Iv ... MYCOTAXON 135(3)

MYCOTAXON

VOLUME ONE HUNDRED THIRTY-FIVE (3) — TABLE OF CONTENTS

Nomenclatural novelties & typifications .......... 0000 c cece cece ee ees vii

ECVIEW CNS Wag ad te. Maat tA nes of Poise Sood ry rere NSS sdk yet By EN as got Bde area ix

COLigeHda cnn oan moat abet, Hie AW RR ee Ne eras id dee Ai ad x

BE POU Pde TE ILO ER Stok Path Pes OM alate te Plas ett ol Ain les alan ddahy LA xi

ZOZOSUDTTISSIOM PNOCCH UTE S.n:. weal Rasier a Rime N,. crete okie cee. ea xiii

TAXONOMY

Lepiota punaensis sp. nov. from Hawai‘i Island,

and a discussion of L. elaiophylla

JEFFERY K. STALLMAN, DON E. HEMMES,

NICOLE A. Hynson, MICHAEL H. SHINTAKU 471

Notes on rust fungi in China 8. Pucciniastrum tiliae life cycle

and new host plants inferred from phylogenetic evidence

JING-XIN JI, ZHUANG Li, Yu L1, MAKOTO KAKISHIMA 490

Guayaquilia gen. nov., typified by Idriella cubensis

FREDDY MAGDAMA, DAYNET SOSA,

FERNANDO ESPINOZA, LIZETTE SERRANO, SIMON PEREZ-MARTINEZ,

ELAINE MALOosso, MARGARITA HERNANDEZ-RESTREPO,

RAFAEL F, CASTANEDA-Ru1z 501

Distoseptispora longispora sp. nov.

from freshwater habitats in China Hal-YAN SONG,

ALY FaraG EL SHEIKHA, ZHI-JUN ZHAI, JIAN-PING ZHOU,

Mrinc-Hutr CHEN, GuANG-Hua Huo, X1-GEN Huane, D1An-MinG Hu 513

Notes on rust fungi in China 9. Puccinia miscanthi life cycle

and morphology confirmed by inoculation

JING-XIN JI, ZHUANG LI, Yu L1, MAKOTO KAKISHIMA 525

Biodiversity of heat-resistant ascomycetes

from semi-arid soils in Argentina STELLA MARIS ROMERO,

ANDREA IRENE ROMERO, ALBERTO MIGUEL STCHIGEL,

ERNESTO RODRIGUEZ ANDRADE, VIVIANA ANDREA BARRERA,

JOSE FRANCISCO CANO, RICARDO COMERIO 535

Cortinarius rapaceoides, a new record for Turkey

MERYEM SENAY SENGUL DEMIRAK & HAKAN IsIK 559

New records of Toninia from China Concconc M1Ao, MEUIE SUN,

XIAO ZHANG, ZHAOJIE REN, LING Hu 569

JULY-SEPTEMBER 2020 ... V

Serendipita sacchari sp. nov. from a sugarcane rhizosphere

in southern China LinG X1g£, YAN- YAN LONG,

YAN ZHANG, YAN-LU CHEN, WEN-LONG ZHANG 579

Five Nolanea spp. nov. from Brazil FERNANDA KARSTEDT,

SARAH E. BERGEMANN, MARINA CAPELARI 589

First record of Trappea darkeri from Turkey

YASIN UzuN, OSMAN BERBER, ABDULLAH Kaya 613

Mesocorynespora sinensis gen. & sp. nov.

from southern China ZHAO-HUuAN Xu,

KAI ZHANG, YOU-QIANG Luo, XIU-GUO ZHANG,

RAFAEL FE. CASTANEDA-RUiZ, JIAN Ma 617

Corynesporopsis hainanensis sp. nov.,

a bambusicolous fungus from southern China ZHAO-HuaNn Xu,

XuU-GEN SHI, WEI-GANG KUANG, XIU-GUO ZHANG,

RAFAEL FE. CASTANEDA-RUiZ, JIAN Ma 623

Hymenochaete longisterigmata sp. nov. from India

NAVPREET Kaur, AVNEET PAL SINGH, GURPAUL SINGH DHINGRA 631

Entoloma conferendum, Hygrocybe coccineocrenata,

and Hypholoma ericaeum new to Montenegro ILINKA CETKOVIG,

ZDENKO TKALCEC, SNEZANA DRAGICEVIC, ANTUN ALEGRO,

VEDRAN SEGOTA, MARGITA JADAN, NEVEN MATOCEC,

IvANA KuSAN, ZELJKO ZGRABLIC, ARMIN ME81¢ 637

First Pakistani report of Erysiphe betae

on the invasive weed Chenopodium ambrosioides

AYESHA ANWAR, NAJAM UL SEHAR AFSHAN,

AAMNA ISHAQ, MARIA RIAZ, ABDUL NASIR KHALID, SIRAJ UDDIN 649

Crustose lichens new to India Rupyyoti GoGo, SILJo JOSEPH,

Manas PRATIM CHOUDHURY, SANJEEVA NAYAKA, FARISHTA YASMIN 657

Chromelosporium re-evaluated,

with Chromelosporiopsis gen. nov. and Geohypha stat. nov.

GREGOIRE L. HENNEBERT 665

VI ... MYCOTAXON 135(3)

PUBLICATION DATE FOR VOLUME ONE HUNDRED THIRTY-FIVE (2)

MYCOTAXON for APRIL-JUNE 2020 (I-XIV + 235-471)

was issued on July 13, 2020

JULY-SEPTEMBER 2020 ...

NOMENCLATURAL NOVELTIES AND TYPIFICATIONS

PROPOSED IN MYCOTAXON 135(3)

Botrytis isabellina Preuss (lectotypified)

[MBT 392406], p. 671

Chromelosporium herbicola (Ellis & Dearn.) Hennebert

[MB 833990], p.673

Chromelosporiopsis Hennebert

[MB 835622], p. 688

Chromelosporiopsis carnea (Schumach.) Hennebert

[MB 835623], p. 689

Chromelosporiopsis coerulescens (Bonord.) Hennebert

[MB 835625], p. 695

Corynesporopsis hainanensis Z.H. Xu, Jian Ma, X.G. Zhang & R.F. Castaneda

[MB 836568], p. 624

Distoseptispora longispora H.Y. Song & D.M. Hu

[MB 826916], p. 516

Geohypha (Fr.) Hennebert

[MB 835628], p. 708

Geohypha terrestris (Fr.) Hennebert

[MB 835629], p. 709

Guayaquilia R.F. Castafieda, Magdama, D. Sosa & Hern.-Restr.

[MB 831849], p. 506

Guayaquilia cubensis (R.F. Castaheda & G.R.W. Arnold) R.F. Castaneda,

Magdama, D. Sosa & Hern.Restr.

[MB 831851], p. 507

Hymenochaete longisterigmata Nav. Kaur, Avn.P. Singh & Dhingra

[MB 832678], p. 632

Lepiota punaensis Stallman

[MB 832104], p. 477

Mesocorynespora Jian Ma, X.G. Zhang & R.E. Castaneda

[MB 836511], p. 618

Mesocorynespora sinensis Jian Ma, X.G. Zhang & R.E Castaneda

[MB 836512], p. 618

Nolanea albertinae Karstedt & Capelari

[MB 826784], p. 596

Nolanea atropapillata Karstedt & Capelari

[MB 805603], p. 600

Nolanea pallidosalmonea Karstedt & Capelari

[MB 805604], p. 602

VII

Vill ... MYCOTAXON 135(3)

Nolanea parvispora Karstedt & Capelari

[MB 805605], p. 603

Nolanea tricholomatoidea Karstedt & Capelari

[MB 805607], p. 606

Polyactis carnea Ehrenb. (lectotypified)

[MBT 392408], p. 689

Puccinia miscanthi Miura (neotypified)

[MBT 388544], p. 531

Serendipita sacchari L. Xie, Y.Y. Long & Y.L. Chen

[MB 836761], p. 582

Trichoderma laeve Pers. (lectotypified)

[MBT 392413], p. 709

JULY-SEPTEMBER 2020 ...

REVIEWERS — VOLUME ONE HUNDRED THIRTY-FIVE (3)

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.

Vladimir Antonin

Flavia Rodrigues Barbosa

Timothy J. Baroni

Maria Virginia Bianchinotti

Uwe Braun

Rafael E Castafieda-Ruiz

Cvetomir M. Denchev

Edit Farkas

Patricia Oliveira Fiuza

André Fraiture

Shouyu Guo

Nils Hallenberg

David L. Hawksworth

R.G.U. Jayalal

Abdullah Kaya

Ali Keles

De-Wei Li

Xiao- Yong Liu

Jian Ma

Eric H.C. McKenzie

Andrew N. Miller

A.R. Niazi

Lorelei L. Norvell

Shaun R. Pennycook

Brian A Perry

Keith Seifert

B.M. Sharma

Silvana Santos da Silva

G.P. Sinha

Ibrahim Tirkekul

Yusuf Uzun

Alfredo Vizzini

Felipe Wartchow

Jiwen Xia

X ... MYCOTAXON 135(3)

CORRIGENDA FOR MYCOTAXON 135-(3)

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

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

been processed.

p. 495, line 10 FOR: A-D READ: 5A-D

p. 496, line 13 FOR: Helongjiang READ: Heilongjiang

p. 531, line 8 FOR: Helongjiang READ: Heilongjiang

p#533,-lines3 FoR: Wuhan READ: Wuchang

JULY-SEPTEMBER 2020 ... XI

FROM THE EDITOR-IN-CHIEF

WHO ‘AUTHORS’ SCIENCE?—Fungal research is now so specialized that the single-

authored paper has become a rare beast indeed. Modern mycological systematics

now rely on contributions from masters from several different disciplines: alpha

taxonomists and collectors well versed in morphological identification, ecologists

able to link environmental requirements and taxa, physiologists and “beta”

taxonomists who craft diagnostic matrices from colonies in Petri dishes, “gene

jockeys” who reliably deliver pristine sequences from targeted DNA regions, and

evolutionary biologists wielding Bayesian analysis and Markov chain Monte Carlo

methods with morphological analytics to generate elegant and reliable phylogenies.

To these may be added those who summarize, analyze, and wrap everything up

in a research paper—the authors. In multi-authored papers, there is usually one

individual who “drives” the paper, although sometimes two authors may be said

to “contribute equally” The first-named (also called “senior”) authors should have

participated in the research since its inception so that they understand all aspects

well enough to create a clear and comprehensible presentation of their research.

Unfortunately lists begin with one name, and often the author with the best English

writing abilities ascends to first position by default, irrespective of the number of

scientific contributions made during research. Alternately, the most fluent English

speakers who are designated corresponding authors may appear anywhere in the

lists. There is also the trend toward ‘anchoring’ authorship by placing the Lab

supervisor in last place; here we devoutly hope that the lab head has read the paper

in its entirety and corrected mistakes well before journal submission.

Your editors recognize the problems encountered by contributing scientists in

deciding which contributors to include as authors and which to recognize in the

Acknowledgments. We definitely expect all authors to gather frequently to discuss

the research while in progress, when being written, and when being prepared for

submission. These discussions are true teaching moments, and authors who have

eliminated mistakes during these stringent vetting processes will have much better

success in publishing their science.

MycoTaxon ART—Your editors are unusually fortunate to be able to present on

our cover such beautiful art created by authors to illustrate their own research. The

illustrations featured on our covers are selected from papers proposing taxa new to

science. However, there are some truly spectacular plates in this issue accompanying

previously named taxa that “escaped” cover selection. Part of the pleasure of opening

a new Mycotaxon is leafing through the beautiful fungal portraits lurking inside.

MYCOTAXON 135(3) contains 18 papers by 92 authors (representing 17 countries) as

revised by 32 expert reviewers and the editors.

The 2020 July-September MycoTAxon proposes FOUR new genera (Chromelo-

sporiopsis, Geohypha, Guayaquilia, and Mesocorynespora) and TEN species new

XII ... MYCOTAXON 135(3)

to science representing Corynesporopsis, Distoseptispora, Mesocorynespora,

and Serendipita from Cuina; Hymenochaete from INp1a; Lepiota from Hawai‘i

(U.S.A.); and Nolanea from BraAziL. We also offer FIVE new combinations in

Chromelosporium, Chromelosporiopsis, Geohypha, and Guayaquilia and FouR newly

registered typifications for Botrytis, Polyactis, Puccinia, and Trichoderma.

New species range extensions are reported for [ascomycetes] five heat resistant

species in an elegant culture study from ARGENTINA; [basidiomycetes] Cortinarius

& Trappea in TuRKEY and Entoloma, Hygrocybe, and Hypholoma in MONTENEGRO;

and [lichens] Bacidia, Malmidea, Porina, and Pyrenula in Inv1a and Toninia in

CHINA. New hosts are cited for Erysiphe in PAKISTAN and Pucciniastrum in China.

We also offer keys to species of Lepiota in Hawati‘l and Toninia s.l. in CuH1Na along

with worldwide keys to Corynesporopsis, Distoseptispora, and Mesocorynespora.

The July-September issue now offers more molecularly supported taxonomic

studies: nine new species representing Distoseptispora, Guayaquilia, Lepiota,

Nolanea, and Serendipita), one Cortinarius range extension, an Erysiphe host

association, and a full life cycle of the rust Pucciniastrum tiliae are all supported by

phylogenetic analyses, while another life cycle is established for Puccinia miscanthi

via culture studies.

Alpha taxonomy is not dead, particularly when supported by a lifetime of close

microscopical scrutiny and systematic research. The issue ends with “my final

paper” by our co-founding editor, GREGOIRE HENNEBERT, who presents a masterful

synopsis of a difficult (and often overlooked) group of fungi and concluding with a

warm memorial of Agriculture Canada’s irreplaceable Stan Hughes, who died last

year at the age of 102.

Warm regards,

Lorelei L. Norvell (Editor-in-Chief)

9 October 2020

JULY-SEPTEMBER 2020... XIII

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MYCOTAXON

July-September 2020—Volume 135, pp. 471-489

https://doi.org/10.5248/135.471

Lepiota punaensis sp. nov. from Hawai'i Island,

and a discussion of L. elaiophylla

JEFFERY K. STALLMAN’, DON E. HEMMEs?’,

NICOLE A. HyNSON?, MICHAEL H. SHINTAKU*

™ Department of Tropical Conservation Biology & Environmental Science Graduate Program,

University of Hawai‘i Hilo, Hilo, Hawai‘i 96720 USA

? Department of Biology, University of Hawai‘i Hilo, Hilo, Hawai‘i 96720 USA

° Pacific Biosciences Research Center, University of Hawai‘i Manoa,

Honolulu, HI 96822 USA

* College of Agriculture, Forestry, and Natural Resource Management

and Department of Tropical Conservation Biology & Environmental Science Graduate Program,

University of Hawai‘i Hilo, Hilo, Hawai‘i 96720 USA

*CORRESPONDENCE TO: jstall@hawaii.edu

ABSTRACT—A new species, Lepiota punaensis, in L. subsect. Helveolinae is described from

the Puna District of Hawai‘i Island. A species closely resembling L. elaiophylla in morphology

but genetically distant from the species paratype is also discussed. Both taxa and additional

Hawaiian collections are analyzed in an nrITS phylogeny, and a key to Lepiota on Hawai‘i

Island is given.

KEY worps—Agaricaceae, Casuarina equisetifolia, cryptic species

Introduction

Lepiota (Pers.) Gray is a large genus of more than 400 species (Kirk & al.

2008) with widespread distribution that continues to see new species described

regularly from temperate (Caballero & al. 2015), subtropical (Qasim & al.

2015), and tropical (Sysouphanthong & al. 2016) environments. Exceptions

occur, but in general the genus is characterized macroscopically by white to

yellow-spored agarics with free gills, evidence of partial and universal veils,

but lacking a volva. Microscopically the lamellar trama are regular, and

472 ... Stallman & al.

basidiospores are thin-walled, without a germ pore, generally dextrinoid,

and not metachromatic in Cresyl Blue. Lepiota has historically been split into

sections based on morphology (e.g., Bon 1993), and more recently, into clades

based on molecular data (Vellinga 2003). Lepiota subsect. Helveolinae Bon &

Boiffard [in L. sect. Ovisporae (J.E. Lange) Kuhner], which contains the two

species closely examined in this manuscript, is characterized morphologically

by ellipsoid to oblong spores with a pileus covering composed of a trichoderm

of predominantly long cylindrical elements and has been consistently recovered

as monophyletic in molecular analyses of Lepiota and its infrageneric sections

(Vellinga 2003; Qasim & al. 2015, 2016; Sysouphanthong & al. 2012, 2016;

Liang & al. 2018; but see Liang & al. 2011). This clade contains species such as

Lepiota subincarnata J.E. Lange and L. brunneoincarnata Chodat & C. Martin,

which are known to contain amatoxins (Benjamin 1995).

Our project documenting the lepiotaceous fungi from Hawai‘i Island

reports a new taxon from L. subsect. Helveolinae, Lepiota punaensis, and a new

record for Hawai‘i, Lepiota cf. elaiophylla. In addition, Lepiota aspera (Pers.)

Quél., Lepiota besseyi H.V. Sm. & N.S. Weber, Lepiota pseudorubella Gubitz,

Lepiota rubrobrunnea Gubitz, and a poorly known taxon with an epithelial

pileus covering (MK412575) are confirmed as occurring on Hawai‘i Island. All

species are found in wet, lowland environments associated with alien vegetation

and are therefore presumed to be introduced.

Materials & methods

Collection and morphologic description

Basidiome collecting occurred from September 2016 to January 2019 on Hawai‘i

Island. Photographs were taken in the field with a Canon Rebel T4i camera and

have not been altered (PLATES 2a; 3a). Morphological details were recorded at the

time of collection. Descriptions of Hawaiian habitats and vegetation zones are from

Gagné & Cuddihy (1999). Macroscopic descriptions are based on terminology from

Vellinga (2001) and color descriptions are from Kornerup & Wanscher (1978).

After collection, specimens were desiccated before microscopic and molecular

analysis, and are deposited at the Joseph F. Rock Herbarium, University of Hawaii

at Manoa, Honolulu, HI, USA (HAW).

Measurement of microscopic structures was performed on material rehydrated

in 10% ammonia (pileus and stipe covering, spores), or 10% ammonia with

1% Congo Red (all other structures). Melzer’s reagent, distilled water, and 1%

Cresyl Blue in distilled water were also used to observe natural colors and color-

changing reactions. Mature spores were measured in side view (excluding the

hilar appendage) using the program Piximétre version 5.9 (http://ach.log.free.

fr/Piximetre/). Measurements represent values falling within a 95% confidence

Lepiota punaensis sp. nov. (Hawai'i USA) ... 473

interval; the annotation [30, 4, 3] indicates that thirty spores from four specimens

in three collections were measured. Micrographs for import to Piximétre were

captured using an Olympus XM10 microscope camera attachment on an Olympus

BX53 microscope with the software cell-sens Dimension version 1.5.

Molecular analysis

The primers ITSIF and ITS4 were used to amplify the nuclear ribosomal

internal transcribed spacer region (nrITS; Schoch & al. 2012) via the polymerase

chain reaction from dried fungal tissue of eleven specimens from Hawai‘i Island.

Amplicons were Sanger sequenced in both forward and reverse directions using

the primers ITSIF and ITS4, and sequences were trimmed, edited, and forward

and reverse reads aligned into consensus sequences in Geneious v. 9.1.8 (Kearse &

al. 2012). Sequences generated were further investigated using the BLAST tool in

GenBank (NCBI 2015), and the top sequences by similarity to Hawaiian taxa, as well

TABLE 1. Sequences newly generated and downloaded from GenBank for phylogenetic

analysis.

TAXON (COLLECTION #) GENBANK # TAXON (COLLECTION #) GENBANK #

C. hetieri AY176459 L. luteophylla AY176475

C. pulverulenta AF391035 L. magnispora AF391005

C. seminuda JF907983 L. pilodes AY 176476

C. sp. (Hawai‘i) (JKS 143) MK412600 L. punaensis (JKS 64) MK412572

C. sp. (Hawai‘i) (JKS 140) MK412604 L. punaensis (JKS 86) MK412577

Ch. molybdites (JKS 96) MK412591 L. punaensis (JKS 87) MK412578

L. andegavensis KP004931 L. rubrobrunnea (JKS 118) MK412583

L. apatelia AY176462 L. rhodophylla AY176480

L. aspera KP843884 L. sp. (O‘ahu) (SS 1) MK412617

L. aspera MK412598 L. sp. (Australia) KP012693

L. brunneoincarnata FJ998395 L. sp. (Australia) KP012854

L. castanea AY176463 L. sp. (Hawai‘i) (JKS 74) MK412575

L. castaneidisca AF391061 L. sp. (Kaua‘i) AY176402

L. cristata U85327 L. sp. (Thailand) HQ647293

L. cf. elaiophylla MK412581 L. sp. (Thailand) JN224825

L. cf. elaiophylla (JKS 71) MK412589 L. sp. (Thailand) JN224826

L. cf. elaiophylla (JKS 77) MK412588 L. sp. (Thailand) JN224828

L. elaiophylla MH979467 L. subincarnata AY176491

L. elaiophylla AF391024 L. thiersii AY176492

L. erminea AY176470 L. tomentella EF080868

L. farinolens AY176368 L. vellingana HE974764

L. felina U85330 L. xanthophylla AY176405

L. helveola MH979466 M. haematospermum KF953545

L. himalayensis HE614898 Uncultured fungus (Australia) FJ528742

Sequences with collection numbers were newly generated; Hawaiian sequences are in bold.

C. = Cystolepiota, Ch. = Chlorophyllum, L. = Lepiota, and M. = Melanophyllum.

474. ... Stallman & al.

as representatives from each of the clades of Lepiota identified in Vellinga (2003)

were included in our phylogenetic analyses (TaBLE 1). Pairwise genetic distances

(TABLE 2) were calculated in Geneious by comparing aligned, overlapping regions,

excluding gaps and ambiguous positions. Lepiota besseyi was excluded from our

phylogenetic analysis and key to Lepiota because it falls outside of Lepiota in the

Leucocoprinus Pat. and Leucoagaricus Locq. ex Singer clade in Agaricaceae Chevall.,

although a formal transfer has not been made (Vellinga & al. 2011).

TABLE 2. Pairwise genetic distance comparison between Lepiota cf. elaiophylla and

closest Lepiota relatives in GenBank.

elaiophylla elaiophylla cf. elaiophylla cf. elaiophylla | cf. elaiophylla

(Canada) (Netherlands) (Hawai‘i) (Hawai‘i) (Hawai‘i)

MH979467 AF391024 MK412581 MK412581 MK412588

elaiophylla

(Netherlands) 92.9%

AF391024

cf. elaiophylla

(Hawai‘i) 99.9% 92.9%

MK412581

cf. elaiophylla

(Hawai ‘i) 99.9% 92.9% 99.9%

MK412589

cf. elaiophylla

(Hawai ‘i) 99.9% 92.9% 99.9% 99.9%

MK412581

L. sp.

(Australia) 98.3% 93.2% 98.3% 98.3% 98.3%

KP012854

Sequences were aligned in Geneious with the default settings of the MAFFT

(Katoh & Standley 2013) plugin, then imported to Molecular Evolutionary Genetics

Analysis (MEGA) version X (Kumar & al. 2018). Based on the “Find Best DNA

Model” tool in MEGA, Maximum Likelihood (ML) phylogenetic trees were created

with all sites with gaps and/or missing data eliminated resulting in 465 total

positions in the final dataset using the following settings: General Time Reversible

(GTR) model (Nei & Kumar 2000) including gamma distributed (G) and invariant

sites (I), moderate branch swap filter, and Subtree-Pruning-Regrafting (Extensive,

level 5) for the ML heuristic method. The initial tree was built using the default

neighbor-joining/bioneighbor-joining method, and the phylogeny was tested with

1000 bootstrap replicates.

The alignment was further analyzed using the Geneious plugin MrBayes

(Huelsenbeck & Ronquist 2001). The tree was constructed with a GTR model

including G and I sites with default settings (four chains for 1,100,000 generations,

sampled every 200 generations with the first 100,000 generations discarded as burn-

in). Tree images were edited in Inkscape version 3 (https://inkscape.org/).

Lepiota punaensis sp. nov. (Hawai'i USA) ... 475

Results & discussion

Our Bayesian and ML phylogenetic analyses generated identical tree

topologies when considering our ML tree with the highest log likelihood

(-4599.22) shown in Fic. 1. The topology of our Lepiota phylogeny is similar to

prior analyses from the last 16 years and does not differ greatly from Vellinga’s

(2003) work using nrITS and nrLSU sequences, Qasim & al’s (2015; 2016)

works using nrITS sequences, Sysouphanthong & al’s (2012; 2016) works using

nrITS sequences, and Liang & al’s (2018) work using nrITS, nrLSU, IGS, and

mtSSU sequences (but see Liang & al. 2011 using nrITS, nrLSU, IGS, and

mtSSU sequences where species with a trichoderm of long elements in clade

2 were dispersed).

Bootstrap support values for clades 2 and 4 (as defined by Vellinga 2003)

are too low to draw general conclusions about the deeper relationships of our

Hawaiian taxa within the genus, but our species clusters do reveal interesting

information about close relatives of Hawaiian Lepiota s.l. species.

Lepiota punaensis is always on a long branch by itself with the closest

nrITS match in GenBank an 87% similar Australian sequence (KP012854).

Lepiota punaensis is sister to the cluster of species from Hawai‘i, Canada,

Australia, and Europe, all of which (except the Australian collection) are

morphologically identified as L. elaiophylla. The Hawaiian L. cf. elaiophylla

is genetically identical (99.9% similar, TABLE 2) to a collection identified as

L. elaiophylla from a greenhouse in Canada, close to (98.3% similar, TABLE

2) a collection from a native Eucalyptus and Acacia swamp in Australia (M.

Barrett, pers. comm.); these three geographically separated collections form a

clade sister to the paratype of L. elaiophylla from the Netherlands, where the

species was originally described. The collection from the Netherlands is 92.9%

similar to the Hawaiian collections based on our alignment (TABLE 2).

Regarding other Hawaiian collections with a trichoderm of predominantly

long elements, L. rubrobrunnea is on a long branch in a poorly supported

group and its closest relatives are unclear. A Lepiota sp. collected on the UH

Manoa Campus on O‘ahu clusters in a well-supported clade with L. vellingana

Nawaz & Khalid, currently known from Pakistan (Nawaz & al. 2013), and

L. farinolens Bon & G. Riousset, from France (Bon 1992). A species collected

on the Island of Kaua‘i clusters in a well-supported clade with three taxa

from Thailand and L. subincarnata, a species known from Europe, North

America, and South Asia (Razaq & al. 2014). It may be conspecific with

sequence JN224826, a species collected in Chiang Mai Province in 2008 that,

according to notes in GenBank, is “close to L. subincarnata?

476 ... Stallman & al.

. Cf. mg a A MK412589

. cf. elaiophylla MK412588

. cf. elalophy lla MK412581

ees a {Canada MH979467

oat (Australlia) KP012854

‘elaiopnytia (Netherlands) AF391024

L. punaensis MK412577

99/10] [ punaensis MK412578

L. punaensis MK412572

99/1.0 rae sp. (Kaua‘i) AY176402

L. sp. AM ) JN224826

subincarnata AE. 76491

sp. (Thailand) JN224825

73/1.0 © sp. (Thailand) JN224828

L. brunneoincarnata FJ998395

1.0 L. farinolens AY 176368

oa vellingana HE974764

6/0.97 98/1.0 (0

op O‘ahu) MK412617

L. helveola mriosba

L. himalayensis HE614898

rubrobrunnea MK412583

86/0.99-— | . [reaopril. AY 176480

clade 2

trichoderm of long elements

o6it10 Thailand) HQ647293 Bo

L. andegavensis KP004931 + 2

89/1.0 L. castanea AY176463 Sie,

eu L. pilodes AY 176476 oa

L. tomentella EF080868

ods L. castaneidisca AF391061

L. cristata U85327

95/1.0 L. apatelia AY176462

L. thiersti AY176492

99/1.0 L. erminea AY176470

aanof— L. magnispora AF391005

*4 L. felina U85330

L. xanthophylla AY176405

L. sp. (Hawai‘i) MK412575

99/1.0; C. sp. (Hawai‘i) MK412604

ano C. sp. (Hawai‘i) MK412600

99/10 Uncultured Fungus (Australia) FJ F 1528742

sp. (Australia) KP012693

C. pulverulenta AF391035

M. haematospermum KF953545

zo C. seminuda JF907983

C. hetieri AY176459

99/1.0, L. aspera MK412598

L. aspera KP843884

L. cf. luteophylla AY176475

Ch. molybdites MK412591 | outgroup

0.03

clade 1

clade 4

Fic. 1. Phylogenetic analysis of selected sequences and species of Lepiota based on nrITS

sequences. The phylogeny was inferred using the Maximum Likelihood method and is drawn to

scale, with branch lengths measured in the number of substitutions per site. Maximum Likelihood

bootstrap support values >70% are shown in bold, and Bayesian posterior probabilities 20.95 are

shown in normal type. Hawaiian taxa are in bold, and islands are indicated for taxa not identified

to species. C. = Cystolepiota, Ch. = Chlorophyllum, L. = Lepiota, and M. = Melanophyllum.

The Hawaiian Cystolepiota species clusters in a well-supported clade

with two unknown taxa from Australia, and these are sister to Cystolepiota

pulverulenta (Huijsman) Vellinga [= C. petasiformis (Murrill) Vellinga], a

purportedly widespread species found in Europe (Vellinga 2001), western

North America (Vellinga 2006), and India (Kumar & Manimohan 2009).

The closest Australian sequence (FJ528742) is 96% similar to the Hawaiian

Lepiota punaensis sp. nov. (Hawai'i USA) ... 477

Cystolepiota species, and its lack of any closer matches in GenBank and

association with native vegetation supports the inference that it is a Hawaiian

endemic (Hemmes & Desjardin 2002). A poorly known Lepiota (MK412575)

that was only collected once in this study is on its own long branch in clade 1

(with only 74% support), and its closest relatives are unclear.

There are few data at this time to support broader conclusions about the

origins and full ranges of Hawaiian Lepiota s.l. taxa. Lepiota pseudorubella

lacks molecular data, L. rubrobrunnea and L. sp. (MK412575) have uncertain

placement in our tree due to low support values, and L. sp. (MK412617) is

on a relatively long branch in a clade with dispersed taxa. Lepiota aspera and

Melanophyllum haematospermum (Bull.) Kreisel (for which no Hawaiian

sequences are available) have widespread distributions, and L. pseudorubella,

L. rubrobrunnea, and L. cf. elaiophylla have all been collected in artificially

warm environments in temperate areas such as Canada and Germany (Gubitz

2008). The only putatively native Hawaiian taxon, Cystolepiota sp., is likely of

Australian origin, while a collection from Kaua‘i (AY176402) clusters with

several species from Thailand. Lepiota punaensis and L. cf. elaiophylla have

a close relative found in native Australian vegetation, but this is far from

conclusive in establishing a location of origin or theory of dispersal.

Taxonomy

Lepiota punaensis Stallman, sp. nov. PLATE 2

MB 832104

Differs from Lepiota elaiophylla by its white lamellae and from L. subincarnata by its

pileus covering containing short and narrowly clavate cells.

Type: USA, Hawai‘i, Hawai‘i Island, Puna District, MacKenzie State Recreation Area,

19°26’11”N 154°51’57”W, in duff under Casuarina equisetifolia L. and Cocos nucifera L.,

9 Dec 2016, coll. JKS 86 (Holotype, HAW-F-00256, GenBank: MK412577).

ETYMOLOGY: from the Puna District of Hawai‘i Island, the only known locality of this

species.

PrLEus 8-24 mm broad, at first hemispherical, becoming obtusely conical to

obtusely broadly conical, then convex to plano-convex in age; pileus covered

in a dense mat of warm brown (6E6 to 7E4-5; occasionally 7F6) fibrils when

young, quickly breaking into minute to small scales, composed of generally

appressed aggregated fibrils, over a white to off-white (6B2) background as

pileus expands, fibrils denser towards the cracked, or remaining intact disc

surface, lessening towards margin, margin aspect straight, shape entire to

eroded in age, often with remnants of the pileus covering adhering when

young, dry, non-hygrophanous; CONTEXT thin (<3 mm), white, unchanging;

478 ... Stallman & al.

LAMELLAE, L = 36,1 = 1-3, free, close, subventricose to ventricose, <3 mm broad,

white to off-white, edge even to eroded in age; stiPE 14-35 x 1-3 (-4 at base)

mm, cylindrical, equal to tapering upwards, off-white to light brown (6B2, 8C2),

covered with aggregated fibrils of similar appearance to pileus covering on

lower 1/2 to 1/3 of stipe that are easily removed and become less conspicuous in

age, appearing glabrous above this zone, but usually with thin covering of white

hyphal strands oriented parallel to the stipe (lens), also easy to remove; stipe

darkening over time to brown (6F5) where handled; ConTExT thin, concolorous

with exterior of stipe, stuffed with white hyphae when young, hollow in age,

base equal to slightly enlarged; BASAL TOMENTUM absent to inconspicuous;

RHIZOMORPHS absent or present, white; ANNULUS present only as annular zone;

SPORE DEPOSIT white; EDIBILITY toxic, likely contains amatoxins; ODOR none

to fungal; TASTE none to mildly fungal.

BasIDIOSPORES [94, 6, 4] (5.4-)6.3-6.6(-7.5) x (3.2-)3.7-3.8(-4.4) um

[x= 6.4 x 3.8 um], Q=(1.4—)1.6-1.8(-2.0) [Qm = 1.7], ellipsoid to subcylindrical

(generally oblong) in side view, usually with a straight adaxial (suprahilar

depression absent) and convex abaxial side; oblong to rarely obovoid in

frontal view, hyaline in H,O and NH,OH, dextrinoid, congophilous, weakly

cyanophilous, non-metachromatic in Cresyl Blue, germ pore absent, mono to

less commonly biguttulate; BAsIp1A [63, 6, 4] (21.2-)25.6-33.3(-34.7) x (5.9-)

6.5-8(-8.3) um [x = 28.9 x 7.2 um], hyaline, thin-walled, narrowly clavate to

subclavate, sometimes with flexuous wall, with 4 sterigmata; LAMELLAR-EDGE

sterile; CHEILOCYSTIDIA [44, 6, 4] (15.6—)22.3-24.6(-31.3) x (5.1-)8.9-10.2

(-14.0) um [x = 23.5 x 9.5 um], narrowly clavate to more commonly clavate or

broadly clavate, hyaline, thin-walled; PILEUS COVERING an intricate trichoderm

of long and shorter inflated cylindrical, sometimes flexuous, golden-colored

hyphae with parietal pigment, thin to slightly thick-walled, shorter elements are

present and common, but not in a distinctive layer, terminal elements narrowly

clavate, cylindrical, inflated cylindrical, or less commonly irregular/flexuose

narrowly clavate, or with a mucronate tip, <160 um long and 12 um diam,

clamp connections common; STIPITIPELLIS a cutis of tightly packed hyaline,

narrowly cylindrical hyphae <5 um diam, scales below annular zone similar

to those on pileus but generally composed of shorter elements, above annular

zone irregularly present and mostly composed of long, narrowly cylindrical,

hyaline, clamped hyphae; ANNULUS as zone only; CLAMP CONNECTIONS present

in all tissues.

ADDITIONAL SPECIMENS EXAMINED—USA, Hawai‘i, Hawai‘i Island, Puna District,

MacKenzie State Recreation Area, 19°26’11”N 154°51’57”W, 17 Sep 2016, coll. JKS

Lepiota punaensis sp. nov. (Hawai'i USA) ... 479

20 um

PLATE. 2. Lepiota punaensis (holotype, HAW-F-00256). a. Basidiomata; b. Basidiospores; c. Basidia

(with four sterigmata; only two shown in this basidial view); d. Cheilocystidia; e. Stipe covering;

f. Pileus covering.

64 (HAW-F-00257; GenBank MK412572); 5 Jan 2017, coll. JKS 87 (HAW-F-00258;

GenBank: MK412578); 1 February 2018, coll. JKS 186 (HAW-F-00255); 2 January 2019,

coll. JKS 204 (HAW-F-00259).

480 ... Stallman & al.

ECOLOGY & DISTRIBUTION — Solitary to scattered on needles and seed pods of

Casuarina equisetifolia in wet, lowland, alien environments. January, February,

September, and December. Currently known only from the Puna District of

Hawai'i Island.

CoMMENTS—Lepiota punaensis is known only from Casuarina equisetifolia

duff in the Puna District of Hawai‘i Island. The species is recognized by its

small size, lack of an annulus, and small, appressed brown fibrils over an

off-white cap context. Its oblong spores and pileus covering of long elements

without a layer of shorter clavate elements (although shorter elements are

present) place this species in L. subsect. Helveolinae with other species that

contain amatoxins.

Although L. punaensis is a small, relatively nondescript species, few

described species provide similar morphological matches, and its nrITS

sequence is unique. Recent studies from around the world such as Thailand

(Sysouphanthong & al. 2012), China (e.g., Liang & al. 2018), Pakistan (Qasim

& al. 2016), and the Dominican Republic (Justo & al. 2015) have described

new Lepiota species from the (neo)tropics, and have included DNA

sequence data that excludes any species from these studies, or other studies

incorporating nrITS sequence data, from providing a match to L. punaensis.

Older literature can be more difficult to interpret, but a review of species

sharing similarities, and newly described species without molecular data also

do not reveal a close match.

Lepiota zalkavritha T.K.A. Kumar & Manim. is a species of small size in

L. sect. Ovisporae described from India with no nrITS sequence (Kumar &

Manimohan 2009). Lepiota zalkavritha differs from L. punaensis in having

occasional apical outgrowths on its cheilocystidia, being more robust, having

darker brown coloration on pileus and stipe, and growth in soil (compared

to Casuarina equisetifolia duff). Lepiota brevipes Murrill is a small species

described from Florida in L. sect. Ovisporae with similar coloration to

L. punaensis, but the stipe is often much wider, and the spores are smaller

at 4.1-5 x 2.5-3.1 um (Akers 1997). Lepiota cinnamomea Cleland is a

small species from Australia in L. sect. Ovisporae (Aberdeen 1992), but the

pinkish-brown and dark-brown color descriptions of the scales and disc, and

shorter elements in the pileus covering (only <68 um long) rule it out. Other

small species described from Australia and stated to be in L. sect. Ovisporae

(Grgurinovic 1998) have annuli (e.g., L. rimosa Murrill) or color-changing

reactions not present in L. punaensis.

Lepiota punaensis sp. nov. (Hawai'i USA) ... 481

Pegler’s work from Ceylon (1972) reveals species in L. sect. Ovisporae that

are much larger, have unique red or yellow colors, or have annuli that do not

match L. punaensis, and his works from Africa (1977) and the Caribbean

(1983) do not reveal close matches. Rick’s (1905, 1907, 1920, 1926, 1930,

1937, 1938, 1961) works from South America present few good matches

for L. punaensis and Pereira’ (1998, 2000, 2001) present none. Rick does

describe a variety of Lepiota species with arachnoid veils (e.g., L. flavipes Rick

and L. hypholoma Rick), but other features such as the coloration or spore

sizes are different.

The closest morphological match appears to be Lepiota subincarnata,

a well-known species described from Europe that is also found in North

America and Asia (Razaq & al. 2014). Lepiota punaensis differs in having a

typically smaller pileus and containing pileus covering elements that can have

flexuose walls, be narrowly clavate, and contain shorter elements (although

not in a distinctive layer), all of which L. subincarnata lacks.

Lepiota cf. elaiophylla Vellinga & Huijser PLATE 3

Piteus (15-)25-40(-50) mm broad, at first paraboloid to narrowly

hemispherical, then convex, expanding to plano-convex or applanate in age,

completely covered in warm brown to dark brown (7F5, 8F4, 8F6, 8F8, 9F5)

fibrils when young, quickly breaking up as the pileus expands into small to

medium, generally appressed scales of aggregated fibrils over a dull yellow (3A4,

3B5, 4A3, 4A2) background; fibrils concentrated at disc (sometimes remaining

unbroken) and gradually lessening towards margin to reveal more of surface;

margin aspect straight, shape entire to eroded in age, sometimes with medium

to large remnants of pileus covering adhering to margin, non-hygrophanous,

dry; CONTEXT <5 mm thick, yellow (3A3, 3A4, 4B4, 4A3) fading to brown or

reddish brown over time; LAMELLAE L = 32,1 = 3 (usually), free, ventricose, up

to 9 mm broad, dull yellow to sulphur yellow (3B6, 3B4), sometimes darkening

in age (3A4, 4B4, 4C5); stripe 20-40 x 3-7 mm, cylindrical (although sometimes

flattening near the pileus) and equal (rarely tapering downwards), yellow (3A3,

3B4, 3A4, 4B3, 4A2, 3B4), with a clear annular zone beginning *% to % from the

bottom of the stipe, above which the stipe appears glabrous, but remnants of

the white, cortina-like partial veil generally remain, uncommonly aggregating

into larger visible structures, below covered in medium to large, horizontally-

oriented brown scales, generally appressed, but more uplifted than those on

the pileus, concolorous with those on the pileus; CONTEXT fleshy-fibrous, color

as in pileus or slightly lighter; when young solid to stuffed with translucent to

482 ... Stallman & al.

white hyphae, hollow in age; RHIZOMORPHS present, white; BASAL TOMENTUM

generally present; SPORE DEPOSIT white; EDIBILITY toxic, likely contains

amatoxins; ODOR none to mildly fungal; TAsTE none to mild.

BASIDIOSPORES [150, 7, 5] (5.8-)7.0-7.2(-8.4) x (3.3-)3.7-3.8(-4.2)

um [x = 7.1-3.8 um], Q = (1.5-)1.9(-2.3) [Qm = 1.9], narrowly ellipsoid to

subcylindrical (usually oblong) in side view with a convex abaxial side and

straighter adaxial side (mild suprahilar depression rarely present), oblong

to subcylindrical in face view, hyaline in H,O and NH,OH, dextrinoid,

congophilous, weakly cyanophilic to cyanophilic and non-metachromatic in

Cresyl Blue, germ pore absent, mono- or (less commonly) bi-guttulate; BAsIDIA

[57,7,5] (20.6-)23.1-28.7(-34.6) x (5.7-)7.2-8.8(-9.2) um [x = 26 x 8 um],

narrowly clavate to clavate, hyaline, thin-walled, 4-spored, rarely 2-spored;

LAMELLAR-EDGE Sterile; CHEILOCYSTIDIA [39, 5, 4] (15.5-)20.1-30.1(-39.3)

x (5.2-)6.1-9.8(-10.3) um [x = 24.5 x 7.9 um], generally narrowly clavate to

clavate, but also cylindrical and somewhat flexuous, sometimes with an apical

extension, narrowly utriform, or less commonly subconical, often septate with

one to two short, cylindrical basal cells, with or without clamp connections,

hyaline, thin-walled; PLEUROCYSTIDIA absent; PILEUS COVERING an intricate

trichoderm of mostly long erect and repent, generally cylindrical hyphae with

brown intracellular and parietal pigments, lacking a layer of short clavate

elements, terminal cells 25-240 x 4-15 um, most commonly 60-100 um in

length, inflated cylindrical to narrowly clavate, often flexuous; STIPITIPELLIS

a cutis of narrowly cylindrical hyaline hyphae 3-7 um diam with stipe scales

below annular zone similar to scales on pileus, above this zone a covering of

thin, hyaline, cylindrical, often flexuous hyphae up to 4 um diam and 100 um

long; ANNULUS as a cortinoid zone only; rarely with stipe covering elements

aggregating to create a medial to superior raised region resembling an

adhering “ring”; CLAMP CONNECTIONS present in all tissues; may be absent in

cheilocystidia.

SPECIMENS EXAMINED—USA, Hawai‘i, Hawai‘i Island, Hilo District, University of

Hawai‘i Hilo Botanical Garden, 19°42’10”N 155°04’56”W, 28 September 2016, coll.

JKS 69 (HAW-F-00244); Hilo, ‘Imiloa Astronomy Center, 19°42’07”N 155°05’16’W,

5 October 2016, coll. JkKS 71 (HAW-F-00245; GenBank MK412589); 6 November

2016, coll. JKS 77 (HAW-F-00246; GenBank MK412588); Puna District, MacKenzie

State Recreation Area, 19°26’22”N 154°51’4”W, 15 November 2017, coll. JKS 174

(HAW-F-00247).

ECOLOGY & DISTRIBUTION—Scattered to gregarious, sometimes cespitose,

in wood chips, on nutrient-rich soil in disturbed settings, or on Casuarina

equisetifolia duff in wet, lowland, alien environments. January, May, and July

Lepiota punaensis sp. nov. (Hawai'i USA) ... 483

Pirate. 3. Lepiota cf. elaiophylla (HAW-F-00246). a. Basidiomata; b. Basidiospores; c. Basidia

(generally with four sterigmata; only two shown in these basidial views); d. Cheilocystidia; e. Pileus

covering.

to December. Currently only known from the Puna and South Hilo Districts

on Hawai‘i Island; L. elaiophylla in the broad sense (see discussion) is reported

484 ... Stallman & al.

from Australia, Europe, North America, South America, India, and possibly

Africa (as L. xanthophylla P.D. Orton).

ComMMENTS—Lepiota cf. elaiophylla occurs on wood chips, nutrient-rich soil

in disturbed environments such as gardens, and under coastal Casuarina

equisetifolia where it can fruit singly, or in large groups and cespitose clusters.

Although many yellow lepiotaceous fungi occur in Hawai‘i, L. cf. elaiophylla

is easily distinguishable by its overall sulphur yellow coloration (including the

gills) with brown scales on the pileus, robust stature (differentiating it from

delicate yellow species in Leucocoprinus), and clear annular zone on the stipe

with cortina-like partial veil remnants above a base covered in brown scales. Its

oblong spores and pileus covering of primarily long elements place this species

in L. subsect. Helveolinae with other species containing amatoxins.

A BLAST search reveals the top match by identity in GenBank to represent

a collection identified as Lepiota elaiophylla by Else Vellinga, the co-describer

of this species, from a greenhouse in Canada at 99.9% sequence similarity

(KP012854). The next closest match (at 98.3% similarity) is MH979467 from a

specimen collected near Darwin Australia growing in a native Eucalyptus and

Acacia swamp habitat (M. Barrett, pers. comm.). Micrographs of the Australian

collection (T. Lebel, pers. comm.) reveal spores that are more ellipsoid than

in most collections of L. cf. elaiophylla, so it is unclear without additional

Australian specimens whether these two taxa are morphologically identical.

Our phylogeny of Lepiota (Fic. 1) shows the Hawaiian L. cf. elaiophylla

and the Canadian collection identified as L. elaiophylla as identical, with the

Australian collection close on its own short branch. The Dutch paratype of

L. elaiophylla is sister to the clade containing the Hawaiian, Canadian, and

Australian collections, and a well-supported (84% bootstrap support, 1.0

Bayesian posterior probability) clade is formed when considering all taxa,

including the paratype collection. See TABLE 2 for a comparison of pairwise

genetic distances between the species discussed.

Lepiota elaiophylla was described from the Netherlands and is reported

throughout Europe (e.g., Holec & Halek 2008), Brazil (Wartchow & al. 2008,

Ferreira & Cortez 2012), India (Kumar & Manimohan 2009), and possibly

Africa (as L. xanthophylla; Pegler 1977). The Hawaiian L. cf. elaiophylla is

clearly closely related to L. elaiophylla, from which it is macroscopically

indistinguishable. The only differences are (morphologically) a more variable

cheilocystidia shape in L. cf. elaiophylla and (molecularly) a different nrITS

sequence, although some European collections outside the Netherlands and

Brazilian collections show variation in cheilocystidia shape.

Lepiota punaensis sp. nov. (Hawai'i USA) ... 485

For a discussion of other Lepiota spp. with yellow lamellae (including the

macroscopic lookalike L. xanthophylla, which is differentiated by containing

a layer of short, clavate elements in the pileus covering and which has not

been recorded from Hawai‘i), see Vellinga & Huijser (1997). Collections of

the European L. xanthophylla and L. elaiophylla are described with differently

colored dried lamellae (yellow in xanthophylla and dirty olive-brown for

elaiophylla). Collections of L. cf. elaiophylla generally have dark yellow lamellae

when dried, although older, poorly dried, or damaged specimens may exhibit

brown colors; so this does not appear to be a useful taxonomic feature at this

time.

More information is needed to distinguish species concepts among L. cf.

elaiophylla, the Australian collection, and L. elaiophylla from the Netherlands

and throughout its reported range. One option is to argue that L. cf. elaiophylla

falls within normal morphological variation (minor cystidia differences) of

L. elaiophylla, a widespread species with high molecular variation and some

morphological variation as evidenced by its presence in India (Kumar &

Manimohan 2009), Brazil (Wartchow & al. 2008, Ferreira & Cortez 2012),

and Australia. This option cannot be proven but falls on the conservative side

of not describing a new taxon without first examining molecular data from

other regions that might reveal whether there is a range of variation in the

nrITS or two distinct taxa. The other option is to argue that L. cf. elaiophylla is

clearly molecularly different from the Dutch L. elaiophylla (over 7% pairwise

distance) and supported by a morphological difference (cheilocystidia

shape). This argument is supported by pointing out that the related species

L. brunneoincarnata and L. subincarnata initially showed little to no molecular

variation across their respective geographic ranges (Europe and Central Asia;

Europe, South Asia, and North America) when examined by Razaq & al.

(2014).

At this time, more molecular data from Europe, South America, India (and

possibly Africa) and morphological data from India and Africa are needed to

clarify the species concept. The Indian collection was not given a full species

description, but collections from South America reportedly have smaller

spores as well as lageniform (Ferreira & Cortez 2012) or subfusoid (Wartchow

& al. 2008) cheilocystidia, while collections from greenhouses in Europe

primarily have narrowly clavate to clavate cheilocystidia (with the exception

of the subglobose, and oval to pyriform cheilocystidia noted by Breitenbach &

Kranzlin 1995). At this time, habitat does not seem to be a useful taxonomic

feature as both Hawaiian and Dutch collections have been found in greenhouse

486 ... Stallman & al.

environments, although only the Hawaiian L. cf. elaiophylla is currently

confirmed from non-artificial habitats in (sub)tropical environments.

Key to the Lepiota of Hawai'i Island

—

Oo OW

Pileus brownish with covering of acute pyramidal scales composed of

elobase on inflated, Clements t..:, 3.) ft os 2h 2 Se Se, sgce th eget Ser arp sca 2

. Pileus color variable (including brownish) with covering lacking

acute pyramidal scales, at least some long (2100 ttm) cylindrical

ClEMIENtS: PRESEHIS. fat. oo eee vate ten Un Mee cet MhrY cag stains Ovid ip ey « 3

. Basidiome with large membranous annulus,

spores oblong to cylindrical, averaging 6.4 x 2.7 um ................ L. aspera

. Basidiome annulus and other features poorly known,

spores ellipsoid to oblong, averaging 4.4 x 2.7m .............. 0.0. eee L. sp.

(MK412575; poorly known; see text)

. Basidiome with ellipsoid spores averaging 5X 3UM..... eee eee eee 4

. Basidiome with ellipsoid to oblong spores averaging 7 x 3-4um _............... 5

. Basidiome with prominent basal stipe ornamentation concolorous

with pileus scales; farinaceous odor when crushed; pileus and stipe

covering of long flexuose hyphae lacking abundant shorter clavate

Clemence s.r hen ween. Mer ety atten Sea eee ent ete eee, L. rubrobrunnea

. Basidiome generally lacking prominent basal stipe ornamentation,

stipe scales present usually lighter than pileus scales; lacking a

farinaceous odor when crushed; pileus and particularly stipe

covering with abundant short clavate elements ............... L. pseudorubella

. Basidiome with brown scales over a dull yellow pileus background,

lamellae and stipe also yellow; arachnoid partial veil usually leaving

only zone on stipe; pileus covering a trichoderm of long (<240 um)

elements lacking shorter cells..................0..000- Lepiota cf. elaiophylla

. Basidiome with warm brown scales over an off-white pileus background

lacking yellow coloration in all tissues; annulus a zone only;

pileus covering a trichoderm of long (<160 um) elements with

shorter, sometimes clavate hyphae interspersed but not in a distinct layer;

shorter hyphae more common in stipe covering, currently known only

from Casuarina equisetifolia duff in the Puna District of Hawai‘i

1S ha (6 hee pes A De nt ye prea, UM Rec Slay. pe Saas Ch ear L. punaensis

Acknowledgments

We would like to acknowledge the National Geographic Society, Puget Sound

Mycological Society, and Sonoma County Mycological Society for funding. Thank

you to Dr. Teresa Lebel and Dr. Matthew Barrett for sharing information on the

Australian collection (GenBank KP012854), to Sean Swift for a collection made on

O‘ahu (GenBank MK412617), and to Dr. Else Vellinga on a variety of topics related

Lepiota punaensis sp. nov. (Hawai'i USA) ... 487

to Lepiota taxonomy in general and L. elaiophylla in particular. We also thank

Dr. Brian A. Perry (Department of Biological Sciences, California State University

East Bay, USA) and Dr. Alfredo Vizzini (Department of Life Sciences & Systems

Biology, University of Torino, Italy) for presubmission review.

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MYCOTAXON

July-September 2020—Volume 135, pp. 491-500

https://doi.org/10.5248/135.491

Notes on rust fungi in China 8.

Pucciniastrum tiliae life cycle and

new host plants inferred from phylogenetic evidence

JING-XIN Jr, ZHUANG L?’, Yu Lr’, MAKOTO KAKISHIMA®?®

‘Engineering Research Center of Chinese Ministry of Education for Edible and Medicinal Fungi,

Jilin Agricultural University, Changchun, Jilin 130118, China

College of Plant Protection, Shandong Agricultural University, Taian 271000, China

>University of Tsukuba, Tsukuba, Ibaraki 305-8572, Japan

* CORRESPONDENCE TO: * 1096314395@qq.com * kakishima.makoto.ga@u.tsukuba.ac.jp

ABSTRACT—The life cycle connection between spermogonial and aecial stages of a rust

fungus found on Abies holophylla and uredinial and telial stages on Tilia mongolica and

T. mandshurica collected in northeast China were confirmed by phylogenetic analyses.

The rust, identified as Pucciniastrum tiliae, was confirmed by morphological observations.

The life cycle of this rust fungus is reported for the first time in China, and A. holophylla

and T: mongolica represent new host plants for the species.

Key worps—phylogeny, Pinaceae, Pucciniastreae, Tiliaceae, tree disease

Introduction

Pucciniastrum tiliae was first described with uredinial and telial stages on

Tilia cordata var. japonica [= T: japonica; Tiliaceae| collected in Hokkaido,

Japan (Hiratsuka 1897). This species has been since reported on T: amurensis

in Korea, China, and Russian Far East; T: americana and T. kiusiana in Japan;

T. mandshurica in China and Russian Far East; T. maximowicziana

[= T. miyabei] in China and Japan; T: platyphyllos in Japan; and T! tuan in

China (Hiratsuka 1936, 1958; Ito 1938; Cummins 1950; Cummins & Ling

1950; Wang 1951; Kim 1963; Tai 1979; Hiratsuka & al. 1992; Azbukina 2005,

492 ... Ji &al.

2015). The life cycle of this fungus was revealed by Kamei in Japan in 1934

through inoculations of basidiospores from teliospores on T: japonica and

T. maximowicziana into needles of Abies sachalinensis var. mayriana (Pinaceae)

(Hiratsuka & al. 1992). Although P tiliae is distributed in northeast China (Tai

1979), there are no reports of a spermogonial and aecial host in the country.

During field investigation of rust fungi in northeast China, we collected

uredinial and telial stages of Pucciniastrum on T. mandshurica and T: mongolica

in the mountains of Jilin and Heilongjiang Provinces. We also observed

occurrences of aecia producing long peridia on Abies holophylla near infected

trees of Tilia. Therefore, we suspected life cycle connections between the rust

on Abies and Tilia. Inoculation of plants by spores are an appropriate method

to clarify rust life cycles (Ji & al. 2017a, b). However, collection and propagation

of plants from the mountains of Maple Valley (Jilin) and Fenix National Park

(Heilongjiang) where we collected specimens are strictly prohibited as they

are nature conservation areas. Therefore, we applied molecular analyses to

resolve these problems and to clarify the life cycle (Liu & Hambleton 2013,

Ji & al. 2016, Padamsee & McKenzie 2017, Scholler & al. 2019). We report

here the results of phylogenetic analyses and morphological observations of

the specimens.

Materials & methods

Molecular analyses

Uredinial and telial stages on Tilia mandshurica and T: mongolica collected in

mountains in Jilin and Heilongjiang Provinces, China, were used for molecular

analyses. The spermogonial and aecial stages on Abies holophylla collected in Jiaohe,

Jilin Province, China, were also used for analyses.

The total genomic DNA was directly extracted from c. 200 spores obtained from

single sori on the leaf of each specimen using similar methods reported by Ji & al.

(2016, 2019). Specimens used in the experiments were deposited in the Herbarium

of Mycology, Engineering Research Center of Chinese Ministry of Education for

Edible and Medicinal Fungi, Jilin Agricultural University, China (HMJAU); all data

sequenced in this experiment were deposited in GenBank as shown in specimens

examined.

The sequences obtained from specimens were aligned following Ji & al. (2019).

ITS and 28S sequences related to P tiliae were retrieved from GenBank and added

to phylogenetic analyses (Ji & al. 2019). The accession numbers of these data are

shown in the phylogenetic trees (Fics 1, 2). Phylogenetic trees were constructed

using Melampsora laricis-populina Kleb. as outgroup according to Ji & al. (2019).

The alignment and trees were deposited in TreeBASE under http://purl.org/phylo/

treebase/phylows/study/TB2:S24909 (Fics 1, 2).

Pucciniastrum tiliae in China ... 493

Pucciniastrum tiliae HMJAU8581 [Tilia mandshurica|UT

Pucciniastrum tiliae HMSJAU8582 [Tilia mongolica|UT

satepii.nd Pucciniastrum tiliae HMJAU8583 [Tilia mongolica|UT

=< Pucciniastrum tiliaeg HMJAU8584 [Tilia mongolica|UT

65\68\0,94| [Pucciniastrum tiliae (AB 221412) [Tilia mandshurica]

Pucciniastrum tiliae (AB 221415) [Tilia japonica]

Pucciniastrum tiliae HMJAU8641 [Abies holophylla|SA

Thekopsora triangula (KF551204) [Cornus macrophylla]

Thekopsora ostryae (KC416004) [Ostrya japonica]

Thekopsora ostryae (KC415993) [Ostrya japonica]

Pucciniastrum agrimoniae (MG787142) [Agrimonia pilosa]

Pucciniastrum agrimoniae (MG787143) [Agrimonia pilosa]

Pucciniastrum agrimoniae (MG787145) [Agrimonia pilosa]

Pucciniastrum agrimoniae (KJ725375)

Thekopsora lanpingensis (KF551208) [Cornus schindleri]

100\100\1;7ekopsora lanpingensis (KF551211) [Cornus walteri]

Thekopsora lanpingensis (KF551213) [Cornus schindleri]

Thekopsora lanpingensis (KF551214) [Cornus officinalis |

Pucciniastrum coronispora (MH780998) [Galium trifidum]

Pucciniastrum coronispora (MG787127) [Galium davuricum]

Pucciniastrum coronispora (MG787128) [Galium boreale]

Pucciniastrum coronispora (MG787132) [Galium bungei|

Thekopsora nipponica (KC416003) [Galium aparine]

L00\100\I-7hekopsora nipponica (KC415995) [Galium aparine]

Pucciniastrum rubiae (MG787136) [Rubia cordifolia]

#1*\0.56| [Pucciniastrum rubiae (MG787139) [Rubia chinensis]

Pucciniastrum rubiae (MG787140) [Rubia chinensis]

Pucciniastrum guttata (KC415999) [Galium odoratum]

Pucciniastrum rubiae (KC616007) [Rubia cordifolia]

Pucciniastrum annulospora (MH781004) [Kalimeris integrifolia]

97 | Pucciniastrum annulospora (MH781003) [Aster tataricus]

Pucciniastrum annulospora (MG787133) [Kalimeris lautureana]

‘Melampsora laricis-populina (MG787151) [Populus sp.]

