIDUENA, SIMON PEREZ-MARTINEZ,

ELAINE MALOosso, BEATRIZ RaAMosS-GARCiA, RAFAEL F, CASTANEDA-Ru1z 151

A comparison of anamorphs of some Pachyphlodes species

and the type of Chromelosporium: are they congeneric?

GREGOIRE L. HENNEBERT & Cony DECOCK 167

Pseudosperma flavorimosum sp. nov. from Pakistan

SANA JABEEN & ABDUL NASIR KHALID 183

Three Rinodina species new to China X1A0-JiA ZHENG & QIANG REN 195

JANUARY-MARCH 2020... V

Gymnopus barbipes and G. dysodes, new records for Pakistan

MALKA SABA, JUNAID KHAN,

SAMINA SARWAR, HASSAN SHER, ABDUL NASIR KHALID 203

Agaricus, Steccherinum, and Typhula species new for Turkey Hakan Istk 213

Tubakia koreana sp. nov. causing Quercus leaf blight

Hyg YOUNG YUN & YouNG Ho Kim 223

MycosBioTa (FUNGA) NEW TO THE MYCOTAXON WEBSITE

Macrofungi from the Hebron and Jerusalem Hills of Palestine (SUMMARY)

Maximus THALER, AYSHA AL-WAHSH,

ALEA Meusrr, ALyssA Rooks, MAzIN QumsrveEH 231

Checklist of Bolivian Agaricales. 2:

Species with white or pale spore prints (SUMMARY)

ELIZABETH MELGAREJO-ESTRADA, DIANA ROCABADO,

Marfa EUGENIA SUAREZ, OSWALDO MAILLARD, BERNARDO ERNESTO LECHNER 233

vI ... MYCOTAXON 135(1)

PUBLICATION DATE FOR VOLUME ONE HUNDRED THIRTY-FOUR (4)

MYCOTAXON for OCTOBER—DECEMBER 2019 (I-xI + 491-740)

was issued on January 13, 2020

JANUARY-MARCH 2020...

NOMENCLATURAL NOVELTIES AND TYPIFICATIONS

PROPOSED IN MYCOTAXON 135(1)

Caliciopsis sambaibae J.L. Bezerra, M.P. Melo & Beserra

[MB 819221], p. 99

Neomyrmecridium asymmetricum R.F. Castaneda, Serrano & D. Sosa

[MB 831330], p. 157

Ochroconis terricola Xin Zhang & Y.L. Jiang

[MB 831137], p. 146

Paramycetinis R.H. Petersen

[IF 555792], p. 9

Paramycetinis austrobrevipes R.H. Petersen

[IF 555793], p. 11

Paramycetinis caulocystidiatus R.H. Petersen

[IF 555794], p. 21

Phlebia wuliangshanensis C.L. Zhao

[MB 830801], p. 111

Pseudocercospora seropedicensis Andrade, Medeiros & Inacio

[MB 819383], p. 121

Pseudocercospora solani-cernui Rembinski, Oliveira & Inacio

[MB 824359], p. 124

Pseudomarasmius R.H. Petersen & K.W. Hughes

[IF 317324], p. 28

Pseudomarasmius efibulatus R.H. Petersen

[IF 555730], p. 31

Pseudomarasmius glabrocystidiatus (Antonin, Ryoo & Ka) R.H. Petersen

[IF 555747], p. 37

Pseudomarasmius nidus-avis (César, Bandala & Montoya) R.H. Petersen

[IF 555746], p. 38

Pseudomarasmius obscurus R.H. Petersen

[IF 555731], p. 50

Pseudomarasmius pallidocephalus (Gilliam) R.H. Petersen

[IF 555745], p.55

Pseudomarasmius patagonianus R.H. Petersen

[IF 555733], p. 68

Pseudomarasmius quercophiloides R.H. Petersen

[IF 555732], p. 73

Pseudomarasmius straminipes (Peck) R.H. Petersen

[IF 555734], p. 83

Pseudosperma flavorimosum Jabeen & Khalid

[MB 823494], p. 187

VII

Vil ... MYCOTAXON 135(1)

Tubakia koreana H.Y. Yun

[MB 814540], p. 225

Umbilicaria subg. Papillophora Davydov, Ahti & Sennikov

[MB 830067], p. 139

= “Umbilicaria subg. Gyrophora” sensu Davydov & al.,

Taxon 66: 1297. 2017.

JANUARY-MARCH 2020...

REVIEWERS — VOLUME ONE HUNDRED THIRTY-FIVE (1)

The Editors express their appreciation to the following individuals who have,

prior to acceptance for publication, reviewed one or more of the papers

prepared for this issue.

Ahmed M. Abdel-Azeem

Vladimir Antonin

Timothy J. Baroni

Jadson D.P. Bezerra

Uwe Braun

Hui Deng

Muhammad Fiaz

Shouyu Guo

Laura Guzman- Davalos

Khalid M. Hameed

Mei-Ling Han

Rosanne Healy

Yuuri Hirooka

Jason M. Karakehian

Abdullah Kaya

Ali Keles

Paul M. Kirk

K.P. Deepna Latha

De-Wei Li

Helmut Mayrhofer

George A. Meindl

Roger Fagner Ribeiro Melo

Josiane Santana Monteiro

Arooj Naseer

Lorelei L. Norvell

Yoshitaka Ono

Clark L. Ovrebo

Shaun R. Pennycook

Christian Printzen

Gerhard Rambold

Gonzalo Romano

Keith A. Seifert

Wen-Xiu Sun

xX ... MYCOTAXON 135(1)

CORRIGENDA FOR MYCOTAXON 135(1)

Cited below are mistakes present in files submitted for PDF conversion in

the current issue but not detected by the authors until after the paper had

gone to press.

p. 30, Fic. 15, label on “green” clade in Pseudomarasmius

FOR: Ps. nitus-avis READ: Ps. nidus-avis

p. 129, line 8 FOR: 2:1-388 READ: 2: 1-388

p. 129, line 21 FOR: V. 13, p. 365-371. READ: 13: 365-371.

p. 224, line 29 FOR: (Maddison & Maddison 2015).

READ: (Maddison & Maddison 2015).

JANUARY-MARCH 2020... XI

FROM THE EDITOR-IN-CHIEF

MyYCOTAXON, RANGE EXTENSIONS, AND TYPE DESCRIPTIONS — ‘There are many

places to publish brief “field guide” descriptions accompanying a photo of a species

collected outside its reported range, but MycoTAxon is not one of them. For each

range extension, MYCOTAXON requires authors to provide a correct scientific name,

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data the species is “supposed” to have but which the specimen lacks), a section

listing the specimens examined and vouchered, illustrations, and a comparison

of the similarities and differences between the new collections and previously

published descriptions.

What we have perhaps NOT stressed enough is that researchers must first closely

compare their find with the TyPE description, even though it may not be possible

to examine the actual type itself. Other critical literature to be consulted includes

type studies and monographic treatments of the putative species. While field guides

(such as the oft-cited Breitenbach & Kranzlin) are helpful in reaching a species

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How BIG IS THAT SPORE AGAIN? — Scale “bars” (lines indicating the relative size of a

specimen or structure ina scientific illustration) should be unobtrusive (i.e. relatively

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dimensions divisible by 5 or 10. Measure macroscopic features in millimeters (using

cm only when a feature is larger than 10 mm) and limit microscopic measurements

to 5 um (small spores), 10 um (larger spores or cells), or 50 or 100 um (complex

hyphal tissues). Scales (and their accompanying structures) should lengthen or

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should group dimensions at the end of each legend for easy reference.

An acceptable legend ends with: “Scale bars: A-C = 500 um; D, F = 10 um;

E,G,H.=5 pm?

An unacceptable legend ends with: “Scale bars: A = 488 um; B = 503.5 um;

C = 425 um; D = 6.8 um; E = 3.2; F= 12.5 um; G = 6.3 um; B = 4.8 um.”

When your editors encounter manuscripts with scale information presented in

the second format, they become very surly indeed.

MyYCOTAXON 135(1) contains 13 papers by 60 authors (representing 13 countries) as

revised by 33 expert reviewers and the editors.

The 2020 January-March MycoTAxon proposes Two new genera (Paramycetinis

& Pseudomarasmius), ONE new subgenus (Umbilicaria subg. Gyrophora), and

12 species new to science representing Caliciopsis & Pseudocercospora from

BRAZIL; Neomyrmecridium from Ecuapor; Ochroconis & Phlebia from CHINA;

xl ... MYCOTAXON 135(1)

Paramycetinis from AUSTRALIA & NEW ZEALAND; Pseudomarasmius from CHILE,

CHINA, and Costa Rica; Pseudosperma from PAKISTAN; and Tubakia from SOUTH

Korea. We also offer four new combinations in Pseudomarasmius.

New species range extensions are reported for [basidiomycetes] Agaricus,

Steccherinum, and Typhula for TuRKEY, Gymnopus for PAKISTAN, and [lichens]

Rinodina for Cutna. Also included are an excellent historical exploration of

the nomenclature (and spelling!) of Umbilicaria spodochroa and a thoughtful

comparison between Pachyphlodes and Chromelosporium anamorphs.

MycotTaxon 135(1) closes with the announcements of two mycobiota [now

posted on www.mycotaxon.com] covering [1] macrofungi in Palestine and [2]

white and pink-spored agarics in Bolivia.

Covib-19—This issue goes to press (late as usual, unfortunately) in the middle of

a global endemic caused by a new corona-virus. The fallout from this plague will

unfortunately alter our lives on many fronts beyond the current deaths and isolation

we now face. Take heart, persevere, and strive ever upward. I close with these lines

by Alicia Jo Rabins:

“Through windows and screens

We sing and raise a glass

Simultaneous but not together.

Oh my friends my beloved strangers

I never knew our closeness

Until it was gone”

For now— work safely apart; stay healthy together.

Lorelei Norvell (Editor-in-Chief)

14 April 2020

JANUARY-MARCH 2020... XIII

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MY COTAXON

January-March 2020—Volume 135, pp. 1-95

https://doi.org/10.5248/135.1

Two new genera of gymnopoid/marasmioid euagarics

RONALD H. PETERSEN & KAREN W. HUGHES

Ecology and Evolutionary Biology, University of Tennessee,

Knoxville, TN 137996-1100, TN

“CORRESPONDENCE TO: repete@utk.edu

ABSTRACT— Twonewgenera, Paramycetinisand Pseudomarasmius, are placed phylogenetically

within the /omphalotaceae among genera allied with Gymnopus. Paramycetinis comprises two

Antipodal taxa related to Mycetinis. Both Paramycetinis species are characterized by luxuriant

rhizomorphs, with basidiomata arising occasionally as side branches but also separately from

rhizomorphs. Pseudomarasmius accommodates four new species plus four others previously

placed in Marasmius. Pseudomarasmius encompasses significant basidiomatal variation, but

nrLSU- and ITS-based phylogenies support its taxa in a monophyletic clade that occupies

a position related to some Rhodocollybia. All Pseudomarasmius and Paramycetinis species

are described (or redescribed) and illustrated. Both genera exhibit characters also found in

other superficially similar lineages, making non-molecular based diagnoses of genera in the

/gymnopus/marasmiellus clades difficult.

KEY worps— biogeography, clamp connections, morphological taxonomy

Introduction

For the last few decades, numerous, small, “white-spored” saprotrophic

mushrooms have undergone a comprehensive reclassification. Especially

edifying has been the introduction of molecule-based phylogenetic analyses

at about the turn of the 21* century. New taxa have been identified at several

nomenclatural ranks, from infraspecific (Aldrovandi & al. 2015; Petersen &

Hughes 2016, 2017a,b) to class (Moncalvo & al. 2002, Matheny & al. 2006,

Dentinger & al. 2016). In many cases, detailed morphological descriptions

have accompanied molecular analyses (or the reverse), but “morphological

taxa’ are still being proposed without phylogenetic placement (Petersen &

2 ... Petersen & Hughes

Hughes 2016a, 2017; Retnowati 2018) when materials are too old or degraded

to produce molecular data or the appropriate instrumentation is unavailable.

Desjardin (1989) summarized the classification and publication history

of Marasmius and its segregation from Gymnopus (sensu Persoon 1801) or

Collybia (sensu Fries 1821). The taxonomic history of Gymnopus (as Collybia)

was outlined by Halling (1983). Subsequently, Antonin & al. (1997) discussed

infrageneric taxonomy within Marasmius and Gymnopus (as Collybia), and

Hughes & al. (2001) segregated Gymnopus (G. fusipes typus generis) from

Collybia s.s. (C. tuberosa typus generis).

Owings & Desjardin (1997) apparently conducted the first research into

the collybioid/marasmioid complexes using DNA sequences based on a

more expanded thesis (Owings 1997; nrITS-based). Their work revealed that

marasmioid fungi were not monophyletic but segregated into three lineages:

present-day Physalacriaceae, Omphalotaceae, and Marasmiaceae. Moncalvo

& al. (2000, 2002; LSU-based) essentially inferred the same. Mata & al. (2004;

ITS- & LSU-based) showed that Marasmiellus juniperinus (typus generis)

belonged in the omphalotoid complex, Wilson & Desjardin (2005) tied the

morphological fragmentation of Marasmius to an nrLSU-based phylogeny

and refined phylogenetic relationships of a wider array of organisms, and

Mata & al. (2007; ITS-based) offered a more comprehensive phylogeny

of /omphalotaceae, with emphasis on Gymnopus. The nomenclatural

consequence of these works was circumscription of two major clades/

families, Marasmiaceae and Omphalotaceae. A further consequence was more

detailed fragmentation within these groups (Petersen & Hughes 2016a, 2017),

in which Gymnopus as summarized by Mata & al. (2007) is clearly para- to

polyphyletic with several related genera (Rhodocollybia, Lentinula, Mycetinis,

Connopus) interspersed and Gymnopanella (Sandoval-Leiva & al. 2016)

also associated. Three genera (Lentinula, Rhodocollybia, Mycetinis) are well-

populated while two (Connopus, Gymnopanella) currently appear monotypic.

If the concepts of Wilson & Desjardin (2005) are advanced to the present day,

two large and complex clades contain traditional gymnopoid/marasmioid

taxa: /Gymnopus incorporates Gymnopus sects. Levipedes, Striipedes, and

Impudici, the traditional Micromphale (including sect. Perforantia), Caripia,

and Marasmius sect. Androsacei; /Marasmiellus (including M. juniperinus,

typus generis) includes Gymnopus sect. Vestipedes as well as Marasmius

(Marasmiellus) ramealis, the type of Collybiopsis (J. Schrot.) Earle (Hughes

& Petersen, ined.). /Marasmiellus) would appear to be a candidate for

fractionation.

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 3

Oliveira & al. (2019) recognized their clade I (called “Gymnopanella”)

based on the taxon described by Sandoval-Leiva & al. (2016) and which

included two provisional names that had appeared in phylogenies by Petersen

& Hughes (2016). We regard those two provisional names as belonging in a

separate generic unit, here proposed as Paramycetinis, based on its placement

as sister to Mycetinis.

The intention of this paper is to propose two new genera, one comprising

two new Antipodal species and the other accommodating several species.

FiGureE | shows the relative placement of the new genera based on nrLSU-

sequence resolution.

Materials & methods

Abbreviations: Pa = Paramycetinis; Ps = Pseudomarasmius; M = Marasmius;

Ma = Marasmiellus; Mi = Micromphale; My = Mycetinis. Colors cited

alphanumerically are from Kornerup & Wanscher (1967) or Munsell (1961), in

quotation marks from Ridgway (1912), and special reference to Maerz & Paul

(1950); BF = bright field microscopy; PhC = phase contrast microscopy (see also

Petersen & Hughes 2017); TFB = Tennessee field-book number, assigned to fresh

collections. In order to track information for collections from TENN, numbers are

the last five digits of the fungal herbarium accession number (in MyCoPortal as

TENN-F-0xxxxx); GSM (also GSMNP) = Great Smoky Mountains National Park,

Tennessee/North Carolina. Percent similarity values within and between clades are

given on the right.

Culture media reported to encourage rhizomorph production followed

directions by Farnet & al. (1999). Two different whole wheat flours (“whole

wheat flour” and “stone-ground whole wheat flour,’ both obtained locally)

produced no difference in rhizomorph occurrence, number, or vigor. The term

“textura” to describe the intricate crust formed by many marasmioid, mycetinoid,

marasmielloid, and gymnopoid isolates in culture has been borrowed from Korf

(1973), who used it to describe the excipular layer of apothecia.

Metrics of microstructures were calculated using a phase-contrast Olympus

BX60 microscope fitted with phase contrast imagery and microphotographs were

produced using an Olympus Qc color camera. Microscope mounts were usually in

3% KOH solution or Melzer’s reagent.

Molecular methods for DNA extractions, PCR and Sanger sequencing of the

nuclear ribosomal ITS and LSU regions follow Aldrovandi & al. (2015). PhyML

phylogenetic analyses were performed in Geneious 11.0.3 (2017). Sequences have

been deposited in GenBank (TABLE 1). Specimen data are available in MyCoPortal

(2018). Nexis files containing nrLSU sequences and alignments were deposited in

the Dryad Data Repository https://doi.org/10.5061/dryad.4081h). ITS sequence

%-similarity is the average of all paired sequences computed by Geneious 11.0.3

(2017) as “percent identity”.

4 ... Petersen & Hughes

Anthracophyllum

DQ457670 Gymnopus contrarius

89 Neonothopanus

<—— | Omphalotus

200 <— | Gymnopus pinophilus MGIII-Gordon

9 85. <— | Gymnopus foliiphilus ‘ :

oy Gymnopus section Perforantia

q Gymnopus perforans (Micromphale sect. Perforantia)

<—_| Gymnopus foetidus (Micromphale sect. Micromphale)

AY639432 Gymnopus vitellinipes

Gymnopus ocior, G. dryophilus, G. bicolor, G. sepiiconicus,

G. aurantiipes, G. earleae

AF223172 Gymnopus acervatus sensu Moncalvo et al.

<=] Gymnopus erythropus, G. spongiosus /C Mata et al. (2007)

<——_] G. hariolorum, G. polyphyllus

—Ice dysodes, G. barbipes, G. iocephalus

<—] G. montagnei and AF261326 G. sp.

/B Mata et al. (2007)

Gymnopus neobrevipes, G. portoricensis (Micromphale sect. Rhizomorphigena)

Petersen and Hughes, 2018

——_] /novae-angliae, /frigidomarginatus, /adventitius

<—] Gymnopus androsaceus (Marasmius sect. Androsacei)

Wilson and Desjardin (2005)

/gymnopus

<=] /inflatotrama

—] Gymnopus fusipes /A Mata et al. (2007)

<——_]} /novomundi MGIIl Gordon

0.06 changes

Paramycetinis (austrobrevipes, caulocystidiosus)

——] Mycetinis (Marasmius) scorodonius My cetinis

<——_| Mycetinis (Marasmiellus) opacus, My. copelandii Wilson and Desjar din (2005)

AY639437 Mycetinis (Marasmius) applanatipes Petersen and Hughes (2017)

— J mycetinis (Marasmius) alliaceus

22 ——_} Connopus acervatus

<i Rhodocollybia

<All Pseudomarasmius (pallidocephalus, glabrocystidiatus)

<—— | Lentinula

=< | Gymnopus dichrous

AY639430 Gymnopus termiticola

<—_| Gymnopus gibbosus

<— _]| Gymnopus luxurians, G. pseudoluxurians

Gymnopus quercophilus

<——_} Gymnopus peronatus

<——_| Gymnopus subnudus

Marasmiellus vaillantii

Gymnopus villosipes,

G. mesoamericanus,

G. biformis,

G. menehune,

G. indoctus,

G. eneficola,

G. confluens

/D Mata et al. (2007)

Gymnopus (Marasmiellus) ramealis

Gymnopus afn. dichrous

Gymnopus melanopus

Gymnopus diminutus,

<<] Marasmiellus synodicus, Gymnopus nonnullus

<——_] Gymnopus juniperinus

/Marasmiellus, Wilson and Desjardin (2005)

<——_] Gymnopus subpruinosus, G. afn. moseri

Fic. 1. PhyML analysis of 191 nrLSU sequences within the Omphalotaceae using 100 bootstrap

replicates. Clades were collapsed to better show broad relationships. Species names in blue

represent /marasmiellus (Wilson & Desjardin 2005). Species names bordered in red represent

/gymnopus (Wilson & Desjardin 2005). The major clades reported in Mata & al. (2007) are

bordered in blue. The genera Paramycetinis and Pseudomarasmius are in green.

Paramycetinis & Pseudomarasmius gen. & spp. nov. ...

TABLE 1. Collections used in phylogenetic analyses

NAME GENBANK # HERBARIUM # FIELD # LOCALITY

[Fic. 2—Gymnopus, Paramycetinis, Mycetinis]

Gymnopus perforans KY026675 TENN-F-61587 TFB13119 Canada, Quebec

KY026719 TENN-F-69059 TFB14395 Canada,

New Brunswick

G. foliiphilus KY026703 TENN-F-68145 TFB14291 USA, Mississippi

G. sequoiae KY026741 TENN-F-69325 TFB14620 USA, California

% woe maked KY026622 TENN-F-50135 —_ TFB4033 Australia

KY026637 TENN-F-53149 TFB3591 Australia

KY026638 TENN-F-53181 TFB3585 Australia

P. caulocystidiatus KY026642 TENN- F-53683 TFB7572 New Zealand

KY026643 TENN-F-53721 TFB7588 New Zealand

KY026644 TENN- F-53725 TFB7589 New Zealand

KY026645 TENN-F-54050 TFB7148 New Zealand

Mycetinis alliaceus AY635776 TENN-F-55620 TFB8960 Russia

DQ450004 TENN-F-55555 TFB8935 Russia

KY696752 TENN-F-55630 TFB8970 Russia

KY696765 TENN-F-67899 TFB14149 Germany

KY696766 TENN-F-67911 TFB14161 Germany

KY696770 TENN-F-69243 TFB14548 Slovakia

KY696771 TENN-F-69244 TFB14549 Slovakia

M. applanatipes KY696775 SFSU DED6628 — USA, California

M. copelandii KY696750 TENN-F-55408 TFB8084h1 USA, California

KY696751 TENN-F-55408 TFB8084h2 USA, California

M. kallioneus KX958397 GB-0150513 — Norway

eae USA, Tennessee,

M. opacus KY696758 TENN-F-60016 TFB11787 GSMNP

KY696767 TENN-F-69190 TFB14499 Mississippi

KY696768 TENN-F-69200 TFB11490h1 Mississippi

KY696769 TENN-F-69200 TFB11490h2 Mississippi

M. querceus KY696785 UPS F740422 — Sweden

M. salalis KX752265 DAOM175251 = Canada, BC,

Vancouver Isl.

M. scorodonius DQ450006 TENN-F-50346 TFB4749 Switzerland

6 ... Petersen & Hughes

NAME

GENBANK #

KY696725

KY696726

KY696727

KY696728

KY696729

KY696730

KY696731

KY696732

KY696733

KY696734

KY696735

KY696736

KY696737

KY696738

KY696739

KY696740

KY696741

KY696742

KY696743

KY696744

KY696745

KY696746

KY696747

KY696748

KY696749

KY696753

KY696754

KY696757

KY696786

KY696787

HERBARIUM #

TENN-F-50343

TENN-F-50352

TENN-F-50369

TENN-F-50377

TENN-F-50447

TENN-F-50515

TENN-F-50522

TENN-F-50533

TENN-F-50689

TENN-F-50696

TENN-F-50763

TENN-F-50809

TENN-F-51233

TENN-F-51442

TENN-F-53465

TENN-F-53466

TENN-F-53467

TENN-F-53468

TENN-F-53469

TENN-F-53470

TENN-F-53471

TENN-F-53472

TENN-F-53474

TENN-F-53568

TENN-F-57663

TENN-F-58260

TENN-F-59615

WTU19061H1

WTU19061H2

FIELD #

TFB4746

TFB4755

TFB4772

TFB4780

TFB4372

TFB3778

TFB3785

TFB3796

TFB3701

TFB3708

TFB3644

TFB3690H1

TFB3690H2

TFB4915

TFB5031

TFB5014

TFB4969

TFB5025

TFB5038

TFB4989

TFB5046

TFB4939

TFB5067

TFB5005

TFB7261

TFB2782

TFB10400

TFB11652

LOCALITY

Switzerland

Mexico

Sweden

USA,

North Carolina

USA, TN GSMNP

USA,

North Carolina

Canada,

Nova Scotia

Canada, Nova

Scotia

USA, New York

Canada,

Nova Scotia

Canada,

Nova Scotia

USA, Maine

USA,

North Carolina

USA, New York

USA,

North Carolina

USA, Maine

Sweden

USA, Georgia

Russia, Western

Russia

USA, Washington

NAME

GENBANK #

HERBARIUM #

FIELD #

[Fic. 15—Gymnopus, Marasmius, Pseudomarasmius, cf. Rhodocollybia, etc.]

Gymnopus glabrocystidiatus

(T)

Marasmius sp.

Pseudomarasmius

quercophiloides

Ps. efibulatus

Ps. patagonianus

Ps. nidus-avis

Ps. pallidocephalus

Rhodocollybia

aff.

R. sp.

Tricholomataceae

NRI152899

KF251072

KR673444

MK268235

MK268234

MK268236

KY352649

KY026646

KY026723

KY026732

KY026733

KY026753

MK020094

MK268237

MK268238

MH016872

KY026635

FJ596762

FJ596763

KY026636

KY026684

KY026685

KY026691

KY026746

FJ475743

KC966049

AY313287

LCO14889

JN890248

JN890219

JN890249

BRNM718676

KFRI1935

KA12-0382

TENN-F-49177

TENN-F-56187

JAC10695

TENN-F-54424

TENN-F-54912

TENN-F-69189

TENN-F-69310

DPL11763

Rhizomorph

culture

FLAS-F-62667

FLAS-F-62668

FLAS-F-60891

TENN-F-52401

TENN-F-59896

TENN-F-52427

TENN-F-65829

TENN-F-66344

no specimen

soil sample

TENN-F-58798

no specimen

soil sample

TFB3162

TFB7070

TFB7364

TFB9087

TFB14498

TFB14607

TFB14606

MES883

MES1477

MES3139

5610 culture

TFB11778h2

TFB11778h1

TFB5698

TFB13933h1

TFB13933h2

SAT11-179-05

5015 culture

TFB10712

8PBLI11

Paramycetinis & Pseudomarasmius gen. & spp. nov. ...

LOCALITY

Korea

South Korea

China, Yunnan

New Zealand,

South Island

New Zealand

Chile

USA, Louisiana

USA, Mississippi

USA, Alabama

USA, Texas

USA, Mississippi

USA, Florida

USA, Idaho

USA, TN GSMNP

USA, Washington

USA, New York

USA, TN GSMNP

Canada, Nova

Scotia

Sweden

Alaska

Greenland

Japan, Nagano

Guyana

8 ... Petersen & Hughes

Paramycetinis

The nrLSU sequence analysis places Paramycetinis within a moderately

supported clade that includes Mycetinis (Fic. 1). A PhyML analysis of

Mycetinis and Paramycetinis nrITS sequences (Fic. 2) places Paramycetinis

as sister to a clade containing several Mycetinis species. The average base pair

identity between Paramycetinis and Mycetinis is 91.72%.

Only sparse literature on marasmioid fungi from Australia (Fuhrer

1985), South Australia (Cleland 1934, Grgurinovic 1997), and Tasmania

(Eygelsheim 1981, Fuhrer & Robinson 1972) preceded introduction of

molecular phylogenetic analyses. Those publications grouped collections

of small marasmioid basidiomata as “Marasmius crinis-equi” or “horse-

hair fungi” according to the presence of obvious rhizomorphs when, in

fact, several discrete taxa qualify under this superficial diagnosis and await

formal proposals. More recently, Desjardin & Horak (1997) revisited earlier

work by Stevenson (1964) and Horak (Horak 1971a,b) to revise numerous

marasmioid fungi from Papua New Guinea, New Caledonia, and New

Zealand, and Desjardin & al. (2000) summarized Marasmius for parts of

Indonesia.

Even less literature dealt with Micromphale (here understood as

Gymnopus sect. Micromphale) on the Australia/New Zealand landmasses

[but see Cooper & Leonard (2013) on micromphalioid Gymnopus taxa].

To our knowledge, only Grgurinovic (1997) has dealt with species of sect.

Perforantia (Agaricus perforans, typus sectionis); she placed Micromphale

australiense, M. rugosum, and M. villosipes (all from South Australia) in

Micromphale based solely on morphological characters.

Owings & Desjardin (1997) first identified three infrageneric phylogenetic

clusters accommodating Marasmius taxa; subsequent phylogenetic studies

refined this early finding. In nrLSU sequence analyses of marasmioid and

gymnopoid fungi, Wilson & Desjardin (2005) identified a clade including

taxa previously segregated into the morphogenus Mycetinis Earle (1909;

Agaricus alliaceus, typus generis). This clade was also resolved by Mata &

al. (2004, 2007) but not nomenclaturally separated from Gymnopus, which

they accepted in a very broad sense. Petersen & Hughes (2016), who also

showed a /mycetinis clade in their overall nrLSU phylogeny of Gymnopus

and associated genera, found that /mycetinis comprised three subclades,

of which the sister subclade to Mycetinis contained sequences of two taxa

known only from Tasmania and New Zealand. This small subclade, sister to

/mycetinis, is taken up here as Paramycetinis.

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 9

/Mycetinis is presently accepted as inclusive of traditionally disparate

morphological taxa. From its re-disclosure by Wilson & Desjardin

(2005; LSU-based), My. alliaceus and My. scorodonius were joined by

Marasmiellus opacus, so Wilson & Desjardin (2005) furnished a discussion

of morphological characters justifying inclusion of Ma. opacus in /mycetinis

and proposed appropriate transfers. Likewise, Mata & al. (2007; ITS-based)

placed Ma. candidus in the same clade although Wilson & Desjardin (2005)

had placed it in /tetrapyrgos, an adjoining clade. At genus rank, Mycetinis

has been accepted by Antonin & Noordeloos (2010) and lately surveyed

by Petersen & Hughes (2017b). One ITS-based PhyML phylogeny (Fic. 2)

shows the relationship of Paramycetinis to Mycetinis.

Oliveira & al. (2019) recognized a small clade (their clade I) called

“Gymnopanella, based on the taxon described by Sandoval-Leiva & al.

(2016). Their clade I included two provisional names that appeared in

phylogenies by Petersen & Hughes (2016). We consider the two provisional

names to belong in a separate generic unit, here proposed as Paramycetinis.

Paramycetinis R.H. Petersen, gen. nov.

IF 555792

Basidiomata diminutive, associated with dominant black rhizomorphs; a monophyletic

clade.

TYPE SPECIES: Paramycetinis austrobrevipes R.H. Petersen

EryMo_oey: reflecting the sister phylogenetic relationship to Mycetinis.

BASIDIOMATA combining characters of Gymnopus sect. Micromphale subsect.

Perforantia, Gymnopus sect. Androsacei, and Marasmius sect. Sicci, minute

to small, arising as rhizomorph branches or separate from rhizomorphs.

PILEus small (3-15 mm broad), some shade of brown or vinaceous brown,

sometimes sulcate-striate, smooth or minutely pruinose; pileus trama thin,

pliant. LAMELLAE adnexed to adnate, sometimes attached to an adherent

pseudocollarium, close to subdistant, from ridge-like to well-developed, usually

with lamellulae in 1-2 ranks, off-white to dull vinaceous gray. ST1PE capillary,

<3 mm broad, terete, more or less equal, solid to stuffed, glabrous-shining to

minutely velutinous, brown to brown-black, especially downward, insititious

(when not a rhizomorph branch). Ro1IzomMoRPHS common, ranging from very

slender to stout, sometimes dominant and forming loose nets, usually black.

PILEIPELLIS commonly involved in slime as a matrix and/or as gelatinizing

hyphal walls, usually including diverticulate hyphae and broom cell-like hyphal

termini. PLEUROCysTIDIA ubiquitous, narrowly to broadly fusiform; contents

10 ... Petersen & Hughes

KY026675 G. perforans

KY026719 G. perforans Gymnopus

KY026703 G. foliiphitus

KY026741 G. sequoiae

KY026643 NZ.

KY026642 NZ

KY026645 NZ Paramycetinis caulocystidiatus

100)

KY026622 Australia

KY026638 Australia

KY026637 Australia

Paramycetinis austrobrevipes

91.72%

KY696765 Germany

KY696771 Slovakia

AY635776 Russia.

KY696770 Slovakia

KY696766 Germany

KY696752 Russia

DOQ450004 Russia.

KY696785 M. querceus

KX958397 M. kallioneus | Mycetinis spp.

75"L. KY696775 M. applanatipes

rea ae ene Mycetinis salalis

kY696750 M. copelandii ae copatandl

971 KY696751 M. copelandii

KY696758 USA, TN

KY696768 USA, MS

KY696769 USA, MS

KY696767 USA, MS

KY696734 USA, TN

KY696736 USA, TN

KY696737 USA, TN

KY696753 USA, GA

KY696732 USA, TN

KY696757 Sweden

KY696757 Russia

KY696725 Switzerland

KY696730 Switzerland

KY696730 Sweden

KY696727 Switzerland Mycetinis scorodonius

KY696749 Sweden

KY696754 Russia

KY696786 USA, WA

KY796787 USA, WA

KY696731 Sweden

KY696729 Mexico

DQ450006 Switzerland

KY696726 Switzerland

KY696744 USA, ME

KY696743 Canada, NS

KY696745 USA, NC

KY696748 USA, ME

KY696741 USA, NY

KY696746 USA, NY

KY696738 USA, NC

KY696740 Canada, NS

0.06 changes KY696747 USA, NC

KY696739 Canada, NS

KY696742 Canada, NS

Mycetinis alliaceus

Mycetinis opacus

Fic. 2. Maximum likelihood analysis of combined ribosomal RNA ITS and LSU sequences.

Bootstrap values are given to the left of the supported node. Numbers are GenBank accession

numbers (See TABLE 1). For collections from the United States and Canada, the state or province

postal code is given. Percent similarity values within and between clades are given on the right.

Paramycetinis & Pseudomarasmius gen. & spp. nov.... 11

more or less homogeneous, occasionally vaguely partitioned. Basrp1a clavate,

4-sterigmate. Basip1ospores thin-walled, smooth, inamyloid, white or off-

white in deposits. CHEILOCYSTIDIA present, arbuscular (stalked, branched)

or broadly clavate, with or without diverticula. STrPE MEDULLARY HYPHAE

strictly parallel, usually involved in a slime matrix. CAULOCYSTIDIA present or

absent, when present gregarious to scattered (not a dense turf), setoid. CLAMP

CONNECTIONS present. Molecular analysis resolving a monophyletic clade

associated with Mycetinis.

Key to taxa of Paramycetinis

1. Rhizomorphs stout (>1 mm diam), forming loose nets; basidiomata arising from

or separate from rhizomorphs; cheilocystidia clavate with clusters of apical

SetwlaeGlasiiaarints: ete ee Ms eet fa et as 2) ee ce Pa Pa. austrobrevipes

1. Rhizomorphs slender (<1 mm diam), extensive; basidiomata arising separate

from rhizomorphs; cheilocystidia clavate, without diverticula; fruiting on

dead leaves and twigs of Nothofagus (New Zealand)......... Pa. caulocystidiatus

Paramycetinis austrobrevipes R.H. Petersen, sp. nov. Figs 3-8

IF 555793

Differs from Paramycetinis caulocystidiatus by its broader rhizomorphs that form loose

nets, by its basidiomata that arise as rhizomorph branches and/or from woody substrate,

and by its broom cell-like cheilocystidia.

Type: Australia, Tasmania, Gordon-Pedder National Heritage Area, Rainforest Nature

Walk vicinity, 8.V1.1991, coll. RHP & KWH, TFB 4033 (Holotype TENN-F-050135).

EryMoLoGey: referring to micromorphological similarities to Gymnopus neobrevipes.

BASIDIOMATA (Fic. 3) marasmioid, diminutive, arising as branches from

rhizomorphs and/or directly from woody substrate. PILEUs 2-12 mm broad,

convex to shallowly conical, sometimes abruptly or broadly umbonate and

even then abruptly umbilicate, deeply sulcate-striate to non-striate, matte,

at high magnification (50x) telltale wispy white hairs can be seen in the

sulcate depressions; disc (including umbo) 5F6 (“Prout’s brown”), 6E3 (“hair

brown’), 9D3 (“benzo brown’), 7C4 (“wood brown’), 6E4 (“fuscous”),

7C6 (“Mikado brown’) to 9C3 (“cinnamon drab”); limb 6C5 (“sayal brown”),

6B5 (“cinnamon”), 17B2 (“light drab”), 7B2 (“tilleul buff”), to 9B2 (“vinaceous

buff”), 8D5 (“army brown”), 9C3 (“cinnamon drab”), 9B3 (“light cinnamon

drab”) to 7C5 (“fawn color”); margin 6A3 (“pinkish buff”) to 4A3 (“cartridge

buff”). LAMELLAE adnexed, subdistant (total lamellae 20-24), about 1 mm

broad, thickish, without anastomoses, 7B2 (“tilleul buff”), 9B2 (“vinaceous

buff”), 6B4 (“cinnamon buff”) to 6A3 (“pinkish buff”), not marginate; dried

12 ... Petersen & Hughes

lamellar trama dark brown and glassy, as though gelatinized. Stripe 18-85

x 0.8-1.2 mm, terete, stuffed (not hollow), appearing glabrous-shining but

minutely pruinose at least upward and downward (35x), black to abruptly

6C5 (“sayal brown”), 6B3 (“cinnamon buff”) or mahogany at very apex (not

concolorous with lamellae), downward totally black; medullary portion

cloud gray, cortical layer thin, nearly black (40x); insertion non-insititious;

vesture sparse, pallid, near 7B2 (“tilleul buff”). Ro1IzoMoRPHs >1 mm broad,

gradually tapering to 0.2-0.5 mm distally, terete or somewhat compressed,

extensive to almost absent, often forming a rudimentary net. ODor negligible;

TASTE negligible or weak of garlic; consistency tough then mealy.

HABITAT & PHENOLOGY: Nothofagus wood (and then densely gregarious)

and twigs, at or near forest floor, Arthrotaxis dead branchlets; to this time,

May-June.

PILEIPELLIS constructed of the following: 1) slender hyphae 2-3.5 um

diam, thick-walled (wall <0.5 um thick), sparsely encrusted (Fic. 4a)

with ornamentation appearing as flakes adhered to outer hyphae wall;

2) slender hyphae 1.5-3 um diam, firm-walled with slender, sometimes

awl-shaped diverticula (FIG. 4B, 54-D), obscurely clamped, forming a

indiscrete layer in a mucoid matrix (i.e. significant debris including bacteria

and collapsed spores, PhC); 3) slender, coralloid hyphal termini (Fic. 5£,F);

4) thicker hyphae 4-12 um diam, refringent (PhC), infrequently branched,

meandering to appear a bit like lattice, thick or gelatinized—walled

(wall <2.5 um thick), with outer profile smooth and clear but inner wall

contour irregular, obscurely clamped; termini occasionally more or

less erect as “pileal hairs.” PLEUROCYSTIDIA (FIGS 6B, 7A—C) common,

26-33 x 5-9 um, fusiform to mucronate, conspicuously clamped,

usually with partitioned contents. BAsipIA 23-28 x 6-9 um, clavate,

4-sterigmate (sterigmata <6 um long, somewhat bowed, stout), obscurely

clamped; contents homogeneous. BASIDIOSPORES (FIG. 6A) 6-9 x 3.5-4.5

(-5) um (Q = 1.502.43; Q™ = 1.92; L™ = 7.47 um), more or less pip-shaped

but often tapered slightly proximally (that is, perhaps marasmioid), smooth,

thin-walled, inamyloid; contents homogeneous. CHEILOCYSTIDIA (FIGS 6D,

7E-H) locally common to scattered among basidia, 17-32(-40) x 5-12 um,

clavate to ventricose-rostrate, obscurely clamped, thin-walled below, firm-

walled above, surmounted by a corona of extremely complex, vermiform

to slender-digitate, refringent (PhC) diverticula; diverticula <7 x 0.71 um,

gnarled, often branched. STIPE MEDULLARY HYPHAE hyaline, 2-5.5 um

diam, firm- to thick-walled, seldom but conspicuously clamped; CORTICAL

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 13

Fic. 3. Paramycetinis austrobrevipes.

Habit. A. (TENN 53181). B. (TENN-F- 050212). Scale bars: 20 mm.

HYPHAE adherent, 3-7 um diam, yellow singly, dark ochraceous brown

and refringent in mass (PhC), involved in a thin film of mucus, with side

branches ranging from simple lobes to caulocystidia. CAULOCYSTIDIA (FIG. 8)

14 ... Petersen & Hughes

Fic. 4. Paramycetinis austrobrevipes. Structures of pileipellis. A. Encrusted hyphae (TENN 50119).

B. Diverticulate hyphae (TENN-F-050135). C. Thick-walled lattice hyphae (TENN-F-050135).

D. Pileal hairs (TENN-F-050119. Scale bars: 20 um.

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 15

Fic. 5. Paramycetinis austrobrevipes (TENN-F-050135). Pileipellis elements. A. Layer of

diverticulate hyphae as embedded within pileipellis. B. Layer of diverticulate hyphae.

C,D. Individual diverticulate hyphae. E, F Broom cell-like hyphal termini. G,H. Hyphae with

gelatinized walls. Scale bars: 10 um.

scattered, <105 x 46 um (at widest point, narrowing to 3-3.5 um diam at base,

2-3 um diam at apex), gregarious but more or less evenly distributed, usually

single, rarely in pairs, as side branches of stipe surface hyphae, apparently

near the terminus of such hyphae (and therefore appearing as though from

an asymmetrical origin, yellow and refringent (PhC), so refringent that wall

thickness is hardly discernible (when wall seems to be discernible, 1-1.5 um

thick), narrowly rounded apically and apparently without adherent debris,

dextrinoid (Melzer’s reagent + PhC) or brown (Melzer’s reagent + BF),

not internally septate.

16 ... Petersen & Hughes

Fic. 6. Paramycetinis austrobrevipes (TENN-F-050135).

A. Basidiospores. B. Pleurocystidium. C. Basidia. D. Cheilocystidia. E. Caulocystidia.

Scale bars: A = 5 um; B-D = 20 um.

ComMENTARY: Pileus shape in Pa. austrobrevipes varies from convex with an

abrupt, distinct umbo to a gradual and shallow umbo to no evidence of an

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 17

Fic. 7. Paramycetinis austrobrevipes (TENN-F-050135).

A-C. Pleurocystidia. D. Basidium. E-H. Cheilocystidia. Scale bars: 10 um.

umbo. In all cases, the disc (with or without umbo) is significantly darker than

the limb.

The pileus micromorphology of Pa. austrobrevipes approaches that of the

North American Marasmius brevipes [= Gymnopus neobrevipes R.H. Petersen &

18 ... Petersen & Hughes

Fic. 8. Paramycetinis austrobrevipes (TENN-F-050135).

Caulocystidia. Scale bars: 10 um.

Hughes 2019)]. Basidiomata of both species arise on woody substrate (usually

twigs with thin bark intact or occasionally as branches from an extensive

rhizomorph net), but G. neobrevipes basidiomata are small and thumb tack-

shaped while those of Pa. austrobrevipes are tall and mycenoid. Pileus colors

are similar but also characteristic of several taxa in Gymnopus sect. Androsacei.

Rhizomorphs and stipes are black, although stipe in G. neobrevipes is glabrous

throughout its length. There is no record of a weak or latent taste of garlic

for G. neobrevipes. Comparison of notes and drawings of G. neobrevipes

specimens indicate that it also has a gelatinizing pileipellis, but with a more

demonstrable “ramealis-structure.’” Gymnopus neobrevipes basidiomata also

occur independently from rhizomorphs as well as branches from rhizomorphs,

and its rhizomorphs are also thick and form an aerial network as well as on

the substrate surface. Despite these similarities, molecular evidence shows

Pa. austrobrevipes and G. neobrevipes to be only distantly related (see Fic. 1).

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 19

The surviving culture of TENN-F-053181 (TFB 3585) formed considerable

aerial mycelium and what appears to be resupinate rhizomorphs against the

side of the MEA-slanted storage test tube. This is different from other cultures

of the same putative species but identical to a culture of TFB 3966, of which

voucher basidiomata are no longer extant.

Collections of Pa. austrobrevipes may be mistaken in the field for

M. crinis-equi. The latter can be distinguished in the field by collariate lamellae,

significantly smaller basidiomata with short, curved stipes and typical

hymeniform pileipellis and cheilocystidia. Grgurinovic (1997) cited collections

of M. crinis-equi from New South Wales and South Australia, and Pegler (1977)

furnished a much wider range for the species.

Close inspection of rhizomorph surfaces shows that when rhizomorphs

branch, whether to produce a basidiome or a sterile branch, there is often a

rupture of the parent rhizomorph cortex—splitting the cortex and revealing

the medulla. This rupture is sparsely covered by a brown mycelium, which also

covers the basal portion (<50 um) of the new branch. Whether sterile or fertile,

the new branch is also vestured with caulocystidia, which peter out in less than

a millimeter distally, leaving the greater portion of the rhizomorph branch (or

stipe) subglabrous.