70\*\0.75

98\97\0,89

79\82\0.86

99\L00\0.

0.04

Fic. 1. Phylogenetic tree constructed by Bayesian method based 28S rDNA sequences. MP/ML

bootstrap values precede Bayesian posterior probabilities (Bpp) on the nodes. Asterisks (*) indicate

bootstrap values <50% or Bpp <0.5. Sample data include species name, (voucher specimen or

GenBank accession number), and [host plant]. Sequences generated in this study are shown in

bold face. SA= Spermogonial and aecial stages; UT = Uredinial and telial stages.

Morphological observations

Light (LM) and scanning electron (SEM) microscopy were used to examine

morphological characters of rust specimens, including the size and shape of sori and

spores following Ji & al. (2019).

Phylogenetic results

The 28S dataset comprised 33 sequences with 496 total characters, of which 394

were constant, 30 parsimony-uninformative variable, and 72 parsimony-informative.

Parsimony analysis yielded one parsimonious tree with TL = 156, CI = 0.7692,

RI = 0.9231, and RC = 0.7101. Bayesian analysis resulted in average standard

deviation of split frequencies of 0.008983. The ITS dataset comprised 39 sequences

of 745 total characters, of which 228 were parsimony-informative. Parsimony

A494 ... Ji & al.

Pucciniastrum coronispora (MH780990) [Galium trifidum]

98\97\1] Pucciniastrum coronispora (MG787102) [Galium davuricum]

Pucciniastrum coronispora (MG787103) [Galium boreale]

1OO\OO Pr cciniastrum coronispora (MG787107) [Galium bungei|

Pucciniastrum nipponica (KC415792) [Galium davuricum]

16\9 1O0\1 00} Mcciniastrum nipponica (KC415794) [Galium aparine]

Pucciniastrum nipponica (KC415793) [Galium aparine]

Pucciniastrum annulospora (MG787108) [Kalimeris lautureana]

00\TOO q Pucciniastrum annulospora (MH780995) [Aster tataricus]

Pucciniastrum annulospora (MH780996) [Kalimeris integrifolia]

Pucciniastrum guttatum (KC415789) [Galium odoratum]

Pucciniastrum guttatum (KC415790) [Galium odoratum]

Pucciniastrum guttatum (KC415791) [Galium odoratum]

Pucciniastrum rubiae (MG787114) [Rubia chinensis]

Pucciniastrum rubiae (MG787115) [Rubia chinensis]

Pucciniastrum rubiae (KC415799) [Rubia cordifolia]

100\9 Pucciniastrum rubiae (KC415802) [Rubia cordifolia]

Pucciniastrum rubiae (KC415801) [Rubia cordifolia]

83\59\0.67 Pucciniastrum rubiae (MG787111) [Rubia cordifolia]

Pucciniastrum tiliae HMJAU8581 [Tilia mandshurica}UT

Pucciniastrum tiliae HMJAU8583 [Tilia mongolica}UT

Pucciniastrum tiliae HMJAU8641 [Abies holophylla]SA

65\*\0.8] Pucciniastrum tiliae HMJAU8582 [Tilia mongolica|UT

Pucciniastrum tiliae HMJAU8584 [Tilia mongolica}UT

9ON\9G\1I| Pucciniastrum tiliae (AB 221453) [Tilia japonica]

SOM Pucciniastrum tiliae (AB 221455) [Tilia mandshurica]

INIG\I Pucciniastrum kusanoi (AB221426) [Clethra barbinervis]

9NIN\I" Pucciniastrum boehmeriae (AB221449) [Boehmeria tricuspis]

Thekopsora triangula (KF551219) [Cornus macrophylla]

Pucciniastrum agrimoniae (MG787117) [Agrimonia pilosa]

100\100\1|Pucciniastrum agrimoniae (MG787118) [Agrimonia pilosa]

99\100 Pucciniastrum agrimoniae (MG787120) [Agrimonia pilosa]

‘Pucciniastrum agrimoniae (KJ486537)

Thekopsora ostryae (KC415796) [Ostrya japonica]

71\85\0.81 Thekopsora lanpingensis (KF551220) [Cornus walteri]

100\100\1| 7hekopsora lanpingensis (KF551222) [Cornus schindleri]

Thekopsora lanpingensis (KF551225) [Cornus schindleri]

Thekopsora lanpingensis (KF551224) [Cornus officinalis |

Melampsora laricis-populina (MG787126) [Populus sp.]

9ON\I\I

Fic. 2. Phylogenetic tree constructed by Bayesian method based on ITS rDNA sequences. MP/

ML bootstrap values precede Bayesian posterior probabilities (Bpp) on the nodes. Asterisks (*)

indicate bootstrap values <50% or Bpp <0.5. Sample data include species name, (voucher specimen

or GenBank accession number), and [host plant]. Sequences generated in this study are shown in

bold face. SA= Spermogonial and aecial stages; UT = Uredinial and telial stages.

analysis yielded one parsimonious tree with TL = 543, CI = 0.8066, RI = 0.9417, and

RC = 0.7596. Bayesian analysis resulted in average standard deviation of split

frequencies of 0.003112. Tree topologies formed by MP, ML, and MCMC methods

were identical among trees. The Bayesian phylogenetic trees are shown in Fic. 1

(28S) and Fie. 2 (ITS).

Both 28S and ITS phylogenetic trees included the uredinial and telial stages on

T. mandshurica (HMJAU 8581) and T: mongolica (HMJAU 8582, 8583, 8584) and the

spermogonial and aecial stages on A. holophylla (HMJAU 8641) in a monophyletic

clade (Fics 1, 2). Genetically identical to one another, they can be considered

the same species. The life connection between spermogonial and aecial stages on

A. holophylla and uredinial and telial stages on T: mandshurica and T: mongolica is

therefore strongly supported by the phylogenetic analyses.

Pucciniastrum tiliae in China ... 495

Taxonomy

The phylogenetic analyses reveal that this rust fungus has a

heteromacrocyclic life cycle, producing spermogonia and aecia on Abies

holophylla (Fic. 3p) and uredinia and telia on Tilia mandshurica and

T. mongolica (Figs 3a-c). LM and SEM observations showed that the

morphological characteristics of spermogonial and aecial specimens on

A. holophylla collected from the field were identical with the descriptions

of Pucciniastrum tiliae reported by Hiratsuka (1936) and Hiratsuka & al.

(1992), except for outer wall surface structure of the peridial cells (Fics 3p,

4A-C, A-D). Under SEM, these walls are minutely verrucose, in contrast to

the smooth surface described by Hiratsuka & al. (1992). Previously, P. tiliae

spermogonial and aecial stages have been reported on Abies species only

from Japan, so this represents the first report of the P. tiliae life cycle from

China. Abies holophylla is a new spermogonial and aecial host plant and

occurs in northeastern China, Korea, and Russian Far East.

After clarifying the life cycle connection of P tiliae with phylogenetic

analyses, we described its morphology based on specimens collected in the

field.

The morphological characters of uredinial and telial specimens on

T. mandshurica and T: mongolica were similar to one another (Fics 34-c,

4D-F, 5E,F) and identical to the descriptions of P. tiliae by Hiratsuka (1936),

Hiratsuka & al. (1992) and Azbukina (2005, 2015). No researcher has

previously reported P. tiliae on T. mongolica (Hiratsuka 1936, 1958; Kim

1963; Tai 1979; Hiratsuka & al. 1992; Azbukina 2005, 2015), so we add

T: mongolica (known from Mongolia, northern China, and eastern Russia)

as a new host for the rust.

Pucciniastrum tiliae Miyabe, Bot. Mag. Tokyo 11: 47, 1897. Figs 3-5

Spermogonia amphigenous, pale yellow to yellowish, subcuticular, type 3

(Cummins & Hiratsuka 2003). Aecia with long peridia, Peridermium-type,

hypophyllous, cylindrical, orange-yellow. Peridia colorless, rupturing at the

sides and apex; peridial cells ovate to ellipsoid, slightly overlapping, inner and

outer walls thin, minutely verrucose. Aeciospores globose to ellipsoid, 15.0-22.0

x 13.0-16.5 um (av. = 18.5 x 15.0 um), walls hyaline, 1.0-2.0 um (av. = 1.5 um)

thick, densely verrucose with annulate verrucae.

Uredinia hypophyllous, scattered to gregarious, subepidermal, small,

round, covered by the epidermis, rupturing with pores, pulverulent. Peridia

hemispherical, delicate, firm, rupturing at the apex, peridial cells small,

496 ... Ji & al.

Fic. 3. Pucciniastrum tiliae. A. Uredinia and telia on lower leaf surface of Tilia mongolica. B. Telia

produced on lower leaf surface of T: mongolica. C. Uredinia produced on lower leaf surface of

T. mongolica. D. Aecia with long peridia produced on needles of Abies holophylla.

irregularly polygonal, walls smooth, thin, ostiolar cells rounded, walls

smooth. Urediniospores subglobose, ellipsoid, obovate, or oblong, 16.5-25.5

x 8.0-15.5 um (av. = 20.5 x 12.5 um), walls echinulate, 0.5-2.0 um (av. = 1 um)

thick. Telia hypophyllous, subepidermal, scattered or in small groups, at

first orange-yellow, then brown to reddish in color. Teliospores forming

under the epidermis, attached laterally, subglobose, oblong, or somewhat

angular, divided into 2-6 cells by vertical or oblique septa, 21.0-37.0 x

5.5-14.0 um (av. = 30.5 x 9.5 um), walls thin, 0.5-1.0 um (av. = 0.7 um)

thick, smooth.

SPECIMENS EXAMINED—CHINA, Jilin Province, Jiaohe, stages 0 & I, on Abies

holophylla Maxim., 16 June 2019 (HMJAU8641, GenBank MN294706, MG787116);

stages II & IH, on Tilia mongolica Maxim., 11 September 2017 (HMJAU8583, GenBank

MG787149, MG787124; HMJAU8584, GenBank MG787150, MG787125); Helongjiang

Pucciniastrum tiliae in China ... 497

Fic. 4. Pucciniastrum tiliae observed with LM. A. Vertical section of spermogonium (type 3).

B. Aeciospores with verrucose surface. C. Vertical section of aecium surround by peridium (P).

D. Urediniospores with echinulate surface. E. Vertical section of uredinium with peridium (P),

opening by central pore. F. Vertical section of telia with laterally attached teliospores, covered by

host epidermis. Scale bars: A, C = 20 um; B = 30 um; D, E= 10 um; F = 15 um.

Province, Wuchang, stages II & III, on T: mongolica, 9 September 2017 (HMJAU8582,

GenBank MG787148, MG787123); stages Il & II, on T: mandshurica Rupr. & Maxim.,

9 September 2017 (HMJAU8581, GenBank MG787147, MG787122).

498 ... Ji & al.

Fic. 5. Pucciniastrum tiliae observed with SEM. A. Aeciospores with densely verrucose surface.

B. Aecium with long peridium. C. Outer walls of peridial cells. D. Inner walls of peridial cells.

E. Urediniospores with echinulate surface. F. Uredinium with peridium (P) and ostiolar cells (O),

opening by central pore. Scale bars: A, E = 5 um; B = 100 um; C, D = 20 um; F = 10 um.

Pucciniastrum tiliae in China ... 499

Hosts & DISTRIBUTION WITHIN CHINA—Abies holophylla, Jilin (new host;

this paper). Tilia amurensis Rupr., Jilin (Ito 1938, Tai 1979). Tilia mandshurica,

Heilongjiang (Ito 1938, Hiratsuka 1958, Tai 1979). Tilia mongolica, Heilongjiang,

Jilin (new host; this paper). Tilia tuan Szyszyt., Guizhou (Hiratsuka 1958, Tai 1979).

Acknowledgments

This work was financed by the Fungal Flora in Jilin Province (20130206073NY).

We thank Dr. E.H.C. McKenzie (Manaaki Whenua Landcare Research, Auckland,

New Zealand) and Prof. C.M. Denchev (Bulgarian Academy of Sciences, Sofia,

Bulgaria) for critical reading of the manuscript and suggestions.

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sori and spores from northeast of China. Mycological Progress 18: 529-540.

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MYCOTAXON

July-September 2020—Volume 135, pp. 501-512

https://doi.org/10.5248/135.501

Guayaquilia gen. nov., typified by /driella cubensis

FREDDY MAGDAMA’, DAYNET SOSA’, FERNANDO ESPINOZA’,

LIZETTE SERRANO’, SIMON PEREZ-MARTINEZ’, ELAINE MALOSSO},

MARGARITA HERNANDEZ-RESTREPO*, RAFAEL F. CASTANEDA-RUIZ>

' Escuela Superior Politécnica del Litoral, ESPOL,

Centro de Investigaciones Biotecnologicas del Ecuador, Campus Gustavo Galindo,

Km. 30.5 Via Perimetral, RO. Box 09-01-5863, Guayaquil, Ecuador

? Universidad Estatal de Milagro (UNEM1I), Facultad de Ingenieria,

Cdla. Universitaria Km. 1.5 via Milagro-Km26. Milagro 091706, Guayas, Ecuador

> Centro de Biociencias, Departamento de Micologia, Universidade Federal de Pernambuco,

Avenida da Engenharia, s/n Cidade Universitaria, Recife, PE, 50.740-600, Brazil

* Westerdijk Fungal Biodiversity Institute, 3508 AD Utrecht, The Netherlands

° 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 genus Guayaquilia is established to accommodate Idriella cubensis based

on morphology and phylogenetic analysis. DNA sequence data place these specimens as

incertae sedis separate from Microdochiaceae (Xylariales), forming a monophyletic lineage

separated from Neoidriella desertorum and phylogenetically distant from Idriella. The novel

genus is characterized by macronematous, tree-like, fasciculate, profuse dichotomously,

alternately, or irregularly branched, brown conidiophores with polyblastic, denticulate,

sympodial extended, intercalary and terminal conidiogenous cells that produce solitary,

sublunate, subnavicular, lunate, inequilateral, (0-)1-septate, hyaline conidia.

KEY worps—asexual fungi, hyphomycetes, Microdochiaceae, taxonomy, tropics

Introduction

Idriella, introduced by Nelson & Wilhelm (1956) with I. lunata as type species,

is distinguished by macronematous, mononematous, unbranched, and basally

502 ... Magdama & al.

inflated and apically slightly geniculate brown conidiophores, mostly reduced

to polyblastic, denticulate, sympodial extended conidiogenous cells. Conidia

are typically lunate or falcate, unicellular, hyaline, and acuminate towards the

apex (Nelson & Wilhelm 1956). Morgan-Jones (1979) updated and provided

brief nomenclatural commentaries on Idriella australiensis, I. couratarii,

I. desertorum, I. lunata, I. mycogonoidea, I. ramosa, I. vandalurensis, and

I. variabilis and proposed two new species, I. angustispora and I. bambusae.

Von Arx (1981), who presented a combined key for Idriella and Microdochium

species, proposed the conidial shape and the habitat as the main characters to

separate the genera; he included the Idriella species treated by Morgan-Jones

(1979), excluded I. tropicalis as similar to I. variabilis, and added five new

combinations (J. bolleyi, I. caespitosa, I. falcata, I. grisea, and I. tainanensis).

Subsequently, 16 new species have been described I. acerosa, I. amazonica,

I. angamosensis, I. asaicola, I. cagnizarii, I. campnospermae, I. cubensis,

I. euterpes, I. jambosae, I. licualae, I. mycophila, I. rara, I. rhododendri,

I. setiformis, I. stilboidea, and I. uncinospora (Castaheda-Ruiz 1985, 1986;

Castaneda-Ruiz & Arnold 1985; Castafteda-Ruiz & Kendrick 1991; Castafieda-

Ruiz & al. 1997; Matsushima 1985, 1995; Rodrigues & Samuels 1990, 1992;

Subramanian & Bhat 1987; Wu & al. 1997).

When the generic concept of Idriella was expanded, it becamea heterogeneous

genus (Seifert & al 2011). In the generic type, I. lunata, conidiophores are

unbranched and mostly reduced to conidiogenous cells producing unicellular

conidia on tiny denticles arranged along the apical and subapical surface of

each conidiogenous cell after several sympodial holoblastic extensions (Nelson

& Wilhelm 1956), but many other Idriella species produce conidiophores

branched in various patterns, sympodial extensions of the conidiogenous

cells confined to a small, nodose area at the apex, and conidiogenous loci as

inconspicuous denticles that produce unicellular or septate conidia. DNA

sequences of several Idriella species revealed the polyphyletic nature of this

genus, showing phylogenetic separation between I. lunata (type species) and

four other Idriella species, which consequently were transferred to four new

genera: Castanediella Hern.-Restr. & al., Idriellopsis Hern.-Restr. & Crous,

Neoidriella Hern.-Restr. & Crous, and Paraidriella Hern.-Restr. & Crous (Crous

& al. 2015, Hernandez-Restrepo & al. 2016).

During a workshop on fungal diversity conservation in Cacao forest areas,

two surveys of microfungi were conducted in Theobroma cacao plantations in

Guayaquil, Guayas Province, Ecuador, and several Idriella-like specimens were

found. LSU and ITS sequence analyses place these specimens as incertae sedis

Guayaquilia cubensis gen. & sp. nov. (Cuba, Ecuador) ... 503

and separate from Microdochiaceae (Xylariales). They are not congeneric with

Idriella lunata (the generic type), and we propose a new genus Guayaquilia to

accommodate them.

Materials & methods

Sampling and fungal strains studied

Samples of decaying plants were placed in plastic bags, transported to the

laboratory, placed in moist chambers, and treated according to Castafeda-Ruiz & al.

(2016). Individual conidia were separated from the plant material under a stereoscope

using an entomological needle and cultured in two different media: V8 (V8: 125 ml

V8 juice, 18 g agar, 1000 ml distilled water, pH 6.3) and Cornmeal agar (CMA: 20

g cornmeal, 18 g agar, 1000 ml distilled water, pH 6.3). Morphological observations

were made from cultures grown on CMA after five days, incubated at 25 + 1°C under

12 h alternating near-UV light and darkness (using Vica, FLB-20W T10 near-UV

lamp in an irradiation box). Colony colors were coded according to Rayner (1970).

Mounts were prepared in lactic acid (90%) or in polyvinyl alcohol-glycerol (8 g PVA

in 100 mL of water + 5 mL of glycerol) and lactofuchsin (0.1 g acid fuchsin, 100 mL

85% lactic acid) (Carmichael 1955). Microscopic characters were measured at x1000

using a Nikon Eclipse Ni-U microscope with DIC optics and a Nikon DS-Fi2 camera.

Pure cultures were deposited at the Microbial Culture Collection of the

Biotechnology Research Center, Guayaquil, Ecuador (CCM-CIBE), and the voucher

specimens and slide preparations were deposited in the Herbarium, Departamento

de Micologia, Universidade Federal de Pernambuco, Recife, Brazil (URM). Also

examined were the holotype and another specimen deposited as Idriella cubensis in

the INIFAT Fungus Collection.

DNA extraction, sequencing, and phylogenetic analysis

Genomic DNA was extracted from fresh mycelia grown on V8 Agar at 25°C using

a modified rapid extraction method for filamentous fungi according to Cenis (1992).

The primer pairs ITS1/ITS4 (White & al. 1990) and LROR/LRS (Vilgalys & Hester

1990) were used for the amplification of the internal transcribed spacers (ITS), and

part of the large subunit of the nuclear ribosomal RNA gene (LSU). PCR cycling

conditions followed Korabecna (2007; ITS) and Yang & al. (2017; LSU). PCR products

were sent to Macrogen Korea for purification and sequencing with the same primers.

Sequences were aligned and edited using MEGA v.6.0. Phylogenetic relationships

among taxa were addressed through Maximum Likelihood (ML) analysis using

MEGA v.6.0 and Bayesian analysis using Beast v1.10.4 (Drummond & al. 2012). Data

for each gene or region was analyzed both individually and together as a combined data

set. jModeltest v.2.1.10 (Darriba & al. 2012) was used to determine the best nucleotide

substitution model for both studies. Congruence between individual gene data sets

was tested using the partition homogeneity test (Farris & al. 1995) implemented in

PAUP v4.0b10 (Swofford, 2001), using a heuristic search option with random taxon

addition and TBR branch swapping with 1000 replicates. Tree topologies from ML

504 ... Magdama & al.

analysis of individual genes were also compared visually for congruence. For the

combined ML study, GTR was specified as the evolutionary model with estimated

proportion of invariables sites and gamma distribution as default parameters.

Number of substitution rates was set to four and topology search changed to SPR

for tree improvement. Nodal support was assessed by bootstrap analysis from 1000

replicates. Bootstrap values equal or higher than 70% were considered significant.

Bayesian inference was run with four Monte Carlo Markov (MCM) chains over 1

million generations with a sampling frequency of 1000 trees using the GTR+I+G

as the best model (Drummond & al. 2012). To detect if the sample distribution has

reached stationarity, convergence and the effective sample size (ESS) were checked

for each run. Posterior probabilities (PP) for the Bayesian analysis were determined

by calculating 50% majority rule consensus tree and added onto congruent nodes of

the ML tree topology. PP equal or above 0.95 were considered significant. Relevant

sequences of ITS and LSU from Hernandez-Restrepo & al. (2016) obtained from

GenBank, were included in the phylogenetic inference for comparison purposes

(TABLE 1).

TABLE I. Sequences of Guayaquilia cubensis and related species used in the

phylogenetic analysis. New sequences are indicated in bold.

SPECIES STRAIN # ITS LSU REFERENCE

Astrocystis IT1612 KP297404 KP340545 Daranagama & al. 2015

concavispora

Castanediella acaciae CBS 139896 KR476728 KR476763 Crous & al. 2015

C. cagnizarii CBS 542.96 KP859054 KP858991 Hernandez-Restrepo &

al. 2016

CBS 101043 KP859051 KP858988 Hernandez-Restrepo &

al. 2016

C. couratarii CBS 579.71 KP859050 KP858987 Hernandez-Restrepo &

al. 2016

Guayaquilia cubensis MUCL 39017 KC775733 KC775708 Becerra-Hernandez &

al. 2016

CCMCIBE-H312 MH777025 MH777024 This Study

CCMCIBE-H320 MH777026 MH777023 This Study

Idriella lunata CBS 177.57 KP859043 KP858980 Hernandez-Restrepo &

al. 2016

CBS 204.56 KP859044 KP858981 Hernandez-Restrepo &

al. 2016

CBS 209.60 KP859045 KP858982 Hernandez-Restrepo &

al. 2016

CBS 736.74 KP859046 KP858983 Hernandez-Restrepo &

al. 2016

Idriellopsis CBS 575.92 KP859052 KP858989 Hernandez-Restrepo &

uncinospora al. 2016

Kretzschmaria deusta CBS 163.93 KC477237 KY610458 Wendt & al. 2018

Guayaquilia cubensis gen. & sp. nov. (Cuba, Ecuador) ... 505

Microdochium CBS 243.83 KP858994 KP858930 Hernandez-Restrepo &

albescens al. 2016

M. citrinidiscum CBS 109067 KP859003 KP858939 Hernandez-Restrepo &

al. 2016

M. lycopodinum CBS 109398 KP859005 KP858941 Hernandez-Restrepo &

al. 2016

M. majus CBS 741.79 KP859001 KP858937 Hernandez-Restrepo &

al. 2016

M. CBS 445.95 KP858997 KP858933 Hernandez-Restrepo &

neoqueenslandicum al. 2016

M. nivale CBS 116205 KP859008 KP858944 Hernandez-Restrepo &

al. 2016

M. seminicola CBS 122707 KP859007 KP858943 Hernandez-Restrepo &

al. 2016

Microdochium sorghi CBS 691.96 KP859000 KP858936 Hernandez-Restrepo &

al. 2016

M. tainanense CBS 269.76 KP859009 KP858945 Hernandez-Restrepo &

al. 2016

M. trichocladiopsis CBS 623 77 KP858998 KP858934 Hernandez-Restrepo &

al. 2016

Neoidriella CBS 985.72 KP859048 KP858985 Hernandez-Restrepo &

desertorum al. 2016

Paraidriella jambosae CBS 374.90 KP859049 KP858986 Hernandez-Restrepo &

al. 2016

Poronia punctata CBS:656.78 KT281904 KY610496 Wendt & al. 2018

Rosellinia aquila MUCL 51703 KY610392 KY610460 Wendt & al. 2018

R. corticium MUCL 51693 KY610393 KY610461 Wendt & al. 2018

R. necatrix CBS 349.36 AY909001 KF7 19204 Pelaez & al. 2008

Sarcoxylon CBS 359.61 KT281903 KY610462 Wendt & al. 2018

compunctum

Selenodriella cubensis CBS 683.96 KP859053 KP858990 Hernandez-Restrepo &

al. 2016

S. fertilis CBS 772.83 KP859055 KP858992 Hernandez-Restrepo &

al. 2016

Xylaria arbuscula CBS:126415 KY610394 KY610463 Fournier & al. 2011;

Wendt & al. 2018

X. hypoxylon CBS 122620 KY610407 KY610495 Wendt & al. 2018

X.polymorpha MUCL 49884 KY610408 KY610464 Wendt & al. 2018

Phylogeny

The LSU-ITS dataset comprised 36 aligned sequences with 1482 positions.

Fic. 1 presents the ML tree including BS and PP values. The partition

homogeneity test showed no significant incongruence between the combined

data sets (p >0.05). Trees obtained from ML and Bayesian analysis of the

individual loci and the combined analysis produced congruent topologies.

Phylogenetic inferences grouped the three strains identified as Idriella

cubensis (MUCL 39017, CCMCIBE-H312, CCMCIBE-H320) together in a

506 ... Magdama & al.

92/1 - Microdochium citrinidiscum CBS 109067

Microdochium sorghi CBS 691 96

100/17 Microdochium seminicola CBS 122707

Microdochium albescens CBS 243 83

76/1 Wl Microdochium tainanense CBS 269 76

Microdochium trichocladiopsis CBS 623 77

89/1] |400/17 Microdochium majus CBS 741 79

Microdochium nivale CBS 116205

Microdochium neoqueenslandicum CBS 445 95

Microdochium lycopodinum CBS 109398

100/1 | Selenodriella fertilis CBS 772 83

Selenodriella cubensis CBS 683 96

-/0.96 Idriella lunata CBS 736 74

100/1 |- !driella lunata CBS 209 60

Idriella lunata CBS 204 56

100/1! Idriella lunata CBS 177 57

Neoidriella desertorum CBS 985 72

“j0.96| Uayaquilia cubensis sp. nov. CCMCIBE-H312_

_ | Guayaquilia cubensis sp. nov. CCMCIBE-H320

Paraidriella jambosae CBS 374 90

Idriellopsis uncinospora CBS 575 92

Castanediella acaciae CBS 139896

Castanediella couratarii CBS 579 71

Castanediella cagnizarii CBS 542 96

99/1 Castanediella cagnizarii CBS 101043

“ft Sarcoxylon compunctum CBS 359 61

Poronia punctata CBS 656 78

86/1

-/t

100/1 Rosellinia corticium MUCL 51693

J 96/0.98 Rosellinia aquila MUCL 51703

oe il Astrocystis concavispora IT1612

78/1 Rosellinia necatrix CBS 349 36

|i Xylaria polymorpha MUCL 49884

86/0.99 Xylaria hypoxylon CBS 122620

-/0.99 Kretzschmaria deusta CBS 163 93

Xylaria arbuscula KY610463

0.050

Fic. 1. Maximum Likelihood (ML) non-rooted tree inferred from the combined LSU and ITS

sequences of Guayaquilia cubensis and related species. Numbers on the branches are support values

>75% from BS and >0.95 from PP. The scale bar shows the expected changes per site.

supported clade and sister to Neoidriella desertorum, both clades separate from,

and basal to, the Microdochiaceae Hern.-Restr. & al. clade containing Idriella

lunata, Microdochium spp., and Selenodriella spp. We therefore propose a new

genus Guayaquilia typified by Idriella cubensis.

Taxonomy

Guayaquilia R.F. Castafieda, Magdama, D. Sosa & Hern.-Restr., gen. nov.

MB 831849

Differs from Neodriella and Paraidriella by its macronematous, tree-like, irregularly

multibranched conidiophores and its intercalary, discrete conidiogenous cells.

Guayaquilia cubensis gen. & sp. nov. (Cuba, Ecuador) ... 507

Type Species: Idriella cubensis R.F. Castahteda & G.R.W. Arnold [= Guayaquilia cubensis

(R.E Castafieda & G.R.W. Arnold) R.F. Castafeda & al.].

EryMo.Loey: Guayaquilia (Latin), referring to Guayaquil city, Ecuador.

ASEXUAL. COLONIES on natural substrate effuse, pulvinate-velutinous,

golden brown to brown. CONIDIOPHORES macronematous, mononematous,

fasciculate, erect, cylindrical, irregularly branched, septate, smooth, brown.

CONIDIOGENOUS CELLS polyblastic, denticulate, sympodial extended,

terminal or intercalary, cylindrical or slightly inflated, pale brown to

brown. Conidial secession schizolytic. Conip1a solitary, acropleurogenous,

sublunate, subnavicular, inequilateral, unicellular or septate, hyaline, smooth.

CHLAMYDOSPORES solitary, bicellular, brown and the apical cell globose, thick

walled, smooth.

Guayaquilia cubensis (R.F. Castafieda & G.R.W. Arnold) R.F. Castafieda, Magdama,

D. Sosa & Hern.-Restr., comb. nov. FIGS 2, 3

MB 831851

= Idriella cubensis R. F. Castaheda & G.R.W. Arnold, Rev.

Jard. Bot. Nac., Univ. Habana 6(1): 50 (1985).

COLONIES on V8 agar at 25°C attaining 60 mm diam after five days, floccose,

pale lavender grey to buff, sporulation poor, sparse. On CMA reaching

50 mm diam, felted, dark brick at the center, whitish toward the periphery.

CONIDIOPHORES macronematous, mononematous, tree-like, fasciculate,

profuse, dichotomously, alternately or irregularly branched toward the apex,

slightly inflated or bulbous at the base, smooth, brown below, pale brown to

subhyaline at the tip of branches, <500 um long, 4-7.5 um wide at the base,

branches 2-3.5 um wide. CONIDIOGENOUS CELLS polyblastic, denticulate,

intercalary and terminal, in branches integrated, cylindrical, or slightly

curved, brown to pale brown or subhyaline, 8-25 x 2-3 um. CONIDIA

solitary, acropleurogenous, sublunate, subnavicular, lunate, inequilateral, (0-)

l-septate, 14-17 x 2.5-3 um. CHLAMyDospoRESs bicellular, the basal cells

subcampanulate, subcuneiform to hemispherical, 5-8 um wide, and the apical

cells globose, 18-22 um diam, brown, thick walled; both cells with lumina

granulose, brown; terminal, smooth, arising from assimilative hyphae.

SPECIMENS EXAMINED: ECUADOR, Guayas PROVINCE, Naranjal, 2°48’S 79°40’W,

on decaying leaves of Theobroma cacao L. (Malvaceae), 8 July 2017, F. Espinoza & S.

Pérez-Martinez (URM 91815 = CCMCIBE-H312; GenBank MH777025, MH777024);

2°41’S 79°36’W, on decaying leaves of Theobroma cacao, 8 July 2017, F. Espinoza & S.

Pérez-Martinez (URM 91815a = CCMCIBE-H320; GenBank MH777026, MH777023).

CUBA, La HABANA PROVINCE, Santiago de Las Vegas, 22°58’N 82°22’W, on decaying

leaves of Calophyllum calaba L. (Clusiaceae), 29 Jun 1983, RF Castaneda Ruiz (holotype,

508 ... Magdama & al.

50pm

50 um

Fic. 2. Guayaquilia cubensis (holotype, INIFAT C83/57-1).

Conidiophores, conidiogenous cells, and conidia.

Guayaquilia cubensis gen. & sp. nov. (Cuba, Ecuador) ... 509

C, D. Conidiophores; E-G. Conidiogenous cells; H-L. Conidia; M, N, Chlamydospores. Scale

bars = 10 um.

INIFAT C83/57-1); PINAR DEL R10 PROVINCE, Los Portales, 22°40’N 83°28’W, on

decaying leaves of Cupania sp. (Sapindaceae), 5 Feb. 1994, RF Castafieda Ruiz, (INIFAT

C94/27 = MUCL 39017).

510 ... Magdama & al.

Note: Guayaquilia shares some morphological characters with the genera

Castanediella, Idriella, Idriellopsis, Neoidriella, and Paraidriella, such as blastic

conidial ontogeny and denticulate conidiogenous loci, but the tree-like,

dichotomously, alternately or irregularly profuse branched conidiophores with

intercalary and terminal conidiogenous cells are present only in Guayaquilia.

Also, the bicellular, globose, brown chlamydospores that arise from assimilative

hyphae in Guayaquilia are another distinctive character. The conidiophore

morphology in Guayaquilia resembles that of some Phaeodactylium species

(P. biseptatum, P. curvularioides, and P. stadleri) described by Castafeda-

Ruiz & al. (2009, 2013) and Matsushima (1980), but these taxa have clavate,

ellipsoidal, to obovoid, brown to subhyaline conidia.

Acknowledgments

We are indebted to Dr. Flavia Rodrigues Barbosa (Instituto de Ciéncias Naturais,

Humanas e Sociais, Universidade Federal de Mato Grosso, Brazil) and Dr. De-Wei Li

(The Connecticut Agricultural Experiment Station Valley Laboratory, USA) for their

critical reviews. The authors are grateful to Escuela Superior 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. P.M. Kirk (Index Fungorum) and Dr. K. Bensch (MycoBank).

Dr. Lorelei Norvell’s editorial review and Dr. Shaun Pennycook’s nomenclature review

are greatly appreciated.

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MYCOTAXON

July-September 2020—Volume 135, pp. 513-523

https://doi.org/10.5248/135.513

Distoseptispora longispora sp. nov.

from freshwater habitats in China

Hal-YAN SONG’, ALY FARAG EL SHEIKHA”?*>*, ZHI-JUN ZHADL,

JIAN-PING ZHOU’, MING-HuI CHEN’, GUANG-Hua Huo},

X1-GEN HuANG?®, Dian-MInG Hu”?

' Key Laboratory of Crop Physiology, Ecology and Genetic Breeding,

Ministry of Education of the P. R. China,

? Bioengineering and Technological Research Centre for Edible and Medicinal Fungi,

’ Jiangxi Key Laboratory for Conservation and Utilization of Fungal Resources, and

* College of Bioscience and Bioengineering,

™4 Jiangxi Agricultural University, 1101 Zhimin Road, Nanchang, Jiangxi 330045, China

° Department of Food Science and Technology, Minufiya University,

Shibin El Kom, Minufiya Government, Egypt

“ CORRESPONDENCE TO: * elsheikha_aly@yahoo.com

®=hxg208@163.com “hudianming1@163.com

ABSTRACT—A novel species, Distoseptispora longispora, from submerged wood in Yunnan

Province, China, is described and illustrated. The fungus is characterized by macronematous,

mononematous, septate conidiophores with monoblastic, integrated, determinate, terminal

conidiogenous cells that produce acrogenous, solitary, obclavate, elongated, distoseptate

conidia. SSU-ITS-LSU sequence analyses were used to infer the phylogenetic relationship

between D. longispora and related species. Both molecular analyses and morphological

data well support D. longispora as an independent taxon. A key to Distoseptispora species is

provided.

KEY worDs—Ascomycota, asexual fungi, Distoseptisporaceae, hyphomycetes

Introduction

Distoseptispora K.D. Hyde & al. was introduced by Su & al. (2016) to

accommodate two species, D. fluminicola McKenzie & al., the type species,

514 ... Song & al.

and D. aquatica Z.L. Luo & al. The genus is characterized by hairy colonies,

macronematous conidiophores, monoblastic conidiogenous cells, and

distoseptate acrogenous cylindrical conidia., Distoseptispora represents

a phylogenetically distinct lineage in Sordariomycetes for which Su & al.

(2016) proposed a new family, Distoseptisporaceae K.D. Hyde & McKenzie.

Subsequently, 13 more species have been added: D. cangshanensis Z.L. Luo

& al., D. dehongensis W. Dong & al., D. guttulata Jing Yang & K.D. Hyde,

D. martinii (J.L. Crane & Dumont) J.W. Xia & X.G. Zhang, D. obpyriformis

Z.L. Luo & H.Y. Su, D. palmarum S.N. Zhang & al., D. phangngaensis Jing Yang

& al., D. rostrata Z.L. Luo & al., D. submersa Z.L. Luo & al., D. suoluoensis Jing

Yang & al., D. thailandica Tibpromma & K.D. Hyde, D. thysanolaenae Goonas.

& al., and D. xishuangbannaensis Tibpromma & K.D. Hyde (Hyde & al. 2019,

Luo & al. 2018, Phookamsak & al. 2019, Tibpromma & al 2018, Xia & al. 2017,

Yang & al. 2018).

Many aquatic fungi have been discovered in China’s freshwater habitats,

which are rich in lignicolous fungi (Hu & al. 2007, 2012a,b; Huang & al. 2016;

Song & al. 2018). Our ongoing survey of aquatic fungi in Yunnan Province

revealed an interesting Distoseptispora species that we describe as new based on

morphological characters and molecular analyses (SSU-ITS-LSU).

Materials & methods

Sample collection & morphological studies

Debarked wood samples from different freshwater habitats in Yunnan Province,

China, were taken to the lab for detection of fungi using dissecting and compound

microscopes. The dried specimens were deposited in the Herbarium of Fungi, Jiangxi

Agricultural University, Nanchang, China (HFJAU).

DNA extraction, sequencing, and phylogenetic analyses

Genomic DNA was extracted from fresh fungal materials found on the natural

substrate following Hu & al. (2012c). We amplified three rDNA regions: internal

TABLE 1. Isolates included in the phylogenetic analyses.

GENBANK ACCESSION NUMBERS

SPECIES STRAIN

ITS LSU SSU

Distoseptispora “adscendens” HKUCC 10820 — DQ408561 —

D. aquatica MFLUCC 15-0374" NR_ 154040 KU376268 —

SNJ14 MH555360 — a

D. fluminicola MFLUCC 15-0417 =NR_154041 KU376270 —

D. guttulata MFLUCC 16-0183 MF077543 MF077554 MF077532

D. “leonensis”

D. longispora

D. martinii

D. multiseptata

D. phangngaensis

D. suoluoensis

D. tectonae

D. tectonigena

Distoseptispora sp.

Dothidea sambuci

Ellisembia bambusicola

E. calyptrata

E. minigelatinosa

Ellisembia sp.

Sporidesmium aquaticivaginatum

S. aquaticum

S. australiense

S. fluminicola

S. macrurum

S. obclavatulum

S. olivaceoconidium

S. pachyanthicola

S. parvum

S. pyriformatum

S. submersum

S. tengii

S. thailandense

S. tropicale

Sporidesmium sp.

Distoseptispora longispora sp. nov. (China) ... 515

HKUCC 10822

HFJAU 0705"

HSAUP myr4280

MFLUCC 15-0609"

MFLUCC 16-1044

MFLUCC 16-0857

MFLUCC 17-1305

MFLUCC 17-0224

MFLUCC 12-0291

GZ25

MFLUCC 12-0292

HLXM 15-1

AFTOL-ID 274

HKUCC 3578

HKUCC 10821

NN47497

HKUCC 10558

MFLUCC 15-0624

MFLUCC 15-0420

HKUCC 10833

MFLUCC 15-0346

HKUCC 2740

HKUCC 10834

MFLUCC 15-0380

HKUCC 10835

HKUCC 10836

MFLUCC 15-06207

MFLUCC 15-0627

MFLUCC 15-0421

HKUCC 10837

MFLUCC 15-0964"

MFLUCC 16-0185

HKUCC 10838

KUFA 0043

MFLUCC 15-0617

MFLUCC 16-0186

MH555359

KU999975

NR_154017

MF077544

MF077545

MF077547

MF077546

KX751711

MH555361

NR_154018

DQ491505

KX710147

KX710144

KX710146

KX710148

MF374361

MF077551

KT824799

MF077550

MF077549

DQ408566

MH555431

KX033566

KX710140

MF077555

MF077556

MF077558

MF077557

KX751713

KX751714

KU376269

AY544681

DQ408562

DQ408564

DQ408567

DQ408565

KX710142

KU376273

DQ408554

KU376271

DQ408555

DQ408556

KX710139

DQ408557

DQ408558

KX710141

KU376272

DQ408559

MF374370

MF077562

DQ408560

MF077561

MF077560

MH555357

KX033537

MF077540

MF077533

MF077534

MF077536

MF077535

MH555358

AY544722

MF077541

MF077542

KX710143

MF077539

MF077538

516 ... Song & al.

transcribed spacer (ITS) with the primer pair ITS1 & ITS4 (White & al. 1990),

partial small subunit (SSU) with NS1 & NS4 (White & al. 1990), and large subunit

(LSU) with LROR & LR6 (Vilgalys & Hester 1990, Rehner & Samuels 1995). PCR

protocols followed the conditions set by Hu & al. (2012b). The PCR products were

purified and sequenced by the same primers used for PCR at Sangon Biotech

(Shanghai) Co. Ltd.

In this study, we generated six novel sequences (MH555360, MH555359,

MH555431, MH555357, MH555361 & MH555358) and retrieved 36 reference

sequences from GenBank (TABLE 1) that were aligned using MAFFT v7.

The ML analyses were produced with RAxML v7.2.6 (Stamatakis & Alachiotis

2010) using a GTRGAMMA substitution model with 1000 bootstrap replicates and

evaluated by bootstrap support (MLBS).

The best-fit evolutionary models were estimated using MrModeltest V2.2.

Posterior probabilities (PP) (Zhaxybayeva & Gogarten 2002) were determined by

Markov chain Monte Carlo sampling (BMCMC) in MrBayes 3.0b4 (Huelsenbeck &

Ronquist 2001). Six simultaneous Markov chains were run 1,500,000 generations,

with trees sampled every 100 generations and the first 25% deleted as burn-

in. Posterior probabilities were calculated based on the remaining trees. The

novel taxonomic descriptions and nomenclature were deposited in MycoBank

(http://www.mycobank.org/).

Phylogenetic results

The six new sequences generated in our study included one LSU, two SSU,

and three ITS sequences. The phylogenetic tree based on the three-locus

analysis (Fic. 1) shows the relationship between the new taxon and other

related species. The concatenated aligned dataset comprised 17 isolates from

13 Distoseptispora spp., 23 isolates from 21 Sporidesmium (or “Ellisembia”)

spp., and one isolate of Dothidea sambuci (Pers.) Fr. (Dothideomycetes) as an

outgroup. The dataset including alignment gaps comprised 2817 characters:

1348 for SSU, 629 for ITS, and 840 for LSU. The combined dataset ML tree

(Fic. 1) with bootstrap support values (MLBS) and Bayesian posterior

probabilities (BPP) indicates several well-supported clades, with the

Distoseptispora longispora strain forming a well-supported clade (MLBS = 94%,

BPP = 0.68) with other Distoseptispora species.

Taxonomy

Distoseptispora longispora H.Y. Song & D.M. Hu sp. nov. Fic. 2

MB 826916

Differs from Distoseptispora aquatica by its larger and more septate conidia; and from

D. tectonae and D. tectonigena by its smooth and brown to yellowish brown conidia.

Distoseptispora longispora sp. nov. (China) ... 517

66/0.99 Distoseptispora tectonae MFLUCC 12-0291

60/0.98|! nistoseptispora tectonae GZ25

Distoseptispora tectonigena MFLUCC 12-0292?

Distoseptispora “adscendens” HKUCC 10820

79/0.96: Distoseptispora fluminicola MFLUCC 15-0417?

Distoseptispora aquatica SNJ14

abet Li Dinseviees basis MFLUCC 15-0374?

73/0.76| | Distoseptispora longispora HFJAU 0705t

v1 Distoseptispora sp. HLXM 15-1

65/0.68 Distoseptispora multiseptata MFLUCC 15-0609T

Distoseptispora multiseptata MFLUCC 16-1044

98/0.67 Distoseptispora phangngaensis MFLUCC 16-0857

Distoseptispora martinii HSAUP myr4280

Distoseptispora guttulataMFLUCC 16-0183

94/0.68] | 100/1)Distoseptispora suoluoensis MFLUCC 17-0224

Distoseptispora suoluoensis MFLUCC 17-1305

63/-

PONS Distoseptispora “leonensis” HKUCC 10822

100/1 >- Sporidesmium olivaceoconidium MFLU! oC

Sporidesmium aquaticivaginatum MI FLUCC

99/0.94 Sporidesmium pyrif

100/1 iE 1! 'Sporidesmium pyriformat,

100/1

93/0.73 ‘Sporidesmiine a c

100/0.85 96/1 =

Sl Sporidesmium sp. MFLUCC 15-0

100/1 Sporidesmium thailandense MFLU

87/0.94, Sporidesmium submersum MFLU u

/0. Ellisembia minigelatinosa NN47497

100/0.93 99/0. Br ON zitisembia bambusicola HKUCC 3578

Ellisembia sp. HKUCC 10558

S Sporidesmium aquaticum MFLUCC 15-0420

100/1/ Sporidesmium tropicale HKUCC 10838

100/0.99 Sporidesmium tropicale MFLUCC 16-0185

64/0.8. Sporidesmium macrurum HKUCC 2740

100/1 Sporidesmium sp. MFLUCC 16-0186

-/67 Ellisembia calyptrata HKUCC 10821

Sporidesmium sp. KUFA 0043

99/1 Sporidesmium tengii HKUCC 10837

100/1 Sporidesmium australiense HKUCC 10833

Sporidesmium obclavatulum HKUCC 10834

Sporidesmium pachyanthicola HKUCC 10835

Dothidea sambuci AFTOL-ID 274

0.1

Fic. 1. Maximum Likelihood (ML) phylogenetic tree of Distoseptispora longispora and allied

species based on concatenated SSU-ITS-LSU data. The RAxML bootstrap support values

(MLBS) and Bayesian posterior probabilities (BPP) are given at the nodes (MLBS/BPP).

Type: China, Yunnan Province: Mengla, a small stream, on submerged wood, 13 Apr

2017, H.Y. Song (Holotype, HFJAU 0705).

EtyMo.oey: longispora, referring to the long conidia of the fungus.

COLONIES on submerged wood superficial, effuse, scattered, hairy, brown to

dark brown. Mycelium mostly immersed, composed of branched, septate,

smooth, subhyaline to pale brown hyphae. CONIDIOPHORES macronematous,

mononematous, 2-4-septate, unbranched, single, straight or slightly

flexuous, erect, cylindrical, smooth, brown to dark brown, 17-37 x 6-10 um.

CONIDIOGENOUS CELLS monoblastic, integrated, determinate, terminal, light

brown to dark brown, cylindrical. Conidial secession schizolytic. CoNIDIA

acrogenous, solitary, dry, obclavate, elongated, straight or slightly curved,

rounded at the apex, truncate at the base, 31—56-distoseptate, smooth, brown

518 ... Song & al.

to yellowish brown, slightly paler towards the apex, 189-297 x 16-23 um

(mean = 204 x 21 um, n= 50).

ADDITIONAL SPECIMEN EXAMINED: CHINA, YUNNAN PROVINCE, Mengla, a small

stream, on submerged wood, 13 Apr 2017, H.Y. Song (HFJAU 0706).

Notes: Distoseptispora longispora is characterized by its macronematous,

mononematous, septate conidiophores, monoblastic, integrated, determinate,

terminal conidiogenous cells, and acrogenous, solitary, dry, obclavate,

elongated, distoseptate conidia, which fit well with the generic concepts of

Distoseptispora (Su & al. 2016). The molecular phylogeny (Fic. 1) clusters the

D. longispora strain well within the Distoseptispora clade and closely related

to D. aquatica but with an ITS six base pair (bp) difference separating it

from D. aquatica (MFLUCC 15-0374). Distoseptispora longispora resembles

D. aquatica, D. tectonae, and D. tectonigena in conidial shape. However, its

larger and more septate conidia separate D. longispora from D. aquatica

(189-297 x 16-23 um; 15-28 septa). In addition, D. tectonae has verruculose,

dark reddish conidia, and D. tectonigena has dark reddish-brown conidia

(Hyde & al. 2016).

The multilocus molecular phylogeny and morphological data fully support

separation of our new species, D. longispora. Morphologically, Distoseptispora

resembles Ellisembia Subram., first established to accommodate the species

with distoseptate conidia in Sporidesmium Link (Subramanian 1992).

However, a molecular phylogeny by Shenoy & al. (2006) showed that Ellisembia

and Sporidesmium were polyphyletic. Su & al. (2016) later clarified differences

between Ellisembia and Sporidesmium, noting that euseptate/distoseptate

characters were not supported by molecular evidence and treating Ellisembia

as a synonym of Sporidesmium. Distoseptispora species differ from Ellisembia

(Sporidesmium) species in having relatively short conidiophores and darker

conidia with slightly paler, but not hyaline, rounded apices, basal cells cut off

by cross walls, and of indeterminate length (Su & al. 2016).

There are various hyphomycetes in freshwater habitats, and several studies

have reported Distoseptispora species, e.g., D. aquatica, D. fluminicola,

D. guttulata, D. multiseptata, D. phangngaensis, and D. suoluoensis (Hu &

al. 2007, 2010a,b; Huang & al. 2018; Hyde & al. 2016; Su & al. 2016; Yang

& al. 2018). Most of these species were identified based on morphology,

which proved to be insufficient for accurate identification (Hu & al. 2017).

Numerous other studies (Réblova & al. 2016, Su & al. 2016, Zhang & al. 2017)

have revealed many different freshwater lineages using molecular analyses.

Distoseptispora longispora sp. nov. (China) ... 519

d b

Fic. 2. Distoseptispora longispora (holotype, HFJAU 0705). a. Colonies on submerged wood;

b, c. Conidia; d. Conidiophore. Scale bars: a = 100 um, b-d = 5 um.

520 ... Song & al.

Key to Distoseptispora species

1. Conidia muriform, having both longitudinal and transverse septa ...... D. martinii

b-@oniciawathy Only transverse septa: 4) ef aia n he Seed ert bs ae natok Sue hes 2

PAC OMIA CUSe Plate .cA Ars Hs AMS sR tee ode gt AE Tee ppc i oan: ood aPag ET NR OVS CP 3

2 COniatacdistosep tate. 7. Reed oe AR ER Ne He see hy wasn aeale pS Ree ea 4

3. Conidia 75-130 x 7-11 um, 11-14(-20)-euseptate, smooth ........... D. guttulata

3. Conidia 80-125 x 8-13 um, 8-10-euseptate, verrucose............. D. suoluoensis

4, Conidiogenous cells polyblastic, conidia 35-180 x 7-ll um ......... D. palmarum

4. Conidiogenous:cellsimonoblastic «5, ose ages axis sean nied omens Panaee ken 5

5. Conidiophores having percurrently proliferating conidiogenous

cellsatthe apex Or comidiophoress:,. <. eactcmtics aha sate Ke sent teen sede oP Seek 6

5 Conidiophores Ot AG ADOVE Bi sts remiaginw Hits te aca hbo aaitess ertticen peta Sabghomem ae 7

6; Conidial PaaS 5 8 Orr pas An. Se, Scene ae tl Rd, Bre mle AN al D. rostrata

Gs ConidianAs 22 51 az ties sea tetiaeale Sat wants. eee D. tectonigena

7 @onidiaverruculoses7..5=9:5-X SiS SYM 25s eens eke aod ete alae os D. tectonae

Fe LOT Cia NOt eTTUCULOSE Pic etn)... Aas 5) ARN oe Sng. cats stata tar Ate so IRB. 8

8. Conidia 3-5-distoseptate, 17-30 x 7.5-10 um................004. D. dehongensis

8. Gonidia with more-than-6. distosepta: «oe. .5 da.04 wedeae vee ee ae cea 9

9. Saprobic on submerged wood in freshwater habitats ...................0000-% 12

9. Not saprobic on submerged wood in freshwater habitats...................-. 10

10. Saprobic on dead culms of Thysanolaena maxima,

conidia 8-14-distoseptate, 21.5-80 x 6.5-12.8 um ........... D. thysanolaenae

10Saprobic-on-deadsleaves*ot Pandanussp. bchie aiuto ag ths to ethuseen nore hoh wht ie 11

11. Conidia reddish brown to brown, 130-230 x 13.5-17 um ......... D. thailandica

11. Conidia green-brown to brown, 160-305 x 8-15 um...... D. xishuangbannaensis

12. Conidia 9-11-distoseptate, 53-71 x 12-16um .................. D. obpyriformis

12. Comidia with-morediant 12 aistos ep ta gets ssi cists sis ae todays aloe fel tte tess 13

13. Conidiogenous cells yellow, conidia 125-250 x 13-15 um ......... D. fluminicola

13¥Gonidiogenous cells: brownish: 22.0202 42-02-50u nessa ee Qo ueeee ee ou eeeeeees 14

14. Conidia brown to yellowish brown, 31-56-distoseptate,

1B oe Pde eo Ad CePA on Ae Roa ee a ai D. longispora

14. Conidia greenish brown or dark olivaceous green ............. 0.0 c cece eee 15

15. Conidiogenous cells subhyaline to pale brown,

conidia 58=1665¢ 10-14 yiml e255 toe tsa pee, Pee ed ee D. cangshanensis

15. Conidiogenous cells brown to dark brown ............ 0.00. eee eee eee eee 16

16. Conidiogenous cells dark brown to olivaceous,

conidia 110-157" 13:5.51:625 im" * 4 254 ak. 7 5 eis ef eal D. aquatica

16.-Gonidiogenious cells brown oo... 4h oi. gee eee ete a ede te deen ce dae 17

Distoseptispora longispora sp. nov. (China) ... 521

17. Conidia dark olivaceous green, multi-distoseptate,

99 = 29096 TP ONE em ae ERS Sin Ss ike ced eS ad D. multiseptata

17. Conidia dark or mid olivaceous to mid or dark brown ...................04- 18

18. Conidia dark olivaceous to mid or dark brown,

16553507% 14 S19 spirrn Vaan ae VR Brn tae eNe Bote Ee, ese ae D. phangngaensis

18. Conidia mid-olivaceous to brown, 95-123 x15-19 um .............. D. submersa

Acknowledgments

The authors express their sincere gratitude to Dr. Rafael EF Castafieda-Ruiz

(Alejandro de Humboldt—INIFAT, Havana, Cuba) and Dr. Jian Ma (College of

Agronomy, Jiangxi Agricultural University, Nanchang, China) for their critical

review of the manuscript, to Dr. Shaun Pennycook for nomenclatural review, and

to Dr. Lorelei L. Norvell for editorial review. Funds for research were provided

by the National Natural Science Foundation of China (NSFC 32070023), the Key

Projects of Youth Fund of Jiangxi Science and Technology Department of China

(20192ACBL21017), Natural Science Foundation of Education Department of

Jiangxi Province of China (GJJ190168).

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MYCOTAXON

July-September 2020—Volume 135, pp. 525-534

https://doi.org/10.5248/135.525

Notes on rust fungi in China 9. Puccinia miscanthi

life cycle and morphology confirmed by inoculation

JING-XIN J’*, ZHUANG LY’, Yu Lt, MAKOTO KAKISHIMA??®

‘Engineering Research Center of Chinese Ministry of Education for Edible & Medicinal Fungi,

Jilin Agricultural University, Changchun, Jilin 130118, China

College of Plant Protection, Shandong Agricultural University, Taian 271000, China

>University of Tsukuba, Tsukuba, Ibaraki 305-8572, Japan

* CORRESPONDENCE TO: * 1096314395@qq.com * kakishima.makoto.ga@u.tsukuba.ac.jp

ABSTRACT— The Heteroecious and macrocyclic life cycle of Puccinia miscanthi is confirmed

for the first time with inoculation experiments in China. The rust produces spermogonial and

aecial stages on Plantago asiatica and uredinial and telial stages on Miscanthus sacchariflorus.

Morphological characters of these stages are described based on field collections and

specimens that confirmed their life cycle connections. The neotype of P miscanthi is

designated.

Key worps—Plantaginaceae, plant disease, Poaceae, Pucciniales, Uredinales

Introduction

Puccinia miscanthi was first described with uredinial and telial stages on

Miscanthus sacchariflorus (Poaceae) collected in northeastern China (Miura

1928). The life cycle of P. miscanthi was reported in Japan by Hiratsuka (1933),

Ito (1934), and Asuyama (1936) under the name of P. eulaliae Barclay. These

authors demonstrated life cycle connection between spermogonial and aecial

stages on Plantago asiatica (Plantaginaceae) and uredinial and telial stages on

M. sinensis Andersson through inoculations. Ito (1950) considered the rust

fungus as producing spermogonial and aecial stages on Plantago and uredinial

and telial stages on Miscanthus and Imperata (as P. eulaliae) and treated

P. miscanthi as a synonym of this species. Later, however, this rust fungus

526 ... Ji & al.

was considered to represent a species different from P. eulaliae because of

morphology and host relations, and it was treated as P. miscanthi (Cummins

1971, Hiratsuka & al. 1992). Sato & Kakishima (1982) reported Lysimachia

clethroides Duby (Primulaceae) as an additional host of the rust’s spermogonial

and aecial stages. This rust fungus has been reported on wide range of host

plants from northeastern Asia (TABLE 1, p. 532).

During investigation of rust fungi in northeastern China, a Puccinia species

producing uredinial and telial stages on M. sacchariflorus was collected in

Heilongjiang Province. Spermogonial and aecial stages of a rust fungus on

Plantago asiatica were also found near infected M. sacchariflorus plants. The

spermogonial and aecial stages were suspected to represent different stages

of Puccinia miscanthi. However, life cycle connection of P miscanthi between

different host plants has not been clarified in China, although PI. asiatica,

Pl. depressa Willd., and Pl. major L. have all been listed as spermogonial

and aecial host plants, and M. floridulus (Labill.) K. Schum. & Lauterb.,

M. sacchariflorus, M. sinensis, Saccharum spontaneum L., and Thysanolaena

maxima (Roxb.) Kuntze as uredinial and telial hosts (Wang & Zhuang 1998,

Cao & Li 1999). Furthermore, Puccinia cynodontis Lacroix ex Desm. also

has been reported to produce spermogonia and aecia on Plantago species

including Pl. asiatica (Cummins 1971, Wang & Zhuang 1998). Worldwide,

uredinial and telial stages of six morphologically similar Puccinia species

have been recorded on Miscanthus (TABLE 1; Cummins 1971, Hiratsuka &

al. 1992, Wang & Zhuang 1998). Among them, P. miscanthi, P. miscanthicola

EL. Tai & C.C. Cheo, P. melanocephala Syd. & P. Syd., and P. erythropus Dietel

have been recorded in China (Tai 1979, Wang & Zhuang 1998). Therefore,

we carried out inoculation experiments to clarify the life cycle connection

between spermogonia and aecia on PI. asiatica and uredinia and telia on

M. sacchariflorus collected in Heilongjiang Province on 2 July 2018. We also

observed morphology of these specimens for identification of species.

Materials & methods

Inoculation experiments

On 2 July 2018 uredinia and telia on Miscanthus sacchariflorus were found at

Wuchang, Heilongjiang Province, China (44°12’22”N 128°03’33”E). Infected plants

were transplanted in plastic pots and placed on the ground at the campus of Jilin

Agricultural University, Changchun, Jilin Province. In late summer, abundant telia

appeared on the plants, and these telia were overwintered on dead leaves. Teliospores

obtained from the telia were observed to germinate and basidiospores produced in

the spring of 2019 were used for inoculations. On 17 June 2019 small pieces of leaves

Puccinia miscanthi life cycle (China) ... 527

- P

XG)

te!

Fic. 1. Puccinia miscanthi on Plantago asiatica (A, C, E) and Miscanthus sacchariflorus (B, D, F).

A. The plants in plastic pots, producing spermogonia and aecia by basidiospore inoculation.

B. Plant transplanted from Heilongjiang Province and cultured at campus of Jilin Agricultural

University. C. Spermogonia produced on leaf surface by basidiospore inoculation. D. Brown

uredinia produced on leaf surface by aeciospore inoculation. E. Aecia on lower leaf surface

produced by basidiospore inoculation. F. Dark brown telia on leaf.

with teliospores were attached to the surfaces of young healthy leaves of Pl. asiatica

for inoculation, using a similar method previously reported by Ji & al. (2017a,b).

The inoculated plants were collected at campus of Jilin Agricultural University and

had been maintained in plastic pots. No natural infection of the field plants had

528 ... Ji&al.

been confirmed prior to inoculation. The inoculated plants were kept in a humid

environment in a plastic box in darkness at 20-23 °C for 3 days and then transferred

to a place near windows at 20-23 °C for observation.

Aeciospores obtained from aecia on the basidiospore-inoculated Plantago asiatica

leaves were dusted onto small pieces of wet filter paper (about 3 mm?’) in a Petri dish.

These papers were then placed on young healthy leaves of M. sacchariflorus, which

had been transplanted from Wuchang, Heilongjiang Province in 2018 and maintained

in plastic pots at campus of Jilin Agricultural University. Plants with new leaves were

used for the inoculation. The inoculated plants were kept in a humid environment in

a plastic box under the same conditions as the basidiospore inoculations, and later

transferred to the place near windows.

Morphological observations

Specimens collected at Wuchang, and specimens obtained by inoculations were

morphologically examined. The spores and size and shape of sori were microscopically

examined using Light (LM) and scanning electron (SEM) microscopes according

to Ji & al. (2019). The specimens are deposited in the Herbarium of Mycology,

Engineering Research Center of Chinese Ministry of Education for Edible and

Medicinal Fungi, Jilin Agricultural University, Changchun, China (HMJAU).

Results & discussion

Life cycle

About 10 days after inoculation of Plantago asiatica with basidiospores from

germinating teliospores, small yellow spots of spermogonia appeared on both

surfaces of the inoculated leaves (Fics 1A, C). These spots were frequently

observed on the leaf, and the leaf tissue around infected veins was sometimes

malformed when many spots were produced. About 10 to 12 days later, orange-

yellow aecia were produced around the spots on lower surfaces of the leaves

(Fic. 1E).

About 5 days after aeciospore inoculation, pale yellow spots appeared on

the leaves of Miscanthus. sacchariflorus, after which then brown uredinia were

produced (Fic. 1D). One to two months after that, blackish telia appeared on

M. sacchariflorus (Fic. 1F). The inoculation results confirmed that the rust

fungus producing spermogonial and aecial stages on PI. asiatica was conspecific

with the rust fungus producing uredinial and telial stages on M. sacchariflorus.

Morphology & identification

Theinoculations show that this rust fungus has an heteromacrocycliclife cycle,

producing spermogonia and aecia on Plantago asiatica and uredinia and telia

on Micanthus sacchariflorus. The uredinial and telial stages of our Heilongjiang

Province rust on M. sacchariflorus are morphologically similar to those of

Puccinia miscanthi life cycle (China) ... 529

ore

seed fl

—-

Fic. 2. Puccinia miscanthi on Plantago asiatica (HMJAU8642: A, B, D) and Miscanthus sacchariflorus

(neotype, HMJAU8643: C, E, FE, G) observed by LM. A. Vertical section of spermogonium (type 4).

B. Aeciospores with verrucose surface. C. Urediniospores with equatorial germ pores. D. Vertical

section of aecium with catenulate aeciospores surrounded by peridium. E. Vertical section of

uredinium mixed with paraphyses. F. Teliospores. G. Vertical section of telia, surrounded by host

epidermis. Scale bars: A, C = 30 um; B, E = 20 um; D = 100 um; F G = 25 um.

Puccinia erythropus, P. melanocephala, and P. miscanthi in urediniospore and

teliospore shapes. However, teliospores of P. erythropus and P. melanocephala

530 ... Ji&al.

are shorter, and their apical walls thinner, than those of our M. sacchariflorus

rust fungus (TABLE 1). Furthermore, although Ito (1950) described paraphyses

(TABLE 1), P. erythropus has been reported to have no paraphysis in uredinia,

whereas, our M. sacchariflorus rust fungus produces abundant paraphyses (Fics

2E, 3D). The morphological characters of our M. sacchariflorus rust fungus are

very close to those previously reported for P miscanthi, but as its teliospores are

only slightly shorter than these descriptions (TABLE 1), we identified our rust

fungus as P. miscanthi. Host range and morphological features of P. miscanthi

reported by different authors as shown in TABLE 1; the differences may be

considered variations within species caused by host plants and geographical

distribution. Puccinia miscanthi has been reported to be genetically very close

to P melanocephala although they are morphologically different (Virtudazo

& al. 2001, Dixon & al. 2010). Phylogenetic analyses with specimens from

different regions and host plants may clarify their relationships.

Spermogonial and aecial stages of Puccinia miscanthi have been reported to

be produced on Plantago and Lysimachia species (Cummins 1971, Hiratsuka

& al. 1992, Wang & Zhuang 1998). However, no confirmation of these stages

of P. miscanthi has been reported in China, although its life cycle connection

was reported in Japan with inoculations. Therefore, Pl. asiatica is the first

host plant confirmed by inoculations of its spermogonial and aecial stages in

China. The morphology of these stages is almost identical with the descriptions

reported from Japan by Ito (1950), Sato & Kakishima (1982), and Hiratsuka &

al. (1992), except for the slightly smaller aeciospores. Wang & Zhuang (1998)

described the rust’s spermogonial and aecial stages on Pl. depressa in China;

however, a connection to its uredinial and telial stages was not confirmed.

Two Plantago species, Pl. asiatica and Pl. lanceolata L., were also reported as

spermogonial and aecial host plants of P. cynodontis without confirmation of

their life cycle connections in China (Wang & Zhuang 1998). Therefore, rust

species identifications of spermogonial and aecial stages on species of Plantago

should be reconsidered. Ono & Azbukina (1997) reported that P. erythropus,

morphologically similar to P miscanthi, produced spermogonia and aecia

on Cynanchum sublanceolatum var. obtusum (Franch. & Sav.) Matsum.

(Apocynaceae), suggesting that this species differs from P miscanthi in its

spermogonial and aecial hosts.

Miura (1928) described Puccinia miscanthi based on a specimen on

Miscanthus sacchariflorus collected in Teikaton, Manchuria (northeast China)

on 22 September in 1919. However, this specimen has not been found in any

herbarium, we therefore designate a neotype. We collected specimens on the

Puccinia miscanthi life cycle (China) ... 531

Fic. 3. Puccinia miscanthi on Plantago asiatica (HMJAU8642: A, B) and Miscanthus sacchariflorus

(neotype, HMJAU8643: C, D) observed by SEM. A. Aeciospores with densely verrucose surface.

B. Aecium with peridium. C. Urediniospore with echinulate surface. D. Uredinium mixed with

paraphyses. Scale bars: A, C = 5 um; B = 40 um; D = 30 um.

same host plant from the same area of China, as reported by Miura (1928) and

used for inoculations. We believe that specimens collected and obtained from

inoculations are morphologically close to the original specimen and describe

its morphology based on these specimens.

Puccinia miscanthi Miura,

Fl. Manch. E. Mong., III Cryptogams, Fungi: 302, 1928. Figs 1-3

Type: Uredinia and telia on Miscanthus sacchariflorus (Maxim.) Franch. (Poaceae),

China, Helongjiang Province, Wuchang, 9 September 2017, leg. M. Kakishima & J-X. Ji

(neotype designated here, HMJAU8643; MBT388544).

Spermogonia amphigenous, surrounded by greenish yellow lesions,

densely grouped, yellow, subepidermal, flask-shaped, type 4 of Cummins

& Hiratsuka (2003). Aecia hypophyllous, densely grouped, yellow, cupulate

with peridia, Aecidium-type. Aeciospores catenulate, globose to subglobose,

Ji & al.

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3 ‘Sgouasqy :— ‘souasarg :+ (7 ‘snAnjaovYyg :q ‘vuavjouvsdy], :], SWinsvyrIvG :S ‘snyguvosIPy 7 ‘vJosadwy :] ssnyguvLag vy (1

sndosyjasa g

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DIOINYJUDISIM J

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‘oy *S avyyjna q =|

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SaIOadS

‘dds snyjuvosip uo payiodas saisads viuioong Jo ela} pue erurpein jo A8ojoydiow saneseduroy “| ATAVI,

Puccinia miscanthi life cycle (China) ... 533

angular, 17.5-23.5 x 14.5-19.0 um (av. = 20.5 x 17. 0 um); walls hyaline,

0.6-1.2 um (av.= 0.9 um) thick, densely verrucose.

Uredinia mostly on abaxial surface, scattered or gregarious, yellow-

brown, subepidermal, erumpent, with abundant paraphyses. Paraphyses

capitate; walls hyaline, thicker at the apical parts. Urediniospores pedicellate,

obovoid to ellipsoid, 27.0-35.5 x 19.5-23.5 um (av. = 30.0 x 21.5 um); walls

brown, echinulate, 0.7-1.5 um (av. = 1.0 um) thick; germ pores mostly 4,

equatorial. Telia mostly on abaxial surface, blackish brown, erumpent,

pulvinate. Teliospores pedicellate, oblong-clavate, 39.5-56.5 x 14.0-22.0 um

(av. = 47.0 x 19.0 um); walls chestnut-brown, 0.5-1.5 um (av. = 1.0 um) thick

at sides, 2.5-6.0 um (av. 4.0 um) thick at apices, smooth; pedicels persistent,

brown, thick-walled, short.

ADDITIONAL SPECIMENS EXAMINED — CHINA, HEILONGJIANG PROVINCE, Wuhan:

Spermogonia and aecia on Plantago asiatica, 2 July 2018, (HMJAU8644); cultured at

Jilin Agricultural University, Changchun, 5 July 2019 (HMJAU8642). Uredinia and telia

on Miscanthus sacchariflorus, 2 July 2018 (HMJAU8645); cultured at Jilin Agricultural

University, Changchun, 5 July 2019 (HMJAU8646).

Acknowledgments

This work was financed by the Fungal Flora in Jilin Province (20130206073NY).

We wish to thank Dr E.H.C. McKenzie (Manaaki Whenua Landcare Research,

Auckland, New Zealand) and Prof. C.M. Denchev (Bulgarian Academy of Sciences,

Sofia, Bulgaria) for critical reading of the manuscript and suggestions.

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(Urediniales). Mycoscience 38: 281-286. https://doi.org/10.1007/BF02464085

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Beijing.

MYCOTAXON

July-September 2020—Volume 135, pp. 535-558

https://doi.org/10.5248/135.535

Biodiversity of heat-resistant ascomycetes

from semi-arid soils in Argentina

STELLA MARIS ROMERO??*, ANDREA IRENE ROMERO’”,

ALBERTO MIGUEL STCHIGEL?, ERNESTO RODRIGUEZ ANDRADE?,

VIVIANA ANDREA BARRERA‘, JOSE FRANCISCO CANO}, RICARDO COMERIO®

' Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales, DBBE,

Intendente Giiiraldes 2160, Pab. II Ciudad Universitaria, C1428EGA, Buenos Aires, Argentina

? CONICET-Universidad de Buenos Aires, Instituto de Micologia y Botanica (InMiBo),

Intendente Giiiraldes 2160, Pab. II Ciudad Universitaria, C1428EGA, Buenos Aires, Argentina

* Mycology Unit, Medical School and IISPV, Universitat Rovira i Virgili (URV),

Sant Lloreng 21, 43201 Reus, Tarragona, Spain

* Instituto Nacional de Tecnologia Agropecuaria (INTA), IMyZA-CICVyA,

N. Repetto y De los Reseros, CC25 1712, Castelar, Pcia. Buenos Aires, Argentina

° Instituto Nacional de Tecnologia Agropecuaria (INTA), EEA Anguil Ing. Agr. G. Covas,

Ruta Nacional N° 5, km 580, CC 11 6326 Anguil, Pcia. La Pampa, Argentina

* CORRESPONDENCE TO— smromero@conicet.gov.ar

ABSTRACT—Artificial thermal shock conducted on 50 soil samples from a semi-arid

geographic region (Catamarca and La Rioja provinces) in northern Argentina yielded 34

heat-resistant fungal strains. These strains were assigned to seventeen taxa in ten ascomycete

genera: Arthrinium (1), Aspergillus (3), Epicoccum (1), Gilmaniella (1), Hamigera (2),

Leiothecium (1), Penicillium (2), Talaromyces (4), Trichocladium (1), and Trichoderma (1). All

strains were identified by phenotypic features, with molecular data additionally obtained for

eleven strains. Five species are reported for the first time in Argentina.

Key worps —Ascomycota, Eurotiales, South America, Monte, Chaco.

Introduction

Temperature is an environmental parameter that plays a key role in the

survival, growth, distribution, and speciation of microorganisms on earth

(Mouchacca 1993). Fungi growing at high temperatures are generally classified

536 ... Romero &al.

in two groups: thermophilic and thermotolerant. Thermophilic fungi are

those able to grow in a temperature range of 20-50 °C or above; while the

maximum temperature for growth of thermotolerant fungi is near 50 °C with

a minimum of below 20 °C (Cooney & Emerson 1964, Mouchacca 2007).

Alternatively, heat-resistant fungi are defined as those capable of surviving

to a thermal shock, e.g., temperatures at or above 75 °C for 30 minutes

or more (Samson & al. 2000). The fungal structures able to survive these

extreme temperatures are mainly ascospores, but other propagules such

as chlamydospores, thick-walled hyphae, and sclerotia, also may survive

such temperature extreme (Scholte & al. 2004). Dijksterhuis & Samson

(2006) expanded the concept of heat-resistant fungi to include most of

fungal structures that survive after a thermal shock between 55-95 °C. Heat

resistance is associated with the ability to maintain the viability of certain

fungal structures in order to overcome natural thermal shock. Thus, heat-

resistant fungi can be mesophilic, thermotolerant, or thermophilic.

Soil, the primary reservoir of fungi, is also the main repository of dormant

propagules of true soil-borne fungi as well as of fungi that grow elsewhere.

Most fungi achieve survival through the production of both dormant

and dispersal spores (Carlile & Watkinson 1994). Latent spores normally

germinate due to slow dormancy decay or in response to a specific (physical

or chemical) stimulus that breaks the dormancy (Carlile & Watkinson 1994),

such as a thermal shock. Constitutive dormancy is a condition in which

the development is delayed by an innate property, such as a barrier to the

penetration of nutrients (including water), a metabolic blockage, or the action

of a self-inhibitory molecule (Dijksterhuis & Samson 2006, Dijksterhuis

2007). Ascospores, the sexual propagules of the Ascomycota, often have a

long survival capability (Dijksterhuis 2007). Ascospores of heat-resistant

fungi exhibit constitutive dormancy and require a robust signal (heat, high

pressure, or exposure to organic compounds, such as dimethylketone) to

break the dormancy (Dijksterhuis 2007, Stchigel 2000). Warcup & Baker

(1963) noted an increase in the number of ascomycete colonies when soil

samples were exposed to temperatures between 50-75 °C for 30 minutes

before culturing, compared with those not exposed to this thermal shock.

The fungal soil communities from arid regions are mainly characterized

by a low population coupled with a high biodiversity (Mouchacca 1993).

During broad research on heat resistant fungi from the semi-arid region

of Northern Argentina 194 isolates were obtained. Most isolates were

related to Aspergillus sect. Fumigati, but 34 isolates represented different

Heat resistant ascomycetes in Argentina ... 537

interesting taxa. This study documents those 34 miscellaneous heat-resistant

ascomycetes isolated from soil.

Materials & methods

Description of the sampling area

Fifty 200-g soil samples were collected in the summer of 2009 and winter of

2011 from several sites in Catamarca and La Rioja provinces, Northern Argentina.

Of the six phytogeographical regions of Catamarca (Morlans 1995), most samples

were collected from the “monte” region and only a few from the “chaquefia’” region;

additionally three collections were sampled from the “monte” region of La Rioja (with

five phytogeographical regions). The monte region in Catamarca, which comprises c.

14,600 km? (Manghi & al. 2005) and extends through the center of La Rioja province,

is characterized by steppe vegetation dominated by xerophilous, psammophilous,

or halophilous shrubs, and characterized by the almost constant presence of

Larrea (Zygophyllaceae) and Prosopis (Leguminosae). The climax community is Larrea

divaricata Cav. (“jarillal”), which develops in the sandy pockets and plains or stony-

sandy soils (Cabrera 1971). Climate in the monte is subtropical, arid, with an annual

150-200 mm precipitation occurring mostly in the summer (60-70% of total rainfall;

Morlans 1995). Temperature shows a relatively broad seasonal and daily variation,

with autumn/winter lows (-30 °C in west mountain region) and spring/summer

highs (45 °C in the east and central regions) (Gobierno de Catamarca 2018, Morlans

1995). The strong winds throughout the year favor environmental dryness. The monte

elevation ranges from 600-3000 m a.s.l. (Morlans 1995).

Fungal isolation

From each sample, c. 5 g of soil was placed into a conical flask, mixed with 100

ml of sterile melted malt extract agar (Oxoid MEA: malt extract 30.0 g, mycological

peptone 5.0 g, agar 15.0 g) plus 50 ppm of chloramphenicol, and heated at 75 °C for

30 min. The suspension was plated into sterile Petri dishes and, once gelled, incubated

at 30 °C <30 d (Samson & al. 2000). All colonies were transferred to individual Petri

dishes containing MEA to obtain pure cultures. The fungal strains were deposited

in the culture collection of the Herbarium, Universidad de Buenos Aires, Argentina

(BAFC) and the culture collection of Faculty of Medicine, Universitat Rovira i Virgili,

Reus, Catalonia, Spain (FMR). Author abbreviations include SMR (S.M. Romero),

AIR (A.I. Romero), AMS (Stchigel), ERA (Rodriguez Andrade), VB (Barrera), and

RC (Comerio). Abbreviations under CULTURE(S) EXAMINED are RN (Ruta Nacional)

and RP (Ruta Provincial).

Morphological study

Preliminary characterization at the genus level was conducted on MEA (Domsch

& al. 2007, Guarro & al. 2012). Species were identified by growing the fungal strains

on different culture media and temperatures (following the bibliography for each

genus mentioned in each description). Vegetative and reproductive structures were

538 ... Romero &al.

microscopically examined by mounting directly into lactic acid from cultures on MEA

and then observed under a Zeiss Axioskop bright field microscope. Mature ascomata

were crushed, coated with gold, and observed and photographed with a Zeiss Supra

40 (extra high tension = 3 Kv; working distance = 3.7-6 mm) scanning electron

microscope.

Molecular study

EXTRACTION: DNA was extracted directly from colonies on potato dextrose agar

(PDA) after 7-10 d at 25 °C in darkness, through the modified protocol of Miiller & al.

(1998). The following genetic markers were sequenced: fragments of -tubulin (BenA),

calmodulin (CaM), RNA polymerase II second largest subunit (rpb2) (Peterson & al.

2010, Samson & al. 2014), and the DNA replication licensing factor (Mcm7) (Schmitt

& al. 2009, Raja & al. 2011). The primers used were Bt2a and Bt2b for BenA, CMD5

and CMD6 for CaM (Glass & Donaldson 1995), RPB2-5F and RPB2-7R for rpb2 (Liu

& al. 1999), and Mcm7-709For and Mcm7-1348Rev for Mcm7 (Schmitt & al. 2009).

PCR products were sequenced in both directions using the same primers at Macrogen

Europe (Macrogen Inc., Madrid, Spain). Sequences were assembled and edited using

Sequencher v.4.1.4. The sequences generated were deposited in the GenBank database

(https:// www.ncbi.nlm.nih.gov/genbank) using the Webin platform of the European

Bioinformatics Institute (EMBL-EBI) (http://www.ebi.ac.uk/ena).

PHYLOGENETIC ANALYSIS: Multiple sequence alignment was conducted using

ClustalW (Thompson & al. 1994) and MUSCLE (Edgar 2004) within MEGA v.6

software (Tamura & al. 2013), with manual adjustments for refinement. The Maximum

Likelihood (ML) phylogenetic method was also run with MEGA v.6, as well as the

estimation of the best nucleotide substitution model. Support of the internal branches

was assessed by the bootstrap method with 1000 replications, where values 270 were

considered significant. The phylogenetic analyses were carried out first individually

for each gene followed by a concatenated study. To support our analyses, sequences

were obtained from GenBank and added to the analyses.

Taxonomy

In our study, 34 heat-resistant fungal strains from the semiarid region in

Argentina were all morphologically characterized and assigned to 17 taxa in

10 genera—Arthrinium (1), Aspergillus (3), Epicoccum (1), Gilmaniella (1),

Hamigera (2), Leiothecium (1), Penicillium (2), Talaromyces (4), Trichocladium

(1), and Trichoderma (1). For 11 strains, the morphological identification was

confirmed by molecular analyses. Five taxa are reported for the first time in

Argentina.

Arthrinium phaeospermum (Corda) M.B. Ellis, Mycol. Pap. 103: 8. 1965.

Cotontes on MEA, 25 °C, 7 d, floccose, covering the whole culture plate,

greyish, with dark specks and areas due to the anamorph presence.

Heat resistant ascomycetes in Argentina ... 539

Conrpi1a lenticular, dark brown, 9-11 x 5-7 um, smooth with a conspicuous

pale equatorial stripe.

CULTURES EXAMINED—ARGENTINA. CaTaMARCA: RP 46, km 70, 28°05’35”S

66°12’55”W, 859 m a.s.l., 24.VHI.2011, leg. SMR, identified by SMR & RC, isolation

22.X.2011 (BAFCcult 4624); 27°35’13”S 66°22’11’W, 996 m a.s.l, 24.VIII.2011, leg.

SMR, identified by SMR & RC, isolation 15.X1.2011 (BAFCcult 4625).

DISTRIBUTION— Worldwide, including Argentina (Broggi & al. 2007, Crous &

Groenewald 2013, Domsch & al. 2007).

Hasitat—Cereals and vegetable materials, dung, fresh water, and soil

(Domsch & al. 2007).

CoMMENTS—Otur isolates agreed morphologically with descriptions by Ellis

(1971), Domsch & al. (2007), and Crous & Groenewald (2013). In Argentina,

this species was isolated from soil in Buenos Aires (Cabello 1986) and cereals

collected in Entre Rios (Broggi & al. 2007, Sacchi & al. 2009).

Aspergillus flavus Link, Mag. Ges. Naturf. Freunde, Berlin 3: 16. 1809.

Cotoniges on MEA, 25 °C, 7 d, plane, granulose, floccose at the centre,

50-54 mm diam., yellowish green, exudate scarce uncolored; asexual sporulation

abundant. On Czapek yeast agar (CYA), 25 °C, 7 d, sulcate, granulose, floccose

at the centre, 60-68 mm diam., yellowish green, exudate scarce uncolored;

asexual sporulation abundant.

CONIDIAL HEADS commonly columnar, radiate, mainly uniseriate. STIPES

roughened, vesicles spherical. Conip1a globose, 3.5-4.5 um, with finely

roughened walls.

CULTURE EXAMINED—ARGENTINA. La Rioja: RN 60, km 1172, 28°40'30’S

66°30'23”W, 653 m a.s.l., 12.1.2009, leg. SMR, identified by SMR, isolation 1.III.2011

(BAFCcult 4615).

DIsTRIBUTION— Worldwide and cosmopolitan (Domsch & al. 2007; Ramirez-

Camejo & al. 2012).

HasitatT—Soil, cultivated or not (Domsch & al. 2007). This is the most

widely cited fungal species as a food contaminant (Klich 2002, Pitt & Hocking

2009).

ComMMENTS— The micromorphology of our isolate, and even its cultural

features, agreed with the descriptions of Klich (2002) and Pitt & Hocking

(2009). Horn & al. (2009) described the sexual stage for this heterothallic

species as Petromyces flavus B.W. Horn &al., but according to the International

Code of Nomenclature (Turland & al. 2018) and following Samson & al.

(2014), the correct name for this species is Aspergillus flavus. In Argentina,

540 ... Romero & al.

it has been isolated several times from soil (Allegrucci & al. 2009, Cabello

1986, Godeas & al. 1977, Mangiaterra & al. 2006, Winitzky 1948), foods

(Bresler & al. 1991, Castellari & al. 2015, Dalcero & al. 1998, Etcheverry & al.

1999, Romero & al. 2005, Sepulveda & Piontelli 2005) and medicinal herbs

(Sanchez & al. 2006).

Aspergillus montevidensis Talice & J.A. Mackinnon,

C.r. Seanc. Soc. Biol. 108: 1007. 1931. BIGiwTASB

CoLonies on MEA, 25 °C, 7 d, plane and dense, 14 mm diam., yellowish;

after 14 incubation d 26-28 mm diam., yellowish, with or without brown

center. On Czapek yeast agar with 20% sucrose, 25 °C, 7 d, velutinous, 35-47

mm diam., yellow, with green center due to the anamorph presence; after 14

incubation d covering the whole culture plate, velutinous to granulose due

to the abundant cleistothecia production, yellow with green center; asexual

sporulation copious; reverse yellowish under cleistothecia and greenish

under anamorph structures. At 37 °C, 7 d, 18-26 mm diam., yellow.

CLEISTOTHECIA superficial, mainly globose, discrete, 90-150 um diam.,

yellow, covered by a hyaline hyphal net. Asci globose, 8-spored, 10-12 x

8-11 um, evanescent. Ascosporss lenticular, 5-4 x 3-4 um, with rough

convex surfaces, furrowed and double-crested along the equator; SEM

images revealed pores distributed in the whole ascospore surface, but mainly

on crests. CONIDIAL HEADS radiate, uniseriate. STIPES 150-210 x 6-8 um,

smooth, vesicles globose, 15-27 um diam. PHIALIDEs ampulliform, 5-8 x 4

um. Conrp1A globose to subglobose, 5-4 x 4-3 um, spinulose.

CULTURES EXAMINED—ARGENTINA. CATAMARCA: RN 46, km 185, 27°45712”S

66°47°59”W, 1069 m a.s.l.. 9.1.2009, leg. SMR, identified by SMR, isolation

21.111.2011 (BAFCcult 4613; GenBank LT964777, LT964778); Papachacra, 2564 m

a.s.l., 10.1.2009, leg. SMR, identified by SMR, isolation 1.[1I.2011 (BAFCcult 4614;

GenBank LT964779, LT964780).

DIsTRIBUTION—Argentina (Winitzky 1952), Australia, Brazil, India, Israel,

Japan, Pakistan, Peru, South Africa, Syria, Tuamotu Archipelago, Turkey

(Domsch & al. 2007).

Hasitat—Soil, stored and/or decaying food products among other

substrates (Domsch & al. 2007, Pitt & Hocking 1997).

ComMMENTS—The morphological features of our strains agreed with

those described by Thom & Raper (1941), Raper & Fennell (1965), Klich

(2002) and Hubka & al. (2013). This species was also known as Eurotium

amstelodami because Thom & Raper (1941) changed the original species

Heat resistant ascomycetes in Argentina ... 541

concept of Mangin (1909), from a species with smooth-walled ascospores to

a species with ornamented ones. Their 1941 published name, E. amstelodami

was, in fact, a later illegitimate homonym. Pitt (1985) noted this confusion

and speculated that Thom & Church (1926) and Thom & Raper (1941)

did not base their description on the Mangin species concept. Hubka & al.

(2013) concluded that the first available and validly published name for the

species matching the concept of Thom & Raper (1941) is A. montevidensis,

which includes both the anamorph and the teleomorph states.

Aspergillus sydowii (Bainier & Sartory) Thom & Church, Aspergilli: 147. 1926.

CoLoniEs on MEA, 25 °C, 7 d, velutinous to slightly floccose, 14-15 mm

diam., blue green, asexual sporulation copious. On CYA, 25 °C, 7 d, sulcate,

velutinous, 20 mm diam., blue green, exudate brown copious; asexual

sporulation abundant; reverse orange brown. On 25% glycerol nitrate agar

(G25N), 25 °C, 7 d, plane and dense, 11-13 mm diam., bluish grey.

CONIDIAL HEADS radiate, biseriate; small and short penicillate heads

produced. Stipes 250-350 um, pigmented. Conip1A globose, 3.5-4 um,

very rough to spinose. HULLE CELLS abundantly produced.

CULTURES EXAMINED—ARGENTINA. Catamarca: Rincon, 28°22’03”S

66°13’39”W, 8.1.2009, leg. SMR, identified by SMR, isolation 4.VI.2009, (BAFCcult

4589); RP 25, 28°22’41”S 66°11'50”W, 1304 m a.s.l., 23.VIII.2011, leg. SMR,

identified by SMR, isolation 10.11.2012 (BAFCcult 4612).

DisTRIBUTION—Argentina, Australia, Austria, Brazil, British Isles, Canada,

Czechoslovakia, Egypt, India, Italy, Japan, Kuwait, Malaysia, Namibia,

Pakistan, Peru, Singapore, South Africa, Spain, Syria, ex USSR, U.S.A.

(Domsch & al. 2007).

Hasitat—Mainly isolated from soil, but also from air (indoor and

outdoor) (Klich 2002). This species has been reported from numerous

foods, especially dried ones (Pitt & Hocking 2009). It has been recovered

also from marine sponges (Ein-Gil & al. 2009).

CoMMENTS— The features of our isolates matched those described by

Raper & Fennell (1965) and Klich (2002). In Argentina, A. sydowii has been

previously isolated from air (Winitzky 1948), amaranth seeds (Bresler &

al. 1995), dried vine fruits (Romero & al. 2005), corn and soybean seeds

(Sepulveda & Piontelli 2005), soil (Allegrucci & al. 2007, Eliades & al. 2006a,

Mangiaterra & al. 2006) and medicinal herbs (Sanchez & al. 2006). This is

the first report for Catamarca province.

542 ... Romero &al.

Epicoccum nigrum Link, Mag. Gesell. Naturf. Freunde Berlin 7: 32. 1816.

Co.ontes on Blakeslee’s MEA, 25 °C, 7 d, floccose, 53-58 mm diam., orange

brown, exudate and soluble pigment absent; reverse orange. On oatmeal agar

(OA), 25 °C, 7 d, lanose to floccose, 60 mm diam., orange with yellowish centre,

exudate absent, soluble pigment yellow; reverse yellowish orange; after 12 d

of incubation brownish green centre due to sporulation. On PDA, 25 °C, 7 d,

23-30 mm diam., orange brown, exudate red caramel, soluble pigment amber;

reverse red brown.

CONIDIOPHORES crowded, even forming pustules. Conrp1A globose to

pyriform 22-30 x 18-29 um, orange brown to dark brown, verrucose, several

septa which divide the conidia in different directions, pale extending cell at the

base.

CULTURE EXAMINED— ARGENTINA. CATAMARCA: Papachacra, 2564 m as.l.,

10.1.2009, leg. SMR, identified by SMR, isolation 1.[II.2011 (BAFCcult 4622).

DISTRIBUTION— Worldwide (Domsch & al. 2007).

Hasitat—Cereals and grains, fruits, soil, among others (Domsch & al.

2007).

CoMMENTS—The morphologic characteristics of our isolate agreed with the

descriptions of Epicoccum nigrum by Ellis (1971) and Domsch & al. (2007). In

Argentina there are numerous records of this species, which were isolated from

soil samples of Jujuy province (Giusiano & al. 2002), wheat and soybean of

Entre Rios province (Broggi & al. 2007), soil and litter of Buenos Aires province

(Allegrucci & al. 2005, 2007), and alpataco fruits from La Pampa province

(Canafoglia & al. 2007), among others.

Gilmaniella humicola G.L. Barron, Mycologia 56: 514. 1964. FIG. 1P,Q

Cotontes on MEA, 25 °C, 7 d, velutinous to slightly floccose, 59-75 mm

diam., grayish, darkening in age; reverse very dark to black. At 30 °C, 7 d,

velutinous to slightly floccose, 79-83 mm diam., grayish, darkening in age;

reverse very dark to black.

Hypuae light brown in age, smooth. Conip14 globose, terminal or lateral,

usually in clusters and provided with a distinct germ pore, smooth, brown

7-9 um.

CULTURES EXAMINED—ARGENTINA. CaTAMaARca: RP 25, 28°15’31”S 66°08’47’W,

1424 m a.s.l., 23.VIH.2011, leg. SMR, identified by SMR & RC, isolation 11.1.2012

(BAFCcult 4610, 4588).

DIsTRIBUTION—Canada, Egypt, England, France, India, Japan, Namibia, Solomon

Islands, The Netherlands, United States of America (Domsch & al. 2007).

Heat resistant ascomycetes in Argentina ... 543

Fic. 1. Aspergillus montevidensis (BAFCcult 4613): A. Colonies on Czapek yeast extract agar

(CYA) with 20% of sucrose, 14 d, 25 °C; B. Ascospore (SEM). Hamigera paravellanea (BAFCcult

4605): C. Colonies on CYA, 7 d, 25 °C; D. Ascospores (SEM); E. Conidiophore and conidia.

Hamigera terricola (BAFCcult 4580): E Colonies on CYA, 7 d, 25 °C; G-H. Conidiophores and

conidia; I. Ascospores (SEM). Leiothecium ellipsoideum (BAFCcult 4598): J. Colony on MEA,

7 d, 25°C; K, L. Ascospores (SEM); M. Chlamydospore. Penicillium capsulatum (BAFCcult 1627):

N. Colonies on CYA, 7 d, 25 °C; O. Conidiophore and conidia. Gilmaniella humicola (BAFCcult

4588): P. Conidia with germination pores (arrows); Q. Colony on MEA 7 d, 30 °C. Trichocladium

pyriforme (BAFCcult 4623): R. Colony on MEA 14 d, 25 °C; S, T. Chlamydospores.

544 ... Romero &al.

Hasitat—Dung, groundnuts, plant debris, soil, among others (Domsch &

al. 2007).

COMMENTS—Otur isolates were phenotypically in accordance with Barron

(1964) and Ellis (1971). Gilmaniella humicola is a good representative example

of a heat-resistant fungus (Domsch & al. 2007), and it was the most thermal-

resistant fungus found by Bollen (1969), tolerating a temperature of 90 °C for

30 minutes. All our isolates were recovered from samples collected in winter.

In Argentina, the fungus was previously reported from soil in a Nothofagus

dombeyi forest in Rio Negro province (Godeas 1977).

Hamigera paravellanea S.W. Peterson, Jurjevi¢, Bills, Stchigel, Guarro & EE. Veja,

Mycologia 102: 852. 2010. FIG. 1C-E

Cotoniges on MEA, 25 °C, 7 d, plane somewhat floccose, 47-67 mm

diam., pale orange to tan; reverse dark, mainly at colony centre. On CYA, 25

°C, 7 d, low, slightly sulcate, 50-70 mm diam., pale orange to tan; reverse dark

reddish-brown.

GYMNOTHECIA globose, discrete, 120-280 um diam., cream coloured,

surrounded by a profuse hyphal net. Asci subglobose to claviform, 8-spored,

15-20 x 11-12 um, evanescent. Ascospores broadly ellipsoidal, 7—-7.5 x 5-6

um, hyaline, thick-walled, echinulate. ConrpIOPHORES penicillate, 120-480

x 3-5 um. Stipes slightly spathulate, smooth or roughened towards their

bases, widest at their apices. METULAE with wide apices, 10-15 x 4-6 um.

PHIALIDES ampulliform, 8-6 x 2-3 um. Conrp1a ellipsoidal, 3-4 x 2-3 um.,

smooth.

CULTURES EXAMINED—ARGENTINA. CATAMARCA: RP 25, 28°15’31”S 66°08’47’W,

1424 ma.s.L., 6.1.2009, leg. SMR, identified by SMR, AMS & ERA, isolation 20.1X.2011

(BAFCcult 4605, FMR 16758; GenBank LT991984, LT991994); 23. VII.2011, leg. SMR,

identified by SMR, AMS & ERA, isolation 11.1.2012 (BAFCcult 4606, FMR 16762;

GenBank LT992033, LT992032).

DIsTRIBUTION— Poland, Spain (Peterson & al. 2010).

Hasitat—Soil, dung (Peterson & al. 2010).

CoMMENTS—Morphological characters observed agreed with the description

of the protologue (Peterson & al. 2010). This is the first report for Argentina.

Hamigera terricola S.W. Peterson, Jurjevi¢, Bills, Stchigel, Guarro & EE. Veja

Mycologia 102: 855. 2010. FIG. 1F-I

COLONIES on MEA, 25 °C, 7 d, low, sulcate, velutinous, 58-70 mm diam.,

ochraceous buff to tan; reverse dark reddish-brown. On CYA, 25 °C, 7 d, low,

Heat resistant ascomycetes in Argentina ... 545

plane, velutinous, 40-43 mm diam., ochraceous buff with yellowish margins;

reverse dark reddish-brown.

GYMNOTHECIA globose, discrete, 80-180 um diam., cream coloured,

surrounded by a profuse hyphal net. Asci subglobose, 8-spored, 15-17.5 x

11-13 um, evanescent. AscosPorEs broadly ellipsoidal, 7-8 x 5-6 um, hyaline,

thick-walled, echinulate. CoNIDIOPHORES penicillate, 170-400 x 3-6 um.

STIPES sometimes spathulate, with apical vesicles 7-9 um wide, smooth or

slightly roughened. METULAE with wide apices, 7-11 x 3-6 um. PHIALIDES

ampulliform, 7-8 x 3 um. Conrp1a ellipsoidal, 3-4 x 2-3 um., smooth.

CULTURE EXAMINED—ARGENTINA. CATAMARCA: RN 60, km 1102, 28°55’15"S

66°08'46’W, 338 ma.s.l., 5.1.2009, leg. SMR, identified by SMR, AMS & ERA, isolation

4.V1.2011 (BAFCcult 4580, FMR 16756; GenBank LT991985, LT991995).

DIsTRIBUTION—Costa Rica, Equatorial Guinea, French Guiana, Guatemala,

Panama (Peterson & al. 2010).

HasitaT—Textile sample in contact with soil (Peterson & al. 2010).

CoMMENTS—In a broad sense, the features of this isolate matched with those

described by Peterson & al. (2010), but ascospores are slightly larger and

the asci smaller than in the original description. This is the first report of

Hamigera terricola for Argentina.

Leiothecium ellipsoideum Samson & Mouch.,

Can. J. Bot. 53: 1634. 1975. FIG. 1J-M

Cotonigs on MEA, 25 °C, 7 d, forming concentric rings, 82-83 mm diam.,

dark, superficial white mycelium present; reverse dark. At 35 °C, covering the

whole culture plate before 7 d; reverse dark due to ascomata abundance. On

OA, 25 °C, 7 d, colonies presenting a continuous layer of ascomata, dark, almost

lacking aerial mycelium. At 35 °C, covering the whole culture plate before 7 d;

subtle annularly developed, less aerial mycelium than on MEA.

VEGETATIVE HYPHAE smooth, 3-11 um diam., hyaline. CLEISTOTHECIA

globose, discrete, 40-90 um diam., dark brown, superficial or semi-submerged

in the substratum, covered by white mycelium supporting ascomata as well;

wall persistent, textura angularis, 5-8 um wide, composed by 15-25 um

diam. dark brown cells. Asci1 globose to subglobose, 14-18 x 13-15 um,

evanescent. Ascosporss ellipsoidal, 7-9 x 5.5-7 um, hyaline, spinulose under

low magnifications, broadly reticulate under high magnifications; young

ascospores presented an internal hyaline sheath which fades with maturity;

SEM observations revealed alveolate plaques on ascospore wall, alveoles

presented conspicuous projections at their edges which look as low frills.

546 ... Romero &al.

CHLAMYDOSPORES isolated, subglobose to ellipsoidal, umbonate, 4.5-11 um

diam., smooth, endogenous, and also terminal in maturity.

CULTURES EXAMINED— ARGENTINA. CATAMARCA: RN 60, km 1146, 28°13’05"S

66°22’41”W, 1051 m a.s.L., 5.1.2009, leg. SMR, identified by SMR, RC & AMS, isolation

13.IX.2011 (BAFCcult 4598, FMR 15064; GenBank LT992254, LT992257); km 1016,

29°30'04”S 65°37'57”W, 237 m a.s.L, 22.VIH.2011, leg. SMR, identified by SMR, RC

& AMS, isolation 12.XI.2011 (BAFCcult 4604, FMR 16765; GenBank LT992253,

LT992256); RP 47, km 35.5, 27°26'50”S 66°24’26”W, 2700 m a.s.l., 24.VHI.2011, leg.

SMR, identified by SMR, RC & AMS, isolation 18.1.2012 (BAFCcult 4609, FMR 16768;

GenBank LT992255, LT992258).

DISTRIBUTION—Canada (Samson & Mouchacca 1975), Greece, Hungary,

Portugal, Spain, United States of America, and Venezuela (CBS-KNAW Culture

Collection 2018).

HaBitTat—Soil, seeds of Capsicum annuum (Samson & Mouchacca 1975),

nest material of Nomia sp., a ground-nesting solitary bee (CBS-KNAW Culture

Collection 2018).

CoMMENTS—Argentinian isolates largely agreed with the description given

by Samson & Mouchacca (1975) and Guarro & al. (2012). However, these

strains grew faster on MEA at 35 °C than the ex-type strain. Regarding

micromorphology, no chlamydospores in chains were observed (solitary

chlamydospores were seen on small hyphal protrusions) and ascomata were

smaller (40-90 um vs. 125 um diam) than those described in the protologue.

This is the first report of Leiothecium ellipsoideum for Argentina.

Penicillium capsulatum Raper & Fennell, Mycologia 40: 528. 1948. Fic. 1N,o

Cotonigs on MEA, 25 °C, 7 d, low, plane, velutinous, 14-18 mm diam.,

white mycelium at the edges, grey green to dull green; conidiogenesis moderate;

exudate and soluble pigment absent; reverse pale to yellowish. On CYA,

25 °C, 7 d, moderately deep, sulcate, slightly fasciculate, 12-15 mm diam.;

conidiogenesis dense, grey green to dull green, white mycelium at the edges;

exudate and soluble pigment absent; reverse pale to yellowish. At 37 °C, 7 d,

funiculose, 18-21 mm diam., with white mycelium at the edges; exudate and

soluble pigment absent; reverse pale to buff. At 5 °C germination absent. On

G25N, 25 °C, 7 d, sulcate, velutinous to floccose, 6 mm diam., white mycelium

at the edges, conidia in mass grey green to dull green; exudate and soluble

pigment absent; reverse pale to yellowish.

TELEOMORPH absent. CONIDIOPHORE monoverticillate, arising from

substrate surface or from superficial mycelium. STIPES somewhat sinuose,

10-40 um long., smooth, mainly non-vesiculate, but sometimes vesicles up to

Heat resistant ascomycetes in Argentina ... 547

4 um diam. PHIALIDEs 5-8, ampulliform to acerose, 6 x 2 um, occasionally up

to 8 um in length, with short collula. Conrp1a ellipsoidal to cylindrical, 3-3.5

x 2 um, smooth-walled.

CULTURE EXAMINED—ARGENTINA. CaTAMARCA: RP 33, 28°42’03”S 65°46'08”W

418 m as.l., 22.VIH.2011, leg. SMR, identified by SMR & RC, isolation 23.X1.2011

(BAFCcult 1627).

DISTRIBUTION—China (Chen & al. 2013), Kiribati (Gilbert Islands), Panama

(Raper & Fennell 1948).

Hasitat—Optical instrument, environmentally exposed canvas and

deteriorated military equipment (Raper & Fennell 1948), and contaminated

beet pulp (Puls & Coughlan 1996).

CoMMENTS—Otr soil-borne isolate matched the Pitt (1979) description. We

also observed vesiculate conidiophores, but phialides were a bit shorter than in

the holotype. Pitt (1979) described the colonies on CYA at 25 °C as depressed

at the center, whereas the Argentinian strains presented a high and fasciculate

center. This is the first report of Penicillium capsulatum for Argentina.

Penicillium citrinum Thom, U.S.D.A. Bur. Animal Industr. Bull. 118: 61. 1910.

CoLonigs on MEA, 25 °C, 7 d, low, plane, centrally umbonate, velutinous,

18-22 mm diam., grey green to dull green; conidiogenesis dense; exudate pale,

soluble pigment absent; reverse pale. On CYA, 25 °C, 7 d, moderately deep,

sulcate, velutinous, 24-30 mm diam., grey green to dull green; conidiogenesis

dense, white mycelium at the edges; exudate pale, soluble pigment yellowish;

reverse yellow to orange. At 37 °C, 7 d, 2-11 mm diam. At 5 °C no germination.

TELEOMORPH absent. CONIDIOPHORES biverticillate, arising from substrate

surface or from superficial mycelium. STIPEs with smooth walls and a

conspicuous divaricate metulae whorl. METULAE of uniform length, 3-5

on each stipe. PH1ALIDES ampulliform, short collula. Conip1a globose to

subglobose, 2.5-3 um diam., smooth-walled.

CULTURE EXAMINED—ARGENTINA. CATAMARCA: Corral Quemado to Papachacra,

27°07'33"S 66°56'36"W, 2152 m a.s.l.: 10.1.2009, leg. SMR, identified by SMR & RC,

isolation 5.III.2011 (BAFCcult 4616).

DisTRIBUTION—Penicillium citrinum is considered a cosmopolitan and

frequently isolated fungus (Domsch & al. 2007).

Hasitat—The fungus has been reported from many substrates, including

food (Domsch & al. 2007).

CoMMENTS—Otmr isolate matched with the description of Penicillium citrinum

provided by Pitt (1979). In Argentina, it has been isolated many times, generally

548 ... Romero & al.

from food, e.g., soybeans (Bonera & al. 1982), amaranth seeds (Bresler & al.

1991, 1995), balanced feed for birds (Magnoli & al. 1998), and dried vine fruits

(Romero & al. 2005), among others.

Talaromyces macrosporus (Stolk & Samson) Frisvad, Samson & Stolk,

Antonie van Leeuwenhoek 57: 186, 1990.

Cotonigs on MEA, 25 °C, 14 d, almost covering the whole culture plate,

moderate deep, plane, granulose, covering the whole culture plate, yellow,

becoming orange at the centre.

CLEISTOTHECIA superficial, mainly globose, commonly confluent but at the

margin discrete, 200-400 um diam., yellow. Asci globose, 8-spored, evanescent;

initials showing thin antheridia coiled around slender clavated ascogonia.

ASCOSPORES ovoidal to broadly ellipsoidal, 5.5-6.5 x 3.5-4 um, pale yellow,

with thick walls and spines. ANAMORPH absent.

CULTURE EXAMINED—ARGENTINA. CaTAMARCA: RP 33, 28°42’03”S 65°46’08”W,

418 m asl, 22.VIL2011, leg. SMR, identified by SMR & AIR, isolation 23.X1.2011

(BAFCcult 4617).

DISTRIBUTION—Ghana, Japan, Korea, Malaysia, New Guinea, Panama, Poland,

South Africa, The Netherlands, United States of America (CBS-KNAW Culture

Collection 2018, Stolk & Samson 1972).

HasitatT—Soil, canned apples (Stolk & Samson 1972), freshly harvested

strawberries (Frison & al. 2012).

CoMMENTS—Our isolate agreed with the description of Talaromyces

macrosporus presented by Stolk & Samson (1972), who recognize two varieties:

T. flavus var. flavus and T. flavus var. macrosporus. Regarding these varieties,

Beuchat (1988) pointed out that the strains with small ascospores (T. flavus

var. flavus) are less thermal resistant than those with larger ones (T. flavus

var. macrosporus). Frisvad & al. (1990) raised them to species rank based on

ascospore size, heat-resistance, and secondary metabolite production. In

Argentina, the species has previously been recorded in soils either as T: flavus

s.l. or as T: flavus var. flavus (Bertoni 1973, Eliades & al. 2006 a,b, Magnoli & al.

1998). Frison & al. (2012) reported T: macrosporus on fresh strawberries.

Talaromyces pinophilus (Hedgc.) Samson, N. Yilmaz, Frisvad & Seifert, Stud.

Mycol. 70: 176. 2011.

CoLoniges on MEA, 25 °C, 7 d, moderately deep, plane, floccose, 34-36

mm diam., green; conidiogenesis sparse, yellow mycelium present, almost

dominating colony colour; exudate pale, soluble pigment absent; reverse

Heat resistant ascomycetes in Argentina ... 549

yellowish. On CYA, 25 °C, 7 d, moderately deep, plane, floccose, 19-20 mm

diam., greenish yellow; conidiogenesis sparse, yellow mycelium present

predominating in colony colour; exudate pale, soluble pigment absent; reverse

brown. At 37 °C, 7 d, 24-26 mm diam., with white mycelium at the edges;

exudate and soluble pigment absent; reverse buff. At 5 °C no germination. On

G25N, 25 °C, 7 d, plane, floccose, 3-5 mm diam., white mycelium; sporulation

absent; exudate pale, soluble pigment absent; reverse buff.

TELEOMORPH absent. CONIDIOPHORES biverticillate, arising from substrate

surface and also from aerial hyphae. Stipes 100-150 um long., smooth.

MetuLae in whorls mainly of 5 elements, 10-12 um long. PHIALIDEs 5-8,

acerose, 8-10 um long., tapered. Conip1A globose to subglobose, 3-4 um

diam., smooth-walled.

CULTURE EXAMINED—ARGENTINA. CATAMARCA: Corral Quemado to Papachacra,

27°07'33"S 66°56”36"W, 2152 m a.s.l., 10.1.2009, leg. SMR, identified by RC, isolation

5.I11.2011 (BAFCcult 4618).

DIsTRIBUTION—Argentina (Magnoli & al. 1998); Australia, France, India,

Papua New Guinea, United States of America (Pitt 1979); Uruguay (Galvalisi

& al. 2012).

Hasitat—Soil, decaying plants, deteriorated material, barley grains,

fermented dry sausages, balanced feed for birds (references as above).

ComMMENTS—Excluding conidia, which in this case were a little bit bigger (in

contrast to 2.5-2.8 x 2.2-2.5 um), we did not find morphological differences

compared to those of Talaromyces pinophilus given by Pitt (1979). In Argentina,

it has been previously reported as Penicillium pinophilum from corn, balanced

feed for birds, and soil (Etcheverry & al. 1999, Magnoli & al. 1998, Nesci & al.

2006, Mangiaterra & al. 2006).

Talaromyces trachyspermus var. macrocarpus J.E. Wright & Loewenb.,

Bol. Soc. Argent. Bot. 15(1): 100. 1973.

CoLonigs on MEA, 25 °C, 14 d, moderately deep, plane, granular, 37-44

mm diam., white to cream-colored; exudate absent; reverse orange.

CLEISTOTHECIA superficial, mainly globose, usually confluent, 400-930

uum diam., at first white becoming cream-colored in age, covered by a network

of hyphal net. Ascr globose to ovoid, 8-spored, 7-8 x 6-7 tm, evanescent.

AscosPpores ellipsoidal, 3-4 x 2-3 um, spinulose. ConrDIOPHORES slender

monoverticillate and biverticillate, short. METULAE 8-11 x 2 um. PHIALIDES

2-5 in verticils, lanceolate, tapering to neck, 9-10 x 2 um. Conrp1a ellipsoidal,

2-2.5 x 2 um, smooth-walled.

550 ... Romero & al.

CULTURE EXAMINED—ARGENTINA. CATAMARCA: RP 46, km 185, 27°45’12”S

66°47'59”’W, 1069 m a.s.l., 9.1.2009, leg. SMR, identified by SMR & AIR, isolation

21.111.2011 (BAFCcult 4621; GenBank LT968851).

DISTRIBUTION—Argentina (Bertoni & al. 1973).

HasitatT—Soil (Bertoni & al. 1973).

COMMENTS—The characteristics of isolate BAFCcult 4621 agreed with the

description of Talaromyces trachyspermus var. macrocarpus by Bertoni & al.

(1973). This variety was not accepted by Pitt (1979); however, we prefer to

recognize it until examination of the holotype, and study of more isolates and

their molecular data. T’ trachyspermus var. macrocarpus is distinguished from

T. trachyspermus by presenting bigger cleistothecia (500-1500 vs 50-350 um)

and ascospores (4-4.5 x 2.8-3.2 vs 3-3.5 x 2-2.5 um).

Talaromyces trachyspermus (Shear) Stolk & Samson, Stud. Mycol. 2: 32 (1973)

var. trachyspermus

CoLonigs on MEA, 25 °C, 14 d, moderately deep, plane, granular, 42-50

mm diam., white to cream-colored; exudate absent; reverse light orange and

orange to brown in the centre.

CLEISTOTHECIA superficial, mainly globose, usually confluent, 150-350

um diam., at first white becoming cream-colored in age, covered by a hyphal

network. Ascr globose to ovoid, 8-spored, 7-9 x 6-8 um, evanescent.

AscosPorss ellipsoidal, 3.5-4 x 2-3 um, spinulose. CONIDIOPHORES slender

monoverticillate and biverticillate, short. METULAE 8-11 x 2 um. PHIALIDES

2-5 in verticils, lanceolate, tapering to neck, 9-10 x 2 um. Conrp1a ellipsoidal,

2-2.5 x 2 um, smooth-walled.

CULTURES EXAMINED—ARGENTINA. CaTAMARCA: RP 46, 27°35'13”S 66°22’11”W,

996 m a.s.l., 9.1.2009, leg. SMR, identified by SMR & AIR, isolation 21.11.2011

(BAFCcult 4619; GenBank LT968849); (BAFCcult 4620; GenBank LT968850);

28°46'22”S 66°09'56”W, 1317 m a.s.l., 24.VIII.2011, leg. SMR, identified by SMR &

AIR, isolation 4.1.2012 (BAFCcult 4629); Belén, 27°07’33”S 66°56'36’W, 2152 m a.s.L.,

25.VIII.2011, leg. SMR, identified by SMR, isolation 27.1.2012 (BAFCcult 4630); RP 47,

km 35.5, 27°26'50”S 66°24’26”W, 2700 m a.s.l., 25.VII.2011, leg. SMR, identified by

SMR, isolation 18.1.2012 (BAFCcult 4631).

DIsTRIBUTION—Argentina, Bangladesh, France, Germany, India, Japan, Nepal,

Pakistan, South Africa, Tahiti, Uganda, United States of America (Domsch &

al. 2007).

Hasitat—Soil, cereals and grains, dung (Domsch & al. 2007).

CoMMENTS—The morphological features of our isolates agreed with the

original description of Talaromyces trachyspermus (Stolk & Samson 1972). In

Heat resistant ascomycetes in Argentina ... 551

Argentina Bertoni & al. (1973) and Godeas (1975) isolated this fungus from

soil.

Trichocladium pyriforme M. Dixon,

Trans. Br. Mycol. Soc. 51: 160. 1968. FIG. 1R-T

Co.tontges on MEA, a 25 °C, 7 d, plane, slightly floccose, 36-40 mm diam.,

grey green to olive, darkening in age; reverse very dark; covering the whole

culture plate in 21 d.

Hypuae hyaline when young, yellow brown in age, 2-5 um wide, smooth.

CHLAMYDOSPORES (holothallic conidia) terminal, pyriform, 3-4 celled, 16-19 x

6-7.5 um, smooth-walled; distal cell dark brown, with an acute apex and

germinative pore.

CULTURES EXAMINED—ARGENTINA. CaTAMaARCA: RN 60, km 934, 29°33’35’S

64°52’56’W, 186 ma.s.l., 5.1.2009, leg. SMR, identified by SMR & AIR, isolation 5.1.2011

(BAFCcult 4623); RN 40, km 4243.5, 26°51’23”S 66°05’52”W, 2031 ma.s.l., 25.VHIL2011,

leg. SMR, identified by SMR & AIR, isolation 12.X1.2011 (BAFCcult 4611).

DISTRIBUTION—Czech Republic (Mantle & al. 2006), Ireland (Dixon 1968).

Hasitat—Soil (Dixon 1968, Goh & Hyde 1999).

CoMMENTS—The morphological features of the Argentinian strains of

Trichocladium pyriforme agreed with those described by Dixon (1968) and by

Goh & Hyde (1999). Domsch & al. (2007) pointed out the remarkable thermal

resistance of the spores of this species, and Dixon (1968) stated that it tolerated

up to 95 °C for 10 min. Bollen (1969) isolated T. pyriforme from greenhouse

soil using a thermal shock at temperatures greater than 55 °C. This is the first

report of this fungus in Argentina.

Trichoderma saturnisporum Hammill, Mycologia 62: 112. 1970.

COLONIES on special nutrient agar (SNA), 72 h, 30 °C, in darkness,

somewhat annellated, 38-44 mm diam., green. At 35 °C, 72 h, 38 mm diam.

On PDA, 72 h, 30 °C, slightly annellated, 60 mm, dull green; at 35 °C, 69 mm

diam. in darkness.

CONIDIOPHORES associated in tufts up to 4 mm diam., branched, short

sterile projections present. PHIALIDES mainly in verticils, cylindrical, 8.5-11

x 3.3-4 um. Conrp1A subglobose to broadly ellipsoidal, 4-4.8 x 2.9-3.7 um,

greenish, walls with few notorious warts. CHLAMYDOSPORES not observed.

CULTURES EXAMINED—ARGENTINA. CATAMARCA: RN 60, km 1016, 29°30'04’S

65°37'57”W, 237 m a.s.l., 5.1.2009, leg. SMR, identified by SMR & RC, isolation

3.VHI.2010 (BAFCcult 4584, 4586, 4587); RP 25, 28°14’36”S 66°08’51”W, 1507 m

a.s.l., 6.12009, leg. SMR, identified by SMR & VB, isolation 1.X1.2010 (BAFCcult 4626);

552 ... Romero &al.

Rincon, 28°22’03"S 66°13’39’W, 8.1.2009, leg. SMR, identified by SMR & VB, isolation

19. VIII.2010 (BAFCcult 4627, 4628.

DIsTRIBUTION—Argentina (Godeas & al. 1977), Australia, Italy, South Africa,

Turkey, United States (Samuels & al. 1998).

HaBitaT—Soil.

CoMMENTS—The morphological features of the examined culture agreed

with those given by Samuels & al. (1998). Trichoderma saturnisporum is easily

recognized by conidia with large warts mimicking wings, which give a saturnian

appearance to the conidia.

Discussion

In previous papers we described two new species of Eurotiales, Talaromyces

systylus S.M. Romero & al. (Romero & al. 2016) and Aspergillus fuscicans S.M.

Romero & al. (Romero & al. 2018) from the same locations and substrate (soil)

as in the present work. ‘The isolates in Aspergillus sect. Fumigati, which were

prevalent in the analyzed samples, are currently under study.