On whole wheat flour agar (Farnett & al. 1999), cultures of Pa. austro-

brevipes produced chiefly submerged mycelium, no textura intricata, and no

rhizomorphs. TFB 3591 (TENN-F-053146) produced a minutely granular

white mycelial mat with ill-defined margins.

Text and illustrations of Paramycetinis species were generously pre-reviewed

by Australian and New Zealand mycologists in order to avoid nomenclatural

duplication of any taxon already described. While no present-day match was

suggested (but see below), five historical names under Marasmius were pointed

out, as follows:

Marasmius eucalypti Berk., in Hooker, Fl. Tasman. 2: 249, 1859 [“1860”].

Pegler (1965: 331) examined the type specimen. While a basidioma

is illustrated as arising as a side branch of a rhizomorph, pileipellis was

described as “hymeniform, composed of diverticulate elements similar to the

cheilocystidia.” Thick-walled caulocystidia are described and illustrated. All of

this indicates identification as a Marasmius, probably M. sect. Sicci.

Marasmius meloniformis Berk., in Hooker, Fl. Tasman. 2: 249, 1859 [“1860”].

Pegler’s (1965: 339) study of the type specimen described surviving material

in poor condition. The “long, setaceous stipes, which arise as branches from

20 ... Petersen & Hughes

creeping rhizomorphs” must resemble the habit of Paramycetinis austrobrevipes.

Conversely, the following characters do not match: 1) pileus 1-2 mm broad,

strongly plicate; 2) lamellae few; 3) epicutis hymeniform; 4) cheilocystidia

broom cell-like. Pegler (1965) concluded: “The species clearly belongs in the

genus Marasmius, sect. Marasmius.’

Marasmius subsupinus Berk., in Hooker, Fl. Tasman. 2: 249, 1859 [“1860”].

From Berkeley's description the following is inferred: Basidiomata

apparently conchate with a very short, curved stipe. Lamellae “so thick and

rigid that this pretty species might almost be placed in Lentinus” “It varies in

color from nearly white to rufous.’

Marasmius emergens Cooke, Handb. Austral. Fungi: 88, 1892.

Protologue: “Very minute, white, suberumpent; pileus convex (1 mm broad);

stem abbreviated, or sometimes elongated, curved, ascending; gills distant, few,

white. On wood. Tasmania.”

Marasmius subroseus Cooke & Massee, Grevillea 21: 37, 1892.

The protologue implies the following: basidiomata caespitose; no mention of

rhizomorphs; “stem becoming a little reddish downwards and clad at the base

with white pubescence;” gills distant. Unless proven otherwise, the organism

can be accepted as a Marasmius.

In addition to these names, an undescribed Australian species is well-known

under the informal name of Marasmius “angina.” Online, numerous photos

and one description (http://www.elfram.com/fungi/fungi_l/marspang_a.html;

https://www.bushheritage.org.au/blog/fungi-and-citizen-science-in-the-liffey-

valley) show an unmistakable resemblance to Paramycetinis austrobrevipes,

except that the basidiomata are distinctly larger and no rhizomorphs appear in

any photo. There is no reason (yet) to consider Marasmius “angina” as anything

but a true Marasmius.

ADDITIONAL SPECIMENS EXAMINED: AUSTRALIA, TasMANIA, Cradle Mountain

National Park, Waldheim Nature Track, 31.V.1991, coll. RHP & KWH, TFB 3977

(TENN-F-053232); Geeveston, Tahune Forest Preserve, 5.VI.1991, coll. RHP & KWH,

TFB 4008 (TENN-F-050195);Gordon-Pedder National Heritage Area, Rainforest Nature

walk vicinity, 8.VI.1991, coll. RHP & KWH, TFB 4038 (TENN-F-050212); Hobart, Mt.

Wellington, gully at 400 m elev., 25.V.1991, coll. KWH, TFB 3591 (TENN-F-053146);

coll. RHP & KWH, TFB 3585 (TENN-F-053181); Lake Pedder National Heritage Area,

26.V.1991, coll. RHP & KWH, TFB 3917 (TENN-F-050119); Lake St. Clair National

Park, Mt. Rufous track, 29.V.1991, coll. RHP & KWH, TFB 3945 (TENN-F-050073);

8-10 kms north of Rosebery, slopes of Mt. Murchison, 30.V.1991, coll. RHP & KWH,

TFB 3949 (TENN-F-051333); coll. RHP, TFB 3966 (TENN-F-053231).

Paramycetinis & Pseudomarasmius gen. & spp. nov.... 21

Fic. 9. Paramycetinis caulocystidiatus (TENN-F-053721).

Basidiomata and rhizomorphs on natural substrate.

Paramycetinis caulocystidiatus R.H. Petersen, sp. nov. Fics 9-14

IF 555794

Differs from Paramycetinis austrobrevipes by its 1) pileipellis with evidence of slime

matrix; 2) high stipe length to pileus breadth ratio; 3) fruiting habit on black beech; and

4) gregarious (but separate) setoid caulocystidia.

Type: New Zealand, North Island, Urewera National Park, track to Waipai Swamp,

35°45'13”S 177°09'17”E, 27.V.1994, coll. RHP, TFB 7148 (Holotype TENN-F-054050).

EryMo toey: referring to caulocystidia of the vestured stipe.

BASIDIOMATA (FIG. 9) slender, of medium stature, marasmioid. PILEUS 2-8

(-20) mm broad, convex at first, later campanulate to applanate with depressed

disc and margin remaining downturned, minutely tuberculate, translucent-

striate to sulcate at margin, surface moist, hygrophanous, glabrous; central dot

8E6 (“Natal brown”), 6D6-6D5 (“snuff brown” near “olive brown”), 5D6-5D5

(“buckthorn brown”), “hazel brown, 6D8-6D4 (“buffy brown”), fading to

5C5-5C4 (“tawny olive”); limb and margin 7C4 (“wood brown”) to 6C5 (“sayal

22 ... Petersen & Hughes

brown”). LAMELLAE adnexed, suppressed and ridge-like to subventricose,

subdistant, <2 mm broad, thickish, total lamellae = 35-50(-75), through

lamellae = 10-17 per pileus with 1-2 ranks of lamellulae between each through-

lamella, not forked or interveined, 4A2 (“pale ochraceous buff”), 5A2 (“pale

cinnamon pink”), 9B2 (“vinaceous buff”), to 5B8 pastel burnt orange to near

6B3 (“tilleul buff” not white); edge entire, rounded (not sharp), not fimbriate,

not marginate. StrpE 30-40(-75) x 1.3-2 mm, terete, equal (stipe base often

slightly swollen), not insititious; surface moist to dry, pruinose, uniformly

vestured as a turf of perpendicular setoid caulocystidia, upward 7C4 (“wood

brown’) to 7D5 (“Rood’s brown”), downward 7E6 (“auburn”), 7E6 (“VanDyke

brown”), 7F5 (“blackish brown (1)”) to black; “sterile stipes” often present,

20-32 x 1-2.5 mm, straight, stiff, vestured, without pileus, occasionally

decapitated (Fic. 9) and then producing rhizomorph extension (black, slender,

glabrous). RHIZOMORPHS <30 x 0.2-0.7 mm (at base), glabrous, gradually

narrowing to flagelliform, mostly black, only slightly paler distally, when

decapitated producing a rhizomorph or stipe (glabrous versus vestured)

extension, but not producing pileate basidiomata from rhizomorphs. ODoR

negligible; TAsTE negligible.

HABITAT AND PHENOLOGY: On dead twigs, leaves and small branches of

Nothofagus solandri; North Island, New Zealand; May-June.

PILEIPELLIS a repent layer of generally radially oriented hyphae embedded

in a thin slime matrix, tightly packed, of the following types: 1) pileal hairs

(Frc. 10) <100 x 3-5 um, perhaps erect, produced as termini of repent hyphae

(Fre. 11) finely encrusted, usually subtly capitulate; 2) repent hyphae 2-8(-13)

um diam, at surface usually of the wider type, firm-walled, coarsely encrusted

in thick scabs; profile calluses <2 um thick, sub-refringent (PhC); crust

material often distributed with suggestion of striped or annular thickenings;

3) repent hyphae 5-10 um diam with gelatinizing walls (wall <2.5 um thick),

variously encrusted; 4) subpellis hyphae usually thicker (6-14 um diam),

smooth or with very fine encrustation appearing gritty on the hyphal surface;

clamp connections ubiquitous. PLEUROCyYsTIDIA (Fic. 12) 24-32 x 6-9 um,

common, broadly fusiform, conspicuously clamped; contents homogeneous

to distinctly partitioned. Basip1a 25-30 x 7-11 um, clavate, often sub-

urniform, 4-sterigmate, clamped; effete basidia (and pleurocystidia) do not

disappear but leave residual lateral walls and shriveled sterigmata (“husking”).

BASIDIOSPORES (5-)6.5-8(-9) x 3.5-4(-4.5) um (Q = 1.43-2.43; Q™ = 1.92;

L™ = 7.41 um), ellipsoid, somewhat flattened adaxially, smooth, thin-walled,

inamyloid. CHEILOocystTipIA (Fic. 13) hardly distinguishable from basidia,

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 23

Fic. 10. Paramycetinis caulocystidiatus (TENN-F-054050).

Pileal hairs. A. Origin of two pileal hairs. B,C. Individual pileal hairs. Scale bars: 10 um.

29-35(-42) x 6-13 um, broadly truncate-clavate, hardly lobed apically,

obscurely clamped. STIPE MEDULLARY HYPHAE 3-9 um diam, strictly parallel,

adherent with minimal slime matrix, thin-walled inward, with gelatinizing walls

(<2.5 um thick) near cortex, obscurely clamped, hyaline, inamyloid inward,

increasingly dextrinoid toward stipe surface (to moderately so in Melzer’s

reagent + BF). STIPE CORTICAL HYPHAE Strictly parallel, probably coherent and

perhaps with thin slime layer, coarsely encrusted, thick-walled, yellow-brown

24 ... Petersen & Hughes

Fic. 11. Paramycetinis caulocystidiatus (TENN-F-054050). Pileipellis elements. A. Heavily

encrusted hypha with gelatinized wall. B,C. Encrusted hyphae with suggestion of striped or annular

ornamentation. D. Hyphae with gelatinized walls. Note anastomosis in “H”-connection. E. Swollen

subpellis hypha. Scale bars: 10 um.

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 25

Fic. 12. Paramycetinis caulocystidiatus (TENN-F-054050). Hymenial elements.

A-D. Pleurocystidia. B. Note partitioned contents. E-H. Basidia. Scale bars: 10 um.

(PhC), dark red-brown in Melzer’s reagent + BE. CauLocystip1a (Fic. 14)

arising as side branches from thick-walled, smooth, superficial cortical hyphae,

stiff, setoid, arising as short lobes (and then hyaline), elongating >70 x 6-9 um

at base, 3-4.5 um diam at apex, narrowly rounded, thick-walled (wall <3 um

thick when visible), sub-refringent and hyaline distally (150x + PhC), strongly

dextrinoid (red-brown in Melzer’s reagent + BF; dark red-brown in Melzer’s

reagent + PhC).

26 ... Petersen & Hughes

Fic. 13. Paramycetinis caulocystidiatus (TENN-F-054050). Cheilocystidia.

A. Grouped at lamellar edge. B-E. Individual cheilocystidia. Scale bars: 10 um.

COMMENTARY: Pileipellis structure is typical of Gymnopus sect. Perforantia.

Hyphal types match quite well, especially gelatinizing hyphal walls and

presence of pileal hairs, and there is evidence of a thin slime matrix. Stipe to

pileus ratio is high, with the stipe universally vestured and minutely barbed.

Dried pileus now somewhat campanulate, with umbo slightly darker than

limb or margin; pileus now appearing somewhat moist-laccate. Basidiomata

are accompanied by thin, ascendant rhizomorphs. The “omnipotent” stipes

and rhizomorphs are somewhat unique, and are quite obvious as abrupt, odd,

asymmetrical joints of such structures (see Fie. 9).

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 27

Fic. 14. Paramycetinis caulocystidiatus (TENN-F-054050). Caulocystidia.

A. Overview of stipe surface; note gregarious but separate individual caulocystidia.

B-E. Individual caulocystidia. Scale bars: A = 20 um; B-E = 10 um.

Paramycetinis caulocystidiatus joins Pa. austrobrevipes in a small clade

sister to that of Mycetinis (Fic. 1). The latter is characterized by a roughly

hymeniform pileipellis of inflated, naked, thin-walled cells and usually a

basidiomatal alliaceous odor and taste. Wilson & Desjardin (2005) showed that

28 ... Petersen & Hughes

Mycetinis taxa were found embedded among taxa of Gymnopus s.l. and assorted

other genera. Petersen & Hughes (2017b) surveyed Mycetinis, with a detailed

ITS-based phylogeny of the genus.

On whole wheat flour agar (Farnet & al. 1999), cultures of Pa. caulocystidiatus

produced no textura intricata, white to off-white felty mycelium, and no

rhizomorphs.

ADDITIONAL SPECIMENS EXAMINED: NEW ZEALAND, NortH ISLAND, Urewera

National Park, Lake Waikaremoana, Black Beech Track, 28.V.1994, coll. A.S. Methven,

TFB 7588 (TENN-F-053721); Lake Waikareiti Track, 27.V.1994, coll. A.S. Methven, TFB

7572 (TENN-F-053683).

Pseudomarasmius

Oliveira & al. (2019) resolved a clade they named /pallidocephalus (not

a genus name). It is this clade that we propose as Pseudomarasmius. They

encouraged generic status for the clade which, they admitted, needed more

research.

Oliveira & al. (2019) proposed Pusillomyces, closely related to Gymnopus

asetosus and G. funalis (their clade B). Pusillomyces manuripioides (typus

generis) was described as lacking clamp connections, a trait virtually

diagnostic for Pseudomarasmius, proposed here, but relatively unrelated to

their clade B.

A PhyML analysis of nrITS sequences for species within Pseudomarasmius

is given in Fic. 15 and broader placement based on the nrLSU region is given

in Fic. 1. Pseudomarasmius comprises several species and an unidentified

clade of fungal soil isolates from Guyana.

Pseudomarasmius R.H. Petersen & K.W. Hughes, gen. nov.

IF 317324

Differs from Marasmius by 1) diverticulate hyphae present in the pileipellis and

2) clamp connections absent.

TYPE SPECIES: Pseudomarasmius pallidocephalus (Gilliam) R.H. Petersen

Erymo .ocy: Referring to its similarity to Marasmius.

Basidiomata of three types: 1) similar in size and stature to those of

Gymnopus sect. Androsacei, with discrete, bristle-like stipe and convex to

applanate cap; rhizomorphs slender and inconspicuous limited to rotting

leaf or needle litter; 2) similar in size and shape to those of Marasmiellus,

with small, curved stipe and shell-like pileus and usually dominant

rhizomorphs which occasionally produce basidiomata as side branches,

similar to this phenomenon in Marasmius crinis-equi; and 3) small,

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 29

slender, pallid yellowish mycenoid basidiomata lacking rhizomorphs,

on rotting needles of Abies or Picea. Microstructures resembling those

of Gymnopus sect. Androsacei and Marasmius sect. Sicci, including:

1) diverticulate hyphae in pileipellis (arranged in a “rameales-structure”);

2) thick-walled, gelatinizing hyphae and/or gelatinizing diverticulate

hyphae in pileipellis and subpellis; 3) without clamp connections or

clamps rare and restricted to pileus trama; 4) basidia (2—)-4-spored;

5) spores similar to those of other gymnopoid genera, ellipsoid, thin-

walled, inamyloid; 6) molecular (LSU, ITS) profiles unique, grouping

into a monophyletic clade related to Rhodocollybia but more distant from

groups exhibiting similar basidiomatal types (Fic. 15).

Key to species of Pseudomarasmius

1. Ratio of pileus breadth to stipe length <2 (“thumb-tack shaped”);

basidiomata small, often arising as side branches of rhizomorphs............ 2

1. Ratio of pileus breadth to stipe length >4; basidiomata marasmioid or mycenoid,

arising as rhizomorph branch or independently on leaf litter or woody

ULES TAT sacks se cchcrcbe-lehoecls AT iel hc rencechcn ces ehcea are a Aries kar 3

2. Southern South America (Isla Chiloé, Chile); rhizomorphs inconspicuous,

not producing basidiomata; fruiting on hardwood substrates;

clamp connections absent; superficially inseparable from

OS PTET SSAV ESN cee itt cnc Ride sala ade oa: Faults se desoy Pagans sme Retded «eRe oh Ps. patagonianus

2. North American Gulf Coastal Plain and subtropical eastern Mexico; rhizomorphs

dominant, long, occasionally branched, occasionally producing basidiomata

as side branches; fruiting on small dead branches and twigs of deciduous trees

usually at some distance from the ground; clamp connections occasional in

rhizomorphs and tramae but otherwise absent ................. Ps. nidus-avis

3. Antipodal (New Zealand); basidiomata minute (pileus 1-3.5 mm broad; stipe

capillary, 6-8 mm long); fruiting on hanging dead terminal branches of

DDGCT VATE Sere! PS ok AMT ae OE ae PTR, AAR soo BTA! ASU MRO Ps. efibulatus

3. Not Antipodal; fruiting substrate leaf or needle litter ......................00.. +

4. Fruiting on conifer needle litter; rhizomorphs usually inconspicuous............ 5

4, Fruiting on hardwood leaf litter (including Castanopsis); rhizomorphs conspicuous,

PESUPINIALE ON Cal SUT LACES od Fe ntl (Oe ahd cae Phone APG eaeead Cai ceee eaten Pte eT shed 7

5. Temperate to northern North America; stipe black; basidiomata like those of

Gymnopus sect. Androsacei (pileus <35 mm broad; stipe 12-43 x 0.2-0.8 mm);

pileus white to off-white; rhizomorphs inconspicuous, usually found deep in

ITUETIRLAS. >. «Sta h'p. «sha Nip tea Mi rata Wie ada bis fost.g be asta iret abe posi anit Ps. pallidocephalus

5. South Korea/eastern North America; basidiomata slender, mycenoid; stipe

Ochtaceous LOrsthawaCOloreds hF 8 Fi ut Atal tna ali sa ha tants a ello arwsalt rk wall tak 6

30 ... Petersen & Hughes

KC966049 Alaska

AY313287 Greenland afn. Rhodocollybia

FJ475743 Soil Sweden

MK268235 China Ps. guercophiloides

KF251072 G. glabrocystidiatus (not type)

JN890249

Soil samples

JN89021 9 Guyana

JN890248

KY352649 Ps. patagonianus Chile

MK268234 NZ

MK268236 NZ

KR673444 Marasmius sp. Korea

| Ps. efibulatus

FJ596763 USA, TN

KY026691 USA, TN

KY026636 USA, WA

KY026635 USA, ID

FJ596762 USA, TN

5015 culture, Canada, NS

KY026684 USA, NY

— Korea G. glabrocystidiatus type

KY026732 USA, AL

MK268237 USA, FL

KY026723 USA, MS

KY026646 USA, LA

KY026733 USA, TX

MH016872 USA, FL

KY026753 USA, MS

MK020094 USA, FL

MK268238 USA, FL

MH560575 Mexico

MH560576 Mexico

MH560577 Mexico

MHS60579 Mexico

MIH560578 Mexico

Ps. pallidocephalus

Pseudomarasmius

0.05 changes

8 3

Ps. nitus-avis

Fic. 15. Maximum likelihood analysis of ribosomal RNA ITS sequences. Bootstrap values are given

to the left of the supported node. Numbers are GenBank accession numbers (See TABLE 1). For

collections from the United States and Canada, the state or province postal code is given. NZ =

New Zealand.

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 31

6. South Korea (and probably Japan); basidiomata minute, fragile

(pileus 48 mm broad; stipe 15-40 x 0.5 mm); pileus dark grey-green;

stipe dark brown to black-brown; rhizomorphs inconspicuous;

fruiting onideadmeedles OPA MICS. feiss. 8 ie eth, cok ented Ps. glabrocystidiatus

6. Eastern North America; basidiomata similar to those of Ps. glabrocystidiatus;

pileus light brown or pale greyish brown; stipe brownish orange to yellow

or straw-colored; on rotting needles of Picea and Abies ......... Ps, straminipes

7. Costa Rica; high-altitude Quercus forests ......... 0... cece eee eee Ps. obscurus

7. South Korea, China (Yunnan); on dead leaves of Castanopsis ... Ps. quercophiloides

Pseudomarasmius efibulatus R.H. Petersen, sp. nov. FIGs 16-20

IF 555730

Differs from Pseudomarasmius taxa by its: 1) minute basidiomata; 2) fruiting habit on

dead Dacrydium branchlets; 3) scattered pyramidal to digitate diverticula on pileipellis

elements; 4) known distribution limited to the New Zealand type location.

Type: New Zealand, South Island, Westland, Fox Glacier, Lake Matheson loop trail,

43°26'22”S 169°57'54’E 13.V.1994, coll. RHP, TFB 7070 (TENN-F-056187).

EryMo_oey: from fibula (Latin) for clamp, buckle + e- (Latin), without. Referring to the

clampless condition.

BASIDIOMATA (FIG. 16A) diminutive. PrLEus 1-3.5 mm broad, convex when

young, becoming everted by maturity, matte, pebbled; disc 7C5 (“fawn color’),

outward 9B2 (“vinaceous buff”) to 6A3 (“pinkish buff”). LAMELLAE adnate,

arcuate, not collariate, thickish, c. 1 mm broad, not marginate, off-white when

fresh, becoming 5A4 (“light ochraceous buff”) over time; lamellulae occasional,

short; total lamellae (10)1218; through lamellae 7-8. Stipe 6-8 x 0.2-0.4

mm, apically 8E6 (“Natal brown”) (and this color extending through lamellar

attachment to pileus flesh), lower 7/8 black, glabrous-shining, insititious.

RHIZOMORPHS (Fic. 164,17) common, <40 x 0.1-0.3 mm, more or less straight

(not curly), rarely branched, arising separately from basidiomata.

HABITAT AND PHENOLOGY: On dead, pendant Dacrydium foliage;

Autumn.

PILEIPELLIS with minimal slime matrix and composed of the following:

1) diverticulate hyphae (Fic. 18A-p) 4.5-10 um diam, thin-walled, with

scattered diverticula <10 x 1.52 um (often c. 4 um diam at base and therefore

more or less pyramidal), often dichotomous or saddle-shaped, not refringent

[in fact appearing darker than the parent hyphae in PhC]; 2) broom cell-

like termini (Fic. 18E-H) 15-25 x 9-16 um, stalked, usually dichotomously

branched, surmounted with rows of diverticula; diverticula 3-8 x 1-1.5 um,

hardly refringent (PhC). Pileus tramal hyphae 3-6 um diam, firm-walled,

long-celled, apparently without clamp connections; wall somewhat obscure,

32 ... Petersen & Hughes

Fic. 16. Pseudomarasmius efibulatus (TENN-F-056187, holotype).

A. Basidiomata and rhizomorphs. B. Basidiospores. Scale bars: A= 5 mm; B = 5 um.

as though semi-gelatinized. Lamellar tramal hyphae similar. Hymenium

apparently involved in minimal gelatinized matrix (excessive hymenial

remnants and other debris). PLEUROCYSTIDIA (FIG. 19A—D) 24-30 x 6-7 um,

fusiform, often irregularly so, without content partitioning or clamp

connection. Basidioles clavate, usually subcapitulate; Bastp1A (FIG. 19E-H)

22-30 x 6-9 um, clavate, 4-sterigmate, without clamp connections.

BASIDIOSPORES (FIG. 16B) (7—)7.5-11 x 3.5-4.5 um (Q = 2.053.14; Q™ = 2.26;

L™ = 8.67 um), elongate-ellipsoid to subcylindrical, more or less marasmioid

(somewhat tapered proximally), flattened somewhat adaxially, thin-walled,

smooth, inamyloid. Lamellar edge apparently fertile. CHEILOCYSTIDIA (FIG.

20) widely scattered, 22-35 x 10-12 um, stalked-clavate to saccate, entire

or with a few slender, digitate apical processes, without clamp connections.

STIPE MEDULLARY HYPHAE 1.5-12 um diam, strictly parallel, involved in a

slime matrix, thin- to thick-walled (wall c. 0.7 um thick, hyaline), distinctly

without clamp connections, relatively short-celled; subcortical hyphae 4-8

um diam, thick-walled (wall c. 1.5 um thick), strongly refringent, yellow

(PhC). STIPE CORTICAL HYPHAE 4-7 um diam, firm-walled, short-celled,

without clamp connections, pigmented yellow-olive (PhC); surface minutely

roughened with dried thin slime. CAuLocysT1p1A absent.

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 33

Fic. 17. Pseudomarasmius efibulatus (TENN-F-056187, holotype).

Dacrydium branchlets with rhizomorphs. Scale bar = 10 mm.

COMMENTARY: Based on ribosomal nrRNA ITS sequences, Ps. efibulatus

is related to but distinct from Ps. pallidocephalus, Ps. glabrocystidiatus, and

Ps. nidus-avis. It is closely related to sequence JAC10695 received from

J. Cooper, Landcare Research, New Zealand (Fic. 15)

A blast of ITS sequence from Ps. efibulatus (TFB 7070, TENN-F-056187)

showed an affinity with ITS sequences of Ps. glabrocystidiatus, a clampless

taxon from South Korea, placed by its authors in Gymnopus sect. Androsacei.

Sequences of Ps. glabrocystidiatus are virtually congruent to those of

Ps. pallidocephalus (see above) found frequently across northern North

America fruiting on dead conifer needles, also the habit of the Asian

G. glabrocystidiatus. The nrITS-based phylogeny produced by Antonin

& al. (2014) placed G. glabrocystidiatus within Gymnopus but on a long,

independent branch described as part of G. sect. Androsacei. The nrLSU-

based phylogeny in Fic. 1 (above) indicates even more significant isolation

of Pseudomarasmius.

Pseudomarasmius efibulatus belongs to a small complex of similar, slender,

gracile basidiomata along with Ps. straminipes and Ps. glabrocystidiatus, but

the basidiomata of all three species are so small and inconspicuous as to be

34 ... Petersen & Hughes

Fic. 18. Pseudomarasmius efibulatus (TENN-F-056187, holotype). Pileipellis elements.

A-D. Diverticulate hyphae. E-H. Broom cell-like hyphal termini. Scale bars: 10 um.

easily overlooked. Pseudomarasmius efibulatus, however, is quite isolated in

substrate and distribution.

The few similarities between Ps. pallidocephalus and Ps. efibulatus include:

1) clampless condition; 2) temperate distribution (North- and South-

Temperate climates); 3) stipe without caulocystidial ornamentation; 4) a

complex “ramealis-structure” of pileipellis; 5) cheilocystidia that are usually

smooth, broadly clavate or ampulliform; and 6) well-developed rhizomorphs.

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 35

Fic. 19. Pseudomarasmius efibulatus (TENN-F-056187, holotype). Hymenial elements.

A-D. Pleurocystidia. E-H. Basidia. Scale bars: 10 um.

Basidiospores are smooth, ellipsoid to elongate pip-shaped, and white in

spore deposits throughout the /gymnopus/marasmiellus clade, with spore

measurements distinguishing the taxa reported as follows: Ps. efibulatus =

(7-)7.5-11 x 3.5-4.5 um (Q = 2.26); G. glabrocystidiatus = (7.7—)8.5-9.5

36 ... Petersen & Hughes

Fic. 20. Pseudomarasmius efibulatus (TENN-F-056187, holotype).

Cheilocystidia. Scale bars: 10 um.

(-10) x 4-5 um (Q = 2); Ps. pallidocephalus = (5-)5.5-7(-9.5) x 3-3.5 um

(QO. = 1,87).

Macroscopically, basidiomata of Ps. nidus-avis and Ps. patagonianus,

superficially resemble numerous Marasmiellus taxa fruiting on dead twigs

and small branches of deciduous trees and exhibiting extensive, robust

rhizomorphs. Although morphologically counterintuitive for inclusion in

Pseudomarasmius, ITS- and LSU-based phylogenies place Ps. nidus-avis and

Ps. patagonianus with the other taxa in the genus.

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 37

Perhaps most surprising about the sole collection of Ps. efibulatus was

the absence of clamp connections. Collections of similar basidiomata found

fruiting on the same somewhat unique substrate demonstrated ubiquitous

clamp connections. Moreover, pileipellis elements of clamped and clampless

organisms compare well: a complex pileipellis composed of both repent

diverticulate hyphae (with diverticula often bifurcate) and broom cell-like

hyphal termini beset with diverticula also often dichotomous. This may be

analogous to the situation in Ps. straminipes (clampless) and its form fibulatus

(clamp connections ubiquitous). DNA sequences not currently extant may

help to clarify this situation.

Pseudomarasmius glabrocystidiatus (Antonin, Ryoo & Ka) R.H. Petersen,

comb. nov. FIGS 16-20

IF 555747

= Gymnopus glabrocystidiatus Antonin, Ryoo & Ka, Mycol. Prog. 13(3):

710. 2014. Republic of Korea, Inje, Baekdam-sa, 27. VI. 2008 leg. V.

Antonin (08.36), R. Ryoo & J.G. Han (Holotype, BRNM 718676).

ADDITIONAL PUTATIVE CONTAXIC COLLECTION (but see below): Culture. Korea, Jeju,

Seonheul-gotjawal, 06.[X.2012, R. Ryoo, K.-H. Ka & H.D. Sou (KFRI 1935).

For detailed description and illustrations see Antonin & al. (2014)

Major distinguishing characters include: 1) basidiomata similar to those of

Ps. straminipes; 2) clamp connections absent; 3) pileus dark (grey-)brown

with paler margin when young, then brownish at center and paler, beige to

pale brown at margin; 4) basidia 2- spored; 5) stipe smooth and glabrous,

dark brown to black-brown; 6) cheilocystidia smooth, clavate or pyriform;

7) pileipellis composed of more or less smooth or diverticulate hyphae and

single broom cells; 8) habitat on dead conifer (Abies) needles; 9) geographic

distribution, South Korea (probably Japan).

Hapsitat: needles of Abies holophylla.

COMMENTARY: In the phylogeny presented by Antonin & al. (2014), two

ITS sequences of putative Gymnopus glabrocystidiatus (GB NR152899,

holotype; KF251072, ancillary sequence from culture) appeared in a

single small clade but on individual short branches. This clade appears

sister to Rhodocollybia, a position similar to that shown on our LSU-based

phylogeny (Fic. 1) for Pseudomarasmius. An ITS sequence from holotype

material of G. glabrocystidiatus differs from those of P. pallidocephalus by

31/694bp (4.46%) and while apparently in the same clade based on nrITS

sequences alone (Fic. 15), nrITS sequence divergences warrant separate

species designations. Further, G. glabrocystidiatus is an Asia taxon while

38 ... Petersen & Hughes

Ps. pallidocephalus is North American in distribution. The description and

illustrations of Ps. glabrocystidiatus furnished by Antonin & al. (2014) are

adequate and require no enhancement here.

Although two GenBank designations for ITS sequences of G. glabro-

cystidiatus were reported in the phylogeny by Antonin & al. (2014),

KP251073 (“NR152898” from holotype material) and KP251072, the

latter can be traced to a culture from an ancillary specimen apparently

misidentified. The two sequences appear only distantly related in our

ITS-based phylogeny (Fie. 15).

The non-type GenBank sequence (KF251072) virtually matches that of

TFB 3162, here proposed as Ps. quercophiloides (q.v.). The latter was collected

in China.

GenBank sequence LC0148839, including the ITS region, was derived from

an environmental sample from leaf litter of Picea jezoensis subsp. hondoensis

(spruce) in Funabashi, Japan (Hagiwara & al. 2015). The sampling site

was in central Honshu Island (35 36’N, 13728’E) and dealt with bleaching

of substrate leaf debris, including conifer (Hintikka 1970; Miyamoto &

al. 2000; Osono 2015). The spruce substrate generally agrees with that of

Ps. glabrocystidiatus and, with the latter distribution in South Korea (Antonin

& al. 2014), the Japanese report seems within the ecological pattern for the

complex.

Pseudomarasmius nidus-avis (César, Bandala & Montoya) R.H. Petersen,

comb. nov. FIGS 21-31

IF 555746

= Gymnopus nidus-avis César, Bandala & Montoya, Mycokeys 43: 25. 2018

Type: Mexico. Veracruz: Municipality of Xalapa, Santuario del Bosque de Niebla,

Instituto de Ecologia A.C., 1343 m a.s.l., gregarious, on fallen twigs of Quercus, 20 April

2016, Cesar 36 (Holotype, XAL).

Diagnostic characters include 1) basidiomata that are small and thumb tack-shaped,

often arising as side branches from rhizomorphs; 2) a short, curved stipe; 3) distribution

in North American Gulf Coast and subtropical Mexico; 4) clamp connections lacking

except rare in stipe medulla.

BASIDIOMATA (Fics 21, 22) arising from woody substrate or (more often)

as side branches of rhizomorphs. PILEus 2-6(2-10) mm broad, convex

to plano-convex, usually somewhat depressed over the disc, occasionally

shallowly umbonate, pulvinate, variously sulcate-striate, matt, uniformly

more or less “wood brown” (7C4), pale brown (7.5 YR 7/4-6; 10YR 7/4;

Munsell) to brown (7.5 YR 5/6; 10 YR 4/4; Munsell); context thin (<1 mm

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 39

Fic. 21. Pseudomarasmius nidus-avis (TENN-F-054912). Basidiomata.

A. Habit photo. B. Basidiomata, macromorphological aspect. Scale bars: 10 mm.

thick), soft, whitish. LAMELLAE (<1 mm broad; 13-26 total lamellae, 4-9

through lamellae, anastomoses lacking), adnate, adnexed to shallowly

subdecurrent, thick with rounded edge, paler than pileus surface, ivory to

off-white when fresh, very pale brown (2.5 Y 8/2; Munsell)], after drying

suffused ruddy brownish outward from stipe (now “wood brown” 7C4, or

“avellaneous” 7B3); lamellar edge entire. STIPE arising either from woody

substratum or as side branches of rhizomorphs; 2—6(-12) x (0.2-)0.7(-1.6)

mm, central to slightly eccentric, terete, equal or tapered downward, black,

glabrous-shining, straight to strongly curved, insititious, reddish-brown

at the apex (2.5 YR 4/6; Munsell), dark brown to black below (1OYR 2/1,

7.5YR 2.5/2; Munsell), brown; medulla light brown (2.5Y 6/4; Munsell;

AO ... Petersen & Hughes

Fic. 22. Pseudomarasmius nidus-avis (TENN-F-047666). Basidiomata.

Scale bars: 10 mm. Photos courtesy David P. Lewis.

basal tuft very sparse). RHIZOMORPHS (FIG. 21, 224,B) <500 x 0.1-0.3 mm,

simple, black, wiry, abundant, black, glabrous-shining. TasTE and opor not

distinctive.

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 41

Fic. 23. Pseudomarasmius nidus-avis (TENN-F-054912). Micromorphology.

A. Pileipellis diverticulate hyphae. B. Pleurocystidium and basidia.

C. Basidiospores. D. Cheilocystidia. Bars: A-C = 10 um; D = 5 um.

HABITAT & PHENOLOGY: Subtropical forest, with dominant rhizomorphs

scattered or gregarious on fallen or aerial twigs of trees, notably Quercus,

with rhizomorphs used by birds as nesting material in Mexico; late spring

and summer; North American Gulf of Mexico Coastal Plain (Alabama,

Florida, Louisiana, Mississippi, Texas).

PILEIPELLIS composed of: 1) pileal hairs (Fic. 244-c) <80 x 2-4 um,

possibly erect, becoming apically subcapitulate, minutely ornamented; 2)

repent diverticulate-ornamented hyphae (Fics 234,25), 2.5-5.5 um diam,

42 ... Petersen & Hughes

Fic. 24. Pseudomarasmius nidus-avis (TENN-F-054912). Micromorphology.

A-C. Pileal hairs. D. Broom cell-like pileipellis hyphal terminus. Bars = 10 um.

thick-walled and diverticulate hyphae termini (Fics 24pD,25); diverticula

1-3 x 1-1.5 um, conical, slightly dextrinoid (fide César & al. 2018); clamp

connections absent; 3) tightly interwoven, repent hyphae 2.5-6 um diam,

thick-walled (wall <1 um thick), ornamented with annular deposits

c. 0.5 um thick (Fic. 264,B); 4) repent hyphae 2-4.5 um diam, thick-walled

(wall <1 um thick) with individual slime sheath. PILEUs TRAMA composed

of interwoven hyphae 2.5-7.5 um diam, weakly dextrinoid (fide César &

al. 2018), individually strongly gelatinized (wall <2.5 um thick, hyaline),

implicitly ornamented with flake-like deposits (seen as carried by individual

slime sheath; Fic. 26c,p); clamp connections absent. Lamellar trama loosely

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 43

Fic. 25. Pseudomarasmius nidus-avis [FLAS-F-60891]. Pileipellis diverticulate hyphae.

Compare with Fics 38-40 (Ps. pallidocephalus). Bars = 10 um.

interwoven; hyphae 2-3.5 um diam, firm-walled, hardly gelatinizing, not

easily disarticulated, inamyloid to weakly dextrinoid (fide César & al. 2018);

clamp connections absent. PLEUROCYSTIDIA (FIG. 23B, 27) common, 24-29

44 ... Petersen & Hughes

Fic. 26. Pseudomarasmius nidus-avis [FLAS-F-60891]. Pileipellis hyphae.

A. Hyphal encrustation in scabs. B. Annular ornamentation. C,D. Individual slime sheaths.

Bars = 10 um.

x (5-)6-7 um, fusiform to plump-fusiform, without clamp connections;

contents multiguttulate; guttules refringent (PhC). Basidioles (Fic. 28a)

clavate becoming subcapitulate, without clamp connections; BASIDIA

(Fic. 28B-pD) (20-)27-32(-41) x (5-)9-11 um, clavate to subcapitulate,

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 45

Fic. 27. Pseudomarasmius nidus-avis [MES 1477 (FLAS)]. Pleurocystidia. Bars = 10 um.

(2-)4-sterigmate, without clamp connections; contents multiguttulate;

guttules refringent (PhC). BastDIOsPORES (FIGS 23C, 28E-H) (7—)8-10.5 x

(3-)4-5.5 um (Q = 1.802.22; Q™ = 1.91; L™ = 9.2 um), ellipsoid to plump

pip-shaped, flattened adaxially, usually somewhat tapered proximally,

46 ... Petersen & Hughes

Fic. 28. Pseudomarasmius nidus-avis [MES 1477 (FLAS)].

A-D. Basidiole and individual basidia. E-H. Basidiospores. Bars = 10 um.

hyaline, thin-walled, inamyloid; contents multiguttulate when mature;

guttules refringent (PhC). CHEILOCysTIDIA (FIGs 23D, 29) 20-39 x 3-8

um, occasional on lamellar edge, hyphal (not inflated), stalked (stalk

2-2.5 um diam, thin-walled), irregularly lobed and/or branched, without

clamp connections; contents homogeneous. STIPE MEDULLARY HYPHAE of

two distinct forms: 1) stout hyphae (Fic. 30), 4-15(-30) um diam, highly

refringent (PhC), apparently aseptate or with occasional cloissons d’retret;

hyphal walls gelatinizing, occasionally strongly so (wall c. 4.5(-12) um thick),

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 47

Fic. 29. Pseudomarasmius nidus-avis [MES 1477 (FLAS)]. Cheilocystidia. Bars = 10 um.

often ornamented with flake-like deposits in vague stripes and carried on

the outside of the gelatinized sheath; and 2) slender hyphae (Fic. 31), 2-4.5

um diam, thick-walled (wall c 0.7 um thick), minutely ornamented, with

occasional but conspicuous clamp connections. STIPE CORTICAL HYPHAE

2.5-7.5 um diam, repent, strictly parallel, strongly pigmented, thick-walled,

minutely encrusted on stipe surface, dextrinoid (red-brown, Melzer’s

reagent + PhC; reddish brown, Melzer’s reagent + BF); pigmented, diver-

ticulate terminal elements 4—19(-—21) x 3-4(-5) um, thin-walled (fide César

& al. 2018).

COMMENTARY: A recent paper (César & al. 2018) described Gymnopus

nidus-avis as clampless (except for scattered clamps in stipe medulla) and

with basidiomata arising as side branches from copious, black rhizomorphs.

48 ... Petersen & Hughes

Fic. 30. Pseudomarasmius nidus-avis [MES 1477 (FLAS)]. Stout stipe medullary hyphae.

A Non-gelatinized hypha. B. Flake-like ornamentation on thin slime sheath.

C. Maximum gelatinization. Bars = 10 um.

ITS sequences from G. nidus-avis were intermixed with sequences deposited

(by our lab) in GenBank as Marasmius brevipes. Independent discovery that

voucher collections of these collections of putative M. brevipes were clampless

confirmed the paraphyletic placement of putative M. brevipes. Typical

Marasmius brevipes (Desjardin & Petersen 1989b) has been transferred

to Gymnopus (as G. neobrevipes; see Petersen & Hughes 2019). Proposal

of G. nidus-avis including several ITS sequences provides a name for the

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 49

Fic. 31. Pseudomarasmius nidus-avis [MES 1477 (FLAS)]. Slender stipe medullary hyphae.

A. Non-gelatinized hypha. B. Flake-like ornamentation on thin slime sheath.

C. Maximum gelatinization plus ornamentation. Bars = 10 um.

non-typical collections of putative M. brevipes. Inherent is a significantly

widened distribution for G. nidus-avis from subtropical Mexico to the North

American Gulf Coast.

A few specimens at FLAS (variously labelled as unidentified or as “aff.

M. crinis-equi”) lack basidiomata but exhibit rhizomorphs of the G. nidus-

avis morphology (black, smooth, very slender).

Murrill (1915) proposed Polymarasmius for basidiomata which arise

as branches of rhizomorphs but noted: “...lamellae adnate to a collar...”

Polymarasmius was typified by Marasmius multiceps Berk. & M.A. Curtis

from Cuba, with the other two species from Belize and Jamaica. Such a

50 ... Petersen & Hughes

diagnosis seems to fit M. crinis-equi of Marasmius subsect. Sicciformes, which

is known from the subtropics of the New World.

SPECIMENS CONSULTED: MEXICO. VERACRUZ, Municipality of Xalapa, Santuario del

Bosque de Niebla, Instituto de Ecologia A.C., 1343 m a.s.l., 18 May 2006, Bandala 4052;

7 July 2016, César 41; 10 Aug 2016, Ramos 682 (all at XAL).

SPECIMENS EXAMINED: UNITED STATES. Atasama, Mobile Co., Univ. South

Alabama, main campus, VII.2014, coll. unknown, det. J.L. Mata, TFB 14607

(TENN-F-69310). FLoripa, Alachua Co., Gainesville, Possum Creek Park, 20.IV.2016,

coll. M.E. Smith [“On suppressed oak understory, fruiting from elevated branches.” ]

MES1477 (FLAS-F-62667: as Marasmius ‘crinis-equi”); Gainesville, Sweetwater

Reserve, 28.VII.18, coll. M.E. Smith, MES3139 (FLAS-F-62668 as Marasmius ‘crinis-

equi’); Gainesville, Rock Creek neighborhood, 1.V.2015, coll. M.E. Smith, MES 884

(FLAS-F-62666; as Marasmius “crinis-equi”); Putnam Co., Ordway-Swisher Biological

Station, near boat ramp of Lake Suggs, 14.VI.2017, coll. R. Healy & al., FLAS-F-60891

as Marasmius cf. brevipes. LOUISIANA, East Feliciana Parish, St. Francisville, James

John Audubon Historic Site, 30°47.84’N 91°18.43’W, 22.V.1997, coll. RHP, TFB 9087

(TENN-F-054912). Mississippi, Harrison Co., Red Creek Wildlife Management Area,

30°40'35”N 88°54’02”W, 11.VII.2014, coll. KWH, TFB 14498 (TENN-F-069189).

TEXAS, Orange Co., Vidor, Virginia Lane, 30°09.24’N 94°00.66W 26.VI.1976, coll.

D.P. Lewis, det. D.E. Desjardin (as M. brevipes), Lewis 276 (TENN-F-047666).

Pseudomarasmius obscurus R.H. Petersen, sp. nov. Figs: 42=35

IF 555731

Differs from Gymnopus pyracanthoides by its 1) marasmioid basidiomata (stipe slender,

black; pileus small, gray, dry); 2) habit on dead, sclerophyllous Quercus leaves; 3) copious

and obvious resupinate rhizomorphs; and 4) lack of clamp connections.

TYPE: Costa Rica, Prov. San José, Jardin de Dota, 3.5 km W of Interamerican Highway

at El Empalme, 9°4’52”N 83°58’28”W, 15.V1.1995, coll. RHP, TFB 7812 (Holotype,

TENN-F-053787).

EryMo_ocy: Referring to the difficulty in observing microscopic structures.

BASIDIOMATA (FIG. 324) diminutive, very slender. PILEus 25 mm broad,

convex at all ages, becoming essentially applanate with downturned margin by

maturity, more or less unicolorous when dry, matte; disc mouse gray; outward

to margin “tilleul buff” (7B2), not at all striate, scalloped. LAMELLAE adnexed

to free, thickish, <1 mm broad, seceding in drying to appear pseudocollariate

when dried, total lamellae 23-28, through lamellae 13-14, off-white; lamellulae

in one rank, often ridge-like. Strrpz 11-22 x 0.3-0.6 mm, insititious, terete

when fresh becoming channeled on drying, appearing superficially glabrous-

shining but under magnification (40x) minutely pruinose upward, apically

dark brown, extending into pseudocollarium, very dark brown downward

to black-brown lower portion. RHIZOMORPHS common, often resupinate on

leaf surface and there anchored by lateral brownish ciliatiform mycelium,

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 91

Fic. 32. Pseudomarasmius obscurus (TENN-F-053787). A. Basidioma and rhizomorphs with inset

showing lamellar attachment. B. Basidioles. C. Pleurocystidium and basidia; D. Basidiospores.

E. Cheilocystidia. F. Caulocystidia. Scale bars: A = 5 mm; B,C,E,F = 20 um; D = 5 um.

<25 mm long where erect, occasionally producing a stellate growth form,

very slender (0.1-0.3 mm diam), glabrous-shining, black, curly, frequently

branched (similar to those of G. pyracanthoides; see Petersen & Hughes 2016).

DISTRIBUTION & PHENOLOGY: At present known from a single collection;

Costa Rica. Fruiting on sclerophyllous Quercus litter with resupinate

rhizomorphs; summer.

52 ... Petersen & Hughes

Fic. 33. Pseudomarasmius obscurus (TENN-F-053787).

Encrusted hyphae of pileipellis. Scale bar = 10 um.

PILEIPELLIS composed of two elements: 1) a tangle of free-form, broadly

diverticulate hyphae (Fie. 34), 3.5-8 um diam, thin-walled, variously lobate

to subsetulose, often septate, without clamp connections; and 2) strongly

encrusted hyphae (Fic. 33) 3-7 um diam, firm- to thick-walled (wall c. 0.7 um

thick, hyaline); crust material in bands or rings with profile calluses c. 1 um

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 53

Fic. 34. Pseudomarasmius obscurus (TENN-F-037987).

Swollen diverticulate hyphae of pileipellis. Scale bars: 10 um.

thick. PLEUROCYSTIDIAL STRUCTURES (FIG. 32c) obscure, 20-28 x 6-8 um,

cylindrical to fusiform, hyaline, thin-walled, without clamp connections.

Basidioles (Fic. 32B) clavate to subampulliform; BAsIDIA (Fic. 32c) 20-30 x

6-8 um, clavate to subcapitulate, 4-sterigmate, without clamp connections.

BASIDIOSPORES (FIG. 32D) 5.5-7 x 3-3.5 um (Q = 1.71-2.33; Q™ = 1.90;

L™ = 6.25 um), ellipsoid to slightly reniform, variable between tapered

proximally and not so, thin-walled, smooth, inamyloid. CHEILOCYSTIDIA

(F1G.32E) very obscure, 16-25 x 3-8 um, cylindrical to clavate, often furcate or

lobate, thin-walled, hyaline, without clamp connections. STIPE MEDULLARY

HYPHAE 4-7 um diam, thick-walled (wall <1 um thick, hyaline to weakly

54 ... Petersen & Hughes

Fic. 35. Pseudomarasmius obscurus (TENN-F-053787).

Caulocystidia. Scale bars: 10 um.

pigmented), strictly parallel, with occasional slender, thin-walled, strangulate-

digitate to branched hyphae, without clamp connections. STIPE CORTICAL

HYPHAE 3.5-7.5 um diam, thick-walled (wall often occluding cell lumen),

heavily pigmented, distal on stipe producing densely scattered caulocystidia.

CAULOCYSTIDIA (Fics 328, 35) limited to distal third of stipe, 22-41 x

8-11 um, variously shaped from pyramidal to furcate to subampulliform,

thick-walled (wall <1.0 um, hyaline), usually with broader base than apex

and usually with narrow hyphal origin.

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 9D

COMMENTARY: Without DNA sequences, accurate placement of TFB

7812 is problematic, but several morphological characters would suggest

Pseudomarasmius. Production of resupinate rhizomorphs is similar to

G. pyracanthoides (Petersen & Hughes 2016), also from Costa Rica, also

fruiting on Quercus leaves and also without clamp connections. The two

differ in pileipellis structure (G. pyracanthoides pileipellis an intricate thatch

of narrow setulose hyphae) and stipe vesture (G. pyracanthoides vesture over

entire stipe and caulocystidia longer).

Pseudomarasmius pallidocephalus (Gilliam) R.H. Petersen,

comb. nov. FIGs 36-42

TF 555745

= Marasmius pallidocephalus Gilliam, 1975 Mycologia 67: 818.

Type: United States, Michigan, Chippewa Co., Gilliam 1165 (MICH, see below).

Diagnostic characters include: Fruiting on dead spruce and fir needles in North America;

2) basidiomata diminutive, exhibiting a white to off-white pileus and off-black, bristle-

like stipe; 3) pileipellis with diverticulate hyphae, free-form hyphal segments and broom

cell-like hyphal termini; 4) caulocystidia occasional, ampulliform.

BASIDIOMATA (FIG. 36A, 37) diminutive, marasmioid; ratio of stipe length

to pileus breadth high (6-8:1). PILEUs 5-24(-35) mm broad, pulvinate to

convex at first, then plano-convex or broadly conic-convex, finally plane to

shallowly concave and often subumbonate or umbilicate, dry or subviscid in

wet weather, dull opaque, pliant or membranous, reviving, smooth, subtly

sulcate-striate, subtuberculate, even or faintly rugulose-striate to disc,

minutely velutinous or matted-fibrillose, dark brown in primordia; disc and

inner limb brown (7D4) when young, fading to light brownish grey (6C3),

soon light yellowish brown (7.3YR/7.0/2.8 Munsell), brown (7E4) to light

brown (7D4) to light greyish brown (7D3) overall, by maturity light grayish

yellowish brown, light pinkish yellowish brown, light yellowish pink 7A2

(“light pinkish cinnamon’), pale orange yellow, 6A2 (“pale pinkish buff”),

light yellowish brown 6B5, 7,3YR/7,0/2.8 (Munsell) (“cinnamon”), dark

brown (Maerz & Paul 16A12) moderate 4.7Y/5.2/4.1 (Munsell) yellowish

brown, fading to greyish orange (6B2); drying about 5A5 (“ochraceous

buff”); outer limb and margin yellowish, at first entire, soon eroded or

crenate, white (Maerz & Paul 9B2), pale orange yellow 6A2 (“pale pinkish

buff”), 4D2 pale yellowish white to off-white to 7B2 (“tilleul buff”), smooth,

brownish grey when young, becoming minutely rugulose-striate, buff in

age — overall pallid coloration in age. Pileus trama thin, yellowish white

(5.5Y/9.3/1.8 Munsell) to light yellowish brown (7.3YR/7.0/2.8 Munsell).

56 ... Petersen & Hughes

Fic. 36. Pseudomarasmius pallidocephalus (TENN-F-063098). A. Basidioma and rhizomorphs.

B-D. Pileipellis elements. B. Encrusted hyphal segment and hypha with slime sheath.

C. Diverticulate hyphae. D. Diverticulate hyphal termini. E. Basidiospores.

Scale bars: A = 20 mm. B-D. = 10 um. E=5 um.

LAMELLAE narrow (c. 1 mm broad), thin, subdistant, total lamellae = 32-37,

through lamellae = 15-17, unequal, adnate at first, becoming adnexed or free

to sinuate in age, or sometimes attached to a partial adnate collar, seceding

upon drying, pliant, entire or minutely fimbriate, thickish, narrow, straight

at first, broader near the stipe in age, not intervenose or obscurely so in age,

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 97

= :

rs ; ' |

Fic. 37. Pseudomarasmius pallidocephalus. Basidiomata. A. Photo from nature (TENN-F-066344).

Courtesy Steve Trudell. B. Field-dried basidiomata (TENN-F-052401). Scale bars: 10 mm.

not forked, off-white at first, yellowish white (5.5Y/9.3/1.8 Munsell), pale

orange yellow 3A2 (“light buff”), or light yellowish brown (7.3YR/7.0/2.8

58 ... Petersen & Hughes

Fic. 38. Pseudomarasmius pallidocephalus (TENN-F-020486). Pileipellis elements. A-D. Repent,

diverticulate hyphae. E-H. Broom cell-like, diverticulate hyphal termini. Scale bars: 10 um.

Munsell), soon buff to pale greyish (5A2), buff to pale orange grey (<5B2),

soon buff to pale greyish (<5A2), 6A2 (“pale pinkish cinnamon’), ca. 5A4

(“light ochraceous buff”), in age mellowing to 9B2 (“vinaceous buff”),

to 6A2 (“pale pinkish cinnamon”), when dried buff to pale orange grey

(<5B2); lamellulae in 12 tiers. StrpE 1243 x 0.20.8 mm thick, insititious,

central or somewhat eccentric, terete, equal or tapering downward, straight

when moist, soon curling and twisting on drying, shining, opaque, hollow,

bristle-like but not tough (thin, stiff and easily cut), equal or tapering

slightly downward, glabrous; apical 12 mm sometimes whitish-pruinose,

more or less concolorous with lamellae (“ochraceous buft”), downward

light yellowish brown (7.3YR/7.0/2.8 Munsell) to moderate yellow brown

(0.7Y/5.2/4.1 Munsell) or dark reddish brown (10.0R/1.5/1.5 Munsell)

to dark brown on the upper half, downward black-brown, brownish grey

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 59

Fic. 39. Pseudomarasmius pallidocephalus (MICH 51323). Pileipellis elements. A,B. Repent

diverticulate hyphae. C-H. Broom cell-like, diverticulate hyphal termini. Note frequency of di- or

trichotomously branched diverticula. Scale bars: 10 um.

(6C3) to greyish brown (7D3), soon dark brown (7F4-8), 5F8 (“bister”), 6F4

(“fuscous black”), 7F8 (“bone brown’), olive-brown [6E4 (“fuscous”), 2F3

60 ... Petersen & Hughes

Fic. 40. Pseudomarasmius pallidocephalus (WTU-F-8911). Free-form pileipellis elements.

A-C. Non-diverticulate individuals. D-I. Diverticulate individuals. Scale bars: 10 um.

(“chaetura black”)], never black downward; when dried glabrous-shining,

delicately ridged, compressed; basal mycelium absent; sterile stipes absent.

RHIZOMORPHS (FIG. 36A) <17 x 0.1-0.5 mm, slender, hair-like, scarce to

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 61

Fic. 41. Pseudomarasmius pallidocephalus.

Pleurocystidia (A-D. MICH 51323. E-H. TENN-F-026266).

Note frequent distal, vague partition of contents. Scale bars: 10 um.

abundant, arising at intervals along the substrate, much branched with

spur-branches often long and flagelliform, twisted and curled, sometimes

forming a loose tangle. Opor negligible; TASTE negligible.

HABITAT & PHENOLOGY: Occasional to common in troops on needles and

debris of conifers, chiefly Picea and Abies in spruce-fir forest, rarely on Thuja

debris: highest elevation of southern Appalachian Mountains, north through

62 ... Petersen & Hughes

New England into and across the continent to the Pacific Coast temperate rain

forest and there accompanied by Pseudotsuga menziesii, Arbutus menziesii,

Quercus garryana, Acer macrophyllum, and occasionally Thuja plicata, with

understory of Alnus, Salix, Shepherdia; apparently a mid-summer fungus.

PILEIPELLIS composed of the following elements embedded in a thin slime

matrix: 1) repent hyphae 3-7.5 um diam, firm- to thick-walled (wall c. 0.5

um thick), hyaline individually, yellowish in mass, coarsely ornamented

variously from stripes with plate-like profile calluses to flake-like with the

flakes very slightly separated from the hyphal outer wall as though with a

very thin slime layer between; 2) scattered free-form hyphal segments

(Fic. 40) as in a “dryophila-structure, but not articulated into a cutis; 3)

repent hyphae (Fics 36B-D, 38A-D, 39) 4-40 x 4-10 um, inflated somewhat,

with diverticulate, papillate to digitate (often forked) processes, 2-6 x

0.7-3 um at base, not thick-walled, sub-refringent, sometimes in pairs or

dichotomously branched (ie. “saddle-shaped”), distributed more or less

randomly (not unilateral); and 4) broom cell-like termini (Fics 36c, D, 38E-H,

39C-H, 40D-1) usually arising from lightly encrusted hyphae, stalked (stalk

15-25 x 3.5-5.5 um), thin-walled, often branched dichotomously and then

often divaricate, beset by numerous diverticula; diverticula lobate to conical

and sometimes with a combination of forms; clamp connections absent.

PILEUS TRAMA loosely interwoven; hyphae 4.5-13 um diam, firm-walled,

without clamp connections, involved in minimal individual slime sheath

but not coherent in tissues, occasionally coarsely ornamented with annular

incrustation <2 um thick, often ornamented with vague, poorly defined

stripes. LAMELLAR TRAMA loosely interwoven; hyphae 2.5-8(-13) um diam,

firm- to thick-walled (wall c. 0.5 um thick, hyaline), involved in minimal

individual slime sheath but not coherent in tissues, occasionally coarsely

ornamented with annular incrustation <2 um thick, often ornamented

with vague, poorly defined stripes, easily disarticulated; clamp connections

absent. PLEUROCYSTIDIA (Fic. 41) 23-33 x 5-8 um, narrowly fusiform to

fusiform with narrowing rounded apex, usually without content partition,

without clamp connections; contents homogeneous but with vague central

vacuolated area (PhC). Basidioles clavate-subcapitate, clampless; contents

vaguely multigranular; BAsIDIA (FIG. 42A-D) (20-)25-34 x (4—)5-8(-10) um,

clavate, (2)4-sterigmate, without clamp connections; contents multigranular.

BASIDIOSPORES (FIG. 36E) (5—)5.5-7(-9.5) x 3-3.5(-6) um (Q = 1.30-2.33;

Q”™ = 1.87; L™ = 6.7 um), ellipsoid to plump ellipsoid to elongate pip-shaped,

flattened adaxially, not tapered proximally, thin-walled, smooth, inamyloid;

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 63

Fic. 42. Pseudomarasmius pallidocephalus. Hymenial structures.

A-D. Basidia (MICH 51323). E-H. Cheilocystidia (TENN-F-026266).

Scale bars: 10 um.

contents homogeneous. CHEILOCYSTIDIA (FIG. 42E-H) scattered, similar

to basidioles, 21-30 x 5-8.5 um, clavate at first, becoming ampulliform by

maturity, without clamp connections. STIPE MEDULLARY HYPHAE (Fic. 43)

64 ... Petersen & Hughes

Fic. 43. Pseudomarasmius pallidocephalus (TENN-F-056761). Stipe medullary hyphae. A. Outer

hyphae, with lines marking hyphal crust depositions. Stipe surface upward in photo. B, C. Individual

inner medullary hyphae. B. Notes septum without clamp connection. C. Note wall thickness.

Scale bars: 10 um.

(inner) (2—)3.5-8.5 um diam, strictly parallel, firm- to thick-walled (wall

<1.2 um thick, hyaline), coherent in tissues (ie. minimal slime matrix),

lacking clamp connections; outer medullary hyphae 1-3.5(-7.5) um, firm-

to thick-walled (wall <1 um thick, hyaline, ornamented with lens-shaped

encrustation often appearing pitted). STIPE CORTICAL HYPHAE moderately to

strongly dextrinoid, 3-5.5 um diam, thick-walled (wall often occluding cell

lumen), smooth or covered with heterogeneous slime with inclusions.

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 65

COMMENTARY: Pseudomarasmius pallidocephalus combines characters seen

in other related complexes. Basidiome stature, size, rhizomorphs, and habitat

all mimic (or are mimicked by) Gymnopus sect. Androsacei and/or G. sect.

Perforantia. The minimal slime matrix of the pileipellis and stipe medullary

hyphae resemble those of Gymnopus sect. Perforantia, while dextrinoid stipe

cortical tissue is like that of some members of /marasmiellus (ss Wilson &

Desjardin (2005). The diverticulate hyphae of pileipellis resemble those of

Mycetinis (including the inflated cells typical of that genus), not those of

Marasmius (siccus-type) and the free-form hyphal segments of pileipellis

are similar to such structures in Gymnopus sect. Levipedes (“dryophilus-

structure”). Lack of clamp connections, however, seems quite unusual in

related complexes. Several taxa in Gymnopus sect. Androsacei have been

described as clampless, mostly by Singer, and they generally have been

found in New- and Old-World tropical forests and none with accompanying

molecular phylogenies.

Desjardin (1989) reported Ps. (as Marasmius) pallidocephalus as “one of

the more commonly collected, early-fruiting litter-decomposing agarics in

the spruce-fir zone of the southern Appalachians.” In these spruce-fir forests,

it has been and is easily misidentified as G. (as Micromphale) perforans, which

usually exhibits a pigmented pileus toward cinnamon or avellaneous and

vestured stipe. Gymnopus androsaceus is encountered only occasionally.

Pleurocystidia in P pallidocephalus are typical of such structures found

widely through gymnopoid/marasmioid fungi. Such structures are fusiform

in shape, with narrowly rounded apex, as opposed to clavate to subcapitulate

shapes of basidioles. Apparently without taxonomic relevance, pleurocystidial

shapes are sometimes accompanied by vaguely partitioned contents. In

Ps. pallidocephalus, however, such structures are often more broadly

rounded apically. Basidioles and basidia, conversely seem to be consistently

clavate from an early stage, and with heterogeneous contents (Fic. 42a-p),

unlike pleurocystidial structures which remain homogeneous in content.

In numerous descriptions (including our publications up to a couple years

ago) the structures termed here as pleurocystidia were included under the

perceived variability of basidioles.

Gilliam (1975) mentioned clamp connections only once, under pileus

tramal hyphae. This observation was erroneous: clamp connections are

consistently absent on all basidiomatal hyphae in Ps. pallidocephalus.

Desjardin (1990) examined cultures of Ps. pallidocephalus and reported the

absence of clamp connections.

66 ... Petersen & Hughes

Past experience (Petersen 1978, Petersen & Hughes 2010) has produced

some links between absence of clamp connections, 2-spored_ basidia,

and oversize basidiospores indicative of haploid state, together with

unexplained sexual compatibility systems. These links seem irrelevant in

Ps. pallidocephalus, where basidiomata lacking clamp connections normally

produce 4-spored basidia. The mating system of this species remains

unknown.

Gilliam (1975) seemed intent on separating Ps. (as Marasmius)

pallidocephalus from M. androsaceus, the often-reported taxon which

fruits on similar substrates in Europe and North America. Her concept

of M. androsaceus was revealed in the concluding lines of her discussion,

as fruiting on a “variety of habitats.” Not only is this observation

understandable given the state of prior literature and the date of her study,

but it may have influenced Desjardin’s (1989) treatment of M. androsaceus

in his dissertation, which also drew no attention to clamp connections.

Currently, phylogenies indicate that M. androsaceus may be split into at least

three species: M. androsaceus fruiting on conifer debris, plus two which fruit

on dead deciduous leaves (Petersen & Hughes, ined.). Other mimics fruit

on pine needles [Gymnopus (Micromphale) pinophilus complex; Petersen &

Hughes 2016a], cedar twigs and redwood litter [Gymnopus (Micromphale)

sequoiae complex]. In addition to presence/absence of clamp connections,

Ps. pallidocephalus lacks conspicuous cheilocystidia [only a few ampulliform

shapes reminiscent of those of G. trabzonensis (Vizzini & al. 2015)] while

G. androsaceus exhibits common cheilocystidial structures. Finally, Gilliam

(1975) reported the lack of “hymenial cystidia” (inclusive of pleuro- and

cheilocystidia) in M. pallidocephalus, perhaps because pleurocystidia are not

conspicuous. Our experience with other taxa indicates that pleurocystidial

structures are present.

Rhizomorph presence and activity are arcane in Ps. pallidocephalus.

Basidiomata are usually formed on dead needles from the previous 1-2

years, but the needle bed on which they are found is usually deeper, with

more decayed needles (differing in color and substance) and these decayed

needles are held together by tangles of slender, branched, black rhizomorphs.

Rhizomorphs at the surface of the needle bed, and therefore accompanying

basidiomata, are usually individual, extremely slender, short, and therefore

easily overlooked.

Examination of numerous marasmioid specimens from Europe has not

revealed a single specimen of Ps. pallidocephalus.

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 67

SPECIMENS EXAMINED: CANADA, BRITISH COLUMBIA, Cariboo Regional District,

Ft. St. James, 54°26’34”N 124°15’04”’W, 13.VII.1940, coll. T.T. McCabe, det. J.W.

Groves (as M. androsaceus), TTM 253 (UC 1319403); Stikine Region, Liard River

Hotsprings Prov. Park, 59°25’06”N 126°05’27’W, 18.VIII.2000, coll. R. Winder, O.

Ceska, det. O. Ceska (as M. androsaceus) (UBC F21036); Metro Vancouver Regional

District, Mt. Seymour Prov. Park, base of chair lift, 49°22’51”N 122°56’24’”W,

27.VIII.1973, coll. S.A. Redhead (as M. androsaceus) (UBC F8178); Capital Regional

District,, Saanich Peninsula, Observatory Hill, 48°31’44”N 123°25’19”W, 17.X.2008,

coll. O. Ceska (as M. androsaceus) (UBC F29843); Observatory Hill, 48°31’44’N

123°25'19”W, 17.X.2008, coll. O. Ceska (as M. androsaceus) (UBC F29843);

Observatory Hill, 48°51.67’N 123°42.20’W, 28.11.2010, coll. & det. O. Ceska (as

M. androsaceus), UBC-F24293; Fraser Valley Regional District, Manning Prov.

Park, Rhododendron Flats, 49°13’N 121°04’W, 11.VI.2006, coll. Paul Kroeger (as

M. androsaceus), PK 4072 (UBC F17578). UNITED STATES, ALAsKaA, Fairbanks,

Fairbanks North Star, Ballane Lake Marge, 64°54.82’N 147°42.31’W, 15.VII.2006, coll.

G.A. Laursen [as Marasmius androsaceus], GAL 19442a (WTU-F-31851). IDAHO,

Bonner Co., Priest Lake, Luby Bay, 48°32’50”N 116°55’24”W, 25.1X.1992, coll. RHP

(as M. androsaceus), TFB 5632 (TENN 52407); Shoshone Co., vic. Wallace, Coeur

d’Alene Nat. Forest., 47°30’N 115°53’W, 24.1X.1992, coll. RHP ( as M. androsaceus),

TFB 5610 (TENN 52401); Maing, Hancock Co., rte 182, vic. Mud Pond and Salmon

Pond, 44°3758’N 68°05'14”W, 21.VII.1992, coll. S.A. Gordon, TFB 4987 (TENN

56761). MICHIGAN, Cheboygan Co., vic. Univ. Michigan Biological Station, Hermit

Swamp, 45°33’31”N 84°40’43” W, 26. V1.1957, coll. R.L. Shaffer (as M. cfr. androsaceus),

RLS 1291 (MICH 51323); Chippewa Co., Tahquamenon Falls State Park, Lower Falls,

46°36'16”"N 85°12’11”’W, 22.VII.1971, coll. & det. M. Gilliam (HOLOTYPE), MSG

1165 (MICH 11402, portion only); Marquette Co., Pine Plains, W of Michigamme

State Forest, “Triple A” road, 46°31’14”N 88°03’14”W, 15.VII.1970, coll. J.-. Ammirati

& S.J. Mazzer, det. M. Gilliam (as M. pallidocephalus), SJM 4479 (MICH 51322).

MINNESOTA, Clearwater Co., Itasca State Park, 9.VIII.1968, coll. M.G. Weaver (as

Marasmius sp.), det. M. Gilliam (as M. pallidocephalus), MGW 1621 (MICH 175913).

NEw York, Franklin Co., Paul Smith’s, NEMF pre-foray trip no. 1, Hazel Pond Rd.,

44°28.89'N 74°16.50’W, 11.VIII.2011, coll. A.S. Methven, TFB 13933 (TENN 65829);

NorTH CAROLINA, Swain Co., vic. Bryson City, GSMNP, Spruce-Fir Nature Trail,

35°34'02”N 83°28'54”W, 28.VIII.2009, coll. RHP (as M. androsaceus), TFB 13664

(TENN 63098); 8.VI.1991, coll. S.A. Gordon, TFB 3623 (TENN 50742); 28. VIII.2009,

coll. RHP, TFB 13664 (TENN 63098); vic. Bryson City, GSMNP, Clingman’s Dome,

35°33’40”N 83°29'45”W, 10.1X.1985, coll. D.E. Desjardin, DED 3412 (TENN 54660);

35°33’47’"'N 83°29'55”W, 27.1V.1954, coll. A.H. Smith & L.R. Hesler (TENN 021343).

TENNESSEE, Blount Co., GSMNP, Appalachian Trail vic Clingman’ss Dome, near old

Indian Gap road, 28.V.2011, coll. Steve Trudell, SAT-11-11-179-05 (TENN 66344);

Sevier Co., GSMNP, Indian Gap, Appalachian Trail, 35°36’35”N 83°26'19”’W,

29.V.2004, coll. RHP (as M. androsaceus), TFB 11778 (TENN 59896); 36°36’35”N

83°26'19’W, 13.V1I.2004, coll. S.C. McCleneghan & E.B. Lickey, TFB 11970 (TENN

59975); Old Indian Gap Road, 28.VI.1986, coll. D.E. Desjardin, DED 3691 (TENN

54667); 19. VII.1988, coll. D.E. Desjardin, DED 4615 (TENN 54650); 11.V1.1986, coll.

D.E. Desjardin, DED 3581 (TENN 54646); WASHINGTON, Jefferson Co., Olympic

68 ... Petersen & Hughes

National Park, Enchanted Valley, 47.6675°N 123.3939°W, coll. J.W. Lennox (as M.

androsaceus), JWL 511 (WTU-F-8911); Lewis Co., Skate Creek Rd., between Horse

Creek and milepost 17, 2.X.1993, coll. RHP (as M. androsaceus), TFB 5698 (TENN

53475).

Pseudomarasmius patagonianus R.H. Petersen, sp. nov. Fics 44-48

IF 555733

Differs from Pseudomarasmius nidus-avis by its 1) slender rhizomorphs that are present

but not dominant; 2) basidiomata arising as branches of rhizomorphs or from rotting

leaves; 3) pileipellis elements usually semi-gelatinized; and 4) distribution in southern

South America (Chilé).

Type: Chilé, Chiloé, Isla Grande de Chiloé, Ancud, 41°52’18”S 73°49’06”W, 9.V.1995,

coll. RHP, TFB 7363 (Holotype, TENN-F-054432).

EryMo_ocy: Referring to the Patagonian origin of the collections.

BASIDIOMATA (Fic. 444) marasmielloid, short-stipitate, similar to those

of Ps. nidus-avis. Piteus <11 mm broad, campanulate with downturned

margin becoming more or less applanate, often abruptly umbilicate, subtly

striate, somewhat pulvinate, matte, from uniformly 6C6 (“tawny”) to disc

6B5 (“cinnamon”) when young, paler in age; outward “pinkish buff” (6A3)

to off-white. LAMELLAE adnate to shortly decurrent, distant, ranging from

well-defined to reduced, usually as shallow (<1 mm broad) pleats, <20 total

folds, 5-8 through folds, 5A2 (“pale cinnamon pink”) to off-white (now 4A2

“pale ochraceous buff”). Stripe 2-8 x 1.5-2.5 mm, terete, equal, straight to

usually strongly curved, black, upward glabrous shining, downward minutely

pruinose, non-insititious with minimal basal tuft. RHIZOMORPHS rare, slender,

black, of unknown length. Opor and TasTE negligible.

HABITAT & PHENOLOGY: A poorly understood taxon, erumpent on

woody substrate, noted as “on hardwood boughs;” Chilean winter.

PILEIPELLIS comprising the following: 1) at pileus margin, repent

diverticulate hyphae 2.5-5.5 um diam, firm-walled, not gelatinized,

hyaline; diverticula stout, <7 x 1-2.5 um at base, gnarled-pyramidal, often

dichotomous; 2) at pileus margin, diverticulate hyphal termini (hardly

broom cell-like) (Fic. 45), resembling repent diverticulate hyphae, perhaps

sometimes erect; 3) over disc and limb, occasional pileal hairs <75 x 3-4.5

um, cylindrical, subtly capitulate, arising as an erect branch of the repent

hypha; 4) unorganized hyphae 3-6.5 um diam, thin-walled, variously

ornamented (Fic. 46) with crust material often coarsely in scabs <2 um

thick (Fic. 468), often appearing annular (Fic. 464,c), easily disarticulated;

5) hyphae 2.5—4.5 um, not easily crushed, thick-walled (wall <0.7 um thick),

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 69

Fic. 44, Pseudomarasmius patagonianus (TENN-F-054432, holotype).

A. Basidiomata (left) habit; (right) longitudinal section.

B. Basidiospores. Scale bars: A = 5 mm; B = 5 um. TFB 7363

often with thin individual slime sheath with flake-like ornamentation

(Fic. 46D). Diverticulate hyphae or hyphal termini not observed over disc

or limb. Lamellar trama loosely interwoven; hyphae 2-5.5 um diam, firm- to

thick-walled (wall<1 um thick), often gelatinizing or with individualthin slime

sheath, without clamp connections. PLEUROCYSTIDIA (Fic. 47) uncommon

or poorly defined, 22-25 x 5-6 um, fusiform, without clamp connections.

Basip1A (Fic. 48) 28-37 x10-12 um, clavate, (2—)4-sterigmate, without

clamp connections; contents heterogeneous when mature, emptying but not

collapsing (“husking”) when effete. BAstprosporEs (F1G.44B) 9-9.5 x 4-5 um

(Q = 1.60-2.25; E™ = 2.07), ellipsoid, thin-walled, not tapered proximally,

inamyloid. CHEILOCysTIDIA absent or undifferentiated. STIPE MEDULLARY

70 ... Petersen & Hughes

Fic. 45. Pseudomarasmius patagonianus (TENN-F-054432, holotype).

Pileipellis diverticulate hyphal termini. Scale bars: 10 um.

HYPHAE 2.55 um diam, thick-walled (wall <1 um thick), strictly parallel in

slime matrix, hyaline, without clamp connections. STIPE CORTICAL HYPHAE

35 um diam, thick-walled, repent in thin slime matrix, roughened outward,

without clamp connections, weakly dextrinoid (reddish tan Melzer’s reagent

+ BF; reddish brown Melzer’s reagent + PhC). CAuLocystTipia <65 x

3.55 um, cylindrical, firm-walled, gnarled and often with rudimentary lobes,

hyaline singly, dull brown in mass, without clamp connections.

COMMENTARY: Collection TFB7363 (TENN-F-054432) comprises several

basidiomata on hardwood substrate. TENN-F-054424, collected in the same

Paramycetinis & Pseudomarasmius gen. & spp. nov.... 71

Fic. 46. Pseudomarasmius patagonianus (TENN-F-054432, holotype). Pileipellis hyphae.

A, B. Encrusted hyphae. A, C. Ornamentation in stripes. B. Ornamentation of scabs -2 um thick.

D. Hypha with individual slime sheath and flake-like encrustation. Scale bars: 10 um.

location at the same time, is a mixed collection composed of several mycenoid

basidiomata and a single marasmielloid basidiome. The ITS-based sequence

deposited in GenBank (KY352649; MF978330, rbp2) derived under this

accession number must have come from the latter basidiome which is

72. ... Petersen & Hughes

(

hag |

Fic. 47. Pseudomarasmius patagonianus (TENN-F-054432, holotype).

Pleurocystidia. Scale bars: 10 um.

Fic. 48. Pseudomarasmius patagonianus (TENN-F-054432, holotype).

Basidia. Scale bars: 10 um.

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 73

morphologically identical to basidiomata in TFB7363: the sequence places

the collection in Pseudomarasmius and the single basidiome matches the

generic diagnosis. The basidiome has been segregated within the specimen.

As surveys of herbarium material and phylogenetic analysis of such

collections are processed, there appears to be a convergent, relatively

common basidiome architecture generally regarded as “marasmielloid,” in

which basidiomata conform to the following: 1) pileus small, rarely exceeding

15 mm broad, with thin, tough consistency; 2) lamellae usually distant,

often reduced and with no anastomoses; 3) stipe central, usually strongly

curved, rarely exceeding 10 x 2 mm, black or nearly so, usually glabrous,

4) fruiting on woody substrate often in vertical position; and 5) rhizomorphs

usually present and sometimes dominant, slender, black. This architecture

might be epitomized by Gymnopus neobrevipes (= Marasmius brevipes),

almost indistinguishable from Ps. nidus-avis and Ps. patagonianus but

distinguishable by presence of clamp connections, dominant rhizomorphs,

more temperate distribution and unique ITS sequence.

ADDITIONAL SPECIMEN EXAMINED: CHILE, Curiof, Isla Grande de Chiloé, 20 km

E Aucud, 41°53’10”S 73°38’52”W, 9.V1.1995, coll. RHP, TFB 7364 (TENN-F-054424).

Pseudomarasmius quercophiloides R.H. Petersen, sp. nov. Figs 49-59

IF 555732

Differs from Gymnopus quercophilus by its 1) lack of clamp connections; 2) abundant

rhizomorphs that are resupinate on sclerophyllous leaves; 3) sulcate-striate pileus with

low umbo; and 4) distribution in interior or subtropical China.

Type: China, Yunnan Prov., Anning Pref., grounds of Southwest Forestry University,

25°03’51"N 102°45'16”E, 10.VHI.1990, coll. RHP, Q. Wu, Li, TFB 3162 (Holotype,

TENN-F-049177).

EryMoLocy: Resemblance to Marasmius quercophilus Pouzar.

BASIDIOMATA (Figs. 494, 50) diminutive. Prteus (Fic. 498) 25 mm broad,

downturned, sulcate-striate with small umbo, matte, “vinaceous buff” (9B2).

LAMELLAE (Fic. 498) not collariate, 19-26 total lamellae, 8-11 through

lamellae, shallow (<1 mm broad), thick with rounded edge, not marginate,

“tilleul buff” (7B2) to “pinkish buff” (6A3). StrPE 8-14 x 0.6-0.9 mm, terete,

dull brown apically and there minutely decorated with hyaline caulocystidia,

downward appearing glabrous-shining but microscopically pruinose, black,

insititious. Rhizomorphs plentiful, resupinate on sclerophyllous hardwood

leaves (Fic. 51), dark copper color with minute fringe of ochraceous rust

color, rarely becoming aerial but then never pileate.

74 ... Petersen & Hughes

Fic. 49. Pseudomarasmius quercophiloides (TENN-F-049179, holotype). A. Basidioma.

B. Pileus, exterior + cross-section. C. Basidiospores. Scale bars: A, B= 5 mm; C= 5 um.

DISTRIBUTION & PHENOLOGY: On at least Castanopsis leaves in

southwestern, subtropical China; summer.

PILEIPELLIS composed of three hyphal types with no evidence of

gelatinization: 1) repent hyphae (Fic. 52a) 3-6.5 um diam, thin-walled,

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 75

bi ee

Pe ak

Fic. 50. Pseudomarasmius quercophiloides (TENN-F-049179, holotype).

Basidiomata on leaf surface. Scale bar = 10 mm.

3148

Fic. 51. Pseudomarasmius quercophiloides (TENN-F-049179, holotype).

Resupinate rhizomorphs on sclerophyllous leaves. Scale bar = 10 mm.

encrusted in scabs not in annular or striped configuration; profile scabs

<1 um thick; clamp connections not observed; 2) diverticulate hyphae

(Fic. 52B,c) 3-6.5 um diam, thin-walled, beset with numerous diverticula;

76 ... Petersen & Hughes

Fic. 52. Pseudomarasmius quercophiloides (TENN-F-049179, holotype). Pileipellis elements.

A. Repent, encrusted hypha. B,C. Diverticulate hyphae. D,E. Broom cell-like hyphal termini.

Scale bars: 10 um.

diverticula digitate (not conical), 1-8 x 0.7-1.1 um, often slightly gnarled,

dense (PhC); and 3) diverticulate broom cell-like hyphal termini (Fic. 52D,.),

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 77

Fic. 53. Pseudomarasmius quercophiloides (TENN-F-049179, holotype).

Pleurocystidia. Scale bars: 10 um.

Fic. 54. Pseudomarasmius quercophiloides (TENN-F-049179, holotype).

A. Basidiole. B—D. Basidia. Scale bars: 10 um.

78 ... Petersen & Hughes

Fic. 55. Pseudomarasmius quercophiloides (TENN-F-049179, holotype).

Clusters of upper stipe caulocystidia. Scale bars: 10 um.

similar to diverticulate hyphae, without clamp connections. No evidence

of gelatinization observed in pileipellis. Pileus trama loosely interwoven;

hyphae 2-5.5 um diam, thin-walled, hyaline, without clamp connections.

Hymenium composed of three elements with no evidence of gelatinization:

1) PLEUROCYSTIDIA (Fic. 53) occasional (not abundant), 19-24 x 5-6

um, fusiform to narrow-fusiform, often with small, subglobose, apical

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 79

Fic. 56. Pseudomarasmius quercophiloides (TENN-F-049179, holotype).

Individual upper stipe caulocystidia. Note caulocystidial origins in D. Scale bars: 10 um.

protuberance (appearing mammilate), without clamp connections;

2) basidioles (Fic. 544) clavate to subcapitulate, without clamp connections;

BASIDIA (FIG. 54B—D) (22—)27-30 x 6-7 um, 4-sterigmate (sterigmata slender,

easily collapsed), without clamp connections; and 3) CHEILOCYSTIDIA not

observed. BAsipIospoREs (FIG. 49c) 6-7 x 3-4(-5) um (Q = 1.40-1.86;

80 ... Petersen & Hughes

Fic. 57. Pseudomarasmius quercophiloides (TENN-F-049179, holotype).

Lower stipe medullary hyphae. A. Stout, thick-walled hypha. B. Gelatinized or shredded stout

hypha. C. Slender hypha. D. Serpentine slender hypha. Scale bars: 10 um.

E™ = 1.53; L™ = 6.60 um), ellipsoid, not tapering proximally, thin-walled,

hyaline, inamyloid. Upper stipe medullary hyphae <12 um diam, strictly

parallel, free (no evidence of gelatinization), hyaline, with no clamp

connections, thick-walled (wall <1.5 um thick). Upper stipe cortical hyphae

4-6 um diam, thick-walled (wall <1 um thick), strongly pigmented (olive

brown, PhC), without clamp connections, decorated with numerous

caulocystidia (Fic. 55); CAuLocysTip1A (Fic. 56) 3-30(-40) x 7-13 um,

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 81

Fic. 58. Pseudomarasmius quercophiloides (TENN-F-049179, holotype).

Individual lower stipe caulocystidia. Scale bars: 10 um.

lobate to thumb-shaped, usually gnarled or constricted, firm- to thick-walled

(wall <1 um thick), weakly pigmented, arising as constricted side branches

of surface cortical hyphae. Lower stipe medullary hyphae strictly parallel, of

two types: 1) 5-8 um diam, thick-walled (wall <2 um thick), occasionally

gelatinized (Fic. 574) or shredded (Fic. 578); and 2) slender, 2-2.5 um

diam, thin-walled, straight (Fic. 57c) to serpentine (Fic. 57p), with dense

contents (PhC). Lower stipe cortical hyphae 3.5-6 um diam, thick-walled

(wall <2 um thick, often obscuring cell lumen), densely pigmented (dark

olive brown, PhC), beset with crowded caulocystidia; CAULOCYSTIDIA

(Fic. 58) 5-30 x 8-13 um, crowded, stout-vermiform, usually gnarled and/

or constricted, thin- to thick-walled (wall <1 um thick), hyaline to weakly

pigmented proximally. Resupinate rhizomorph anatomy similar to that of

lower stipe. Cortical hyphae with superficial film of gelatinous material

(Fic 59a) acting as adhesive to the leaf surface. Rhizocystidia (Fic. 59B-E)

widely scattered, similar to caulocystidia but thick-walled (wall <1.5 um

thick) and strongly pigmented.

82 ... Petersen & Hughes

Fic. 59. Pseudomarasmius quercophiloides (TENN-F-049179, holotype).

Rhizomorph anatomy. A. Surface of cortex with suggestion of superficial gel.

B-E. Rhizocystidia. Scale bars: 10 um.

COMMENTARY: Similarities with G. quercophilus: 1) abundant

rhizomorphs, not black (in G. quercophilus chestnut brown to red-

brown), dominantly resupinate on sclerophyllous hardwood leaves;

2) pleurocystidial general shape and size; 3) pileus sulcate-striate with small

umbo; and 4) basidiomata arising from leaf surface not from rhizomorphs.

Dissimilarities to G. quercophilus include: 1) clamp connections lacking;

2) absence of cheilocystidia; 3) presence of pruinose caulocystidia.

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 83

The ITS sequence of Ps. quercophiloides is largely identical to GenBank

sequence KF251072, deposited as Gymnopus glabrocystidiatus (not holotype)

from South Korea (2 bp difference in 636bp). Geographical proximity and

sequence homology suggest that these likely represent the same taxon.

ADDITIONAL SPECIMEN EXAMINED: CHINA, YUNNAN PrRov., Jinhong Pref., vic.

Menghai, Xishuangbanna Nature Reserve, 8.VIII.990, coll. RHP, Q Wu, Li, TFB 3148

(TENN-F-049179)

Pseudomarasmius straminipes (Peck) R.H. Petersen, comb. nov. FiGs 60-64

IF 555734

= Marasmius straminipes Peck, Bull. Buffalo Soc. Nat. Sci. 1:59.

= Chamaeceras straminipes (Peck) Kuntze, Revis. Gen. P. 3(2): 457.

Ho.ortyPe (implicit, Peck): United States, New York, Albany Co, Center (= Kamer),

42°39'10”"N 73°45'52”W, Oct. 1872. C.H. Peck (NYS!).

Diagnostic characters include: 1) stipe capillary, buff above, ochraceous downward,

glabrous-shining; 2) rhizomorphs usually common, pallid; 3) fruiting on fallen conifer

needles, usually spruce/fir; 6) distribution in eastern North America.

BASIDIOMATA (Fic. 60A) diminutive, marcescent, reviving. PILEuS (1.5-)

3-7 mm broad, hemispherical or convex, expanding to plano-convex,

occasionally with suggestion of low umbo, dull, opaque, glabrous or minutely

suede-like, light brown (7D4-5) or pale greyish brown (7D3) overall when

young, soon fading to pale brownish orange (7C3) or pale brownish grey

(6B2, 6C3); disc smooth or weakly rugulose; margin rugulose-striate or

rugulose but not striate, soon fading to greyish orange (6C2) or pale greyish

orange (5B2), in age with a hint of grey and buff; context thin, buff, often

fading to whitish, buff or pinkish buff overall. LAMELLAE adnate, subdistant,

without anastomoses or interveining, narrow, whitish or buff when young,

becoming greyish buff or pale brownish grey (6C3) in age; edges even or

crystalline-fimbriate; lamellulae in 1-2 series. Stipe 10-35 x 0.2-0.5 mm,

terete, equal, tough, rigid or wiry, glabrous-shining, sometimes tacky,

hollow, instititious, brownish orange (5C4, especially downward) to yellow

(4A5-6) or straw-colored overall when young or with a slightly paler apex,

darkening overall with age to stramineous, becoming slightly more brownish

at maturity, never dark brown or black. RH1IZoMoRPHs (FIG. 60A) extensive,

with short resupinate lengths on needle surface, then loosely curly, <20 x

0.1-0.3 mm, repeatedly branched with branches often long and flagelliform,

glabrous, dull yellow-brown in basal parts, champagne colored upward and

finally off-white. Opor and TasTE negligible.

84 ... Petersen & Hughes

Fic. 60. Pseudomarasmius straminipes (TENN-F-026266).

A. Basidiomata and rhizomorphs. B. Basidiospores. Scale bars: A = 10 mm; B = 5 um.

HABITAT & PHENOLOGY: Gregarious to scattered to dead conifer needles

including Picea/Abies and Pinus (especially P strobus); eastern North

America [reported by Desjardin & Petersen (1989a) from Alabama, New

Jersey, New York, North Carolina, Ohio and Tennessee]; summer—autumn.

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 85

Fic. 61. Pseudomarasmius straminipes (TENN-F-047638).

Pileipellis hairs. Note production of abortive side branch. Scale bars: 10 um.

PILEIPELLIS without evidence of slime matrix, a well-developed

Rameales-structure, of the following elements: 1) pileal hairs (Fic. 61)

densely scattered, <80 x 4-6 um, firm-walled, cylindrical, without clamp

connections, often with aborted side branch, hyaline, usually subcapitulate;

2) repent hyphae 4-8.5(-15) um diam, thin- to firm-walled, without clamp

86 ... Petersen & Hughes

Fic. 62. Pseudomarasmius straminipes (TENN-F-047642). Pileipellis elements.

A-C. Diverticulate hyphae. D-K. Broom cell-like hyphal termini. Scale bars: 10 um.

connections, strongly to weakly encrusted (crust material in scabs with

vaguely annular to spiral patterning but occasionally protruding and then

resembling diverticula); 2) scattered repent diverticulate hyphae (Fic. 62A-c)

irregular in outline, often lobed, 2.5-8(-12) um diam, hyaline to pale

brownish, smooth or weakly encrusted, inamyloid; diverticula 1-8 x 1-3

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 87

~~ *

Fic. 63. Pseudomarasmius straminipes (TENN-F-047638). Hymenial structures.