Some of the fungal species found have been previously cited as heat-resistant,

such as Arthrinium phaeospermum (Kashiwagi & al. 2009, Pitt & Hocking

2009), Gilmaniella humicola (Bollen 1969, Jesenska & al. 1992), Leiothecium

ellipsoideum (Samson & Mouchacca 1975), Talaromyces macrosporus (Beuchat

1988, Frison & al. 2012) and Trichocladium pyriforme (Bollen 1969, Dixon

1968).

It was not possible to obtain information about the heat-resistance of

Aspergillus montevidensis; however, Aspergillus glaucus (L.) Link, another

species in A. sect. Aspergillus, was isolated from contaminated jellies and

grape marmalades, and its thermal resistance was analyzed (Splittstoesser &

al. 1989). No information about the heat resistance was found for Trichoderma

saturnisporum; however, chlamydospore production (Samuels & al. 1998)

would explain its survival after a thermal shock performed in the lab.

Byssochlamys nivea, perhaps the most well-known heat-resistant fungal

species, was not isolated during this study. This could be attributed to our

thermal treatment (75 °C, 30 min), since other authors have determined that

its ascospores can survive at 70 °C for 60-75 min but not at 80 °C for 1 min

(Pieckova & al. 1994). The same authors also mentioned that at 80 °C the

ascospores of Hamigera avellanea Stolk & Samson survive 120 min, whereas

the conidia of Gilmaniella humicola survive between 8-14 min. Although no

data were found on the thermal resistance of H. paravellanea and H. terricola

ascospores, given the heat resistance described for H. avellanea (Pieckova & al.

Heat resistant ascomycetes in Argentina ... 553

1994, Scaramuzza & Berni 2014), it can be assumed that other species of the

genera would possess a similar capacity.

Acknowledgements

S.M. Romero and A.I. Romero thank the Consejo Nacional de Investigaciones

Cientificas y Técnicas (CONICET-Argentina), PIP1086 and PICT-2018-03781 for

the financial support to perform the present study. In addition, the Instituto de

Microbiologia y Zoologia Agricola (Instituto Nacional de Tecnologia Agropecuaria)

is particularly recognized for the provision of supplies and facilities to carry out

this work. The authors acknowledge Dr. Maria Virginia Bianchinotti (CERZOS-

CONICET, Universidad Nacional del Sur, Argentina) and Dr. Andrew Miller (Illinois

Natural History Survey, University of Illinois, U.S.A.) for reading and improving the

manuscript as pre-submission reviewers. The insightful and careful review of the

Nomenclature Editor, Dr. Shaun R. Pennycook, is especially appreciated. The design

and preparation of the plate by the technician Mariana Valente (InMiBo-CONICET)

is held in high esteem.

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MYCOTAXON

July-September 2020—Volume 135, pp. 559-568

https://doi.org/10.5248/135.559

Cortinarius rapaceoides, a new record for Turkey

MERYEM SENAY SENGUL DEMIRAK * & HAKAN IsIk?

'Department of Molecular Biology and Genetics, Faculty of Arts and Sciences,

Tokat Gaziosmanpasa University, Tasliciftlik Campus, 60250, Tokat, Turkey

?Tokat M. Emin Sarac Anatolian Religious High School, 60030, Tokat, Turkey

*CORRESPONDENCE TO: senay.sengul@gop.edu.tr

ABSTRACT—A macrofungus collected from Akbelen village in Tokat has been identified

based on molecular evidence and morphology as Cortinarius rapaceoides, a new record from

Turkey. Sequence data generated from the nuclear ribosomal internal transcribed spacer

and ribosomal large subunit regions were deposited in GenBank and used to determine

phylogenetic relationships within Cortinarius subg. Phlegmacium. A morphological

description, illustrations, and phylogeny of the Turkish C. rapaceoides are provided.

Key worps—Cortinariaceae, ITS, LSU, taxonomy

Introduction

The ectomycorrhizal basidiomycete genus Cortinarius (Agaricales) is

the world’s most speciose mushroom genus. Ectomycorrhizal associations

formed with trees (e.g., Nothofagus, Quercus, Pinus) and herbaceous plants

are important for forest ecosystem (Moser & Horak 1975, Breitenbach &

Kranzlin 2000, Stensrud & al. 2014, Itoo & al. 2015). Morphological features

such as cortina covering young gills, rusty brown mature gills, ochraceous-

rusty to deep rusty spore print, and bulbous stipe distinguish this genus.

Species separation within Cortinarius is greatly complicated by a high

degree of inter- and intra-specific morphological variation.

Molecular markers have become important in helping resolve and

refine Cortinarius taxonomic classification. Previous studies have used

molecular markers derived from ribosomal gene loci containing the internal

560 ... Sengiil Demirak & Isik

transcribed spacer regions (ITS1 and ITS2), ribosomal large subunit (LSU),

and the two largest subunits of RNA polymerase II (RPB1, RPB2) (Garnica

& al. 2003, 2009; Froslev & al. 2005, 2007; Niskanen & al. 2009; Liimatainen

& Niskanen 2013; Stensrud & al. 2014; Liimatainen & al. 2017; Sesli &

Liimatainen 2018). Among those, ITS data strongly supports phylogenetic

delimitation of species among Cortinarius sections and our understanding

of the evolution of the genus (see Garnica & al. 2009). Most studies stress the

necessity of combining sequence data with morphological studies to resolve

the relationships among Cortinarius species.

Cortinarius species have been previously recorded in Turkey (Sesli &

Denchev 2014; Akata & al. 2015; Giingor & al. 2015; Sesli & al. 2015, 2016;

Sesli 2018; Sesli & Liimatainen 2018; Kalmer & al. 2019). Our research

provides the first identification of C. rapaceoides in Turkey with detailed

description of morphological characteristics and evidence at the molecular

level.

Materials & methods

Morphological studies

Fresh Cortinarius basidiomes were collected from Akbelen village (Tokat) in

the autumn of 2018, photographed in the field, and important morphological and

ecological features noted. Basidiomes were stored in paper bags and kept in a box.

The fresh collection was transported to the laboratory in paper bags, and collection

numbers were assigned. The dried basidiomes were placed into polyethylene bags

for storage. Tissues were mounted in Melzer’s reagent, distilled water, Congo red,

and KOH prior to examination under a light microscope. At least 25 measurements

from each microscopic structure were obtained from a mature basidiome, which

was initially identified based on its morphological features with the help of Boisselet

(2012a) and Tanchaud (2016). The identified collection was deposited in the

Fungarium of the Department of Biology, Tokat Gaziosmanpasa University, Tokat,

Turkey (GOPUF).

Molecular studies

Genomic DNA was extracted from lamella using the ZR Fungal/Bacterial DNA

MiniPrep kit as described by the manufacturer's protocol. The primer pair ITS4-

ITS5 (White & al. 1990) was used to amplify ITS1-5.8S-ITS2 region and the primer

pair LROR-LRS (Vilgalys & Hester 1990) was used to amplify 28S LSU rRNA gene

region. The DNA was amplified in a 30 ul volume mixture containing 3 ul 10X buffer,

3 ul dNTP mix, 3 ul degenerate primer pair (final concentration of 1 uM each), 0.3 ul

Dream Taq DNA polymerase (Thermo), 10 ul gDNA and 7.7 ul sterile ddH,O. PCR

amplification for ITS region was set as: A negative control reaction using only sterile

ddH,O was also run in parallel. 5 min initial denaturation at 95°C + 40 denaturation

Cortinarius rapaceoides new for Turkey ... 561

cycles (95°C for 30 sec) + annealing (53°C for 30 sec) + extension (72°C for 1 min)

and a final extension for 10 min. The LSU PCR protocol comprised: a 3 min initial

denaturation at 95°C + 40 denaturation cycles (95°C for 30 sec) + annealing (48°C

for 30 sec) + extension (72°C for 1 min) and a final extension for 10 min.

PCR products were run in a 1% agarose gel electrophoresis and positive PCR

products were gel purified by using Wizard SV Gel and PCR Clean-up System.

Purified PCR products were sequenced in both directions using the same primer

pairs by BM Labosis Inc. (Ankara).

Sequence and phylogenetic analysis

Chromatograms were checked for any nucleotide errors for each genomic

sequence generated from both ends and errors were corrected manually. Assembled

rDNA sequences were examined using Basic Local Alignment Search Tool (BLAST)

programme to select the most closely related ITS and LSU sequences in the database.

Representative ITS and LSU sequences of Cortinarius species were retrieved from

GenBank for phylogenetic analysis. The sequence alignments and phylogenetic trees

for each genomic region were analysed using MEGA 6.0 (Tamura & al. 2013). In the

CLUSTALW alignment, sequences were trimmed at both ends. Phylogenetic trees

were constructed using the maximum likelihood (ML) and maximum parsimony

(MP) methods. Tamura-Nei model (Tamura & Nei, 1993) was used to construct

the ML tree with bootstrap support of 1000 replicates and default settings. The

bootstrap support values >50% were marked on the branches of the tree.

Taxonomy

Cortinarius rapaceoides Bidaud, G. Riousset & Riousset,

Micologia 2000: 68 (2000) Fic. 1

PitEus 50-100(-120) mm diam., at first convex then plane-convex to

plane; surface dull when dry, lubricous when moist; colour overall yellowish

to ochre-yellow, in the centre orange-yellow with darker brown squamules.

LAMELLAE pale lilac, soon rust-brown, broad, crowded, edges smooth to

slightly crenate. Stipe whitish, sometimes gray-purplish at the apex, fragile,

solid, cylindrical, base with a marginate bulb, surface longitudinally whitish-

fibrillose when young, brownish when old. FLEsH whitish sometimes bluish

in the upper and lower stipe. KOH negative on the flesh, amber yellow colour

on the pileal surface. ODorR of chocolate.

BASIDIOSPORES (9.6—)10-13(-13.8) x (6.0-)6.5-8(-8.5) um, elliptical to

amygdaliform, strongly verrucose, yellow-brown. BasIpIA, (30—)35-40(-44.6) x

(9-)10.5-12(-12.4) um, with 2-4 sterigmata, clavate.

SPECIMENS EXAMINED: TURKEY, TOKAT, Akbelen village, 40°27’0”N 36°39'16’E,

1050 m, among dead leaves of Quercus sp, 8 November 2018, leg. Isik 798 (GOPUF;

GenBank MN094878, MN099705).

562 ... Sengiil Demirak & Isik

Cortinarius rapaceoides new for Turkey ... 563

HABITAT & DISTRIBUTION—Among fallen leaves on calcareous soils in

deciduous woods. Previously recorded from France and Italy (Liimatainen

& al. 2014).

Molecular phylogeny

The ITS1-5.8S-ITS2 (666 bp) and LSU rRNA (996 bp) sequences were

deposited in GenBank. ITS sequence MN094878 BLAST results matched

closely with the sequences representing C. caroviolaceus (EU057049),

C. rapaceoides (KF732407, ex-holotype), and C. saporatus (DQ663413),

which are included in the /Caroviolacei clade (Garnica & al. 2009).

The remaining BLAST hits were also matched with species within the

same clade. Accordingly, 67 sequences in the /Calochroi & Fulvi clade of

Cortinarius subg. Phlegmacium (Liimatainen & al. 2014) were selected and

ITS sequences were retrieved from GenBank for phylogenetic analyses. The

outgroup Cortinarius aureifolius and C. caerulescens were used to root the

phylogeny (Froslev & al. 2005, Garnica & al. 2009).

Although fewer in number, LSU sequences for genus Cortinarius were

also selected to construct a phylogenetic tree, but the LSU-based phylogeny

was not well resolved due to low bootstrap values and no phylogenetic

separation at the subgenus level was observed for LSU regions.

Our two constructed MP and ML phylogenetic trees showed similar

topologies for each genomic region. We present only the ML tree to indicate

phylogenetic relationship of the studied species based on the ITS region.

The sequence from our Cortinarius specimen clustered with sequences

derived from C. rapaceoides described from Italy and France (including

the ex-holotype sequence KF732407) and C. caroviolaceus (EU057049)

from Italy with high bootstrap support (Fic. 2). Liimatainen & al. (2014)

suggested that C. caroviolaceus sequence actually represented C. rapaceoides

based on their ITS sequence phylogeny; and our phylogeny also supports it

as conspecific with C. rapaceoides.

Discussion

Cortinarius rapaceoides, originally described from France and collected

under Quercus ilex in a deciduous forest with calcareous soil, is characterized

by a cream to yellow color, smooth convex pileus with a rolled margin,

Fic. 1. Cortinarius rapaceoides (GOPUF - Isik 798): A. basidiomata; B. basidia and basidioles;

C. basidia; D. basidiospores. Scale bars: A = 3 cm; B-D = 10 um.

564 ... Sengiil Demirak & Isik

cream to purple lamellae, blue, cream or purple colored cylindrical stipe

with a marginate bulb, 10-13 x 6-7 um verrucose spores, and a hardwood

mycorrhizal association (Boisselet 2012a, Tanchaud 2016).

‘The species is morphologically similar to C. aleuriosmus Maire [= C. caro-

violaceus P.D. Orton] and C. saporatus Britzelm. These fungi may be

confused with each other due to their similar ecological features; however,

C. aleuriosmus can be distinguished from C. rapaceoides by its whitish pileus

and lamellae, smaller, almond-to lemon-shaped rough spores, abundant

white veil, and farinaceous taste and odor (Orton 1955, Moser 1983, Phillips

1981, 2013; Boisselet 2012a, 2012b; Tanchaud 2016). Cortinarius saporatus

differs in its bigger pileus and stipe, dark brown colour changes with KOH

on the pileus, its yellowish colour at the stipe base, and an odor that is

initially fruity but later more sour (Phillips 1981, 2013). Although some

spores of Turkish C. rapaceoides are larger, the morphological characters

are very similar and agree with French C. rapaceoides (Boisselet 2012a,

Tanchaud 2016).

Sequence analyses also support identification of Turkish collection as

C. rapaceoides. It belongs in Cortinarius subg. Phlegmacium, a polyphyletic

subgenus (Peintner & al. 2004, Garnica & al. 2005). Although Cortinarius

species are morphologically quite variable, their ITS sequences show a

high sequence identity and are reliably used for species delimitation and

identification (e.g. Froslev & al. 2005, 2007; Ortega & al. 2008; Garnica &

al. 2009; Niskanen & al. 2009). We found the ITS region more useful than

the LSU region for inferring the phylogenetic relationships and were able to

determine independent phylogenetic lineages with high boostrap support.

Our ITS sequence clustered in Cortinarius sect. Calochroi and showed

100% sequence identity with C. rapaceoides from France and Italy without

any intraspecific sequence polymorphisms.

Our first record of C. rapaceoides in Turkey is well supported by both

morphological and molecular evidence.

Acknowledgments

We would like to thank Dr. Ibrahim Turkekul for critically reviewing this article.

We also thank the referees, Drs Abdullah Kaya (Gazi University, Ankara, Turkey)

Fic. 2. Phylogeny of selected Cortinarius species inferred from ITS sequences using the

ML method. The diamond [@] indicates the Turkish C. rapaceoides identified in this study.

Ex-type sequences are annotated with [T]. Cortinarius aureifolius and C. caerulescens represent

the outgroup. Bootstrap support values =>50% from ML analysis are shown on the branches.

Bar = 0.02 expected changes per site per branch.

Cortinarius rapaceoides new for Turkey ... 565

67 Cortinarius caesiocinctus DQ663239 [T]

Cortinarius caesiocinctus DQ663241

63 | Cortinarius cobaltinus KF673472

Cortinarius cobaltinus KF673471

Cortinarius cobaltinus KF673470 [T]

Cortinarius bigelowii KF732265 [T]

97, Cortinarius metarius KF732347 [T]

Cortinarius barbarorum DQ663236

Cortinarius flavipallens KF732554 [T]

Cortinarius olympianus KF732364 [T]

74' Cortinarius olympianus KF732553

99 | Cortinarius sublilacinopes KF732561 [T]

Cortinarius sublilacinopes DQ663434

Cortinarius pseudogracilior KF732394 [T]

Cortinarius calojanthinus KF732272 [T]

73' Cortinarius corrosus DQ663281

Cortinarius arenicola KF732252 [T]

Cortinarius frondosophilus KF732562 [T]

96! Cortinarius platypus KF732563 [T]

Cortinarius magnivelatus EU056976

Cortinarius spectabilis DQ663425

Cortinarius spectabilis DQ663426

99 Cortinarius sannio KF732536

Cortinarius sannio KF732537

76, Cortinarius pseudoglaucopus DQ663395

97| | Cortinarius pseudoglaucopus AY669573

Cortinarius elotoides KF732300 [T]

Cortinarius rapaceoides DQ663417

Cortinarius caroviolaceus EU057049

99| | Cortinarius rapaceoides KF732407 [T]

74 | Cortinarius rapaceoides NR130253

@ Cortinarius rapaceoides MN094878

Cortinarius cacodes KF732270 [T]

80 ; Cortinarius saxamontanus KF732421 [T]

Cortinarius saxamontanus EU057026

Cortinarius cf. aureofulvus KF732544

Cortinarius cf. aureofulvus KF732545

Cortinarius cf. aureofulvus KF732543

Cortinarius sulphurinus var. fageticola DQ663439

: Cortinarius sulphurinus KC842431

Cortinarius sulphurinus DQ663437

Cortinarius luteicolor KF732546

Cortinarius luteicolor KF732547

Cortinarius luteicolor FJ717511

Cortinarius citrinipedes KF732281 [T]

Cortinarius osmophorus DQ663368

Cortinarius evosmus KF732302 [T]

9 Cortinarius osmophorus AY174815

Cortinarius cupreorufus KF732548

Cortinarius cupreorufus KF732549

98 | Cortinarius cupreorufus KF732550

Cortinarius cupreorufus KF732294 [T]

96 Cortinarius flavobulbus EU057017

Cortinarius flavobulbus KF732305 [T]

98 , Cortinarius viridirubescens KF732476 [T]

Cortinarius viridirubescens EU057007

Cortinarius dibaphus DQ663286

79, Cortinarius alnobetulae EU655672

Cortinarius alnobetulae KF732246 [T]

Cortinarius amnicola KF732249 [T]

Cortinarius subpurpureophyllus KF732450 [T]

Cortinarius subpurpureophyllus KF732557

79| Cortinarius napus GU363492

Cortinarius napus KC842428

Cortinarius cf. meinhardii KF732551

96 | Cortinarius meinhardii AY174840

Cortinarius parafulmineus EF014269

99! Cortinarius parafulmineus KF732552 [T]

Cortinarius caerulescens AY174863

Cortinarius aureifolius AF268893

566 ... Sengiil Demirak & Isik

and Yusuf Uzun (Van Yuzuncu Yil University, Turkey), for their helpful comments

and suggestions. We greatly appreciate Dr. Lorelei Norvell’s editorial review and

Dr. Shaun Pennycook’s nomenclatural review.

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MYCOTAXON

July-September 2020—Volume 135, pp. 569-578

https://doi.org/10.5248/135.569

New records of Toninia from China

CoNGCONG Mi1Ao*', MEIJIE SUN*’,

XIAO ZHANG!, ZHAOJIE REN’, LING Hu?

' Key Laboratory of Plant Stress Research, College of Life Sciences,

Shandong Normal University, Jinan, 250014, P. R. China

? Shandong Museum, Jinan, 250014, P. R. China

3 Institute of Environment and Ecology,

Shandong Normal University, Jinan, 250014, P. R. China

“ CORRESPONDENCE TO: ccmjy123@163.com

ABSTRACT—Four Toninia taxa (T. albilabra, T: poeltii, T. tristis subsp. Arizonica, and T. tristis

subsp. fujikawae) are reported for the first time from China. Descriptions with morphological

and chemical characters and known distribution are given for each taxon. A key to the species

of Toninia s.l. in China is also provided.

KeEyYworps—lichen-forming fungi, Lecanorales, Ramalinaceae, taxonomy

Introduction

Toninia, widely distributed in drought and arid areas, is an important

component of desert lichen communities (Timdal 1991, 2002). The current

delimitation of the genus is based mainly on the features of the asci and

paraphyses (Meijie & al. 2019) and molecular genetic characters (cf. Kistenich

& al. 2018, Ekman 2001, Ekman & al. 2008). Toninia is characterized

by a crustose to squamulose and epruinose to densely pruinose thallus;

lecideine, epruinose to densely pruinose apothecia; dark brown to colourless

hypothecium; usually colourless, grey, green, or brown epihymenium and

exciple that changes colour in K and N; clavate, Biatora-type, 8-spored asci;

and elliptical to spindle-shaped, colourless, 1-8-celled ascospores. Toninia

* ConGCONG Miao & MEWIE SUN contributed equally to this work.

570 ... Miao, Sun & al.

was previously placed in Lecideaceae (Massalongo 1852; Timdal 1991;

Zahlbruckner 1890). With the advent of the molecular age, phylogenetic

analysis of lichens is carried out in combination with their morphological

and anatomical features; currently, Toninia is classified in Ramalinaceae

(Timdal 2002; Ekman 2001).

In this paper we contribute to the knowledge of Toninia in China and

add accurate data for the Lichen Flora of China project by reporting on four

taxa new to the country: T. albilabra, T: poeltii, T: tristis subsp. arizonica and

T: tristis subsp. fujikawae.

Materials & methods

The specimens studied are preserved in the Lichen Section of Botanical

Herbarium, Shandong Normal University, Jinan, China (SDNU) and the

Lichen Herbarium of the Kunming Institute of Botany, Kunming, China

(KUN). The morphological and anatomical characters of the specimens were

examined under a COIC XTL7045B2 stereomicroscope and an Olympus CX41

polarizing microscope. Thallus and medulla were tested with K (a 10% aqueous

KOH solution), Cl (a solution of aqueous NaOCl), and I (Lugol’s iodine) for

identification. Lichen substances were identified using standardized thin

layer chromatography (TLC) techniques with system C (Orange & al. 2010).

Photographs were taken with Olympus SZX16 and BX61 microscope with a

DP72 camera.

Taxonomy

Toninia albilabra (Dufour) H. Olivier, Bull. Géogr. Bot. 21: 196 (1911) Fic. 1

THALLUS squamulose, indeterminate. Squamules <2(-4) mm diam.,

scattered or contiguous to irregularly imbricate, orbicular or sometimes

slightly lobed, weakly concave to weakly convex. Upper side reddish brown,

usually epruinose, shiny, with regular and usually deep fissures in the cortex,

pores and pseudocyphellae absent. Margin partly to entirely white pruinose.

Underside white to pale brown. Algal layer continuous. Lower cortex poorly

developed or absent.

APOTHECIA $1.5(-2.5) mm diam., weakly concave to weakly convex,

usually persistently (but narrowly) marginate, densely white pruinose or

more rarely epruinose. Proper exciple reddish brown throughout or with an

additional grey pigment near the rim, lacking crystals. Hypothecium medium

brown in upper part, pale brown to colourless in lower part. Hymenium 60-70

um high; epithecium grey, K+ violet, N+ violet, containing crystals of calcium

oxalate. Spores broadly to narrowly fusiform, 1-septate, 13.5-22.5 x 3-4 um.

Toninia taxa new for China... 571

aN Baa Dh. fh «

ao iy,

‘ > P

Why

Fic. 1. Toninia albilabra (15-49721 KUN).

A. Thallus; B. Epihymenium; C. Purple K reaction of epihymenium; D. Ascospores.

PYCNIDIA not seen.

CHEMISTRY— Terpenoids detected by TLC.

SPECIMENS EXAMINED—CHINA. TIBET, Baju County, on the way from Bangda

to Bangda Airport, alt. 4129 m, 17 Sep. 2014, Wang Lisong & al. 14-47025 (KUN);

Jiangda County, Tongpu Township, 317 national highway side slope, 31°38’33”N

98°26'10’E, alt. 3940 m, 5 Oct. 2016, Wang Lisong & al. 16-51356 (KUN). YUNNAN,

Zhongdian, Benzilan Town, 28°10.47’N 99°23.01’E, alt. 2115 m, 27 Aug. 2006,

Wang Li Song & al. 06-26674 (KUN); Deqin, Baima Snow Mountain, 28°19.47’N

99°05.13’E, 2 Nov. 2015, alt. 4350 m, Wang Lisong & al. 15-49721 (KUN).

DisTRIBUTION—On rock or soil. Toninia albilabra has been reported in

Europe, North Africa, Macaronesia, and the Middle East (Timdal 1991).

New to China.

COMMENTS— The Chinese material closely matches the previously published

description by Timdal (1991). Toninia albilabra is morphologically similar to

T: sedifolia, which differs mainly by having a smooth upper cortex, a thinner

epinecral layer, and farinose pruina. The white edge of the squamules in

572 ... Miao, Sun & al.

T. sedifolia is caused by pruina on the cortex surface and is not sharply

delimited from the less pruinose inner part of the upper side. The white edge in

T: albilabra, however, is caused by the white medulla being visible outside the

sharply delimited, usually entirely epruinose, upper cortex (Timdal 1991).

Fic. 2. Toninia poeltii (16-53317 KUN).

A. Thallus and apothecia; B. Epihymenium; C. Purple K reaction of epihymenium; D. Ascospores.

Toninia poeltii Timdal, Opera Bot. 110: 85 (1991) Fic. 2

Thallus squamulose, indeterminate. Squamules <1 mm diam. More or

less continuous crust. Upper side dark greyish brown, epruinose, dull, with

a few shallow fissures in the cortex, lacking pores and pseudocyphellae.

Margin concolorous with upper side, epruinose. Upper cortex 30-80 um

thick, lacking crystals. Algal layer continuous. Medulla lacking crystals.

Lower cortex <40 um thick, brown pigment hyphae.

Apothecia <0.8 mm diam., plane and indistinctly marginate when young,

later more or less convex and immarginate, epruinose. Proper exciple

olivaceous green in the rim, colourless in inner part, K+ violet or brown,

N-, lacking crystals. Hypothecium colourless. Hymenium 60-70 um high;

Toninia taxa new for China... 573

epithecium olivaceous green, K+ violet or brown, N-. Spores bacilliform,

often slightly curved, 1-3-septate, 12-21.5 x 4-5 um.

Pycnidia not seen.

CHEMISTRY—Terpenoids detected by TLC.

SPECIMENS EXAMINED—CHINA. TIBET, Mangkang, on the way from Lawushan to

the town of Rumei, alt. 4290 m, 20 Sep. 2016, Wang Lisong & al. 16-53317 (KUN);

Dingri County, Alpine meadow, alt. 5250 m, 22 Aug. 2007, Wang Lisong & al. 07-

28528 (KUN). YUNNAN, Deqin County, Sola Mountain, 28°38.19’N 98°36.30’E, alt.

4800 m, 10 Sep. 2012, Niu Dongling & al. 12-35865 (KUN).

DiIsTRIBUTION—On soil or rock. Reported only in the Himalayas (Timdal

1991). New to China.

COMMENTS— The Chinese material closely matches the previously published

description by Timdal (1991). Toninia poeltii is morphologically similar to

T. squalida (which has not been recorded for China), but differs in having

an olivaceous green, K+ violet/brown, N- pigment in the epithecium and

exciple rim. Toninia poeltii has at most 3-septate spores, but T. squalida has

3-7-septate spores (Timdal 1991).

Toninia tristis subsp. arizonica Timdal, Opera Bot. 110: 112 (1991) Fia. 3

THALLUS squamulose, squamules <6(-8) mm diam., scattered to adjacent,

rounded, bullate but often with an irregular depression. Upper surface

castaneous brown to dark brown, epruinose, dull, smooth in the cortex, lacking

pseudocyphellae. Upper cortex <160 um high, lacking crystals.

APOTHECIA <6 mm diam., plane and weakly convex, distinctly marginate,

epruinose. Proper exciple brown to dark brown, colourless in inner part.

Epithecium brown, partly with a green tinge. Spores simple, narrowly ellipsoid

to fusiform, 8-17 x 4-4.5 um. Orange pigment present in the upper part of the

hypothecium and in the lumina of many paraphyses, asci, and spores; yellow

pigment present in the upper part of the hypothecium.

CHEMISTRY—Terpenoids and fatty acid detected by TLC.

SPECIMENS EXAMINED—CHINA. INNER MONGOLIA, Urad Rear Banner, Huhe

Bash GeTusheng, alt. 1600 m, 19 Aug. 2011, Tong Debao 20123675 (SDNU).

NINGXIA, Jingyuan County, Migang Mountain, alt., 2300 m, 20 Jun. 2011,

Cheng Yuliang 20116001 (SDNU). Sicnuan, Ningnan County, Baihetan Town,

alt. 701 m, 18 Oct. 2013, Wang Lisong & al. 13-39450 (KUN); Jinsha River,

alt. 1550 m, 20 Apr. 2014, Wang Lisong & al. 14-43367 (KUN). TIBET, Naqu

County, 317 country road side slope, alt. 4680 m, 30 Sep. 2016, Wang Lisong

& al. 16-51961 (KUN). YUNNAN, Yunmin County, Jinsha River Valley, alt. 920

m, 27 Nov. 2014, Wang Lisong & al. 14-46464 (KUN); Dongchuan, Sandstone

from Dongchuan to Qiaojia Road, alt. 1080 m 11 May 2017, Wang Lisong & al.

17-55053 (KUN).

574 ... Miao, Sun & al.

Fic. 3 Toninia tristis subsp. arizonica (16-53317 KUN).

A. Thallus; B. Epihymenium; C. Ascus; D. Ascospores.

DISTRIBUTION—On soil. Arizona, USA (Timdal 1991). New to China.

ComMMENTS—The Chinese material closely matches the previously

published description by Timdal (1991). Toninia tristis subsp. arizonica

produces orange asci similar to T! tristis subsp. asiae-centralis, which differs

in having 1-septate spores (Timdal 1991). In its possession of simple spores,

the Chinese material closely matches previously published descriptions

of T. tristis subsp. pseudotabacina, which differs in its absence of orange

pigments (Timdal 1991).

Toninia tristis subsp. fujikawae (M. Satd) Timdal,

Opera Bot. 110: 113 (1991) Fic. 4

Thallus squamulose, squamules <4(-5) mm diam., continuous, bullate,

often forming cushions. Upper side castaneous brown to dark brown,

lacking pseudocyphellae. Margin concolorous with upper side, epruinose.

Upper cortex not containing crystals. Algal layer continuous. Medulla

Toninia taxa new for China... 575

lacking crystals. Lower cortex resembling upper cortex, but stainable layer

often dark reddish brown.

Apothecia <4 mm diam., plane and weakly convex, distinctly marginate,

epruinose, Proper exciple brown to dark brown, colourless in inner part, K-,

N-, lacking crystals. Hypothecium pale brown, 60-70 um high; epithecium

olivaceous green to bright green, K+ violet or brown, N-. Spores simple,

ellipsoid, 7-12 x 3-5 um. Orange and yellow pigments lacking.

CHEMISTRY—Terpenoids and fatty acid detected by TLC.

SPECIMENS EXAMINED—CHINA. SICHUAN, Huili County, Jinsha River, alt. 1550 m, 20

Apr. 2014, Wang Lisong & al. 14-43349; 14-43368 (KUN); TiBET, Wuqi County, next to

the 214 state road, 31°09’51”N 96°37'54’E, alt. 3840 m, 2 Oct. 2016, Wang Lisong & al.

16-51390 (KUN); Baqing County, Laxi village, 317 national road side slope, 31°50’17”N

94°23’24’E, alt. 4180 m, 1 Oct. 2016, Wang Lisong & al. 16-53934 (KUN). YUNNAN,

Luquan County, alt. 2540 m, 19 Apr. 2014, Wang Lisong & al. 14-43234 (KUN);

Deqin County, Benzilan, 28°1135.54”N 99°21’08’E, alt. 2112 m,19 Aug. 2018, Wang

Chunxiao & al. 20180326 (SDNU).

DiIsTRIBUTION—On rock or soil. Japan (Timdal 1991). New to China.

CoMMENTS— Toninia tristis subsp. fujikawae may be confused with T. tristis

subsp. canadensis in having green epithecium, but the latter subspecies has

mainly 1-septate ascospores (Timdal 1991). Although also characterized

by simple spores, T! tristis subsp. arizonica is distinguished by a brown

epithecium (Timdal 2002).

Key to the species of Toninia s.|. in China

Kistenich & al. (2018) place some species below in Thalloidima (T. opuntioides,

T: physaroides, T. sedifolia) or other genera (e.g., T: gobica, T. poeltii). Here we follow

Timdal (1991) and treat all species in Toninia s.1.

ie Thalliis proinose (arinose.or-cranulaty .) wees lie ete scorn ae eels as ae 2

i thallusseprumeseto faintly prtimoses £2 4 5.53 2 5.x Alay ton eschg eo nes oaoraae eget 14

22 Epitheciom: prays PIO NG VAIOlet th. nres WN An Reh cba eieee MO sear ece 2 3

2. Epithecium brown, K+ red, or epithecium dark green to bright green,

Ka, N+ Violet® oes iG cage ee eee! eens Hae bios ate ee ne eae eee 13

SRASGOSPOLeSzat LEASt AN Sep tate’ Seo see Steeirerd aca Shepton agit ree A aoa scl eRe or AR Biss 9s 4

3. ASCOSPOres OU ESeptate®.... oe eck che sah ile eee fend a eee ee Dee eae T. toniniana

A) Aseospotes.maily. s-septater... Strachan s.r tet cc ohne get kes ARS Aen een ct S)

Av Ascosporesiexclusively.lusepiate s+ %. 18 As dchaa sort. Set Ake, BB eee ne: Book 6

5. Ascospores 1-3-septate, 13.5-22.5 3.5-5 UM ..... 2. ce eee ee cece ee T. superioris

5. Ascospores mainly 3-septate, 23.5-33 x 3-4 Um... 6... eee eee eee T. alutacea

6. Thallts pruingse; pruinatarinose® 42.2 oni we ee ne eee sca pet we ee ees Yi

6. Thallus:prutnhose, pruina. granulate) 0-4 iu oh este sict wide SIE Pack andi 10

576 ... Miao, Sun & al.

7. Thallus rosulate, pruina exposed surface of thallus .................... T. candida

7: Thallusnot-rosulate, priuina-patehy.en thallus: ../ 5.2 eee5 an se ee ee 8

SeThalluswvithpsetidocyphellae %) 1.34.0 2048 oer sek bat teed T. physaroides

Sathallusswithout pseudocyphellae ¢ .. GoW akg ete poe he Gear a eS 9

9. Squamules bullate and partly vertically flattened, more or less imbricate,

containing an unknown substance (TLC solvent system C Rf.21) ... T. opuntioides

9. Squamules weakly convex to bullate but not vertically flattened or imbricate; not

containing an unknown substance (TLC solvent system C Rf. 21) ... T. sedifolia

10. Squamules margin usually densely with white pruina ............... T. albilabra

10. Pruina not only on the edge of squamules ............... 00. e eee e eee eee 11

11. Ascosporeés: stall (835-13 4-5: (ith) att we ole PE ee ina eae es T. nordlandica

LL-Ascospores larger(12—24-% 335 punt) ee ge Ae arte papa ah ped eka s Panam aiaite 12

12;-Thallus-rosulate; apothecia S4omime. aoe ky Been ee ae eee T. rosulata

12Atnallasnoetrosulate;apothecia S157 tien. sho: opt Max doar eee T. diffracta

13, Epithecttimred-brown, K+ red) ni. c. gered be Sedo Vee Pes ee ees T. lutosa

13. Epithecium green, K-, N+ purple; ascospores 3-septate ................ T. gobica

14, Thallus epruinose-or slighthypruinose <3 * 2; 2s cajos 53556652 ba Rees oS: 15

WAS. Phalitis<e premise” af Seen. eh Mea ents fein tiis eared Meta Ye tae tae Bae Rwy 17

15. Thallus pale brown; epithecium dark brown ..................... T: sculpturata

15. Thallus of different colour; epithecium not dark brown...................06- 16

16. Thallus crustose; epithecium gray, K+ purple, N+ purple ............. T. pennina

16. Thallus squamulose, epithecium olive green to bright green, K-,

NEF ipurplesascosperes las-sepiate Mg... 2. s Cok ah maker Ges T aromatica

17. Epithecium olive green to bright green, K-, N+ purple ....................04. 18

17. Epithecium red-brown, K+ red; ascospores (1-)7(-9) -septate ........ T. ruginosa

18. ‘thallus :crustose:ascospores-luseptate: +... nee Be ee ee T. philippea

18. Thallus squamulose, or crustose to subsquamulose ............... 22.0020 eee 19

19. Thallus crustose to subsquamulose; epithecium olive green;

ASCOSPOLES 3S + SCWUALE ac iso's, ote Cecio E es Shope Mites) oot ew Aten Lane T. coelestina

19. Thallus squamulose; epithecium olive green to bright green .................. 20

20. Ascospores simple to 1-septate; epithecium bright green to dark brown .. T. tristis

20. Ascospores 1 to more septate; epithecium olivaceous to bright green .......... 21

21. Epithecium olivaceous to bright green (K-),

ascospores 1(—3)=septate s..sess02. Gs vas tees ee seee tom euesnes T’ cinereovirens

21. Epithecium olivaceous (K+ purple to brown), ascospores 1-(-3)-septate ... T. poeltii

Key to the subspecies of Toninia tristis

Ik -ASCUSIOLAM OC GE PEGE tote Metta totale Stas Phas Ue BRR OR MRE 51 .C cea ale Geet ah IRE se aE CP 2

LPASCUSHIO LOPAN CGPI oh aa. ov hge bers tee OS Sane Makes: Sng shne tees hy SMe Pa hah Sr cots 3

Toninia taxa new for China... 577

Zi ASCOSPOLeS L-SEPtate,. shea srs oihtetier as map Ue eleye a! ath wen odes subsp. asiae-centralis

D SASEOSPOLES SUN pLeT ae. Y ye rik. toree 4g. C28 Se he sees eee oe ates subsp. arizonica

J MEpit DE CWT AO FEEN Bc i at Te rguee eet Siete Ay eee See a ON dan ee Ue as 4

Deve PIC Ve CAUTION TA. ta heel = Leche iay ens coisas eater os bSK oheaeraaih sant Tae Males SEA» ene Ns nce 7

At ASCOSPOLES. Ue SEDIALE bo ah cttlaliee ek SAP ace te Sethe 8 PeRSO Nees, Lei bee We. /e SG subsp. canadensis

Al PASCOS POLES SLIP agen give eo trend rer ase he ape te, ates Pe SE AR MB Cha 5

5, sxscospores largerd OES 5: WIT) tena Ms es anos Nae es subsp. pseudotabacina

d. Ascosporesismall(Sa12-Minn): Wier ics kone. nee oe es ae eRe nee «Ae 6

G. No, chemical substance Ss. s.enzo se Seo ook oR a at. Seek oe subsp. thalloedaemiformis

6: Containine-chemical substance: e's ont ee hg ete co eons ie subsp. fujikawae

FacNSCOSPOTES STAPLE Maes 42h. harks tee agers ten Saitek a GME e eke. aah subsp. scholanderi

7, EXSCOS POLES! | -SEPTAte a eo ule satan, sleet ics, as telns a edz “niet Beles ase, hn 0 Dhow de onde Gaagaet 8

Se OGM ATMS Se NCO IN TaNaIN 81a: PS Rt a ec dT es aT MI otc tetne dd tay tla subsp. tristis

8. Squamules <3(—4) mim! ps esas Feec dw ead Gee eee awa ee eee ae subsp. coahuilae

Acknowledgments

We thank Dr Edit Farkas (MTA Centre for Ecological Research, Vacratot,

Hungary) and Dr Shou-Yu Guo (State Key Laboratory of Mycology, Institute of

Microbiology, Chinese Academy of Sciences, Beijing, China) for presubmission

review. The authors would also like to thank Lisong Wang and Xinyu Wang

(Kunming Institute of Botany, CAS, China) for assistance during this study. This

work was supported by the Emergency Management Project of National Natural

Science Foundation of China (31750001), the National Natural Science Foundation

of China (31900010).

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Scopoli JA. 1760. Flora carniolica. Wien, Sumptibus Joannis Thomae Trattner.

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Sun MJ, Yan SK, Tang R, Wang CX, Zhang LL. 2019. New records of Bilimbia and Toninia from

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578 ... Miao, Sun & al.

Timdal E. 1991. A monograph of the genus Toninia (Lecideaceae, Ascomycetes). Opera Botanica

110. 137 p.

Timdal E. 2002. Toninia. 488-501, in: TH Nash & al. (eds). Lichen Flora of the Greater Sonoran

Desert Region. Vol. 1. Tempe, Lichens Unlimited, Arizona State University.

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K6niglichen Naturhistorischen Hofmuseums, Wien 5: 20-48.

MYCOTAXON

July-September 2020—Volume 135, pp. 579-587

https://doi.org/10.5248/135.579

Serendipita sacchari sp. nov.

from a sugarcane rhizosphere in southern China

LING XIE?*, YAN- YAN LONG?”*, YAN ZHANG’”,

YAN-Lu CHEN’”, WEN-LONG ZHANG?*

'Plant Protection Research Institute, Guangxi Academy of Agricultural Science,

Nanning 530007, China

? Microbiology Research Institute, Guangxi Academy of Agricultural Science,

Nanning 530007, China

*CORRESPONDENCE TO: chris1933@126.com

ABSTRACT—We isolated a new species, proposed here as Serendipita sacchari, from a

sugarcane rhizosphere in Guangxi Province, China. This species is characterized by its

unstable nucleus numbers (1-15) in its chlamydospores versus their regular distribution in

hyphal cells. ITS rDNA and combined LSU+ TEFl-a sequence analyses also support the

uniqueness of this new plant symbiont.

Key worps—molecular phylogeny, Sebacinales, Serendipitaceae, taxonomy

Introduction

Serendipita P. Roberts (Basidiomycota, Sebacinales, Serendipitaceae), typified

with S. vermifera (Oberw.) P. Roberts, originally comprised seven species

(Roberts 1993). Two additional new species S. lyrica Trichiés (Trichiés 2003),

and S. herbamans K. Riess & al. (Riess & al. 2014) have been proposed in this

genus, and two anamorphic species in Piriformospora Sav. Verma & al. have

been recombined as S. indica (Sav. Verma & al.) M. Weiss & al., and S. williamsii

(Zuccaro & M. Weiss) M. Weiss & al. (Verma & al. 1998; Basiewicz & al. 2012;

WeifS & al. 2016). Serendipita currently contains 11 species and DNA barcodes

are widely accepted as an important tool in delineating species (Schoch & al.

2012; Riess & al. 2014).

* LING XIE & YAN-YAN LonG contributed equally to this work.

580 ... Xie, Long & al.

During an investigation of plant endophytes in sugarcane rhizosphere in

Guangxi Province, China, an interesting strain has been morphologically and

phylogenetically confirmed as a new species of Serendipita and is described

herein.

Materials & methods

Soil samples were collected from a 5-15-cm sugarcane rhizosphere layer

in Tanluo Town, Nanning City, Guangxi Province, China, in Feb 2011. Fungal

isolates were obtained by baiting with Chinese cabbage as described by Narisawa

& al. (1998), cultured on PDA media, and deposited in the China General

Microbiological Culture Collection Center (CGMCC3.19906). A pure culture of

the fungus was air-dried and frozen to prepare the holotype specimen, which is

curated in the Mycological Herbarium, Institute of Microbiology, Chinese Academy

of Sciences, Beijing, China (HMAS). Observations and measurements were made

with an Olympus BX53 Ci-L light microscope and Tescan-vega3 LMU scanning

electron microscope (SEM).

DNA extraction, amplification, sequencing

Genomic DNA was extracted using the Sangon Biotech Rapid Fungi Genomic

DNA Isolation Kit from mycelia grown at 28 °C for 10 d on oatmeal medium

(OA, 10 g oatmeal, 10 g bacto agar, 1 g MgSO,e7H,O, 1.5 g H,PO,, 1 g NaNO,,

for 1 L distilled water). The internal transcribed spacer of ribosomal DNA (ITS

rDNA), large subunit of ribosomal DNA (LSU), and translation elongation factor

1-a (TEF1-a) were amplified with fungal specific primer pairs ITS1/ITS4, LROR/

LRS, and EF1-983f/EF1-2218r1, respectively (Vilgalys & Hester 1990, Basiewicz & al.

2012). PCR reaction mixture and conditions followed the protocol of 2xEasyTaq

PCR SuperMix. The DNA was amplified in 50 uL volumes containing PCR buffer

[20 mM KCl, 10 mM (NH,),SO,, 2 mM MgCl , 20 mM Tris-HCl, pH8.4], 200 uM

of each dNTP, 15 pmol of each primer, 100 ng DNA template, and 2.5 units of Taq

DNA polymerase (Biocolor BioScience & Technology). Thermal cycling began with

5 min of denaturation at 94 °C, followed by a 35 cycles of denaturation (94 °C for

40 s) + annealing (56 °C for 40 s) + extension (72°C for 60 s), and finalized with

a final extension at 72 °C for 10 min. A negative control using sterilized distilled

water instead of template DNA was included in the amplification process. The

PCR products were examined by electrophoresis at 75 V for 2 h in 0.8 % (W/V)

agarose gel in 1xTAE buffer (0.4 M Tris, 50 mM NaOAc, 10 mM EDTA, pH 7.8)

and visualized under an ultraviolet light after staining with ethidium bromide (0.5

ug mL). The PCR products were purified using MultiScreen® PCRw96 Cleanup

Filter Plates according to the manufacturer’s protocol. Purified PCR products were

directly sequenced with primer pairs mentioned above in an ABI 3730-XL DNA

sequencer. The sequences were deposited at GenBank (http://www.ncbi.nlm.nih.

gov) and compared through a BLAST search.

Serendipita sacchari sp. nov. (China) ... 581

TABLE 1. Taxa with GenBank ITS accession numbers

SPECIES VOUCHER GENBANK NO.

Cantharellus avellaneus 1217/ER KX857081

Chaetospermum camelliae MFLUCC12-0433 KF516965

C. chaetosporum CBS:154.59[T] KJ710461

Craterocolla cerasi TUB 020203 KF061265

Ditangium altaicum LE 231836 [T] NR 163760

Efibulobasidium albescens — AF384860

Globulisebacina rolleyi — AY509550

Helvellosebacina concrescens TUB 019706 JQ665516

H. helvelloides TUB 019983 KF000415

Paulisebacina allantoidea — AF490396

RoKi 179 KF061266

Sebacina candida TUB 020331 KF061278

S. pallida TUB 019650 JQ665562

Serendipita herbamans S1[T] KF061285

S. indica DSM 11827[T] KF061284

S. sacchari CGMCC3.19906 [T] KY496808

S. williamsii DAR 29830 KY509323

Tremellodendron ocreatum TH8577 KT339265

Tremelloscypha dichroa VB4212 KF061281

TABLE 2. Taxa with GenBank LSU and TEF1-a accession numbers

GENBANK NO.

SPECIES VOUCHER LSU TEF1-a

Serendipita. indica DSMZ 11827[T] AY505557 AJ249911

S. sacchari CGMCC3.19906 [T] KY496809 MF196313

S. vermifera MAFF 305835 DQ983814 JN211111

MAFF 305838 DQ983816 JN211116

MAFF 305828 DQ520096 JN211115

S. williamsii DAR 29830 AY505556 JN211110

Phylogenetic analyses

The three DNA loci were separately aligned using Clustal-X (1.83). Two

neighbour-joining trees were derived from ITS1-5.8S-ITS2 and the combined

LSU+ TEF1-a datasets using MEGA v. 4 (Tamura & al. 2007)(TABLES 1,2; Fics 2,3).

Bayesian analyses of the aligned DNA sequence dataset were conducted with

MrBayes v. 3.1.2 (Huelsenbeck & Ronquist 2001) following Sun & Guo (2010). The

best-fit evolutionary model was determined for each dataset by comparing different

evolutionary models via MrModeltest v. 2.3 (Nylander 2008). Four simultaneous

Markov Chain Monte Carlo chains were run by starting from random trees and

sampling every 100 generations. The analyses were halted at 4,000,000 generations

after the calculation reached stationarity. Of the 40,000 trees generated, 25% were

582 ... Xie, Long & al.

excluded as “burn in” when calculating the posterior probabilities. Bayesian posterior

probabilities were obtained from the remaining 50% majority rule consensus trees.

Significant support was defined by clades contained in >95% of the sampled trees.

Taxonomy

Serendipita sacchari L. Xie, Y.Y. Long & Y.L. Chen, sp. nov. Fig. 1

MB 836761

Differs from other Serendipita species by its variable nucleus numbers (1-15 nuclei) in

the chlamydospore cells.

Type: China. Guangxi Province: Nanning City, Tanluo Town, in sugarcane rhizosphere,

22°55’30"N 107°58’34’E, alt. 96 m, Apr. 2011 (holotype, HMAS 247074; ex-type culture

CGMCC3.19906; GenBank KY496808, KY496809, MF196313).

EtryMo.oey: sacchari, referring to the plant from whose rhizosphere the species was

first collected.

Colonies reaching 50 mm in diam on PDA medium at 28 °C after 2 weeks, light

yellow, circular (or nearly circular), less aerial hyphae. Hyphae hyaline to light

yellow-green, septate, 1.2-3.4 um (2.2 + 0.5 um, n = 50), each cell regularly

containing 1-2 nuclei. Chlamydospores abundant, spherical to pear-shaped,

singly on top of hypha, aseptate. Spores 11.1-17.6 um (13.5 + 1.3 um, n = 55)

in diameter, variably containing 1-15 nuclei. Neither conidiophores nor sexual

structures observed.

Sequences & the phylogenetic analyses

The obtained ITS (610 bp), LSU (945 bp), and TEF1-a (1131 bp) sequences

were deposited in GenBank. To determine the phylogenetic position of

Serendipita sacchari, all available ITS, LSU, and TEFl-a sequences from

Serendipita and related genera were downloaded from GenBank (TABLES

12s

The ITS sequences from one isolate of Serendipita sacchari, five

Serendipitaceae, nine Sebacinaceae, three Sebacinales incertae sedis, and

one outgroup Cantharellus avellaneus, were included in the phylogenetic

analysis. In the alignment of these 19 sequences, the data matrix comprised

591 characters. The alignment dataset was analyzed using MrBayes, applying

the GTR+G model selected by MrModeltest as the best-fit model. The

prior probability density is a flat Dirichlet (all values = 278 1.0) with both

Revmatpr and Statefreqpr as default settings. Posterior probability (BPP)

and bootstrap (BS) values are shown on the branches of the ITS Bayesian

tree (Fic. 2). The ITS phylogeny places S. sacchari in Serendipitaceae and in

a clade with S. indica (BPP = 0.83; NJ support = 97%).

Serendipita sacchari sp. nov. (China) ... 583

Fic. 1. Serendipita sacchari (ex-type, CGMCC3.19906). A: Colony; B, C: Bright field microscopic

image of chlamydospores; D-F: Scanning electron microscopic image of chlamydospores;

G, H: DAPI-stained chlamydospores; I: DAPI-stained hyphae. Scale bars: B-I = 10 um.

A combined LSU+TEF1-a dataset comprising one Serendipita sacchari

isolate, five Serendipita isolates, and the outgroup Sebacina incrustans was

included in a second phylogenetic analysis. These seven sequence pairs

(LSU+TEF1-a) aligned in a 1990-character data matrix. The alignment

dataset was analyzed in MrBayes, applying the GTR+G model selected

by MrModeltest as the best-fit model. The prior probability density is a

flat Dirichlet (all values = 278 1.0) with both Revmatpr and Statefreqpr as

default settings. Here also BPP and BS values are shown on the branches of

the combined Bayesian tree (Fic. 3).

584 ... Xie, Long & al.

Sebacina pallida JQ665562

0.86/88

0.97/81 | Tremelloscypha dichroa KF061281

Sebacina candida KF061278

0.89/* . :

1.00/100 Helvellosebacina concrescens JQ665516

0.90/*

0.85/*

Helvellosebacina helvelloides KF000415 :

Sebacinaceae

Tremellodendron ocreatum KT339265

)-S1/* 4 men

ee Globulisebacina rolleyi AY 509550

0.94/100 Craterocolla cerasi KF061265

Ditangium altaicum NR 163760

Paulisebacina allantoidea AF490396

1.00/100

0.97/99 Paulisebacina allantoidea KF061266

Serendipita herbamans KF061285

aT / . Serendipitaceae

Serendipita sacchari KY 496808

0.83/97

Serendipita indica KF061284

1.00/100

Serendipita williamsii KY 509323

0.56/* Chaetospermum chaetosporum KJ710461

0.80/88 Efibulobasidium albescens AF384860

Sebacinales incertae sedis

Chaetospermum camelliae KF516965

Cantharellus avellaneus KX857081

Fic. 2. Phylogenetic tree based on ITS sequences of Serendipita and related Sebacinales species.

Cantharellus avellaneus was included as outgroup. The numbers at each branch represented

Bayesian posterior probabilities (left) and bootstrap support calculated from 1000 replicates (right).

The sequence derived from the proposed new species is in bold. Bar = 0.1 expected changes per site.

In the LSU+TEF1-a phylotree, S. sacchari formed a clade with S. williamsii

with 0.95 BPP support and 100% NJ support. A comparison of the ITS,

LSU and, TEFl-a sequence dataset indicates that S. sacchari differs from

S. williamsii in 12/530 bp (2.3%, ITS), 15/930 bp (1.6%, LSU), and 60/1068

bp (5.6%, TEF1-a); and from S. indica in 22/552 bp (4.0%, ITS), 21/938 bp

(2.2%, LSU), and 63/1132 bp (5.6%, TEF1-a).

COMMENTS. Four species in Serendipita—S. herbamans, S. indica, S. williamsii,

and S. vermifera—were closely related to and allied with S. sacchari.

Neither conidiophores nor sexual structures were observed in these four

taxa. Serendipita sacchari can be easily distinguished from S. herbamans

Serendipita sacchari sp. nov. (China) ... 585

Serendipita vermifera DQ983814+JN2 11111

Serendipita vermifera DQ983816+JN211116

Serendipita vermifera DQ520096+JN2 11115

Serendipita sacchari KY 496809+MF196313

Serendipita williamsti AY 505556+JN2 11110

1.00/100

Serendipita indica AY 505557+AJ2499 11

Sebacina incrustans FJ6445 13+KF3 13907

Q.']

Fic. 3. Phylogenetic tree based on the combined LSU+TEF 1-a sequence data of Serendipita species.

Sebacina incrustans was designated as outgroup. The numbers at each branch point represented

Bayesian posterior probabilities (left) and bootstrap support calculated from 1000 replicates (right).

The sequence derived from the proposed new species is in bold. Bar = 0.1 expected changes per site.

and S. vermifera by its bigger (11.1-17.6 um diam.) chlamydospores (3-5

um x 4-5 um in S. herbamans; approximately 8 um diam. in S. vermifera;

Riess & al. 2014, Ray & Craven 2016). The main difference among

S. sacchari, S. indica, and S. williamsii is the distribution and number of nuclei

in the cells (Basiewicz & al. 2012). Nuclear number in the chlamydospores

of Serendipita sacchari (1-15 nuclei) distinguishes the new species from

S. indica (8-25 nuclei) and S. williamsii (<10 nuclei) (Basiewicz & al. 2012).

Moreover, hyphal cells of S. sacchari have fewer nuclei and exhibit a more

regular distribution compared with S. williamsii (2-6 nuclei irregularly

distributed) (Verma & al. 1998; Basiewicz & al. 2012). Morphological traits

and phylogenetic data both support the recognition of Serendipita sacchari

as an independent species.

Serendipita herbamans is a common species in plant roots and widely

distributed across agricultural and grassland ecosystems. It has the potential

to promote the growth of herbaceous plants (Riess & al. 2014). Serendipita

586 ... Xie, Long & al.

indica (= Piriformospora indica) is widely distributed and well known as

a symptomless root endophyte that colonizes bryophytes, pteridophytes,

gymnosperms and angiosperms. It has been reported to occur in four

continents, is extremely versatile in its mycorrhizal associations, and

is known for its ability to promote plant growth (Varma & al. 2012).

Serendipita williamsii (= Piriformospora williamsii) was isolated from the

spore of arbuscular mycorrhizal fungal in Australia (Basiewicz & al. 2012).

Serendipita vermifera was isolated from the Australian orchid Cyrtostylis

reniformis (Warcup 1988). It is reported to induce beneficial effects on plant

performance including growth promotion, increase nutrient uptake, enhance

seed production, and increase resistance against different biotic and abiotic

stresses (Ray & Craven 2016). Serendipita sacchari was from sugarcane

rhizosphere soil, and symbiotic with Chinese cabbage and sugarcane as a

symptomless root endophyte (data not shown). All five Serendipita species

are plant symbiotic microbionts.

Acknowledgements

This work was supported by the National Natural Science Foundation of China

(No. 31460016), Guangxi Natural Science Foundation (No. 2015GXNSFBA139083),

and Basic Scientific Research Special Project of Guangxi Academy of Agricultural

Sciences (No. 2015YT80). We appreciate the thoughtful reviews of Dr. Xiaoyong Liu

(Institute of Microbiology, Chinese Academy of Sciences) and Dr. Jiwen Xia (College

of Plant Protection, Shandong Agricultural University).

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phenotypic characterization of Sebacina vermifera strains associated with orchids,

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Serendipita sacchari sp. nov. (China) ... 587

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analysis (MEGA) software version 4.0. Molecular Biology and Evolution 24: 1596-1599.

https://doi.org/10.1093/molbev/msm092

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autres hétérobasidiés notables de Lorraine (France). Bulletin trimestriel de la Société 118

(4): 351-379.

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P. 1998. Serendipita indica, gen. et sp. nov., a new root-colonizing fungus. Mycologia 90:

896-903. https://doi.org/10.2307/3761331

Varma A, Bakshi M, Luo B, Hartmann A, Oelmueller R. 2012. Serendipita indica: a novel

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https://doi.org/10.1111/nph.13977

MYCOTAXON

July-September 2020—Volume 135, pp. 589-612

https://doi.org/10.5248/135.589

Five Nolanea spp. nov. from Brazil

FERNANDA KARSTEDT*, SARAH E. BERGEMANN?, MARINA CAPELARI’

* Nucleo de Pesquisa em Micologia, Instituto de Botanica,

Caixa Postal 68041, 04045-902 Sao Paulo, SP, Brazil

* Biology Department, Middle Tennessee State University,

PO Box 60, Murfreesboro, TN 37132, USA

" CORRESPONDENCE TO: fernanda.karstedt@gmail.com

ABSTRACT—Five new species of Nolanea (Entolomataceae, Agaricales)—N. albertinae,

N. atropapillata, N. pallidosalmonea, N. parvispora, and N. tricholomatoidea—were

collected from the Sao Paulo metropolitan region. Phylogenetic analyses based on partial

sequences of the mitochondrial small subunit (mtSSU), nuclear large subunit (nLSU)

and second largest RNA polymerase subunit (rpb2), confirm that all these species belong

to Nolanea. Morphological descriptions, comments, illustrations, scanning electron

microscopy and color photos are included.

Key worps—basidiomycetes, Entoloma, Neotropical fungi, South America, taxonomy

Introduction

Currently, Nolanea has two main circumscriptions, one according to

Largent (1994) and Henkel & al. (2014) as Nolanea (Fr.) P. Kumm., and the

other according to Noordeloos (1992, 2004) and Noordeloos & Gates (2012)

as Entoloma subg. Nolanea (Fr.) Noordel. Both circumscriptions agree that

Nolanea comprises species often mycenoid with the pileus surface typically

hygrophanous, usually translucent-striate and with angled basidiospores

with five, six or seven angles in profile view, which can be subisodiametric or

heterodiametric. The main difference between the accepted circumscriptions

of the genus Nolanea is in the pileipellis structure. Largent (1994) considers

Nolanea to encompass only species that have a pileipellis composed of two

layers, with a suprapellis composed of a cutis or a repent entangled hyphae

590 ... Karstedt, Bergemann, Capelari

layer overlying a subpellis of inflated hyphae, whereas Noordeloos accepts

species in Entoloma subg. Nolanea in which the repent, cylindrical hyphae of

the pileipellis often (but not necessarily always) overlie a distinct differentiated

subpellis of inflated cells.