A-D. Pleurocystidia. E. Basidiole. F-H. Basidia. Scale bars: 10 um.

um, knob-like, often dichotomous and in fascicles, obtuse, thin-walled; and

3) broom cell-like hyphal termini (Fic. 62p-K) common, stalked (stalk 4-8 x

3.5-5.5 um, without clamp connections), usually branched, distally variously

lobed to diverticulate; diverticula 1-5 x 1-1.5 um, nodulose, subrefringent,

often appearing dichotomous. Pileus trama interwoven; hyphae 28-11)

88 ... Petersen & Hughes

Fic. 64. Pseudomarasmius straminipes (TENN-F-047638).

Cheilocystidia. Note suggestion of dichotomous setulae. Scale bars: um.

um diam, inflated or not, hyaline, inamyloid, thin-walled, smooth or

weakly incrusted; incrustations granular or often helical and yellowish to

pale brownish. Lamellar tramal hyphae similar to pileus tramal hyphae,

but hyaline and non-encrusted throughout, without clamp connections.

PLEUROCYSTIDIA (FIG. 63A4-D) abundant, 21-26(-29) x 5-8 um, fusiform

to fusiform-rostrate, without clamp connections; contents more or less

homogeneous. Basidioles (Fic. 63E) clavate; BASIDIA (FIG. 63F-H) 19-28 x

6.4-10 um, clavate, (2—)4-sterigmate, without clamp connections; contents

more or less homogeneous; effete basidia emptying but not collapsing

(“husking”). BASIDIOSPORES (FIG. 60B) 6.5-9.5 x 3.2-4.5 um (Q = 1.70-2.40;

E™ = 2.00; L™ = 8.00 um), ellipsoid, slightly tapered proximally, smooth,

hyaline, inamyloid; spore deposit whitish. CHEILOCysTIDIA (Fic. 64)

scattered to abundant, 13-—23(-33) x 10-15 um 4.5-8(-11) um, stalked (stalk

<2-6 x 3.5-8 um, thin-walled, without clamp connections), unbranched

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 89

or once-branched (and then branches short, stout), often lobed, setulose,

hyaline, firm-walled; setulae 1-4 x 0.5-2 um, sub-refringent, often saddle-

shaped to dichotomous). STIPE MEDULLARY HYPHAE 3.5—7(-10) um diam,

strictly parallel, firm- to thick-walled (wall <0.5 um thick, hyaline, Melzer’s

reagent) without evidence of slime matrix (Melzer’s reagent, PhC), without

clamp connections. inamyloid. STIPE CORTICAL HYPHAE 1.5-6.5 um diam,

strictly parallel, thick-walled (walls <2.5 um thick, non-encrusted), smooth,

yellowish or pale brownish orange, moderately dextrinoid (Melzer’s reagent

+ BF). RHIZOMORPH tissue similar to that of the stipe, with dextrinoid

cortical hyphae. Clamp connections absent on all tissues.

COMMENTARY: Protologue (Peck 1873 Bull. Buffalo Soc. Nat. Sci. 1(2): 59.):

“Pileus membranaceous, hemispherical or convex, smooth, striate, whitish;

lamellae distant, unequal, white; stem corneous, smooth, shining, filiform,

inserted, pale straw colored. Plant 1”—2” [inches] high, pileus 1”—3” [lines]

broad. Fallen leaves of pitch pine, Pinus rigida. Center [N.Y.] October.”

After examination of the type specimen of M. straminipes, Gilliam

(1976: 136) opined that: “The type specimen is closest to Marasmiellus,

and reviewed some putative diagnostic characters. In the same publication,

Gilliam (1976: 131) reported on another Peck taxon, M. filopes Peck,

basidiomata of which are similar to those of M. straminipes, and stated:

“There is too little left of the specimen to section,’ but concluded that

“M. filopes is nearest Marasmiellus’ Transfers of neither species

(M. straminipes, M. filopes) to Marasmiellus, however, were formally

accomplished. Redhead (1980), in his treatment of Marasmiellus filopes,

did not mention or compare M. straminipes (validly published by Peck

1873, repeated in 1874), but, like Gilliam (1976), considered M. filopes,

M. thujinus Peck, and M. piceina Kauffman as taxonomic synonyms. In a

later discussion of clampless Marasmius taxa, Redhead (1984) rejected

Gilliam’s (1976) opinion of M. straminipes as Marasmiellus, retaining the

species in Marasmius sect. Androsacei, but without mention of Marasmiellus

filopes and its synonyms. Redhead (1984) mentioned the clampless state of

M. straminipesby way ofcomparison with M. pallidocephalus. Desjardin (1989)

examined Peck’s type specimens of M. straminipes and M. filopes. Still later, in

a paper redescribing Marasmius straminipes (Desjardin & Petersen 1989b),

M. straminipes was again reportedasclampless and Redhead’s (1984) retention

of M. straminipes in Marasmius sect. Androsacei was accepted, but M. filopes

was not mentioned. In addition to the characters dictating placement of

M. straminipes in Marasmius for Redhead (1984), Desjardin & Petersen

90 ... Petersen & Hughes

(1989b) added the dextrinoid reaction on stipe medullary hyphae in

M. straminipes, theretofore unknown in Marasmiellus.

Description of basidiomata exhibiting clamp connections and those

lacking clamps as forms or varieties of the same species is commonly

found in the literature. Often, the clampless state is found accompanied by

2-sterigmate basidia and somewhat larger basidiospores than expected (see

such forms in Hymenopellis, Petersen & Hughes 2010; and in Mycena, Kithner

1938, Lamoure 1957a,b, 1959, 1960). Upon further study, such forms may

be found to be technically anamorphic (asexual) structures reproducing

without meiosis and existing in a haploid state. In clampless forms in

Gymnopus subg. Gymnopus, however, basidia appear to be 4-sterigmate

and basidiospore metrics conform to the norms expected. Causes for

such correlation remain opaque. In background research resulting in this

paper, Ps. efibulatus was first noted as a clampless form of an undescribed

Marasmius species. Marasmius straminipes (clampless) was thought to have

a variety with clamp connections (M. straminipes var. fibulatus Desjardin

& R.H. Petersen 1989. Bull. Torrey Bot. Club 49: 184.) With the proposal

of Pseudomarasmius, a different avenue is offered, with molecular analyses

important. DNA sequences of the analogous forms might clarify taxonomy,

but so far such sequences have not been produced.

The proposed transfer of M. straminipes to Pseudomarasmius is based

on morphological characters: unfortunately, sequences derived from

M. straminipes basidiomata have consistently been “dirty,” with more than

one simultaneous sequence overlapping one another. The close resemblance

to Ps. glabrocystidiatus in form, ecology, and microstructure, however,

makes our transfer relatively comfortable.

SPECIMENS EXAMINED: UNITED STATES, NorTH CaRo.Lina, Macon Co., vic.

Otto, Coweeta Hydrologic Laboratory, Ball Creek area, 35°02’58”N 83°26’20’W,

13.VIII.1987, coll. D.E. Desjardin, DED 4458 (TENN 47642); vic. Highlands, Blue

Valley, ca 3 mi up Clear Creek Rd., 9.VII.1991, coll. D.E. Desjardin, DED 5149

(TENN-F-050018, SFSU); Swain Co., vic. Bryson City, GSMNP, Clingman’s Dome,

1.VIII.1989, coll. S.A. Gordon, TFB 2125 (TENN-F-049143). TENNESSEE, Sevier

Co., GSMNP, Old Indian Gap Road off Clingman’s Dome Road, 11.VI.1986, coll.

D.E. Desjardin, DED 3578 (TENN-F-047638); Indian Gap, 35°36’34”N,83°26'47” W,

12.VI.1952, coll. L.R. Hesler (as Marasmius sp.), det. D.E. Desjardin (TENN-F-026266).

General discussion

The clampless state has always raised questions about nuclear behavior in

the basidial primordium and basidiospores (see Petersen 1995). Often, the

clampless state is accompanied by 2-sterigmate basidia, with the ploidy of

Paramycetinis & Pseudomarasmius gen. & spp. nov. ... 91

parent mycelium, dikaryotization, meiosis and subsequent mitosis unclear

(Petersen 1978). In Pseudomarasmius, however, the clampless state seems

coupled with 4-sterigmate basidia (rarely 2-sterigmate in Ps. pallidocephalus

and Ps. nidus-avis). This allows some conjecture that parent mycelium

was dikaryotic, with diploidization in the basidial primordium probable,

although nuclear segregation in the basidium and nuclear number in the

basidiospores remains unknown. Clarifying experiments in this situation

cannot be assumed by culturing single basidiospores and pairing resultant

cultures, for the nuclear condition of the “monokaryons” is unknown until

observed Cytogenetic evidence must be gathered directly, usually with

the aid of DAPI staining and phase contrast microscopy or some more

cumbersome stains (Restivo & Petersen 1976, Hubbard & Petersen 1979).

In an entirely different direction, if any doubt remains, morphological

identification and especially generic (or infrageneric) placement has

become almost impossible in large groups of marasmioid fungi. Gymnopus

in the sense of Wilson & Desjardin (2005), especially, has been repeatedly

shown to form a monophyletic clade but inclusive of disparate basidiomatal

morphology, ranging from G. androsaceus (traditionally type of Marasmius

sect. Androsacei) and G. perforans (traditionally type of Micromphale sect.

Perforantia) rather typical of marasmioid stature, to G. fusipes (typus generis)

forming much larger, coarser and fasciculate basidiomata, and Caripia

which forms only a subclavarioid hymenophore. Typical Micromphale

(M. venosum typus generis), with gelatinized tissue layer in the pileus trama,

also is now found in the Gymnopus clade (see Fie. 1).

In another different taxonomic direction, Mycetinis (and Paramycetinis,

see above), Rhodocollybia, Lentinula, Connopus, and Pseudomarasmius (see

above), all traditionally within larger genera, are now known to resolve as

monophyletic clades (see Fie. 1).

Acknowledgments

We acknowledge NSF DEB-1354802 awarded to M.E. Smith and P.B. Matheny,

through which sequence data was obtained for Ps. patagonianus. The authors

recognize and appreciate several National Science grants (especially two PEET

grants) which supported fieldwork and provided experience basic to this effort.

Equally, the Hesler Endowment Fund of the University of Tennessee provided funds

for travel. Thanks are extended to Dr. Scott Redhead and Dr. Else Vellinga for help

with nomenclatural issues. Dr. Victor Bandala provided valuable information on Ps.

nidus-avis. Jerry Cooper (New Zealand), Genevieve Gates (Tasmania, Australia),

and Tom May (Melbourne, Australia) pre-reviewed materials on Paramycetinis

92 ... Petersen & Hughes

for possible nomenclatural repetition. David Lewis contributed photos. Vladimir

Antonin and Clark Ovrebo are thanked for providing presubmission reviews.

The editors of Mycotaxon have been particularly helpful in seeing this paper through

the press.

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MY COTAXON

January-March 2020—Volume 135, pp. 97-102

https://doi.org/10.5248/135.97

Caliciopsis sambaibae sp. nov. from the Brazilian Cerrado

JosE Luiz BEZERRA*, MARUZANETE PEREIRA MELO’,

JosE EVANDO AGUIAR BESERRA JR”, ELLIOT WATANABE KITAJIMA3,

SAMARA RAQUEL SOUSA’, CRISTIANE DUARTE SANTOS?

"Centro de Ciéncias Agrarias, Ambientais e Bioldgicas, Universidade Federal

do Reconcavo da Bahia, Cruz das Almas BA, 44380-000, Brazil

? Departamento de Fitotecnia, Universidade Federal do Piaui,

Teresina PI, 64049-550, Brazil

° Departamento de Fitopatologia, Universidade de Sado Paulo,

Escola Superior de Agricultura Luiz de Queiroz/USP

Piracicaba SP, 13418-900, Brazil

“CORRESPONDENCE TO: evando@ufpi.edu.br

ABSTRACT—A new species, Caliciopsis sambaibae, is described associated with Davilla

elliptica plants. The collections were made in the Cerrado of Sete Cidades National Park, in

the municipality of Piracuruca, Piaui State, Brazil.

Key worps—Ascomycota, Coryneliaceae, Coryneliales, Eurotiomycetes, sambaiba

Introduction

The Cerrado, designated as a neotropical plant diversity hotspot (Simon & al.

2009), is the main vegetation type of five Brazilian states— Minas Gerais, Goias,

Mato Grosso do Sul, Maranhdo, and Tocantins—and also occurs in six other

states—Bahia, Mato Grosso, Piaui, Sao Paulo, Parana, and Rond6énia (Beuchle

& al. 2015). This vegetation covers about 23% of Brazil and incorporates

c. 12,000 species of angiosperms (Zappi & al. 2015). Davilla Vand., one of the

most diverse genera within Dilleniaceae Salisb., comprises about 30 neotropical

species of lianas and erect or climbing shrubs (Fraga & Stehmann 2010). Plants

of this genus are locally known as “sambaiba’ and are commonly used in

traditional medicine in anti-inflammatory treatments (Lima & al. 2014).

98 ... Bezerra & al.

The survey of fungi associated with plants from the Cerrado has revealed

many new fungi (Dornelo-Silva & Dianese 2004, Dornelo-Silva & al. 2007)—

c. 100 new species and 20 new genera of fungi (mainly ascomycetes) have

been described (Dianese 2000, Hernandez-Gutiérrez & Dianese 2008, Pereira-

Carvalho & al. 2009, Armando &al. 2015, Firmino & al. 2016, Melo & al. 2017).

Caliciopsis (Coryneliaceae), represented by c. 30 species and found in

temperate and tropical regions of North and South America, Eurasia,

and Australia (Crous & al. 2016, Pratibha & al. 2010, Fitzpatrick 1920), is

characterized by long black perithecia lacking paraphyses, evanescent asci,

and brown ascospores. Some species produce an asexual morph, which is

rarely observed because the conidia are produced within stromatic pycnidia

(Garrido-Benavent & Pérez-Ortega 2015). Some species are associated

with tree cankers or leaf spots (Jordal & al. 2004, Pratibha & al. 2010);

severe cankering is caused by the type species C. pinea Peck (on Pinus),

C. calicioides (Fr.) Fitzp. (on Populus), and C. pleomorpha P.A. McGee &

Pascoe (on Eucalyptus) (Benny & al. 1985, Fitzpatrick 1920, 1942; Munck &

al. 2015, Pascoe & al. 2018).

The symptoms on our collections of Davilla elliptica leaves began with

yellow spots, which turned necrotic and brown. In the lesions, we noted

numerous fungal structures similar to black spines corresponding to the

rostrate perithecial necks characteristic of a Caliciopsis fungus. As the

morphology and biometric data of our specimens do not correspond with

any described Caliciopsis, we propose a new species, Caliciopsis sambaibae.

Materials & methods

During 2014-16, Davilla elliptica leaves bearing signs of fungal infection were

collected in Sete Cidades National Park in the municipality of Piracuruca, Piaui State,

Brazil. The leaves were examined under a stereoscopic microscope and fragments

and hand sections of the ascomata were mounted between slide and coverslip using

Amann’ lactophenol. Leaf samples were submitted to scanning electron microscopy

(SEM) in the Electron Microscopy laboratory of the Escola Superior de Agricultura

Luiz de Queiroz, Universidade de Sao Paulo, as follows: leaf fragments containing the

fungal structures were placed on filter paper moistened in a Petri dish, along with

a small plastic container containing ca. 1 mL of 2% OsO, in aqueous solution. The

container was wrapped with Parafilm and foil and left overnight (Kitajima & Leite

1999). The plate was then opened, and the leaf fragment was removed, air dried,

mounted in the scanning electron microscope stub port with carbon double-sided

adhesive tape, and covered with a thin layer of gold on a Bal-tec SCD050 sputter

coater. The sample was examined and digitally recorded with a LEO 435 VP scanning

electron microscope at 20 kV and a 10-mm working distance.

Caliciopsis sambaibae sp. nov. (Brazil) ... 99

Specimens were conserved in the herbarium, Universidade Federal de Vicosa,

Minas Gerais, Brazil (VIC).

Taxonomy

PxaTE 1. Caliciopsis sambaibae (holotype, VIC 44133) on Davilla elliptica. A-C. leaf symptoms;

D-F: perithecia on leaves (LM). Scale bars: D = 30 um, E, F = 100 um.

Caliciopsis sambaibae J.L. Bezerra, M.P. Melo & Beserra, sp. nov. Figs 1, 2

MB 819221

Differs from other Caliciopsis spp. by its perithecia having scars on their middle third

and a basal dilation where ascospores are stored, and by its dilleniaceous host.

Type: Brazil, Piaui, Piracuruca 4°02’08”S 41°40’45’”W, on living leaves of Davilla

elliptica A.St.-Hil. (Dilleniaceae), 20 May 2016, M.P. Melo (Holotype, VIC 44133).

Erymo ocy: Referring to “sambaiba’, the common name of the host.

Spots yellowish, becoming brown and necrotic with age. COLONIES

amphigenous, rarely epiphyllous. Srromata irregular to circular in outline,

superficial mycelium absent; internal mycelium intercellular; stromata black,

formed on the leaf surface from which the perithecia and pycnidia are inserted.

SEXUAL MORPH: PERITHECIA ellipsoid, with long necks, simple or forked,

straight or curved, 550—730 x 30-60 um, with a swollen ascogenous region near

the base; ring-like scars and apical filaments on the necks of mature perithecia;

ASCI bitunicate with eight ascospores, formed in the ascogenous swellings,

evanescent, 17-20 x 5—7 um (n = 20); ascospores 1-celled, smooth-walled,

100 ... Bezerra & al.

PLATE 2. Caliciopsis sambaibae (holotype, VIC 44133) on Davilla elliptica. A, B. perithecia

(SEM); C. asci and ascospores (LM). D, E. scars (red arrows) on face of perithecia (SEM);

FE apical filaments (red arrow) on the neck of mature perithecium (SEM); G. neck of immature

perithecium (SEM); H. pycnidia in the basal stroma (SEM); I. cross sections of pycnidia (SEM).

Scale bars: A, B = 1 mm; C = 10 um; D, E = 200 um; F, G = 20 um; H = 200 um; I = 10 um.

globose to lenticular, brown, 3-6 x 3—5 um. pycnidia cespitose, developed at

the perithecial base, irregular, stromatic, 23-20 um diam.; CONIDIOGENOUS

CELLS holoblastic, nearly inconspicuous; conrp1A 1-celled, smooth, hyaline,

ellipsoid, 3-4 x 2 um.

ADDITIONAL SPECIMEN EXAMINED: BRAZIL, Prauf, Piracuruca, on living leaves of

Davilla elliptica, 11 October 2014, J.E.A. Beserra Jr. (VIC 44132).

Discussion

Caliciopsis sambaibae produces ellipsoid, spine-shaped perithecia with

ascogenous swellings near the bases where the asci are formed. Sections

in this region revealed asci in formation, and ascogenous swellings were

not observed in immature perithecia. The apex of the perithecial ostiole

is somewhat dilated, and hyphal filaments may assist in the transfer of

ascospores to this region. Due to their evanescent nature, only a few intact

asci could be observed. Possibly for this reason, asci have been observed

only in C. indica J. Pratibha & Bhat, C. valentina Garrido-Ben. & Pérez-Ort.,

Caliciopsis sambaibae sp. nov. (Brazil) ... 101

C. calicioides, and C. beckhausii (Korb.) Garrido-Ben. & Pérez-Ort. (Garrido-

Benavent & Pérez-Ortega 2015, Jordal & al. 2014, Pratibha & al. 2010). After

disintegration of the asci, passive ascospore liberation and agglomeration

occur at the extremities of the perithecial necks. The presence of pycnidia

immersed in plant tissue separates C. arrhiza (Pat. & Gaillard) Bat. &

J.L. Bezerra and C. confusa Bat. (Batista & Maia 1964, Batista 1956) from

C. sambaibae. The presence of scars in the middle or near the apex of the

ostiole, as well as bifurcated ostioles distinguish C. sambaibae from other

Caliciopsis species. The pycnidia of C. sambaibae are generally produced

in groups inserted in the basal stroma, a morphological feature shared

with C. beckhausii. Although most Caliciopsis species are associated with

tree cankers (Jordal & al. 2004, Benny & al. 1985, Fitzpatrick 1942), we did

not observe cankers in Davilla elliptica plants. Caliciopsis indica is the only

species associated with leaves (Pratibha & al. 2010).

Acknowledgments

The authors thank CAPES (Coordenacao de Aperfeigoamento de Pessoal de

Nivel Superior) and CNPq (Conselho Nacional de Desenvolvimento Cientifico e

Tecnoldgico) for funding through grants to authors. They are also grateful to Roger

Fagner Ribeiro Melo (Universidade Federal de Pernambuco, Brazil) and Jadson Diogo

Pereira Bezerra (Universidade Federal de Goias, Brazil) for helpful comments and

review.

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MYCOTAXON

January-March 2020—Volume 135, pp. 103-117

https://doi.org/10.5248/135.103

Morphological and molecular identification of

Phlebia wuliangshanensis sp. nov. in China

Ruo-X1A HUANG?”, KAI-YUE Luo?, RuI-XIN MA3, CHANG-LIN ZHAO???

' Key Laboratory of State Forestry Administration for Highly Efficient Utilization of

Forestry Biomass Resources in Southwest China,

’ Key Laboratory for Forest Resources Conservation and Utilization in the

Southwest Mountains of China, Ministry of Education,and

* College of Biodiversity Conservation,

'23Southwest Forestry University, Kunming 650224, PR. China

" CORRESPONDENCE TO: fungichanglinz@163.com

ABSTRACT —A new white-rot fungus, Phlebia wuliangshanensis, is proposed based on a

combination of morphological features and molecular evidence. The species is characterized

by an annual growth habit, resupinate basidiocarps with a smooth to tuberculate hymenial

surface, a monomitic hyphal system with thin- to thick-walled generative hyphae bearing

simple septa, presence of cystidia, and narrow ellipsoid to ellipsoid basidiospores (5-6 x

3-3.7 um). Our phylogenetic analyses of ITS and LSU nrRNA sequences performed with

maximum likelihood, maximum parsimony, and Bayesian inference methods support

P. wuliangshanensis within a phlebioid clade in Meruliaceae (Polyporales). ITS+nLSU

sequence analyses of additional Phlebia taxa strongly support P. wuliangshanensis within a

monophyletic lineage grouped with P. chrysocreas and P. uda.

Key worps—Basidiomycota, Ceriporiopsis, taxonomy, wood-inhabiting fungi, Yunnan

Province

Introduction

Phlebia Fr. (Meruliaceae, Polyporales) is typified by P. radiata Fr. (Fries

1821). Basidiocarps are resupinate or rarely pileate with a subceraceous to

subgelatinous consistency when fresh and membranaceous to coriaceous

when dry. The hymenophore may be smooth, tuberculate, phlebioid,

104 ... Huang & al.

odontoid, merulioid, or poroid. Microscopic characters include a monomitic

(rarely dimitic) hyphal structure with clamp connections or simple-septa,

narrowly clavate basidia, and hyaline basidiospores that are thin-walled,

smooth, allantoid to ellipsoid, acyanophilous, and negative in Melzer’s reagent

(Bernicchia & Gorjon 2010). So far about 100 species have been accepted in the

genus worldwide (Fries 1821; Ginns 1969; Nakasone & Burdsall 1984, 1995;

Dhingra 1989; Nakasone 1997, 2002, 2003, 2009; Roberts 2000; Gilbertson

& Hemmes 2004; Duhem & Michel 2007; Duhem 2009, 2013; Bernicchia &

Gorjon 2010; Singh & al. 2010; Westphalen & al. 2018; Gorjon & Greslebin

2012; Kaur & al. 2017; Shen & al. 2018).

Recent, molecular studies have elucidated the classification of Phlebia

among corticioid homobasidiomycetes (Larsson & al. 2004; Larsson 2007;

TomSsovsky & al. 2010; Binder & al. 2013, Justo & al. 2017; Shen & al. 2018).

Larsson & al. (2004) showed that Phlebia clustered into a phlebioid clade and

grouped with Ceriporia Donk and Gloeoporus Mont. Larsson (2007) emended

part of Polyporales and demonstrated that Phlebia was polyphyletic and nested

within Meruliaceae. A phylogenetic study of European taxa of Ceriporiopsis

Domanski showed that Phlebia radiata and the generic type of Ceriporiopsis,

C. gilvescens (Bres.) Domanski, grouped closely on the basis of combined

nuclear ribosomal large subunit RNA (nLSU) and mitochondrial ribosomal

small subunit rRNA (mtSSU) gene sequences (TomSovsky & al. 2010). Binder

& al. (2013) multi-gene sequence analyses placed P. radiata within a phlebioid

clade and apparently grouped with Ceriporiopsis and Climacodon P. Karst.

Also using multi-gene datasets, Justo & al. (2017) revised the family-level

classification of Polyporales, including eighteen families. They showed that

P. radiata belonged to Meruliaceae and grouped with Aurantiporus Murrill

and Ceriporiopsis gilvescens. Shen & al. (2018) described a new Phlebia

species based on morphological characters and rDNA sequences. This species

belonged within the phlebioid clade and was related to P. radiata.

During our investigations of wood-inhabiting fungi in southern China, we

found an additional taxon that could not be assigned to any described species.

In examining the taxonomy and phylogeny of this new species, we employed a

two-gene molecular phylogenetic approach using internal transcribed spacer

(ITS) and long subunit (nLSU) plus an expanded sampling of Phlebia isolates.

Materials & methods

The specimens studied are deposited at the herbarium of Southwest Forestry

University (SWFC). Macro-morphological descriptions are based on field notes.

Colour terms follow Petersen (1996). Micro-morphological data were obtained

TABLE 1. Species, specimens, and sequence data used in this study.

New sequences in bold.

SPECIES NAME

Abortiporus biennis

Antrodia albida

A. heteromorpha

Antrodiella americana

A. semisupina

Ceriporiopsis gilvescens

Climacocystis borealis

Coriolopsis caperata

Dacryobolus karstenii

Daedalea quercina

Earliella scabrosa

Fomitopsis pinicola

E rosea

Fragiliporia fragilis

Ganoderma lingzhi

Gelatoporia

subvermispora

Gloeoporus dichrous

G. pannocinctus

Grammothelopsis

subtropica

Heterobasidion

annosum

Hornodermoporus

martius

Hydnophlebia

chrysorhiza

Hypochnicium

lyndoniae

Junghuhnia nitida

Obba rivulosa

O. valdiviana

Phanerochaete

chrysosporium

P. velutina

Perenniporia

medulla-panis

Perenniporiella neofulva

Phlebia acanthocystis

P. acerina

SAMPLE NO.

TFRI 274

CBS 308.82

CBS 200.91

Gothenburg 3161

FCUG 960

BRNM 710166

KH 13318

LE(BIN)-0677

KHL 11162

HHB 8735

PR 1209

CCBAS 536

ATCC 76767

Dai 13080

Dai 13559

Dai 13561

Wu 1006-38

BRNU 592909

KHL 11173

BRNM 709972

Cui 9041

PFC 5252

MUCL 41677

FD-282

NL 041031

KHL 11903

KCTC 6892

FF 503

BKM-F-1767

HHB-15343

MUCL 49581

MUCL 45091

FP 150571

FD-301

HHB-11146

FP-135252

DR-60

Phlebia wuliangshanensis sp. nov. (China) ... 105

GENBANK ACCESSION NO.

ITS

EU232187

DQ491414

DQ491415

JN710509

EU232182

FJ496684

JQ031126

AB158316

EU118624

FJ403214

JN165009

FJ608588

DQ491410

KJ734260

KJ734261

KJ734262

JQ781858

FJ496694

EU118627

EU546099

JQ845094

KC492906

FJ411092

KP135338

JX124704

EU118638

FJ496693

HQ659235

HQ188436

KP135184

FJ411087

FJ411080

KY948767

KP135378

KP135372

KP135371

KP135375

LSU

EU232277

AY515348

AY515350

JN710509

EU232266

FJ496720

JQ031126

AB158316

EU118624

FJ403214

JN164793

DQ491410

KJ734264

KJ734265

KJ734266

FJ496706

EU118627

FJ496708

JQ845099

KC492906

FJ393859

KP135216

JX124704

EU118638

FJ496710

HQ659235

GQ470643

FJ393875

FJ393852

KY948844

KP135378

KF691615

REFERENCES

Unpublished

Kim & al. 2007

Binder & al. 2013

Binder & al. 2005

Tomsovsky & al. 2010

Binder & al. 2013

Tomsovsky & al. 2010

Binder & al. 2005

Lindner & al. 2011

Binder & al. 2005

Unpublished

Kim & al. 2007

Zhao & al. 2015a

Zhao & al. 2013

Tomsovsky & al. 2010

Binder & al. 2005

Tomsovsky & al. 2010

Zhao & al. 2013

Binder & al. 2013

Robledo & al. 2009

Floudas & Hibbett 2015

Unpublished

Binder & al. 2005

Miettinen &

Rajchenberg 2012

Miettinen &

Rajchenberg 2012

Wu & al. 2010

Floudas & Hibbett 2015

Robledo & al. 2009

Justo & al. 2017

Floudas & Hibbett 2015

Table 1, continued on next page

106 ... Huang & al.

Table 1, continued

SPECIES NAME

P. ailaoshanensis

P aurea

P. centrifuga

P. chrysocreas

P floridensis

P. fuscoatra

= Mycoacia fuscoatra

P. hydnoidea

P. lindtneri

P livida

P. ludoviciana

P. nantahaliensis

P. nothofagi

= Mycoacia nothofagi

P radiata

P rufa

P. setulosa

P. subochracea

P. subserialis

P. uda

P. wuliangshanensis

SAMPLE NO.

CLZhao 3996

CLZhao 4036

DLL 2011-100

FCUG 2767

NH-14434

RLG-5075

HHB-9239

L-15541

GB-1013

HHB-6333

HHB 3946

FP-102161

KUC 20121123-24

HHB-9905

HHB-6466

HHB-7175

FP-102562-T

HHB-10782

HHB 15354T

HHB 18642

FP-102173

KHL 13275

HHB-1993

GB-501

FCUG 2189

FD-427

HHB-2816

HHB-4273

HHB-6906

HHB- 12067

KHL 13750

UBCF 19726

AFTOL-ID 484

FD-85

FD-121

HHB-14924

HHB-6891

AH 31879

PH 11749

HHB 8715

FCUG 1434

FP-101544

FCUG 2452

USDA Kropp-1

CLZhao 3475 T

CLZhao 3639

GENBANK ACCESSION NO.

ITS

MH784926

MH784927

KJ140614

HQ153409

AY463445

KY948759

KP135380

KP135381

KP135379

KP135358

KP135357

AY219367

KJ668482

KP135383

KP135385

KP135384

KP135386

KP135364

KP135367

KP135366

KP135364

JN649352

KY948778

KY948772

AF141624

KP135342

KY948777

KP135369

KP135368

KP135370

GU480000

HQ604797

AY854087

KP135377

KP135376

KP135374

KP135382

GQ259417

GU461312

KY948770

AF141631

KP135361

AF141614

AB084621

MK881787

MK881788

LSU

MH784936

MH784937

HQ153409

AY586691

KY948918

KP135262

KP135263

AY586695

KJ668335

KP135264

KP135265

KP135363

JN649352

KY948853

KY948847

AF141624

KY948852

KP135266

GU480000

HQ604797

AF287885

KP135377

KX065989

KP135267

GQ259417

KY948846

AF141631

KP135232

MK881897

MK881898

REFERENCES

Shen & al. 2018

Unpublished

Binder & al. 2013

Larsson & al. 2004

Justo & al. 2017

Floudas & Hibbett 2015

Binder & al. 2005

Floudas & Hibbett 2015

Justo & al. 2017

Floudas & Hibbett 2015

Tomsovsky & al. 2010

Justo & al. 2017

Tomsovsky & al. 2010

Floudas & Hibbett 2015

Justo & al. 2017

Floudas & Hibbett 2015

Tomsovsky & al. 2010

Binder & al. 2013

Binder & al. 2005

Justo & al. 2017

Floudas & Hibbett 2015

Justo & al. 2017

Binder & al. 2005

Floudas & Hibbett 2015

Tomsovsky & al. 2010

Floudas & Hibbett 2015

Unpublished

Suhara & al. 2002

Present study

CLZhao 3645 MK881789 MK881899 Present study

Phlebia wuliangshanensis sp. nov. (China) ... 107

Table 1, concluded

GENBANK ACCESSION NO.

SPECIES NAME SAMPLE NO. Torr REFERENCES

ITS LSU

Piloporia sajanensis Mannine 2733a HQ659239 HQ659239 TomSovsky & al. 2010

Podoscypha venustula CBS 65684 JN649367 JN649367 Sjoekvist & al. 2012

Polyporus tuberaster CulTENN 10197 AF516596 AJ488116 Binder & al. 2013

Postia guttulata KHL 11739 EU11865 EU11865 Kim & al. 2007

Pouzaroporia subrufa BRNM 710164 FJ496661 FJ496723 TomSsovsky & al. 2010

Sebipora aquosa Miettinen 8680 HQ659240 HQ659240 Miettinen &

Rajchenberg 2012

Skeletocutis amorpha Miettinen 11038 FN907913 FN907913 TomSovsky & al. 2010

S. jelicii H 6002113 FJ496690 FJ496727 Tomsovsky & al. 2010

S. portcrosensis LY 3493 FJ496689 FJ496689 TomSovsky & al. 2010

S. subsphaerospora Rivoire 1048 FJ496688 FJ496688 TomSsovsky & al. 2010

Steccherinum KHL 11905 EU118668 EU118668 TomSovsky & al. 2010

fimbriatum

S. ochraceum KHL 11902 JQ031130 JQ031130 Tomsovsky & al. 2010

Stereum hirsutum NBRC 6520 AB733150 AB733325 TomSovsky & al. 2010

Truncospora ochroleuca MUCL 39726 FJ411098 FJ393865 Robledo & al. 2009

Tyromyces chioneus Cui 10225 KF698745 KF698756 Zhao & al. 2013

Xanthoporus syringae Gothenburg 1488 JN710607 JN710607 Tomsovsky & al. 2010

from the dried specimens and observed under light microscopy following Dai

(2012). Abbreviations are: KOH = 5% potassium hydroxide, CB = Cotton Blue,

CB- = acyanophilous, L = mean spore length (arithmetic average of all spores), W =

mean spore width (arithmetic average of all spores), Q = variation in the L/W ratios

among the specimens studied, n (a/b) = number of spores (a) measured from given

number (b) of specimens.

We extracted genomic DNA from dried specimens using HiPure Fungal DNA

Mini Kit II (Magen Biotech Co.) according to the manufacturer's instructions with

some modifications. A small piece (c. 30 mg) of dried fungal material was ground

to powder with liquid nitrogen. The powder was transferred to a 1.5 ml centrifuge

tube, suspended in 0.4 ml of lysis buffer, and incubated in a 65 °C water bath for 60

min, after which 0.4 ml phenol-chloroform (24:1) was added to each tube and the

suspension was shaken vigorously. After centrifugation at 13,000 rpm for 5 min, 0.3

ml supernatant was transferred to a new tube and mixed with 0.45 ml binding buffer.

The mixture was then transferred to an adsorbing column (AC) for centrifugation

at 13,000 rpm for 0.5 min. Then, 0.5 ml inhibitor removal fluid was added in AC for

centrifugation at 12,000 rpm for 0.5 min. After washing twice with 0.5 ml washing

buffer, the AC was transferred to a clean centrifuge tube, and 100 ml elution buffer was

added to the middle of adsorbed film to elute the genomic DNA. The ITS region was

amplified with primer pairs ITS5 and ITS4 (White & al. 1990). Nuclear LSU region

was amplified with primer pairs LROR and LR7 (Vilgalys 2018). The ITS was amplified

by initial denaturation at 95 °C for 3 min, followed by 35 cycles at 94 °C for 40 s, 58 °C

for 45 s, and 72 °C for 1 min, and a final extension of 72 °C for 10 min. The nLSU was

amplified by initial denaturation at 94 °C for 1 min, followed by 35 cycles at 94 °C for

108 ... Huang & al.

30 s, 48 °C 1 min, and 72 °C for 1.5 min, and a final extension of 72 °C for 10 min. The

PCR products were purified and directly sequenced at Kunming Tsingke Biological

Technology Limited Company. All newly generated sequences were deposited in

GenBank (TABLE 1).

DNA sequences were edited using Sequencher 4.6. Sequences were aligned

in MAFFT 7 (Katoh & Toh, 2008) using the “G-INS-I” strategy and manually

adjusted in BioEdit (Hall 1999). The sequence alignment (ID 24249) was deposited

in TreeBase. In Fic. 1, Heterobasidion annosum (Fr.) Bref. and Stereum hirsutum

(Willd.) Pers. were used as outgroup to root the tree following Binder & al. (2013). In

Fic. 2, Hydnophlebia chrysorhiza (Eaton) Parmasto and Phanerochaete velutina (DC.)

P. Karst. were used as outgroup to root the tree following Floudas & Hibbett (2015).

Phylogenetic analyses of the ITS+nLSU sequences were performed using maximum

parsimony, maximum likelihood, and Bayesian inference methods. Maximum

parsimony (MP) analyses followed Zhao & Wu (2017), and tree construction was

performed in PAUP* version 4.0b10 (Swofford 2002). All characters were equally

weighted and gaps were treated as missing data. Trees were inferred using the heuristic

search option with TBR branch swapping and 1000 random sequence additions. Max-

trees were set to 5000, branches of zero length were collapsed and all parsimonious

trees were saved. Clade robustness was assessed using bootstrap (BP) analysis with

1,000 replicates (Felsenstein 1985). Tree statistics tree length (TL), consistency index

(CI), retention index (RI), rescaled consistency index (RC), and homoplasy index

(HI) were calculated for each Maximum Parsimonious Tree generated. Sequences

were analyzed using Maximum Likelihood (ML) with RAxML-HPC2 through the

Cipres Science Gateway (Miller & al. 2009). Branch support (BS) for ML analysis was

determined by 1000 bootstrap replicates.

MrModeltest 2.3 (Posada & Crandall 1998; Nylander 2004) was used to determine

the best-fit evolution model for each data set for Bayesian inference (BI). Bayesian

inference was calculated with MrBayes_3.1.2 using a general time reversible (GTR+G)

model of DNA substitution and a gamma distribution rate variation across sites

(Ronquist & Huelsenbeck 2003). Four Markov chains were run for 2 runs from

random starting trees for 8 million generations (ITS+nLSU) in Fie. 1, for 5 million

generations (ITS+LSU) in Fic. 2 and trees were sampled every 100 generations. The

first 25% of the generations were discarded as burn-in. A majority rule consensus

tree of all remaining trees was calculated. Branches that received bootstrap support

for maximum likelihood (BS), maximum parsimony (BP) and Bayesian posterior

probabilities (BPP) greater than or equal to 75 % (BP) and 0.95 (BPP) were considered

significantly supported, respectively.

Molecular phylogeny

The first ITS+nLSU dataset (Fic. 1) included sequences from 57 fungal

specimens representing 49 species of Polyporales plus the outgroup had an

aligned length of 2104 characters, of which 1236 characters were constant,

248 variable and parsimony-uninformative, and 620 parsimony-informative.

Phlebia wuliangshanensis sp. nov. (China) ... 109

ofulve

Gecaoed ochroleuca MUCL 39726

Grammothelopsis subtropica Cui 9041 {

sani ai i Wu 1006-38 core polyporiod

‘Hornodermoporus martius MUCL 41677 lad

“Perenniporia Sella ATM MUCL 49581 clade

Earliellascabrosa PR 1209

CECE caperata LE(BIN)-0677

orus tuberaster CulTENN 8976 _

Outgroup

Fic. 1. Maximum Parsimony strict consensus tree illustrating the phylogeny of Phlebia

wuliangshanensis and related species in Polyporales based on ITS+nLSU sequences. Branches

are labeled with maximum likelihood bootstrap >70%, parsimony bootstrap proportions >50%

and Bayesian posterior probabilities >0.95. Clade names follow Binder & al. (2013).

Maximum parsimony analysis yielded 3 equally parsimonious trees (TL = 4783,

CI = 0.307, HI = 0.693, RI = 0.526, RC = 0.162). Best model for this dataset

estimated and applied in the Bayesian analysis: GTR+I+G, lset nst = 6, rates

= invgamma; prset statefreqpr = dirichlet (1,1,1,1). Bayesian analysis and ML

analysis produced a similar topology as MP analysis (average standard deviation

of split frequencies = 0.003241 (BI). The tree clustered the 57 polypore species

into seven major clades, placing our new species, Phlebia wuliangshanensis, in

the phlebioid clade (Fie. 1).

The second ITS+nLSU dataset (Fic. 2) comprising sequences from 51 fungal

specimens representing 22 Phlebia species plus the outgroup had an aligned

length of 2102 characters, of which 1588 characters were constant, 142 variable

110... Huang & al.

100/100/1.00

90/89/0.99

99/100/1.00

98/100/1.00.

-/6510.98 81/100/1.00

84/78/0.99

-/65/0.99

Phlebiafloridensis HHB-6466

Phlebia floridensis HHB-7175

Phlebiafloridensis HHB-9905

Phlebiafloridensis FP-102562-T

Phlebia acerina FD-301

Phlebia acerina HHB-11146

Phlebia acerina FP-135252

Phlebia acerina DR-60

Phlebiaradiata AFTOL 484

Phlebiaradiata F 19726

Phiebia radiata FD-85

Phlebia radiata FD-121

Phlebia rufa HHB-14924

Phlebia lindtneri GB-501

100/99/1.00 |Phlebia setulosa HHB-6891

-/79/-

91/90/1.00_| 'Phiebiasetulosa AH 31879

Phlebiasetulosa PH 11749

Phlebia centrifuga L-15541

-/65/0.99

99/100/1.00 | ' Phlebia centrifuga HHB-9239

Phlebia centrifuga GB-1013

93/90/1.00

Phlebiahydnoidea HHB-1993

Phlebianantahaliensis HHB-2816

Phlebia fuscoatra HHB-10782

g99/100/1.00 -PAlebia fuscoatra HHB-15354

Phlebia fuscoatra HHB-18642

Phlebia fuscoatra FP-102173

Phlebia nothofagi HHB-12067

Phlebianothofagi HHB-6906

Phlebia nothofagi HHB-4273

Phlebia aurea RLG-5075

94ni00.00| 'Phlebia aurea NH-14434

Phlebia aurea FCUG 2767

Phlebia aurea DLL2011-100

90/82/-

98/100/1.00

-/70/-

-/69/-

Phlebiasubserialis FCUG 1434

Phlebiauda FP-101544

Phlebiauda FCUG 2452

Phlebiauda USDA Kropp-1

Phlebia chrysocreas HHB-6333

'Phlebia chrysocreas HB-3946

100/100/1.00 -Phlebia chrysocreas FP-102161

Phlebia chrysocreas KUC 20121123-24

Phlebia wuliangshanensis CLZhao 3639

Phlebia wuliangshanensis CLZhao 3475

Phlebia wuliangshanensis CLZhao 3645

Phlebia ailaoshanensis CLZhao 4036

Phlebia ailaoshanensis CLZhao 3996

Phlebialudoviciana FD-427

Phlebia subochracea HHB-8715

Phlebia acanthocystis FP 150571

Hydnophlebia chrysorhiza FD-282

Phanerochaete velutina HHB-15343

100/100/1.00

-/6910.99

100/100/1.00

100/100/1.00 100/100/1.00

/100/1.00

99/100/1.00

85/81/1.00

Fic. 2. Maximum parsimony strict consensus tree illustrating the

phylogeny of Phlebia

wuliangshanensis and related species in Phlebia based on ITS+nLSU sequences. Branches are

labeled with maximum likelihood bootstrap >70%, parsimony bootstrap proportions >50%

and Bayesian posterior probabilities >0.95.

and parsimony-uninformative, and 372 parsimony-informative. Maximum

parsimony analysis yielded 10 equally parsimonious trees (TL = 1113, CI =

0.518, HI = 0.482, RI = 0.873, RC = 0.403). Best model

for the ITS+nLSU

dataset estimated and applied in the Bayesian analysis: GTR+I+G, lset nst = 6,

Phlebia wuliangshanensis sp. nov. (China) ... 111

rates = invgamma; prset statefreqpr = dirichlet (1,1,1,1). Here also Bayesian

analysis and ML analysis generated a similar topology as MP analysis (average

standard deviation of split frequencies = 0.002221 (BI). The phylogeny

(Fic. 2) inferred from the combined ITS+nLSU sequences from 49 Phlebia

isolates grouped the new species with P chrysocreas (Berk. & M.A. Curtis)

Burds. and P. uda (Fr.) Nakasone.

Taxonomy

Fic. 3. Phlebia wuliangshanensis (holotype, SWFC 003475). Scale bar = 1 cm.

Phlebia wuliangshanensis C.L. Zhao, sp. nov. Figs 3, 4

MB 830801

Differs from Phlebia chrysocreas by its narrower ellipsoid basidiospores and from

P. uda by its smooth to tuberculate hymenium and generative hyphae with simple septa.

Type: China. Yunnan Province: Puer, Jingdong County, Wuliangshan National Nature

Reserve, on angiosperm trunk, 2 October 2017, CLZhao 3475 (Holotype, SWFC

003475; GenBank MK881787, MK881897).

Erymo.ocy: ‘The specific epithet wuliangshanensis (Lat.) refers to the locality

(Wuliangshan) of the type specimen.

BASIDIOMATA annual, resupinate, easily separable from the substratum,

ceraceous to gelatinous, without odor or taste when fresh, becoming

membranaceous upon drying, <12 cm long, 200-700 um thick. Hymenial

112... Huang & al.

gts

A B

20,4

Ot Ok

o) 0 £03

: +l y tN C

: es, | : i |

is Oper i aaa

Paley)

(| py Sor se

\/ vu pro VP >Q }

\2\ i? .. “aff

a, | Losey] fa fe (SS,

J) ge is i % jp ?

es Ret) | bes

Ip S p ! tS

Fic. 4. Microscopic structures of Phlebia wuliangshanensis (drawn from the holotype, SWFC

003475). A. Basidiospores; B. Basidia and basidioles; C. Cystidia; D. Basidiocarp section;

E. Hymenium section; F Hyphae from subiculum. Scale bars: A = 5 um; B-F = 10 um.

Phlebia wuliangshanensis sp. nov. (China) ... 113

surface smooth to tuberculate, white to cream to pale brown when fresh,

cream to pale brown upon drying. Sterile margin distinct, white.

HyYPHAL STRUCTURE monomitic; generative hyphae with simple septa,

negative in Melzer’s reagent, CB-; tissues unchanged in KOH.

SUBICULUM generative hyphae hyaline, thin- to thick-walled, branched,

3-5.5 um in diam.

HyYMENIUM cystidia of two kinds: 1) hyaline, cylindrical, numerous, thick-

walled, strongly encrusted, 20-55 x 6-10.5 um; 2) hyaline, lanceolate, few,

thin-walled, 12-18 x 3-4.5 um; basidia barrel-shaped, with four sterigmata

and a simple basal septum, 10-16 x 4-5 um; basidioles dominant, in shape

similar to basidia, but slightly smaller.

BASIDIOSPORES narrowly ellipsoid to ellipsoid, more or less curved,

hyaline, thin-walled, smooth, negative in Melzer’s reagent, CB-, 5-6(-6.5) x

3-3.7 um, L = 5.42 um, W = 3.32 um, Q = 1.64-1.73 (n = 180/3).

TYPE OF ROT: white.

ADDITIONAL SPECIMENS EXAMINED: CHINA. YUNNAN PROVINCE. Puer: Jingdong

County, Wuliangshan National Nature Reserve, on angiosperm trunk, 2 October

2017, CLZhao 3639 (SWFC 003639; GenBank MK881788, MK881898), CLZhao 3645

(SWFC 003645; GenBank MK881789, MK881899).

Discussion

We describe a new species, Phlebia wuliangshanensis, based on phylogenetic

analyses and morphological characters.

Previously, seven clades were found in Polyporales: /antrodia, /core polyporoid,

/fragiliporia, /gelatoporia, /phlebioid, /residual polyporoid, and /tyromyces

(Binder & al. 2013, Zhao & al. 2015). Our combined ITS+nLSU sequence

analysis (Fic. 1) strongly supports Phlebia wuliangshanensis (100% BS,

100% BP, 1.00 BPP) within the phlebioid clade and related to the type

species, P. radiata.

Phlebia wuliangshanensis is closely related to P. chrysocreas and P. uda in

the ITS+nLSU phylogenetic tree (Fic. 2). Morphologically, P chrysocreas

differs from P. wuliangshanensis by having ochraceous-buff to yellow ochre

color hymenophore and narrowly ovoid basidiospores (4-6 x 2-2.5 um,

Lombard & al. 1975), while P uda presents an odontoid hymenophore,

generative hyphae bearing clamp connections, and smaller basidiospores

(5-5.5 x 2-2.5 um, Bernicchia & Gorjon 2010).

Phlebia wuliangshanensis morphologically resembles other Phlebia

species: P. bispora (Stalpers) Nakasone, P capitata Bernicchia & Gorjon,

P. coccineofulva Schwein., P. femsjoeensis (Litsch. & S. Lundell) J. Erikss.

114... Huang & al.

& Hjortstam, P. livida (Pers.) Bres., P. nothofagi (G. Cunn.) Nakasone,

P. radiata, P. rufa (Pers.) M.P. Christ., P segregata (Bourdot & Galzin)

Parmasto, and P. subserialis (Bourdot & Galzin) Donk. These species differ

from P. wuliangshanensis as follows: P. bispora by a hydnoid hymenophore,

dimitic hyphal system, and smaller basidiospores (4—5 x 2.5-3 um, Nakasone

2002); P. capitata by an odontoid hymenophore, generative hyphae with

clamps, and the presence of capitate cystidia (Bernicchia & Gorjén 2010);

P. coccineofulva by a granular hymenophore with vivid yellow radiating

margin and metuloid cystidia (Schweinitz 1832); P femsjoeensis by orange

to violaceous basidiocarps (Eriksson & al. 1981); P livida by its reddish

hymenophore and a monomitic hyphal system with generative hyphae with

clamps (Bernicchia & Gorjén 2010); P nothofagi by effused basidiocarps

with a hydnoid hymenophore, a monomitic hyphal system with generative

hyphae with clamps, and narrower basidiospores (4—6 x 2-3 um, Nakasone

1997); P. rufa by its effused basidiocarps with the reticulate or subporoid

hymenophore (Christiansen 1960); P segregata by its smooth hymenophore

and cylindrical basidiospores (6-7 x 2-2.5 um, Parmasto 1967); and

P. subserialis by a white hymenium, clamped generative hyphae, and

allantoid basidiospores (6-7 x 1.5-2 um, Bernicchia & Gorjon 2010).

Although wood-rotting fungi are an extensively studied group in

Basidiomycota (Gilbertson & Ryvarden 1987, Nufiez & Ryvarden 2001,

Bernicchia & Gorjoén 2010, Dai 2012, Ryvarden & Melo 2014, Dai & al. 2015),

Chinese wood-rotting fungi diversity is still not well known, especially

in the subtropics and tropics. The new species Phlebia wuliangshanensis

is from the Chinese subtropics, where many new taxa in the Polyporales

have been described (Cui & al. 2007, 2009, 2010, 2011; Cui & Dai 2008;

Du & Cui 2009; Li & Cui 2010; He & Li 2011; Jia & Cui 2011; Yu & al. 2013;

Yang & He 2014; Chen & al. 2015; Zhao & Wu 2017; Zhao & Ma 2019).

We anticipate that more new polypore taxa will be found in China after

further investigations and molecular analyses.

Acknowledgments

Special thanks are due to Jason Karakehian (Harvard University, USA) and

Dr. Mei-Ling Han (Langfang Normal University, P.R. China) who reviewed the

manuscript. The research was supported by the National Natural Science Foundation

of China (Project No. 31700023) and the Key Laboratory of State Forestry

Administration for Highly Efficient Utilization of Forestry Biomass Resources in

Southwest China (Southwest Forestry University) (Project No. 2019-KF10).

Phlebia wuliangshanensis sp. nov. (China) ... 115

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MY COTAXON

January-March 2020—Volume 135, pp. 119-130

https://doi.org/10.5248/135.119

Pseudocercospora seropedicensis &

P. solani-cernui spp. nov. on Solanum from Brazil

KERLY M. ANDRADE, PETER S. MEDEIROS, JESSICA REMBINSKI,

JUCIMAR M. OLIvErRA, Cartos A. INACIO

Plant Pathology Sector, Dept? de Entomologia e Fitopatologia,

Inst. Ciéncias Biologicas e da Saude, Universidade Federal Rural do Rio de Janeiro,

BR 465 - Km 7 - Campus, Rural Zone, 23851-970, Seropédica, Rio de Janeiro, Brazil

* CORRESPONDENCE TO: inacio@ufrrj.br

ABSTRACT— Using stereo- and light microscopy, two leaf-spotting cercosporoid fungi,

collected from areas of the Brazilian Atlantic Forest in the states of Minas Gerais and Rio

de Janeiro, were shown to represent two new species (Pseudocercospora seropedicensis on

Solanum asperum and P. solani-cernui on S. cernuum), which are herein described and

illustrated.

Key worps—cercosporoid fungi, hyphomycetes, morphology, mycodiversity, Solanaceae

Introduction

Cercosporoid fungi, which are found worldwide, are especially abundant

and diverse in tropical and subtropical areas of Africa, Asia, Australia, and

Central and South America (Beilharz 2002; Braun & Freire 2003, 2004, 2006;

Castaneda & Braun 1989; Hernandez-Gutiérrez & Dianese 2008, 2009; Inacio

& al. 1996; Silva & al. 2016). These fungi have been studied by several authors in

Brazil (Batista & al., cf. Silva & Minter 1995; Braun & Freire 2004; Chupp 1954;

Crous & Braun 2003; Crous & Wingfield 1997; Crous & al. 1999; Fernandes &

al. 2013; Hernandez-Gutiérrez & Dianese 2008, 2009; Hernandez-Gutiérrez &

al. 2014, 2015; Inacio & Dianese 1998, 1999, 2006; Inacio & al. 1996; Pereira &

Barreto 2006; Silva & al. 2016; Viégas 1945). They have been reported as plant

pathogens on several hosts including different economically important crops,

120 ... Andrade & al.

and some cercosporoid fungi are also used for biological control of weeds

(Farr & Rossman 2019, Groenewald & al. 2013, Hanada & Gasparotto 2002,

Mendes & Urben 2019, Mendes & al. 1998, Pereira & Barreto 2006, Pons 1987,

Spegazzini 1910).

Pseudocercospora Speg. (introduced by Spegazzini 1910 for the type species

P. vitis (Lév.) Speg.) contains endophytic, pathogenic, or saprotrophic species,

some of which are used for weed control (Breeyen & al. 2006). The genus now

comprises synnematal fungi previously assigned to Phaeoisariopsis Ferraris

(Ferraris 1909) and fungi with slightly pigmented conidiogenous structures

and with paracercospora-like loci, although most Pseudocercospora species are

characterized by unthickened, non-pigmented conidiogenous loci and conidial

hila (Braun 1995, Deighton 1976). Sexual morphs are unknown for most

species, but when present they are mycosphaerella-like, previously allocated in

Mycosphaerella, now a heterotypic synonym of Ramularia Unger (Braun & al.

2013, Crous & al. 2013).

Solanaceae comprises about 3000 species in 150 genera, distributed primarily

in tropical and subtropical South America. In Brazil there are 350 species

in 32 genera (Souza & Lorenzi 2008). Solanum, the most abundant genus,

encompasses about 1500 species and occurs in tropical and subtropical areas

worldwide (Sarmento-Silva & al. 2003). Several economically important species

also belong in Solanaceae: pepper (Capsicum), potato (Solanum tuberosum),

tomato (S. lycopersicum), and tobacco (Nicotiana tabacum) (Martins & Costa

1999). Cercosporoid fungi occur on Solanum species (Silva & al. 2016), and

recently Braun (2017) published an identification key for Pseudocercospora

species on Solanum hosts with descriptions and illustrations for some species.

Here we describe and illustrate two new Pseudocercospora species found on

leaves of Solanum asperum and S. cernuum.

Materials & methods

Plant material showing necrotic leaf spots was collected during July 2013-October

2017 in the States of Rio de Janeiro (UFRRJ campus in Paracambi Natural Park and the

National Forest in Seropédica) and Minas Gerais (town of Liberdade).

Fresh and dried symptomatic leaves were examined with a dissecting microscope

to select fungal structures for mounting on slides with Cotton Blue / lactoglycerol or

phloxine / KOH glycerol. Material was also hand-sectioned and mounted on slides.

Tissues were microscopically observed and photographed using an Olympus BX41

light microscope coupled to a Canon Power Shot ELPH 100 HS camera. Structures

were measured using a calibrated ocular micrometer. The exsiccates were registered

and deposited in the Plant Pathological Herbarium “Verlande Duarte Silveira’ at the

Rural University of Rio de Janeiro (UFRJ).

TABLE 1. Selected Pseudocercospora species on Solanum

SPECIES

P. atromarginalis

Pseudocercospora seropedicensis & P. solani-cernui spp. nov. (Brazil) ... 121

Hosts

Solanum spp.;

S. americanum,

S. lycopersicum

DISTRIBUTION

Cuba, Taiwan,

Korea, USA

REFERENCES

Atkinson (1892),

Castafieda & Braun (1989),

Choi et al. (2015), Deighton (1976),

Guo & Hsieh (1995)

P. marcelliana S. nudum, Spain, Venezuela Chupp (1954),

S. torvum var. Crous & Braun (2003),

hartwegianum, Braun (2017)

S. micranthum

P. seropedicensis S. asperum Brazil This work

P. solani-cernui S. cernuum Brazil This work

P. solani- S. pseudocapsicum Brazil Silva et al. (2016)

pseudocapsicicola

P. solani-torvicula S. torvum Taiwan Guo & Hsieh (1995)

P. trichophila Solanum spp. Asia, Central and Stevens (1917),

var. trichophila South America, Chupp (1954),

North America, Braun (2017)

Oceania

P. trichophila S. hirtum Venezuela Braun & Urtiaga (2012)

var. punctata

P. venezuelae Solanum sp., Brazil, Venezuela, Chupp (1954),

S. argenteum, South America Crous & Braun (2003),

S. “aculeatum’, Braun & Freire (2003; 2006),

S. aethiopicum Braun (2017)

[= S. gilo]

Taxonomy

Pseudocercospora seropedicensis Andrade, Medeiros & Inacio, sp. nov. PL. i

MB 819383

Differs from Pseudocercospora solani-asperi |= P. trichophila var. trichophila] by its much

longer conidia and conidiophores and its larger, amphigenous. stromata.

Type: Brazil, Rio de Janeiro, Seropédica, road to cattle farm, near Instituto de Florestas,

22°45'31”S 43°41'57”W, on leaves of Solanum asperum Rich. (Solanaceae), 07 May 2016,

C. A. Inacio 122, (Holotype, UFRJ 12.390).

ErymMoLocy: honoring the municipality, Seropédica, where the species was first

discovered.

LESIONS <50 mm diam., epiphyllous, sparse, sometimes gregarious, confluent,

circular to irregular, reddish brown to light brown in the center, later becoming

necrotic, grayish, with dark brown border. CoLoNiIEs amphigenous, mainly

hypophyllous, caespitose, sparse. SUPERFICIAL MYCELIUM colorless to pale

brown, septate, branched, amphigenous, mostly hypophyllous, sometimes

emerging through the stomata, bearing conidiophores; hyphae 2-4(-7) um

122 ... Andrade & al.

diam.; CONIDIOPHORES 5-42(-45) x 3-4 um, 1-septate, sometimes curved,

slightly geniculate due to sympodial proliferation, not branched, borne

terminally or (usually) laterally on superficial hyphae, olive to olive brown,

rounded at apex. INTERNAL MYCELIUM colorless to olivaceous brown or pale

brown, septate, branched, penetrating deep into the mesophyll and spreading

through leaf tissue; hyphae 2-5(-6) um diam. STROMATA 25-80(-85) x

20-140(-160) um, brown, amphigenous, subepidermal, erumpent, textura

angularis; cells 4-7 um wide, giving rise to a fascicle of conidiophores;

CONIDIOPHORES 20-70 x 1-4 um, 1—-5-septate, numerous, macronematous,

mononematous, olive to olive brownish, smooth, densely caespitose, arising

from stromata, straight or slightly sinuous. CONIDIOGENOUS CELLS integrated,

polyblastic, usually monoblastic and terminal, sympodial, geniculate, with

inconspicuous conidiogenous loci. Conrp1A solitary, 20-145(-226) x 2-6 um,

1-16-septate, straight, sometimes sinuous to curved, subcylindrical to

obclavate, smooth, hyaline, subhyaline to olivaceous brown, attenuated at apex,

obconically truncate at base, hila neither thickened not darkened, 1-2 um wide.

COMMENTS—Several Pseudocercospora species on solanaceous hosts are

morphologically close to P. seropedicensis. Pseudocercospora atromarginalis

(G.F. Atk.) Deighton (Deighton 1976), P marcelliana (Chupp) U. Braun &

Crous (Crous & Braun 2003; Braun 2017), P. solani-torvicola Goh & W.H.

Hsieh, P. trichophila var. trichophila (F. Stevens) U. Braun [as P. solani-asperi

(R.E.D. Baker & & W.T. Dale) Deighton] (Stevens 1917; Baker & Dale 1951;

Chupp 1954; Deighton 1976; Crous & Braun 2003; Braun 2017), P. trichophila

var. punctata U. Braun & Urtiaga, P. venezuelae (Chupp) Deighton, and P

solani-pseudocapsicicola Meir. Silva & al. are compared in TaBLEs 1-2.

Pseudocercospora atromarginalis lacks superficial mycelia with secondary

conidiophores; collections from Taiwan with conidiophores (10-65 x 3-5

um/0-4-septa) and close to our specimen differed in lacking stromata

(Guo & Hsieh 1995, Choi & al. 2015). Pseudocercospora marcelliana differs

by shorter conidia (15-70 um) and conidiophores (5-25 um), an absence

of superficial mycelium, and occurrence on other hosts or in different

locations (Braun 2017; Crous & Braun 2003; Chupp 1954, as Cercospora).

Pseudocercospora venezuelae differs by shorter conidiophores (10-70 um),

conidia (15—)30-65 wm), and lacks superficial mycelium; and P solani-

torvicola differs by smaller (<30 um) stromata, shorter and wider primary

conidiophores (35-60 x 4.0-5.5 um), shorter secondary conidiophores

(10-25 um), and shorter conidia (20-115 um) with fewer septa (1-11).

Pseudocercospora seropedicensis & P. solani-cernui spp. nov. (Brazil) ... 123

PLATE 1. Pseudocercospora seropedicensis (holotype, UFRJ 12.390) on leaves of Solanum

asperum. A. Branch of S. asperum. B. Leaf with lesions. C. Detail of fructifications on lesions.

D-EF Conidiophores and conidia as seen under dissecting microscope. G-I. Stroma (vertical

section). J. Secondary mycelia on leaf undersurface. K-M. Conidia. Scale bars: A= 5 cm; B= 1 cm;

C= 1 um; D-I = 20 um; J, K= 10 um; L, M=5 um.

124 ... Andrade & al.

Pseudocercospora trichophila var. punctata has smaller dark brown to blackish

stromata (10-60 um diam) (Braun 2017). Finally, although P solani-asperi

[= PB. trichophila var. trichophila] (Braun 2017) has been reported in Brazil

on the same host species (Chupp 1954 as Cercospora solani-asperi, Crous &

Braun 2003, Deighton 1976 as Pseudocercospora solani-asperi, Fernandes &

al. 2013), it produces shorter 20-110(165) conidia with fewer 1-10(15) septa;

P. solani-asperi also usually shows hypophyllous caespituli and lacks a stroma,

clearly distinguishing it from the new species found now in the Atlantic

Forest. Pseudocercospora solani-pseudocapsicicola, a new species recently

reported on leaves of Solanum pseudocapsicum from Brazil (Silva & al. 2016),

clearly differs in lacking stromata, shorter (42-128 um) conidia with fewer

(2-6) septa, and hypophyllous conidiophores.

Pseudocercospora seropedicensis is the first report of a Pseudocercospora

species on leaves of Solanum asperum in the State of Rio de Janeiro and the

second in Brazil.

ADDITIONAL SPECIMENS EXAMINED—BRAZIL. RIO DE JANEIRO: Paracambi, Parque

Natural Municipal do Curid, 22°35’44”S 43°42’18”W, 9.XII.2013, C. A. Inacio 61, (UFRJ

12015); 19.III.2014, C. A. Inacio 74 (UFRJ 12050). Seropédica, Campus UFRRJ, Jardim

Botanico near the road to Agricultural Institute, 22°45’53”S 43°41’35”W, 11.VHI.2017,

J. M. Oliveira 32 (UFRJ 12.495); 11.VIII.2017, J. D. Almeida 03 (UFRJ 12.496); near

Bus Station in front of Female Students Dormitory, 22°45’59”S 43°41’24”W, 22.1X.2017,

J. Rembinski 35, J. (UFRJ 12.594); National Forest (Fiona), 22°43’46”S 43°42’36”W,

29.V1.2013, C. A. Inacio 79 (UFRJ 12064).

Pseudocercospora solani-cernui Rembinski, Oliveira & Inacio, sp. nov. Pl. 2

MB 824359

Differs from P. seropedicensis by its narrower conidiophores, its larger stromata, and its

lack of secondary mycelium.

Type: Brazil, Minas Gerais, Liberdade, Sitio Carcara, Zona Rural, Bairro Taquarucu-

Rodovia Joanito Balieiro Km 9, 22°04’26”S 44°23’48”W, on leaves of Solanum cernuum

Vell. (Solanaceae); 14.Oct. 2016, J.C. Oliveira 29 (Holotype, UFRJ 12.422).

EryMoLoey: referring to the host species, Solanum cernuum.

LESIONS <25 mm diam., amphigenous, mainly epiphyllous, sparse,

sometimes confluent, circular, later irregular, initially visible as yellowish

spots becoming reddish brown to light brown or grayish brown in the

center with yellowish margin at the adaxial face and shown as pale areas

PLATE 2. Pseudocercospora solani-cernui (holotype, UFRJ 12.286) onleaves of Solanum cernuum.

A, B. Affected leaf (A: upper surface; B: under surface). C. Leaf spots on upper surface. D.

Fructifications on upper surface. E. Conidia and conidiophores seen under stereomicroscope.

Pseudocercospora seropedicensis & P. solani-cernui spp. nov. (Brazil) ... 125

FE. Stroma with conidiophores showing textura angularis (vertical section). G. Stromata

(vertical section). H. Stroma and conidiophores. I-K. Conidia. Scale bars: A, B = 5 cm;

C=10mm; D =5 mm; E£, F = 20 um; G = 50 um; H = 5 um; I, J =10 um; K = 5 um.

126 ... Andrade & al.

at the abaxial face of the leaf, becoming slightly brownish at the center,

corresponding to the upper leaf spots. CoLonigs epiphyllous, caespitose,

sparse. MycELium: internal, colorless to pale brown, septate, branched,

penetrating the mesophyll; hyphae 2-4 um diam. Stromata 10-70 x

30-90(-150) um, brown, epiphyllous, subepidermal, erumpent, textura

angularis, with brownish angular cells; cells 4-7 um wide, forming a fascicle

of conidiophores. CONIDIOPHORES 20-46 x 3-6 um, 1-3-septate, numerous,

macronematous, mononematous, brown, smooth, densely caespitose,

arising from stromata, straight or slightly sinuous. CONIDIOGENOUS

CELLS integrated, polyblastic, mostly monoblastic and usually terminal,

sympodial, geniculate, with inconspicuous conidiogenous loci. CONIDIA

solitary, 30-145 x 2-5 um, 2-12-septate, straight, sometimes sinuous

to curved, subcylindrical to obclavate, smooth, subhyaline to brown

olivaceous, attenuated at the apex, obconically truncate at the base, hila

neither thickened nor darkened, 1-2 um diam.

TABLE 2. Morphological comparison of selected Pseudocercospora species on Solanum

SPECIES STROMATA (um) CONIDIOPHORE/ —- CONIDIA (tm) / REFERENCES

SEPTA (um) SEPTA

P. atromarginalis Absent or only a 15-65(100) x 20-95 x Guo & Hsieh (1995)

few brown cells 3-5 / 0-4 3.5-5 / 3-11

P. marcelliana 15-75 5-30 x 15-110 x Braun (2017)

1.5-4/ 1-2 2-357 (3-14

P. seropedicensis 25-80 (-85) x 1’—20-70 x 20-140(226) x This work

20-140 (-160) 1-4/ 1-5; 2-6 / 1-16

2’—5-42(45) x

3-5/ 0-1

P. solani-cernui 10-60 x (15)30-46 x 30-145 x This work

30-90 (-150) 2-4 / <3 2-5 / 2-12

P. solani- Lacking 10-35 x 42-128 x Silva et al. (2016)

pseudocapsicicola 3-5 / 0-3 2-3.5 / 2-6

P solani-torviculaz <30 1’—35-60 x 20-115 x Guo & Hsieh (1995)

4.0-5.5 / 1-3 4.5-6/ 1-11

2’—10-25 x

4-5/ 0-1

P. trichophila 10-60 10-70 x (10)20-90(110) x Braun & Urtiaga

var. punctata 3-7 / 0-4 (2.5)3-5 (6) / (2012)

(1)2-10(12)

P. trichophila Lacking to few 5-80 x 20-110(165) x Braun (2017)

var. trichophila swollen hyphal 2.5-5 / 0-4 3-5(7) / 1-10(15)

cells

P. venezuelae 15-40 10-70 x (15-)30-65 Braun (2017)

2-4 / 0-1(2) x 3-5 / (0-)3-6

Pseudocercospora seropedicensis & P. solani-cernui spp. nov. (Brazil) ... 127

CoMMENTS—Pseudocercospora solani-cernui is closely related to P. trichophila

var. trichophila, P. solani-pseudocapsicicola, and P. seropedicensis (TABLES 1-2).

Although showing similar conidial dimensions (20-110(165) x 3-5(-7) um)

and occurring on various Solanum species, P. trichophila var. trichophila is

distinguished by much smaller (<5 mm) leaf spots, absence of a stroma, and

longer (45-80 um) conidiophores (Deighton 1976). Pseudocercospora solani-

pseudocapsicicola (reported on S. pseudocapsicum in Brazil) differs in lacking

stromata and narrower conidia (42-128 x 2-3.5 um) with fewer (2-6) septa

(Silva & al. 2016); P. seropedicensis exhibits similar sized stromata ((20-)140

(-160) um) but is clearly distinguished from the new species on S. cernuum by

its much longer (20-60(-70) um) conidiophores, conidia with more (1-16)

septa, and superficial mycelium bearing secondary conidiophores.

The above differences morphologically support the Pseudocercospora

specimen on S. cernuum as a new species, here designated as P. solani-cernui.

ADDITIONAL SPECIMENS EXAMINED—BRAZIL. RIO DE JANEIRO: Paracambi,

Parque Natural Municipal do Curid, 22°35’44’S 43°42’18”W, 05.11.2016, J.M. Oliveira

14 (UFRJ 12.271); 10.VI.2016, J.M. Oliveira 17 (UFRJ 12.290). Seropédica, Campus

UFRRJ, reserve in front of the Instituto de Florestas UFRRJ, 22°45’30”S 43°41'54” W,

26.X.2017, C.A. Inacio 160 (UFRJ 12.553). Minas GERAIS, Liberdade: Sitio

Carcara, Zona Rural, Bairro Taquarugu—Rodovia Joanito Balieiro Km 9, 22°04’26”S

44°23’48”W, 28.X.Oct. 2015, J.C. Oliveira 5 (UFRJ 12172).

Key to Pseudocercospora species on Solanum from Brazil

IMyceliuni dntennalarid external «x! «Sica! 9 sits wi ioltia wiseltia wibeltia weenie w podea Wied a Wie 2

DALI isCelititia COMSIStEIIU Yat RL Ra i eg Mase seo UMS nee EASE ogo VASE ogo VA go Ve ng Gea Ona 4

2 Leaf spots lacking or indistinct, stroma absent ........ P. trichophila var. trichophila

2’ Leaf spots amphigenous, stroma well developed................. P. seropedicensis

3 Stromata lacking or small as small substomatal clumps of swollen hyphae ......... 4

3° Stromata well-developed; often forming sporodochial conidiomata .............. 5

4 Conidia 42-128 um long; on S. pseudocapsicum ........ P. solani-pseudocapsicicola

4’ Conidia shorter (<110(-120) um); on Solanum spp. ............ P. atromarginalis

5 On Solanum cernuum; conidia 30-145 um long, <12-septate ...... P. solani-cernui

5° On Solanum aculeatum; conidia (15-)30-65 um long, <6-septate.... P. venezuelae

Acknowledgments

The authors acknowledge CAPES, CNPq, FAPERJ for financial support and EMBRAPA

and UFRRJ for facilities. Thanks are also given to Dr. Uwe Braun (Martin Luther

University, Germany), Dr. Paul Kirk (Royal Botanical Garden Kew, UK), and Prof. José

Dianese (Universidade de Brasilia, Brazil) for reviewing our manuscript and to Jonas

Dias de Almeida (UFRRJ) and Ernandes Silva Barbosa (UFRRJ) for technical assistance.

128 ... Andrade & al.

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MYCOTAXON

January-March 2020—Volume 135, pp. 131-141

https://doi.org/10.5248/135.131

The nomenclatural history of Umbilicaria spodochroa

and nomenclatural corrections in Umbilicariaceae

Evceny A. Davybov’, TEUvo AHTI’, ALEXANDER N. SENNIKOV”?

‘Altai State University, Lenin Avenue, 61, Barnaul, 656049, Russia

*Botanical Museum, Finnish Museum of Natural History,

PO. Box 7, 00014 University of Helsinki, Finland

*Herbarium, Komarov Botanical Institute of Russian Academy of Sciences,

Prof. Popov str. 2, 197376 St. Petersburg, Russia

* CORRESPONDENCE TO: eadavydov@yandex.ru

ABSTRACT— The name Umbilicaria spodochroa is currently applied to a species with an

oceanic distribution in Europe and East Asia. The upper surface of its thallus is grey to

dark brown, apothecia are omphalodiscs with prominent central umbilicus-like buttons.

Its designated type and other original material are referred to U. hirsuta. Conservation is

required to retain this name in current use. The nomenclatural history of U. spodochroa is

presented to serve as the background for its conservation. The subgeneric nomenclature

of Umbilicaria is revised and one new name (U. subg. Papillophora) is proposed to

replace the illegitimate U. subg. Gyrophora. The status of many new names published by

G.E. Hoffmann in his “Deutschlands Flora” (1796) is discussed and their nomenclatural

validity is supported.

Key worps—conservation, Ehrhart, historical collections, Hoffmann, typification

Introduction

Species of Umbilicariaceae Chevall. are predominantly saxicolous

lichens mostly found in regions of higher latitudes or altitudes worldwide.

Multilocus phylogenies resulted in a new generic concept of the family,

which currently includes three genera comprising together about one

hundred species with mostly umbilicate growth habit (Bendiksby & Timdal

2013, Davydov & al. 2017).

132 ... Davydov, Ahti, Sennikov

Morphological details of the upper and lower thallus surfaces as well

as traits of the rhizinomorphs were mostly taken into consideration by

earlier lichenologists to distinguish species of Umbilicaria Hoftm. However,

these characters have not always been easily recognizable. For this reason,

the circumscriptions of some of the early described species have been

controversially discussed. One species with a rather complicated taxonomic

and nomenclatural history is Umbilicaria spodochroa which, in the 18" and 19"

centuries, was often confused with another early described species, U. hirsuta

(Sw.) Ach.

Umbilicaria spodochroa, with an oceanic distribution in Europe and East Asia

(Wei & Jiang 1993), is characterized by a large thallus with a grey to dark brown

upper surface and pale brown to black papillate lower surface with abundant

rhizinomorphs and characteristic omphalodisc-type apothecia with prominent

central umbilicus-like buttons. By such characterization, this species is easily

recognizable, and specimens thereof were frequently distributed in exsiccatal

series (see the list in Llano 1950).

Umbilicaria hirsuta, a Holarctic species, is characterized by producing

parasoredia on the marginal part of the upper thallus surface, unlike the

closely related U. grisea Hoffm. in which the parasoredia clump and become

farinose with variously sized granules (Codogno & al. 1989). The lower surface

of U. hirsuta is beige to almost black with sparse to abundant rhizinomorphs.

Apothecia, which are rarely produced, belong to the gyrodisc type.

As currently defined (Frey 1933, Llano 1950, Codogno &al. 1989, Wei & Jiang

1993), Umbilicaria hirsuta and U. grisea can be unambiguously distinguished

by their diagnostic characters as stated above. Both species names are currently

accepted (Poelt & Vézda 1981, Hitch & Purvis 2009, Stenroos & al. 2016),

and the species were assigned to the recently resurrected “Umbilicaria subg.

Gyrophora (Ach.) Frey” (Davydov & al. 2017), which corresponds to a rather

large clade of 13 phylogenetically analysed species. The core of this clade is

constituted by the Umbilicaria vellea group, which includes U. spodochroa and

U. hirsuta.

In spite of the current advances in the taxonomy of Umbilicariaceae,

the nomenclature of several species is still to be clarified. In the present

contribution we aim at unravelling the old confusion concerning the species

name Umbilicaria spodochroa.

Figure 1. Type collection of Umbilicaria spodochroa, voucher from Gottingen (GOET 019934).

A. printed label of Ehrhart’s exsiccata; B. upper surface of thallus; C. lower surface of thallus.

Photo: Marc Appelhans.

Umbilicari i

ia subg. Papillophora subg. nov. & U.

. NOV. . spodochroa .

oettingen

j | Herb. Goett

wna Mte

316. Lichen fpadochrous Ehrh. j

Upfalia 320. Spherocephalus {

7 Herrenhaufit.

ee —

a Wn jplitarra Pinsnda (Fe 7

133

134 ... Davydov, Ahti, Sennikov

Materials & methods

The protologue of Umbilicaria spodochroa and the relevant historical literature was

examined to uncover the history and the original material of the name. Herbarium

collections and high resolution digital photographs of Umbilicaria from GOET, H,

LE, LINN, and MW were studied de visu, via online portals (https://plants.jstor.org),

or provided by curators. Taxonomic literature was screened for relevant treatments.

Historical background

A species of Umbilicaria with the specific epithet “spodochrous” was

first introduced by Ehrhart (1793), who distributed a specimen named

“Lichen spadochrous Ehrh?” among his exsiccatae (Fic. 1). The specimen was

accompanied with a printed label, but no description or diagnosis of the species

was provided.

The species epithet is controversial because in its original spelling it is

meaningless and has long been considered linguistically erroneous (e.g., Schade

1955). Nylander (1861: 115; 1869: 11) stated that the epithet “spadochrous”

seems to be a misspelling because it is apparently derived from the Greek word

omodoc (cinder), meaning “ashes” Since the species epithet was incorrect,

either as a typographic or orthographic error, under ICN (Shenzhen Code)

Art. 60.1 its spelling may be corrected to “spodochrous,’ which is currently in

common use.

Acharius (1794) referred Ehrhart’s specimen of Lichen “spadochrous” to

L. polyrrhizos L. This was done under a broad species concept and had no

practical influence on the further taxonomy or nomenclature.

Valid publication and protologue

Hoffmann (1796) revised species of cryptogams (ferns, mosses, and

lichens) known from Germany. Since the available knowledge was very

uneven and several taxa were not sufficiently understood at that time, he

treated the taxa differently as he explained in the Preface:

“Varietaten, Halbarten (Subspecies), auch Arten, welche ich als solche

aufzufihren noch unentschlossen war, findet man entweder in Klammern

.. den Anmerkungen, oder ohne Bezifferung der nachstverwandten Art

beigestellt” [= Varieties, subspecies, also species that I was still undecided

to list as such, are to be found either in parentheses ( ) placed with the

annotations, or, without numeration, with the most closely related species]

(Hoffmann 1796: [Vorbericht: 4]).

In the Index, according to our interpretation of this work, Hoffmann

(1796) implicitly listed accepted names and their basionyms in italics and

Umbilicaria subg. Papillophora subg. nov. & U. spodochroa ... 135

synonyms in the regular font. In the taxonomic part of his work, he listed

many species without numbers, yet with binomial names in the accepted

genus. Several of such species were new to science. In annotations placed

in brackets, Hoffmann mentioned quite a number of species names

published by previous authors, with their original generic assignments. He

also mentioned several varieties in these annotations, usually unnamed

or under old polynomials, sometimes with previously published species

names; according to the Index, such species names were listed as synonyms.

Hoffmann’s use of subspecies was very sparse, and we are not aware of any

name that he may have applied at this rank.

Isoviita (1966) considered new names in Hoffmann’s treatments of

“undecided” taxa invalidly published because of the presumed absence

of explicit acceptance by the author [ICN (Shenzhen Code) Art. 36.1].

Contrary to his opinion, we consider the internal evidence in Hoffmann

(1796) (typesetting of the Index and explanations in the Preface) to be

sufficient to dispel doubts about Hoffmann’s acceptance of such taxa.

Hoffmann (1796) was the first to provide a description for Ehrhart’s lichen

under the name Umbilicaria “spadochroa”; although this species name was

left unnumbered in the synopsis, it was listed as accepted in the Index and

therefore was validly published in spite of any doubts that Hoffmann may

have had at that time.

The species description provided by Hoffmann inadequately distinguishes

between the Umbilicaria species in their current circumscription. Hoffmann

distinguished U. spodochroa from its presumed closest relative U. hirsuta

mostly by the colour of the upper thallus surface (bluish grey vs. grey)

and the lower thallus surface (light brown vs. brownish grey), and also by

rhizinomorph density (scarce vs. abundant). These characters are variable in

both species in that the colour of the lower thallus surface varies from beige

to black-brown and the rhizinomorphs are scarce to abundant. According

to the current species concept, Hoffmann’s description fits both taxa,

U. spodochroa and U. hirsuta, so that it can be applicable either to a species

different from U. hirsuta or it may indicate a phenotypic variation within the

same species.

Further treatments

A later author who treated this lichen species was Acharius (1799), who

validly published the combination Lichen spodochrous (as “spadochrous’),

which he explicitly accepted and accompanied by a species description.

136 ... Davydov, Ahti, Sennikov

Acharius broadened the limits of this taxon and, disregarding priority,

included one previously described species, Umbilicaria cirrosa Hoftm.; this

name has not yet been typified but the figures accompanying the description

by Hoffmann (1789) suggest that it may be a synonym of U. vellea.

Acharius (1799, 1803) specified the character of the lower surface of

the thallus as dark hirsute (“subtus ater hirsutus”). Later, Acharius (1810:

673) emphasized that his Gyrophora vellea (L.) Ach., G. spodochroa, and

G. crustulosa Ach. can be optimally distinguished by their apothecial

morphologies. In this work, Acharius was the first to mention one of the

most important diagnostic characters of Umbilicaria spodochroa, i.e., the

prominent central button on the apothecia. Finally Acharius (1814) lowered

the rank of the taxon to the varietal level but maintained its diagnostic

characters.

It is rather obvious that the works of Acharius were essential in

establishing the current concept of U. spodochroa. Nevertheless, despite the

characters indicated in his descriptions, the specimens labelled by Acharius

as “Gyrophora vellea B G. spadochroa” belong to U. vellea (H-ACH 581) or

U. vellea and U. cinereorufescens (Schaer.) Frey (H-ACH 580). The specimens

referable to U. spodochroa were identified by Acharius as “Gyrophora vellea”

(H-ACH 576), in accordance with the illustration of the latter species in

Acharius (1794: Tab. II], fig. 3).

Stenhammar (1825) described the same type of apothecia, with thick

margin and a central verruca, for Umbilicaria vellea var. spodochroa, although

the respective herbarium specimen was referred to U. vellea (Merrill 1906).

The other important diagnostic characters of the species were unknown at

that time.

Nylander (1861) established an additional character that can diagnose

Umbilicaria spodochroa: ascospores that are simple and colourless to

submuriform and brown. However, he treated U. spodochroa broadly

and included other Umbilicaria taxa with submuriform ascospores:

U. cinereorufescens, U. cirrosa, U. crustulosa, and U. depressa (Ach.) Duby

(Nylander 1869).

This broad concept of Umbilicaria spodochroa prevailed until Frey (1933)

recognized the four aforementioned species as separate and provided an

artificial key and detailed descriptions, which are still accepted. Frey’s

treatment became the basis for all subsequent interpretations of U. spodochroa

and established the current application of its name.

Umbilicaria subg. Papillophora subg. nov. & U. spodochroa ... 137

Type designation

Hoffmann (1796) cited two collections in the protologue, which are

therefore syntypes. The first collection is Lichen “spadochrous” of Ehrhart

(1793), which was erroneously cited under no. 317 instead of no. 316. The

distribution of Ehrhart’s exsiccatae was so limited (Gubanov & Balandina

2000) that this incorrect citation was reproduced in the great majority of

subsequent taxonomic publications.

We were able to locate three specimens of Ehrhart’s “Lichen spadochrous”

at GOET, LINN-HS, and MW. In addition we checked B, BM, G, HAL, LE,

M, and UPS (the herbaria in which some Ehrhart’s collections are known to

be located), but without success.

The specimen at GOET (barcode 019934) was studied by Arnold (1880)

and Schade (1955), who referred it to Umbilicaria hirsuta. We agree with

this identification because of its prominent marginal farinose-granular

parasoredia.

The specimen at LINN-HS (1703.19.3) belongs to the Herbarium of Sir

James Edward Smith, to which it went through the collection of Edmund

Davall in 1802 (Beer 1947). According to the annotations, Smith referred it

to “Lichen polyrrhizos,” although its correct identity is Umbilicaria hirsuta.

The specimen at MW belongs to the personal collection of F. Ehrhart,

which was owned by Hoffmann who left its part, including sets of the

exsiccatae, to the Moscow Branch of the Military Medical-Surgical Academy,

from which the collections were transferred to the Moscow University after

the Academy was closed in 1842 (Sokoloff & al. 2002). Some specimens of

Ehrhart’s cryptogams were purchased by the Botanical Museum of the St.

Petersburg Academy of Sciences (now the Komarov Botanical Institute)

(Karavaev & Barsukova 1968) but the specimen of Lichen “spadochrous”

was left in Moscow (Gubanov & Balandina 2000). This specimen was

certainly examined by Hoffmann and apparently was the main basis for the

original description and his concept of Umbilicaria spodochroa. The traits

of his specimen do match the original description well, but it evidently also

belongs to U. hirsuta.

The second collection mentioned by Hoffmann (1796) is a specimen

of “Lichen polyrrhizos,” which was communicated by Smith. We cannot

recognise this specimen among the lichen collections of Hoffmann, which

were purchased from him by Moscow University (Hoffmann 1825). However,

a suitable specimen in Smith’s herbarium (LINN-HS 1273.212), identified as

“Lichen polyrrhizos” and originating from Ehrhart’s collection, matches the

138 ... Davydov, Ahti, Sennikov

protologue of Umbilicaria spodochroa. This specimen, which was received

by Smith most likely in 1793 (as evident from annotations on other similar

specimens in this collection, e.g. LINN-HS 39.34) and may have been shared

with Hoffmann prior to 1796, could be part of the gathering mentioned by

Hoffmann in the protologue. This specimen also belongs to U. hirsuta.

The original description of Umbilicaria spodochroa and the relevant

herbarium material convincingly demonstrate that this name was applied

by Hoffmann to a variant of U. hirsuta, not to the species known as

U. spodochroa in the current use. Arnold (1880) studied Ehrhart’s lichen

collections and stated that Ehrhart’s specimen of Lichen “spadochrous”

was mixed and the typical U. spodochroa was also present under no.

316; however, we have found no evidence for this statement. Moreover,

U. hirsuta is common in the vicinity of Uppsala (Shah & Coulson 2018), the

type locality of U. spodochroa, whereas U. spodochroa in its current concept

is absent from Uppsala proper, although it occurs fairly close to the town,

mostly along the coast and to some extent along the shores of Lake Malaren

(S. Ekman, pers. comm.).

Llano (1950: 101) rather mechanically cited “Ehrhart ... Crypt. Exs. 317”

as the type of Umbilicaria spodochroa, thus fulfilling conditions for effective

type designation [ICN (Shenzhen Code) Art. 7.11, 9.17]. This typification

is formally correct but has an undesirable effect that a familiar species

name would change its application because the type collection belongs to

U. hirsuta although the species name has been widely and persistently used

in the sense of U. spodochroa since Frey (1933). Llanos type designation is

referable to a gathering rather than a specimen, since he failed to specify the

herbarium in which the type is housed, but we refrain from the second-step

typification of the name as unnecessary in view of a conservation proposal

currently under review (Hestmark, submitted).

Umbilicaria spodochroa Hoftm., Deutschl. Fl. 2: 113. 1796, [as “spadochroa”|

= Lichen spodochrous (Hoffm.) Ach., Lichenogr. Suec.

Prodr.: 149. 1799 [“1798”; as “spadochrous”]

= Gyrophora spodochroa (Hoftm.) Ach., Methodus: 108. 1803 [as “spadochroa” |

= Gyrophora vellea var. spodochroa (Hoffm.) Ach., Syn.

Meth. Lich.: 68. 1814 [as “spadochroa”]

= Umbilicaria vellea var. spodochroa (Hoftm.) Stenh., Sched.

Crit. Lichen. Suec. 5-6: 4. 1825 [as “spadochroa” |

= Omphalodiscus spodochrous (Hoftm.) Schol., Nyt Mag. Naturvid. 75: 26. 1934

LECTOTYPE (designated by Llano 1950: 101)—Sweden. Uppsala, F. Ehrhart in Plantae

Cryptogamae Linn. no. 316 (GOET [image!], LINN-HS [image!], MW [image!]).