In general, the few phylogenetic studies that include Nolanea species

do not define which circumscription is the most appropriate for the group;

additionally, they suggest Nolanea can be polyphyletic (Co-David & al. 2009,

Baroni & Matheny 2011). However, Nolanea species are always in the Nolanea-

Claudopus clade, independent of the combination of loci used: mtSSU, nLSU,

and rpb2 (Co-David & al. 2009, Baroni & Matheny 2011) or ITS, nLSU, and

rpb2 (Kinoshita & al. 2012). Also, it is usually possible to visualize two separate

branches within the Nolanea—Claudopus clade, with a clade of Nolanea species

separate from a clade comprising Claudopus species.

More recent molecular studies of Entoloma subg. Nolanea (Vila & al. 2013,

Raj & al. 2014, Raj & Manimohan 2016) examined Nolanea species using

only ITS sequences, with special emphasis on taxa delimitation and new taxa

proposition. The phylogeny produced in these studies supports, even with only

ITS sequences, the recognition of Nolanea as a good phylogenetic group and

would seem to support Nolanea as an autonomous genus in the Entolomataceae.

Nolanea (with about 250 names in MycoBank and Index Fungorum) is

considered a common and speciose genus, especially in temperate regions

(Noordeloos 1992, 2004, Largent 1994, Horak 2008), but it is also present in

tropical and subtropical regions (Romagnesi & Giles 1979). In Brazil of the 23

Nolanea species recorded (Rick 1919, 1920, 1930, 1961, Bresadola 1920, Singer

1953, Horak 1977, Capelari 1989, Putzke & Cavalcanti 1997, Pegler 1997, de

Meijer 2001, 2006, Rosa 2002), eleven have been cited and/or described by

Rick (1919, 1920, 1930, 1961, Singer 1953) and are considered as incertae sedis

(Horak 1977). Many other species have been reported in dissertations (Capelari

1989, Rosa 2002), an identification key (Pegler 1997), and lists (Singer 1953, de

Meijer 2001, 2006), but only three have been published with a full description

(Putzke & Cavalcanti 1997).

In this work, we describe five new species found in the Sao Paulo metropolitan

region accompanied by molecular information and phylogenetic analyses.

Materials & methods

Basidiomata collection and morphological observations

The studied materials were collected mainly at the Parque Estadual da Cantareira

and the Parque Estadual das Fontes do Ipiranga in Sao Paulo City and Reserva

TABLE I. Collections used in the phylogenetic analyses

Cl. = Claudopus; E. = Entoloma; I. = Inocephalus; L. = Leptonia; P. = Pouzarella;

Nolanea spp. nov. (Brazil) ... 591

Rh. = Rhodocybe; R. = Richoniella; T. = Trichopilus; Sequences generated in this study in bold.

NCBI spECIES NAME

Cl. minutoincanus

Cl. viscosus

E. abortivum

E. albidoquadratum

E. bloxamii

E. cephalotrichum

E. conferendum

E. cf. conferendum

E. gasteromycetoides

E. haastii

E. hebes

E. nidorosum

E. pallideradicatum

E. parasiticum

E. porphyrescens

E. procerum

E. pygmaeopapillatum

Entoloma readiae

E. sericatum

E. sericionitidum

E. serrulatum

E. sinuatum

E. strictius

E. strictius var.

isabellinus

E. turbidum

E. undatum

RPB2

HQ731517

HQ731518

GU384642

GQ289222

GQ289223

GQ289226

GQ289229

GQ289231

GQ289235

GQ289238

GQ289241

GU384643

GQ289247

GQ289248

GQ289253

GQ289254

GQ289256

GQ289257

GQ289260

EF421016

GQ289263

GQ289264

GU384641

GU384656

GQ289270

LSU

HQ731514

HQ731516

GU384616

GQ289150

GQ289151

GQ289154

GQ289157

GQ289160

JQ320115

GQ289164

GQ289167

GQ289170

GU384617

GQ289176

GQ289177

GQ289182

GQ289183

GQ289185

GQ289186

GQ289189

AF261315

GQ289192

GQ289193

AF042620

GU384618

GU384630

GQ289202

GENBANK ACC. NO.

mtSSU

HQ731511

HQ731513

GU384595

GQ289290

GQ289291

GQ289294

GQ289297

GQ289300

GQ289304

GQ289307

GQ289310

GU384596

GQ289316

GQ289317

GQ289322

GQ289323

GQ289325

GQ289326

GQ289329

EF421098

GQ289332

GQ289333

GU384594

GU384603

GQ289342

VOUCHER

DLL 9871

DLL 9788

TB6693

den Bakker 92

Manimohan 667-T

Noordeloos

200442

Ulje 1997-08-01

Noordeloos

200313

HKAS 48953

Gates E2031

Noordeloos

2004055

Hartman 1992-

10-28

TB9971

Hausknecht

Noordeloos

200330

Noordeloos

2004113

Noordeloos

2004070

Noordeloos

200364

Noordeloos

2004050

Noordeloos

200328

TB7144

Noordeloos

2004062

Wisman 2003-

09-19

M96/10

1TB7710

TB6949

Noordeloos 200327

ORIGIN

Australia,

NSW

Australia,

QLD

USA, NY

Canada

India, KL

Austria

Netherlands

Belgium

China, SC

Australia,

TAS

Australia,

TAS

Netherlands

USA, NY

Austria

Belgium

Australia,

TAS

Australia,

TAS

Slovakia

Australia,

TAS

Slovakia

USA, NY

Australia,

TAS

Netherlands

USA, TN

USA, NY

Belgium

592 ... Karstedt, Bergemann, Capelari

GENBANK ACC. NO.

NCBI sPECIES NAME RPB2 LSU mtSSU VOUCHER ORIGIN

E. valdeumbonatum GQ289271 GQ289203 GQ289343 Meusers E4565 [T] Germany

I. murrayi GU384637 GU384620 GU384590 VHAs02.02 —

I. sp. MCA2479 GU384640 GU384622 GU384593 MCA2479 —

L. serrulata GU384634 GU384624 GU384588 VHAs01.02 —

L. sp. MCA1486 GU384635 GU384623 GU384589 MCA1486 —

N. albertinae — KF738937 KF738925 FK1731 Brazil, RJ

KF771345 KF738938 KF738926 FK1732 Brazil, RJ

KF771344 KF738936 KF738924 FK0935-T Brazil, SP

N. atropapillata — KF738940 KF738929 FK0898-T Brazil, SP

N. cetrata KF771346 KEF738942 KF738927 DLL9531 USA, CA

— AF261319 — TB7382 USA, NY

N. conferenda KF771351 KF738946 KEF738935 ~—11CA014 USA, CA

N. conferenda — AF261321 — TB7660 USA, NY

N. hirtipes — AF261320 — K1171992

N. pallidosalmonea — KF738941 KF738930 _—_‘-FK0891 Brazil, SP

N. parvispora KF771348 KF738943 KF738931 FK1140 Brazil, SC

— — KF738932 FK2135 Brazil, SC

N. sericea DQ367435 DQ367423 EF421099 VHAs 03/02 —

GQ289262 GQ289191 GQ289331 Noordeloos Slovakia

200329

KF771349 KF738944 KF738933 11CA055 USA, CA

KF771350 KF738945 KF738934 11CA056 USA, CA

— AF261318 —_— TB6506 USA, CA

N. strictior EF421017 — EF421100 DUKE-JM96/10

N. tricholomatoidea KF771347 KF738939 KF738928 FK1049-T Brazil, SP

P. albostrigosa HQ876513 HQ876535 HQ876557 __Largent 9641 Australia

P lasia HQ876507. HQ876529 HQ876555 _Largent 9662 Australia

P. nodospora _ AF261308 — TB5716 USA, SC

P. setiformis HQ876503 HQ876525 HQ876525 _ Largent 9809 Australia

Rhodocybe trachyspora GU384658 GU384629 GU384605 TB5856 USA, CA

Richoniella asterospora JF706311 JF706310 — PBM3268 USA, TN

iG dar ee Le AF261290 — TB6957 USA, NC

OUTGROUP

Clitocybe dealbata DQ825407 = AF223175 AF357138 IE-BSG-HC95cp3 —

Lyophyllum IE-BSG-

DQ367434 AF223202 AF357101 =

leucophaeatum HAe251.97

Nolanea spp. nov. (Brazil) ... 593

Bioldgica de Paranapiacaba in Santo André City, in remnants of the Atlantic Forest

in Sao Paulo State, Southeast Brazil.

Revived sections of tissues from dried basidiomata mounted in 5% KOH, 5%

NH,OH, or 1% aqueous Congo Red dye used to stain hyaline structures were

examined microscopically. All the micrographs were drawn with the aid ofa drawing

tube. ‘Q’ represents the minimum and maximum length/width quotient, and ‘Qn

represents the mean length/width quotient of the total sample of measured spores.

The pileipellis structure was examined from the middle portion of the pileus and not

the disc or margin. The specimens were deposited in the herbarium of the Instituto

de Botanica (SP) and of the Royal Botanic Gardens (K). The samples were prepared

for SEM following Baroni (1981) and scanning electron micrographs (SEMs) were

made using a LEO 435 VP scanning electron microscope.

DNA extraction, amplification, sequence analyses

The DNA was extracted with hexadecyltrimethylammonium bromide (CTAB)

(Ferreira & Grattapaglia 1995) using lyophilized tissues from basidiomata ground

to a fine power in liquid nitrogen. The sample was resuspended in 50 uL of TE and

stored at —20 °C. The partial sequences of the mitochondrial small subunit (mtSSU)

were amplified via the polymerase chain reaction (PCR) primed by MS1 and MS2

(White & al. 1990); the 28S nuclear large subunit (nLSU) rDNA was amplified

using LROR and LR5 (Moncalvo & al. 2000); the ITS1-5.8S-ITS2 was amplified

using ITS1-F and ITS4 (White & al. 1990, Gardes & Bruns 1993); and the second

largest subunit of the RNA Polymerase II (rpb2) using rpb2-i6F and rpb2-i7R (Co-

David & al. 2009) or rpb2-F1 and rpb2-R1b (Largent & al. 2013). The PCRs were

performed in 25-50 uL reaction volumes, where the reaction concentration was 1x

for PCR Buffer, 2 mM for MgCl2, 0.2 mM for each dNTP, 0.5 uM for each forward

and reverse primer, and 0.025 U of Taq polymerase. Products from multiple

reactions were pooled for weak PCR amplifications. The reaction was performed

using the following cycling parameters: 94 °C for 2 min, 34 cycles at 94 °C for 45 s,

50 °C (54C? for rpb2) for 1 min and 10 s and 72 °C for 2 min, and then 72 °C for 10

min. PCR amplifications products were purified using the AxyPrep PCR Clean-up

Kit (Axygen Biosciences, Union City, USA) or alternatively, using the ExoSapIT kit

following the protocols outlined in Largent & al. (2011). DNA sequencing reactions

were performed with BigDye Terminator v3.1 Cycle Sequencing Kit. The sequences

were run in a ThermoFisher 3730 DNA Analyzer using Sequencing Analysis 5.3.1

and Base Caller KB. The forward and reverse sequences for each sample were

aligned and edited using CodonCode Aligner or Sequencher.

The phylogenetic analysis was conducted using sequences obtained in this study

and sequences found in GenBank of species traditionally accepted as Nolanea

[Nolanea—Claudopus clade sensu Co-David & al. (2009) and Baroni & Matheny

(2011)] (TaBLeE I). Lyophyllum leucophaeatum (P. Karst.) P. Karst. and Clitocybe

dealbata (Sowerby) P. Kummer were selected as outgroups based on phylogenetic

analyses performed by Co-David & al. (2009). The sequences were manually

aligned using Se-Al Sequence Alignment Editor; spliceosomal introns in the rpb2

594 ... Karstedt, Bergemann, Capelari

were delimited using Augustus ver. 2.4 webserver (Stanke & al. 2008) and omitted

along with ambiguous sites in mtSSU. For these phylogenetic analyses, sequences

from the three genes including missing data were assembled. The final alignment

after the exclusion of introns and ambiguous sites was 3162 bp including gaps:

592 bp for the mtSSU, 1390 bp for the nLSU and 1180 bp for the rpb2 (Tree-BASE

database ID14850).

Phylogenetic analyses were performed using maximum likelihood (ML),

Bayesian Inference (BI), and maximum parsimony (MP). For the BI, the best-fit

model of nucleotide substitution was performed using the Akaike information

criterion (AIC) in TOPALI v.2 (Milne & al. 2009). ML analysis was performed using

RAXML 7.0.4 with a GTR + G model according to recommendations (Stamatakis

2006, Stamatakis & al. 2008). Support for nodes was tested using 1000 bootstrap

replicates. For the BI phylogeny, the best-fit model of nucleotide substitution for

each gene partition and codon for the rpb2 (GTR + G for LSU, mtSSU and codons

and 3 of rpb2, and GTR + G + I for first codon of rpb2). The BI was performed with

MrBayes 3.2.2 (Huelsenbeck & Ronquist 2001, Ronquist & Huelsenbeck 2003)

specifying four chains (one cold, three heated) for 2,000,000 generations (sampling

trees every 1000 generations), with a burn-in of 30,000 generations. MP heuristic

searches were performed using the program TNT (Goloboff 1999, Nixon 1999,

Goloboff & al. 2008). Tree searches were conducted with equal weights and treating

gaps as a fifth character state. Branch support was calculated with 1000 bootstrap

replicates. A bootstrap value of 70% was considered significant for ML and MP. The

Bayesian posterior probability (BPP) statistical support values of branches were

considered informative if = 0.95.

Phylogenetic results

The DNA sequences obtained, including partial sequences of the nLSU,

mtSSU,andrpb2maresummarizedin TABLE I. Additionally, sequences obtained

from Nolanea albertinae (FK0912: KF679349, FK0935 [holotype]: KF679348),

N. atropapillata (FK0898 [holotype]: KF679354), N. pallidosalmonea (FK0891

[holotype]: KF738923), N. parvispora (FK1140 [holotype]: KF679353) and N.

tricholomatoidea (FK1049 [holotype]: KF679352) were deposited in GenBank.

The phylogenetic analyses placed all five species studied into /Nolanea-

Claudopus (Co-David & al. 2009, Baroni & Matheny 2011, Kinoshita & al.

2012), an unsupported clade based on ML, MP, and BPP. The phylogenies

place all five species into one of two strongly supported internal clades (Fie. 1)

in the /Nolanea clade with other species of Nolanea.

Taxonomy

Five new species are described and illustrated with line drawings,

macroscopic photographs and SEMs of basidiospores.

Nolanea spp. nov. (Brazil) ... 595

Entoloma cf.conferendum HKAS48953

Nolanea cf. conferenda11CA014

Entoloma conferendum MEN200313

Nolanea conferendaTB7660

Nolanea albertinae FK1731

Nolanea albertinae FK1732

Nolanea albertinae FK0935

Nolanea tricholomatoideaFK1049

Entoloma cephalotricum CU19970801

Nolanea atropapillata FK0898

Nolanea pallidosalmoneaFK0891

Entoloma strictus var.isabelina TB7710

Nolanea cetrata DLL9531

Nolanea cetrata TB7382

Entoloma pallideradicatumWU189010

Nolanea parvispora FK1140

Nolanea parvispora FK2135

Entoloma pygmaeopapillatum MEN200364

Nolanea sericea VHAs03/02

Entoloma strictius M9610 lI

Nolanea sericea MEN200329

"/ Nolanea sericea 11CA055

Nolanea sericea11CA056

Entoloma valdeumbonatum MME4565

sya Nolanea sericea TB6506

Nolanea hebes CH19921028

Nolanea hirtipes K1171992

Nolanea strictior DUKEJM9610

Entoloma readiae MEN2004050

Entoloma arbortivum TB6693

Entoloma abortivum Bakker92

Entoloma sericeonitidumTB7144

Entoloma undatum MEN200327

Claudopus minutoincanus DLL9871

Entoloma parasiticum MEN200330

Claudopus viscosusDLL978 Claudopus

Inocephalus sp. MCA2479

Inocephalus murrayi VHAs02/02

Entoloma procerum MEN2004070

Entoloma albidoquadratum PM667

Entoloma porphyrescens MEN2004113

Trichopilus porphyrophaeusTB6957

Entoloma gasteromycetoides GGE2031

Entoloma serrulatum MEN2004062

Leptonia sp. MCA1486

Leptonia serrulata VHA01/02 Inocephalus-Cyanula

Pouzarella albostrigosa DLL9641

Pouzarella lasiaDLL9662

Pouzarella setiformis DLL9809

Pouzarella nodospora 1B5716 Pouzarella

Richoniella asterospora PBM3268

adie *t00°_ Entoloma nidorosum 1B9971

100 100 / 100) Entoloma sinuatum JW20030919 J

ae ae sericatum MEN200328 Entoloma s.s. / Rhodopoloid

Entoloma haastii MEN2004055

Entoloma turbidum TB6949

| Rhodocybe trachyspora TB5856

Entoloma bloxamii MEN200442

Clitocybe dealbata HC95cp3

Lyophyllum leucophaeatum HAe25197

Nolanea-Claudopus

93/81

100

100 497.

100

Fic. 1. Phylogram of maximum likelihood (ML) phylogenetic reconstruction using rpb2, nLSU,

and mtSSU sequences. Support is indicated with ML/MP bootstrap percentage above and

Bayesian posterior probabilities (BPP) below branches. The names of species studied in this

paper are in boldface. I. Species with heterodiametric basidiospores. II. Species with iso- to

subisodiametric basidiospores.

596 ... Karstedt, Bergemann, Capelari

Nolanea albertinae Karstedt & Capelari, sp. nov. FIGS. 2A-E, 3, 8A,B

MB826784

Differs from Entoloma belouvense by its smaller basidiospores and presence of refractive

hyphae.

Type—Brazil, Sao Paulo State, Santo André: Reserva Bioldgica de Paranapiacaba,

23°46’S 46°18’W, trail 15, 18.V.2006, F. Karstedt & M. Capelari FK0935 (Holotype: SP;

GenBank KF771344, KF738936, KF738924).

EtymMoLoGy—an honour to Albertina Teresinha Karstedt.

PitEus 14-18 mm diam., broadly conic, conico-convex, convex, plano-convex

or plane, often slightly papillate, rarely with the center plane or depressed, pale

brown, brown, dark brown (chocolate-colored), or gray-brown, center often

darker than the margin, surface glabrous or with a few appressed, concolorous

radial fibrils, when humid strongly translucent-striate, when dry, opaque,

hygrophanous, margin slightly or strongly eroded. PILEUS CONTEXT very

thin, whitish. LAMELLAE adnexed or sinuate, pale pinkish brown, subdistant,

with 1-5 lamellulae (3 series), margin slightly eroded. ST1PE 36-71 x 1-4 mm,

often cylindrical, rarely tapered above or subbulbous over base, brownish

beige, pale brown, gray-brown or ocher brown, rarely yellowish brown or

concolorous with pileus, apex always paler than the base, sometimes beige

or whitish, surface glabrous, often with the apex slightly pruinose, fibrillose-

twisted, hollow, base of stipe often with a white tomentum. SPORE PRINT not

recorded.

BASIDIOSPORES 7.5-10(-11.2) x 5-7.5 um (Q = 1.16-1.75, Qm = 1.36),

heterodiametric, strongly angled, with 5 or 6 angles in profile, slightly pinkish

brown, thin-walled. Basip1A 23-37 x 8.7—15 um, clavate, hyaline, thin-walled,

tetrasporic. LAMELLAE EDGES heterogeneous. CHEILOCYSTIDIA always

present, dispersed or present [in portion of] on the lamellae edges near the

stipe, 26-68 x 5-7.5 um, cylindrical, often apex capitate, hyaline, thin-walled.

PLEUROCYSTIDIA absent. LAMELLAR TRAMA subparallel, with hyphae 5-40 um

diam., cylindrical or fusiform, hyaline, thin-walled; subhymenium ramose.

PILEITRAMA composed of radially arranged and parallel hyphae, with hyphae

8.7-25 um diam., inflated or fusiform, hyaline, thin-walled. PILEIPELLIs

a cutis, made up of cylindrical hyphae, with brown intracellular pigment,

hyphae thin-walled, 3.7-7.5 um diam., with cylindrical or cylindrical-clavate

terminal elements; subpellis made up of inflated or fusiform, hyaline, thin-

walled hyphae, 10-37 um diam. STIPITIPELLIs a cutis of straw yellow, thin-

walled hyphae 5-15 um diam. CAULocysTIDIA absent. CLAMP CONNECTIONS

present, but inconspicuous. REFRACTIVE HYPHAE present at pileipellis.

Nolanea spp. nov. (Brazil) ... 597

Fic. 2. Basidiomata. Nolanea albertinae: a. FK735; b. FK883; c. FK912; d. FK935 (holotype);

e. FK1111. N. atropapillata: f. FK649; g. FK898 (holotype). N. pallidosalmonea: h. (FK892,

holotype). N. parvispora: i. FK664 (holotype); j. FK689; k. FK794. N. tricholomatoidea: |. (FK1049,

holotype). Scale bars = 1 cm. Photos: a, b, d-j, 1: F. Karstedt; c, k: M. Capelari.

Hasitat—Solitary or sometimes gregarious, on litter or in soil with

fragmented organic material.

ADDITIONAL SPECIMENS EXAMINED—BRAZIL, SAO PauLo STATE: Sao Paulo,

Parque Estadual da Cantareira, Nucleo Engordador, 23°20’S 46°41’W, Macuco

trail, 27.1V.2006, EF Karstedt & M. Capelari FK630 (SP); 29.VI.2006, F. Karstedt &

M. Capelari FK0690 (SP); 19.IX.2006, E Karstedt & M. Capelari FK0735 (SP);

Cachoeira trail, 20.III.2007, F. Karstedt & al. FK0883 (SP); 29.V.2007, F. Karstedt &

al. FK0912 (SP); 13.III.2008, F. Karstedt FK1101 (SP); Parque Estadual das Fontes do

Ipiranga, 23°39’S 46°37’W, Nascente trail, 20.V1.2006, F. Karstedt & al. FK0682 (SP);

28.VIII.2008, FE. Karstedt & K. Patekoski FK1111 (SP); Santo André, Reserva Bioldgica

de Paranapiacaba, 23°46’S 46°18’W, trail 15, 18.V.2006, F. Karstedt & M. Capelari

FK0651 (SP), FK0652 (SP); trail 2, 18.V.2006, F. Karstedt & M. Capelari FK0656 (SP);

trail 3, 22.VI.2006, F. Karstedt FK0685 (SP); trail 1, 23. VIII.2006, F. Karstedt FK0727

598 ... Karstedt, Bergemann, Capelari

(SP). SANTA CATARINA STATE: Gaspar, #1111 Pedro Schmitt Street, hill behind the

Joao Pedro Lenfers’ house, 02.VIII.2008, F. Karstedt & A.T. Karstedt FK1110 (SP).

RIO DE JANEIRO STATE: Parque Nacional do Itatiaia, 11.1.2011, EF Karstedt, PB.

Schwartsburd & J.B. Pereira FK1731 (SP; GenBank KF738937, KF738925), FK1732

(SP; GenBank KF771345, KF738938, KF738926).

CoMMENTS—WNolanea albertinae apparently isa common species in the Atlantic

Forest of southern and southeastern Brazil; additionally it is usually solitary

and almost undetectable in the litter, considering that both have similar color.

Nolanea albertinae is similar to N. atripes Dennis and N. pseudopapillata Pegler,

which are both described from Central America (Horak 1977, Pegler 1983). It

also resembles Entoloma dissimile (Singer) E. Horak (described from Argentina;

Singer 1969, Horak 1977), N. fuscifolia (Peck) Sacc. (described from the United

States (Peck 1874, Hesler 1963) but also found in Argentina; Singer 1969) and

Entoloma novum E. Horak, (described from Chile; Horak 1977). These species

have a similar stature including the mycenoid habit; a pileus that is brownish,

conical to plane, papillate and hygrophanous; a similar pileipellis with brown

pigmented repent hyphae; and similarly shaped and sized basidiospores,

7-10 x 5-7.5 um. All also can be separated morphologically besides lacking the

cylindrical to capitate cheilocystidia that characterize N. albertinae.

Nolanea atripes has a darker pileus, no cheilocystidia, and pileipellis with a

vacuolar pigment (Pegler 1983). Nolanea pseudopapillata, Entoloma dissimile

and Entoloma novum differ (from N. albertinae) by the absence of cheilocystidia

and clamp connections and by the presence of encrusted pileipellis hyphae

(Singer 1969, Horak 1977, Pegler 1983). Nolanea fuscifolia grows on wood

(instead soil or litter), has a dark brown pileus, lacks cheilocystidia, and

has clamp connections; additionally, N. fuscifolia was originally described

from New York (Peck 1874, Hesler 1963). Nolanea fuscifolia sensu Singer

(1969) from Argentina, is closer to Nolanea albertinae in its very infrequent

cheilocystidia and not (or only slightly) encrusted pigmentation; however it

is distinguished by its growth on dead wood and by the presence of fusoid-

ventricose caulocystidia.

The species most closely related to Nolanea albertinae is Entoloma belouvense

Noordel. & Hauskn. from the Republic of Seychelles, an island archipelago

in the Indian Ocean off the east coast of Africa (Noordeloos & Hausknecht

2007). Entoloma belouvense has similar macroscopic features and scattered

cheilocystidia that are cylindrical and often capitate. However, in addition to

its different distribution, N. albertinae has distinctly smaller basidiospores

(7.5-10 x 5-7.5 um versus 10-12 x 6-7,5 um) and the presence of refractive

hyphae at pileipellis.

Nolanea spp. nov. (Brazil) ... 599

Fic. 3. Nolanea albertinae (holotype, FK935):

a. Basidiospores; b. Basidia; c. Cheilocystidia; d. Pileipellis.

Scale bars: a-c = 10 um; d = 20 um.

600 ... Karstedt, Bergemann, Capelari

Nolanea atropapillata Karstedt & Capelari, sp. nov. FIGS. 25,G, 4, 8C,D

MB805603

Differs from N. albertinae, N. atripes, N. pseudopapillata, and Entoloma dissimile by its

bisporic basidia, by its intracellular and encrusted pigment in pileipellis hyphae, and by

the absence of clamp connections.

Type—Brazil, Sao Paulo State, Santo André: Reserva Bioldgica de Paranapiacaba,

23°46’S 46°18’W, Estrada Frederico Carlos Hoehne, 12.IV.2007, F. Karstedt & al. FK0898

(Holotype: SP, GenBank KF738940, KF738929).

EtryMoLocy—refers to the color of the papillae that are darker than the pileus.

PILEus 9-25 mm diam., convex, plano-convex or plane, papillate or umbonate,

brown or dark brown, darker in the center compared to the edge, turning

grayish brown or pale gray-brown with blackish papilla, surface smooth or

sometimes slightly squamulose-fibrillose at center, striate-translucent, strongly

hygrophanous then silky, margin slightly eroded or split and upturned with

age. PILEUS CONTEXT very thin. LAMELLAE almost free, narrowly adnexed,

pink or pink-brown, subdistant, with 1-5 lamellulae (2 tiers), margin smooth

or slightly eroded. St1pzE 20-55 x 1-7 mm, cylindrical, with subbulbous base

or slightly tapering upwards from swollen base, nearly white, pale beige or very

pale brown, apex that is paler than the base, fibrillose-twisted, hollow, base of

stipe often with a white tomentum. SPORE PRINT not recorded.

BASIDIOSPORES 8.7-12.5 x 6.2-8.7 um (Q = 1.16-1.6, Qm = 1.4),

heterodiametric, strongly angled, with 5-6 angles in profile, slightly brownish

pink, thin-walled. Basrp1a 20-35 x 7.5-10 um, clavate, hyaline, thin-walled,

bisporic. LAMELLAE EDGE fertile. CHEILOCYSTIDIA and PLEUROCYSTIDIA

absent. LAMELLAR TRAMA made up of regular cylindrical or fusiform, hyaline,

thin-walled hyphae, 6.2-26 um diam.; subhymenium ramose. PILEITRAMA

composed of radially arranged and parallel cylindrical or fusiform, hyaline,

thin-walled hyphae, 3.7-37 tm diam. PILEIPELLIS a cutis, suprapellis made

up of cylindrical hyphae, with brown intracellular pigment, sometimes

with brown encrusted pigment, hyphae thick or thin-walled, 2.5-18 um

diam.; terminal elements repent or anticlinal and then almost a trichoderm,

cylindrical or cylindrical-clavate at center of pileus; subpellis composed of

inflated or fusiform, hyaline or slightly straw yellow, thin-walled hyphae,

13.7-44 um diam. STIPITIPELLIS a cutis made up of hyaline or slightly straw

yellow, thin-walled hyphae, 2.5-10 um diam. CAULOCysTIDIA absent. CLAMP

CONNECTIONS absent. REFRACTIVE HYPHAE not observed.

HasBitaT—Solitary or dispersed in litter.

ADDITIONAL SPECIMENS EXAMINED—BRAZIL, SAO PAuLo State, Santo André:

Reserva Bioldgica de Paranapiacaba, 23°46’S 46°18’W, trail 12, 18.V.2006, F. Karstedt

Nolanea spp. nov. (Brazil) ... 601

Fic. 4. Nolanea atropapillata (holotype, FK898): a. Basidiospores; b. Basidia; c. Pileipellis.

Scale bars: a, b = 10 um; c = 20 um.

& M. Capelari FK0649 (SP), FK0650 (SP); trail 6, 18.V.2006, F. Karstedt & M. Capelari

FK0655 (SP); Estrada Frederico Carlos Hoehne, 12.V.2007, E. Karstedt & al. FK0899

(SP).

CoMMENTS—Nolanea atropapillata resembles N. albertinae, N. atripes,

N. pseudopapillata, and Entoloma dissimile from South America and

N. fuscifolia from the United States. All of these species have papillate, pale

brown pilei with a darker center and a translucent-striate surface and

heterodiametric basidiospores in the 8-13 x 6-9 um range. However, all

differ from N. atropapillata in producing tetrasporic basidia and clamp

connections. Nolanea atripes is further distinguished by its slender basidioma,

membranaceous pileus, and black stipe (Dennis 1961). The description of

N. atripes by Pegler (1983) differs from the Dennis's type description, in which

Dennis (1961) emphasized the contrast between the black stipe and gray-

brown pileus, while Pegler (1983) described the stipe as pale brown to dark,

and the basidiomata illustrated is much more robust.

Nolanea albertinae also differs by the presence of cheilocystidia and absence

of encrustations in the pileipellis hyphae, and Entoloma dissimile differs by

the pileipellis as a cutis without a differentiated subpellis. Nolanea fuscifolia

is distinguished by its subnodulose basidiospores (Hesler 1963, Singer 1969)

and N. pseudopapillata by its strongly heterodiametric basidiospores (Q = 1.61,

Pegler 1983).

602 ... Karstedt, Bergemann, Capelari

Nolanea pallidosalmonea Karstedt & Capelari, sp. nov. FIGS. 2H, 5, 8E,F

MB805604

Nolanea pallidosalmonea differs from other Nolanea spp. by its delicate and slender

basidiomata, by a pale pinkish salmon pileus with salmon pink striations and slightly

brownish center, by 5-6-angled heterodiametric basidiospores, by a pileipellis composed

of repent hyphae, and by the presence of refractive hyphae.

Type—Brazil, Sao Paulo State, Santo André: Reserva Bioldgica de Paranapiacaba,

23°46’S 46°18’W, trail 1, 23.11.2007, F. Karstedt & al. FK0892 (Holotype, SP; isotype K).

EtryMoLoGy—the name refers to the pale salmon pink coloration of the basidioma.

PitEus 4-15 mm diam., convex, slightly papillate, when young with more

pronounced papillae, very pale salmon pink or whitish salmon pink, darker

at the center and strongly translucent-striate when in fresh condition,

hygrophanous and then whitish from the margin towards the center and

finally whitish salmon pink and when dry, sometimes slightly brownish at

center, translucent-striate, after opaque and silky-fibrillose, margin entire.

PILEUS CONTEXT thin, translucent. LAMELLAE adnexed or sinuate, pale pink,

ventricose, subdistant, with 2 tiers of lamellulae. Stipe 25-54 x 1-3 mm,

cylindrical or slightly tapering, pale salmon pink or light brownish, hollow,

fibrous, with a small amount of white tomentum at the base. SPORE PRINT not

recorded.

BasIDIosPpoREs 8.7-10 x 6.2-7.5 um (Q = 1.16-1.6; Qm = 1.41),

heterodiametric, strongly angled, with (5)6-7 angles in profile view, hyaline

or slightly pinkish brown, thin-walled. Basip1a 25-37 x 8.7-11.2 um, clavate,

hyaline, thin-walled, tetrasporic. PLEUROCYSTIDIA and CHEILOCYSTIDIA

absent. LAMELLAE EDGE fertile. LAMELLAR TRAMA subparallel, hyphae with

5-20 um diam., cylindrical or slightly inflated, sometimes fusiform, hyaline,

thin-walled; subhymenium ramose. PILEITRAMA trama composed of regular

cylindrical, inflated or fusiform, hyaline, thin-walled hyphae, 11.2-26 um

diam. PILEIPELLIS a cutis made up of cylindrical, hyaline, thin-walled hyphae,

2.5-8.7 um diam.; subpellis made up of inflated or fusiform, hyaline, thin-walled

hyphae, 10-21 um diam. STIPITIPELLIs a cutis of hyaline, thin-walled hyphae,

3.7-12.5 um diam. CauLocysTipia not observed. CLAMP CONNECTIONS

absent. REFRACTIVE HYPHAE abundant in pileipellis.

HaBITAT— Gregarious or scattered, on soil.

ADDITIONAL SPECIMEN EXAMINED—BRAZIL, SAo PAULO STATE, Santo André:

Reserva Bioldgica de Paranapiacaba, 23°46’S 46°18’W, trail 1, 23.11.2007, FE. Karstedt &

al. FK0891 (SP; GenBank KF738941, KF738930).

ComMMENTS—Nolanea pallidosalmonea resembles Entoloma quadratum (Berk.

& M.A. Curtis) E. Horak, cited from Borneo, Singapore, Costa Rica, United

Nolanea spp. nov. (Brazil) ... 603

JOYSHOLOHOEGOOLY

Fic. 5. Nolanea pallidosalmonea (holotype, FK892): a. Basidiospores; b. Basidia; c. Pileipellis.

Scale bars: a, b = 10 um; c = 20 um.

States, Madagascar, Malaysia and Papua New Guinea, in its conical pileus and

a smooth surface (a cutis) that is salmon colored. However, E. quadratum is

clearly separated from N. pallidosalmonea by its cuboidal basidiospores and

clavate cheilocystidia (Horak 1975, 1977, Baroni & Halling 2000).

Nolanea minuta P. Karst., described from Europe, is another species with

pale and pinkish basidiomata, a silky-fibrillose pileus surface, heterodiametric

basidiospores, and absence of cheilocystidia. However, N. minuta differs from

N. pallidosalmonea in basidiospores with 5-7 angles in profile view, and a

pileitrtama composed of hyphae with intracellular and encrusted pigments

(Noordeloos 1980).

Nolanea parvispora Karstedt & Capelari, sp. nov. FIGS. 21-K, 6, 8G-I

MB805605

Nolanea parvispora differs from other Nolanea spp. by its lignicolous habitat, pale beige

pileus with dark center, stipe with a dark base and pale apex, sinuate lamellae, small

(6.2-7.5 x 6.2-7.5 um) isodiametric basidiospores, a lack of cystidia, and a pileipellis

composed of repent hyphae with dispersed groups of anticlinal hyphae.

TypE—Brazil, Sao Paulo State, Santo André: Parque Estadual da Cantareira, 23°20’S

46°41’W, Nucleo Engordador, Cachoeira trail, 25.V.2006, E Karstedt & M. Capelari

FK0664 (Holotype, SP).

EtyMoLoGcy—refers to the small size of the basidiospores.

PiLteEus 13-23 mm diam., convex, plano-convex or plane, sometimes with a

slightly or prominent papilla or slightly depressed at the center, whitish beige

604 ... Karstedt, Bergemann, Capelari

or pale brownish beige or light pinkish beige, brown at the center (insertion

of stipe), smooth and sometimes pruinose at the center, glabrous, translucent-

striate, margin revolute, even, or sometimes lobate. PILEUS CONTEXT very

thin. LAMELLAE sinuate, white or pinkish beige, abundant, with 5-7 lamellulae

(2 series), margin even. STIPE 25-42 x 2-4 cm, cylindrical or flattened,

brown or brownish translucent, beige at the apex, surface smooth, base with

white tomentum. SPORE PRINT pinkish. BASIDIOSPORES 6.2-7.5 x 6.2-7.5

um (Q = 1-1.2; Qm = 1.1), isodiametric, with 5-6 angles in profile view,

slightly pinkish, thin-walled. Bastp1a 27-43 x 8.7-10 um, clavate, hyaline,

thin-walled, tetrasporic. PLEUROCYSTIDIA and CHEILOCYSTIDIA absent.

LAMELLAE EDGE fertile. LAMELLAR TRAMA subregular, cylindrical, inflated

or fusiform, hyaline, thin-walled hyphae 3.7-12.5 um diam.; subhymenium

ramose. PILEITRAMA regular, made up of cylindrical, inflated or fusiform,

hyaline, thin-walled hyphae, 3.7-25 um diam. PILEIPELLIS a cutis composed

of cylindrical, hyaline or pale straw yellow, thin-walled hyphae, 6.2-13.7 um

diam., with dispersed or clustered thin-walled cylindrical or clavate, hyaline or

pale straw yellow anticlinal terminal elements; subpellis made up of inflated or

fusiform, hyaline, thin-walled hyphae 10-20 um diam. STIPITIPELLIs a cutis,

made up of hyaline or pale straw yellow, sometimes with encrusted (external)

yellow-brown pigment, thin or thick-walled hyphae, 6.2-16.2 um diam.

CAULOCYSTIDIA rare, cylindrical, clavate or ventricose, with pale straw yellow

plasmatic pigment, thin-walled, 28-75 x 2.5-7.5 um. CLAMP CONNECTIONS

absent. REFRACTIVE HYPHAE present in pileipellis.

HasitatT—Solitary or gregarious, on hardwood.

ADDITIONAL SPECIMENS EXAMINED—BRAZIL, SAO PAULO STATE, Sao Paulo:

Parque Estadual da Cantareira, Nucleo Engordador, 23°20’S 46°41’W, Macuco trail,

29.V1.2006, F. Karstedt & M. Capelari FK689 (SP); Cachoeira trail, 29.VI.2006, F.

Karstedt & M. Capelari FK0696 (SP); 25.VII.2006, E. Karstedt & M. Capelari FK0723

(SP); 24.X.2006, E. Karstedt & M. Capelari FK794 (SP); 30.X.2007, E Karstedt & al.

FK0926 (SP). SANTA CATARINA STATE, Joinville: Reserva Particular de Patrimdénio

Natural Caetezal, 02.1].2009, F. Karstedt FK1140 (SP; GenBank KF771348, KF738943,

KF738931); Corupa, Reserva Particular de Patriménio Natural Emileo F. Battistella,

falls route, 12.XII.2011, FE Karstedt FK2135 (FLOR; GenBank KF738932).

CoMMENTS—Rhodophyllus avellanicolor Romagn. & Gilles, Nolanea

mazophora (Berk. & Broome) Pegler, Entoloma myceliosum E. Horak, and

Entoloma imbecille (E. Horak) E. Horak ex Segedin & Pennycook [= E. fragile

E. Horak, nom. illegit.] are among the few species that resemble Nolanea

parvispora based on pale basidiomata and isodiametric basidiospores

measuring c. 7 x 7 um. Rhodophyllus avellanicolor, described from Africa, and

Nolanea spp. nov. (Brazil) ... 605

506909HQGO6O

Fic. 6. Nolanea parvispora (holotype, FK664):

a. Basidiospores; b. Basidia; c. Caulocystidia; d. Pileipellis.

Scale bars: a-c = 10 um; d = 20 um.

Nolanea mazophora, described from Sri Lanka, are terrestrial. Rhodophyllus

avellanicolor has clamp connections (Romagnesi & Gilles 1979) and

N. mazophora has a convex pileus with prominent papillae (Pegler 1977,

Horak 1980).

Entoloma myceliosum, described from Chile, is also lignicolous but differs

from N. parvispora by having a white pileus and stipe and a pileus surface

densely covered by white fibrils (Horak 1977). Entoloma imbecille, found in

New Zealand (Horak 1973, as “E. fragile”) and Argentina (Horak 1977, as

“E. fragile”), differs by a pileus that is initially brown and then turns beige, brown

encrusted pigments in the pileipellis hyphae, and abundant clamp connections.

606 ... Karstedt, Bergemann, Capelari

Nolanea tricholomatoidea Karstedt & Capelari, sp. nov. FIGS. 2L, 7, 8J,K

MB805607

Nolanea tricholomatoidea differs from other Nolanea spp by its robust stature, a

mostly beige pileus that turns brownish beige at the center; its whitish beige stipe,

its often pentagonal basidiospores; its basidial basal clamp connections at the base of

the basidia; and the encrusted pigment in the subpellis hyphae.

Type—Brazil, Sao Paulo State, Santo André: Parque Estadual da Cantareira, Nucleo

Engordador, 23°20’S 46°41’W, Cachoeira trail, 31.1.2008, F. Karstedt, L.A. Silva

Ramos & M. Capelari FK1049 (Holotype, SP; GenBank KF771347, KF738939,

KF738928).

ETyMoLoGy—the name refers to the tricholomatoid stature.

PiLEuS 80-92 mm diam., conico-campanulate or plane, umbonate, pale

beige, beige or brownish beige at center, surface smooth or slightly pruinose

at center, opaque, hygrophanous, translucent-striate at margin. PILEUS

CONTEXT white. LAMELLAE adnexed (almost free), beige, ventricose,

subdistant, with 2 tiers of lamellulae. Stipe 100-116 x 8-12 mm, and 39 mm

at base, subclavate, beige or whitish, fibrous and fibrillose, with white basal

mycelium. Opor indistinct. SPORE PRINT not recorded.

BASIDIOSPORES 8.7-10 x 7.5-8.7 um (Q = 1-1.33; Qm = 1.2), iso-

diametric or subisodiametric, strongly angled, with 5-6 angles in profile

view, with an obvious hilar appendix, hyaline or slightly brownish pink,

thin-walled. Basrp1a 37-51 x (8.7—)10-15 um, clavate with a narrow base,

hyaline, thin-walled, tetrasporic. LAMELLAE EDGE fertile. PLEUROCYSTIDIA

and CHEILOCYSTIDIA absent. LAMELLAR TRAMA regular and hyaline, with

hyphae 3.7-17.5 um diam., cylindrical or slightly inflated, hyaline, thin-

walled, septate; subhymenium ramose. PILEITRAMA regular, composed

of cylindrical or inflated, hyaline, thin-walled hyphae 3.7—27 um diam.

PILEIPELLIS a cutis, made up of cylindrical, hyaline, thin-walled hyphae

2.5-8.7 um diam., with rare anticlinal elements; subpellis composed

of inflated, hyaline, thin-walled hyphae 16.2-46 x 50-120 um diam.

STIPITIPELLIS a cutis made up of straw-yellow, hyaline, thin-walled hyphae,

3.7-7.5 um diam. CAULOCYSTIDIA absent. CLAMP CONNECTIONS present in

the hymenium. REFRACTIVE HYPHAE present in the pileipellis.

HABITAT—Solitary, on soil.

ADDITIONAL SPECIMEN EXAMINED—BRAZIL. Rio DE JANEIRO STATE, Nova

Iguacu: Reserva Bioldgica de Tingua, Ouro road, 8 Dec 2004, M.P. Albuquerque &

A.A. Carvalho-Jr. 29-VI (RB415680).

COMMENTS—Nolanea tricholomatoidea was collected in Rio de Janeiro

State but reported as Entoloma lividoalbum (Kihner & Romagn.) Kubicka

Nolanea spp. nov. (Brazil) ... 607

0O9HCO0G000

Fic. 7. Nolanea tricholomatoidea (holotype, FK1049):

a. Basidiospores; b. Basidia; c. Pileipellis.

Scale bars: a, b = 10 um, c = 20 um.

(Albuquerque & al. 2007). The robust stature is reminiscent of a feature

common to Entoloma sensu stricto, and based mainly on its stature, its

umbonate pileus that is sometimes translucent-striate, and its white stipe,

N. tricholomatoidea might seem related to species of Entoloma s.s., such as

E. lividoalbum and E. prunuloides (Fr.) Quél.

Entoloma lividoalbum, described from Europe and recorded from

the United States, differs in its pileus that varies from dark brown, dark

yellowish brown, pale yellowish white to pale grayish beige, and clamp

connections that are present in all tissues (Largent 1994, Noordeloos &

Polemis 2008). Entoloma prunuloides, recorded from Europe, differs in its

smaller basidiospores (6.5-8 x 6.5-8 um), an ixocutis-type pileipellis, and

the presence of clamp connections in all tissues (Noordeloos 1992).

Although a tricholomatoid stature is uncommon in Nolanea, the

pileipellis structure in N. tricholomatoidea, comprising a subpellis with

inflated elements and a suprapellis with thin elements, is a feature shared

among many Nolanea spp.

608 ... Karstedt, Bergemann, Capelari

Discussion

The phylogenetic analyses as supported by ML, MP, and BPP cluster the

species described here and other species traditionally accepted as Nolanea

within a single clade, the /Nolanea clade (Fia. 1).

Four of the species described in this paper—Nolanea albertinae,

N. atropapillata, N. pallidosalmonea, and N. tricholomatoidea—occur in

litter or soil and have heterodiametric basidiospores. They cluster in clade

I. Nolanea parvispora, the only lignicolous species and characterized by

isodiametric basidiospores, clusters in clade II.

Species that are placed in Nolanea often have a mycenoid habit (Henkel

& al. 2014, Largent 1994, Noordeloos 1992, 2004, Noordeloos & Gates

2012). Nolanea tricholomatoidea described herein and the two species

classified as Entoloma s.s. (E. pallideradicatum Hauskn. & Noordel. and

E. valdeumbonatum Noordel. & Meusers) have a tricholomatoid habit and

yet are included in the /Nolanea clade. These species share with Nolanea

species a pileipellis with a subpellis composed of inflated hyphae and

heterodiametric basidiospores.

Curiously, all species studied and almost all species (that have been

published with a detailed description of the pileipellis) in the /Nolanea

clade have a pileipellis composed of a suprapellis with narrow hyphae and

a subpellis with inflated hyphae. We recommend paying careful or close

attention to this character in the future to determine if these features are

phylogenetically informative.

Acknowledgments

The authors are grateful to Dr. David L. Largent and Ms. Kerri Kluting for the

donation of collections that were included in DNA amplification and the phylogenetic

analyses. A special thanks to Dr. David L. Largent and Dr. Genevieve M. Gates

for their suggestions, and Dr. Timothy J. Baroni and Dr. Felipe Wartchow for the

reviews. This study was supported by the grants FAPESP 2006/58549-4 to Fernanda

Karstedt (master’s grant) and FAPESP 2004/04319-2 to Marina Capelari provided by

Fundacao de Amparo a Pesquisa do Estado de Sao Paulo. M. Capelari also thanks the

CNPg - Conselho Nacional de Desenvolvimento Cientifico e Tecnoldgico.

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MYCOTAXON

July-September 2020—Volume 135, pp. 613-616

https://doi.org/10.5248/135.613

First record of Trappea darkeri from Turkey

YASIN UZUN', OSMAN BERBER’, ABDULLAH KAYA?

2 Department of Biology, Science Faculty, Karamanoglu Mehmetbey University,

70100, Karaman, Turkey

* Department of Biology, Science Faculty, Gazi University, 06560, Ankara, Turkey

“ CORRESPONDENCE TO: kayaabd@hotmail.com

ABSTRACT—The truffle-like basidiomycete genus Trappea is reported as a new record from

Turkey and western Asia, based on a collection of T: darkeri from Nigde province. This

is also the first record of the family Trappeaceae for the country. A brief description and

photographs of the collection is provided.

Key worps—Basidiomycota, biodiversity, Hysterangiales, taxonomy

Introduction

Trappea Castellano is a truffle-like fungi genus in the family Trappeaceae

(Kirk & al. 2008). The genus was proposed by Castellano (1990), who

segregated T: darkeri and T: phillipsii from Hysterangium Vittad. based on

their smooth bacilloid basidiospores and layer of sterile locules below the

peridium. Trappea species are characterized by subglobose to irregularly lobed

fruit bodies with one or more rhizomorphs emerging from the base; a thin

white peridium covering a zone of sterile chambers; an olive- brown to bright

olive-green gleba with small empty chambers and a dendroid, gelatinous to

cartilaginous columella; and ellipsoid to oblong basidiospores.

Index Fungorum (www.indexfungorum.org accessed 25 June 2019) lists

four Trappea species, none of which has been reported from Turkey (Acar

& al. 2019, Sesli & Denchev 2014, Solak & al. 2015, Turkekul 2017). Here

we contribute to the Turkish mycobiota and report T: darkeri as the first

representative of the genus and family in Turkey.

614 ... Uzun, Berber, Kaya

Materials & methods

The Trappea specimen was collected from Ulukisla district, Nigde province, in

2019. The basidiomata were photographed in their natural habitat, where ecological

and morphological observations were taken. Dried material was examined and

photographed microscopically in the fungarium using a Nikon Eclipse Ci-S

trinocular compound microscope with Nikon DS-Fi2 camera and a Hitachi

SU5000 scanning electron microscope. The specimen was identified by consulting

the literature (Castellano 1990, Desjardin & al. 2014, Gémez-Reyes & al. 2014,

Montecchi & Sarasini 2000, Ruini 1990, Zeller 1939). The collection is preserved in

the fungarium of Department of Biology, Science Faculty, Karamanoglu Mehmetbey

University, Karaman, Turkey (KMU).

Taxonomy

Trappea darkeri (Zeller) Castellano, Mycotaxon 38: 3 (1990) FIG. 1

BasIDIOMA 40 mm in diam, hypogeous to semi-hypogeous, globose,

surface glabrous, with basal rhizomorphs at the base, whitish, light brownish

when rubbed. PERIpIUM whitish in section, not separable, membranous-

leathery. GLEBA firm, bright olive-green, olive gray to light olive-brown,

white near the peridium, spongy gelatinous. COLUMELLA distinct, gelatinous,

translucent, typically dendroid and reaching almost throughout the

sporocarp. Odor unpleasant. Basip1A 19-25 x 5-8 um, irregularly cylindrical

to clavate, (4-)6(-8)-spored, clamp connections present. CysTipIA not

observed. BASIDIOSPORES 4-5 x 2-3 um, ellipsoid to cylindrical, smooth,

hyaline, sessile or with a short pedicel.

SPECIMEN EXAMINED—TURKEY, NiGDE: Ulukisla, Ciftehan village, 37°30’N 34°47’E,

930 m, in soil among needle-litter in pine-oak mixed forest, 23.03.2019, O.Ber 342

(KMU).

CoMMENTS— Trappea darkeri has a hypogeous to semi-hypogeous habit with

long rhizomorphs at the base and is distributed in high altitudes of 1000-

3000 m (Castellano 1990, Desjardin & al. 2015, Gomez-Reyes & al. 2014).

It is distinguished by a layer of sterile locules just under the peridium, and

small, cylindrical spores (G6mez-Reyes & al. 2014). Morphologically it may

be confused with Hysterangium species with rubbery-gelatinous olive-green

gleba, which differ in their easily separable peridia, absence of sterile locules

next to the peridium and larger, spindle-shaped spores (Desjardin & al.

2015, Siegel & al. 2019). Ecologically and morphologically similar, Trappea

pinyonensis States can easily be distinguished from T. darkeri by the staining

reactions of the peridium and rhizomorphs and its longer (5-6.2 x 2-2.5 um)

spores (States 1991).

Trappeaceae newly recorded for Turkey... 615

. ow ;

Fic. 1. Trappea darkeri (KMU - Ber 342). a. Basidiomata; b. Basidia and basidiospores;

c-e. Basidiospores (SEM). Scale bars: b = 10 um; c-e = 3 um.

Although our specimen was collected at a slightly lower elevation

(930 m) than cited elsewhere, its morphological characters agree with those

cited in the literature (Castellano 1990, Desjardin & al. 2015, Gomez-Reyes

& al. 2014, Montecchi & Sarasini 2000, Ruini 1990, Zeller 1939).

Acknowledgments

The authors would like to thank Dr. Ali Keles (Yuziincti Yil University, Van,

Turkey), Prof. Dr. [brahim Tiirkekul (Gaziosmanpasa University, Tokat, Turkey)

and Nomenclature Editor Dr. Shaun Pennycook for their helpful comments and

careful review.

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616 ... Uzun, Berber, Kaya

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from Hysterangium. Mycotaxon 38: 1-9.

Desjardin DE, Wood MG, Stevens FA. 2014. California mushrooms, the comprehensive

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Sesli E, Denchev CM. 2014. Checklists of the myxomycetes, larger ascomycetes, and

larger basidiomycetes in Turkey. 6th ed. Mycotaxon Checklists Online. 136 p.

http://www.mycotaxon.com/resources/checklists/sesli-v106-checklist.pdf

Siegel N, Vellinga EC, Schwarz C, Castellano MA, Ikeda D. 2019. A field guide to the rare fungi

of California’s National Forests. Bookmobile: Minneapolis, MN. 313 p.

Solak MH, Isiloglu M, Kalmus E, Alli H. 2015. Macrofungi of Turkey, checklist, vol. 2. Izmir,

Turkey: Universiteliler Ofset [in Turkish].

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127-133.

Trappe M, Evans E Trappe J. 2007. Field guide to North American truffles. Berkeley:

Ten Speed Press.

Tirkekul I. 2017. New Calbovista, Mycena, Rhizopogon, Stictis, and Symphyosirinia records

from Turkey. Mycotaxon 132: 503-512. https://doi.org/10.5248/132.503

Zeller SM. 1939. New and noteworthy gasteromycetes. Mycologia 31(1): 1-32.

https://doi.org/10.2307/3754429

MYCOTAXON

July-September 2020—Volume 135, pp. 617-622

https://doi.org/10.5248/135.617

Mesocorynespora sinensis gen. & sp. nov.

from southern China

ZHAO-HUAN XU’, KAI ZHANG’, YOU-QIANG LUO’,

Xi1u-Guo ZHANG?, RAFAEL F. CASTANEDA-RuiIz‘, JIAN Ma‘*

' College of Agronomy, Jiangxi Agricultural University, Nanchang, Jiangxi 330045, China

? Department of Landscaping, Shandong Yingcai University, Jinan, Shandong 250104, China

> Shandong Provincial Key Laboratory for Biology of Vegetable Diseases and Insect Pests,

College of Plant Protection, Shandong Agricultural University,

Taian, Shandong 271018, China

‘Instituto de Investigaciones Fundamentales en Agricultura Tropical Alejandro de Humboldt

(INIFAT), Académico Titular de la Academia de Ciencias de Cuba,

Calle 1 Esq. 2, Santiago de Las Vegas, C. Habana, Cuba, C.P. 17200

* CORRESPONDENCE TO: majian821210@163.com; jxaumj@126.com

ABSTRACT—A new anamorphic genus and species, Mesocorynespora sinensis collected on

decaying culms of bamboo in China, is described and illustrated. The fungus is distinguished

by short, unbranched, clavate conidiophores with monotretic, conidiogenous cells that

produce solitary, acrogenous, obclavate, euseptate conidia. A key to Mesocorynespora and

morphologically similar genera is provided.

KEY worps—asexual fungi, hyphomycetes, taxonomy

Introduction

Lushan Mountain, located in the northern part of Jiangxi Province, China,

covers approximately 302 km’ and exhibits a unique geography with complex

terrains and unusual climate. The mountain's distinctive vegetation has favored

the survival and multiplication of various microbial species, yet only relatively

little information has been published on saprobic hyphomycetes recorded in

China (e.g., Ma & Zhang 2015, Ma 2016, Ma & al. 2016, Xu & al. 2017, Ai &

al. 2019). During our continuing surveys of asexual fungi in this region, an

618 ... Xu &al.

interesting fungus was collected that differed remarkably from all previously

described hyphomycetes (Seifert & al. 2011). Thus, it is described here as a new

genus and species.

Taxonomy

Mesocorynespora Jian Ma, X.G. Zhang & R.F. Castaneda, gen. nov.

MB 836511

Differs from Solicorynespora by its short, determinate, clavate conidiophores and multi-

euseptate conidia; and from Corynespora by its euseptate conidia.

TYPE SPECIES: Mesocorynespora sinensis Jian Ma & al.

ErymMo.Loey: Greek, meso- meaning middle- + Latin, -corynespora, referring to the

genus Corynespora.

CONIDIOPHORES short, single or in groups, clavate, unbranched, brown.

CONIDIOGENOUS CELLS monotretic, integrated, terminal, clavate, determinate.

Conidial secession schizolytic. Conrp1A solitary, acrogenous, dry, obclavate,

multi-euseptate.

Mesocorynespora sinensis Jian Ma, X.G. Zhang & R.F. Castafieda, sp. nov. Fics 1, 2

MB 836512

Differs from Solicorynespora spp. by its short, determinate conidiophores with multi-

euseptate conidia, and from Corynespora spp. by its euseptate conidia.

Type: China, Jiangxi Province: Lushan Mountain, on decaying culms of bamboo,

9 November 2014, J. Ma (holotype, HJAUP M0296).

ETyMoOLoGy: refers to China where the fungus was collected.

COLONIES on natural substrate effuse, brown to dark brown. Mycelium

superficial and immersed, composed of branched, septate, pale brown to

brown, smooth-walled hyphae. ConipiopHorgs short, single or in groups,

erect, straight or flexuous, clavate, unbranched, brown, smooth, 1-3-septate,

25-40 x 3.5-5 um. CONIDIOGENOUS CELLS monotretic, integrated, terminal,

determinate, clavate, brown, smooth, 9-14.5 x 5-6.5 um. Conidial secession

schizolytic. Conrp14 solitary, dry, acrogenous, obclavate, smooth, brown

to dark brown, rounded and golden yellow at the apex, truncate at the base,

20-35-euseptate, 72-171 x13.5-16 um, tapering to 7-10.5 um diam. at the

apex, 4-5 um diam. at the base.

Discussion

Mesocorynespora sinensis is unique in having short, clavate, unbranched

conidiophores, and solitary, acrogenous, obclavate, multi-euseptate conidia

Mesocorynespora sinensis gen. & sp. nov. (China) ... 619

wig]

Fic. 1. Mesocorynespora sinensis (holotype, HJAUP M0296).

A. Developing conidia; B. Conidiophores with developing conidia;

C. Conidiophores, and conidiogenous cells.

620 ... Xu &al.

20um

Fic. 2. Mesocorynespora sinensis (holotype, HJAUP M0296).

A-C. Conidiophores, conidiogenous cells, and conidia; D. Conidia.

that secede schizolytically from the monotretic, integrated, terminal,

determinate conidiogenous cells. We initially thought of assigning it to the

genus Solicorynespora R.F. Castafteda & W.B. Kendr. (Castafeda-Ruiz &

Kendrick 1990), but Solicorynespora species have long, cylindrical to obclavate

conidiophores with percurrently extending conidiogenous cells and conidia

with fewer septa.