Umbilicaria subg. Papillophora subg. nov. & U. spodochroa ... 139

Subgeneric nomenclature

Gyrophora Ach. was published as an explicit substitute [replacement name,

ICN (Shenzhen Code) Art. 6.11] for the illegitimate Umbilicaria Hoftm.

1789 (non Fabr. 1759), even though the illegitimacy of Hoffmann’s genus

was not realized at that time (e.g., Leighton 1856). Acharius (1803: 100) cited

Hoffmann’s name in synonymy and stated that he changed the latter because

he considered it “not optimal’, thus making both names homotypic [ICN

(Shenzhen Code) Art. 7.4]. Since the generic name Umbilicaria Hoffm. was

illegitimate prior to its conservation in 1996 [ICN (Shenzhen Code) Art.

14.15], the autonym “Umbilicaria subg. Umbilicaria” cannot be established

[ICN (Shenzhen Code) Art. 22.5]; because of the illegitimacy of the generic

name, ICN (Shenzhen Code) Art. 22.2 does not apply and the combination

Umbilicaria subg. Gyrophora (Ach.) Frey was validly published for a subdivision

of the genus that includes the type of the generic name. When Davydov & al.

(2017) accepted U. subg. Gyrophora but excluded from its circumscription

the type of Umbilicaria (which is also the type of U. subg. Gyrophora (Ach.)

Frey), also providing a description of this subgenus and a type designation,

they created an illegitimate later homonym [ICN (Shenzhen Code) Art. 48.1]

which, however, was not validly published under ICN (Shenzhen Code) Art.

E5.1. This subgenus is formally named here with the same type and a reference

to the validating description in Davydov & al. (2017).

Two other subgeneric names accepted by Davydov & al. (2017) are revised

with their corrected nomenclature as follows.

Umbilicaria subg. Agyrophora Nyl. [Flora 61: 247. 1878, nom. nud.] ex Cromb.,

Monogr. Lich. Britain 1: 323. 1894

TyPE (designated by Llano 1950: 49)—Umbilicaria atropruinosa Schaer.

[= Umbilicaria leiocarpa DC.]

Umbilicaria subg. Lasallia (Mérat) Frey, Hedwigia 71: 106. 1931

= Lasallia Mérat, Nouv. Fl. Env. Paris, ed. 2, 1: 202. 1821

TypeE—Unmbilicaria pustulata (L.) Hoftm.

Umbilicaria subg. Papillophora Davydov, Ahti & Sennikov, subg. nov.

MB 830067

= “Umbilicaria subg. Gyrophora” sensu Davydov & al., Taxon 66: 1297. 2017.

TypeE—Unmbilicaria vellea (L.) Ach.

DESCRIPTION—see Taxon 66: 1297. 2017, under Umbilicaria subg. Gyrophora “(Ach.)

Frey”.

EryMoLoGy—The name refers to the papillose lower surface and rhizinomorphs, the

characteristic trait for the majority of species in the subgenus.

140 ... Davydov, Ahti, Sennikov

Acknowledgments

Norbert Kilian (Berlin) kindly commented on the complicated taxonomy

and nomenclature of Hoffmann (1796) and provided his English translation of

the quoted passage. We are grateful to Marc Appelhans (Gottingen) for scanned

images and a photocopy of the printed matter of Ehrhart’s exsiccatae from GOET.

Mikhail Kozhin (Moscow) supplied a photograph from Ehrhart’s collection at

MW. The following curators kindly reported the absence of Ehrhart’s specimens:

Stefan Ekman (UPS), Uwe Braun (HAL), Andreas Beck (M), Robert Lticking (B),

Philippe Clerc (G), Len Ellis (BM). Stefan Ekman is also thanked for information

on the occurrence of the species of Umbilicaria around Uppsala. A photograph of

Umbilicaria spodochroa from GOET is reproduced with kind permission from the

Georg-August- Universitat Gottingen. We thank Christian Printzen (Senckenberg

Research Institute Frankfurt, Germany) and Gerhard Rambold (University of

Bayreuth, Germany) for expert presubmission review.

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MY COTAXON

January-March 2020—Volume 135, pp. 143-150

https://doi.org/10.5248/135.143

Ochroconis terricola sp. nov. from China

XIN ZHANG, KUN- YING WANG, PENG-PENG REN, YU-LAN JIANG

Department of Plant Pathology, Agriculture College, Guizhou University,

Guiyang, 550025, China

*CORRESPONDENCE TO: yljchsd@163.com

ABSTRACT—A new species, Ochroconis terricola, was isolated from soil in Guizhou Province,

China. Morphology and phylogenetic analyses of the combined sequence data of nuclear

ribosomal DNA genes (ITS, LSU, SSU) revealed the strain as different from other Ochroconis

species.

Key worps—Dothideomycetes, hyphomycetes, Sympoventuriaceae, taxonomy, Venturiales

Introduction

Ochroconis, typified by O. constricta (E.V. Abbott) de Hoog & Arx, was

morphologically separated from Scolecobasidium E.V. Abbott (Abbott 1927)

by de Hoog & von Arx (1974), for species with unbranched, ellipsoidal to

cylindrical conidia, while Scolecobasidium was retained for species having

T- or Y-shaped conidia. However, the taxonomic status of Scolecobasidium

was questioned due to ambiguity of its type species, S. terreum E.V. Abbott,

and species with lobed conidia similar to S. terreum in Scolecobasidium

have been combined in Ochroconis on phylogenetic grounds (Samerpitak

& al. 2014). Historically, Ochroconis was characterized by sympodial

conidiogenesis and septate, mostly rough-walled conidia that are liberated

rhexolytically (Samerpitak & al. 2015, Ellis 1971). A recent multigene

(nSSU, nLSU, mtSSU, RPB2) phylogenetic analysis places Ochroconis in

Sympoventuriaceae (Venturiales, Dothideomycetes) (Machouart & al. 2014).

By combining molecular phylogeny, morphology, and ecology, Samerpitak

& al. (2014) revised the taxonomy of the Ochroconis lineage, introducing

144 ... Zhang & al.

Verruconis Samerp. & al. as a new genus for a group of thermophilic species

around O. gallopava (W.B. Cooke) de Hoog [= Verruconis gallopava (W.B.

Cooke) Samerp. & de Hoog]. (Ochroconis species are mesophilic.) ITS and

LSU nuclear ribosomal sequences as well as the conserved SSU gene are all

suitable for identifying species in Ochroconis and Verruconis, but the ITS

and LSU regions are recommended as the best DNA barcoding candidates,

due to the high variability found even in conserved markers (Samerpitak

& al. 2014, 2015). Presently, 31 species are accepted in Ochroconis

(http://www.mycobank.org).

During our 2018 investigation of dematiaceous hyphomycete diversity,

we isolated one Ochroconis-like strain from soil in Guizhou Province,

China. Now we propose a novel species, Ochroconis terricola, based on

morphological characters and phylogenetic analyses of ITS, LSU, and SSU

nuclear ribosomal sequences.

Materials & methods

Fungal isolates and morphological studies

Soil samples were collected from Guizhou Province, China, in 2018 and transported

to the laboratory in sterilized, zip lock polyethylene bags. All isolates are conserved in

the Herbarium of Department of Plant Pathology, Guizhou University (HGUP). The

taxon is described from cultures grown for 2 weeks at 25°C on potato dextrose agar

(PDA). Conidia and conidiophores were placed in a drop of 60% lactic acid, examined,

and photographed at 100x magnification using a Nikon 90i microscope.

DNA extraction, amplification, sequencing

Genomic DNA was extracted from colonies grown on PDA using the Fungal

gDNA Kit GD2416 following the manufacturer's instructions. ITS, LSU and SSU

genes were amplified via PCR using primers ITS4/ITS5 (White & al. 1990), LROR/LR5

(Rehner & Samuels 1994, Vilgalys & Hester 1990), and NS1/NS4 (White & al. 1990),

respectively. The PCR products were purified and sequenced by Sangon Biotech. The

new sequences were deposited in GenBank, and other sequences were obtained from

GenBank (TABLE 1).

Phylogenetic analyses

Sequences were aligned using ClustalX 1.81 (Thompson & al. 1997) and edited

manually using BioEdit (Hall 1999). The alignments were concatenated as a Fasta

document using MEGA 6.0 (Tamura & al. 2011). Phylogenetic analyses of combined

ITS, LSU, and SSU rDNA sequences were computed using maximum likelihood (ML)

analysis and Bayesian Inference (BI). The ML tree was generated using RAxML-

HPC2 on XSEDE v.8.2.8 (Stamatakis 2014) via the CIPRES Science Gateway platform

(Miller & al. 2010) with 1000 bootstrap replicates. Bayesian analysis was performed

via MrBayes v3.0b4 (Huelsenbeck & Ronquist 2001) using Markov Chain Monte

Ochroconis terricola sp. nov. (China) ... 145

TABLE 1. Sequences used in phylogenetic analyses.

GENBANK ACCESSION NO.

SPECIES STRAIN

ITS SSU LSU

Mycosisymbrium cirrhosum GUFCC 18012 KR259883 KR259885 KR259884

Ochroconis bacilliformis CBS 100442 KP798632 KP798638 KP798635

O. constricta CBS 202.27(T) MH854929 KF156072 KF282652

CBS 211.53 HQ667519 KF282671 KF282653

NH 1234 —_ = LC187202

FMR 3906 LM644509 — LM644552

NBRC 9375 DQ307327 AB564608 AB564619

O. cordanae CBS 475.80(T) KF156022 KF282672 KF282654

O. gamsii CBS 239.78(T) KF156019 KF156088 KF156150

O. icarus CBS 536.69(T) HQ667524 KF156084 KF156132

O. macrozamiae CBS 101179 KF156020 KF156091 KF156151

CBS 102491 KF156021 KF156092 KF156152

O. minima CBS 423.64 = KF282680 KF282666

CBS 119792 KF156027 KF156086 KF156133

CBS 510.71(T) HQ667522 KF156087 KF156134

O. phaeophora CBS 206.96 KP798631 KF282675 KF282660

O. ramosa UTHSC 121082(T) LM644524 LM644551 LM644567

O. robusta CBS 112.97(T) KP798633 KP798639 KP798636

O. sexualis PPRI 12991(T) KF156018 KF156089 KF156118

O. terricola HGUP1808(T) MK377301 MK377071 MK377073

Pleospora herbarum CBS 191.86 KC584239 GU238232 GU238160

Scolecobasidium cateniphorum CBS 769.83 KF156013 KF156044 KF156153

S. excentricum CBS 469.95(T) HQ667543 KF156096 KF156105

S. fusarioideum CBS 210.95 — KF156043 KF156154

S. fusiforme CBS 586.82 KF156012 KF156101 KF156155

Scolecobasidium sp. NH503 — AB564606 AB564617

S. terreum PO43 — EU107356 EU107306

S. tricladiatum POS1 — EU107354 EU107286

S. tropicum CBS 380.87 ard KF156042 KF156102

Venturia asperata ATCC 34052 — EF114736 EF114711

V. inaequalis CBS 594.70 KF156040 GU296205 GU301879

V. populina CBS 256.38 MH855959 GU296206 GU323212

V. pyrina ATCC 38995 — EF114739 EF114714

Veronaeopsis simplex CBS 588.66(T) KF156041 KF156095 KF156103

Verruconis calidifluminalis CBS 125818(T) MH875239 KF156046 KF156108

V. gallopava CBS 437.64(T) HQ667553 KF282674 KF282656

V. verruculosa CBS 119775 KF156014 KF156055 KF156106

New sequence is set in bold font. Type strains are marked by T.

Carlo method. Pleospora herbarum (CBS 191.86) was used as an outgroup. The

phylogenetic tree was viewed in Treeview v. 1.6.6 and the layout was completed in

Adobe Illustrator CS5.

146 ... Zhang & al.

Taxonomy

Ochroconis terricola Xin Zhang & YL. Jiang, sp. nov. Fia. 1

MB 831137

Differs from Ochroconis macrozamiae by its dark brown conidiophores and subhyaline

to pale brown conidia, from O. gamsii by its flexuous conidiophores and bigger conidia,

and from O. sexualis by its 1-septate and much smaller conidia.

Type: China, Guizhou Province, Leishan County, from forest soil, March 2018, X. Zhang

(Holotype, HGUP1808; ex-type culture, HGUP1808; GenBank MK377301, MK377071,

MK377073).

Erymo oey: referring to living on the soil.

Cotoniges on PDA effuse, flat, velvety, olivaceous, reverse dark brown,

growing slowly, attaining 9 cm diam. at 25° after 4 weeks. MycELIUM

immersed and superficial, hyphae hyaline to pale brown, smooth.

CONIDIOPHORES unbranched, 1-6 septa, flexuous, thick-walled, dark

brown, 2-4 um diam., with sympodially proliferating conidiogenous cells

bearing one or more denticles in the apical region. CONIDIOGENOUS CELLS

terminal, integrated, 1-3 um long, polyblastic, cylindrical, subhyaline to

medium brown. Conipia rhexolytic secession from conidiophores, solitary,

subhyaline to pale brown, coarsely verrucose, 1-septate, sometimes slightly

constricted at the septum, ellipsoidal to cylindrical or fusiform, apex mostly

rounded, base narrowly truncated with minute marginal frills, 7-12 x

2—4 um.

Phylogenetic analysis

The final alignment contained three genes for 36 isolates and 2568 (765

ITS + 827 LSU + 976 SSU) characters with Pleospora herbarum (CBS 191.86)

as outgroup. The ML phylogenetic tree is shown (Fic. 2). Empirical base

frequencies with 1,000 bootstrap inferences are 0.249211 (pi A), 0.221282

(pi C), 0.295026 (pi G), and 0.234480 (pi T). Bayesian analysis generated

a similar phylogenetic tree (one million generations; average standard

deviation of split frequencies = 0.009724). The Ochroconis terricola (HGUP

1808) sequences cluster together in a clade with O. macrozamiae Crous &

R.G. Shivas, O. gamsii de Hoog, and O. sexualis Samerp. & al. (Fic. 2) and

form a single branch sister to the other three species with high bootstrap

support (ML/BI = 84/1).

Discussion

In this study, one Ochroconis strain (HGUP1808) isolated from soil

was identified based on morphological characters and molecular data.

Ochroconis terricola sp. nov. (China) ... 147

Fic. 1. Ochroconis terricola (holotype, HGUP1808). A, B: Colony on PDA at 14 days; C: Mycelia on

the colony; D-F: Conidiophores and conidia; G-M: Conidia. Scale bars = 5 um.

148 ... Zhang & al.

100/1 Scolecobasidium cateniphorum CBS 769.83

84/1 Scolecobasidium fusarioideum CBS 210.95

Scolecobasidium fusiforme CBS 586.82

Scolecobasidium tropicum CBS 380.87

Venturia asperata ATCC 34052

Venturia pyrina ATCC 38995

3/1 100/1

100/1 | - Venturia populina CBS 256.38

oB/t Venturia inaequalis CBS 594.70

88/1 Veronaeopsis simplex CBS 588.66

98/1 Scolecobasidium excentricum CBS 469.95

100/1 Mycosisymbrium cirrhosum GUFCC 18012

Scolecobasidium tricladiatum P051

1o0/1 100/1 Verruconis gallopava CBS 437.64

Verruconis calidifluminalis CBS 125818

96/1 971-- Ochroconis constricta NH1234

99/1 Ochroconis constricta NBRC 9375

Verruconis verruculosa CBS 119775

100/1 Ochroconis cordanae CBS 475.80

Ochroconis phaeophora CBS 206.96

79/1 Ochroconis sexualis PPRI 12991

100/1 Ochroconis gamsii CBS 239.78

a 100/1- Ochroconis macrozamiae CBS 101179

a Ochroconis macrozamiae CBS.102491

Ochroconis terricola HGUP1808

Ochroconis constricta CBS 202.27

ma toon | 2Colecobasidium sp. NH503

Ochroconis constricta CBS 211.53

98/1 Ochroconis constricta FMR 3906

Scolecobasidium terreum P043

1001 ) Ochroconis icarus CBS 536.69

89/1 Ochroconis minima CBS 423.64

oo, Ochroconis minima CBS 510.71

Ochroconis minima CBS 119792

Ochroconis ramosa UTHSC 121082

100/1 Ochroconis bacilliformis CBS 100442

Ochroconis robusta CBS 112.97

Pleospora herbarum CBS 191.86

0.2

Fic. 2. Maximum Likelihood (ML) tree based on combined LSU, ITS, and SSU data of Ochroconis

species and other related genera species, with Pleospora herbarum (CBS 191.86) as outgroup.

Branch support is shown as: Maximum Likelihood bootstrap support values >75% / Bayesian

posterior probabilities >0.95. The new species is in bold.

SSU, ITS, and LSU multi-gene analyses support phylogenetic separation

of O. terricola from known Ochroconis species within a single clade in

the ML tree. Although Ochroconis terricola is phylogenetically close to

O. macrozamiae, O. gamsii, and O. sexualis in the phylogram (Fic. 2),

there are obvious morphological differences separating O. terricola and the

three species. Ochroconis macrozamiae is distinguished by its red-brown

conidiophores and conidia that constrict at the middle septum (Crous & al.

2014); O. gamsii differs by its erect conidiophores and curved or unilaterally

flattened, smaller conidia (6-9 x 2.4-2.8 um; de Hoog 1985); and O. sexualis

differs by its 1-3-septate, larger conidia (15-22 x 3.5-5.0 um; Samerpitak &

al. 2014). The combined morphological and phylogenetic analysis supports

O. terricola as a new taxon.

Ochroconis terricola sp. nov. (China) ... 149

Acknowledgments

The authors are grateful for pre-submission comments and suggestions provided

by Drs. Hui Deng (Institute of Agricultural Resources and Regional Planning, Chinese

Academy of Agricultural Sciences, Beijing), Wen-Xiu Sun (Bioengineering, College of

Life Sciences, Changjiang University), and Shaun Pennycook (Nomenclature Editor,

Mycotaxon). Yun Chen is thanked for valuable help. This project was supported by the

National Natural Science Foundation of China (no. 31660006).

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https://doi.org/10.1016/B978-0-12-372180-8.50042-1

MY COTAXON

January-March 2020—Volume 135, pp. 151-165

https://doi.org/10.5248/135.151

Neomyrmecridium asymmetricum sp. nov. from Ecuador

LIZETTE SERRANO’, DAYNET SOSA’?’, FREDDY MAGDAMA', FERNANDO

ESPINOZA’, ADELA QUEVEDO’', MARCOS VERA’, MIRIAM VILLAVICENCIO’,

GABRIELA MARIDUENA’, SIMON PEREZ-MARTINEZ’, ELAINE MALOSSO?,

BEATRIZ RAMOS-GARCIA4, RAFAEL F. CASTANEDA-RUuIz?

' Escuela Superior Politécnica del Litoral, ESPOL,

Centro de Investigaciones Biotecnologicas del Ecuador, Campus Gustavo Galindo,

Km. 30.5 Via Perimetral, PO. Box 09-01-5863, Guayaquil, Ecuador

? Universidad Estatal de Milagro (UNEMI), Facultad de Ingenieria,

Cdla. Universitaria Km. 1.5 via Milagro-Km26. Milagro 091706, Guayas, Ecuador

> Centro de Biociéncias, Departamento de Micologia, Universidade Federal de Pernambuco,

Avenida da Engenharia, s/n Cidade Universitaria, Recife, PE, 50.740-600, Brazil

‘Instituto de Investigaciones Fundamentales en Agricultura (INIFAT),

Tropical Alejandro de Humboldt, OSDE, Grupo Agricola,

Calle 1 Esq. 2, Santiago de Las Vegas, C. Habana, Cuba, C.P. 17200

* CORRESPONDENCE TO: dasosa@espol.edu.ec

ABSTRACT—A new species Neomyrmecridium asymmetricum, found on decaying leaves of

Theobroma cacao, is distinguished by grouped conidiophores and polyblastic production of

narrow clavate to subclavate, 1-septate, asymmetrical, and yellowish or subhyaline conidia.

An ITS- and LSU-based phylogenetic analysis, description, and illustrations are provided.

A key and illustrations to Neomyrmecridium species are also presented.

Key worps—asexual fungi, Myrmecridiaceae, taxonomy, tropics

Introduction

Crous & al. (2018a) introduced Neomyrmecridium Crous for three species:

N. septatum Crous (type species), N. asiaticum Crous, and N. sorbicola

(Crous & R.K. Schumach.) Crous. Neomyrmecridium is distinguished by

macronematous, unbranched, subcylindrical, multiseptate, smooth, brown

152 ... Serrano & al.

conidiophores with polyblastic, terminal, subcylindrical, denticulate, pale

brown conidiogenous cells. The conidia are solitary, fusoid-ellipsoid, obovoid,

hyaline or subhyaline (becoming pale brown with age), septate, smooth,

and sometimes encased in mucoid tunica (Crous & al. 2018a). The diversity

of microfungi in the Ecuadorian rainforests has received little attention,

especially in cacao plantations. During a survey of hyphomycetes associated

with plant litter in the Balao cacao plantation, Guayas province, Ecuador

(Fic. 1), we collected a Neomyrmecridium specimen that differs remarkably

from all previously described taxa (Crous & al. 2018a) and for which we

propose a new species.

Materials & methods

Collections

Samples of decaying plant materials were collected and placed in plastic bags for

transport to the laboratory, where they were washed, treated according to Castaneda-

Ruiz & al. (2016), and placed in moist chambers. Pure cultures were obtained by

transferring single conidium using a flamed needle to solidified media (with pH

adjusted to 6.3) containing corn meal extract mixed 1:1 with carrot extract plus 15

g agar (CMC) or V8 according to Crous & al. (2009). Plates were incubated at 25 °C.

Color notations in parentheses are from Kornerup & Wanscher (1984). Mounts were

prepared in PVL (polyvinyl alcohol, lactic acid) and measurements made at 1000x

magnification. Microphotographs were obtained with an Olympus BX51 microscope

equipped with bright field and Nomarski interference optics. The type specimen was

deposited in the Herbarium of Universidade Federal de Pernambuco, Recife, Brazil

(URM) and cultures obtained from the type specimen were deposited in the Culture

collections of Microorganism CIBE (CCM-CIBE), Guayaquil, Ecuador.

DNA extraction, sequencing, and phylogenetic analysis

Isolates CCMCIBE-H304 and CCMCIBE-H304-A were cultured on PDA

in darkness for 7 days at 25 °C. DNA was extracted using a modified protocol

from Cenis (1992). The primers ITS1/ITS4 were used to amplify ITS regions,

including the 5.8S gene (Manter & Vivanco 2007), and LROR/LRS5 to amplify

the D1/D3 domain of the LSU nrDNA (White & al. 1990). PCR products were

sent to Macrogen Inc. (South Korea) for purification and sequencing. Consensus

sequences assembled and edited using Geneious (ver. 10.1.2) were later compared

with those of the National Center for Biotechnology Information (NCBI) using

the Basic Local Alignment Search Tool (BLAST).

Each data set was aligned in MEGA 6.0 (Tamura & al. 2013) using

ClustalW (Thompson & al. 1994) and refined with MUSCLE (Edgar 2004).

The alignment included our strain sequences and those from different genera

in the Myrmecridiaceae obtained from NCBI (TABLE 1). ITS- and LSU-based

Neomyrmecridium asymmetricum sp. nov. (Ecuador) ... 153

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Myrmecndium flexuosum CBS 398.76

Myrmecridium fluviae CNUFC-YR61-2

Myrmécridium phragmitis CBS 131311

100 Myrmecnidum schulzeri CBS 100.54

u Myrmecnidium banksiae CBS 132536

93 ' Myrmecndum sparti CPC 24953

Ed Myrmecridaum montsegurinum JF 13180 Myrmecridiaceae

Neomyrmecnoum sorbdicola CBS:143433

700 Neomyrmecndium asiatcumn CBS:145080

Neomyrmecridium septatum CBS 145073

Neomyrmecridium asymmetrycum CCMCIBE H304

10+ Neomyrmecridium asymmetrycum CCMCIBE H304-A

mn |

Woswasia atropurpurea WU 32007

Pynculariomyces asan CPC 27444

Pynculana urashimae CPC 29414

Neopyncularia commelimicola CBS 128303 4

Pseudopyricularia iraniana [RAN 2761C Pyricula “

Pseudopyrculana bothnochioae CPC 21650

Pseudopyncularia hagahagae CPC 26635

- siosphacria ruguios

Vermiculanopsielia acaciae CPC 26291

Vermxulanopsiella dichapetal CBS: 143440

Vermicutariopsicla immersa MUCL39135

TCC MYA-257!

Vermiculariopsieila eucalypticola CBS. 143442 Vermiculariopsiellaceae

Vermiculanopsielia eucalypti CPC 25525

Vermiculariopsieia lauracearum CBS:145055

Vermiculanopseia pedculsta CBS 132484

a portoric: NFCC

Castanedelia acacwse CPC 24969

03 - Castanediela cagnizani CBS 101043

7 Castanediella tereticomis CBS:145088

Castanediells eucalypti CPC 24746 Castanediellacea

Castanediella couratani CBS 579.71

85 — Castanediella malaysiana CPC 24918

— Saccharata proteae CBS:119218

Fic. 2. The tree derived from the phylogenetic analysis using concatenated sequences of the

LSU and ITS of Myrmecridiales revealed that Neomyrmecridium asymmetricum and N. septatum

CBS:145073 were nested in a well-supported subclade (bootstrap value of 79).

phylogenies were generated using Maximum Likelihood (ML) with the best

nucleotide substitution model in MEGA 6.0 (Tamura & al. 2013).

Best models used were (for LSU) Tamura-Nei with Gamma distribution and

(for ITS) Kimura 2-parameter with Gamma distribution and Invariant sites (G+I).

The best nucleotide substitution model for the combined LSU + ITS analysis

was the General Time Reversible with Gamma distribution and Invariant sites

(G+I). Bootstrap analysis of 1,000 replicates was used to assess the reliability

Neomyrmecridium asymmetricum sp. nov. (Ecuador) ... 157

of the reconstructed phylogenies. ML bootstrap values =70% were considered

significant. DNA sequences generated in this study were deposited in GenBank

(TABLE 1).

Phylogeny

The BLAST query revealed that the CIBE H304 and CIBE H304-A LSU

sequences of N. asymmetricum showed a 97% similarity with N. septatum

CBS145073 and N. asiaticum CBS145080. However, they showed sequence

identity of 94-96% with LSU sequences of Myrmecridium species, also

belonging to the Myrmecridiaceae. The CIBE H304 and CIBE H304-A ITS

sequences showed a 90 % similarity with N. sorbicola CBS143433, the highest

value matched after blast search.

We carried out individual and combined analyses of the LSU and ITS loci

to assess relationships with members of the Myrmecridiales (Sordariomycetes)

(Fic. 2). The final concatenated analysis encompassed 48 sequences and

comprised 1118 bp (ITS 555 bp, LSU 563 bp). The ML tree nested both

N. asymmetricum isolates (CIBE H304 and CIBE H304-A) and N. septatum

CBS145073 in a well-supported subclade (bs = 79%), with N. asiaticum CBS

145080 as the closest sister species (FIG. 2).

Taxonomy

Neomyrmecridium asymmetricum R.F. Castafieda, Serrano & D. Sosa,

sp. nov. Fries 3-5

MycoBank MB 831330

Differs from Neomyrmecridium sorbicola by its narrow clavate to subclavate, 1-septate,

asymmetrical conidia.

Type: Ecuador, Guayas Province, Guayaquil, Balao, 2°48 S 79°40 W, on decaying

leaves of Theobroma cacao L. (Malvaceae), 8 July 2017, F. Espinoza & S. Pérez-Martinez

(Holotype, URM 90896; ex-type cultures, CCMCIBE-H304, GenBank MN014057,

MN014055; and CCMCIBE-H304-A, GenBank MN014058, MN014056).

ETYMOLOGY: asymmetricum- (Latin), meaning asymmetric.

CoLonigs on the natural substrate hairy, effuse, amphigenous, yellowish

brown. Mycelium superficial and immersed, composed of branched, 1-2.5

um diam, smooth, brown hyphae. CoNIDIOPHORES macronematous,

mononematous, grouped, slightly fasciculate, erect, straight, cylindrical,

unbranched, 3-15-septate, brown, pale brown toward the apex, 40-210 x

3.5-9 um, smooth. CONIDIOGENOUS CELLS polyblastic, terminal, integrated,

cylindrical or subcylindrical, indeterminate, with several sympodial

extensions, denticulate, with tiny cylindrical denticles, pale brown, 8-35 x

158 ... Serrano & al.

Fic. 3. Neomyrmecridium asymmetricum (cultures ex holotype, URM 90896).

A-D. Colonies and reverses on CMC (A-B) and V8 agar (C-D).

3.5-5 um. Conidial secession schizolytic. Conrp1 solitary, acropleurogenous,

narrow clavate or subclavate, 1-euseptate, asymmetrical, 12-15 x 2-3 um,

basal cell 7-10 um long, apical cell 4-6 um long, yellowish or subhyaline,

smooth-walled.

CULTURE CHARACTERISTICS: Colonies on CMC attaining 22 mm in 7 days

at 25°C, flat, subfelted, olive (3/E5). Mycelium mostly immersed, scarcely

aerial toward the entire margin. Reverse dark green (30/F6). Hyphae septate,

Neomyrmecridium asymmetricum sp. nov. (Ecuador) ... 159

Fic. 4. Neomyrmecridium asymmetricum (holotype, URM 90896).

A-C. Conidiophores and conidiogenous cells. D. Conidiogenous cell.

160 ... Serrano & al.

2 @ #~-es & @ '

‘

Fic. 5. Neomyrmecridium asymmetricum (holotype, URM 90896).

A-C. Conidia. D. Conidiogenous cell and conidia. E-F. Conidiogenous cells.

Neomyrmecridium asymmetricum sp. nov. (Ecuador) ... 161

Fic. 6. Representative conidia of Neomyrmecridium spp. (re-drawn from the literature).

A. N. asiaticum (Crous & al. 2018a). B. N. septatum (Crous & al. 2018a).

C. N. sorbicola (Crous & al. 2018b). Scale bars = 10 um.

subhyaline to pale olivaceous-brown, smooth, 1.5-2 um diam. Sporulation

occurred after 4 days, producing conidia similar to those observed from nature.

Colonies on V8 agar attaining 12 mm in 7 days at 25°C, flat concentric near

cottony center, filamentous-filiform toward margin, yellowish orange (4/A7).

162 ... Serrano & al.

Reverse deep orange (5/A8) at the center, light orange (5/A6) toward the

margin. Sporulation poor and sparsely after 5 days, conidia similar to those

observed from nature.

Note: Neomyrmecridium sorbicola (Crous & al. 2018a,b) is superficially

similar to N. asymmetricum in producing 1-septate conidia, but N. sorbicola

differs in its pale brown obovoid conidia with median regions surrounded

by a mucoid tunica (Fic. 6). Neomyrmecridium septatum is separated by its

conidia, which are fusoid-ellipsoid, mostly 3-septate, guttulate, pale brown,

and with a mucoid tunica encasing the upper two thirds (Crous & al. 2018a)

(FIG. 6).

Morphological and phylogenetic analyses support N. asymmetricum as a

new species in Myrmecridiaceae.

Key to Neomyrmecridium species

WeOnidta AOS tvs Sep CALC sna hoainanr arte eyettns e eyttente Sache atcha eee hp tie etal nar 2

1. Conidiaanostly more than 1-sepfates< ¢ int. os aed oben} Mine et ote net 3

2. Conidia clavate or subclavate, 12-15 x 2-3 um, asymmetrical,

basal cell 7-10 um long, apical cell 4-6 um long,

vellowishvorsubhyaline ty. 5 awed 56, gb wn ute ary ale coded 08 go N. asymmetricum

2. Conidia obovoid, (7—)8-10(-15) x 4(-5) um, initially hyaline

(pale brown in age), (0O-)1(-3)-septate, with a 1-2 um thick

mucoid tunica surrounding the medial region ................ N. sorbicola

3. Conidia fusoid-ellipsoid, (12—)14—16(-20) x (3.5-)4(-5) um, hyaline

(pale brown in age), (1-)3-septate, guttulate, with a 1-2 um thick

mucoid tunica encasing the upper two thirds ................. N. septatum

3. Conidia ellipsoid to obovoid, (13—)15-17 x 4-5 um, pale brown,

(2—)3-septate, guttulate, surrounded by a 0.5 um thick

DelANOUS HUMIC Ags file, 52. che 5 Mec ene iene oh aM SM acs maa acto ee N. asiaticum

Acknowledgments

We are indebted to Dr. Josiane S. Monteiro (Museu Paraense Emilio Goeldi, Belém,

Brazil) and Dr. De-Wei Li (The Connecticut Agricultural Experiment Station Valley

Laboratory, Windsor CT, USA for their critical reviews. The authors are grateful to

Escuela Superior Politécnica del Litoral (ESPOL), CIBE for financial support and

the International Society for Fungal Conservation for facilities. RFCR is grateful

to the Cuban Ministry of Agriculture. We acknowledge the websites provided by

Dr. Paul Kirk (Index Fungorum) and Dr. Konstanze Bensch (MycoBank). Dr. Lorelei

Norvell’s editorial review and Dr. Shaun Pennycook’s nomenclature review are

greatly appreciated.

Neomyrmecridium asymmetricum sp. nov. (Ecuador) ... 163

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MY COTAXON

January-March 2020—Volume 135, pp. 167-182

https://doi.org/10.5248/135.167

A comparison of anamorphs of some Pachyphlodes species

and the type of Chromelosporium:

are they congeneric?

GREGOIRE L. HENNEBERT! & CONY DECOCK?

'Rue de l'Elevage 32, B 1340 Ottignies-Louvain-la-Neuve, Belgium

?MUCL, Place Croix du Sud, 1348 Louvain-la-Neuve, Belgium

CORRESPONDENCE TO: hennebertg@scarlet.be

ABSTRACT— There is a question of whether or not the name Chromelosporium competes with

the name Pachyphlodes given that both genera exhibit similar conidiogenesis. We address

here the question through a comparative study of their anamorphs. The type specimen of

C. ochraceum (generic type of Chromelosporium), has been studied and compared to newly

collected specimens of two Pachyphlodes species, P. nemoralis and P. citrina (whose identity

was confirmed through comparison of ITS DNA sequences) and an unidentified species from

the Pachyphlodes—Plicariella lineage. Comparison of the morphology of the Pachyphlodes—

Plicariella lineage anamorphs with Chromelosporium ochraceum reveals discriminating

characters that may support a generic distinction.

Keyworps—Ascomycota, Glischroderma, Pezizaceae, molecular analyses

Introduction

Corda (1833) described Chromelosporium in Sturm’s DEUTSCHLANDS

Fiora fora fungus collected in 1827 on dead stems of Allium (Amaryllidaceae)

and Hemerocallis (Xanthorrhoeaceae) and characterized by ochre spores and

hyphae immersed in a gel. Subsequent authors, however, overlooked the

presence of the gelatinous matrix reported by Corda (1833). Chromelosporium

has had a confusing history due to similarities of conidiogenesis found in

Hyphelia Fr., Ostracoderma Fr., and Glischroderma Fuckel.

Juel (1920) noted that the conidiogenesis in collections of Hyphelia

terrestris Fr. and in a fungus that he collected on soil in Mésseberg in western

168 ... Hennebert & Decock

Gotland, Sweden, considered to represent Ostracoderma pulvinatum Frt.,

were similar. Consequently, Juel reduced Ostracoderma to a synonym of

Hyphelia. No original specimen of Ostracoderma pulvinatum, from Fries or

Juel, were found in the S herbarium, impeding attempts to confirm whether

the peridium described by Fries for Ostracoderma as “peridio rotundato

minime villoso sed crustaceo” and the one supposedly observed by Juel

(1920) are made of a definite pseudotissue of dense intricate hyphal cells

enveloping the totality of the conidioma as in Glischroderma cinctum Fuckel

or comprises just a superficial agglutination of hyphae embedded in a gel

that upon drying becomes a crust-like thin layer, covering only parts of the

hyphal mat—a “peridium indeterminatum e villo in pelliculam cohaerentem

context,’ as observed by Fries in Hyphelia and by Korf (1994).

Because Fries based his generic type, Hyphelia rosea, on “Trichoderma

roseum Pers. (non Trichothecium Link), Hughes (1958) [having

identified Trichoderma roseum Pers. and Trichothecium roseum (Pers.)

Link as homotypic] considered the name Hyphelia no longer available.

Therefore, Hughes (1958) rejected Juel’s synonymy and, following Juel’s

observations, adopted Ostracoderma for species with similar conidiogenesis,

such as Hyphelia terrestris and Chromelosporium ochraceum, making

Chromelosporium a synonym of Ostracoderma. Hennebert (1973) segregated

the non-peridiate Chromelosporium (with C. ochraceum and other species)

from the peridiate Ostracoderma and Glischroderma.

Korf (1994) reported several Chromelosporium-like specimens from

soil and litter in New York State with a superficial gelatinous mass, which

he placed in Glischroderma, a genus described by Fuckel in 1870 and

characterized by a superficial sticky gel (glischros = viscous). Norman and

Egger (1999) sequenced one of Korf’s anamorphs and demonstrated that

it was affiliated with the Pachyphlodes-Plicariella |as Scabropezia] lineage.

In examining Korf’s specimens identified as Glischroderma, Healy (2015)

recognized additional epigeous anamorphs of Pachyphlodes species and

molecularly linked Pachyphlodes pfisteri, a truffle with a Chromelosporium-like

anamorph. Hennebert (2017) subsequently distinguished Chromelosporium

from Glischroderma”.

“) The list of specimens examined under Pachyphlodes pfisteri in Hennebert (2017)

erroneously cites specimens received on loan from CUP instead of citing specimens identified

as Pachyphlodes pfisteri (represented by CUP 62646, 62650, 62651, 62653, 63540, 63648)

based on conidial size and ornamentation and hyphal width; CUP 62652, 62654, 63647, 63695

represent undefined species.

Are Chromelosporium & Pachyphlodes congeneric? ... 169

The presence of gel embedding the conidiogenous cells in

Chromelosporium incited the senior author to search for fresh samples for

further investigations. Fourteen samples of Chromelosporium-like spore

mats were then collected on bare or mossy soils or mixed with organic

debris in forests in Belgium, some of them bearing a few gel droplets. These

Chromelosporium-like specimens were identified by ITS barcoding as two

species of Pachyphlodes and an undetermined species of the Pachyphlodes-

Plicariella lineage. Their anamorphs are described below and compared to

the type specimen and original description of Chromelosporium ochraceum.

This comparative morphological study revealed clear distinctions from

Chromelosporium ochraceum.

Materials & methods

SPECIMENS. Fourteen specimens were collected in the forest, accessioned, and

deposited in MUCL. The type specimen of Chromelosporium ochraceum (A.C,].

Corda 155414) was borrowed from the Herbarium Kryptogamologicum Musei

Nationalis Pragae in Prague (PRM). Microscopical examinations and drawings

were carried out using a 1960 phase contrast Olympus FH microscope equipped

with a Wild drawing tube. The photographs used in figure 6 were made with an

Olympus BX50 microscope equipped with Olympus SC100 numerical camera and

Olympus screen. All drawings and photographs are made from samples mounted in

lactic acid with Cotton blue.

SEQUENCING. DNA was extracted from fragments of five freshly collected

samples in lysis buffer, using innuPREP Plant DNA kit following the manufacturer’s

recommendations. Sequences were determined for the nuc rDNA ITS1-5.8S-ITS2

region (ITS) with the primer pair ITS5 and ITS4 (White & al. 1990). PCR conditions

are as described in Gordillo & Decock (2018). Amplicons were sequenced in both

directions by Macrogen Inc. (Korea) using the same primers. MK prefixes designate

sequences deposited in GenBank.

Taxonomy

Anamorph of Pachyphlodes nemoralis Hobart, Bona & A. Paz,

Ascomycetes.org. 7(6):363, 2015 FIGS 1, 6A

HypHAL MATS forming cushion-shaped or flattened patches, irregular

in outline, grouped, 3-23 mm in diam., 1-3 mm high, white when young

discoloring to pale yellow ochre when mature, the surface appearing

fluffy to compacted, with a few superficial, small droplets of translucent,

glutinous, liquid gel, drying orange and flattened, small, brittle crust-like,

the margin remaining fluffy and powdery.

170 ... Hennebert & Decock

Fic. 1. Pachyphlodes nemoralis anamorph (MUCL 56524).

A. Synnema. B. Young branching. C. Open branching. D. Compact terminal cells.

E. Conidiogenous branches. F. Conidia (Scale bars = 10 um).

Are Chromelosporium & Pachyphlodes congeneric? ... 171

HypuHae at first loosely interwoven, smooth, hyaline, 4-8 um diam.,

becoming fasciculate, forming hyaline, erect synnemata <35 um diam.,

made of hyphae with 2—20 septa, with lateral branches at wide angles, all

hyphae turning into apically branched conidiogenous cells, with a repeated

asymmetrical bifurcating pattern, coralloid, at variable angles, openly

disposed to very compact, sparsely septate, each branch 15-50 um long,

and <10 um diam. at the inflated apical section, covered at maturity along

their entire length with conidia.

CONIDIOGENEsIS holoblastic, more or less synchronous with minute

denticles, 1-2 um long.

Conrp1A globose to subglobose, hyaline, 4-6.5 um, averaging ~5 um

diam. excluding warts, rarely ovoid or pyriform in moist conditions, 4 x

6.5(—8) um, the wall cyanophilic, 0.5 um thick, warted or tuberculate all

over, warts, <0.3 um high, evenly-spaced, 12—14 in median section.

Hasirat. On soil, moss, and organic debris along paths under Quercus

and Fagus sylvatica trees in forest.

SPECIMENS EXAMINED: BELGIUM, WALLOON BRABANT, Ottignies-LLN, Lauzelle

forest, on soil, moss, and organic debris along paths under Quercus and Fagus

sylvatica, October 2017, coll. GL Hennebert (MUCL 56520, GenBank MK714923;

MUCL 56524, GenBank MK7 14924; MUCL 56529, GenBank MK71492).

Anamorph of Pachyphlodes citrina (Berk. & Broome) Doweld,

Index Fungorum 31: 1, 2013 FIGS 2, 6B

HyYpPHAL MATS forming cushion-shaped or flattened patches, irregular

in outline, somewhat in groups, 2-25 mm in diam., 1-2 mm high, white

when young, discoloring to pale yellow ochre when mature, with a dense

surface and fluffy margin, the largest patches bearing superficially droplets

of translucent, glutinous, liquid gel, becoming orange flattened crust-like

and brittle when dried.

HypHAe at first loosely interwoven and branched, smooth, hyaline,

3—6(—8) um diam., with 2-15 septa, becoming fasciculate in hyaline, erect

synnemata, <25 um wide, the hyphae frequently anastomosed, laterally

and apically branched, furcating repeatedly and asymmetrically, sparsely

septate, each part 10-25 um long and terminally inflated <8 um wide,

covered at maturity all along their length with conidia.

CoNIDIOGENEsIS holoblastic, more or less synchronous with minute,

denticles, 1 um long.

Conrp1A4 globose to subglobose, hyaline, pale yellow ochre in mass,

4.5—7 um, averaging ~5.2 um diam., excluding warts, with a cyanophilic

172 ... Hennebert & Decock

Fic. 2. Pachyphlodes citrina anamorph (MUCL 56652). A. Synnema. B. Anastomosis. C. Young

branching. D. Developed branches. E. Conidiogenous branches. F. Conidia (Scale bars = 10 um).

Are Chromelosporium & Pachyphlodes congeneric? ... 173

wall 0.5 um thick, coarsely warted or baculate all over, warts cylindrical,

apically blunt, <0.8 um high, evenly spaced, 12—14 in median view.

Hapsirat. On soil along path near Castanea and Larix in forest.

SPECIMEN EXAMINED: BELGIUM, WALLOON BRABANT, Ottignies-LLN, Lauzelle

Forest, on soil along path near Castanea and Larix in Lauzelle forest, October 2017,

coll. GL Hennebert (MUCL 56652, GenBank MK714926).

Anamorph of undetermined Plicariella sp. or Pachyphlodes sp. FIGS 3, 6C

HypHAL mats forming cushion-shaped patches, determinate, irregular

in outline, grouped, 1-5 mm in diam., 1-2 mm high, white when young

to yellow ochre in color when mature, with surface fluffy to compacted

and granulose, a few fresh mature patches bearing a droplet of translucent

glutinous gel, forming a local orange flattened brittle crust when dried, the

margin, if not the entire patch, remaining fluffy and granulose.

HypuHaeE at first loosely interwoven, smooth, hyaline, 3-7 um diam.,

2—10-septate, becoming fasciculate in hyaline erect synnemata <20 um

wide, becoming terminally branched through repeated and asymmetric

bifurcations to form coralloid branch tips that are openly disposed or

compacted and sparsely septate. Each part of branch is 10—70 um long and

inflated <8 um, covered at maturity all along their length with conidia.

CONIDIOGENESIS holoblastic more or less synchronous with minute

denticles 1—1.5 um long.

Conip1A globose, hyaline, (3-)3.5-4.5 um, most 4 um diam., with a

finely punctate wall.

Hasitart. Soil, moss, and organic debris along path under mixed trees in

forest.

SPECIMEN EXAMINED: BELGIUM, WALLOON BRABANT, Ottignies-LLN, Lauzelle

forest, on soil along path in, October 2017, coll. GL Hennebert (MUCL 56522,

GenBank MK714927).

Original descriptions by Corda (1833)

Chromelosporium Corda and C. ochraceum Corda, Sturm’s Deutschlands Flora.

Abbildungen nach Natur., Abt. III Die Pilze, Bd. 3, p. 81, Tab. 41, 1833.

“CHROMELOSPORIUM Corda

Char. Gen. Sporae continuae coloratae, in gelatinae nidulantes, floccis heterogeneis

destructis ramosis, articulatis, hyalinis inspersae. Acervuli effuse colorati.

[Gen. char. Spores simple, colored, embedded in gel, hyphae dispersed, hyaline,

heterogeneous, articulate, branches fragmented; mats extended, colored. ]

174 ... Hennebert & Decock

Fic. 3. Plicariella sp. or Pachyphlodes sp. anamorph (MUCL 56522).

A. Tip of synnema, B. Branching, C-E Conidiogenous hyphae. F. Conidia. (Scale bars = 10 um).

Are Chromelosporium & Pachyphlodes congeneric? ... 175

CHROMELOSPORIUM OCHRACEUM Corda

Ocherfarbige Chromelosporie. Tab. 41

Ch. acervulis effusis ochraceis pulverulentis; sporis copiosissimis globosis

ochraceis, gelatinae coloratae immersis; floccis hyalinis grosse-articulatis albis.

[Fungus cushion-like, effuse, ochre in color, pulverulent; spores very abundant,

globose, ochre in color, immersed in a colored gel, dispersed in hyphae hyaline,

white, branched and broadly articulate. ]

Chr. mit verbreiteten ocherfarbigen, bestaubten Haufchen, sehr haufigen

ocherfarbigen, mit gefarbten Schleim eingehiillten, kuglichen Sporen, und

durchsichtigen, weissen, grossgegliederten Faden.

Wohnt auf faulenden Stengeln der Liliaceen, verziiglich der Hemerocallis

und Allium, 1827.

Die grossgegliederten, und nicht durch Scheidewande (septa) getheilten

Faden unterscheiden diese Gattung von Sporotrichum, so wie auch die gefarten

Sporen, da Sporotrichum flocci septati (non articulati) und sporae homogeneae

besitzt.

Die glashellen, weissen, grossgliedrigen, niederliegenden und verworrenen

Faden, sind wenig 4stig, und im Vergliech zu ihren Sporen sehr gross und

sparsam.

Die Sporen selbst sind nicht aus den Faden entstanden, daher heterogen,

sind kuglich, klein, sehr zahlreich, intensive ocherfarb, fast ziegelroth gefarbt.

Die sind einfach, und nicht getheilt, auch sind sie durch einen im Wasser leicht

ldsslichen und gefarbten Schleim Haufchenartig an die Faden befestigt.

[Ch. in extended, ochre, powdery mats, very often covered by a colored gel, spores

globose, hyphae hyaline, in broad segments.

Habitat on rotten stems of Liliaceae, such as Hemerocallis and Allium. 1827.

The articulate and not septate hyphae distinguish this genus from Sporotrichum,

which also has colored spores, septate hyphae and homogeneous spores.

The hyphae are hyaline, articulate, prostrate, interwoven, sparsely branched

and, compared to the spores, broad and sparse.

The spores themselves are not formed from the hyphae, thus heterogeneous.

They are globose, small, very abundant, deep ochre, near reddish colored. They are

simple, not septate; they are also fixed to the hyphae forming a kind of head by the

colored gel slightly dissolved in water]

Tab. 41. A. Nattirliche Grosse. B. Faden mit Sporen. C. Sporen noch starker

vergrossert. A. J. Corda.”

CORDA’S SECOND GENERIC DESCRIPTION (1842)

In ICONES FUNGORUM HUJUSQUE COGNITORUM, V, p. 8, Prague 1842,

Corda gives a shortened description of Chromelosporium with a new

comment:

176 ... Hennebert & Decock

tf.

’

: Chrmeley

ava vtec €

PLATE 4. Chromelosporium ochraceum Corda,

Sturm’s Deutschl. FI., III (Pilze), 3(13): 81, tab. 41, 1833

Are Chromelosporium & Pachyphlodes congeneric? ... 177

“CHROMELOSPORIUM Corda, Sturm FI. III, 13, p. 81.

Sporae continuae, coloratae, gelatinae immersae, floccis heterogeneis, articulatis,

repentibus, ramosis, hyalinis inspersae.

K. Wir haben eine deutliche Schleimmasse gefunden, welche die Sporen und

Flocken einhiillte”

[Spores simple, colored, embedded in gel, among hyphae heterogeneous, articulate,

prostrate, branched, hyaline.

CoMMENTS. We have found a conspicuous mass of gel covering the spores and

hyphae. |

Chromelosporium ochraceum re-examined

Chromelosporium ochraceum Corda,

Sturm’s Deutschl. FI., III (Pilze), 3(13): 81, tab. 41, 1833. FIGS 5, 6D

Ho.otype: Czechoslovakia, Prague, “Chromelotrichum|(crossed out|sporium

ochraceum Ca. ramulis furcis, Myrinema generis” [scr. & del. A.C.J. Corda]. On Allium

dead stem. (Herb. Corda in PRM 155414).

COLONIES in small patches <10 mm, velvety, at first white then ochraceous

when sporulating, made of two layers, one of conidiophores under one of

terminal conidiogenous branches.

CONIDIOPHORES erect, mononematous, emerging from creeping hyphae,

3-5 um diam. in the substratum, made of a bulbous basal cell, 28-35 x

12-15 wm, ochraceous, extending into a cylindrical, septate stipe,

100-400 um long, light to ochraceous, the individual cells 40-95 x 10-14

um, repeatedly branched, terminating with up to five dichotomous branches,

each branch most often with a basal septum, and a second one next to

the subsequent dichotomous branches, their length decreasing from one

dichotomy to the next, usually the first basal dichotomy 40-90 x 9-10 um,

the second 30-70 x 6-10 um, the third 30-50 x 6-9 um, but in some cases

dichotomies very short, decreasing from 20-10 um, the range of angles in

the dichotomies from 35-45°.

CONIDIOGENOUS CELLS comprising up to four terminal dichotomies,

together 70-100 x 6-9 um, often apically clavate to 11 um diam., bearing

conidia all along with small, denticles 2 x 1 um, collapsing after conidial

release, seceding at their basal septum, rarely leaving a frill on the last

conidiophore cell.

Conip1A holoblastic, borne singly on denticles, at maturity globose or

subglobose, thick-walled, cyanophilic, verrucose, ochraceous to salmon in

mass, 4—5.5 um diam., <6 um including warts, warts prominent and blunt,

12-18 in median view.

178 ... Hennebert & Decock

tt=

—c— | eae 9 =f Sines Tq

Fic. 5. Chromelosporium ochraceum Corda (Holotype PR 155414). A. Conidiophores stipes

with basal cell and dichotomous branching. B. Terminal conidiogenous dichotomies. C. conidia

(Scale bars = 10 um).

Are Chromelosporium & Pachyphlodes congeneric? ... 179

ComMMENTS— The terms used by Corda interpreting his observations illustrated

in Fic. 4 (Corda’s fig. 41b) may raise some questions. The hyphae described as

broadly articulate (“grosse-articulatis, grossegegliederten”) appear in the figure

to be hyphae distantly septate in long cells, drawn in their natural dichotomous

position. Also the “ramis destructis” might refer to collapsed and disappeared

conidiogenous branches after conidial release That would explain why Corda

did not report attachments of the conidia to hyphae, which he said were

heterogeneous (“floccis heterogeneis, sporen nicht aus den Faden entstanden,

daher heterogen’). The gel reported as colored (“gelatinae coloratae, gefarbten

Schleim”’), is translucent and may appear colored because of the embedded

ochre spores. Our observations suggest that Corda noted those characters from

an over-mature collection.

Discussion

Corda (1833, 1842) described Chromelosporium ochraceum with conidia

embedded in a gel. However, this character was not considered significant

by Hughes (1958) or Hennebert (1973) until Korf (1994) collected

Chromelosporium-like specimens with a superficial gel and Healy (2015)

noticed gel on anamorphic growth of Pachyphlodes spore-mats when crushed.

The gel, if noticed on fresh anamorphs in humid conditions, may appear

on dried herbarium specimens as an orange spot, membranous and brittle,

variable in size (as small as 2 mm) or may leave no traces, as is the case for the

type specimen of Chromelosporium ochraceum. After the very dry summer of

2017, we observed gel droplets on only a few samples in shady places. In our

opinion, the presence of gel is not a reliable, stable character, and its degree

of development is probably circumstantial and dependent on environmental

conditions.

The anamorphs of P nemoralis and P. citrina are not yet described

(R. Healy, pers. comm.). The anamorph of Pachyphlodes pfisteri Tocci &

al., an endemic North American taxon, was described and illustrated

with macro- and micro-photographs by Healy & al. (2015) and drawn by

Hennebert (2017). Its conidiogenesis is in all respects identical to that of

P. nemoralis and P. citrina (Fics 1, 2).

The anamorphs of the two Pachyphlodes and one undetermined

Pachyphlodes-Plicariella species here described show distinctive features,

particularly in hyphal width, conidiophore branching, and conidial size

and ornamentation (Fic. 6). These characters should be examined in more

Pachyphlodes and Plicariella species to evaluate their diagnostic value.

180 ... Hennebert & Decock

Although they have the same type of conidiogenesis, Chromelosporium

ochraceum, the generic type species, differs significantly from Pachyphlodes

and Plicariella in its anamorphic morphology. Chromelosporium, as

demonstrated by C. ochraceum, has mononematous, erect apically

branched conidiophores with regular dichotomies, each branch delimited

by a septum, with up to four terminal dichotomies bearing holoblastic,

synchronous conidia. Pachyphlodes and Pachyphlodes—Plicariella as shown

here differ from Chromelosporium by creeping and erect, intermixed

synnematous conidiophores, with lateral and apical coralloid branches that

are sparsely septate and bearing holoblastic synchronous conidia along the

entire length.

Conclusion

Differences found here between the anamorphs of Pachyphlodes species

and the Pachyphlodes-—Plicariella lineage and the anamorphs and the type

species of Chromelosporium include synnematous versus mononematous

conidiophores, apical and lateral branching versus solely apical branching,

and spore production along most branches of the spore mat versus being

limited to terminal hyphal branches. These differences might provide

morphological evidence for maintaining Chromelosporium as a distinct

genus from Pachyphlodes.

It remains to compare the morphologies of the still unknown anamorphs

of P ligerica (Tul. & C. Tul) Zobel, the type species of Pachyphlodes

Zobel, and of Plicariella radula (Berk. & Broome) Rehm, the type species

of Plicariella (Sacc.) Rehm, with the description provided here for

C. ochraceum to confirm the morphological distinctiveness of the

anamorphs of these lineages. The most expedient solution to answering the

question of relatedness would be molecular analyses of C. ochraceum in its

fresh anamorphic and/or sexual phases (yet to be discovered) and of the

types of Pachyphlodes and Plicariella.

Another group of species presently named in Chromelosporium,

C. coerulescens (Bonord.) Hennebert and C. carneum (Pers.) Hennebert,

do not produce gel but form erect synnematous conidiophores, coralloid

branching, and similar conidiogenesis (Hennebert 1973). These appear to

be morphologically closer to the anamorphs of Pachyphlodes-Plicariella

than to Chromelosporium. Genetic analysis should decide their taxonomic

position in the Pezizaceae.

Are Chromelosporium & Pachyphlodes congeneric? ... 181

PiaTE. 6. Conidial ornamentation: A. Pachyphlodes nemoralis (MUCL 56524). B. P. citrina

(MUCL 56652). C. Plicariella sp. or Pachyphlodes sp. (MUCL 56522). D. Chromelosporium

ochraceum (Type PR 155414).

Acknowledgments

We are very thankful to the revisers of the manuscript, Dr Rosaria Ann Healy

(Department of Plant Pathology, Florida University) and Dr Keith Seifert (Agriculture

and Agri-Food Canada, Ottawa Research Center) for their very appreciated

annotations. Dr Jan Holec (Director, Mycological Herbarium, Prague) is greatly

thanked for the loan of the precious holotype of Corda’s Chromelosporium ochraceum,

which allowed us to fulfill the purpose of this paper. We thank Stephanie Huret

(MUCL genetic service) who conducted the sequence analyses. Also we are thankful

to the online Biodiversity Heritage Library for providing with color reproduction of

Sturm’s Deutschlands Flora as sent by Prof Kathie Hodge, Cornell University. Cony

Decock gratefully acknowledges financial support received from courtesy of the

Belgian Federal Science Policy and the BCCM™ research program).

182 ... Hennebert & Decock

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the First Whetzel-Wescott-Dimock Lectureship, Cornell Univ. Oct. 1994, 19 p.

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https://doi.org/10.1016/B978-0-12-372180-8.50042-1

MY COTAXON

January-March 2020—Volume 135, pp. 183-193

https://doi.org/10.5248/135.183

Pseudosperma flavorimosum sp. nov. from Pakistan

SANA JABEEN? & ABDUL NASIR KHALID?

‘Department of Botany, Division of Science and Technology, University of Education,

Township, Lahore, Punjab, Pakistan

*Department of Botany, University of the Punjab,

Quaid-e-Azam Campus, Lahore, Punjab, Pakistan

“ CORRESPONDENCE TO: sanajabeenue@gmail.com; sanajabeen@ue.edu.pk

ABSTRACT—Pseudosperma flavorimosum is a new species described from Khyber

Pakhtunkhwa province, Pakistan. It is delimited based on morphological characters

combined with a molecular phylogeny inferred from nuclear ribosomal DNA internal

transcribed spacer (ITS) sequence analyses. The ITS-based phylogeny supports the

independence of the new species, which is also is morphologically distinct from closely

related taxa.

Key worps—Agaricales, Basidiomycota, Inocybaceae, taxonomy

Introduction

Inocybaceae Julich (Basidiomycota, Agaricales) is one of the larger

families of agaric fungi with more than 700 species distributed worldwide

(Matheny & al. 2009; Kropp & al. 2010; Bougher & Matheny 2011; Bougher

& al. 2012; Fan & Bau 2013, 2014; Braaten & al. 2014; Esteve-Raventos & al.

2015; Jabeen & al. 2016; Farooqi & al. 2017; Matheny & Bougher 2017; Latha

& Manimohan 2017; Ullah & al. 2018; Matheny & al. 2019). The number

of species has increased considerably with the exploration of tropical and

southern temperate areas. Many representatives of genera of Inocybaceae

have been reported from Asia (Kobayashi & Onishi 2010; Fan & Bau 2013,

2014; Horak & al. 2015; Latha & Manimohan 2015, 2016, 2017; Saba & al.

2015; Jabeen & al. 2016; Pradeep & al. 2016; Farooqi & al. 2017; Naseer &

184 ... Jabeen & Khalid

al. 2017; Liu & al. 2018; Ullah & al. 2018), with 24 species reported from

Pakistan (Ahmad & al. 1997, Ilyas & al. 2013, Saba & al. 2015, Jabeen & al.

2016, Faroogi & al. 2017, Naseer & al. 2017, Liu & al. 2018, Ullah & al. 2018).

All these species were previously placed in Inocybe (Fr.) Fr., but the recent

classification by Matheny & al. (2019) distributes these taxa among separate

genera.

Infrageneric classifications of Inocybe have been based on the morphology

of the basidiospores, cystidia, and stipe (Kuyper 1986), with various

morphological classifications proposed by different workers. Traditionally,

I. sect. Rimosae was placed in I. subg. Inosperma (Kithner 1980, Kuyper

1986, Stangl 1989, Larsson & al. 2009). Larsson & al. (2009), who studied

the phylogeny within I. sect. Rimosae using multigene DNA sequence

analyses, found that I. sect. Rimosae comprised two strongly supported

clades: /maculata and I. sect. Rimosae s. str. (sensu Larsson & al. 2009) The

species in /maculata clustered with those in I. sect. Cervicolores and these

two groups collectively represent I. subg. Inosperma s. str. Inocybe sect.

Rimosae s. str. [corresponding with /pseudosperma of Matheny (2005)]

has emerged as an independent clade, well distinguished from I. subg.

Inosperma s. str. species of I. sect. Rimosae s. str. are characterized by the lack

of metuloids and pleurocystidia and the presence of cylindrical to clavate

cheilocystidia; smooth, radially appressed-fibrillose to rimose pilei; and

smooth, elliptical to indistinctly phaseoliform basidiospores. In their recent

revised classification, Matheny & al. (2019) recognize I. sect. Rimosae s. str.

as an independent genus, Pseudosperma Matheny & Esteve-Rav., which we

recognize here.

During 2012-2014 we wished to analyze fungal communities in coniferous

forests of Khyber Pakhtunkhwa, Pakistan, that host a highly diverse mycota

including mycorrhizal and non-mycorrhizal species. Here we identify one

species in Inocybaceae based on morphological and molecular analyses and

reveal the phylogenetic relationships among taxa.

Materials & methods

Morphological analysis

Basidiomata were collected from two administrative divisions of Pakistan.

The first sampling site—Khanian, District Mansehra, Hazara Division, Khyber

Pakhtunkhwa province—lies immediately south of the main Himalayan range

with typical moist temperate climate and is dominated by Cedrus deodara along

Pseudosperma flavorimosum sp. nov. (Pakistan) ... 185

with Abies pindrow (Royle ex D. Don) Royle, and Pinus wallichiana. (Siddiqui &

al. 2013). The second site is Mashkun, a high mountainous region in the Swat

district of Malakand division, Khyber Pakhtunkhwa province. The climate is dry

temperate, and forests are dominated by C. deodara and P. wallichiana, with some

A. pindrow also present (Champion & al. 1965).

Basidiomata were collected and photographed in their natural habitat.

Morphological characters were recorded from fresh specimens. Color codes

follow Munsell (1975). Each collection was preserved using hot air dryers.

Tissues from dried basidiomata were rehydrated in 5% KOH prior to anatomical

observation under a Meiji MX4300h light microscope. Crush mounts were

prepared and stained with Congo red. The abbreviation (n/m/p) represents

n number of basidiospores measured from m number of fruit bodies and p number

of collections. Basidiospores were measured in lateral view. The dimensions were

recorded as (a—)b-—c(—d), where (a) = extreme minimum value, range b-c contains

minimum of 90% of the calculated values and (d) = extreme maximum value,

Q indicates |/w ratio of the spores and avQ = average Q of all spores. Other hyphal

measurements are given as ranges. The examined specimens are deposited in the

herbarium (LAH), Department of Botany, University of the Punjab, Quaid-e-Azam

Campus, Lahore, Pakistan.

Molecular analysis

Genomic DNA was extracted and amplified according to White & al. (1990),

Gardes & Bruns (1993), and Bruns (1995). The PCR products were purified and

sequenced by Macrogen Inc. (Korea). The newly generated sequence was deposited

in GenBank.

Sequences generated using forward and reverse primers were combined in

the BioEdit software version 7.2.5 (Hall 1999) to obtain a consensus sequence.

A sequence homology search was conducted using the BLAST algorithm

(http://www.ncbi.nlm.nih.gov) on 25 May 2019. Complete ITS sequences that

showed the maximum similarity and sequences of the putative closest relatives

of our species according to the published literature were included in the final

matrix (Larsson & al. 2009, Kropp & al. 2013, Latha & Manimohan 2017, Liu &

al. 2018, Matheny & al. 2019) to reconstruct phylogeny. Auritella foveata C.K.

Pradeep & Matheny (GU062740) served as outgroup following Larsson & al.

(2009) and Matheny & al. (2019). Multiple sequences were aligned using the online

webPRANK at EMBL-EBI (https://www.ebi.ac.uk/goldman-srv/webprank/).

Maximum likelihood analysis was performed using General Time Reversible

model (Nei & Kumar 2000) in MEGA version 6 (Tamura & al. 2013) at 1000

bootstrap pseudoreplicates by finding best-fit substitution model. A discrete

Gamma distribution was used to model evolutionary rate differences among sites

(5 categories (+G, parameter = 0.9359)). The rate variation model allowed for some

sites to be evolutionarily invariable ([+I], 0.0000% sites).

186 ... Jabeen & Khalid

shit

52 KJ546158 Pseudosperma mimicum

400 tk cntion Pseudosperma mimicum

KF056319 Pseudosperma mimicum

FJ904124 Pseudosperma mimicum

96 -—— FJ904134 Pseudosperma arenicola

Ge | FJ904133 Pseudosperma arenicola

ido FJ904126 Pseudosperma ‘dulcamaroides'

66 KJ726737 Pseudosperma mimicum

60 JX630909 Pseudosperma ‘dulcamaroides'

FJ904127 Pseudosperma '‘dulcamaroides'

JQ408750 Pseudosperma breviterincamatum

100 JQ408753 Pseudosperma breviterincarnatum

JQ408754 Pseudospermma breviterincarnatum

JQ408751 Pseudosperma breviterincarnatum

78, JF908162 Pseudosperma squamatum

100 |' AM882780 Pseudosperma squamatum

56 | FJ904136 Pseudosperma squamatum

FJ904132 Pseudosperma squamatum

56, JF908260 Pseudosperma spurium

100 | Fj904139 Pseudosperma spurium

JQ408794 Pseudosperma spurium

1 Er Pseudosperma flavellum

JQ724026 Pseudospema flavellum

JQ724025 Pseudosperma flavellum

JQ724027 Pseudospema flavellum

100 , MF588965 Pseudospemma pakistanense

'— MF575849 Pseudosperma pakistanense

100 , AM882769 Pseudosperma obsoletum

98 AM882770 Pseudosperma obsoletum

58 400 , AM882772 Pseudosperma perlatum

AM882771 Pseudosperma perlatum

100; NR 153171 Pseudosperma luteobrunneum

KX073581 Pseudospemma luteobrunneum

78 96> NR 153172 Pseudosperma brunneosquamulosum

ge): NR 153173 Inocybe rubrobrunnea*

| NR 152346 Pseudosperma araneosum Pseudosperma

100 | KJ729878 Pseudosperma araneosum

100 | KY440094 Pseudosperma keralense

96 NR 160442 Pseudosperma keralense

— & NR 153169 Pseudosperma griseorubidum

, MH216093 Pseudosperma sp.

u 00! 1216091 Pseudosperma sp.

MH216090 Pseudosperma sp.

100 ,MH578031 Pseudosperma sp. 7

98 MH212073 Pseudospema sp.

32 isl + KP636864 Pseudosperma sp.

| _—— @ MG495391 Pseudosperma flavorimosum

MH578027 Pseudosperma sp.

60 ; FJ904166 Pseudosperma umbrinellum

100 || FJ904163 Pseudosperma umbrinellum

FJ904165 Pseudosperma umbrinellum

oe Pseudosperma bulbosissimum

100

76 |'FJ904160 Pseudosperma bulbosissimum

FJ904158 Pseudosperma bulbosissimum

66! 99 |) _FJ904159 Pseudosperma bulbosissimum

| AM882777 Inocybe rimosa*

JF908172 Inocybe rimosa*

-- MH734760 Pseudosperma himalayense

88 + HG796995 Pseudosperma himalayense

MH745138 Pseudosperma himalayense

MH745140 Pseudosperma himalayense

HQ604610 Pseudosperma sororium

100 |HQ604617 Pseudosperma sororium

HQ604618 Pseudosperma sororium

HQ604607 Pseudosperma sororium

AM882844 Inocybe rimosa*

94 | |! AM882761 Inocybe rimosa*

6 AM882762 Inocybe rimosa*

FJ904147 Inocybe rimosa*

92, FJ904148 Pseudosperma melliolens

| FJ904149 Pseudosperma melliolens |

100 FJ936169 Inosperma quietiodor |

90 —___— KY616964 /nosperma shawarensis

ol JQ408762 Inosperma lanatodiscum

100 ; MH578017 Inosperma maculatum

'MH578013 Inosperma maculatum

GU062740 Auritella foveata ]

Inosperma

Out group

'————

0.05

Fic. 1. Molecular phylogenetic analysis of Pseudosperma flavorimosum based on ITS sequences.

Sequence generated during this study is marked by @. Scale bar = nucleotide substitutions per site.

The names of the taxa are written according to Matheny & al (2019). Those represented in quotes

lack type specimens. * represents taxa that are not discussed in Matheny & al (2019) and given

names according to their publication source and GenBank record.

Pseudosperma flavorimosum sp. nov. (Pakistan) ... 187

Phylogeny

The NCBI BLAST search revealed that the Pseudosperma flavorimosum

sequence (MG495391) shared 89-90% identity with unidentified “Inocybe

sp. sequences (MH578031, MH578027, MH212073) from USA. Closely

related ITS sequences were retrieved from GenBank for generating a

phylogeny for the Pakistani taxon based on 77 nucleotide sequences. The final

ITS matrix comprised a total of 1403 positions of which 734 were conserved,

442 were variable, 337 were parsimony informative, and 99 were singletons.

The P. flavorimosum sequence forms a sister clade with unidentified Inocybe

sequence KP636864, and both sequences clustered on the same branch with

two more unidentified sequences (MH212073, MH578031) in the same clade

as unidentified Pseudosperma sequence MH578027 with 82% bootstrap

support and were separated from the clade containing P bulbosissimum

(Kihner) Matheny & Esteve-Rav., P. himalayense (Razaq & al.) Matheny

& Esteve-Rav., P. melliolens (Kihner) Matheny & Esteve-Rav., P. rimosum

(Bull.) Matheny & Esteve-Rav., P. sororium (Kauffman) Matheny & Esteve-

Rav., and P. umbrinellum (Bres.) Matheny & Esteve-Rav. Our phylogenetic

tree (Fic. 1) places the above Pseudosperma species in the same relationships

as shown in Larsson & al. (2009, there as Inocybe) and Matheny & al. (2019).

Pseudosperma flavorimosum Jabeen & Khalid, sp. nov. FIGs. 2, 3

MB 823494

Differs from Pseudosperma sororium by its smooth ellipsoidal to slightly amygdaliform

basidiospores with a broad base and narrow apex.

Type: Pakistan. Khyber Pakhtunkhwa province: Hazara division, Mansehra district,

Kaghan valley (Khanian), 2500 m a.s.l., on soil under Pinus wallichiana A.B. Jacks., 3

Aug 2014, Muhammad Burhan SJ103 (Holotype, LAH35042; GenBank: MG495391).

EryMmo.ocy: The specific epithet (Lat.) refers to the yellowish and rimose pileus.

Piteus 45-5.1 mm wide, initially conico-convex with a prominent umbo,

becoming broadly conico-convex with a prominent umbo at maturity, umbo

conical when young, becoming more obtuse in maturity, surface yellowish

brown (10YR8/8) with dark brown (10R4/12) fibrillose streaks, dry, fibrillose,

radially rimose towards the margin, margin incurved. LAMELLAE adnate to

adnexed, moderately close, <2.5 mm deep, narrow, initially yellowish brown

(10YR7/8), becoming dark brown (10R6/14) at maturity, edges fimbriate.

LAMELLULAE in different lengths. Stripe 75 x 7 mm, central, cylindrical,

narrower towards the apex and wider towards the base, apex furfuraceous,

base splitting, surface dry, context off white under a yellowish (10YR8/8) to

dark brown (5YR2/6) cuticle.

188 ... Jabeen & Khalid

BASIDIOSPORES [40/2/2] (7.8—)9.5-12.8(-13.31) x (5.7—)6.3-7.8(-8.9) um,

Q = (1.21-)1.39-1.91(-1.98), avQ = 1.62, elliptical, amygdaliform with

broad apex and narrow base, smooth, uniguttulate. Basrp1a (14.8-)

26.4-31.2(-36.7) x (10.0-)10.5-10.8(-11.8) um, clavate, 4-spored, thin-

walled, guttulate. CHEILOCYSTIDIA (18.5—)19.2—21.7(-25.3) x (7.5-)7.7-8.6

(-9.1) um, broadly clavate, thin-walled, frequently arranged on lamellar

edges. PLEUROCYSTIDIA absent. PILEIPELLIS hyphae (3.8—)4.9-6.3(-6.6) um

diam., intricate trichoderm, non-encrusted, frequently septate, filamentous,

fusoid terminals, clamped septa common, hyaline. StiprriPELLis hyphae

(4.5-)4.7-8.7(-9.2) um diam., trichoderm, septate, filamentous, rarely

branched, clamp connections not observed.

ECOLOGY & DISTRIBUTION—among conifers in Himalayan moist and dry

temperate forests; known only from Pakistan.

ADDITIONAL SPECIMEN EXAMINED—PAKISTAN. KHYBER PAKHTUNKHWA PROVINCE:

Malakand division, Swat district, Mashkun, 2500 m a.s.l., on soil under Cedrus deodara

(Roxb. ex D. Don) G. Don, 5 Sep 2013, Sana Jabeen MTI (LAH35043).

Discussion

Pseudosperma flavorimosum is characterized by its yellowish brown pileus

with dark brown striations and radially rimose margins and a stipe with a

rimose base and losing its stipitipellis over time. It is characterized with

smooth ellipsoidal to slightly amygdaliform basidiospores with a broad base

and narrow apex and broadly clavate cheilocystidia. ITS sequence analysis

clusters P flavorimosum with other Pseudosperma species. Taxa within

the genus separated into several clades, with P flavorimosum clustering in

subclade A. Pseudosperma melliolens, described from France, also looks like

a typical P. rimosum (Bull.) Matheny & Esteve-Rav. (Bon 1997).

Pseudosperma flavorimosum shares morphological similarities with

P. sororium, as originally described by Kauffman (1924). But the elliptical

or elongate-ellipsoid basidiospores of P. sororium are not truly subreniform,

sub-inequilateral and are obtuse at both ends, distinguishing from

P. flavorimosum.

Pseudosperma himalayense, a recently described taxon from Pakistan,

differs morphologically from P. flavorimosum by its paler pileus (varying from

white to cream, pale yellow, olive yellow, and light brown to camel brown)

with dentate margins, ellipsoid to slightly amygdaliform basidiospores, and

clavate to cylindrical cheilocystidia (Liu & al. 2018).

Pseudosperma umbrinellum bears yellowish to reddish brown pileus with a

dark centre and strongly rimose and lighter periphery while P. flavorimosum

Pseudosperma flavorimosum sp. nov. (Pakistan) ... 189

bie =

+ sate

Fic. 2. Pseudosperma flavorimosum (LAH35042, holotype). Basidiomata. Scale

Photos by Sana Jabeen

bars = 1 cm.

190 ... Jabeen & Khalid

Ds —_— Le

Fic. 3. Pseudosperma flavorimosum (LAH35042, holotype). A. Basidiospores; B. Cheilocystidia; C.

Basidia; D. Stipitipellis; E. Pileipellis. Scale bars = 10 um. Drawings by Sana Jabeen.

Pseudosperma flavorimosum sp. nov. (Pakistan) ... 191

bears yellowish pileus with brown striations, but the anatomical features

are identical to P. rimosum. The taxa adjacent to P. flavorimosum in the

phylogenetic tree are undescribed morphologically but form separate

lineages within subclade A of Pseudosperma with a strong bootstrap value

supporting P. flavorimosum as a distinct taxon.

Acknowledgments

This work was financially supported by Higher Education Commission (HEC)-

Pakistan under Indigenous PhD Fellowship. Sincere thanks to Dr. K.P. Deepna

Latha (Fungal Diversity Division, Department of Botany, University of Calicut,

Kerala, India) for valuable comments and suggestions, which greatly improved the

manuscript. Thanks are also due to Dr. Arooj Naseer (Centre for Undergraduate

Studies, University of the Punjab, Lahore, Pakistan) for also making corrections as

presubmission reviewer. We are also grateful to Dr. Abdul Rehman Khan Niazi and

Dr. Najam ul Sehar Afshan, Muhammadah Khalid, Abdul Rehman, Muhammad

Burhan, Fatima-tu-Zahra, Muhammad Umar Khan Niazi, and all laboratory fellows

for accompanying the field trips to different areas of Pakistan.

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MY COTAXON

January-March 2020— Volume 135, pp. 195-201

https://doi.org/10.5248/135.195

Three Rinodina species new to China

X1A0-JIA ZHENG & QIANG REN*

College of Life Sciences, Shandong Normal University,

Jinan 250014, China

" CORRESPONDENCE TO: rendagiang@hotmail.com

ABSTRACT—Rinodina ascociscana, R. subminuta, and R. trevisanii are reported as new to

China. A detailed description of each species is provided, and comparisons with related

species are made.

Key worps—Ascomycota, lichenized fungi, Physciaceae

Introduction

Rinodina (Physciaceae, Caliciales), which comprises more than 300 species

worldwide (Sheard & al. 2017), occurs on rock, bark, mosses, soil, or debris

and is characterized by a crustose thallus, mostly lecanorine apothecia with

brown to black discs, a hyaline hymenium, a red-brown or (rarely) blue-grey

epihymenium, clavate asci, and brown, primarily 1-septate (rarely 3-septate)

to submuriform ascospores with + unequally thickened walls (Mayrhofer &

Moberg 2002; Sheard 2004, 2010).

Although many have recently contributed to crustose lichen diversity in

China (e.g., Kou & al. 2013, Ren & Zhao 2014, Ren 2017, Ren & al. 2018, Zhou

& Ren 2018), knowledge of Rinodina species remains poor in this country.

Forty-five species and three forms have been reported from China (e.g.,

Mayrhofer 1984; Wei 1991; Abdulla & Wu 1994, 1998; Aptroot & Seaward

1999; Aptroot & Sipman 2001; Aptroot & Sparrius 2003; Joshi & al. 2014;

Kondratyuk & al. 2016; Sheard & al. 2017). During a survey of Rinodina

specimens, we detected three species new for China: Rinodina ascociscana,

R. subminuta, and R. trevisanii.

196 ... Zheng & Ren

Materials & methods

The specimens examined in this study were housed in SDNU (the Plant

Herbarium of Shandong Normal University, Jinan, China) and HMAS-L (Herbarium

of Mycology, Chinese Academy of Sciences, Beijing, China).

Morphological and anatomical characters were observed using an Olympus

SZ51 stereo microscope and an Olympus CX21 light microscope. Lichen substances

were detected by thin layer chromatography (TLC) with solvent systems A, B,, and

C (Elix 2014). Photos were taken using Olympus SZX16 and BX61 microscopes

with a DP72 digital camera. We identified the specimens primarily from Sheard &

al. (2017).

Species

Rinodina ascociscana (Tuck.) Tuck., Gen. Lich.: 124 (1872). FIG:

THALLUS crustose, thick, well-developed, surface continuous, ochraceous

to brownish; prothallus absent; vegetative propagules absent.

APOTHECIA lecanorine, not erumpent, 0.5-1.2 mm in diam, scattered,

sessile, narrowly attached; pisc brown to black, epruinose, persistently

plane, thalline margin concolourous with thallus, often with radially cracked,

sometimes becoming flexuous; excipular ring present, confluent, prominent,

darker than thalline margin. THALLINE EXCIPLE 50-80 um wide; proper

exciple hyaline, 15-25 um wide; HYMENIUM hyaline, 80-110 um high,

paraphyses 2-3 um in diam.; EPIHYMENIUM red-brown, 10-20 um high;

HYPOTHECIUM hyaline, 20-40 um deep. Asci clavate, 8-spored; ASCOSPORES

brown to dark brown, type A development, Physcia-type, ellipsoid, 1-septate,

27.5-35 x 12.5-15 um, torus absent, ornamented walls not evident. PYCNIDIA

not seen.

CHEMISTRY—No lichen substances detected by TLC.

SPECIMENS EXAMINED: CHINA. JiLin: Antu County, c. 30 km from Erdaobaihe

town toward Changbaishan mountain and lake, alt. c. 1100 m, on bark of conifer tree,

2011.09, Sohrabi with Ghobad-Nejhad 16644 (HAMS-L 120853); Helong City, Mt.

Zengfeng, alt. 1600 m, on bark of deciduous trees, 2011.08.19, Y.L. Cheng 20119406A,

20119440B, 20119844 (SDNU).

SUBSTRATE & DISTRIBUTION—Rinodina ascociscana occurs on bark of

conifers or deciduous trees in mixed forests with Acer, Abies, and Larix. It was

previously known from North America (Sheard 2010) and north-eastern Asia

(Japan, Siberia, and South Korea; Sheard & al. 2017); new from China.

CoMMENTS—Our material closely matched the descriptions of Sheard

(2010) and Sheard & al. (2017). Rinodina ascociscana is characterized by the

often radially cracked thalline margin, the large Physcia-type ascospores

Rinodina spp. new to China... 197

Fic. 1. Rinodina ascociscana (SDNU, Cheng 20119844). A. Thallus with substratum; B. Apothecia

showing radially cracked thalline margin; C. Section of apothecium; D. Immature Physcia-type

ascospore; E. Physcia-type ascospore mature. Scale bars: A = 1 mm; B = 0.5 mm; C = 100 um;

D, E= 10 pm.

with type A development, and the absence of lichen substances. Rinodina

dolichospora Malme differs by its thalline margin that is not radially cracked

and Pachysporaria-type I ascospores (Sheard 2010).

Rinodina subminuta H. Magn., Bot. Not. 1947: 44 (1947). FIG. 2

THALLUS crustose, thin to endophloeodal, continuous to rimose, surface

matte or glossy, yellowish or light to dark grey; prothallus absent; vegetative

propagules absent.

APOTHECIA lecanorine, erumpent; pisc black, epruinose, plane,

sometimes convex, 0.4-0.65 mm in diam; thalline margin concolourous

with thallus, excipular ring absent. THALLINE EXCIPLE 50-80 um wide;

proper exciple hyaline, 15-25 um wide; HYMENIUM hyaline, 60-80 um high,

paraphyses 2.5-4.5 um in diam.; EPIHYMENIUM red-brown; HYPOTHECIUM

hyaline, 30-60 um deep. Asci clavate, 8-spored, 45-70 x 20-30 um;

198 ... Zheng & Ren

Fic. 2. Rinodina subminuta (HMAS-L 073131). A. Thallus with substratum; B. Section of

apothecium; C. Mature ascospore. Scale bars: A = 0.5 mm; B = 50 um; C = 10 um.

ASCOSPORES brown, type A development, ellipsoid, 1-septate, Physcia-

type, 17-20(-22.5) x 8-9.5(-10) um, torus present, ascospore walls not

ornamented. PYCNIDIA not seen.

CHEMISTRY—Zeorin, + one unknown substance with UV+ blue (TLC:

A=f, B=7,.C=7):

SPECIMENS EXAMINED: CHINA. JILIN: Mt. Changbai, Tianchi, alt. 1850 m, on Betula

ermanii Cham. (Betulaceae), 1984.08.14, X.D. Lu 848410-1 (HMAS-L 0141286); Hot

spring, on Betula ermanii, 1994.08.06, Wei & al. 94423 (HMAS-L 073131); South Slope,

alt. 1900 m, on Betula ermanii, 1983.08.03, J.C. Wei & J.B. Chen 6586-1 (HMAS-L

0141288); on Betula ermanii, 2003.8.20, H.M. LiCBS114-1 (HMAS-L 0141273); North

Slope, alt. 1810 m. on Sorbus sp. (Rosaceae), 2018.08.19, Q. Ren 7119, 7120 (SDNU);

on Betula ermanii, 2018.08.19, Q. Ren 7029, 7040 (SDNU).

SUBSTRATE & DISTRIBUTION— Rinodina subminuta occurs on the bark of Betula

ermanii and Sorbus sp. in open sites of deciduous forests. It was previously

known from North America and Siberia (Sheard 2010), and from north-eastern

Asia (Japan, Siberia, and South Korea; Sheard & al. 2017); new from China.

Rinodina spp. new to China... 199

COMMENTS—Otur material closely matches the description in Sheard (2010).

Rinodina subminuta is characterized by a thin thallus, erumpent apothecia,

Physcia-type ascospores with type A development, and the presence of

zeorin. Rinodina orientalis Sheard differs by strongly convex discs of mature

apothecia, smaller ascospores (16-18 x 7-8.5 um), and the absence of

lichen substances (Sheard & al. 2017).

Rinodina trevisanii (Hepp) K6rb., Parerga Lichenol.: 70 (1859) FIG. 3

THALLUS crustose, very thin, surface continuous to rimose, brownish;

prothallus absent; vegetative propagules absent.

APOTHECIA lecanorine, not erumpent, 0.4-0.6 mm in diam, sessile,

broadly or narrowly attached; pisc brown to black, epruinose, plane,

frequently becoming convex, thalline margin concolourous with thallus,

often becoming excluded or margin biatorine, excipular ring not prominent.

THALLINE EXCIPLE 50-70 um wide; proper exciple hyaline, c. 15 um wide;

HYMENIUM hyaline, 65-80 um high, paraphyses 1.5-2.5 um in diam.;

EPIHYMENIUM red-brown, 10-30 um high; HyPOTHECIUM hyaline, 30-110

um deep. Asc clavate, 8-spored, 50-60 x 17.5-30 um; ASCOSPORES brown,

type A development, Physconia-type, ellipsoid, 1-septate, 18-—22.5(-23.5) x

8-9.5(-10) um, torus sometimes present but not prominent, ascospore walls

not ornamented. PYcNIDIA not seen.

CHEMISTRY—+Zeorin.

SPECIMENS EXAMINED: CHINA. HEILONGJIANG: Heihe City, Mt. Xifeng, alt. 220

m, on bark, 2009.08.17, Q. Ren 1192 (SDNU); Mohe City, Mt. Guanyinshan, alt.

554 m, on bark, 2009.08.13, Q. Ren 1179 (SDNU); Tahe City, Mt. Baikalu, alt. 800

m, on bark, 2011.08.25, Y.L. Cheng 20125237B (SDNU); Daxing/anling, alt. 800

m, on tree trunk, 1984.08.07, X.Q. Gao 105 (HMAS-L 072400). Jrt1n: Antu City,

Mt. Changbai, alt. 1750 m, on Larix sp. (Pinaceae), 1984.08.28, X.D. Lu 848421-5

(HMAS-L 076349); North Slope of Mt. Changbai, alt. 1810 m. on Betula ermanii,

2018.08.19, Q. Ren 7118 (SDNU).

SUBSTRATE & DISTRIBUTION— Rinodina trevisanii occurs on bark of Larix

sp., Betula ermanii, and other trees in mixed forests. It was previously known

from Europe, Asia (Caucasus, Turkey, Siberia), western North America

(Mayrhofer & Sheard 2007, Sheard 2010), Kazakhstan (Hauck & al. 2013a),

and western Mongolia (Hauck & al. 2013b); new from China.

COMMENTS—Our material closely corresponds with the description of

Sheard (2010). Rinodina trevisanii is characterized by a thin thallus, mostly

scattered apothecia with frequently convex discs and margins often biatorine,

and Physconia-type ascospores with type A development. Rinodina archaea

200 ... Zheng & Ren

Fic. 3. Rinodina trevisanii (HMAS-L 076349). A, B. Thallus with substratum; C. Immature

ascospores; D. Mature ascospore. Scale bars: A = 1 mm; B = 0.5 mm; C, D = 10 um.

(Ach.) Arnold differs by its relatively thick, typically areolate thallus, often

compressed (hence angular) apothecia, persistently plane discs of mature

apothecia, and larger ascospores (Mayrhofer & Sheard 2007).

Acknowledgments

HMAS-L is thanked for loan of specimens. We are very grateful to Prof.

H. Mayrhofer (Institute of Biology, University of Graz, Austria) and Dr. S.Y. Guo

(State Key Laboratory of Mycology, Institute of Microbiology, Chinese Academy

of Sciences, Beijing, China) for reviewing the manuscript. This project is supported

by the National Natural Science Foundation of China (31750001).

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MY COTAXON

January-March 2020—Volume 135, pp. 203-212

https://doi.org/10.5248/135.203

Gymnopus barbipes and G. dysodes,

new records for Pakistan

MALKA SABA", JUNAID KHAN’, SAMINA SARWAR},

HASSAN SHER’, ABDUL NASIR KHALID‘

“Department of Plant Sciences, Quaid-i-Azam University,

Islamabad, 45320, Pakistan

*Center for Plant Sciences and Biodiversity, University of Swat, Swat, Pakistan

*Department of Botany, Lahore College for Women University, Lahore, Pakistan

‘Department of Botany, University of the Punjab,

Quaid-e-Azam Campus, Lahore, 54590, Pakistan

*CORRESPONDENCE TO: rustflora@gmail.com; msaba@qau.edu.pk

AsBstTRACT— Morphological and phylogenetic analyses were conducted to identify Gymnopus

species collected in Pakistan during 2013-14. Phylogenetic analysis was performed using

internal transcribed spacer region (ITS) of ribosomal DNA sequences. Among the collected

taxa, we identified Gymnopus barbipes and G. dysodes, represent new records for Pakistan.

Their detailed descriptions and illustrations are also provided.

Key worps —Khyber Pakhtunkhwa, Omphalotaceae, pine forests

Introduction

Gymnopus (Pers.) Roussel (Omphalotaceae, Agaricales) is a large fungal

genus, which consists of c. 300 species distributed worldwide (Jang & al.