Several other genera including Corynespora Giissow, Corynesporella Munjal

& H.S. Gill, Hemicorynespora M.B. Ellis, Corynesporopsis P.M. Kirk, and

Corynesporina Subram. (Giissow 1905, Munjal & Gill 1961, Ellis 1972, Kirk

1981, Gams & al. 2009) have the same conidiogenesis as Mesocorynespora, but

Mesocorynespora differs in having clavate, determinate conidiogenous cells

Mesocorynespora sinensis gen. & sp. nov. (China) ... 621

formed from short, clavate, unbranched conidiophores; and differs further

from Corynespora, Corynesporella, and Corynesporina by its euseptate conidia

and from Corynesporopsis by its solitary conidia. Conidia in Hemicorynespora

are 0-1-septate, an obvious difference from Mesocorynespora.

Key to Mesocorynespora and morphologically similar genera

1. Conidiophores short, conidiogenous cells

Wwithoutpercurtent extension § 2.25. 51.5 aed ng ee eee Mesocorynespora

1. Conidiophores long, conidiogenous cells with percurrent extension............ 2

2sConidig-aseptate oreuseptate 6 hic. wie tae ok ete whom ce ery ee pA eg 3

DAG QIN IC AGH SEOSCDLALEY tsi age ber sel gia cite ages ses soneet vara a wtbeneed ac seet astt 9 wiica byeUe Ey arise AAs ae 5

8 Conidia-catenates si 4 ous. trons Hd nh Fed Wee Ek ee OREO Corynesporopsis

SiS onidia-solitaryes tan te shaes bot atethe ue ae kone ane tena ee ec eaece 4

4. Conidia aseptate or with a single septum ..................... Hemicorynespora

A ONIdia tWO*.t6-pIMPISeptaler sce. anesthe a ants cs Ee oh Solicorynespora

5. Conidiogenous cells formed as terminal and lateral branchlets

On The CONIMIGPROTES srecse.t, sto. ste str somese neat ob Hen ge ae eeaE ste taba cae Corynesporella

5. Conidiogenous cells integrated, terminal on the conidiophores ................ 6

6. Gonidia formed'singlé or in.acropetal chains’ 2: ..0.25 4.5 orc mesees Corynespora

6. Conidia solitary and in basipetal chains .....................4.. Corynesporina

Acknowledgments

The authors express gratitude to Dr. Patricia Oliveira Fiuza (Programa de Pos-

graduacao em Sistematica e Evolu¢gao, Universidade Federal do Rio Grande do

Norte, Brazil) and Dr. De-Wei Li (The Connecticut Agricultural Experiment Station

Valley Laboratory, USA) for serving as pre-submission reviewers and to Dr. Shaun

Pennycook for nomenclatural review and Dr. Lorelei L. Norvell for editorial review.

This project was supported by the National Natural Science Foundation of China

(Nos. 31970018, 31360011, 31870016), and the Education Department of Jiangxi

Province of China (No. GJJ160357).

Literature cited

Ai CC, Ma J, Zhang K, Castaneda-Ruiz RF, Zhang XG. 2019. Cordana meilingensis and

C. lushanensis spp. nov. from Jiangxi, China. Mycotaxon 134: 329-334.

https://doi.org/10.5248/134.329

Castafieda-Ruiz RE, Kendrick B. 1990. Conidial fungi from Cuba: II. Univ. University of

Waterloo Biology Series 33. 61 p.

Ellis MB. 1972. Dematiaceous hyphomycetes. XI. Mycological Papers 131. 25 p.

Gams W, Seifert KA, Morgan-Jones G. 2009. New and validated hyphomycete taxa to resolve

nomenclatural and taxonomic issues. Mycotaxon 110: 89-108. https://doi-org/10.5248/110.89

Giissow HT. 1905 [”1904”]. Notes on a disease of cucumbers, II. Journal of the Royal Agricultural

Society of England 65: 271-272.

622 ... Xu &al.

Kirk PM. 1981. New or interesting microfungi IH. Dematiaceous hyphomycetes from

Esher Common, Surrey. Transactions of the British Mycological Society 77: 279-297.

https://doi.org/10.1016/S0007-1536(81)80031-9

Ma J. 2016. Corynesporopsis obclavata and Stanjehughesia jiangxiensis spp. nov. from Lushan

Mountain, China. Mycotaxon 131: 583-588. https://doi.org/10.5248/131.583

Ma J, Zhang XG. 2015. A preliminary report of dematiaceous hyphomycetes from

dead branches in Jiangxi Province. Biological Disaster Science 38(4): 290-293.

https://doi.org/10.3969/j.issn.20953704.2015.04.003

Ma J, Zhang XG, Castafieda-Ruiz RE 2016. Podosporiopsis, a new genus of synnematous

hyphomycetes from China. Mycotaxon 131: 773-780. https://doi.org/10.5248/131.773

Munjal RL, Gill HS. 1961. Corynesporella: a new genus of hyphomycetes. Indian Phytopathology

14(1): 6-9.

Seifert K, Morgan-Jones G, Gams W, Kendrick B. 2011. The genera of hyphomycetes. CBS

Biodiversity Series 9. 997 p.

Xu ZH, Hu DM, Luo YQ, Ma J. 2017. Three species of Linkosia and Spadicoides new to China.

Mycotaxon 132: 243-250. https://doi.org/10.5248/132.243

MYCOTAXON

July-September 2020—Volume 135, pp. 623-630

https://doi.org/10.5248/135.623

Corynesporopsis hainanensis sp. nov.,

a bambusicolous fungus from southern China

ZHAO-HUAN Xu’, XU-GEN SHI’, WEI-GANG KUANG’,

X1u-Guo ZHANG’, RAFAEL F. CASTANEDA-RuiIz}, JIAN Ma'*

' College of Agronomy, Jiangxi Agricultural University, Nanchang, Jiangxi 330045, China

? Department of Plant Pathology, Shandong Agricultural University,

Taian, Shandong 271018, China

3 Instituto de Investigaciones Fundamentales en Agricultura Tropical Alejandro de Humboldt

(INIFAT), Académico Titular de la Academia de Ciencias de Cuba,

Calle 1 Esq. 2, Santiago de Las Vegas, C. Habana, Cuba, C.P. 17200

* CORRESPONDENCE TO: majian821210@163.com jxaumj@126.com

ABSTRACT—A new anamorphic species, Corynesporopsis hainanensis, is described and

illustrated from dead bamboo culms collected in the tropical mountain rainforest of

Jianfengling, Hainan, China. The fungus is characterized by its terminal monotretic

conidiogenous cells with catenate, obclavate, brown to pale brown, smooth,

(3-)5-6(-10)-euseptate conidia. A dichotomous key and a synoptic table to Corynesporopsis

species are provided.

KEY worps—asexual fungi, hyphomycetes, saprobes, taxonomy, Xylariales

Introduction

Taking into account its large area, varied geographical conditions and

advantageous natural environments, China is considered an important

reservoir of the original bamboo diversity (Yang & Xue 1998). Its bamboo

forest ecosystems offer favorable habitats for survival and multiplication of

bambusicolous fungi. However, our knowledge of bambusicolous fungi is

scant, and relatively little information about their taxonomy has been published

in China (e.g. Zhou & al. 2001, Xu & al. 2006, 2007, Ma 2016). During our

ongoing surveys of saprobic microfungi associated with plant debris, we

624 ... Xu &al.

collected on dead bamboo culms an interesting hyphomycete with the typical

morphological features of Corynesporopsis P.M. Kirk (Kirk 1981a). The fungus,

significantly different from previously described taxa, is proposed here as new

to science.

Materials & methods

Samples of litter were placed in paper and plastic bags, taken to the laboratory, and

prepared according to Castafeda-Ruiz & al. (2016). Mounts were prepared in PVL

(polyvinyl alcohol and lactic acid) and measurements were made at a magnification

of x1000. Micrographs were obtained with a Nikon Eclipse 80i microscope equipped

with bright field and Nomarski interference optics. The type specimen was deposited in

the Herbarium of Jiangxi Agricultural University, Nanchang, Jiangxi, China (HJAUP).

Taxonomy

Corynesporopsis hainanensis Z.H. Xu, Jian Ma, X.G. Zhang &

R.F. Castafieda, sp. nov. Fig. 1

MB 836568

Differs from Corynesporopsis curvularioides, C. iberica, and C. rionensis by its

(3-)5-6(-10)-euseptate obclavate conidia; from C. iberica by its wider conidia; and from

C. curvularioides and C. rionensis by its longer and narrower conidia.

Type: China, Hainan Province, Jianfengling National Nature Reserve, on dead culms of

bamboo, 18 April 2014, J. Ma (Holotype, HJAUP M0377).

EryMo.oey: refers to the province where the type was collected.

CoLonigs on dead wood effuse, dark brown to black, hairy. Mycelium partly

superficial, partly immersed in the substratum, composed of branched, septate,

pale brown to brown, smooth hyphae. ConrpiopHor:s differentiated, single,

erect, unbranched, straight or slightly flexuous, brown to dark brown, smooth,

septate, 45-62 x 5.5-6.5 um, thick-walled. CONIDIOGENOUS CELLS monotretic,

integrated, terminal, determinate, cylindrical, brown, smooth, 10-13 um long,

4.5-5.5 um wide. Conidial secession schizolytic. CONIDIA acrogenous, catenate,

in unbranched, acropetal chains, obclavate, straight or curved, brown to pale

brown, smooth, (3-)5-6(-10)-euseptate, 17-48 um long, 6.5-8 um diam. in the

broadest part, 2-3 um diam. at the truncate base.

Dissussion

Corynesporopsis was established by Kirk (198la) with Corynespora

quercicola Borowska [= Corynesporopsis quercicola (Borowska) P.M. Kirk]

as the type species, and was mainly characterized by acrogenous, catenate,

euseptate conidia seceding schizolytically from monotretic, integrated,

Corynesporopsis hainanensis sp. nov. (China) ... 625

20 pm

Fic. 1. Corynesporopsis hainanensis (holotype, HHAUP M0377).

A, B. Conidiophores, conidiogenous cells, and conidia; C. Conidiophores; D. Conidia.

terminal, determinate or rarely percurrently extending conidiogenous cells.

It is similar to Corynespora Giissow, Corynesporella Munjal & H.S. Gill,

Hemicorynespora M.B. Ellis, Solicorynespora R.F. Castaheda & W.B. Kendr.,

and Corynesporina Subram. in having monotretic conidiogenous cells (Giissow

1905, Munjal & Gill 1961, Ellis 1972, Castafeda-Ruiz & Kendrick 1990, Gams

& al. 2009). However, Corynesporopsis differs from Hemicorynespora and

Solicorynespora by its catenate conidia and from Corynespora, Corynesporella,

626 ... Xu &al.

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Corynesporopsis hainanensis sp. nov. (China) ... 627

and Corynesporina by its euseptate conidia. Corynesporopsis is also similar to

Heteroconium Petr. (Petrak, 1949) in having catenate, euseptate conidia, but

Heteroconium is separated by monoblastic conidiogenesis.

Although Corynesporopsis was placed in Xylariales (Sordariomycetes) based

on ITS and LSU rDNA based phylogenetic studies of C. acaciae R. Kirschner,

its family remains incertae sedis due to its tretic conidiogenesis, previously

unrecorded for Xylariales (Kirschner 2015). Teleomorphs and phylogenetic

affinities with other Corynesporopsis species are not yet known.

Among the known species of Corynesporopsis, only C. curvularioides J.W.

Xia & X.G. Zhang (Xia & al. 2013), C. iberica R.F. Castafieda & al. (Castaneda-

Ruiz & al. 2011), and C. rionensis Hol.-Jech. (Holubova-Jechova & Mercado

Sierra 1986) are comparable to C. hainanensis. However, C. curvularioides

differs in its obclavate to ellipsoidal, shorter and wider conidia with 1-5

eusepta; C. iberica differs in its cylindrical, narrower conidia with (2-)3-7

eusepta; while C. rionensis differs in its wider fusiform, broadly fusiform, or

ellipsoidal conidia with 3—4(-5) eusepta.

With the addition of C. hainanensis, Corynesporopsis currently contains 17

species. A synopsis for comparisons of conidia and substrates of these accepted

species is presented in TABLE 1. The conidia of these 17 Corynesporopsis

species are mostly ellipsoidal, cylindrical, obclavate, or fusiform, but conidia

may also be obovoid, naviculiform, sub-oblong, sub-pyriform, or oval. The

conidia are mostly unicolored, but C. antillana R.F. Castaheda & W.B. Kendr.,

C. inaequiseptata Matsush., C. quercicola, and C. variabilis R.F. Castaneda & al.

produce versicolored conidia. Eight species produce predominately 1-septate

conidia, usually thickened and darkened at the septa—C. acacia, C. cylindrica

B. Sutton, C. indica P.M. Kirk, C. isabelicae Hol.-Jech., C. liquidambaris Jian Ma

& X.G. Zhang, C. uniseptata P.M. Kirk, C. inaequiseptata, and C. variabilis—

while the other Corynesporopsis species produce conidia with more septa.

All species were found as saprobes on bark, decaying leaves, rotten wood,

or dead palm rachides in terrestrial environments, except for C. variabilis on

submerged dead wood. Fourteen species definitely produce smooth-walled

conidia; the conidial ornamentation of the other three species (C. uniseptata,

C. curvularioides and C. excoecariae J.W. Xia & X.G. Zhang) is inexplicably

omitted in their original descriptions (Kirk 1981b, Xia & al. 2013, 2014).

Keys to species of Corynesporopsis have been provided by Castafieda-

Ruiz & al. (2011) and Hernandez-Restrepo & al. (2014), based on conidial

morphology. The following expanded key includes all 17 accepted

Corynesporopsis species.

628 ... Xu &al.

Key to Corynesporopsis species

lConidiazpredomirianthyslvetiseptat]e mo icnst etree Bl Me inte ge Pee ls kos 2

t.. Conidia: predominantly 2 or more septas a... nx esses eu ekg Adages aed wits 9

2. Conidia versicoloraus 1... benaeahoaceles dee hee bodes ee Se ee oe 3

ae Conidia cOnCOlOKOUS *. dasisrect othe eristhy tds oryp raat agrees: Setincapeds So aghast 4

3. Conidia obclavate, 17-25 x 4-5.5 um ....... eee eee eee C. inaequiseptata

3. Conidia variable ellipsoid, obclavate, sub-oblong, cylindrical,

sub-pyriform to-ovakl 1-16 x 5-8-2 ets 2 sages Ae C. variabilis

4, Conidia (14—)17-24(-27) x 8-12(-14) um ..... eee eee C. indica

4, COnidia-Mot.exceecin et: Ziaafltiy CIA. 04,5 ween Ra thane Game cae ee Re ees 5

5. Conidia (24-)27-43.5 x 4-6.4 um,

elongate fusiform or elongate naviculiform ....................-. C. isabelicae

5; Gonidia net exceedine 20%s:1itin lone: 8:2 534/22 Renee a Ss ee Age Soe Feasts St 6

6; Gonidia stricthreyiindnicaltorcellipsoidal *: .e Sacg td non pate ae Ad «oes 7

6. Conidia ellipsoidal to cylindrical, or naviculiform to ellipsoid .................. 8

7: Gonidia.cylindrical, 12-5520. 56 —Z, Sat eects ag cess ata PAS Coane C. cylindrica

7. Gonidiaellipsoidal, 12-16 537M «bso ot ox eels fee eae C. uniseptata

8. Conidia ellipsoidal to cylindrical, (10-)11-13 x (4.5-)5(-5.5) um ...... C. acaciae

8. Conidia naviculiform to ellipsoid, 10-16.5 x 3.5-5.5 um ......... C. liquidambaris

OF OMIA VEESICOlOTOUS Ss. bet oc. 2 percus dete cee P es oe oa scene eee SC Pu, ye 2 es. AR 10

9 Conidia concolorous. .4)..2 je eres eget epee beg ee ee ee 11

10. Conidia ellipsoidal, 21-28(-33) x 5-8 um, (3-)5(-6)-euseptate ...... C. antillana

10. Conidia broadly ellipsoid to cylindrical,

(12-)14-18 x 6-8(-9) um, 2-euseptate ............... 0... eee C. quercicola

WiGonidiacylindri¢al™ 2.52.6. hae tse ee tae twee He ee ee crows es 12

11. Conidia fusiform, ellipsoidal, obclavate, or obclavate to ellipsoidal ............ 14

12. Conidia 15-48(-59) x 3-4 um, (2-)3-7-euseptate ................00.. C. iberica

2 Conidia-aVleastfpmiewide, 2.2 Filet ce feta en hse ide, Wet eh Pane ence ik eee 13

13. Conidia 18-33 x 7-9 um, mostly 2-euseptate ...................0.. C. biseptata

13. Conidia 15-40 x 7.5-10.5 um, 1-3-euseptate .................... C. excoecariae

14:-Conidia strictly obclavate: «+3 tesco nay thtmadia ds apie angccedempa eae ghneme 4 15

14. Conidia fusiform, ellipsoidal, or obclavate to ellipsoidal ..................... 16

15. Conidia 17-48 x 6.5-8 um, (3-)5-6 (-10)-euseptate .............. C. hainanensis

15. Conidia 27-61 x 6-12 um, 5-9-euseptate .......... eee ee eee C. obclavata

16. Conidia obclavate to ellipsoidal,

1539: 7511 um, J SFetseptate: 8.6 era ee Fa thane vale ones C. curvularioides

16. Conidia fusiform or ellipsoidal,

24-36 x 8-11 (mostly 9-10) um, 3-4(-5)-euseptate .............. C. rionensis

Corynesporopsis hainanensis sp. nov. (China) ... 629

Acknowledgments

The authors express gratitude to Dr. De-Wei Li (The Connecticut Agricultural

Experiment Station Valley Laboratory, USA) and Dr. Silvana Santos Da Silva

(Departamento de Ciéncias Bioldgicas, Laboratério de Micologia, Universidade

Estadual de Feira de Santana, Brazil) for serving as pre-submission reviewers and

to Dr. Lorelei L. Norvell for final editorial review and Dr. Shaun Pennycook for

nomenclatural review. This project was supported by the National Natural Science

Foundation of China (Nos. 31970018, 31760513, 31360011) and the Education

Department of Jiangxi Province of China (No. GJJ160357).

Literature cited

Castafieda-Ruiz RE, Kendrick B. 1990. Conidial fungi from Cuba: II. University of Waterloo

Biology Series 33. 61 p.

Castafieda-Ruiz RF, Silvera-Sim6n C, Gené J, Guarro J, Minter DM, Stadler M, Saikawa M.

2011 [“2010”] A new species of Corynesporopsis from Portugal. Mycotaxon 114: 407-415.

https://doi.org/10.5248/114.407

Castaneda-Ruiz RF, Heredia G, Gusmao LFP, Li DW. 2016. Fungal diversity of Central and

South America. 197-217, in: DW Li (ed.). Biology of Microfungi. Springer International

Publishing. https://doi.org/10.1007/978-3-319-29137-6_9

Ellis MB. 1972. Dematiaceous hyphomycetes. XI. Mycological Papers 131. 25 p.

Giissow HT. 1905 [“1904”]. Notes on a disease of cucumbers, II. Journal of the Royal

Agricultural Society of England 65: 271-272.

Gams W, Seifert KA, Morgan-Jones G. 2009. New and validated hyphomycete

taxa to resolve nomenclatural and taxonomic issues. Mycotaxon 110: 89-108.

https://doi.org/10.5248/110.89

Hernandez-Restrepo M, Gené J, Castafieda-Ruiz RF, Kirk PM, Guarro J. 2014. A new species

of Corynesporopsis from Spain. Mycotaxon 127: 155-160. https://doi.org/10.5248/127.15

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MYCOTAXON

July-September 2020—Volume 135, pp. 631-635

https://doi.org/10.5248/135.631

Hymenochaete longisterigmata sp. nov. from India

NAVPREET KAUR, AVNEET PAL SINGH*, GURPAUL SINGH DHINGRA

Department of Botany, Punjabi University, Patiala-147002, Punjab, India.

*CORRESPONDENCE TO: avneetbot@gmail.com

ABSTRACT—A new hymenochaetoid species, Hymenochaete longisterigmata, is described

from Himachal Pradesh, India.

Keyworps—Basidiomycota, Hymenochaetales, Hymenochaetaceae, Himalaya

Introduction

During the 2014 fungal forays conducted in the Narkanda area of district

Shimla, Himachal Pradesh, India, Navpreet & Dhingra collected a fungal

specimen on the branch of a living tree of Picea smithiana. This fungus was

again collected (2016) in the Jalori Pass area of district Kullu, Himachal Pradesh,

India, by Navpreet & Avneet on the branch of a living Quercus semecarpifolia.

Based on the comparison of morphological characters (Cunningham 1963,

Sharma 1995 & 2012, Natarajan & Kolandavelu 1998, Bernicchia & Gorjon

2010, Dai 2010, Kaur & al. 2015, Sharma & Mishra, 2015, Mycobank 2019), we

referred the specimens to Hymenochaete Lév. (Hymenochaetaceae) and close

to H. corrugata (Fr.) Lév. based on its resupinate, adnate basidiomata, heavily

cracked hymenial surface, and presence of abundant setae with encrusted

apices. We now propose the fungus as a new species H. longisterigmata, named

in reference to its exceptionally long sterigmata.

Materials & methods

The present study is based on fungal samples collected during the field trips

conducted in Narkanda (District Shimla) and Jalori Pass (District Kullu) forest areas of

Himachal Pradesh during September 2014 and 2016. Colours terms follow Kornerup

632 ... Kaur, Singh, Dhingra

& Wanscher (1978). After removing any extraneous matter, we dried the specimens

in the sun or with a portable electric drier. The dried specimens were deposited at the

internationally recognized herbarium of Department of Botany, Punjabi University,

Patiala, India (PUN). The arrangement, size and shape of hyphae, sterile structures,

basidia, and basidiospores were studied by making crush mounts and free hand

sections in water, KOH (3%/5%/10%), 1% Congo red, 1% phloxine, cotton blue (15

in Lactophenol), and Melzer’s reagent. We examined the material microscopically

at 100x, 400x and 1000x magnifications using a compound microscope and drew

structural line diagrams on white paper sheets with the aid of a camera lucida.

Photomicrographs of important structures were taken using a Nikon 80i Eclipse

microscope.

Taxonomy

Hymenochaete longisterigmata Nav. Kaur, Avn.P. Singh &

Dhingra, sp. nov. PLATE 1

MB 832678

Differs from Hymenochaete corrugata by its thick-walled hyphidia, its larger basidia

with long sterigmata, and its broadly ellipsoid basidiospores.

Type: India, Himachal Pradesh, Shimla, Narkanda, about 3 Km from Narkanda towards

Baghi, on branch of living tree of Picea smithiana (Wall.) Boiss., 4 September 2014,

Navpreet & Dhingra 9689 (Holotype, PUN).

Etrymo.oey: The specific epithet refers to the long sterigmata.

Basidiomata annual, resupinate, <2 mm thick in section, adnate, soft and

corky when collected, hard and corky after drying; hymenial surface smooth

to somewhat aculeate under lens, cracked, greyish orange to greyish brown

when collected, no significant change after drying; margins thinning,

indeterminate, paler concolorous, sterile portion <0.5 mm. Hyphal system

monomitic. Subicular hyphae brown, heavily encrusted, up to 6.3 um wide;

subhymenial hyphae subhyaline to yellowish brown, up to 3.4 um diam.

Setae subulate, apices straight with crystalline encrustation, dark brown,

abundant, thick-walled, 68-92 x 11.4-20 um; arising from the subhymenium,

extending through hymenium, projecting <52 um above the hymenium.

Hyphidia subfusiform with moniliform apices, yellowish brown, abundant,

thick-walled, 20-34 x 4.2-5.2 um; arising from the top of subhymenium and

base of hymenium. Basidia narrowly clavate to subcylindrical, subhyaline,

thin-walled, tetrasterigmate, 22-30 x 2.8-4.6 um; sterigmata <17.2 um long.

PLATE 1. Hymenochaete longisterigmata (holotype, PUN - Navpreet & Dhingra 9689).

1. Basidiocarp showing hymenial surface; 2. Basidiospores; 3. Vertical section through

basidiocarp showing: a. basidia; b. hyphidia; c. setae; d. subhymenial hyphae; 4. Subicular hyphae;

5. Basidium; 6. Setae.

Hymenochaete longisterigmata sp. nov. (India) ... 633

el [p> ow

ty NX Ey. xX) [=

N/Zty

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634 ... Kaur, Singh, Dhingra

Basidiospores broadly ellipsoid, subhyaline, smooth, thin-walled, 4-5.1 x 2.8-

3.4 um, inamyloid, acyanophilous.

ADDITIONAL SPECIMEN EXAMINED: INDIA, HIMACHAL PRADESH, Kullu, about 3 km

from Jalori Pass towards Shoja, on branch of living tree of Quercus semecarpifolia Sm., 3

September 2016, Navpreet & Avneet 9688 (PUN).

Discussion

Hymenochaete (Hymenochaetaceae) is a widespread genus with c. 350

published species names and c. 150 currently accepted species (Kirk & al.

2008, Index Fungorum 2019, MycoBank 2019). The earliest record of this

genus from India was by Montagne (1842), who described Stereum rheicolor

[= H. rheicolor| from Gudalur, Nilgiri Hills, Tamil Nadu. Until our report,

40 Hymenochaete taxa were reported from India.

Wagner & Fischer (2002) inferred phylogenetic relationships from DNA

sequence data for several Hymenochaete species. Their phylogeny supporting

traditional Hymenochaete species in two genera, they described the new genus

Pseudochaete T. Wagner & M. Fisch. The two genera are not easily distinguished

by morphology alone. The new species described here has not been sequenced

and is therefore referred to Hymenochaete s.1.

Acknowledgments

Thanks are due to Head, Department of Botany, Punjabi University, Patiala,

for providing research facilities and University Grants Commission, New Delhi,

for financial support under UGC DRS-SAP, DSA-I programme. We also thank

Dr. Nils Hallenberg (Professor Emeritus, University of Gothenberg) for expert

comments and peer review and Prof. B.M. Sharma (Department of Plant Pathology,

COA, CSKHPAU, Palampur, India) for peer review.

Literature cited

Bernicchia A, Gorjoén SP. 2010. Corticiaceae s.l. Fungi Europaei 12. Edizioni Candusso.

Alassio, Italia. 1008 p.

Cunningham GH. 1963. The Thelephoraceae of Australia & New Zealand. Bulletin of the New

Zealand Department of Scientific and Industrial Research 145. 359 p.

Dai YC. 2010. Hymenochaetaceae (Basidiomycota) in China. Fungal diversity 45: 131-343.

https://doi.org/10.1007/s13225-010-0066-9

Index Fungorum. 2019. Search Index Fungorum. Accessed September, 2019:

http://www.indexfungorum.org/names/Names.asp

Kaur N, Sharma J, Singh AP, Dhingra GS. 2015. Additions to genus Hymenochaete Lév. from

Himachal Pradesh. International Journal of Advanced Research 3(5): 836-843.

Kirk PM, Cannon PEF, Minter DW, Stalpers JA. 2008. Ainsworth & Bisby’s

dictionary of the fungi, 10" ed. CAB International, Wallingford, UK. 770 p.

https://doi.org/10.1079/978085 1998268.0000

Hymenochaete longisterigmata sp. nov. (India) ... 635

Kornerup A,Wanscher JH. 1978. Methuen’s handbook of colour, 3rd ed. Methuen & Co. Ltd.

London. 252p.

Montagne JPFC. 1842. Cryptogamae Nilgherienses. Annales des Sciences Naturelles, Botanique

sér., 18: 12-23.

Mycobank. 2019. Hymenochaete. Website accessed 11/03/2019:

http://www.mycobank.org/Biolomics.aspx? Table=Mycobank&Rec=56728&Fields=All

Natarajan K, Kolandavelu K. 1998. Resupinate Aphyllophorales of Tamil Nadu, India. Centre for

Advanced Study in Botany, University of Madras, Chennai. 133 p.

Sharma JR, Mishra D. 2015. A synoptic mycoflora of wood-rotting fungi of Andaman. Nelumbo.

57: 1-30.

Sharma JR. 1995. Hymenochaetaceae of India. Botanical Survey of India, Ministry of

Environment & Forests, Calcutta. 219 p.

Sharma JR. 2012. Aphyllophorales of Himalaya (Auriscalpiaceae - Tremellodendropsis).

Botanical Survey of India, Ministry of Environment & Forests, Kolkata. 590 p.

https://doi.org/10.20324/nelumbo/v57/2015/87118

Wagner T, Fischer M. 2002. Classification and phylogenetic relationships of Hymenochaete

and allied genera of the Hymenochaetales, inferred from rDNA sequence data and

nuclear behavior of the vegetative mycelium. Mycological Progress 1: 93-104.

https://doi.org/10.1007/s11557-006-0008-9

MYCOTAXON

July-September 2020—Volume 135, pp. 637-647

https://doi.org/10.5248/135.637

Entoloma conferendum, Hygrocybe coccineocrenata,

and Hypholoma ericaeum new to Montenegro

ILINKA CETKOVIG ', ZDENKO TKALGEC 2’, SNEZANA DRAGICEVIG *,

ANTUN ALEGRO 3, VEDRAN SEGOTA 3, MARGITA JADAN 2, NEVEN MATOCGEC ?,

IVANA KUSAN 2, ZELJKO ZGRABLIG 4, ARMIN MESIG 2

' Natural History Museum of Montenegro,

Trg Vojvode Becir bega Osmanagica 16, 81000 Podgorica, Montenegro

? Ruder Boskovicé Institute, Bijenicka cesta 54, 10000 Zagreb, Croatia

> Department of Botany, Faculty of Science, University of Zagreb,

Marulicev trg 20/II, 10000 Zagreb, Croatia

* Croatian Research Forest Institute, Research Centre for Forest Ecosystem Goods &

Services Josip Ressel, 154. brigade Hrvatske vojske 2, 52000 Pazin, Croatia

* CORRESPONDENCE TO: Ztkalcec@irb.hr

ABSTRACT—As a result of the first research on macrofungi on peat moss habitats

in Montenegro, three species new to the Montenegrin mycobiota are presented and

described: Entoloma conferendum, Hygrocybe coccineocrenata, and Hypholoma ericaeum.

The descriptions are accompanied by photographs of the basidiomata on site and some

microscopic characters. Additionally, the second Montenegrin record of Bovista paludosa

is presented and described, for the first time with locality and habitat data. Species

identifications were checked by comparing ITS rDNA sequences from samples with the

sequences in GenBank.

Key worps—Agaricales, Basidiomycota, biodiversity, taxonomy

Introduction

Sphagnum (peat moss) is a genus with several hundred species that live in

wet habitats and can store water in their cells. The most important habitats

are peat bogs, where due to constant water availability and oxygen reduction,

peat sediments form by the accumulation of the partially decomposed plant

remnants. Peat bogs are characterised by acidic soils, low nutrient content,

638 ... Cetkovié & al.

and dominance of peat mosses. These habitats, which form in humid and

cold climates, are widely distributed in northern parts of Asia, Europe, and

North America. However, despite a still favourable climate, natural peatlands

are vanishing from northern Europe due to human activities such as peat

extraction, drainage, and land development. In the western Balkans, primarily

due to unfavourable climate conditions, peat bogs are exceptionally rare and

relict habitats, occupying only small highly isolated areas. They are particularly

sensitive to climate changes, which can cause them to dry out and become

overgrown by other vegetation types. Therefore, peat bogs are one of the

most endangered ecosystems in Montenegro. Some macrofungal species live

exclusively in this type of habitat, but a certain number of fungi with a wider

habitat tolerance can also be found in peat bogs. Peat mosses can also live in

wet, acidic habitats without forming peat, mostly in coniferous forests.

Due to the small number of researchers, Montenegrin fungi, including

agarics, are rather poorly explored. Kasom (2013) lists 839 macro-basidiomycete

species for Montenegro, 444 of which belong to the Agaricales. Bubak (1915),

who conducted the first mycological research on Montenegrin peat bogs

in 1904, reported several microfungi from the Barno jezero peat bog (Mt.

Durmitor). However, until our research, no macrofungi had been reported in

peat bogs of Montenegro. During the summer and autumn of 2017 and 2018

we surveyed macrofungi of mountain peat bogs and peat moss carpets on Mt.

Sinjajevina, Mt. Durmitor, and Mt. Hajla. We describe three Agaricales species

new to Montenegrin mycobiota, as well as one previously reported rare species

with the first data on its locality and habitat in Montenegro.

Materials & methods

Collected basidiomata were photographed on the site, described, preserved by

drying and deposited in the mycological collections of Natural History Museum of

Montenegro, Podgorica (NHMP) and/or Croatian National Fungarium in Zagreb

(CNE).

The morphological descriptions are entirely based on the collected material.

Microscopic features are described from dried material mounted in 2.5% potassium

hydroxide (KOH) and observed with a light microscope under magnification up to

1500x. Basidiospore measurements were made by Motic Images Plus 2.0 software on

30 randomly chosen mature basidiospores from calibrated digital images. Arithmetic

means of basidiospore length and width are shown in italic font in the centre of the

measurements. The length/width ratio of all measured basidiospores is given as the

“Q” value (min. — av. — max.).

Genomic DNA was extracted from dried specimens using DNeasy Plant Mini

kit (Qiagen) according to the manufacturer’s instructions. PCR primers ITS1F and

Three agarics newly recorded for Montenegro ... 639

ITS4 (Gardes & Bruns 1993) were used for ITS region amplification and sequencing

purposes. The ITS rDNA regions were amplified in a thermocycler (Eppendorf) with

a final volume of 25 uL containing approximately 50 ng DNA template, 0.2 uM of

each primer, 0.2 mM dNTP, 1.5mM MgCl, 1x buffer, and 1 unit of Taq polymerase

(Invitrogen). The cycling protocol was: 94 °C for 5 min, 30 cycles (94 °C for 45,

60 °C for 45 s, 72 °C for 45 s), followed by a final extension of 72 °C for 10 min. The

PCR products were resolved on a 1% agarose gel stained with ethidium bromide

and viewed under ultraviolet light. PCR products were purified and sequenced by

Macrogen Inc. (The Netherlands). Sequences were aligned in both directions and

edited using Sequencher v.4.1.4 and submitted to GenBank.

Taxonomy

Hygrocybe coccineocrenata (P.D. Orton) M.M. Moser,

Rohrlinge & Blatterpilze, 3. Aufl.: 68 (1967). Fic. 1 A,B

PILEUS 6-23 mm broad, convex at first, later applanate to concave with a

depressed centre and mostly inflexed and somewhat crenate margin, scarlet

red, orange red or reddish orange, densely covered with small squamules or

tufts which are reddish brown to grey towards the centre and concolorous

towards the margin, not translucently striate, dry. LAMELLAE decurrent,

distant, broad, white when young, wax yellow at maturity. STIPE 25-70 x

2-5 mm, cylindrical or tapering downwards, scarlet red to orange red,

orange near the base, whitish in the base, glabrous, hollow, dry. CONTEXT

thin, pale orange red. SMELL and TASTE indistinct. SPORE-PRINT white.

BASIDIOSPORES 9.2-11-12.1 x 5.3-6.1-6.8 um, Q = 1.51-1.81-2.13,

ellipsoid to oblong, often slightly phaseoliform in side view, sometimes

with slight median constriction, thin-walled, hyaline. Basip1a 4-spored,

narrowly clavate. CysTIDIA absent. PILEIPELLIS at centre and in squamules a

trichoderm composed of ascending hyphae constricted at septa, with brown

intracellular pigment. CLAMP CONNECTIONS present.

SPECIMEN EXAMINED—MONTENEGRO, KOoLaSIN MUNICIPALITY, Mt. Sinjajevina,

Semolj saddle, 42.9092°N 19.2731°E, 1580 m as.l, in peat bog, among dense

Sphagnum subsecundum Nees peat moss with Carex lasiocarpa Ehrh. sedge, 18 July

2017, leg. S. Dragicevi¢ (NHMP 368/6986; GenBank MN082022).

ECOLOGY & DISTRIBUTION—his waxcap occurs mostly in peat bogs among

or near peat moss, but also in wet meadows. Widespread (but generally rare)

in Europe, known also from North America and Asia (Tkalc¢ec & al. 2008,

Boertmann 2010). In the surrounding countries, H. coccineocrenata is recorded

in Croatia and Serbia, and it is a strictly protected species in both countries.

New to Montenegro.

640 ... Cetkovié & al.

CoMMENTS—Hygrocybe coccineocrenata is characterised by decurrent

lamellae, predominantly red pileus and stipe (at least in young basidiomata),

darker small squamules on the pileal surface, and by living in open, wet

habitats (mostly peat bogs). It has often been confused with H. turunda

(Fr.) P. Karst., which also has darker squamules on pileus, but which differs

in its orange to orange yellow pileus and stipe ground colours and growth

in rather dry grasslands and heathlands (Boertmann 2010). Our material

fits well with the modern descriptions (e.g., Boertmann 2010, Ludwig

2012) of H. coccineocrenata. The GenBank BLAST analysis shows that our

ITS sequence is identical with one unpublished sequence (FM208899, as

“H. turunda;” Query Cover 87%), and very close to another sequence from

the same authors (FM208865, Identity > 99%, Query Cover 87%; deposited as

“H. cantharellus, but quite different from the other H. cantharellus (Schwein.)

Murrill sequences in the database). We assume that both sequences were

derived from misidentified samples. Our Montenegrin sequence is the only

GenBank entry deposited as H. coccineocrenata.

Hypholoma ericaeum (Pers.) Kihner,

Bull. Trimest. Soc. Mycol. Fr. 52: 23 (1936) FIG. 1 C-G

PiLEUs 17-60 mm broad, paraboloid to hemispherical at first, later

convex to plano-convex, often subumbonate or with central papilla,

hygrophanous, not translucently striate, brown to dark reddish brown

when moist, orange brown to ochre brown on drying, often paler near

margin in young basidiomata, surface dry to greasy, often with fine,

whitish, fibrillose veil remnants near margin in young basidiomata.

LAMELLAE narrowly adnate, subventricose to ventricose, moderately

crowded, broad, pale brownish grey when young, then mottled grey

brown, finally mottled dark purplish brown, edge whitish. STIPE 35-78 x

2-7 mm, subcylindrical, often with broadened base (up to 10 mm), solid

to fistulose, pale to medium brown, whitish at apex in young basidiomata,

surface dry, pruinose at apex, fibrillose-furfuraceous below, mostly with

white basal tomentum. CONTEXT brown in the narrow zone beneath the

pileus surface and above the lamellae, pale brown in the inner part of the

pileus, whitish to dark red brown in the stipe. SMELL fungoid. TasTE slightly

bitter. SPORE-PRINT violaceous brown. BASIDIOSPORES 12.7—13.7-15 x

7.5-8.2-8.9 um, Q = 1.56-1.67-1.83, ellipsoid, moderately thick-walled to

thick-walled, smooth, with distinct and central germ-pore, yellow brown

in KOH, light red brown in H,O. Basrp1a 4-spored, narrowly clavate or

Three agarics newly recorded for Montenegro ... 641

Fic. 1. Hygrocybe coccineocrenata (NHMP 368/6986). A. basidiomata; B. basidiospores.

Hypholoma ericaeum (NHMP 614/9034). C. basidiomata; D. basidiospores; E. cheilocystidia;

FE, G. pleurocystidia. Scale bars: A = 10 mm; C = 20 mm; B, D-G = 10 um.

642 ... Cetkovié & al.

with median constriction. CHEILOCYSTIDIA (leptocystidia type) densely

packed, forming a sterile lamellar edge, 20-45 x 5-12 um, lageniform,

narrowly utriform or subcylindrical (often with somewhat broadened

apex), sometimes forked, thin-walled, hyaline. PLEUROCYSTIDIA

(chrysocystidia type) abundant, 30-56 x 11-18 um, clavate, mostly with

mucronate apex, thin- to moderately thick-walled, hyaline with yellowish

amorphous body in alkali solution. PILEIPELLIS a cutis composed of thin-

walled, hyaline, repent hyphae. CLAMP CONNECTIONS present.

SPECIMENS EXAMINED—MONTENEGRO, KOLASIN MUNICIPALITY, Mt.

Sinjajevina, Semolj saddle, 42.9074°N 19.2758°E, 1572 ma.s.L, among Sphagnum

moss and grass on the edge of the peat bog, 15 September 2018, leg. Z. Tkaléec &

al. (CNF 1/7623, NHMP 614/9034; GenBank MN082025); (CNF 1/7628).

ECOLOGY & DISTRIBUTION— The species is saprotrophic and occurs on

peaty acid soil (mostly among mosses) and in wet poor grasslands and

moist heaths (Watling & Gregory 1987, Noordeloos 2011, Vesterholt &

Rald 2012). It is widespread all over the boreal and temperate regions of

Europe and North America (Noordeloos 2011), but usually (rather) rare.

In the surrounding countries, H. ericaeum is recorded only in Croatia

(Tkaléec & MeSi¢ 2003). New to Montenegro.

CoMMENTS—Hypholoma ericaeum is characterised by long basidiospores

(>13 um) and a hygrophanous pileus that is neither translucently striate nor

viscid. Only three other European Hypholoma species have similarly long

basidiospores (H. eximium (C. Laest.) Rald, H. myosotis (Fr.) M. Lange, and

H. udum (Pers.) Quél.), but their spore lengths reach at least 17 um and

their pilei are viscid or subviscid. Our collections correspond relatively well

with recent descriptions of H. ericaeum (Watling & Gregory 1987, Ludwig

2001, Noordeloos 2011, Vesterholt & Rald 2012). However, the maximum

stipe width of 7 mm exceeds the 4.5 mm maximum reported by Watling

& Gregory (1987), the maximum cheilocystidia width of 12 um exceeds

the 7.5 um maximum reported by Noordeloos (2011), and the maximum

pleurocystidia size of 56 x 18 um exceeds the 45 x 15 um maximum reported

by Noordeloos (2011). Moreover, Watling & Gregory (1987) described

distinctly narrower cheilocystidia (30-40 x 4-5.5 um) and pleurocystidia

(30-40 x 8-10 um). Noordeloos (2011) described somewhat flattened

basidiospores, but the spores in our collections and in the descriptions of

other cited authors are not flattened. According to the BLAST analysis,

our ITS sequence does not match any sequence in GenBank database.

Three agarics newly recorded for Montenegro ... 643

The closest sequence (96.91% identity) is derived from H. dispersum Quél.

(HQ604746). The only GenBank ITS sequence annotated as H. ericaeum

(MH856009) is genetically much more distant (84% identity). It is possible

that MH856009 was sequenced from a misidentified sample, but it is also

possible that two morphologically close species are not yet recognized as

different; further research is required.

Entoloma conferendum (Britzelm.) Noordel.,

Persoonia 10: 446 (1980) FIG. 2 A,B

= Entoloma nothofagi G. Stev., Kew. Bull. 16: 234 (1962)

= Entoloma staurosporum (Bres.) E. Horak, Sydowia 28: 222 (1976)

PiLEus 10-30 mm broad, paraboloid at first, expanding to plano-conical

or plano-convex with subumbonate centre, hygrophanous, brown with

black centre when young, later greyish brown at centre and gradually fading

to light brown towards the margin when moist, pale brownish when dry,

translucently striate up to 2/3 of the radius when moist, surface dry, glabrous.

LAMELLAE free, ventricose, broad, moderately crowded, white when

young, pink at maturity. StrpE 35-85 x 2-4 mm, cylindrical with slightly to

distinctly broadened base, densely silvery striate on pale to light grey brown

background, dry. CoNTExT thin and fragile, light grey brown. SMELL and

TASTE farinaceous. SPORE-PRINT dirty pink. BAsIDIosPORES 9.3-10.8-11.8 x

8.1-9.4-10.4 um, Q = 0.95-1.15-1.31, cruciform-stellate, thin-walled, hyaline

to pinkish. Basrp1A 4-spored, narrowly clavate. CysT1p1A absent. PILEIPELLIS

a cutis with transition to a trichoderm at centre, with intracellular brown

pigment. CLAMP CONNECTIONS absent.

SPECIMEN EXAMINED—MONTENEGRO, Rozaje MuNICcIPALITY, Mt. Hajla, right

bank of the Ibar river, 42.7981°N 20.1083°E, 1170 m a.s.l, on dense carpet of peat

moss Sphagnum quinquefarium (Lindb.) Warnst., on very steep (almost vertical)

siliceous rocks, surrounded by mixed forest of spruce (Picea abies), fir (Abies alba)

and beech (Fagus sylvatica), 7 September 2017, leg. A. Alegro (NHMP 426/7230;

GenBank MN082023).

ECOLOGY & DISTRIBUTION— This species occurs in various habitats from the

lowlands to arcto-alpine zone, including grasslands, marshes, peat bogs, damp

places in deciduous and coniferous forests. It is a very common saprotroph

with a wide distribution, including most temperate zones on both hemispheres

(Noordeloos 1992). In the surrounding countries, E. conferendum is recorded

only in Croatia (Mesi¢ & Tkaléec 2003). New to Montenegro.

CoMMENTS—Entoloma conferendum is characterised by a_ brown

translucently striate pileus, cruciform-stellate basidiospores (unique among

644. ... Cetkovié & al.

European Entoloma species), and absence of cystidia and clamp connections.

Noordeloos & Gates (2012) place it in E. sect. Staurospora in E. subg.

Nolanea with species having predominantly cuboid basidiospores. Entoloma

staurosporum is currently accepted as a synonym of E. conferendum. Horak

(1976, 1980) synonymised E. nothofagi, described from New Zealand, with

E. staurosporum, but subsequently reverted to his original opinion (Horak

1971, 1973), treating E. nothofagi as a separate species (Horak 2008).

Noordeloos & Gates (2012), who later revised the type of E. nothofagi,

found only minimal morphological differences from E. conferendum, and

therefore regarded the two species as probably conspecific. This conclusion

is supported by our GenBank BLAST analysis showing the RPB2 sequence of

Slovakian collection of E. conferendum (KC710191) as almost identical with

the RPB2 sequence of E. nothofagi from New Zealand (MH190134), with

only a single base pair difference between them. There is no ITS sequence

available for E. nothofagi. Another similar species is E. brevispermum G.M.

Gates & Noordel. from Australia, which differs by shorter (<9 um long)

basidiospores. Our material fits well with the modern descriptions of the

species (Noordeloos 1992, Ludwig 2007, Noordeloos & Gates 2012), and

our ITS sequence matched well (identity > 99%) with most E. conferendum

sequences in GenBank.

Bovista paludosa Lév., Ann. Sci. Nat., Bot., Sér. 3, 5: 163 (1846) FIG. 2 Cc, D

BASIDIOMATA pyriform, 23-48 mm high and 16-24 mm broad, sometimes

with white rhizoids. ExoPERIDIUM when young white and smooth or with

small, very thin, adherent, greyish, polygonal patches, later breaking up

into irregular, rather large plates. ENDOPERIDIUM smooth, white at first,

becoming brown to olivaceous. GLEBA white when young, then yellowish

brown, becoming olive brown at maturity. SUBGLEBA well developed,

<15 mm high, compact, white when young, olive to grey brown at maturity.

SPORE-PRINT Olive brown. BASIDIOSPORES 4.6-5.1-5.6 x 4.2-4.7-5.4 um,

Q = 1-1.07-1.19, globose to subglobose (rarely broadly ellipsoid), almost

smooth to asperulate, moderately thick-walled, pale yellow brown in KOH,

with sterigmal remnant <15 um long. CAPILLITIUM Bovista-type, non-

poroid, thick-walled, rarely septate.

SPECIMEN EXAMINED—MONTENEGRO, ZABLJAK MUNICIPALITY, Mt. Durmitor,

Barno jezero lake, 43.1582° N, 19.0932° E, 1495 m a.s.l, among Aulacomnium

palustre (Hedw.) Schwagr., Bryum pseudotriquetrum (Hedw.) P. Gaertn. & al., and

Campylium stellatum (Hedw.) C.E.O. Jensen mosses on the outer zone of the peat bog

(less acidic), 9 July 2017, leg. I. Cetkovié (NHMP 425/7229; GenBank MN082024).

Three agarics newly recorded for Montenegro ... 645

Fic. 2. Entoloma conferendum (NHMP 426/7230). A. basidiomata; B. basidiospores.

Bovista paludosa (NHMP 425/7229). C. basidiomata; D. basidiospores. Scale bars: A, C = 10 mm;

B, D= 10 um.

ECOLOGY & DISTRIBUTION— This puffball lives as a saprotroph in a rather

wide range of open wet mossy habitats, most often in alkaline to neutral fens,

bogs, mires, wet meadows, and heaths, but avoiding oligotrophic and strongly

acidic mires (Kaltucka 2019). The species is rather widespread but rare in Europe

(Red Listed in most countries where reported), known also from Asia (China,

Russia, India) and North America (Canada) (Jeppson 2018). The only earlier

646 ... Cetkovié & al.

Montenegrin record of B. paludosa from 1904 (Bubak 1915) was published

without locality and habitat data. In the surrounding countries, it is recorded

only in Serbia (Katucka 2019).

COMMENTS—Bovista paludosa is characterised by its occurrence on wet mossy

habitats and by the presence of a well-developed subgleba, long sterigmal

remnants on basidiospores, and a Bovista-type capillitium lacking pores. The

most similar species, B. cretacea T.C.E. Fr., differs in a very poorly developed

(<2 mm high) subgleba and regularly septate capillitium. Our material agrees

with the modern descriptions (Pegler & al. 1995, Sarasini 2005, Jeppson 2018).

According to the BLAST analysis, our ITS sequence matched well with all three

sequences of B. paludosa in GenBank database (Identity > 99%).

Acknowledgements

We are very grateful to Dr. André Fraiture (Botanic Garden Meise, Belgium)

and Dr. Vladimir Antonin (Moravian Museum, Brno, Czech Republic) for their

critical review of the manuscript. The research received financial support from the

Montenegrin Ministry of Science and Croatian Ministry of Science and Education

through the project “Inventory of biodiversity and conservation status of the

representative peat bogs on the area of Montenegro and Croatia.” Also, this work

has been partially supported by Croatian Science Foundation under the project

ForFungiDNA (IP-2018-01-1736).

Literature cited

Boertmann D. 2010. The genus Hygrocybe, 2nd revised edition. Fungi of Northern Europe 1.

Svampetryk, Tilst.

Bubak FE. 1915. Dritter Beitrag zur Pilzflora von Montenegro. Botanikai K6zlemények 14(3-4):

39-83.

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

Horak E. 1971. A contribution towards the revision of the Agaricales (Fungi) from New Zealand.

New Zealand Journal of Botany 9: 403-462. https://doi.org/10.1080/0028825x.1971.10430193

Horak E. 1973. Fungi agaricini Novazelandiae I-V. Beihefte zur Nova Hedwigia 43. J. Cramer,

Lehre.

Horak E. 1976. On cuboid-spored species of Entoloma (Agaricales). Sydowia 28: 171-236.

Horak E. 1980. Entoloma (Agaricales) in Indomalaya and Australasia. Beihefte zur Nova

Hedwigia 65. J. Cramer, Vaduz.

Horak E. 2008. Agaricales of New Zealand 1: Pluteaceae - Entolomataceae. The fungi of New

Zealand 5. Fungal Diversity Research Series, vol. 19. Fungal Diversity Press, Hong Kong.

Jeppson M. 2018. Puffballs of northern and central Europe. SMF, Goteborg.

Katucka IL. 2019. Bovista paludosa Lév. The Global Fungal Red List. Viewed online on 22 May

2019: http://iucn.ekoo.se/iucn/species_view/209589/

Three agarics newly recorded for Montenegro ... 647

Kasom G. 2013. Basidiomycota macrofungi of Montenegro. PhD ‘Thesis, University of

Montenegro, Faculty of Science and Mathematics, Podgorica. (in Montenegrin).

https://fedora.ucg.ac.me/fedora/get/0:334/bdef:Content/get (viewed online on 22 May 2019)

Ludwig E. 2001. Pilzkompendium, Band 1. IHW-Verlag, Eching.

Ludwig E. 2007. Pilzkompendium, Band 2. Fungicon-Verlag, Berlin.

Ludwig E. 2012. Pilzkompendium, Band 3. Fungicon- Verlag, Berlin.

MeSi¢ A, Tkaléec Z. 2003. Preliminary checklist of Agaricales from Croatia IV: Families

Bolbitiaceae, Coprinaceae, Entolomataceae and Pluteaceae. Mycotaxon 87: 283-309.

Noordeloos ME. 1992. Entoloma s.l. Fungi Europaei 5. Libreria editrice Giovanna Biella,

Saronno.

Noordeloos ME. 2011. Strophariaceae s.1. Fungi Europaei 13. Edizioni Candusso, Alassio.

Noordeloos ME, Gates GM. 2012. The Entolomataceae of Tasmania. Springer, New York -

London. https://doi.org/10.1007/978-94-007-4679-4

Pegler DN, Lzessge T, Spooner BM. 1995. British puffballs, earthstars and stinkhorns. Royal

Botanic Gardens, Kew.

Sarasini M. 2005. Gasteromiceti epigei. A.M.B., Trento.

Tkaléec Z, Mesi¢ A. 2003. Preliminary checklist of Agaricales from Croatia V: Families

Crepidotaceae, Russulaceae and Strophariaceae. Mycotaxon 88, 279-314.

Tkaléec Z, MeSi¢ A, Matocéec N, KuSan I. 2008. Red book of Croatian fungi. Ministry of Culture

- State Directorate for Protection of Nature, Zagreb (in Croatian).

Vesterholt J, Rald E. 2012. Hypholoma (Fr.) P. Kumm. 942-946, in: H Knudsen, J Vesterholt

(eds). Funga Nordica, Nordsvamp, Copenhagen.

Watling R, Gregory NM. 1987. Strophariaceae & Coprinaceae p.p.: Hypholoma, Melanotus,

Psilocybe, Stropharia, Lacrymaria and Panaeolus. British Fungus Flora: Agarics and Boleti

5. Royal Botanic Garden, Edinburgh.

MYCOTAXON

July-September 2020—Volume 135, pp. 649-655

https://doi.org/10.5248/135.649

First Pakistani report of Erysiphe betae

on the invasive weed Chenopodium ambrosioides

AYESHA ANWAR™, NAJAM UL SEHAR AFSHAN’, AAMNA ISHAQ3,

MarRIA RIAz?, ABDUL NASIR KHALID’, SIRAJ UDDIN'

' Department of Botany, University of Peshawar, 25120, Pakistan

? Department of Botany, University of the Punjab,

Quaid-e-Azam Campus, Lahore, 54590, Pakistan

> Department of Botany, Faculty of Fisheries and Wildlife,

University of Veterinary and Animal Sciences-Lahore,

Ravi Campus, Pattoki, 55300, Pakistan

" CORRESPONDENCE TO: Ayeshaanwar556@gmail.com

AxssTRACT—During September-October 2017, powdery mildew symptoms were observed

on both surfaces of leaves of Chenopodium ambrosioides in Abbottabad, Malakand, and

Upper Dir districts of Pakistan. The causal agent was identified as Erysiphe betae, based on

its asexual morphology and chasmothecia, and its identity was confirmed by molecular data.

This is the first Pakistani report of Erysiphe betae on this host.

Key worps—Amaranthaceae, Erysiphaceae, mexican tea, new record

Introduction

Fungi belonging to Erysiphales are widely distributed all over the world

and cause serious economic damage on wild and cultivated plants. This order

comprises the single family Erysiphaceae (powdery mildews) represented by

17 genera and c. 820 species (Braun & Cook 2012). Powdery mildews infect

9839 angiosperm species in 1617 genera, 196 families, and 44 orders (Amano

1992, Takamatsu 2004), including numerous economically important plants.

Various powdery mildews may cause serious economic damage on wild

and cultivated plants in Pakistan (Burni & al. 2010, Mukhtar & al. 2012,

650 ... Anwar & al.

2013). Nineteen of the 56 powdery mildew species reported from Pakistan

represent the most common genus, Erysiphe, which infects 52 hosts in 19

plant families (Ahmad & al. 1997, Burni & al. 2010, Mukhtar & al. 2013). This

includes Erysiphe betae on Chenopodium botrys L. and Spinacia oleracea L.

(Amaranthaceae; Amano 1986).

In 2017, a powdery mildew appeared on the leaves of Chenopodium

ambrosioides in the districts Abbottabad, Malakand, and Upper Dir, Pakistan.

The infected leaf surfaces were fully covered with white powdery mycelial

masses, asexual conidia, and sexual chasmothecia indicating progressed

stages of infection and severity.

Materials & methods

Sample collection

During phytopathological surveys during September-October 2017, we observed

powdery mildew infections on leaves and stems of Chenopodium ambrosioides in the

districts Abbottabad, Malakand, and Upper Dir, Khyber Pakhtunkhwa, Pakistan. The

infected plants were shade dried on blotting paper and protected in brown envelopes

for future use. The specimens are deposited at the herbarium of the University of

Peshawar, Khyber Pakhtunkhwa, Pakistan (PUP) and Department of Botany,

University of the Punjab, Lahore, Punjab, Pakistan (LAH).

Morphology

We first examined infected leaves under a Labomed CSM2 stereomicroscope,

and then prepared using lactic acid. We examined the hyphae on the host; hyphal

appressoria; conidial, conidiophore, and chasmothecial shapes and sizes; and

ascospores and asci under a Nikon YS 100 microscope. Measurements (20 repetitions

per structure) were recorded using a Lomo Filar Eyepiece Micrometer AM9-2-15x on

a Zeiss microscope. Micrographs were made using a HDCE-5X digital camera.

DNA extraction, PCR amplification, phylogenetic analysis

Dried powdery mildew infections were scraped off from fresh fungal specimens

with sterile razor blades, ground in liquid nitrogen, and stored in Eppendorf tubes at

-18 °C. DNA was extracted using GeneJET Plant Genomic DNA Purification Mini

Kit #K0791 according to the manufacturer’s instructions. The Internal Transcribed

Spacer (ITS) region was amplified using PMITS1/PMITS2 primers (Cunnington

& al. 2003) and then commercially sequenced by Tsingke in China. Raw sequence

data were edited using BioEdit (Hall 1999). The ITS sequences were BLAST searched

against the GenBank database (www.ncbi.nlm.nih.gov) and aligned using Muscle

E multiple alignment tool within MEGA v. 7.0 (Kumar & al. 2016). Twenty ITS

sequences were analyzed using the maximum likelihood (ML) method based on the

Tamura 3-parameter model (Tamura 1992). The initial trees for the heuristic search

were obtained by applying the Neighbor-Joining method to a matrix of pairwise

Erysiphe betae new on Chenopodium ambrosioides in Pakistan ... 651

KY660905 Erysiphe ludens

35| KY660901 Erysiphe ludens

KY660739 Erysiphe trifoliorum

KY660903 Erysiphe trifoliorum

KJ845646 Erysiphe sp.

KJ845645 Erysiphe sp.

KY926844 Erysiphe castaneigena

59 89' KY926843 Erysiphe castaneigena

KY660931 Erysiphe cruciferarum

KY660884 Erysiphe cruciferarum

KY660750 Erysiphe pisi

LC270861 Erysiphe viciae-unijugae

79 KY660922 Erysiphe buhrii

KY660881 Erysiphe buhrii

82] | @ MN368297 Erysiphe betae

71'LC009946 Erysiphe betae

MG938639 Erysiphe berberidicola

MG938640 Erysiphe berberidicola

AB080464 Phyllactinia angulata

LC108847 Phyllactinia leveilluloides

0.05

Fic. 1. Phylogenetic analysis of the ITS region for 18 Erysiphe sequences, with Phyllactinia angulata

and P. leveilluloides as outgroup. The evolutionary history was inferred by using the Maximum

Likelihood method based on the Tamura 3-parameter model. The tree with the highest log

likelihood (-470.6292) is shown, and the amplified sequence from Pakistan is denoted by ¢.

distances estimated using the Maximum Composite Likelihood (MCL) approach.

Evolutionary analyses were conducted in MEGA6 (Tamura & al. 2013). in the

phylogenetic analyses. After elimination of gaps and missing data, the final dataset

comprised 134 positions, and Phyllactinia angulata (E.S. Salmon) S. Blumer and P

leveilluloides Moreno-Rico & U. Braun served as outgroup. All sequences were aligned

and trimmed at conserved sites from both 5’ and 3’ ends. The phylogenetic tree with

the highest log likelihood (-470.6292) is shown in Fic. 1; the percentage of trees in

which the associated taxa clustered together is shown next to the branches, and the

tree is drawn to scale, with branch lengths measured in the number of substitutions

per site.

652 ... Anwar & al.

Fic. 2. Erysiphe betae (PUP Bot.01). A. Infected plant, Chenopodium ambrosioides; B. Fungal

mycelium under stereomicroscope; C. Conidia; D. Conidiophore; E. Chasmothecium;

E Germinating Conidium; G. Asci with ascospores; H. Ascospores; I. Chasmothecial appendage.

Seale‘bars: A= 1 cm, B =2am,'C, D, BG= 15pm; EH, L = 10 pm.

Erysiphe betae new on Chenopodium ambrosioides in Pakistan ... 653

Taxonomy

Erysiphe betae (Vanha) Weltzien, Phytopathol. Z. 47: 127 (1963) BIG 72

Mycelium amphigenous, forming dense, thick, white patches; hyphae

hyaline, smooth, thin-walled; hyphal appressoria nipple-shaped to

somewhat lobed, 2-5 um diam. Conidiophores arising from the top of

mother cells, 43-87 x 3.5-7.5 um; foot cells cylindrical, straight or sometime

curved, 22-61 x 4.5-8 um, constricted at the basal septum, followed by 1-2

shorter cells; conidia formed singly, ellipsoid-ovoid to cylindrical-doliiform,

14.5-51 x 7-12 um; germ tubes arising from one end, tips slightly swollen;

Chasmothecia scattered to gregarious, globose to subglobose, light to dark

brown, 48.5-104 um diam.; peridium cells small, 12-27 um diam., irregularly

polygonal; appendages numerous, mycelioid, irregularly branched, septate,

thin-walled, at first hyaline but pigmented when chasmothecia mature,

29-119 x 2.5-4.5 um; asci 3-5, ellipsoid-obovoid, 28-49.5 x 19.5-29.5 um,

stalked, 4-5-spored; ascospores globose-ellipsoid to ovoid, 8.5-18.5 x

5.5-10.5 um, colorless.

SPECIMENS EXAMINED: PAKISTAN, KHYBER PAKHTUNKHWA, Malakand Division,

Malakand district, 844 ma.s.l., on Chenopodium ambrosioides L. (Amaranthaceae), 19

September 2017, asexual morph, A. Anwar AA#4 (PUP Bot.01; GenBank MN368297);

Upper Dir district, 844 m a.s.l., on C. ambrosioides, 29 October 2017, asexual and

sexual morphs, A. Anwar AA#5 (PUP Bot.02); Hazara Division, Abbottabad district,

Ayubia National Park, 2438 m a.s.l, on C. ambrosioides, 30 October 2017, asexual

morph, N.S. Afshan NSA#34 (LAH 35658).

Phylogenetic results

The NCBI BLASTn analysis showed that our ITS sequence (MN368297

Erysiphe betae) closely matched (99.54% identity) a sequence of E. betae

(LC009946) on Beta vulgaris from Japan (Takamatsu & al. 2015). The 20

nucleotide sequences used in the phylogenetic analysis included Phyllactinia

angulata (AB080464) and P leveilluloides (LC108847) as outgroup. The

final aligned data set contained 818 positions of which 567 were conserved,

202 variable and parsimony uninformative, and 49 parsimony informative

sites. Maximum likelihood (ML) analysis clustered the Pakistan E. betae

collection (MN368297) with the Japanese E. betae collection (LC009946)

with a 71 bootstrap value. Based on these results, we assigned the name

Erysiphe betae to the powdery mildew pathogen on Chenopodium

ambrosioides in Pakistan.

654 ... Anwar & al.

Discussion

Worldwide, three Erysiphe spp. (including E. betae), two Leveillula spp.,

and three anamorphic powdery mildew species have been reported on

Chenopodium ambrosioides (Fungus-Host Database https://nt.ars-grin.gov,

29 May, 2018, Braun & Cook 2012). From Pakistan, only “Erysiphe communis

(Wallr.) Schltdl.” (nom. rej.) on Chenopodium botrys and Spinacia oleracea

and Leveillula cylindrospora U. Braun and L. taurica (Lév.) G. Arnaud on

Chenopodium murale L. have been reported. Chenopodium ambrosioides is

reported here as a new host record for Erysiphe betae in Pakistan.

Previously, only asexual morphs of Erysiphe have been reported on

Chenopodium species from Asia and Europe, whereas sexual fruiting bodies

(chasmothecia) have been reported only from India and Japan (Braun & Cook

2012). We now report both asexual and sexual morphs of Erysiphe betae from

Pakistan. We found only the asexual morph in the Abbottabad and Malakand

districts, but additionally collected the chasmothecia in the Upper Dir

district. Previous reports from Pakistan of Erysiphe on Chenopodium botrys

(as “E. communis;” Amano 1986) is probably allocable to E. betae. Francis &

al. (2007), who analysed ITS rDNA sequences retrieved from European (UK)

and North American (USA) collections of E. betae on sugar beet, confirmed

the involvement of a single recognizable Erysiphe species more closely allied

to Erysiphe heraclei DC. than to E. polygoni DC. Takamatsu & al. (2015)

included in their phylogenetic analyses a sequence obtained from E. betae

on Beta vulgaris L. with corresponding results, but sequences retrieved from

E. betae on Chenopodium ambrosioides are not yet available and still in need

of molecular confirmation. Our research represents the first phylogenetic

analysis of E. betae on Chenopodium ambrosioides, which clusters our ITS

sequence in a strongly supported clade with a sequence of E. betae on

Beta vulgaris (Fic. 1).

Acknowledgements

We are really thankful to Prof. Dr. Uwe Braun (Martin-Luther-Universitat,

Institut fiir Biologie, Germany) for his help in the identification of Erysiphe betae

on Chenopodium ambrosioides and peer review of the manuscript. The authors are

also thankful to Dr. Abdul Rehman Khan Niazi, Department of botany, University of

Punjab, Lahore, for his valuable suggestions to improve this manuscript.

Literature cited

Ahmad §, Iqbal SH, Khalid AN. 1997. Fungi of Pakistan. Sultan Ahmad Mycological Society of

Pakistan, Lahore, PK.

Amano K. 1986. Host range and geographical distribution of the powdery mildew fungi. Japan

Scientific Societies Press, Tokyo.

Erysiphe betae new on Chenopodium ambrosioides in Pakistan ... 655

Amano K. 1992. Notes on the host range and geographical distribution of Podosphaera.

Transactions of the Mycological Society of Japan 33: 139-148.

Braun U, Cook RTA. 2012. Taxonomic manual of the Erysiphales (powdery mildew). CBS

Biodiversity Series 11. 707 p.

Burni T, Jamil F, Jabeen M. 2010. Erysiphales of Peshawar valley, Khyber Pakhtunkhwa, Pakistan.

Pakistan Journal of Plant Sciences 16(1): 11-14.

Cunnington JH, Takamatsu S, Lawrie AC, Pascoe IG. 2003. Molecular identification of

anamorphic powdery mildews (Erysiphales). Australasian Plant Pathology 32: 421-428.

https://doi.org/10.1071/AP03045

Francis SA, Roden BC, Adams MJ, Weiland J, Asher MJ. 2007. Comparison of ITS sequences

from UK and North American sugar-beet powdery mildews and the designation of Erysiphe

betae. Mycological Research 111(2): 204-212. https://doi.org/10.1016/j.mycres.2006.10.010

Hall TA. 1999. BioEdit: a user-friendly biological sequence alignment editor and analysis

program for Windows 95/98/NT. Nucleic Acids Symposium Series 41: 95-98.

https://doi.org/10.1017/S0953756203008517

Kumar S, Stecher G, Tamura K. 2016. MEGA7: Molecular Evolutionary Genetics Analysis

version 7.0 for bigger datasets. Molecular Biology and Evolution 33(7): 1870-1874

https://doi.org/10.1093/molbev/msw054.

Mukhtar I, Khurram R, Hannan A, Hayat Z. 2012. First report of powdery mildew of Cucurbita

moschata Duch. caused by Golovinomyces cichoracearum DC. in Neelum Valley, Azad

Kashmir. Plant Disease 96(6): 906. https://doi.org/10.1094/PDIS-08-11-0642-PDN

Mukhtar I, Mushtaq S, Khokhar I. 2013. First report of powdery mildew on Dahlia (Dahlia

variabilis) caused by Golovinomyces cichoracearum in Lahore, Pakistan. Australasian Plant

Disease Notes 8(1): 1-3. https://doi.org/10.1007/s13314-011-0025-7

Takamatsu S. 2004. Phylogeny and evolution of the powdery mildew fungi (Erysiphales,

Ascomycota) inferred from nuclear ribosomal DNA sequences. Mycoscience 45: 147-157.

https://doi.org/10.1007/s10267-003-0159-3

Takamatsu S, Ito H, Shiroya Y, Kiss L, Heluta V. 2015. First comprehensive phylogenetic

analysis of the genus Erysiphe (Erysiphales, Erysiphaceae) 1. The Microsphaera lineage.

Mycologia 107: 475-489. https://doi.org/10.3852/15-007

Tamura K. 1992. Estimation of the number of nucleotide substitutions when there are strong

transition-transversion and G + C-content biases. Molecular Biology and Evolution 9:

678-687. https://doi.org/10.1093/oxfordjournals.molbev.a040752

Tamura K, Stecher G, Peterson D, Filipski A, Kumar S. 2013. MEGA6: Molecular Evolutionary

Genetics Analysis version 6.0. Molecular Biology and Evolution 30: 2725-2729.

https://doi.org/10.1093/molbev/mst197

MYCOTAXON

July-September 2020—Volume 135, pp. 657-663

https://doi.org/10.5248/135.657

Crustose lichens new to India

RUPJYOTI GOGOT’, SILJO JOSEPH?, MANAS PRATIM CHOUDHURY’,

SANJEEVA NAYAKA’, FARISHTA YASMIN’*

' Department of Botany, Nowgong College,

Nagaon, Assam - 782001, India

? Lichenology Laboratory, CSIR-National Botanical Research Institute,

Rana Pratap Marg, Lucknow - 226001, India

* CORRESPONDENCE TO: farishtayasmin4rs@gmail.com

ABSTRACT—Bacidia pycnidiata (Ramalinaceae), Malmidea nigromarginata (Malmideaceae),

Porina malmei and P. nuculastrum (Porinaceae), and Pyrenula laetior and P. wrightii

(Pyrenulaceae) are reported for the first time from India. The specimens were collected

from the state of Assam. Taxonomic descriptions, distributions, and illustrations are

provided for each species.

Key worps—biodiversity, lichenized Ascomycota, Nagaon, taxonomy, Tezpur

Introduction

Lichens constitute a significant part of India’s biodiversity, and 2714

lichen species are known from the country (Sinha & al. 2018). Although

lichens occur everywhere in India, ecological studies reveal that Northeast

India, Western Ghats, Western Himalaya, and Andaman & Nicobar Islands

are the major centres of diversity (Nayaka & Asthana 2014). Assam, a state of

Northeast India, possesses the heterogeneous climatic conditions that make

it suitable for rich floral, faunal, and fungal diversity. Over 325 species (in

83 genera and 27 families) have been reported from Assam (Gupta & Sinha

2018, Gogoi & al. 2019). Here we record six additional lichen species (in

Malmideaceae, Porinaceae, Pyrenulaceae, and Ramalinaceae) new to Assam

and to India.

658 ... Gogoi & al.

Materials & methods

The present study is based on the freshly collected lichen samples from Assam.

The lichen specimens were dried, preserved, and deposited in the herbarium at CSIR-

National Botanical Research Institute, Lucknow, India (LWG). The lichen thallus

was morphologically examined under a Leica EZ4 stereo zoom microscope. Thin

hand-cut sections of ascus and thallus were mounted in distilled water, Lactophenol

Cotton Blue (LCB), 5% KOH, or Lugol’s iodine solution for observation under

a Leica DM 2500 compound microscope. The thallus and ascomatal tissue were

spot-tested with K (5% KOH aqueous solution), C (aqueous Ca(ClO)2 solution)

and P (0.5 g of para-phenylenediamine dissolved in 5 ml of ethanol) following

Orange & al. (2001). Thin Layer Chromatography was performed in solvent system

C (toluene: acetic acid; 85: 15 ml) following Orange & al. (2001).

Taxonomy

Bacidia pycnidiata Czarnota & Coppins, Lichenologist 38: 407. 2006 Pie ika

Thallus epilithic, pale yellowish to greenish, slightly granular, cracked.

Apothecia few, 0.2-0.4 mm diam. Excipulum prominent, colourless, 90-140

um wide. Hymenium hyaline, 40-55 um high, I+ blue. Asci cylindrical,

Bacidia-type, 35-41 x 4.5-5.8 um. Ascospores acicular, 3(-—7)-septate, 25-38 x

1.2-2.8 um. Pycnidia very few, having round flask shaped bottom with long

neck, 0.1-0.2 mm diam., constricted at base, concolorous with apothecia,

neck slightly paler than bottom, ostiole present at the tip of the beak; conidia

hyaline, 3-5-septate, straight to slightly curved, filiform, 37-62 x 0.4-1.0 um.

CHEMISTRY— Thallus K-, C-, P-, UV-; no chemicals detected in TLC.

SPECIMEN EXAMINED—INDIA, Assam, Tezpur, Ouguri Hills, 26°37’03”N 92°46'37”E,

elev. 113 m, on rock, 16 May 2015, Manas P. Choudhury 15027001 (LWG 35885).

REMARKS— The ascospore length of the Indian specimen is slightly shorter

than that given in the protologue of Bacidia pycnidiata (35-52 um; Czarnota

& Coppins 2006). Further, the Indian specimen had occasional 7-septate

ascospores rather than the usual 3-septate, which has not been reported

previously for B. pycnidiata. In all other aspects the Indian specimen agrees

with the published description of B. pycnidiata.

DISsTRIBUTION—Belgium, Czech Republic, Estonia, Finland, Lithuania,

Mordovia, North Caucasus, Poland, and Ukraine (Urbanavichene &

Urbanavichus 2014). This is a first report of B. pycnidiata for India.

Malmidea nigromarginata (Malme) Licking & Breuss,

Lichenologist 47: 19. 2015. Pr lb

Thallus crustose, epiphloeodal, smooth, grayish black, medulla

ochraceous, K+ yellow; borderline blackish. Apothecia mostly solitary,

Bacidia, Malmidea, Porina & Pyrenula spp. new for India ... 659

PLATE 1. Habit. a. Bacidia pycnidiata (LWG 35885); b. Malmidea nigromarginata (LWG 35887);

c. Porina malmei (LWG 35886); d. Porina nuculastrum (LWG 35888); e. Pyrenula laetior (LWG

35889); f. Pyrenula wrightii (LWG 35890). Scale bars = 1 mm.

660 ... Gogoi & al.

rarely 3—4 aggregated, margin black, disc flesh coloured. Excipulum without

algal cells, lacking medullary layer, hyphae radiating and conglutinated.

Hymenium hyaline, 70-90 um high, I+ blue. Hypothecium dark brownish to

brown black, 70-100 um thick. Asci 6-8-spored. Ascospores simple, hyaline,

8.9-13.4 x 4.5-5.4 um.

CHEMISTRY—Thallus K—, C—, P-, UV-; no lichen substances detected

im ELE.

SPECIMEN EXAMINED—INDIA, AssaM, Nagaon district, Samaguri, Chapanalla,

26°19'13”N 92°54’16’E, elev. 119 m, on bark, 18 February 2018, Rupjyoti Gogoi

NCLH 030 (LWG 35887).

REMARKS—The ascospore sizes of the Indian specimen fit well within the

range of Malmidea nigromarginata (10-14 x 4-6 um) as mentioned in the

world key to the genus by Breuss & Licking (2015).

DIsTRIBUTION—Nicaragua (Breuss & Liicking 2015) and Puerto Rico

(Mercado-Diaz & al. 2015). This is a first report of M. nigromarginata for India.

Porina malmei P.M. McCarthy, Biblioth. Lichenol. 52: 70. 1993. Baeale

Thallus saxicolous, medium to dark grey, areolate, ecorticate. Perithecia

emergent, hemispherical to subglobose, dull black, 0.1-0.3 mm diam.

Ostioles apical, conspicuous. Involucrellum brownish, continuous with the

exciple. Excipulum black, 20-50 um thick. Centrum 0.15-0.25 um wide.

Hymenium clear. Asci elongate-cylindrical. Ascospores hyaline, fusiform,

(3-)6-7-septate, 20-27 x 4-6 um. Pycnidia not seen.

CHEMISTRY— Thallus K-, C-, P-, UV-; no chemicals detected in TLC.

SPECIMEN EXAMINED—INDIA, Assam, Tezpur, Rudrapad Temple, bank of River

Brahmaputra, 26°36’55”N 92°46’10”E, elev. 65 m, on rock, 16 March 2015, Manas P.

Choudhury 15027003 (LWG 35886).

REMARKS— The Indian specimen agrees with the description of Australian

Porina malmei in most features, and ascospore lengths fit well within the

17-30 um range cited in the protologue (McCarthy 1993).

DisTRIBUTION—Brazil, north-eastern Australia (McCarthy 2000). This is

a first report of P malmei for India.

Porina nuculastrum (Mill. Arg.) R.C. Harris, More Florida Lichens,

incl. 10 Cent Tour Pyrenol. (New York): 174. 1995. Bred

Thallus crustose, epiphloeodal, olivaceous, slightly rimose, prothallus black.

Perithecia erumpent to + immersed, mostly solitary, rarely grouped into 2-3,

perithecia concolorous with the thallus except in the peri-ostiolar and ostiolar

regions, convex to sub-globose 0.4-0.65 mm diam.; ostiole inconspicuous or

Bacidia, Malmidea, Porina & Pyrenula spp. new for India ... 661

in a shallow depression; peri-ostiolar area 0.12-0.18 mm diam. Involucrellum

enclosing the exciple yellowish to blackish. Exciple ca. 30 um thick. Hymenium

clear. Asci elongate-cylindrical. Ascospores narrowly ellipsoidal to fusiform,

submuriform to muriform with 6-10 transverse septa and (0-)1(-3)

longitudinal septa, 49.8-67.6 x 13.5-22.2 um. Pycnidia not seen.

CuHEMISTRY— Thallus K-, C—, P-, UV-; no chemicals detected in TLC.

SPECIMEN EXAMINED—INDIA, Assam, Nagaon district, Samaguri, Suang Reserve

Forest, 26°19’20”N 92°54’18’E, elev. 142 m, on bark, 18 February 2018, Rupjyoti

Gogoi NCLH 031 (LWG 35888).

ReMARKS—'The Indian specimen closely matches the Australian and

Vietnamese descriptions of Porina nuculastrum (McCarthy 2001, Joshi & al.

2019), except that our ascospores have fewer transverse septa.

DIsTRIBUTION—Neotropics, Madagascar, the Philippines, Hong Kong

and neighbouring countries (McCarthy 2001), Vietnam (Joshi & al. 2019).

This is a first report of P nuculastrum for India.

Pyrenula laetior Mill. Arg., Bot. Jahrb. Syst. 6: 413. 1885. PL. le

Thallus crustose, corticate, epiphloeodal, yellow, without external

pigments. Perithecia black, solitary, 0.4-0.9 mm diam. Ostiole apical

without pigment. Hamathecium inspersed with oil droplets. Asci 8-spored.

Ascospores brown, 3-septate, eciliate, distoseptate, lumina in a straight line,

13.8-19.2 x 5.8-7.7 um, central lumina not strongly elongated, terminal

lumina separated from the exospore wall by endospore thickenings, old

ascospores without orange oil.

CHEMISTRY— Thallus K—, C—, P-, UV-; no chemicals detected in TLC.

SPECIMEN EXAMINED—INDIA, Assam, Nagaon district, Samaguri, Suang Reserve

Forest, 26°19’20”N 92°54’18’E, elev. 143 m, on bark, 18 February 2018, Rupjyoti

Gogoi NCLH 032 (LWG 35889).

REMARKS—Aptroot (2012) cites ascospores ranging from 13-18 um for

Pyrenula laetior, while those of the Indian specimen are slightly longer. The

Indian material agrees with P Jaetior in all other characters.

DISTRIBUTION—Neotropics (Aptroot 2012), Vietnam (Joshi & al. 2018).

This represents a first report of P laetior for India.

Pyrenula wrightii (Mill. Arg.) R.C. Harris,

More Florida Lichens, incl. 10 Cent Tour Pyrenol.: 111. 1995. Prt

Thallus corticolous, epiphloeodal, olive green with pseudocyphellae.

Perithecia black, without external pigments, solitary to mostly aggregated

with separated ostioles, 0.2-0.5 mm diam. Ostiole apical without pigment.

662 ... Gogoi & al.

Hamathecium not inspersed with oil droplets. Asci 8-spored. Ascospores

brown, 2-3-septate, 27.3-33.2 x 12.5-14.5 um, lumina in a straight

line, external lumina separated from the exospore wall by endospore

thickening.

CHEMISTRY— Thallus K—, C—, P-, UV-; no lichen substance detected

bia es bs oe

SPECIMEN EXAMINED—INDIA, Assam, Nagaon district, Doboka Circle, Jamunamukh,

26°05'54’"N 92°44’44”E, elev. 40 m, on bark, 5 February 2017, Rupjyoti Gogoi NCLH

033 (LWG 35890).

REMARKS— The Indian specimen agrees with the published description of

Pyrenula wrightii by having pseudocyphellae and in ascospore characteristics

(Aptroot 2012). The ascospores of the Indian specimen are slightly longer

than the measurements provided in the basionym protologue (27-30 um;

Miller 1885).

DisTRIBUTION—Cuba (Aptroot 2012). This represents a first report of

P. laetior for India and the first outside the type country.

Acknowledgments

The authors thankfully acknowledge the financial support of Department of

Biotechnology (DBT, Govt. of India) through Institutional Biotech Hub (No. BT/04/

NE/2009) for establishing the Lichenology Laboratory at Nowgong College, Assam,

to provide laboratory facilities for this study. The authors thank the Director of CSIR-

NBRI for providing the laboratory and herbarium facilities and Dr. D.K. Upreti for

his encouragement and help with the identification of the Pyrenula and Porina

specimens. We thank Drs. G.P. Sinha (Botanical Survey of India, Allahabad, India)

and Udeni Jayalal (Sabaragamuwa University of Sri Lanka, Belihuloya, Sri Lanka)

for reviewing the manuscript. One of the authors (SJ) is grateful to DST-SERB for

providing financial assistance under the NPDF scheme (PDF/2016/002054).

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44(1): 5-33. https://doi.org/10.1017/s0024282911000624

Breuss O, Liicking R. 2015. Three new lichen species from Nicaragua, with keys to the known

species of Eugeniella and Malmidea. Lichenologist 47(1): 9-20.

https://doi.org/10.1017/S0024282914000565

Czarnota P, Coppins BJ. 2006. A new Bacidia with long-necked pycnidia from Central Europe.

Lichenologist 38(5): 407-410. https://doi.org/10.1017/s0024282906005986

Gogoi R, Joseph S, Nayaka S, Yasmin F. 2019. Additions to the lichen biota of Assam State,

India. Journal of Threatened Taxa 11(6): 13765-13781.

https://doi.org/10.11609/jott.4642.11.6.13765-13781

Gupta P, Sinha GP. 2018. Lichen flora of Assam. Bishen Singh Mahendra Pal Singh, Dehra

Dun, India.

Bacidia, Malmidea, Porina & Pyrenula spp. new for India ... 663

Joshi S, Upreti DK, Hur JS. 2018. Key to the lichen families Pyrenulaceae and Trypetheliaceae

in Vietnam, with eight new records. Mycotaxon 132(4): 957-969.

https://doi.org/10.5248/132.957

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McCarthy PM. 2001. Trichotheliaceae. 105-157, in: PM McCarthy (ed.). Flora of Australia,

Volume 58A, Lichens 3. ABRS/CSIRO Australia, Melbourne.

Mercado-Diaz JA, Gould WA, Gonzalez G, Liicking R. 2015. Lichens in Puerto Rico:

an ecosystem approach. General Technical Report IITF-GTR-46. San Juan, PR: US.

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https://doi.org/10.2737/iitf-gtr-46

Miller J. 1885. Pyrenocarpeae Cubenses a cl. C. Wright lectae. Botanische Jahrbiicher fiir

Systematik, Pflanzengeschichte und Pflanzengeographie 6: 375-421.

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lichenogeographic regions. 79-96, in: T Marimuthu & al. (eds). Biodiversity Conservation

— Status, Future and Way Forward. National Academy of Biological Science, Chennai,

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Orange A, James PW, White FJ. 2001. Microchemical methods for the identification of lichens.

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Sinha GP, Nayaka S, Joseph S. 2018. Additions to the checklist of Indian lichens

after 2010. Cryptogam Biodiversity and Assessment, Special Volume: 197-206.

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MYCOTAXON

July-September 2020—Volume 135, pp. 665-718

https://doi.org/10.5248/135.665

Chromelosporium re-evaluated, with

Chromelosporiopsis gen. nov. and Geohypha stat. nov.

GREGOIRE L. HENNEBERT

Rue de l'Elevage 32, B 1340 Ottignies-Louvain-la-Neuve, Belgium

CORRESPONDENCE TO: hennebertg@scarlet.be

ABSTRACT—Chromelosporium-like asexual morphs of the subterraneous Pezizaceae in

the /Pachyphlodes clade are not congeneric with the type of Chromelosporium. Some

Chromelosporium morphs are of species in the /Pezizaceae clade (the /Peziza clade), and

the genus Chromelosporium as it has been defined is polyphyletic. A diagnostic character

that distinguishes these two groups is the presence or absence of conidiophore fasciculation.

Mononematous conidiophores characterize Chromelosporium and species related to the

/Peziza clade. Synnematous conidiophores characterize Pachyphlodes and other asexual

species and define the new genus Chromelosporiopsis, to be excluded from Chromelosporium.

Hyphelia terrestris, long misapplied to Chromelosporium, is revaluated, lectotypified,

redescribed and recombined as Geohypha terrestris.

Keyworps—hyphomycete, nomenclature, taxonomy

Introduction

Collections of Chromelosporium-like hyphomycetes made by R.P. Korf

around Ithaca, NY, which he tentatively named Glischroderma because of

their production of gel (Korf 1994), were revaluated by Healy & al. (2015)

and identified as conidial morphs in the truffle genus Pachyphlodes. Fresh

Chromelosporium-like specimens collected on bare soil in woods in Belgium,

identified by DNA sequencing as Pachyphlodes nemoralis and P. citrina,

provided the necessary asexual material to compare with and segregate from

the asexually typified species, Chromelosporium ochraceum (Hennebert &

Decock 2020).

666 ... Hennebert

The main distinction that emerged between the asexual morphs in

C. ochraceum and the two Pachyphlodes species is conidiophore fasciculation.

This criterion, applied to species already named in Chromelosporium

(Hennebert 1973), segregates them into two genera. It seems generally

accepted that these fungi are asexual morphs of Pezizales. As long as their

type species are not known to be congeneric with species with a known

sexual morph, however, it seems sensible to keep these genera separate.

Mononematous species define Chromelosporium. Synnematous species

are excluded and are accommodated in the new genus Chromelosporiopsis.

Names such as Botrytis epigaea, Phymatotrichum species, and others are

discussed as doubtful taxa. The conidial ornamentation observed in the

numerous herbarium specimens that might contribute to the diagnosis of

unidentified Chromelosporiopsis species is also analysed.

Hyphelia terrestris, currently misclassified in Chromelosporium, is re-

evaluated and recombined as the type of Geohypha [= Hyphelia sect.

Geohypha elevated to generic rank].

The asexual morph of Plicaria endocarpoides, previously supposed to

show similarities with Chromelosporium, is described.

The illustrated descriptions of the asexual morphology of these taxa

provide the rationale for the present paper.

Materials & methods

Species descriptions are based on observations made on herbarium material

borrowed from many herbaria and some fresh collections made during 1960-62 while

working as a postdoctoral researcher under the late Dr S.J. Hughes at the Agriculture

& Agri-Food mycology group in Ottawa (DAOM). This material is complemented

with some contemporary collections from Europe. A 1960 Olympus FH microscope

equipped with Olympus positive low phase contrast objectives and an adapted

Wild drawing tube was used for the microscopic examinations. All samples were

mounted in lactic acid with Cotton blue. Slides made from the examined specimens

are preserved in the Canadian National Mycological Herbarium, Ottawa, Canada

(DAOM) and/or Mycotheque, Université Catholique de Louvain, Louvain-la-Neuve,

Belgium (MUCL), and accession numbers are listed for each specimen in square

brackets. Information provided in the Examined specimen sections duplicates the

herbarium labels with bold numbers separating each specimen.

Taxonomy

Chromelosporium

This section includes the mononematous species considered morphologically

congeneric with the type species Chromelosporium ochraceum, which currently

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 667

delimits the genus Chromelosporium. In addition, one Peziza and one Plicaria

species with similar asexual morphs are each described.

Chromelosporium Corda, in Sturm’s Deutschl. Fl., Abt. 3 (Pilze), 3(13): 81, 1833.

TYPE SPECIES: Chromelosporium ochraceum Corda??

Ascomycota, Pezizaceae, sexual morph unknown.

HyPHAE septate, creeping on the substratum.

CONIDIOPHORES mononematous, erect, septate, apically branching in

successive symmetrical dichotomies, septate, apically obsuse.

CONIDIOGENUS CELLS subterminal and terminal branches forming

synchronous conidia on denticles along their lengths, collapsing after

rhexolytic secession of conidia.

Conrp14_ holoblastic, one-celled, globose or subglobose, with wall

ornamented from finely to coarsely verrucose, colored in mass.

HasitTat: on plant fragments.

COMMENTS—Hennebert (1973) made a clear distinction between

Chromelosporium and Ostracoderma Fr., so Chromelosporium should not be

placed as a synonym of Ostracoderma as cited in the 10" edition of Ainsworth

& Bisby’s Dictionary of the Fungi (Kirk & al. 2008) and in Index Fungorum.

Chromelosporium is also incorrectly listed as a synonym of Peziza in Baral’s

contribution to Jaklitsch & al. (2016). Such synonymy is indeed impossible—

Peziza being clearly polyphyletic and Chromelosporium an “orphan” genus.

Pfister & al. (2016) did not mention the case when evaluating competing

sexually and asexually typified names. Indeed, as the type species is not

connected to any sexual morph, the generic name Chromelosporium cannot

be competing so far. Also, Chromelosporium should not have been ignored

in the recent “Outline” of fungal genera (Wijayawardene & al. 2020). It is

possible that the four species described in the genus below, beside the type

species, might also be revealed as polyphyletic.

Chromelosporium ochraceum Corda,

in Sturm’s Deutschl. Fl., Abt. 3 (Pilze), 3(13): 81, 1833. FIGS 1, 2

= Sporotrichum ochraceum (Corda) Sacc. Syll. Fung. 4: 105, 1886.

= Ostracoderma ochraceum (Corda) S. Hughes, Canad. J. Bot. 36: 792, 1958.

= Botrytis dichotoma Corda, Icon. Fung. 1: 18, 1837.

= Campsotrichum dichotomum (Corda) Bonord., Handb. Mykol.: 102, 1851.

= Botrytis isabellina Preuss, Linnaea 25: 75, 1852.

= Ostracoderma isabellinum (Preuss) S. Hughes, Canad. J. Bot. 36: 792, 1958.

668 ... Hennebert

Fig. 1. Chromelosporium ochraceum. A. Conidiophore with conidiogenous cells (drawing

by S.J. Hughes, DAOM 83330). B, D. Conidiophore and conidia (holotype, PRM 155414), C.

conidiophore after secession of the conidiogenous cells (DAOM74695, Mycoth. Ven. 1080).

Scale bar = 10 um.

= Polyactis crystallina Bonord., Abh. Naturf. Ges. Halle 8: 95, 1864.

= Botrytis crystallina (Bonord.) Sacc., Syll. Fung. 4: 135, 1886.

= Botrytis spectabilis Harz, Bull. Soc. Imp. Nauralistes Moscou 44(1): 144, 1871.

= Clonostachys spectabilis (Harz) Oudem. & Sacc., Ned. Kruid. Arch., 2. ser., 4: 539, 1886.

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 669

Type: Chromelotrichtm [crossed out] sporium ochraceum Ca. ramulis furcis, Myrinema

generis” [scr. et del. A.C.J. Corda]. On Allium dead stem, Prague, Czechoslovakia.

Herb. Corda (Holotype: PRM 155414; isootypes: [DAOM 40946, MUCL.2895]

CoLonigs in small patches, <10 mm diam., velvety, at first white, then

ochraceous when mature.

HyPpHAE 3-5 um wide, creeping in the substratum.

CONIDIOPHORES mononematous, erect, emerging from hyphae in

substratum, with a bulbous basal cell, 28-35 x 12-15 um, ochraceous,

extending into a cylindrical, septate stipe, 100—400 um long, pale ochraceous,

the individual cells 40—95 x 10-14 um, terminally repeatedly branched with

up to five dichotomies, each branch with a basal septum and a second septum

before the subsequent dichotomy, the length of branch decreasing from one

dichotomy to the next, usually the first dichotomy 40-90 x 9-10 um, the

second 30—70 x 6-10 um, the third 30—50 x 6—9 um and so on, but in some

cases dichotomies being very short decreasing from 20 to 10 um, the angle of

the dichotomies ranging from 30° to 45°.

CONIDIOGENOUS CELLS comprising the last four and five terminal

dichotomies, together 70-100 x 6—9 um, apically clavate, <11 um, covered

with synchronous conidia borne on a denticle, and collapsing after conidial

release, seceding or (rarely) remaining as a frill on the last conidiophore cell.

Conrp1A holoblastic, borne simultaneously on denticles 2 x 1 um, one-

celled, globose or subglobose, ochraceous to salmon-colored in mass,

4—6.5 um, wall 0.5 um thick, cyanophilic, verrucose, warts prominent 0.3 um

high and blunt, with 12—14 warts in median view, seceding rhexolytically.

HasirtaT: on plant fragments.

DISTRIBUTION: Europe: Italy, France, Belgium, England, Africa: Maroc,

Uganda, Asia: Sumatra.

CoMMENTS— The name Chromelosporium remained unused until Hughes

(1958) brought the genus to light. Corda’s 1833 description of an overmature

collection was so unclear that even Corda (1837) himself did not recognize

the conspecificity of his later taxon, Botrytis dichotoma. His figure (Fic. 3A)

shows globose ochraceous conidia accumulated along the sides and the top

of the obtuse dichotomous branches.

Korf & Hennebert (1975) stated that Botrytis spectabilis was a synonym

of Chromelosporium ochraceum based on an original illustration from Harz

reproduced here (Fics. 3B-C) and preserved in MUCL 1072.

The names Botrytis spectabilis and Polyactis crystallina (another synonym

of Chromelosporium ochraceum) have been misapplied to the asexual morph

670 ... Hennebert

Fig. 2. Chromelosporium ochraceum (DAOM 46357). A. Conidiophores under low magnification.

B. Dichotomies of conidiophore. C-D. Conidiogenous dichotomies. E. Conidiophore after

secession of conidiogenous cells. F Verrucose conidia.

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 671

=. —

i 2 ” * . i 7 b =

é _

4B. dicholemie. A Nes

Fig. 3. Illustrations of A. Botrytis dichotoma Corda, Icon. Fung. 1: 18, fig. 244, 1837.

B-C. Botrytis spectabilis Harz, original drawings (MUCL 1072).

of Peziza ostracoderma Korf (Hennebert & Korf 1975). For that reason, both

Botrytis spectabilis and Polyactis crystallina have been erroneously listed as

synonyms of Peziza ostracoderma in Index Fungorum. Chromelosporium

ochraceum is also erroneously cited as a synonym of C. fulvum (Link)

McGinty& al., typified by the asexual morph of Peziza ostracoderma in

Seifert & al. (2012, p. 139).

The lectotype collection of Botrytis isabellina and the authentic collection

distributed in Rabenhorst’s KLoTzscHi] HERBARIUM VIVUM MYCOLOGICUM

N° 1570, were described in sufficient detail by Preuss (1853) as “Floccis erectis

crassis repetito ramosis dichotomis vel trichotomis, apice verrucosis sporis

irregulariter accumulatis, globosis, hilo instructo, episporio verrucoso” to

confirm its synonymy with Chromelosporium ochraceum.

SPECIMENS EXAMINED

TYPE & AUTHENTIC MATERIAL. EUROPE: Czecuia: (2) Botrytis dichotoma,

Herb.Corda (PRM 155393 type), [DAOM 84706, MUCL 2896]. GERMANY: (3)

Botrytis isabellina Preuss 1850, auf [rotten] nadelholz, Herb Preuss 451 (B)

(designated here as lectotype MBT 392406) [MUCL 5999]. (4) Botrytis isabellina Pr.,

ad lignum abietinum humi jacentem, prope Hoyerswerda [Germany], Preuss [1851],

672 ... Hennebert

Rabenh., Klotzschii Herb. Viv. Mycol. 1570 (B, PR, BM, BR, PAV, L) (isolectotypes)

[DAOM 51762, MUCL 1810]. (5) Botrytis spectabilis Harz, original drawing by Harz

(isolectotype 1975) (MUCL 1072).

OTHER SPECIMENS. EUROPE: ITALY: (6) Botrytis epigaea Link f. ochracea Sacc,

ad terram argillosam in horto botanico, Padova. Aug. 1902. D. Saccardo, Mycotheca

italica Mucedinaceae. [1903, Cent. XII] n° 1178 (PR 184863, BM, L 910.224-158)

[DAOM 74898, MUCL 1803]. [The name has been listed in Michelia I :102, 1877].

(7) Botrytis dichotoma Corda, in sarmentis, foliis, capsulis putrescentibus, in Horto

botanico ticinensi. Autumno, coll. Cavara. Micol. Lomb. n° 137, tab. 1, fig. 20-24

[1894]. Cavara, Fungi Longobardiae exsiccate. 195, (PAD, BM, PAV, BPI, NY, B, L)

[DAOM 83330, MUCL 1801]. (8) Botrytis fulva Link. in scapis Allii cepae, Consiglio

(Treviso). Sept. 1897. D. Saccardo Mycotheca italica Mucedinaceae. Cent. II.186, (K-

M) [DAOM 74699, MUCL 1802]. (9) Botrytis fulva Lk. Polyactis fulva Bon. in truncis

putridis. Selva (Treviso) Oct. 1876. Saccardo PA. Mycotheca veneta. Cent. XI. 1897.

n°1080 (K-M) [DAOM 74695, MUCL 1812]. (10) Botrytis fulva Link, ad corticem

ramorum, Valtaro, Sept. 1923, coll. Renji, det. G. Bresadola, (BPI) [DAOM 83884,

MUCL 2413]. (11) Botrytis fulva Link f. argillacea, ad Salicem petandram, Eichler,

Polachia, 1894. [scr. Bresadola] Herb. Bresadola, [In Bresadola Fungi polonici a

cl. Viro B. Eichler lecti. Ann. Mycol. 1, 2: 97-131, p. 127(1903), forma argillacea of

Botrytis fulva Link is not described] (S) [MUCL2531]. UK: (16) Botrytis fulva Link.

Queen’s Cottage grounds, Kew, Nov. 1898. G.E. Massee. (K-M) [MUCL 2295]. (17)

Phymatotrichum, on Fagus sylvatica rotten wood, Sheepleas, Horsley, Surrey, R.W.G.

Dennis, March 2 1952. (K-M-IMI 49481) [MUCL 3547]. (18) Botrytis splendida

Schwein., on mossy Salix bark, Wheatfen Broad, Norfolk, E.A. Ellis , (K-M-IMI

26983) [MUCL 3568]. (16) Phymatotrichum, on Petasites ovalis, Forge Valley,

Yorkshire, coll. I.M.I., det. S.J. Hughes. (K-M-IMI 8160) [MUCL 3581]. (19) Botrytis

(Phymatotrichum), on Heracleum, Forge Valley, Yorkshire, Sept. 16 1946, coll. I.M.L.,

det. S.J. Hughes. (K-M-IMI 6851) [MUCL3592]. (20) Botrytis, Phymatotrichum, on

Petasites dead petioles, Forge Valley, Yorkshire, Oct. 1945, coll. I.M.L, det. S.J. Hughes,

(K-M-IMI 1586) [MUCL 3595]. (21) Phymatotrichum, on Eupatorium cannabinum,

Farwath, Newton Dale, Pikering, Yorkshire, Aug. 6 1960, W. Bramley. (K-M-IMI

83016a) [MUCL 3622]. (22) Phymatotrichum, on Heracleum, Forge Valley, Yorkshire,

Sept. 16 1946, coll. I.M.L., det. S.J. Hughes. (K-M-IMI 7341) [MUCL 3582]. FRANCE:

(12) Botrytis fulva Link, in ligno putrido, Céte d’Or, Sept. 1889, coll Fautrey n°9 (PC)

[DAOM 83390, MUCL 2378]. (13) Botrytis fulva Bon., sur tricot de coton pourri

pres dun lavoir, Lyon, coll. J.J. Therry. Cryptogames du Lyonnais. J.J. Therry n° 6474,

(G) [MUCL 2398]. (14) Botrytis carnea Schum., ad culmos Paludi, Meudon, Nov.

3 1893, coll. E. Roussel]. (PC) [MUCL 2706]. BELGrum: (23) Phymatotrichum, on

herbaceous stems and leaves of compositae, Forét de Soignes, Sept. 16 1956, coll. M.B.

Ellis. (K-M-IMI 69956) [MUCL 3542].

AFRICA: Morocco: (24) Phymatotrichum, sur souche de Quercus ilex, Forét

dAzrou, Moyen Atlas, Maroc, 17 Nov. 1965, G. Malencon. Herb. Cryptog. G.

Malencgon 5805, (PC) [MUCL 8041]. UGANDA: (25) Botrytis, on dead wood,

Kampala, 4000; May 1936, G. Hansford. Herb. Mycol. Dep. Agric. Uganda n° 1769

(K-M-IMI 7899) [MUCL 3613].

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 673

ASIA: INDONESIA: (26). Botrytis? op aarde, bladeren, takjes, Sumatra, Kampong

Baroe bij Medan, Juli 1927, K.B. Boedijn. n° 4619 (Bot. Museum Utrecht 179892)

[MUCL 5381].

Fig. 4. Chromelosporium herbicola (isotype NY, MUCL 5889). A-C. Conidiophores missing the

seceded conidiogenous cells. D. Verrucose conidia. Scale bar = 10 um.

Chromelosporium herbicola (Ellis & Dearn.) Hennebert, comb. nov. Fic. 4

MB 833990

= Rhinotrichum herbicola Ellis & Dearn., Proc. Canad.

Inst., n.s. 1: 90, 1897 [(as “herbicolum” |.

Type: Rhinotrichum herbicolum E|llis] & Dearness, on Solidago virga-aurea golden rod

stems, Aug. 13 1895, D.V.S. Valley, London, Canada, J. Dearness. Herb. Dearness 2314

(Holotype: DAOM [MUCL2321]; isotype NY [MUCL 5889]).

674 ... Hennebert

COLONIES in small patches, 3-5 mm wide, about 1 mm high, ochraceous.

HyPpHAE 5—15 um wide, creeping on the substratum.

CONIDIOPHORES mononematous, erect, emerging single from creeping

hyphae, as a stipe 100-150 um high, 9-15 um wide, ochraceous, slightly

constricted at septa in cells 25-35 um long, terminally branching in

successive dichotomies, at 10-45° angles, branches septate at the base and

sometimes over their length, first dichotomies 30-50 um long, second

dichotomies 20—25 um long,

CONIDIOGENUS CELLS presumably the third and fourth dichotomies

collapsed and fallen away, not observed in the material but similar to those

of Chromelosporium fulvum after Sumstine (1911).

Conip1a presumably holoblastic, borne simultaneously all along the

conidiogenous cell as described by the original authors, globose to obovate,

one-celled, (6—)7—9(—11) um, ochraceous, thick-walled and verrucose, with

18—20 warts in median view, seceding rhexolytically.

Hasirat: On decaying herbaceous plant stems, similar to Solidago.

DISTRIBUTION: Canada: Ontario.

CoMMENTS — The original description of Rhinotrichum herbicola by Ellis &

Dearness reads: “Effuse, light yellow, becoming brown in the center. Hyphae

coarse, septate, branched, nearly hyaline, 8—10 u, thick, Fertile hyphae sub-

undulate above, tips swollen and bearing the globose, sub-hyaline, finely

echinulate, 7—9 u, conidia. On dead stems of Solidago canadensis, London,

Can., Aug. 1895. Herb. Dearness n°2314

Sumstine (1911) examined the isotype specimen in NY and observed

that the “spores clustered at the apex of the fertile branches” Although

“the specimen proved to be poor’, he concluded that “it resembles in

many respects Botrytis fulva Link.’ This comment joined to the observed

dichotomous branching and the verrucose globose conidia leads to transfer

the species to Chromelosporium.

The species differs from Chromelosporium ochraceum by its shorter

conidiophores and conidial size and ornamentation.

Chromelosporium canadense Hennebert, Persoonia 7: 196, 1973. FIGS 5, 6

= Rhinotrichum carneum Ellis & Everh., J. Mycol. 1(7): 93, 1885

(non Chromelosporium carneum (Pers.) Hennebert

TyPE: Chromelosporium canadense Hennebert, on rotten wood in forest, Gatineau

Park, Gatineau, Quebec, Canada, 25.11.1960, leg. S.J. Hughes & G.L. Hennebert,

(Holotype: DAOM 71947; isotype: MUCL 1689).

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 675

Fig. 5. Chromelosporium canadense (holotype, DAOM 71947). A. Young conidiogenous cells

on terminal dichotomies. B. Primordia of the conidiogenous cells. C. Conidiogenous cells.

D. Dichotomous branching of the conidiophore. E. Mature conidia. Scale bar = 10 um.

676 ... Hennebert

COLONIES in patches <15 mm wide and <3 mm thick, effuse, loose, first

white, then tawny to cinnamon, composed of mixed hyphae and branched

conidiophores bearing clusters of conidia.

HypuHAe on substrate, thin, 4-7 um, evanescent.

CONIDIOPHORES mononematous, erect, single, tawny, stipes 100-300 x

6-10 um, then branching in regular and repeated dichotomies of different

lengths, branches divergent at 20—60° angle, the basal ones 160-200 um

long, the distal ones shorter (16—48 um), 10—12(—18) um wide, the stipe

and each branch septate at its base, the last dichotomy bearing two clusters

of 4—8 primordial conidiogenous cells.

CONIDIOGENUS CELLS formed as two groups of quadri-furcate terminal

primordia that elongate up to 30—45 x 12—22 um, slightly clavate, becoming

conidiogenous over their entire length.

Conip1A4 holoblastic, borne simultaneously on denticles, one-celled,

globose, (3.5—)4—5(—7) um, tawny, verrucose with 10-18 warts in median

view, seceding rhexolytically.

HABITAT: on rotten wood and bark, branches, mosses, plant stems and

herbs, in gardens and forests.

DISTRIBUTION: Canada and USA.

CoMMENTS—Growth on PDA was obtained from young conidiophore

hyphae taken after 3 days at 4°C from fresh material of four collections

(MUCL 1689 T, 1690, 1691, 1692) but not from conidia. Mycelium in

culture is effuse, ochre, forming dense dispersed patches; hyphae are regular,

septate, branching at an angle of c. 45°, or very irregular, closely septate in

swollen cells. In two cases, small clusters of dark brown cells were produced,

suggesting primordia of apothecia.

Herbarium specimens often lack the conidiogenous cells which are

collapsed or absent; their presence is inferred from the branching pattern

and the conidial characters.

SPECIMENS EXAMINED:

TYPE & AUTHENTIC MATERIAL: USA: OREGON: (2) Rhinotrichum carneum Ell. &

Ev., on bark, Oregon, 1885, W.S. Carpenter n° 125 [scr. Ellis] (NY).) [DAOM 84659,

MUCL 2694]. (holotype) NEw York: (3) Rhinotrichum carneum Ell. & Everh., on

rotten wood, Macoun [scr. Ellis]. Ellis Collection (NY) [DAOM 84677, MUCL 2839].

(4) Rhinotrichum carneum Ell. & Everh. [no substrate, no date], Macoun [scr. J.B.

Ellis]. Ellis Collection (NY) [MUCL 2840].

OTHER SPECIMENS: CANADA, Ontario (5) Hyphelia terrestris, on Tilia,

Nashville, York Co., Oct. 25 1953, coll. det. RF. Cain (TRT 30178) [DAOM 84684,

MUCL 2858]. (6) Hyphelia terrestris, on hardwood, Nashville, Nov. 14 1954, coll.

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 677

Fig. 6. Chromelosporium canadense (holotype, DAOM 71947). A. Conidiophores at low

magnification. B-D. Young stages of conidiogenous cells. C. Half and fully mature conidioma. E.

Wide angle of the dichotomies. F. Terminal bunch of mature conidiogenous cells.

det. R.E. Cain (TRT 31311) [DAOM 84683, MUCL 2857]. (7) Hyphelia terrestris, on

Populus Nashville, Oct. 22 1955, coll. det. RE Cain (TRT 31699, BPI) [MUCL 2419].

(8) Hyphelia terrestris, on decayed log, New Durham, Brant Co., Oct. 14 1956 (TRT

678 ... Hennebert

32870) [DAOM 84688, MUCL 2862]. (9) Botrytis fulva Link, on dead wood, Oct.

20 1896, Macoun [Scr. J.B. Ellis]. Herb. Ellis (BPI) [DAOM 83886, MUCL 2415].

(10) Botrytis (Phymatotrichum) fulva Link, on decayed wood, N. Bathurst street

Toronto, Nov. 17 1934, H.S. Jackson, det. D.H. Linder, Herb. R.E Cain 8207 (TRT

8065, FH, DAOM 81409) [MUCL 2889)] (11) Botrytis (Phymatotrichum) fulva Link,

on rotten wood, S. Aurora, Oct. 23 1932, H.S. Jackson, det. D.H. Linder. Herb. R.E.

Cain 8210 (TRT 8060, DAOM 81410) [MUCL 2890]. (12) Ostracoderma, on soil

of lawn, North Toronto, June 17 1960, S.J. Hughes. (DAOM 83325) [MUCL 2255].

(13) Chromelosporium, on bark and plant decay, South March, April 10 1962, G.L.

Hennebert (MUCL 3067). QUEBEC (14) Ostracoderma on bark, Gatineau Park,

Nov. 25 1960, S.J.Hughes. (DAOM 71946) [MUCL 1688]. (15) Ostracoderma, on

wood, Gatineau Park, Nov. 25 1960, S.J.Hughes. (DAOM 71948) [MUCL 1690]. (16)

Ostracoderma, on wood, Gatineau Park, Nov. 2(5, 1960, S.J.Hughes. (DAOM 71949)

[MUCL 1691]. (17) Ostracoderma, on wood, Gatineau Park, Nov. 25, 1960, S.J. Hughes.

(DAOM 71950) [MUCL 1692]. (18) Ostracoderma, on wood and bark, Gatineau Park,

Oct. 31, 1960, S.J. Hughes. (DAOM 71399) [MUCL 1689]. (19) Ostracoderma, on stem

of Veratrum viride, Yoho Valley, Aug. 8 1960, S.J. Hughes. (DAOM71326) [MUCL

1513]. (20) Chromelosporium, on Betula papyrifera rotten wood, Gatineau Park, Oct.

31 1960, G.L.Hennebert. (DAOM 71545, MUCL 1590). (21) Chromelosporium, on

rotten wood, Gatineau Park, Oct. 7 1960, G.L.Hennebert. (DAOM 71419, MUCL

1505). MANITOBA (22) Botrytis near phymatotricha Sacc. prov. cfr. lateritia, C.E.

Fairman suggests Botrytis carnea Schum, on old board, Winnipeg, Sept. 16 1928,

G.R. Bisby 4140 [scr. Dearness]. Herb. Dearness 6732 (DAOM 84789) [MUCL 3070].

USA, OunI0 (23) Mycobiota of Ohio. Botrytis isabellina Preuss, on rotten wood, The

7 Caves, Highland Co., coll. det. D-H. Linder 2667, Sept. 6 1933, Herb. W. Bridge

Cooke (NY, FH, DAOM 84679) [MUCL 2847]. Iowa (24) Hyphelia laxa Schw., on

decorticated conifer wood. (IA) [DAOM 83317, MUCL 2260]. SouTH CAROLINA:

(25) Botrytis epigaea Link var. on clay surface after rain, Sept. 1883, H.W.R. [scr. H.W.

Ravenel]. Herb. U.W. Ravenel, (K-M, DAOM 83332) [MUCL 2281]. MASSACHUSETTS

(26) Botrytis? Hyphelia spectabilis Harz, on coniferous bark, York, Sept. 14 1891.

Herb. Thaxter 137 (FH) [DAOM 84702, MUCL 2886]. (27) Botrytis fulva Link, on

hardwood branches on the ground, Beaver Brook reserve, Belmont, Oct. 3 1937, coll.

G.D. Darker 64-90, det. D.H. Linder. (FH) [DAOM 84703, MUCL 2888]. NEw YorRK

(28) Ostracoderma canadense, on stem of fern in a pond, Forest Home, Ithaca, May 1

1962, G.L. Hennebert. (MUCL 3120). NEw JERSEY (29) Botrytis epigaea Pers., on old

pine boards partly buried in the ground, Newfield, N.J., Sept. 1881, Ellis. Ellis North

American Fungi 827 (WIS, CUP, K-M, B, G, L) [MUCL 1804].

Chromelosporium arenosum Hennebert, Persoonia 7: 196, 1973. FIG. 7

Type: Flora Venezuelae 2474, on rotten wood of Espeletia, 3550 m, alt., Mucudaji,

Sanide Santo Domingo, Estada Mesida, July 22 1958, coll. R.W.G. Dennis. (K-M)

(Holotype: MUCL 2298 [DAOM 83359]; isotype IMI 75582 [MUCL 5889]).

EryMo.Locy: from the Latin arenosus or sand-like, suggesting the spreading

granulose aspect of the sporulating conidiophores of ochraceous color seen under

low magnification.

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 679

Fig. 7. Chromelosporium arenosum (holotype, DAOM 83359). A. Young conidiophore branching.

B. Erect conidiophore with conidiogenous cells. C. Mature conidiophore with conidia borne on

aerial hypha. D. Conidiophore after secession of conidiogenous cells. E. Conidia. Scale bar = 10 um.

COLONIES superficial, sparse, delicate, cobweb-like white mycelium, becoming

arenose ochraceous with mature conidiophores appearing like minute,

scattered grain.

Hypuae repent, hyaline, single, 7—9 um wide, regular, straight, branched,

thin-walled, smooth, septate with cells 7-12 um long, sometimes regenerating

after collapse from septum by internal narrow hyphal proliferations 3—4 um

wide.

CONIDIOPHORES arising singly, mononematous, as erect branches of the

repent hyphae, short, 70-100 um high, 8—9 um wide, stipes with the basal

cell bulbous 11—13 x 6—8 um,, constricted at septa, cells 15-50 um, apically

branching in dicho- or trichotomies forming 2—8 terminal cells 40-50 um

long, obtuse, clavate up to 13 um wide and conidiogenous.

680 ... Hennebert

Conip1a holoblastic, borne simultaneously on denticle 1 x 0.5 um, one-

celled, globose or ovoid, 3.5—5.5 x 3.5—6.5(—8.1) um, mostly 4.5 um wide,

0.5—1 um thick-walled, the outer wall hyaline, the inner wall subhyaline to

ochraceous and coarsely verrucose, with 6—10 globoid or elongate warts in

median view, seceding rhexolytically.

HABITAT: on rotten bark of Espeletia sp. in the Tropics.

DISTRIBUTION: Venezuela.

Chromelosporium macrospermum Hennebert, Persoonia 7: 197, 1973. Fic. 8

Type: living and dried cultures from Peziza sp. ind. on sterilized soil in greenhouse,

Heverlee, Belgium, 2.04.1960, leg. G.L. Hennebert (Holotype: MUCL 1116; isotype

DAOM 67492 = dried culture of MUCL 1116).

COLONIES on malt agar substrate thin, white to rust-color when sporulating,

comprising interwoven and prostrate hyphae, hyaline to subhyaline,

producing laterally large spherical cells, 40-50 um diam, thin-walled.

CONIDIOPHORES mononematous, erect, pale to rust-colored, septate,

stipes 400-600 x 15-18 um, apically bi- or quadri-furcate, branches 20 um

long, few septate, the distal ones erect, radiate, cylindrical to clavate 130-160

x 20-25 um, conidiogenous.

Conip1A holoblastic, borne on a denticle, more or less simultaneous,

globose, rust-colored, 15-23 (av. 19.7) um, thin-walled, finely punctate,

seceding rhexolytically.

Hapsirat: sterilized greenhouse or nature soil

DISTRIBUTION: Europe: Belgium, Oceania: Australia.

ComMENnts — This unique strain MUCL 1116 was found in one ascospore

germinating from several apothecia then identified as Peziza ostracoderma,

collected in a tropical greenhouse of the Faculty of Agronomy UCL in

Heverlee, Belgium in 1960. Korf differentiated the size and ornamentation of

the ascopores preserved in lactic acid mounted slides of the original Peziza

apothecia (Hennebert & Korf 1975) but in the absence of the single original

apothecium, he could not circumscribe the species. The same species has been

isolated from soil in South Australia by Dr. G.C. Hansford at the University

of Adelaide in 1953 and preserved as living culture IMI 54710 in the culture

collection at Egham and as dried culture in K-M—IMI 54710 under the name

Phymatotrichum tax. sp. 7. The culture is C. macrospermum mixed with

conidia of P. ostracoderma and unidentified hyaline chlamydospores.

The species might be a mutant Peziza ostracoderma or a distinct

undetermined species of Peziza.

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 681

Fig. 8. Chromelosporium macrospermum (ex holotype, MUCL 1116). A-B. Conidiophores at

diverse stages. B. Punctate conidia. D. Swollen cells in culture. Scale bar = 10 um.

682 ... Hennebert

Species of Pezizaceae with known sexual morphs and

similar mononematous conidiophores and conidiogenesis

Peziza ostracoderma Korf, Mycologia 52: 650, 1961 [“1960”]. Fic. 9

= Plicaria fulva R. Schneid., Zentralbl. Bakteriol. Parasitenk. Infektionskrankh.

Hyg., 2. Abt., 108: 147, 1954 [non Peziza fulva Pers. 1822, nom. sanct.].

= Dematium ollare Pers., Syn. Meth. Fung. 2: 697, 1801.

= Botrytis fulva Link, Spec. Plant., ed. 4, 6(1): 58, 1824, nom. illeg. [superfluous].

= Sporotrichum fulvum Fr., Syst. Mycol. 3: 418, 1832, nom.

sanct. [not “(Link) Fr’; non Link 1809].