2016). It is characterized by a collybioid basidioma; a convex to applanate

or slightly concave pileus; free, emarginate or adnate and crowded to fairly

distant lamellae; a central stipe; a white spore print; inamyloid or non-

dextrinoid hyphae with clamp connections; ellipsoid to oblong, thin-walled,

hyaline, inamyloid basidiospores. Cheilocystidia are often present while

pleurocystidia are usually absent (Antonin & al. 2013, Jang & al. 2016, Saba &

Khalid 2014). Seven species of Gymnopus—worldwide a relatively common

204 ... Saba & al.

saprobe on leaf litter and dead wood—have been reported in Pakistan

(Ahmad 1980, Iqbal & Khalid 1996, Shibata 1992, Sultana & al. 2011, Saba &

Khalid 2014). In view of the high worldwide diversity, we would expect the

diversity of Pakistani Gymnopus to be much greater than currently reported.

To explore the diversity of indigenous Gymnopus spp., we collected

specimens in Pakistan, which we identified through morphological and

phylogenetic analyses. Here we report Gymnopus barbipes and G. dysodes as

new records for Pakistan.

Materials & methods

Morphological evaluation

Basidiomata were collected, photographed, characterized morphologically,

dried, and deposited in the University of Swat Herbarium (SWAT) in Pakistan and

Farlow Herbarium of Harvard University in USA (FH). Color designations are from

Munsell (1975). Microscopical observations were made from slide preparations from

dried specimens mounted in 5% potassium hydroxide (KOH) or Melzer’s reagent

under a Meiji Techno MX4300H biological microscope. Dimensions of anatomical

features (basidiospores, basidia, cystidia, hyphae) were calculated from at least 25

measurements using an ocular micrometer and 100x oil-immersion objective with

x = arithmetic means of spore lengths and widths for all spores measured. Line

drawings were made with a camera lucida.

DNA extraction, PCR amplification, DNA sequencing

Genomic DNA was extracted from a small piece of basidioma by a modified

CTAB method (Lee & al. 1988) using the Qiagen DNeasy Plant Mini Kit (cat. no.

69104). The entire rDNA internal transcribed spacer region (ITS = ITS1+5.8S+ITS2)

was targeted with primer pair ITS1F/ITS4 (White & al. 1990, Gardes & Bruns

1993) and then amplified using Econo Taq DNA Polymerase following the PCR

cycling parameters: initial denaturation (94 °C for 1 min), 35 cycles (94 °C for 1

min, 53 °C for 1 min, 72 °C for 1 min), and final extension (72 °C for 8 min). The

PCR amplicons were sent to Macrogen (Republic of Korea) for purification and

bidirectional sequencing.

Sequence alignment & phylogenetic analysis

The ITS sequence of MSM0034 comprised 700 base pairs and that of ING-21

comprised 691 base pairs. We retrieved closely related ITS sequences via a BLAST

search of GenBank for subsequent phylogenetic analysis, according to Saba &

Khalid (2014). Sequences used in phylogenetic construction are shown in TABLE 1.

Sequences were manually edited and assembled using BioEdit (www.mbio.

ncsu.edu/bioedit/bioedit.html). Following Dentinger & al. (2011) for complete ITS

sequences, all sequences were trimmed with the conserved motifs 5’-(...GAT) CATTA-

and -GAccT (CAAA...)-3’ with the enclosed aligned portions analysed. Sequences of

Gymnopus spp. new for Pakistan ... 205

TABLE 1: Gymnopus and Omphalotus ITS sequences analyzed.

New sequences in bold.

TAXON COUNTRY GENBANK # COLLECTION/ VOUCHER #

G. barbipes Pakistan MK450334 MSM 0034

USA KJ416265 TENN 69173

USA KJ416266 TENN 69173

USA KJ416269 TENN 67858-T

G. biformis Costa Rica DQ450064 TFB7843

Costa Rica DQ450063 TFB7820

G. confluens Russia AY256697 TENNS58242

— HM240527 UBC F19677

G. cylindricus Costa Rica AY256696 TENNS58024

Costa Rica DQ450057 TFB10091

G. dryophilus Italy JX536157 BRNM707149

G. dysodes Pakistan MT114698 SWAT 001355

USA AF505778 TENN 59141

USA KY026666 TENN F-61125

Czech Republic JX536158 BRNM712600

G. erythropus Slovakia DQ449996 SAV X12002

USA DQ449998 WTU JFA12910

G. fibrosipes Costa Rica AF505763 TENNS56660

= AY842953 PR23TN

G. fusipes France AY256710 TENNS59217

Austria AF505777 TENN59300

G. gibbosus — AY 263437 AWW66

— AY 263438 AWW95

G. impudicus Russia KJ416263 TENN 60094

Russia KJ416264 Culture

G. luxurians Pakistan KF803760 MSM 001

Pakistan KF803761 MSM 002

USA AY256709 TENN57910

Switzerland DQ450022 TENNS50619

G. menehune Pakistan KF803762 MSM 003

_— JN182864 _—

— AY263426 DED5866

G. mesoamericanus Costa Rica DQ450036 TENN58106

Costa Rica AF505768 NYBG REH7379

G. polygrammus Puerto Rico DQ450028 TENN56589

= AY 842954 PR 2542TN

G. readiae New Zealand DQ450034 TENN53687

New Zealand HQ533036 PDD95844

G. subcyathiformis Puerto Rico DQ450041 TENNS58130

Dominican Republic DQ450042 TENN59550

G. subpruinosus USA DQ450026 TENNS6242

USA DQ450027 TENN59477

O. illudens USA AY313271 TENN54507

O. olearius France AY313277 Culture 9061b

O. olivascens USA AY313281 TENN55337

206 ... Saba & al.

Omphalotus spp. were used as outgroup based on results reported by Moncalvo &

al. (2002) and Mata & al. (2004).

We aligned our new sequences with GenBank sequences from related taxa

using ClustalX (Thompson & al. 1997) and manually edited where necessary using

BioEdit (Hall 1999). A maximum likelihood analysis was performed via CIPRES

Science Gateway (Miller & al. 2010) employing RAxML-HPC v.8 and Rapid

bootstrap analysis/search for best scoring, with the topology assessed by 1000

bootstrap replicates.

Taxonomy

Gymnopus barbipes R.H. Petersen & K.W. Hughes,

N. Amer. Fung. 9(3): 2 (2014). Fic. 1

Pileus 25 mm diam., broadly convex, expanding with age to plane,

flat, thin; margin straight; surface matte, smooth; central disc brownish

orange (5YR5/8) to strong brown (5YR4/6) or light brown (5YR8/8).

Lamellae adnexed or free, subdistant, light orange (2.5YR8/8), edges

even, concolorous. Lamellulae 1-2 tiered. Stipe 40 x 5 mm, central, equal,

cylindrical, hollow, sheathed in a thin, off-white, felty covering, brownish

orange (5YR5/8). Odor & taste not distinctive.

Basidiospores 5-7 x 2.3-4 um [x = 5.6 x 3.0 um, Q = 1.2-1.6], ellipsoid

in profile view, slightly flattened adaxially, rarely ovoid, smooth, thin-

walled, aguttulate, hyaline in KOH, inamyloid. Basidia 23-28 x 5-6 um,

clavate, 4-spored (rarely 2-spored) basidia, thin-walled, hyaline in KOH;

sterigmata 3-3.7 um. Basidioles subclavate, abundant. Pleurocystidia absent.

Cheilocystidia abundant, sometimes constricted at neck. Pileipellis a cutis,

repent hyphae cylindrical, 5-14 um, thin-walled, hyaline in KOH. Stipe

hyphae cylindrical, 5-9 um, non-encrusted, hyaline in KOH. Caulocystidia

cylindrical, knobby, abundant, hyaline, thin-walled, 9-14 um in diam.

Clamp connections present in all tissues.

MATERIAL STUDIED: PAKISTAN, KHYBER PAKHTUNKHWA, Shangla, Yakh Tangay,

under Pinus wallichiana A.B. Jacks., 2 September 2013, Malka Saba & Abdul Nasir

Khalid, MSM#0034 (FH00304574; GenBank MK450334).

CoMMENTS—Our Pakistani specimen agrees morphologically with the

original protologue of G. barbipes except for its broader pileus (cited as 5-15

mm diam. in Petersen & Hughes 2014), robust stipe, and presence of ovoid

basidiospores in addition to ellipsoid basidiospores. Gymnopus barbipes

was described from southeastern North America and characterized by

its occurrence on leaf litter and absence of well-developed pleurocystidia

and cheilocystidia (Petersen & Hughes 2014). Our Pakistani collection

Gymnopus spp. new for Pakistan ... 207

Fic. 1. Gymnopus barbipes (MSM 0034). A, B. Basidiomata; C. Basidia; D. Cheilocystidia;

E. Basidiospores; F. Caulocystidia; G. Pileipellis. Scale bars: A, B = 10 mm; C, D = 10 um;

E=5 um; KR G= 25 um.

208 ... Saba & al.

represents the first occurrence of G. barbipes reported since the Tennessee

collection was described.

Gymnopus dysodes (Halling) Halling, Mycotaxon 63: 364 (1997). Fia. 2

= Collybia dysodes Halling, Mycol. Mem. 8: 79 (1983).

Pileus 15-30 mm across, conic to campanulate at first, later plano-convex

to plane, color light pinkish brown (5YR 6/4) to pale reddish brown (SYR

4/4) with a darker (7.5YR 1/2) disc, surface dry, dull, disc smooth, otherwise

sulcate, pileus margin crenate or eroded in some specimens, context thin,

1-2 mm at the disc, <lmm at the margins, flesh pinkish brown (7.5YR

9/4), unchanging when cut. Lamellae adnexed, seceding, narrow (1.5-2.5

mm) wide, distant, initially creamy pinkish (SY 9/4), becoming light brown

(7.5YR 7/4) and finally dark brown (5Y 3/4). Lamellulae present, rare, short,

mostly in single tier. Stipe 10-20 x 4—7 mm, cylindrical in young specimens,

compressed in mature specimens, equal to thickening downward, surface

reddish brown (7.5 YR 9/4), smooth or slightly scaly with hirsute base,

interior hollow at the center, pith at apex and in base whitish fibrillose,

concolorous with stipe surface, unchanging when cut. Odor of garlic. Taste

not noted.

Basidiospores 7.3-8.6 x 3.4-3.9 um, [x = 7.9 x 3.7 um, Q = 1.8-2.4],

narrowly ellipsoid to oblong, smooth, thin-walled, hyaline in both KOH

and Congo Red, inamyloid. Basidia 22-25 x 4-6 um, clavate, with four

sterigmata, thin-walled, hyaline in KOH. Basidioles subclavate to cylindrical,

numerous. Cheilocystidia 25-35 x 5-7 um, variously shaped, cylindrical,

narrowly utriform or fusoid, flexuous, bottle shaped, horned/rostrate, thin-

walled, clamped at bases, hyaline in KOH. Pleurocystidia absent. Pileipellis

a cutis, individual hyphae 5-10 um diam., terminals mostly acute, some

cylindrical, others with irregular terminals, thin-walled.

MATERIAL STUDIED: PAKISTAN, KHYBER PAKHTUNKHWA, District Swat, village

Ingaro Dherai, in clusters on decaying woodchips of deciduous trees, 23 July 2014,

Junaid Khan, ING-21 (SWAT 001355; GenBank MT114698).

CoMMENTS—Our Pakistani specimen agrees morphologically with the

original protologue of Collybia dysodes by Halling (1983). Gymnopus

dysodes falls within G. sect. Impudici, characterized by an unpleasant smell

resembling rotten cabbage, onion, garlic, etc.

Gymnopus dysodes, a North American species, is characterized mainly by

its lignicolous habit, dark reddish brown to cinnamon brownish basidiomata,

cinnamon brown lamellae, dark to light reddish brown stipe, a pungent

Gymnopus spp. new for Pakistan ... 209

Fig. 2. Gymnopus dysodes (ING-21). A-C. Basidiomata; D. Basidiospores; E. Basidiole and

basidium; F. Cheilocystidia; G. Pileipellis hyphae (with clamped septa). Scale bars: A-C = 5 mm;

D, E=5 um; F = 10 um; G= 15 um.

garlic-like odour, and cylindrical to flexuous cheilocystidia (Halling 1983).

Our collection was also collected on decaying wood chips of deciduous

trees; both morphology and sequence analysis support its identification as

210 ... Saba & al.

vs , polygrammus AY 842954

95 . polygrammus (002

100 |G. subpruinosus DQ450027

88 G. subpruinosus DQ450026

95 G. fibrosipes AY 842953

G. fibrosipes AF505763 Clade|

92 sop G: /uecurians DQ450022 ede:

G. luxurians AY256709

100 °G. hecurians KF803760

G. luxurians KF803761

100 |G. gibbosus AY263437

G. gibbosus AY263438

100 G. readiae DQ450034

G. readiae HQ533036

72 100 G. confluens HM240527

aa 7 con anit iris DOA San

subcyathiformis

100 G. subcyathiformis DQ450042

99 G. menehune AY263426

100 99 4G. menehune KF803762 Clade Il

G. menehune JN182864

100 | G. mesoamericanus AF505768

G. mesoamericanus DQ450036

100 100 G. cylindricus AY256696

81 G. cylindricus DQ450057

100 G. biformis are

G. by for ‘mis DQ4500

G. opliles JX536158

99 G. dryophilus herent

= ce Se fiver BO49096

. erythropus

= G MTI1

100

S8G. dysodes AF505778

98 G. dysodes KY 026666

100 G, impudicus KJ416263

100 100 G. impudicus KJ416264 Clade lV

100 450334

09 G. <G: barbpes KJ416266

G. barbipes K3416269 T

G. barbipes KJ416265

100 G. fusipes AY256710

G. fusipes AF505777 CladeV

100 Omphalotus illudens AY313271

73 O. olearius AY313277 Outgroup

O. olivascens AY313281

Clade Ill

Fic. 3. Molecular phylogenetic analysis of Gymnopus sequences by the Maximum Likelihood

method. The bootstrap consensus tree inferred from 1000 replicates implies the evolutionary

history. The percentage of trees in which the taxa cluster together is shown next to the branches.

New sequences reported in this study are in bold; T = holotype.

G. dysodes. There are no previous reports of G. dysodes from the country,

which makes this species an addition to the funga of Pakistan.

Gymnopus impudicus (Fr.) Antonin &al., a similar species, can be separated

by its darker basidiomata (dark brown pileus and almost black stipe base),

fetid (not garlic-like) odour, smaller basidiospores (5.5-7.5 x 3.5-4) um, and

moniliform to coralloid cheilocystidia (20-40 x 3-9 um; Noordeloos 1995).

Phylogeny

A total of 45 nrITS sequences were used in the molecular phylogenetic

analysis, 42 representing 18 Gymnopus taxa and three representing three

Omphalotus in-group taxa (TABLE 1). The final dataset had 935 sites of which

475 were conserved, 454 were variable, and 423 were parsimony informative.

Gymnopus spp. new for Pakistan ... 211

Maximum likelihood analysis clustered the included nucleotide sequences

of Gymnopus into five well supported clades. Species within the Clades I

and II are members of Gymnopus subsect. Vestipedes. Clade III correlates

to G. subsect. Levipedes, clade IV to G. subsect. Impudici, and clade V to

G. sect. Gymnopus. Both Pakistani species fall within clade IV (G. subsect.

Impudici). Gymnopus dysodes and G. barbipes clustered with sequences from

USA with strong bootstrap support (100%).

Acknowledgments

This work was financed by Higher Education Commission (HEC), Pakistan

under Phase IJ, Batch I, Indigenous PhD Fellowships Program for 5000 scholars

and through the International Research Support Initiative Program (IRSIP).

We are thankful to Dr. Vladimir Antonin (Moravian Museum, Brno, Czech

Republic) and Dr. Muhammad Fiaz (Hazara University, Mansehra, Pakistan) for

critically reviewing the manuscript.

Literature cited

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Antonin V, Sedlak P, TomSovsky M. 2013. Taxonomy and phylogeny of European Gymnopus

subsection Levipedes (Basidiomycota, Omphalotaceae). Persoonia 31: 179-187.

https://doi.org/10.3767/003158513X674043

Dentinger BTM, Didukh MY, Moncalvo JM. 2011. Comparing COI and ITS barcode

markers for mushrooms and allies (Agaricomycotina). PLoS One 6(9): e25081.

https://doi.org/10.1371/journal.pone.0025081

Gardes M, Bruns TD. 1993. ITS primers with enhanced specificity for Basidiomycetes:

application to the identification of mycorrhizae and rusts. Molecular Ecology 2: 113-118.

https://doi.org/10.1111/j.1365-294X.1993.tb00005.x

Hall TA. 1999. Bioedit: a user-friendly biological sequence alignment editor and analysis

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Halling RE. 1983. The genus Collybia (Agaricales) in Northeastern United States and adjacent

Canada. Mycological Memoirs 8. 148 p.

Iqbal SH, Khalid AN. 1996. Materials for the fungus flora of Pakistan. I. Checklist of Agarics,

their distribution and association with the surrounding vegetation. Science International

(Lahore) 8(1): 51-64.

Jang S, Jang Y, Lim YW, Kim C, Ahn BJ, Lee SS, Kim JJ. 2016. Phylogenetic identification of

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G. subnudus. Mycobiology 44(3): 131-136.

Lee SB, Milgroom MG, Taylor JW. 1988. A rapid, high yield mini-prep method for

isolation of total genomic DNA from fungi. Fungal Genetics Reports 35(1): Article 11.

https://doi.org/10.4148/1941-4765.1531

Mata JL, Halling RE, Petersen RH. 2004. New species and mating system reports in Gymnopus

(Agaricales) from Costa Rica. Fungal Diversity 16: 113-129.

Miller MA, Pfeiffer W, Schwartz T. 2010. Creating the CIPRES Science Gateway for inference of

large phylogenetic trees. 2010 Gateway Computing Environments Workshop (GCE), New

Orleans, Louisiana 2010, 1-8. https://doi.org/10.1109/GCE.2010.5676129

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and seventeen clades of Euagarics. Molecular Phylogenetics and Evolution 23: 357-400.

https://doi.org/10.1016/S1055-7903(02)00027-1

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Petersen R, Hughes K. 2014. New North American species of Gymnopus. North American

Fungi 9(3): 1-22. https://doi.org/10.2509/naf2014.009.003

Saba M, Khalid AN. 2014. New reports of Gymnopus from Pakistan based on ITS sequences.

Mycotaxon 129(1): 63-72. https://doi.org/10.5248/129.63

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145-164.

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valley — 1. Pakistan Journal of Botany 43(3): 1777-1787.

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https://doi.org/10.1016/B978-0-12-372180-8.50042-1

MY COTAXON

January-March 2020—Volume 135, pp. 213-222

https://doi.org/10.5248/135.213

Agaricus, Steccherinum, and Typhula species new for Turkey

HAKAN ISIK

Tokat M. Emin Sarac Anatolian Religious High School, 60030, Tokat, Turkey

CORRESPONDENCE TO: hakanbiyoloji@gmail.com

ABSTRACT—Basidiomycetes collected from Tokat and Yozgat provinces during field trips

during 2012-18—identified as Agaricus dulcidulus (Agaricaceae), Steccherinum oreophilum

(Meruliaceae), and Typhula erythropus (Typhulaceae)—were recorded for the first time for

the Turkish mycota. Short descriptions, illustrations, GPS coordinates, and comparisons with

similar taxa are provided.

Key worps—Agaricales, fungal diversity, morphology, Polyporales, taxonomy

Introduction

Fungi, among the most diverse organisms in the world, have been

estimated to represent approximately 1.5 million species (Hawksworth

2001). Those producing fruitbodies visible to the naked eye are referred

as macrofungi, most of which are assigned to two phyla, Ascomycota and

Basidiomycota. Basidiomycetous macrofungi are an important group

comprising saprophytic, mycorrhizal symbiotic, and parasitic species

that produce fruitbodies in such varied forms as truffles, corals and clubs

(clavarioid fungi), jellies, bird’s nests, puffballs, conks (shelf fungi), and

agarics (gilled fungi) (Andrew & al. 2013; Kinge & al. 2017).

Research on the macrofungal diversity of Turkey, which was first

conducted by Vlaev (1915) in European Turkey, has continued to the

present day. Sesli & Denchev (2014) reported that 1943 basidiomycetous

macrofungal species had been recorded for Turkish mycota up through

the end of 2014. Other scientists have also contributed to basidiomycetous

macrofungal diversity of Turkey and increased this number (Kaya 2015;

214... Isik

Sesli & al. 2015, 2016; Akata & Sesli 2017; Isik & Turkekul 2017, 2018a,b;

Sesli & Vizzini 2017; Turkekul 2017; Uzun & Demirel 2017; Uzun & al. 2017,

2018a,b; Akata & Giirkanli 2018; Akata & al. 2018; Sesli 2018; Uzun & Acar

2018; Uzun & Kaya 2018a,b; Keles 2019a,b).

This study should contribute additional knowledge of basidiomycetous

macrofungal diversity in Turkey.

Materials & methods

Fresh basidiomata were carefully collected from Akdagmadeni (Yozgat) district

and Yaylacik Mountain (Tokat) during 2012-18. The specimens were photographed

in the field, and their macroscopic features and ecological features were noted.

The fresh specimens were brought to the laboratory, assigned collection numbers,

dried thoroughly with an electric heater, and placed into polyethylene bags for

further study. Microscopical structures (basidia, basidiospores, cystidia, hyphae)

were examined under a compound light microscope in mountants prepared with

Melzer’s reagent, distilled water, Congo red, methylene blue solution, and KOH. The

specimens were identified based on their macroscopic, microscopic, and ecological

features by consulting the literature (Moser 1983; Breitenbach & Kranzlin 1986,

1991; Niemela & Saarenoksa 1985; Niemela 1998; Noordeloos & al. 2001; Phillips

1981, 2013; Lzssoe 1998; Sanyal & al. 2016; Loizides 2017; Putzke & Putzke

2017). The examined specimens were deposited in the Fungarium of the Biology

Department, Gaziosmanpasa University, Tokat, Turkey (ISIK).

Taxonomy

Agaricus dulcidulus Schulzer, Icon. Sel. Hymenomyc. Hung. 2: 29 (1874) Fic. 1

PiLtEus 20-70 mm diam., ovate at first, later plane to sometimes

umbonate, depressed when old, white when young with brown squamules,

later lilaceous to vinaceous brown fibrillose with vinaceous brown or

purplish brown disk on white ground, white at margin. LAMELLAE free,

close, pinkish brown at first, later dark brown with whitish edge. STIPE (20-)

30-85(-90) x (2—)4-8(-10) mm, central, cylindrical to clavate, bulbous

at base, white, yellowish towards base, smooth to fibrillose, annulate.

ANNULUS white, membranous, pendant. SPORE PRINT brown.

BASIDIOSPORES (4.2—)5-6(-6.5) x (3.1-)4.5-5 um, ellipsoidal, smooth,

inamyloid. Basrp1A, (19-)22-23(-23.2) x (6-)6.5-6.8(-7) um, clavate,

4-spored. CHEILOCYSTIDIA abundant, pyriform to broadly clavate or

clavate. PLEUROCYSTIDIA absent. CLAMP CONNECTIONS absent.

Fic. 1. Agaricus dulcidulus (ISIK 225): A. basidiomata in situ; B. cheilocystidia and basidium;

C. basidiospores; D. hyphae of pileipellis. Scale bars: A = 3 cm; B, C = 10 um; D = 20 um.

New Agaricus, Steccherinum, and Typhula records for Turkey ... 215

216... Isik

SPECIMEN EXAMINED: TURKEY, YozGat, Akdagmadeni, 39°39’95”'N 35°56'04’E, 1580

m, among needle litter in Pinus sylvestris L. (Pinaceae) forest, 27 May 2012 (ISIK 225).

CoMMENTS—Although some spores in our specimens are larger, the macro-

and micromorphological characters otherwise agree with those cited by

Moser (1983), Noordeloos & al. (2001), Phillips (2013), and Putzke & Putzke

(2017).

Agaricus dulcidulus (Agaricaceae) is morphologically close to A. porphyrizon

P.D. Orton and A. semotus Fr., but A. porphyrizon is separated by its bigger

pileus, longer stipe, and habitat preference, growing mostly in deciduous

forests (Noordeloos & al. 2001, Phillips 2013). Agaricus semotus, which also

grows in deciduous forests or among grasses, differs from A. dulcidulus by

its white to light brown pileus with lilac to reddish-violet fibrillose scales

(especially in the centre), double ring, smaller spores, and purple-brown

spore print (Phillips 1981, Breitenbach & Kranzlin 1991, Lacheva &

Radoukova 2014).

Steccherinum oreophilum Lindsey & Gilb., Mycologia 69(1): 194(1977) Figs 2,3

PiLEus white, effused-reflexed, resupinate, glabrous, stipeless, 6-10 mm

broad, margin sinuate when mature, hymenophore odontoid, concolorous

with basidiocarp surface. ACULEI irregularly flattened and sometimes

subulate, confluent or simple, 0.4-2.3 mm long, creamy whitish. HyPHAL

SYSTEM dimitic; generative hyphae with clamp connections, thin-walled;

skeletal hyphae <4(-4.5) um diam., aseptate, thick-walled.

BASIDIOSPORES (5.5—)6-6.5(-7) x (3-)3.2-3.5(-3.9) jm, ellipsoid,

smooth, inamyloid. SkELETOCysTIDIA long, deep-rooting and tapering

(carrot-like), apically encrusted, (79-)85-120(-123) x (6.2-—)7-7.5(-9.8) um.

BasIDIA (9.6—)11.5-16.5(-17.2) x (4-)4.2-4.8(-5) um, clavate, 4- spored,

with basal clamp connections.

SPECIMEN EXAMINED: TURKEY, Toxat, Yaylacik mountain, 40°31’12”N 36°40’36’E,

1211 m, on fallen and rotten branches of Fagus sp. (Fagaceae), 27 June 2018 (ISIK 769)

CoMMENTS—The specimen we identified as Steccherinum oreophilum

(Meruliaceae) agrees with the literature (basidiospores: 6-6.8 x 3.8 um,

Niemela & Saarenoksa 1985; 5-6.4 x 2.4-3 um, Sanyal & al. 2016; 5.5-7 x

2.5-3.5 um, Loizides 2017).

Morphologically, S. oreophilum is similar to S. ochraceum (Pers. ex

J.E Gmel.) Gray, S. bourdotii Saliba & A. David, and Irpex lacteus (Fr.) Fr.

Matted pileal surface, cylindrical spines, and smaller basidiospores separate

S. ochraceum from S. oreophilum. In addition to the tomentose pileal surfaces

New Agaricus, Steccherinum, and Typhula records for Turkey ... 217

C. basidiospores. Scale bars: A = 1 cm; B, C = 10 um.

of S. bourdotii and I. lacteus, distinguishing characters include slightly smaller

and cylindric spores, an absence of clamp connections, and less strongly

encrusted cystidia for I. lacteus, while the cylindrical spines and subglobose

spores separate S. bourdotii from S. oreophilum (Niemela & Saarenoksa 1985,

Niemela 1998).

218... Isik

Fic. 3. Steccherinum oreophilum (ISIK 769):

D. skeletocystidia; E. hypha with clamp. Scale bars = 10 um.

Typhula erythropus (Pers.) Fr., Observ. Mycol. 2: 297 (1818) FIG. 4

FRUITING Bopy (5—) 10-30 mm, comprising a fertile head and a long stalk.

FERTILE HEAD white to creamy white, cylindrical to clavate, smooth. STALK

0.1-0.3 mm thick and 10-20 mm long, filiform, bended, slender pubescent,

cartilaginous, hollow, deep reddish, lighter red towards the apex, arising from

Fig. 4. Typhula erythropus (ISIK 751): A. basidiomata; B. stipe hypha with clamp;

C. stipe hair; D. basidia; E. basidiospores. Scale bars: A = 1 cm; B, D, E= 10 um; C = 40 um.

New Agaricus, Steccherinum, and Typhula records for Turkey ... 219

220°.2 Isik

a buried blackish sclerotium. Harrs 260-290 x 25-27 um, pointed at apex,

wide at base. FLESH white, transparent especially in fertile head, soft, very

thin. SPORE PRINT white.

BASIDIOSPORES (5-)5.5-9(-9.7) x (2.1-)2.5-3(-3.8) m., elliptical

to oblong, sometimes elongated, smooth. CHEILOCysTIDIA not found.

Cau.LocystTip1A4 hyaline, thick-walled, subulate, septate. Basip1a slightly

clavate, 25-40 x 5-6(-6.7) um, 4-spored, with basal clamp.

SPECIMEN EXAMINED: TURKEY, Tokar, Yaylacik mountain, 40°31’07”N 36°39'24’E,

1298 m, on fallen leaves and dead branches of Alnus sp. (Betulaceae), 4 November 2017

(ISIK 751).

CoMMENTS— The features of our collected specimen are compatible with

published descriptions of Typhula erythropus (Phillips 1981, Lzessoe 1998,

Breitenbach & Kranzlin 1986).

Typhula erythropus (Typhulaceae) is easily recognizable by its white club

and its red-brown stalk. T: erythropus and T. phacorrhiza (Reichard) Fr.

may be confused with each other due to their similar ecological features,

but T. phacorrhiza is distinguishable by its longer and honey-yellow fruiting

body, absence of an evident club, and bigger basidiospores (Breitenbach &

Kranzlin 1986).

Acknowledgments

I would like to thank Dr. Abdullah Kaya, Dr. Ali Keles, and Dr. Shaun Pennycook

for reviewing this article.

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MYCOTAXON

January-March 2020—Volume 135, pp. 223-229

https://doi.org/10.5248/135.223

Tubakia koreana sp. nov.

causing Quercus leaf blight

Hyz YOUNG YUN & YOUNG Ho KIM’

Department of Agricultural Biotechnology,

Research Institute of Agriculture and Life Sciences,

Seoul National University, Seoul 151-921, Korea

“CORRESPONDENCE TO: yhokim@snu.ac.kr

ABSTRACT — A new species, Tubakia koreana (Diaporthales, Ascomycota), isolated in

Korea from blight symptoms on Quercus leaves, is described and illustrated. The species is

morphologically distinct from other Tubakia species and causes a progressive necrotic leaf

blight that differs from leafspot symptoms caused by other species. Phylogenetic analysis

of ITS sequences supports a monophyletic T. koreana clade independent of other Tubakia

species.

Key worps — leaf death, Quercus acutissima, Quercus mongolica, sawtooth oak

Introduction

Oaks (Quercus spp.), which are widely distributed in the northern

hemisphere, are the most common tree species in Korea, occurring

in ~27% of the mountainous area. Four species of Tubakia have been

reported in Korea: T: japonica (Sacc.) B. Sutton (on Castanea crenata,

Quercus acutissima, and Q. x alienoserratoides), T. rubra (T. Yokoy. &

Tubaki) B. Sutton (on Q. serrata), and an undescribed Tubakia sp. (on

Q. rubra; Cho & al. 2004); and Tubakia seoraksanensis H.Y. Yun (on

Q. mongolica; Yun & Rossman 2011). In search of more Tubakia species,

we surveyed various regions in Korea; in the mountain localities of

several South Korean provinces leaf blight symptoms were observed from

September 2009 through November 2014 on four species: Q. acutissima,

224 ... Yun & Kim

Q. x alienoserratoides, Q. mongolica, and Q. serrata. There had been no

previous report of severe foliar blight symptoms of oaks in Korea. The

symptoms were similar to those of bur oak blight in U.S.A., caused by

T. iowensis T.C. Harr. & McNew (Harrington & al. 2012). In this study, the

causal agent of the Korean oak leaf blight was examined to determine its

morphological and molecular genetic characteristics. It was compared with

other Tubakia species for identification of the fungus, which is described

below as T: koreana.

Materials & methods

For fungal isolation, conidiomata on leaves of Quercus spp. with blight

symptoms were detached from the adaxial leaf areas along the necrotic mid- and

lateral veins and cultured on malt extract agar (MEA, 1.5% malt extract, and 2%

agar) at 25°C in the dark. Fungal hyphae growing out of the conidiomata were

harvested and sub-cultured on fresh MEA at 25°C for 14 days, at which time

mycological characteristics were examined. For detailed morphological study,

microscopic structures of the fungus growing on MEA were examined under

a Zeiss Axiophot compound light microscope. Color of cultures was described

using Munsell soil color charts (Munsell 2000). Voucher specimens were

deposited in the Herbarium of the Korean Forest Research Institute, Seoul, South

Korea (HKFRI), and cultures were conserved in Jengeup-si, Jeollabuk-do, South

Korea (KCTC).

For DNA extraction, PCR, and DNA sequencing, fungal cultures grown on

MYEA (2% malt extract, 0.2% yeast extract, and 1.5% agar) at room temperature

for 14 days were used. Methods for DNA extraction, PCR (including primers),

and sequencing of ITS rDNA followed Yun & Rossman (2011). ITS sequences

were deposited in GenBank. The ITS sequences for our isolates along with other

known Tubakia species were preliminarily aligned using Clustal X version 2.1

under default settings (Thompson & al. 1997). The alignment was edited using

Mesquite 3.03 beta 2 (Maddison & Maddison 2015). Greeneria uvicola was used

as outgroup for Tubakia since this species was among the closest to genus Tubakia

based on BLAST search at the National Center for Biotechnology information

(NCBI, https://www.ncbi.nlm.nih.gov). Maximum parsimony analysis (MP) was

conducted using PAUP version 4.0b10 (Swofford 2002), using a heuristic search

with the starting tree obtained via stepwise addition with random addition of

1000 replicates, tree—bisection—reconnection (TBR) as the branch swapping

algorithm, and MULTREES off. All characters were unordered, whereas equal

weights and gaps were treated as a fifth character. Stability of the clades (bootstrap)

was assessed with 1000 replicates using the same MP settings. Bootstrap values

greater than or equal to 70% were considered significant (Hillis & Bull 1993).

Pathogenicity tests of Tubakia koreana on Quercus acutissima were performed

using inoculation protocols modified from those of Harrington & al. (2012).

Tubakia koreana sp. nov. (Korea) ... 225

Fic. 1. Tubakia koreana (HKFRI4072) on Quercus acutissima:

Sign of the blight, black crustose conidiomata. Scale bars: A = 1 mm; B = 0.5 mm.

Taxonomy

Tubakia koreana H.Y. Yun, sp. nov. PLATEs 1. 2

MB814540

Differs from Tubakia dryina by its brown colony color and its bigger spore size.

Type: South Korea, Gyeonggi: Mt. Geom-dan, Hanam-si, Sinjang-dong, living tree of

Q. mongolica Fisch. ex Ledeb., 18 July 2012, HY Yun, HY946 (Holotype, HKFRI 4073;

ex-type culture, KCTC 46072; GenBank KP886837).

EryMo.oecy: koreana refers to the country where the species was collected.

The fungus was consistently isolated from leaves with blight symptoms

produced in nature and by artificial inoculation. Small crustose conidiomata

with radiate scutella formed on the necrotic tissues of mostly the upper

side or occasionally on both sides of leaf surfaces. The fungal colonies fully

grown on malt extract agar (MEA) at 25°C were felt-like and very pale

brown to brown in color with concentric rings of dense aerial mycelium

and a scalloped margin. Scutella that formed on leaf surfaces were 60-180 x

55-120 um, membranous, composed of thick-walled, pale brown to brown,

septate hyphae radiating, typically bifurcating up to three times from

a central disc that consisted of a single hyaline cell towards the margin,

acute and cornuted at the margin that was fringed and unattached to the

substrate. Conidia were 8.5-14.5 x 6-10 um, blastic, subglobose, broadly

226 ... Yun & Kim

Fic. 2. Tubakia koreana (HKFRI4073). A. Colony grown on malt extract agar at 25°C for 14

days; B. Scutella; C. Scutella and conidia (arrows); D. Conidia. Scale bars: B = 50 um; C = 20 um;

D=10um.

ellipsoid to ellipsoid, and hyaline. They were turning pale yellowish brown,

and the walls were smooth. Microconidium was not observed.

ADDITIONAL SPECIMENS EXAMINED: SOUTH KOREA: CHUNGBUK, Buk-myeon,

Inje-gun, living tree of Quercus mongolica, 5 July 2012, HY Yun, HY882 (HKFRI4083);

Jeungpyeong-gun, Jeungpyeong-eup, Miam-ri, living tree of Q. acutissima Carruth.,

18 July 2012, HY Yun, HY1095 (HKFRI4090, KCTC46103), HY1096 (HKFRI4091,

Tubakia koreana sp. nov. (Korea) ... 227

KCTC46104), HY 1098 (HKFRI4081, KCTC46105); living tree of Q. serrata Murray,

18 July 2012, HY Yun, HY1097 (HKFRI4082). GANGwon, Wonju-si, Halla Univ.,

Heungeop-myeon, Heung-eop-ri, living tree of Q. mongolica, 6 October 2012, HY

Yun, HY134 (HKFRI4086); Sokcho-si, Mt. Seorak, Seorak-dong, living tree of

Q. x alienoserratoides T.B. Lee, 28 October 2011, HY Yun, HY719 (HKFRI4071,

KCTC46044, GenBank KP886836); Seoraksan National Park, Seorak-dong, living

tree of Q. acutissima, 31 Aug. 2009, HY Yun, US227 (HKFRI4085); GYEONGGI,

Namyangju-si, Mt. Un-gil, Songchon-ri, Joan-myeon, on living tree of Q. mongolica,

27 July 2012, HY Yun, HY989V (HKFRI4089, KCTC46092); JEonBUK, Muju-gun,

Mt. Deog-yu, Samgong-ri, Seolcheon-myeon, living tree of Q. mongolica, 10 July

2012, HY Yun, HY190 (HKFRI4070; KCTC46031; GenBank KP886834); JEONNAM,

Gwangyang-si, Ongnyong-myeon, living tree of Q. acutissima, 24 February 2012,

HY Yun, HY107 (HKFRI4084); SEOUL: Seoul, Mt. Kwan-ak, Gwanak-gu, Daehak-

dong, living tree of Quercus acutissima, 23 June 2012, HY Yun, HY816 (HKFRI4072,

KCTC46051, GenBank KP886835).

Discussion

Initial symptoms were small pale-brownish and dehydrated areas

on leaves that spread through leaf veins and blades until the entire leaf

or large areas were blighted with necrotic mid and lateral veins. Those

symptoms sometimes preceded premature defoliation. The mycological

characteristics of the present pathogen were distinct from those of other

Tubakia species; i.e., colony color (very pale brown to brown) differing

from T: dryina (Sacc.), B. Sutton (light gray), T: iowensis (light and dark

gray), and T. seoraksanensis (whitish to pale yellow); conidial size smaller

than T. seoraksanensis; and an absence of microconidia unlike their

occasional presence in T’ iowensis (Harrington & al. 2012, Yun & Rossman

2011). Also, ITS rDNA sequence analysis of the four isolates from diseased

oak trees showed they had identical sequences. In addition, phylogenetic

analysis placed this fungus in a monophyletic lineage independent of other

Tubakia species (PLATE 3).

Maximum likelihood tree comparison revealed it is a unique

monophyletic clade, distinct from other Tubakia species. Also, the

observed symptom of severe necrosis is different than the symptom of

leaf spots observed in T: japonica. The Korean Tubakia species has much

smaller conidia than T: japonica. These observations all suggest that the

causal agent of oak leaf blight in the present studies is a new Tubakia species

with different mycological and genetic characteristics from other Tubakia

species distributed worldwide. The species epithet is named koreana to

indicate a pathogen that is prominent throughout the mountains of South

Korea. The disease caused by this fungus should be named oak leaf blight,

228 ... Yun & Kim

Tubakia koreana KP886834

oN Tubakia koreana KP886835

Tubakia koreana KP886836

Tubakia koreana KP886837 [T]

Tubakia iowensis JF704196

Tubakia japonica CBS191.71

Tubakia seoraksanensis HM991734

Tubakia seoraksanensis HM991737

Tubakia seoraksanensis HM991735

100

Dicarpella sp. HM855225

ie Dicarpella sp. HM855226

Tubakia dryina CBS114919

Tubakia dryina AY853240

Tubakia rubra CBS192.71

Tubakia castanopsidis CBS124732

Tubakia subglobosa CBS193.71

100

Tubakia iowensis JF129017

Tubakia iowensis JF704194

Greeneria uvicola JN547718G

— 5changes

Fic. 3. Maximum likelihood tree produced from selected ITS rDNA sequences, showing the

relations of Tubakia koreana isolates with other Tubakia species, ['T]: holotype. The tree was

obtained using PhyML 3.0. Numbers above the branch points are bootstrap values obtained

from 1000 replicates.

Tubakia koreana sp. nov. (Korea) ... 229

rather than oak blight, since twig and branch dieback symptoms were

rarely found, as in the bur oak blight reported in the U.S.A. (Harrington &

al. 2012). In pathogenicity tests, leaf blight symptoms similar to naturally

occurring symptoms developed on all oak trees wound-inoculated (mid

vein) with conidial suspensions of T: koreana 15 days after inoculation;

the most severe symptoms were observed on Q. acutissima. Oak trees are

common tree species in Korea. Therefore, this disease could be a potential

threat to the stable ecological succession of Korean forest vegetation.

Acknowledgments

We would like to express our gratitude to our reviewers, Yuuri Hirooka and

Ono Yoshitaka, for their expertise and thank the Korea Forest Service for financially

supporting our research (KFS 500-20130153).

Literature cited

Cho WD, Shin HD (eds). 2004. List of plant diseases in Korea, fourth edition. Korean Society

of Plant Pathology, Seoul, Korea.

Harrington TC, McNew D, Yun HY. 2012. Bur oak blight, a new disease on Quercus

macrocarpa caused by Tubakia iowensis sp. nov. Mycologia 104(1): 79-92.

https://doi.org/10.5248/115.369

Hillis DM, Bull JJ. 1993. An empirical test of bootstrapping as a method for assessing

confidence in phylogenetic analysis. Systematic Biology 42: 182-192.

https://doi.org/10.1093/sysbio/42.2.182

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Version 3.03 http://mesquiteproject.org.

Munsell AH. 2000. Munsell color charts: year 2000 revised washable edition. GretagMacbeth,

New Windsor, NY.

Swofford DL. 2002. PAUP*: phylogenetic analysis using parsimony (*and other methods).

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interface; flexible strategies for multiple sequence alignment aided by quality analysis tools.

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Yun HY, Rossman AY. 2011. Tubakia seoraksanensis, a new species from Korea. Mycotaxon 115:

369-373. https://doi.org/10.3852/11-112

MYCOTAXON

January-March 2020—Volume 135, p. 231

https://doi.org/10.5248/135.231

Regional annotated mycobiota new to the Mycotaxon website

SUMMARY—In February, MycoTaxon added the 136th annotated species distribution

list to our previously posted fungae. The 23-page “Macrofungi from the Hebron and

Jerusalem Hills of Palestine” by Maximus Thaler, Aysha Al-Wahsh, Alea Meuser, Alyssa

Rooks, and Mazin Qumsiyeh may be downloaded (for no charge) from our website via

http://www.mycotaxon.com/mycobiota/index.html

MID-EAST

Palestine

Maximus THALER, AYSHA AL-WAHSH, ALEA MEUSER, ALyssA ROOKS,

MAZIN QumsIYEH. Macrofungi from the Hebron and Jerusalem Hills

of Palestine. 23 p.

ABSTRACT— This study is based on specimens of macrofungi collected from

biodiversity hotspots in the southern West Bank of occupied Palestine during a

four-week survey period in December and January 2018-19. We identified 27

macrofungi species representing 39 genera from six field sites. Samples were

collected, photographed, and archived in the Palestine Museum of Natural

History herbarium. Species descriptions, field site details, and a cladogram

of the observed fungi are provided. The importance of citizen science and

accessible taxonomic inquiry is also discussed.

KEY worps—diversity, mycota, taxonomy

MYCOTAXON

January-March 2020—Volume 135, p. 233

https://doi.org/10.5248/135.233

Regional annotated mycobiota new to the Mycotaxon website

ABSTRACT—MycotTaxon is pleased to add a new annotated species distribution

list to our 136 previously posted free-access fungae. The 29-page “Checklist of

Bolivian Agaricales. 2: Species with white or pale spore prints” by Melgarejo-Estrada,

Rocabado, Suarez, Maillard, and Lechner may be downloaded from our website via

http://www.mycotaxon.com/mycobiota/index.html

SOUTH AMERICA

Bolivia

ELIZABETH MELGAREJO-ESTRADA, DIANA ROCABADO, MARIA EUGENIA

SUAREZ, OSWALDO MAILLARD, BERNARDO ERNESTO LECHNER.

Checklist of Bolivian Agaricales. 2: Species with white or pale spore

prints. 29 p.

ABSTRACT—We _ provide a literature-based checklist of Agaricales

reported from Bolivia. In this second contribution, 264 species

belonging to 55 genera and 15 families (Agaricaceae, Clavariaceae,

Cyphellaceae, | Hygrophoraceae, Hymenogastraceae, | Marasmiaceae,

Mycenaceae, Omphalotaceae, Physalacriaceae, Pleurotaceae, Pterulaceae,

Schizophyllaceae, Tricholomataceae, and Typhulaceae) are listed. Of these,

Marasmiaceae, Omphalotaceae, and Hygrophoraceae were the most species-

abundant families. Some new local distribution records are documented in

this checklist. The lichens Cora and Dictyonema are also included in the

present checklist.

Key worps—Basidiomycota, distribution, diversity, fungi, Neotropics,

South America