= Trichosporum fulvum (Fr.) Fr., Summa Veg. Scand. 2: 492, 1849.

= Polyactis fulva (Fr.) Bonord., Handb. Mykol.: 115, 1851 [not “(Link) Bonord.”].

= Chromelosporium fulvum (Fr.) McGinty, Hennebert & Korf,

Mycologia 67: 216, 1975 [not “(Link) McGinty et al.”].

= Chromelosporium ollare (Pers.) Hennebert, Persoonia 7: 197, 1973.

= Haplaria nitens Delacr., Bull. Soc. Mycol. France 6: 140, 1890.

= Botrytis luteobrunnea Krzemien. & Badura, Acta Soc. Bot. Poloniae 23: 727, 1955 (1954).

= Mycotypha dichotoma F.A. Wolf, J. Elisha Mitchell Sci. Soc. 71: 217, 1955.

= Ostracoderma dichotomum (F.A. Wolf) Matsush., Icon.

Microfung. Matsush. Lect.: 103, 1975.

Types: Plicaria fulva R. Schneid., on damp sterilized soil in greenhouse, Berlin-

Dahlem, Germany, 25.3.1953 (Holotype: B; authentic material in CUP R.P.K. 4114).

Dematium ollare Pers.: dried culture from living culture of Plicaria fulva R.

Schneider, on damp sterilized soil, greenhouse, Berlin-Dahlem, Germany, March

1953, received in Febr. 1954 from R. Schneider (Neotype: MUCL 1112, designated

in Hennebert 1973; isoneotypes: dried culture DAOM 81809, living cultures CBS

382.54, CCRC 36608, IMI 059206).

COLONIES velvety to floccose, white when young, yellow to fulvous or

cinnamon when sporulating, rapidly growing.

HypHAE mostly prostrate, interwoven, branched, septate, anastomosing,

hyaline to pale fulvous.

CONIDIOPHORES mononematous, solitary or gregarious, erect and

divergent, arising from prostrate or aerial hyphae; stipe 130-600 x 8-17 um,

hyaline to fulvous, sparsely septate, attenuate at the base, apically 1-3 times

dichotomously branched, dichotomies short, 10—25 x 5-13 um, septate at

the base, the terminal ones clavate, slightly inflated <15 um, conidiogenous.

CONIDIOGENUS CELLS producing conidia along their length, collapsing

after release and seceding away at basal septa, leaving the stipe or the lowest

dichotomies as a stump.

Conip1a holoblastic, borne simultaneously on a denticle 3 x 1 um, densely

and irregularly spaced, globose or napiform, 5—9(—14) um (mean = 7.5 um),

fulvous, wall thin, finely punctate, seceding rhexolytically. Conidia

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 683

Fig. 9. Peziza ostracoderma (neotype MUCL 1112). A-C. Conidiophores at diverse stages.

D. Punctate conidia. Scale bar = 10 um. The magnification of the line-drawings of conidia in

figures 8 and 9 is half that in other figures.

germinate and colonies sporulate profusely on malt agar and diverse organic

poor culture media.

Hasitat: On sterilized soil in greenhouses, on mushroom beds, on

cardboard, paper pots, paper, and old textiles in moist chambers, on humid

plaster walls in houses, on moist hay, on dung of rodents, and on forest soil.

DISTRIBUTION: Europe, North America, Asia.

ComMMENTs—Korf (1961) knew the species in both its asexual and sexual

morphs in the greenhouses at Cornell when he received the type and a living

culture of Plicaria fulva. The species has elliptical ascospores like most Peziza

species. Seeing the sanctioned name Peziza fulva Pers., he created the new

684 ... Hennebert

name Peziza ostracoderma, referring to Hughes’ opinion that the conidial

morph could be referred to Ostracoderma. Hennebert (1960) reported and

described the same fungus observed in a tropical greenhouse of the Faculty

of Agronomy UCL in Heverlee, Belgium, in 1960.

Hennebert (1973) segregated Chromelosporium from Ostracoderma,

which Hughes (1958) had considered synonymous. He named the conidial

morph of P. ostracoderma, Chromelosporium ollare,, with Dematium ollare

as basionym, which he neotypified with GLH 1112 (MUCL 1112) in the

absence of an original authentic specimen from Persoon.

Hennebert & Korf (1975) cited Dematium ollare as the earliest post-

Linnean name for the conidial morph common in greenhouses. Link (1824)

transferred the species to Botrytis as Botrytis fulva, an illegitimate name with

Dematium ollare cited as its unique synonym. Fries (1932a) excluded D. ollare

from his genus Dematium, treating it, after Link, as a synonym of B. fulva,

both of which he transferred into Sporotrichum as S. fulvum, a sanctioned

name.

Searching for the earliest possible name to combine in Peziza in

accordance with the 2017 Shenzhen ICN, it is apparent that the earliest name

of the fungus, Dematium ollare, cannot be used, because of Peziza ollaris Fr.

(Fries 1822), a different species, the name of which is sanctioned over Peziza

ollaris Schaeff. (Fungorum in Bavaria et Palatinatu circa Ratisbonam Icones

4:126, 1774) and Peziza ollaris Pers. (Mycol. Eur. 1: 299, 1822), which might

be the present species.

Despite what is posted on Index Fungorum, the names Alytosporium

fulvum (Link) Link and Nodulisporium fulvum S. Hughes, being based on

Sporotrichum fulvum Link, are synonymous with neither Dematium ollare

nor Botrytis fulva.

Haplaria nitens, added here as a synonym, was collected on sterilized oak

barks from a tannery used as mulch in a warm glasshouse of the Laboratoire

de Pathologie végétale de l'Institut national Agronomique in Paris. Its conidia

are minutely verrucose.

The variable conidiophore length and variable number and length of

the dichotomies of the species might explain its frequent confusion with

Chromelosporium ochraceum. ‘The primary diagnostic differences reside in

conidial size and ornamentation and the ecological habitat.

In an attempt to clarify the relationship with Chromelosporium

macrospermum, the variability of conidial size in monoascospore strains

of Peziza ostracoderma was studied on different culture media (malt-agar,

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 685

potato-dextrose-agar, carrot-agar, and jiffy-agar, a mixture of soil and peat).

Although conidial size varied between 5-9 um and one strain produced

conidia <12 um and a different strain produced conidia <14 um, none

reached 15—23 um, the conidial size reported for C. macrospermum of.

TYPE SPECIMEN EXAMINED. EUROPE: FRANCE: (2). Haplaria nitens Del. n. sp. Paris

(scr. Delacroix). (holotype, PC) [isotypes, DAOM 59453, MUCL2700]

OTHER SPECIMENS EXAMINED See Hennebert & Korf (1975).

Plicaria trachycarpa (Curr.) Boud.,

Hist. Class. Discom. Eur. 50, 1907, var. trachycarpa Figs 10, 11

= Peziza trachycarpa Curr., Trans. Linn. Soc. London 24: 493, 1864.

= Discina trachycarpa (Curr.) P. Karst., Acta Soc. Fauna Fl. Fenn. 2(6): 113, 1885.

= Detonia trachycarpa (Curr.) Sacc., Syll. Fung. 8: 105, 1889.

= Curreyella trachycarpa (Curr.) Massee, Brit. Fung.-Fl. 4: 401, 1895.

= Lamprospora trachycarpa (Curr.) Seaver, Mycologia 6: 19, 1914.

= Plicariella trachycarpa (Curr.) Velen., Monogr. Discomyc. Bohem.: 342, 1934.

= Galactinia trachycarpa (Curr.) Le Gal, Bull. Trimestriel

Soc. Mycol. France 78: 212, 1962.

= Rhinotrichum trachycarpum F.A. Wolf, J. Elisha Mitchell Sci. Soc. 74: 166, 1958.

= Chromelosporium trachycarpum Hennebert, Persoonia 7: 197, 1973.

Types: Peziza trachycarpa Curr., supra solum deustum, Ascot com. Surrey, Nov. 1862

(Holotype: K(M) 29980).

Chromelosporium trachycarpum Hennebert, asexual morph of Peziza

trachycarpa, on burned area, Camp II, Allegany State Park, USA: June 11, 1961, coll. &

isol. G.L. Hennebert, det. R.P. Korf (Holotype, DAOM 83324; isotype, MUCL 2197).

CoLonigs fast growing on malt agar medium, pellicular, white to pale

ochraceous.

Hypuas thin, hyaline, interwoven and prostrate.

CONIDIOPHORES mononematous, erect, very short, 50-100 x 8-10 um,

repeatedly dichotomously or assymetrically branched, irregularly septate,

branches cylindrical, short, 20-35 x 8—10 um, widely divaricate, becoming

totally conidiogenous, forming compact globose conidial heads.

Conrp1A holoblastic, borne simultaneously on denticle, yellow-ocher,

one-celled, globose or shortly ovate, 5—7 x 5—9 um, wall finely verrucose,

with 10—15 warts in median view, seceding rhexolytically.

Hasirat: in moist burned area in forests.

DISTRIBUTION: Northern Hemisphere.

CoMMENTS— The drawings were made while I was in Canada from an

ascospore culture of the species collected in Ontario (MUCL 23339) identified

686 ... Hennebert

Fig. 10. Plicaria trachycarpa. A-B. Branched conidiophore. C. Terminal conidiogenous cells.

D. Much septate conidiophore on porcupine dung (MUCL 2933). E. Punctate conidia.

(A-C and E on malt agar MUCL 2339). Scale bar = 10 um.

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 687

Fig. 11. Peziza trachycarpa. A. Initiation of conidiophores. B-D. Globular branched conidiophores

showing the dichotomies. E. Initiation of conidia. (A-E, MUCL 2339). E Mature conidiogenous

cells (DAOMC 199631 = MUCL 57201 on malt-agar).

688 ... Hennebert

as identical to DAOM 83324 but not after examination of K(M) 29980.

A living strain labelled as Plicaria trachyspora var. muricata (= Plicaria

carbonaria (Fuckel) Fuckel 1870) was recently received as DAOMC 199631

that, on 2 % malt agar, developed similar clusters of dichotomous branches

close to the substrate and bearing mature conidia.

SPECIMENS EXAMINED;

NORTH AMERICA: USA, New York (2) Peziza trachycarpa Currey, on burned

area, State Line Run, near Onoville, Cattarangu Co., June 10 1961, coll. Kumi T Korf, det.

R.P. Korf (MUCL 2198). (3) Chromelosporium, on porcupine dung (in moist chamber)

from Stoddard Brook Road, Allegany State Park, June 11 1961, coll. & isol. G.L.

Hennebert. (DAOM 89363, MUCL 2933). CANADA, Ontario (4) Peziza trachycarpa

Currey, on moist clay soil in mixed woods, Bell’s Corners, July 31 1961, coll. & isol.

G.L. Hennebert 2339. (DAOM 83382, MUCL 2339). (5) Plicaria trachycarpa (Currey)

Boudier var. muricata Grelet on burned litter in burned mixed forest, Renfrew Co., Ont.,

Sept. 2 1979, K.N. Egger 0281, dried and culture (CCF 6918, DAOMC 199631, MUCL

57201).

EUROPE: UK (6) Peziza trachycarpa (Curr.) Boud., conidia on cardboard flower

pot., Evesham, Worcs., March 25 1955, col. R.E. Taylor, det. R.W.G. Dennis. Dried

culture IMI 59800) [MUCL 3514].

Chromelosporiopsis

Chromelosporium-like species with synnematous conidiomata resemble

mononematous Chromelosporium species in having the same conidiogenesis

and ornamented globose conidia but differ in the conidiophore

fasciculation and asymmetric sparsely septate dichotomous branching (a

branching pattern described here as coralloid). This concerns two asexual

species named by Hennebert (1973)—Chromelosporium carneum and

C. coerulescens—both with sexual morph unknown. For these species, the

new generic name Chromelosporiopsis is proposed. Similar asexual morphs

have also been described in some Pachyphlodes species of the Pezizaceae.

Chromelosporiopsis Hennebert, gen. nov.

MB 835622

Differs from Chromelosporium by its synnematous conidiophores and irregular

successive bifurcate branching.

TYPE SPECIES: Chromelosporiopsis carnea (Schumach.) Hennebert

EryMo.ocy: Chromelosporium + -opsis, similar to.

Ascomycota, Pezizaceae, sexual morph unknown.

HyPHAE septate, branched and anastomosed, fasciculating in synnemata,

forming defined or effuse cushions, diversely colored (white, ochre, yellow,

rose, flesh red, blue, or violet).

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 689

CONIDIOPHORES synnematous, laterally and/or apically branching in

irregular successive dichotomies called here a coralloid pattern, sparsely

septate, apically slightly inflated.

CONIDIOGENUS CELLS subterminal and terminal branches forming

synchronously along their lengths holoblastic conidia, each on a denticle;

secession rhexolytic.

Conip1A one-celled, globose or subglobose, with thick ornamented

(finely to coarsely verrucose) wall, colored in mass. seceding rhexolytically.

HABITAT: on organic soil.

Chromelosporiopsis carnea (Schumach.) Hennebert, comb. nov. _ FIGS 12-14, 15A

MB 835623

= Botrytis carnea Schumach., Enum. Pl. 2: 238, 1803, nom. sanct.

= Isaria carnea Pers., Ann. Bot. (Usteri) 15: 13, 1795; Syn. Meth. Fung. 2: 689, 1801.

= Chromelosporium carneum (Pers.) Hennebert, Persoonia 7: 196, 1973.

= Polyactis carnea Ehrenb., Sylv. Mycol. Berol. 25, 1818.

= Mucor carneus (Ehrenb.) Link, Spec. Plant., ed. 4, 6(1):

88, 1824, nom. illeg., non Schaeff. 1774

= Botrytis carnea (Ehrenb.) Spreng., Syst. Veg., ed. 16, 4(1):

551, 1827, nom. illeg., non Schumach. 1803.

= Sporodinia carnea (Ehrenb.) Wallr., Fl. Crypt. Germ. 2: 317, 1833.

= Ostracoderma carneum (Ehrenb.) S. Hughes, Canad. J. Bot. 36: 792, 1958.

= Botrytis rosea Link, Mag. Ges. Naturf. Freunde Berlin

7: 36, 1815, nom. illeg., non DC. 1805.

= Botrytis linkii Duby, Bot. Gall, 2: 919, 1830 [as “linckii”].

= Ostracoderma linkii (Duby) S. Hughes, Canad. J. Bot. 36; 792, 1958.

= Campsotrichum splendidum Schwein.,Trans. Amer. Philos. Soc., n.s. 4: 283, 1832.

= Botrytis splendida (Schwein.) Sacc., Syll. Fung. 4: 123, 1886.

= Rhinotrichum opuntia Berk. & Broome, Ann. Mag. Nat. Hist., ser. 2, 13: 462, 1854.

= Botrytis carnea f. foliicola Roum., Fungi. Sel. Gall. Exs., Cent. 14: no. 1367, 1881.

Types—Polyactis carnea Ehrbg.[scr. Ehrenberg] on leaves of Fagus sylvatica, Herb.

Schwaeningrichen (STR) (MBT 392408, here designated as lectotype [DAOM

83897, MUCL 2440]). Polyactis carnea Ehrbg. [scr. Ehrenberg] on leaves of Fagus

sylvatica, Herb. Nees (STR) (Isolectotypes: DAOM 83896, [MUCL 2439]; Herb.

Persoon (L 910-262-747) [MUCL 2464]).

Chromelosporium carneum (Pers.) Hennebert on leaves of Quercus pedunculata

and Fagus sylvatica, Forest de Soignes, Tervueren, Belgium, Aug. 18 1960, leg. G.L.

Hennebert (Epitype: MUCL 1208, designated as “neotype” by Hennebert, 1973;

isoepitype: DAOM 74697).

CoLonligs superficial, fast growing and evanescent, in scattered tufts of hyphae,

3-15 mm wide, 15-2 mm high, comprising <15 erect divergent synnemata

690 ... Hennebert

Fig. 12. Chromelosporiopsis carnea. A Synnemata (MUCL 1806) .B. Coralloid branching of the

conidiophores. C. Conidiogenous cells. D. Verrucose conidia (B-D, isolectotype MUCL 2464).

Scale bar = 10 um.

bearing conidia in their upper half, white when young, flesh-colored when

mature on the field, ochraceous salmon when dried.

HypHae repent, sparse, delicate, hyaline to ochraceous, single or aggregated,

(9—)12—15(—23) um wide, thin-walled, smooth, septate, branched, often

anastomosing, with short cells.

SYNNEMATA arising from the basal mat of hyphae, erect columnar, <1.5 mm

high, 50 um wide, composed of <30 growing conidiophores.

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 691

Fig. 13. Chromelosporiopsis carnea, cluster of synnemata with isarioid conidial head (MUCL 6278).

CONIDIOPHORES stipes contiguous, 11-17 um wide, anastomosing, septate

at 50—80-um intervals, thin-walled, smooth, slightly bent outwards laterally

and apically in the upper half of the synnemata and branching in a coralloid

pattern of asymmetric dichotomies, one branch of each dichotomy developing

first and sometimes becoming septate at the base,

CONIDIOGENOUS HEAD formed of 4—9 last successive bifurcations, the

branches being 15-50 x 6-10 um, apically obtuse, slightly clavate, 7-12 um

wide and often bent, all branches conidiogenous, forming an isarioid head.

Conip1A holoblastic, borne simultaneously on denticle 1 x 0.5 um, 5-11

um distant from each other, one-celled, globose, occasionally subglobose,

4.8-6.6(—7.8) um (most 5.4-6 um), walls 1-1.5 um thick, the inner wall

ochraceous, the outer wall cyanophilic and coarsely verrucose, bearing 8-12

rounded warts in median view, seceding rhexolytically.

HasiTaT: on dead leaves of Fagus sylvatica and Quercus pedunculata,

also on other leaves, bark, mosses, and organic debris on soil, in forests, from

August to October.

DISTRIBUTION: Europe: Austria, Belgium, France, Denmark, Germany,

Great Britain, Netherlands, Sweden.

CoMMENTS— The earliest description and illustration of Chromelosporiopsis

carnea is by Persoon (1795) under the name Isaria carnea, with conidiophores

sticking together (“conferta’), at first white, then flesh-colored (“primo albida

dein carnea’) and evanescent (“evanescens”), illustrated in Persoon (1796:

Observ. Mycol. 1, tab. 2 figs 6-7) by fine color paintings (Fic. 15A). The

692 ... Hennebert

Fig. 14. Authentic specimens of Isaria carnea Pers. A-C. Isaria carnea [written by Persoon].

A. Isaria carnea [written by Persoon]. A. Three pieces of Fagus leaves. B. Enlarged immature

colony. C. Enlarged collapsed synnemata. D-E Isaria carnea var? [written by Persoon].

D. Patch of moss. E-F. Enlargements of a small patch of the fungus showing young white isarioid

synnemata. (Photos J. Nuytinck and Dr M. Scherrenberg, Naturalis Museum, Leiden).

authentic specimens of Isaria carnea in Persoon's herbarium, signed by Persoon

but with no date, are here documented by the excellent photographs made

by Dr Nuytinck and M. Scherrenberg in Leiden (Fic. 14). Schweinitz (1832)

showed that Isaria carnea has fascicles of hyphae (synnemata) diverging from

a shared base, pale brown, apically floccose and bearing abundant fleshy-red

spores (‘sporulis carneo-rubris’) spread along the hyphae. Isaria carnea was

treated by Fries (1832a,b) as a synonym of Dactylium macrosporum Fr., nom.

sanct. [= Botrytis macrospora Ditmar, nom. illeg.], but it is really a different

fungus, and and its epithet is therefore potentially available for use.

However the epithet is not truly available, as Fries (1832a) accepted and

sanctioned the species under the name Botrytis carnea based on Schumacher’s

description and his examination of Ehrenberg’s specimens of Polyactis carnea.

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 693

Botrytis carnea is described as “cespitosa, stipitibus brevissimis, dichotomis,

ramosissimis, albidis, sporulis congestis spicatis carneis; in lingo putrido

Betulae albae, Septembr,’ while Polyactis carnea shows “floccis suberectis

dichotomis capitatis albis, sporidiolis primum albis, dein carneis, globosis. In

foliis pinorum. Pluvio freq. In fossis. Septembr.’ As neither Schumacher nor

Ehrenberg cited Persoon, they are two distinct names for the same species,

not obligate synonyms. As the name Botrytis carnea is sanctioned it must

serve as the basionym for the species as understood here.

No original material of Botrytis carnea is available, but three good

specimens collected by Ehrenberg and preserved in STR and L under the

name Polyactis carnea contain synnemata, bifurcate branching hyphae, and

verrucose conidia.

Campsotrichum splendidum Schwein. n° 2695 is described with synnemata

(“coalitis floccis”), furcate branches emerging along the upper half, and

bearing reddish orange verrucose conidia (“pulcherrime decorate”). This is a

possible synonym of C. carneum.

accarde | Fungi alates

Huphomycetene

th bey li CQ ebiiaeas

Costa, /Hettorie) on Rene

Sut. WPL.

788. Jon 1881 683. Jan. 1881

Fig. 15. A. Persoon’s illustration of Isaria carnea in Observ. Mycol. 1: tab. II, fig. 6-7, 1796.

B-C. Saccardo Fungi Autografice Delineati (1881), fig. 688: Botrytis carnea Schum. and fig. 689:

Botrytis epigaea var. rosea Sacc.

Saccardo (1881) illustrated Botrytis carnea (Fic. 15B) as mononematous

and dichotomous with verruculose, rose-isabella conidia without indication

of synnemata. However, he drew synnemata and asymmetric bifurcations

in Botrytis epigaea var. rosea, another synnematous Chromelosporium-like

taxon (Fic. 15C). Saccardo (1886) also described B. carnea var. quercina

with smaller conidia, 4.5—5 um, asperulate and rose-colored. Both of these

varieties show characters of C. coerulescens.

694 ... Hennebert

SPECIMENS EXAMINED.

AUTHENTIC SPECIMENS: EUROPE: (5), Isaria carnea [scr. Persoon] on leaves of

Fagus leaves, Herb. Persoon (L 910-258-645). (6) Isaria carnea var? [scr. Persoon]

on a patch of moss. Herb. Persoon (L 910-258-663) [the specimen bears immature

smooth conidia]. GERMANY, (7) Botrytis rosea Link, on small pieces of leaves of Fagus

sylvatica, Rostock. Herb. Link (B) [DAOM 51764, MUCL2451]. UK, (8) Rhinotrichum

decolorans Forden, on cupules of Fagus sylvatica, Forden 312. Herb. M.C. Cooke 1885

(K-M) [MUCL 3495].

OTHER SPECIMENS: EUROPE, UK (9) Phymatotrichum, on bare soil and dead

leaves, Ashridge, Herts., England, July 17 1955, D.A. Reid (K-M) [MUCL 2304]. (10)

Phymatotrichum sp. on soil and vegetable debris, Windsor Great Park, Windsor, Oct.

3 1962, D.A. Reid & R. McNabb (K-M) [MUCL 3133, MUCL 3494]. (11) Botrytis

carnea, on Epilobium hirsutum, Witham Park, Oxfordshire, Sept. 17 1949, M.B. &

J.PEllis (K-M-—IMI60211) [MUCL 3517]. (12) Botrytis aff. splendida Schw., on mossy

Salix bark, Wheatfen Broad, Nrf., M.B. Ellis (K-M-IMI 26983) [MUCL 3568]. (13)

Phymatotrichum sp. on petioles of Petasites ovatis, Masham, Yorks., Oct. 13 1947, S.J.

Hughes (K-M-IMI 19231b) [MUCL 3572]. (14) Phymatotrichum, bark of Quercus,

Kinclaven, Perthshire, Sept. 1953, M.B. Ellis (K-M-—IMI63926) [MUCL 3505]. (15

Phymatotrichum sp., on bark and earth ina deep hole, Ashridge, Hertford. July 17 1956,

D.A. Reid, det. M.B. Ellis (K-M-—IMI 60584) [MUCL 3520]. (16) Phymatotrichum

sp., on rotten wood, Ruislip Woods, Oct.14 1955, C. Booth, det. A.H.S.. Brown.

Herb.(K-M-IMI 61356) [MUCL 3523]. (17) Phymatotrichum, on moss on beach,

Burnham, Beeches, Oct. 20 1956, A.H.S. Brown. (K-M-IMI 69690) [MUCL 3541].

(18) Phymatotrichum., oak and hazel leaf litter, Park Wood, Ruislip, Oct. 15 1949,

P.K.C. Austwick (K-M-IMI 37957) [MUCL 3559]. (19) Chromelosporium carneum,

decaying Quercus wood, Pett near Hastings, Sussex, Aug. 29 1965, P.C. Holland,

det. W. Gams (MUCL 7929). (20) Chromelosporium carneum, on stem debris,

Ashdown Forest, Forest Row, Sussex, Oct. 1 1967, D. Mitchell & P.C. Holland (MUCL

11290). NETHERLANDS (21) Botrytis fulva Link ex Fr. on herbaceous stems under

Fagus sylvatica. Bussum, Oct. 24 1929, WJ. Liittjeharms (L) [DAOM 83937, MUCL

2515]. AUSTRIA (22) Botrytis carnea Schum. ad folia et cortices putridos, sylvis

umbrosis, autumno, L. Fuckel.. Fuckel Fungi. Rhen. Exs. 146 (G, K-M, GRO, NY)

[DAOM 74694, MUCL 1808]. DENMARK (23) Phymatotrichum, on leaves, wood

and cupules of Fagus sylvatica, Gris Skov, Sealand, Oct. 2 1955, S.J. Hughes (DAOM

51702). (24) Phymatotrichum, on wood chips of Fagus sylvatica, Gris Skov, Sealand,

Oct. 2 1955, S.J. Hughes (DAOM 51690). PoLAND:(25) Botrytis carnea Schum., in

trunco Juniperi, Eichler, in herb. Bresadola 184 (S) [DAOM 83948, MUCL 2532].

(26) Chromelosporium carneum, on rotten wood, Biatowiesga forest, Pologne, Sept.5

1966, W. Gams (MUCL 9298). (27) Chromelosporium carneum, on rotten wood,

Augustow Reserve, Starozym, Pologne, Sept.3 1966, W. Gams (MUCL 9299). FRANCE

(28) Botrytis fulva Link écorce de Quercus et Corylus, Lyon, Sept. 1879, J. Therry. C.

Roumeguere Fungi Gall. Exs. (BR) [DAOM74695, MUCL 1809]. (29) Botrytis carnea.

feuilles de chéne [Quercus] pourrissantes, Tassin, Lyon. Sept. 1880, Cryptogames du

Lyonnais, J.J. Therry. 5092 (PAD) [MUCL 1870]. (30) Phymatotrichum, on mossy

bark of Quercus, Forét de Jupilles, France, Sept.15 1952, E.M. Wakefield, det. M.B.

Ellis (K-M-—IMI 50643) [MUCL 3598]. BELG1um (31) Botrytis rosea Link, in foliis

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 695

Quercus robur, coll. Libert (BR) [DAOM 74693, MUCL 1806]. (32) Chromelosporium

carneum, on dead leaves of Fagus sylvatica and Quercus pedunculata in woods, Parc

d'Arenberg, Hevelee, Sept. 1 1960, G.L. Hennebert (MUCL 1218, DAOM 74698). (33)

Chromelosporium carneum, on twig of Fagus sylvatica, Parc d’Arenberg, Hevelee, Aott

21 1964, G.L. Hennebert (MUCL 6278). (34) Chromelosporium carneum, on bare

humid soil under Fagus sylvatica, in Lauzelle forest, Louvain-la-Neuve, Oct. 2017,

G.L. Hennebert (MUCL 56651).

Chromelosporiopsis coerulescens (Bonord.) Hennebert, comb. nov. _— Fics 16-18

MB 835625

= Polyactis coerulescens Bonord., in Fresenius, Beitr. Mykol. 1: 14, 1850.

= Botrytis coerulescens (Bonord.) Sacc., Syll. Fung. 4: 132, 1886.

= Chromelosporium coerulescens (Bonord.) Hennebert, Persoonia 7:197, 1873.

= Hyphelia purpureospadicea Fuckel, Jahrb. Nassauischen. Vereins Naturk. 23-24: 363, 1870.

= Botrytis purpureospadicea (Fuckel) Sacc., Syll. Fung. 4: 121, 1886.

= Botrytis epigaea subsp. rosea Sacc., Michelia 2(8): 544, 1882.

= Botrytis epigaea var. rosea (Sacc.) Sacc., Syll. Fung. 4: 136, 1886.

= Botrytis carnea var. quercina Sacc. Syll. Fung. 4: 119, 1886.

Types: C.[Chromelosporium] rhodianthinum n.sp.-[prov.name] on rotting leaves and

humic debris of Acer saccharum, Betula lutea and Tsuga canadensis in mixed woods,

Bell’s Corners, Ontario, Canada, 18.07.1961, leg. G.L. Hennebert (Neotype: DAOM

83371, designated in Hennebert 1973; Isoneotype: MUCL 2323).

COLONIES in tufts, appearing and rapidly evanescent, with <20 synnemata

arising from a basal mat of hyphae, at first white to bluish, sky-blue turning

to rose-lilac, finally vinaceous ocher at maturity in fresh conditions and

vinaceous brown when dried.

HypHAE superficial, sparse, delicate, hyaline, thin-walled, smooth,

forming mats 1-5 mm wide, 1 mm high, hyphae interwoven 11-29 um wide,

septate into 32—46 um long cells.

SYNNEMATA erect, divergent, composed of <25 hyphae, septate and

anastomosing, the central hyphae growing first <800 um high, the outermost

ones shorter, each hyphae serving as a conidiophore stipe.

CONIDIOPHORES stipes hyaline, 6—9 um wide, septate, with 36—54 um long

cells, thin-walled, smooth, all branching at the top, in successive, asymmetric

dichotomies in a coralloid pattern, forming a globose or somewhat conical

conidial head.

CONIDIOGENOUS HEAD comprising all branches of the conidiophore except

the first ones, 15—75 (mostly 30-60 um) um long, 9-12 um wide, the distal

cells clavate <15 um wide, obtuse, forming conidia almost simultaneously.

ConipiA holoblastic, on denticle 1 x 0.5 um, 4—9 um spaced, one celled,

globose, 4.2—6.6 um, most 5.4 um, walls 1 um thick, subhyaline, the outer

696 ... Hennebert

Fig. 16. Chromelosporiopsis coerulescens (neotype DAOM 83371).

A. Synnemata B. Asymmetric dichotomies or coralloid branching. C. Conidiation.

D. Finely verrucose conidia. Scale bar = 10 um.

wall finely punctate (‘minutissime asperulis, Saccardo 1886 p. 132), with

12-18 small warts in median view, seceding rhexolytically.

HABITAT: on mosses, rotten wood debris, tree leaves, conifer needles in

humid forest.

DISTRIBUTION: North America: Canada, USA, Europe: France, Belgium.

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 697

Fig. 17. Chromelosporiopsis coerulescens (neotype DAOM 83371). A. Fungus mat with

emerging young synnemata. B. Top of growing synnema. C. Head of branches. D. Asymmetric

dichotomous branching. E. Conidial head.

COMMENTS—Fresenius (1852: 74), who received a specimen and a description

of the species from Bonorden, noticed that “die sporen sind rund, feinwarzig,

nicht glatt wie Bonorden angibt”. Indeed Bonorden (1851) described the spores

as “glatt”. Fresenius also illustrated the conidiogenesis of the species.

Chromelosporiopsis coerulescens differs mainly from C. carnea by the

colour when young, the synnematal development, the narrower conidiophore

hyphae, and the smaller, punctate conidia.

Bonorden (1851) described Polyactis coerulescens as “zuerst weisse, dann

hellblau, zuletzt duch die Sporen braunlich” The pale blue (hellblau) young

698 ... Hennebert

Fig. 18. Chromelosporiopsis coerulescens. A. Young blue cushions and white young synnemata.

B. Rose-lilac nearly mature synnemata (specimens collected and photographed on rotting

trunk on the ground, A. 14.06.2014, Sainte-Cécile-de-Masham, QC and B. 19.08.2018, Orleans,

Ont, Canada, by Jonathan Mack, Mycoquebec.org).

stage of the colonies changes during maturation to rose-lilac-mauve and

then vinaceous ochre-brown.

Fuckel (1870) described under Hyphelia the new species H. purpureo-

spadicea. It is here a synonym of C. coerulescens after examination of the

holotype.

Labbé (2015) describes the species as “remarquable par ses colorations

bleu cristal puis rose-violet et ressemblant presque a des cristaux en

forme daiguilles.” The species is illustrated in its change of colour in the

photographs by Jonathan Mack (Fig. 16) in J. Landry on mycoquebec.org

and in photographs by Cornell Mycology (Hodge 2017).

SPECIMENS EXAMINED

TYPE & AUTHENTIC SPECIMENS EUROPE: GERMANY (2) Hyphelia purpureospadicea

Fuckel, ad terram arenosam humidam in sylvis acerosis, raro, Autumno, circa

Budenheim, Fuckel. Fungi Rhen. Exs. 2214 (G, holotype; BR, GRO, PAV, FH, isotypes)

[DAOM 83395, MUCL 2396].

OTHER SPECIMENS. NORTH AMERICA: CaAnapa (3) Botrytis ?, on humus,

Morgan's woods, MacDonald College, near Montreal, Qué., Aug. 27 1941, R.E Cain

12979 (DAOM 80136) [MUCL 2276]. USA (4) Botrytis dichotoma, on ground and

rotting wood, Fall Creek, Ithaca, N.Y., June 18 1894, G.E Atkinson. Herb. Atkinson

1153 (CUP) [DAOM 84693, MUCL 2876]. (5) Botrytis terrestris, on humic ground

under conifers, Shelburne, New Hampshire, Sept. 1891, W.G. Farlow (FH) [DAOM

84699, MUCL 2883]. (6) Botrytis terrestris, on dead leaves, trunks, dungs, etc. in

conifer woods, Shelburne, W.G. Farlow (FH) [DAOM 84700, MUCL 2884]. (7) Botrytis

epigaea, on rotten wood and mosses, Hadley Lake, Machias, Sept. 21 1898, W.G.

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 699

Farlow 36 (FH) [DAOM 84698, MUCL 2882]. (8) Botrytis epigaea Link, on mosses,

Magnolia, Mass. July, W.G. Farlow (FH, UPS, CUP) [DAOM 84912, MUCL2484].

(9) Botrytis epigaea, on Hydnum, Gerrish Island, Kittery Pt., Maine, R. Thaxter (FH)

[MUCL 2892]. (10) Botrytis (Polyactis) coerulescens (Bon.) Sacc., on bare soil in forest,

Lingle Valley, Mifflin Co., Pa., July 3 1938, L.O. Overholts 21176, D.H. Linder (FH)

[MUCL 2891]. (11) Hyphelia terrestris, on decaying leaves, Ringwoods, N.Y., Sept. 6

1952, R.E Cain (TRT 24143) [MUCL 2863]. (12) Ostracoderma, on Quercus rotten

wood, Pennsylvania state, July 4 1957, E.A. Atwell n° 5779 (DAOM 59233). (14)

Ostracoderma, on lawn, College Park, Maryland, Sept. 15 1961, A.M. Golden, C.R.

Benjamin (BPI) [MUCL 2957].

EUROPE: Beteium (15) Hyphomycete n° 517, sur sol de jardin, Rue Berchmans,

Bruxelles, June 21 1917, M. P. C. Beeli (BR). (16) Botrytis violacea/lilacina Schw, sur

argile, Forét de Soignes, Sept. 23 1933, Bommer (BR).

Species of Pezizaceae with known sexual morphs and similar synnematous

conidiophores and conidiogenesis

A similar Chromelosporiopsis asexual morphology was found in

specimens collected by Korf (1994) and identified by Healy & al. (2015)

as Pachyphlodes pfisteri Tocci & al. (Pezizaceae), and briefly described by

Hennebert (2017).

In October 2017, some samples of synnematous Chromelosporiopsis

conidial mats were collected on bare soil in forests around Louvain-la-

Neuve in Belgium. The DNA sequencing in MUCL identified three samples

as Pachyphlodes nemoralis Hobart & al., and one sample as P. citrina (Berk.

& Broome) Doweld. Their conidial morphs are clearly distinguished by the

synnematal structure and hyphal width, the hyphal length and width of the

conidiophore coralloid branches, and particularly by the conidial size and

the conidial wall ornamentation (Hennebert & Decock 2020).

Pachyphlodes nemoralis has globose conidia, 4—6.5 um wide, with 0.5 um

thick walls covered with <0.3 um high tuberculate warts (12-14 in median

view).

Pachyphlodes citrina has globose conidia, 4.5—7 um wide, with 0.5 um

thick walls covered with <0.8 um high baculate warts (12-14 in median

view).

3. Doubtful and excluded taxa

Most synnematous Chromemosporium-like herbarium specimens

examined were labeled as Botrytis epigaea, Polyactis epigaea, Hyphelia

terrestris, or Phymatotrichum, among others. These names are considered

below.

700 ... Hennebert

Botrytis epigaea Link, Spec. Plant., ed. 4, 6(1): 52, 1824.

= Polyactis epigaea (Link) Bonord., Handb. Mykol.: 115, 1851.

= Phymatotrichum epigaeum (Link) Vasyag., in Shvartsman &

al., Flora Sporov.. Rast. Kazakhst. 8: 263, 1973.

No authentic material of Botrytis epigaea or Polyactis epigaea was retrieved

from relevant herbaria. Link (1824) originally described Botrytis epigaea as:

“Botrytis epigaea, thallo effuso, floccis sporodiferis subramosis brevibus,

sporidiis globosis minutis. Habitat in terra humida sylvarum Germaniae.

Lecta Berolini im Thiergarten (Lk.) (v.v.)” This nine-word description being

applicable to a number of fungi, the name is doubtful in absence of authentic

specimen. Fries (1832a,b) did not accept the species in Sporotrichum because

he considered it as only mycelium.

Bonorden (1851) interpreted Link’s species under Polyactis epigaea in his

Fig. 161 as a fungus showing bifurcate conidiophores covered with conidia,

adding that the colour of the mature mass of spores is gray or greyish yellow

(“grauen oder graugelben Pulver”). This might suggest a Chromelosporium-like

species, but no authentic material is extant.

The earliest specimen received as Botrytis epigaea was collected by Fuckel in

1861. This synnematous Chromelosporium-like fungus with punctate conidia

does not possess enough for identification.

Saccardo (1886) considered Botrytis epigaea similar to Hyphelia terrestris

sensu auct.

Phymatotrichum Bonord., Handb. Mykol.: 116 (1851).

Bonorden, who defined his genus with conidiophores branched tree

or shrub-like, not umbellate, bearing on their distal swollen branches

pecidellate conidia (“gestielte Sporen”), described three species:

P. gemellum, P. pyramidale, and P. laneum (Fie. 19).

While P pyramidale Bonord. was transferred to Botryosporium,

Saccardo interpreted P laneum as Botrytis laneus (Bonord.) Sacc. and

P. gemellum as B. gemella (Bonord.) Sacc.; Hennebert (1973), who

interpreted P gemellum as Botrytis cinerea Pers., deduced that

Phymatotrichum was a synonym of Botrytis.

But none of the researchers considered the pedicellate conidia (“gestielte

Sporen”) mentioned by Bonorden as a diagnostic generic criterion, which

is neither a character of Botrytis nor of Phymatotrichopsis (Fic. 20).

Phymatotrichum gemellum is described with some fasciculate superficial

hyphae (“verbundenen hypha”) bearing short tufts ending in unequal

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 70.1

182

138

000

Fhymatotrichum gemellurm Frymatotrichum pyramidale

Fiymato trichum laneum

Fig. 19. Copy of the Bonordens illustrations of the species of Phymatotrichum.

bifurcate apical swollen fertile cells (“Endaste erweitern zich zu zwei

ungleich grossen Basidien”), characters that possibly suggest species of

Chromelosporiopsis. But Bonorden commented that the species possessed

characters of both Polyactis and Botrytis, but absent any authentic material,

the genus remains doubtful.

Fig. 20. Conidiogenesis of Phymatotrichopsis omnivora (Shear) Hennebert from dried sporemat

on soil in cotton field, Paris, Texas, Sept. 18, 1915, BMD, received from Missouri Bot. Garden

June 1916 (FH) [MUCL 2868] (Marek et al. 2009).

702 ... Hennebert

Phymatotrichum silvicola Taubenh. & G.M. Watkins,

Amer. J. Bot. 24: 390, 1937 [as “silvicolum”].

SPECIMENS EXAMINED: (1) Phymatotrichum silvicolum Taubenhaus and Watkins, on post

oak wood on sandy soil near Bryan, Brazos Co, Texas, November 1936 (NY) [DAOM

84656, MUCL 2690]. (2) Phymatotrichum silvicolum Taubenhaus and Watkins, on post

oak wood on sandy soil near Bryan, Brazos Co, Texas, Sept.8, 1937 (BPI) [DAOM 84657,

MUCL 2691].

Two authentic specimens of Phymatotrichum silvicola were received. The

fungus is described from Texas on sandy soil in forest, as “hyphal mats

more or less circular, 0.5-4 cm wide, white becoming greyish yellow, with

synnemata of conidiophores arising from them, with bifurcate branching,

the apical branches clavate, 5-8 um wide and covered with subglobose one-

celled conidia 2-4.6 um wide on denticles”. The specimens are ambiguous,

containing globose (3.5-5.2 um) punctate conidia mixed with napiform to

globose (6.5-8.7 um) verrucose conidia.

Rhinotrichum thwaitesii Berk. & Broome, Ann. Mag. Nat. Hist., ser. 2, 7: 177, 1851.

SPECIMEN: Rhinotrichum thwaitesii Berk. & Broome, Clifton, Berkeley. Herb. Berk. (K-

M), holotype, with included watercolour plate by Berkeley [MUCL 2952].

The specimen contains conidiophores branched in dichotomies bearing

globose conidia on denticules with very verrucose walls, but no synnemata

have been seen. Rhinotrichum thwaitesii remains a doutful species in view of the

discrepancies between the fungus and Berkeley's accompanying watercolour

plate and the published diagnosis of the name with ovoid and larger spores.

Isaria thyrsoidea Penz. & Sacc., Malpighia 15: 251, 1902.

Isaria thyrsoidea, found on rotting leaves,

insects and dung, in Tjibodas, Java, 1897, is : ; |

described as white tufts of erect cylindrical |. \ is alt ZG .- BE

synnemata 1.5 mm high, 40-50 um wide, made rAd

of septate hyphae that emerge laterally and

apically from the upper half of the synnemata

as bi- and tri-furcated conidiogenous branches,

saat ae

5.5—6 um wide, sparsely septate and covered

on denticle (“subsessile”) with smooth globose

conidia 3.5-4 um wide (Fic. 21), The fungus

is turns ash-grey at maturity. Are the conidia

really smooth? Despite the colour of the Fig. 21. Isaria thyrsoidea Penzig &

Sacc., Malpighia 15: 251, 1902

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 703

fungus and the smooth conidia, the description suggests a Chromelosporiopsis

species. The original material needs to be located and re-studied.

Chromelosporium terrestre (Fr.) M.B. Ellis, More Demat. Hyphom.: 154, 1976.

= Hyphelia terrestris Fr., Syst. Mycol. 3(1): 213, 1829, nom. sanct.

Ellis (1976) proposed the combination Chromelosporium terrestre for

the basionym H. terrestris in order to correct the then illegitimate name

Chromelosporium tuberculatum (Hennebert 1973). But Hyphelia terrestris

is not a Chromelosporium-like species as shown below. The fungus

described and illustrated by Ellis under that name is an unidentified species

of Chromelosporiopsis and belongs to the set of collections considered

hereunder.

Unidentified herbarium material with features of Chromelosporiopsis

Beside the presence of synnemata, a generic character, the specific

microscopic characters of Chromelosporiopsis (observed when possible in the

following material) are the hyphal widths (diameters), the branching patterns,

the condial size, and the ornamentation of the conidial wall. The conidial

wall ornamentation serves as an important criterion distinguishing between

asexual morphs of Pachyphlodes species (Hennebert & Decock 2020) and is

used here as a basis for a preliminary grouping of the specimens. But a good

characterization of the wall ornamentation requires high quality equipment

without which the present groups are based only on the number of warts

visible on median view of the conidia and not on the wall morphologies. The

specimens are classified according to the conidial wall surfaces ranging from

punctate to coarsely warty. To be diagnostic, the number of median warts

must be combined with wart morphology.

SPECIMENS EXAMINED

CONIDIA PUNCTATE (218 WARTS, MEDIAN VIEW)

AUTHENTIC SPECIMEN EUROPE: (3) Trichoderma laeve ? Pers., frequens in

sylvulis prope Parisios [scr. Pers.] Herb. Pers. (L 910.267.31) The fungus is immature

and fragmentary. [DAOM 83899, MUCL 2461].

OTHER SPECIMENS. EUROPE: GERMANY (4) Botrytis epigaea Link, Polyactis

epigaea Bon., Giesen, Oct. 22.1861, Herb. Fuckel in herb. Barbey-Boissier (G 005439)

MUCL 2400)]. (5) Botrytis epigaea Link, ad terram Bavaria, Killermann. Herb.

Bresadola (BPI) [DAOM 83891, MUCL 2428]. SWEDEN (6) Hyphelia terrestris, ad

terram nudam, Stollsbachen, Uppsala, Aug. 7 1932, Seth Lundell. Flora Suecica (UPS)

[DAOM 83921, MUCL 2494]. (7) Hyphelia terrestris, at the lower course of the rivulet

Skytebacken, Halsungland, Farila parish, Skyte, Aug. 2 1956, J.A. Nannfeldt. Flora

Suecica 14770 (UPS) [DAOM83905, MUCL 2770]. (8) Hyphelia terrestris, Zogenern

Wald, b. Liindensitz, Aug. 1844 (PR 181907) [DAOM 84707, MUCL 2907]. UK (9)

704 ... Hennebert

Ostracoderma, on wet soil, Wotten under Edge, Gloucestershire, Aug. 1961, R.W.G.

Dennis (K-M-IMI 89255) [DAOM 83951, MUCL 2541]. (10) Botrytis splendida

Schw., on soil, Skircoat Green, Halifax, Yorkshire, Nov. 1955, R.Watling, det. M.B.Ellis

(K(-M-IMI 61440) [MUCL 3524]. FINLAND (11) Hyphelia terrestris = Botrytis epigaea

Link var. rosea Sacc. on soil, Mustiala, Aug. 25 1869, P.A. Karsten, W Nylanders,

Herb. P.A. Karsten (H) [DAOM 83903, MUCL 2470]. (12) Hyphelia terrestris, supra

terram, Mustiala, Aug. 1865, P.A.Karsten. Flora Fennica (UPS) [DAOM 83913,

MUCL 2485]. NETHERLANDS (13) Ostracoderma (? Tomentella granulata Bref.), op

naakte bodem in loofbos lenig humensegrond, Ulvenhout, Ulvenhoutse bos, Sept. 29

1959, R.A. Maas Geesteranus 13006 (L) [DAOM 83935, MUCL 2513]. PoRTUGAL

(14) Botrytis carnea Schum., ad terram, Portugal, Torrend 21, Herb. Bresadola (S)

[DAOM 83946, MUCL 2528]. BELGruM (15) Hyphelia, on bare soil, Forét de Soignes,

Tervueren, Aug. 16 1960, G.L. Hennebert (MUCL 1214) [DAOM 83904]. DENMARK

(16) Ostracoderma, ad terram in sylva faginea, Sjaelland, Sonnerup sogn., Ovdrup

skov, Sept. 13 1952, Mykologisk Kongres, J.A. Nannfeldt. Flora Danica 12552 (UPS)

[DAOM 83915, MUCL 2488].

NORTH AMERICA: CANADA (17) Ostracoderma, on garden soil, in gras,

Ottawa, Oct.15 1960, V.J. (MUCL 1534). (18) Botrytis epigaea, on soil in barley field,

Glen Williams, Halton Co., Ont., Sept. 9 1956, R.F. Cain (TRT 32547) [DAOM 84686,

MUCL 2860]. USA (19) Rhinotrichum thwaitesii B. & Br., on ground in woods, south

side of Fall Creek, Ithaca, June 25 1959, G.F. Atkinson, Herb Atkinson 22860 (CUP)

[DAOM 84695, MUCL 2878]. (20) Hyphelia terrestris on ground, Nashville, York Co,

Ont. Sept. 29 1956, R.E Cain (TRT 32443) [DAOM 84685, MUCL 2859]. USA (21)

Botrytis isabellina Pr., on bark of Pinus sylvestris, Juamby Lawn, Boston, Nov. 1906,

H.C. Hawley (BM) [MUCL 2280]. (22) Hyphelia terrestris, on damp ground in woods,

Bois Mallet, West La. May 23 1886, A.B. Langlois 412 (BPI) [DAOM 83892, MUCL

2429]. (23) Hyphelia terrestris, on sandy soil in field, Nashville, York. Co. Ont. Oct. 9

1955, R.E Cain, Herb. RFC 31601 (WU) [MUCL 2408]. (24) Botrytis epigaea Link,

on moist humus and soil, Altamonte, Florida, Aug. 1 1957, P.O. Schalbert, P.L. Lentz

(BPI) [MUCL 2960].

AUSTRALASIA: AusTRALIA (25) Phymatotrichum, on dead leaves, twigs and

woody fruits, Brisbane, Australia, June 1951, R.E Langdon 817 (K-M-IMI 54841)

[MUCL 3504]. NEw ZEALAND (26) Botrytis terrestris, snow white patch on soil, New

Zealand, W. Odenso. Herb. J.B. Ellis (NY) [DAOM 84680, MUCL 2851].

CONIDIA VERRUCOSE (12-16 WARTS, MEDIAN VIEW)

EUROPE: Austria (1) Hyphelia terrestris var. flava, ad terram humidam in

dumestis, non raro, Autumno, Oestrich, Fuckel. Fuckel Fungi Rhen. Exs. Supp.

1641 (Symb. Mycol.: 363 1869) (G, GRO, BM, FH) [DAOM 83327, MUCL 2258].

Irauy: (2) Botrytis epigaea Link. Padova, ad terram udam umbrosam, December.

1874. Saccardo P.A. Mycotheca veneta [Cent. IV, 1876] 360. [MUCL 2283] (3)

Botrytis epigaea Link, a) ad terram nudam in Horto Botanica Ticinensi, b) ad terram

herbosam sub cupuliferis, ibidem. Ipse lege. Autumno. Forma b rubella ad var.

roseam Sacc. ferendo mihi videtur, quamvis conidia paulum majora sint. Cavara F.

Fungi Longobard. Exs. 145 (B, K-M, S, L 910.224-184) [DAOM 83335, MUCL 1805

= 2285]. (3) (4) Botrytis epigaea Link, ad terram, Florentiae, June 1891, N. Martely,

Herb. Bresadola (S) [DAOM 83939, MUCL 2516]. GERMANY (5) Hyphelia terrestris

ad vias nemorum, Grossen Garten, prope Dresden. Rabenh. Klotzschii Herb. Viv.

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 705

Mycol. I, 1846 (PR, PAV, BR) [DAOM 83895, MUCL 2432]. (6) Hyphelia terrestris,

in parnecio Tyrois, Aug. 29 1859, PA. Karsten (H) [DAOM 83902, MUCL 2469].

SWEDEN (7) Hyphelia pulvinata (Fr.) Juel, pa sandjord utmed en vag I baarskog,

Uppland, Uppsala, Stadsparken nara Norby, Oct. 1933, Seth Lundell. Flora Suecica

1068 (UPS) [DAOM 83910, MUCL 2482]. (8) Hyphelia pulvinata (Fr.) Juel, pa

naken jord I vagkant, Upplannd, Estuna, Ljushammaren, Malmé, Sept. 27 1955,

Seth Lundell. Fungi Suecici (UP) [DAOM 83917, MUCL 2490]. (9) sine nomine, auf

nackter Erde, in Park, Uppland, par. Solna, Aug. 1906, L. Romell, Herb. Mykol. Lars.

Romell 17715 (S) [DAOM 83944, MUCL 2523]. (10) Polyactis epigaea (Link) Bon,

?Hyphelia, auf nackter Erde auf einen Pfaden Upl. Stockholm: Ekbacken, Jul. 22 1894,

L. Romell. Herb. L Romell (S) [DAOM 83945, MUCL 2526]. NETHERLANDS (11)

Ostracoderma, op de gronde in loofbos, Ulvenhout, Ulvenhouter bos, Oct. 6 1959,

R.A. Maasgesteranus 13025 (L) [DAOM 83936, MUCL 2514]. FRANCE: (12) Hyphelia

terrestris, in umbrosis viis, Montmorency, St Germain, Fontainebleau Sept. 1842-

1848, Roussel, Herb. E Roussel (PC) [DAOM 84661, MUCL 2703]. BELGrum (13)

Chromelosporium, sur le sol dun chemin en forét, Bois de Hevelee, Heverlee, Brab.,

G.L. Hennebert (MUCL 6701). (14) Hyphelia?, sur soil humide, au bord dun chemin

ombragé, Forét de Soignes, Groenendael, Aug. 16 1960, G.L.H. (MUCL 1205, DAOM

74900).

NORTH AMERICA: CANADA: ONTARIO (15) Ostracoderma, Bells Corners,

Ont. Sept. 10 1961, G.L. Hennebert (DAOM 83922, MUCL 2496). USA: VIRGINIA

(16) Botrytis epigaea Lk. var. rosea Sacc., on dead leaves, Limberlost, Shenandoah

National Park, Va., Sept. 9 1937, J.A. Stevenson and V.K. Charles (BPI) [MUCL 2409].

(17) Botrytis epigaea Link, on soil under leaves, Arlington cementary, Virginia, May

22 1932, C.L. Shear (BPI) [DAOM 83870, MUCL 2427]. CONNECTICUT (18) Botrytis

fulva Link, near B. epigaea, on soil and grass, New Haven, Conn. July 6 1889, R.

Thaxter, Herb. Atkinson (CUP) [DAOM 84696, MUCL 2879]. MASSACHUSETTS (19)

Botrytis spectabilis, on rotten wood, Prospect Hill, Waltham, Oct. 1901, WG. Farlow

(FH) [DAOM 84701, MUCL 2885]. (20) Botrytis epigaea, near Hyphelia terrestris, on

rotten wood in humic litter, Bedford, Mass., Sept. 1901, W.G. Farlow (FH) [DAOM

84697, MUCL 2881]. NEw Mexico (21) Rhinotrichum roseum, on buried dead

leaves, Chirlehunt, Sept. 1854. Herb. Currey (K-M) [MUCL 2300].

CONIDIA VERY VERRUCOSE (6-12 WARTS, MEDIAN VIEW)

EUROPE: SwEDEN (2) Hyphelia terrestris, on soil, Kronoparken, Uppsala,

Uppland, Aug. 1916, H.O. Juel. Flora Suecica (UPS) [DAOM 83911, MUCL 2483].

(3) Hyphelia pulvinata (Fr.) Juel, pa naken jord I strandsnaret, Upland, Naturpark

Bondkyrka , Vardsiatra, Jul. 18 1930, Set Lundell. Flora Suecica 0798 (UPS) [DAOM

83909, MUCL 2481]. (4) Hyphelia, ad terram nudam sub Corylus, dland, Persmas

parish, Legends, Aug. 3 1953, J.A. Nannfeldt. Flora Suecica 13327 (UPS) [DAOM

83906, MUCL 2477]. DENMARK (5) Ostracoderma, ad terrram in fageto, Sjealland,

Hvaloo skov, Sept. 14 1952, Mykologisk Kongress, J.A. Nannfeldt. Flora Danica 12603

(UPS) [DAOM 83919, MUCL 2492]. UK (6) Ostracoderma pulvinatum Fr. Sibbertoft,

1873, Berkeley, Herb. Berk (K-M) (conidia with prominent blunt warts, drawn by

Berk.) [MUCL 3497]. (7) Phymatotrichum, on burn ground and wood, Ashridge,

Herts., England, Oct. 9 1955, D.A. Reid (K-M) [MUCL 2303]. Germany (8) Botrytis

epigaea Link., ad terram humidam, non frequens. Aestate. circa Schlangenbad. L.

Fuckel. Fuckel Fungi Rhen. Exs. 2301, 1871 (G, K-M, B, K, GRO) [MUCL 2282].

706 ... Hennebert

FRANCE (9) Hyphelia terrestris, ad vias in sylvis Fontainebleau, Aug. 4 1869, Roussel,

Herb. E. Russel (PC) [DAOM 84662, MUCL 2704]. BELG1um (10) Chromelosporium,

sur le sol en forét, Bois de Bonsecours, Blaton, Hainaut, G.L. Hennebert (MUCL

6332).

CONIDIA SMALL, COARSELY VERRUCOSE (4 —6 WARTS, MEDIAN VIEW)

EUROPE. SweEpDEN (1) Hyphelia, on bare soil under stormfelled spruce,

Gastrikland, Hills in Tolfforskogen, near Tolffors, Aug. 15 1954, J.A. Nannfeldt. Flora

Suecica 11065a (UPS) [DAOM 83908, MUCL 2480].

NORTH AMERICA: CANADA: QUEBEC (2) Botrytis?, on humus, Morgan’s

woods, MacDonald College, near Montreal, Qué. Aug. 27 1941, R.E Cain 12979

(DAOM 80136) [MUCL 2276]. USA: NEw York (3) Phymatotrichum, on soil, Lloyd

Cornell Preserve, Slatterville, NY, Sept. 6 1952, W.W. Diehl (BPI) [DAOM 83889,

MUCL 2426].

Re-evaluation of Hyphelia terrestris

Identification of the majority of the examined herbarium specimens

found to be synnematous Chromelosporium-like species as Hyphelia

terrestris is based only on the short macroscopical description and habitat

information by Fries (1829): “effuse, strigoso-tomentosa, candida, medio

evanescens, sporidiis subargillaceis ... in terra humosa, umbrosa, humida

and denudata” and ignores Fries’s synonymy.

‘The signed and dated specimens among the specimens examined show

that this interpretation was accepted as early as in 1842 by Roussel, 1846 by

Rabenhorst, 1861 by Fuckel, 1865 by Karsten, 1886 by Saccardo, 1901 by

Farlow, 1920 by Juel, and by many afterwards. Taking into account Fries’s

protologue for Hyphelia terrestris, this traditional interpretation is actually a

misapplication of the name.

Juel (1920) observed that the macroscopical description of Hyphelia

terrestris in Fries (1829) matched well the collection he made near Uppsala

in 1916 and a similar collection made in Mustiala, Finland in 1865 by

P.A. Karsten, who identified it as Hyphelia terrestris (UPS). Convinced that

the observed conidiogenesis in these specimens had to be that of Hyphelia

terrestris, Juel emended the genus Hyphelia (for the part regarding Hyphelia

terrestris) adding “rami conidiophori, apice non inflati, conidiis rotundis

brevissime pedicellatis undique tecti” to the Fries description.

Hennebert (1973) accepted Juel’s interpretation of Hyphelia terrestris

as a Chromelosporium-like species and interpreted (erroneously) Juel’s

publication as a lectotypification of Hyphelia terrestris. Actually Juel did

not typify the name from any authentic Friesian material or from the two

specimens that he cited. Hennebert named the fungus Chromelosporium

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 707

tuberculatum (Pers.) Hennebert, Trichoderma tuberculatum being the first

synonym cited by Fries but for which no type exists. As Hyphelia terrestris had

been sanctioned (Fries 1829), the name was corrected to Chromelosporium

terrestre (Fr.) M.B. Ellis.

Fries (1825) originally introduced the generic name Hyphelia for

Trichoderma roseum Pers. [= Trichothecium Link, fide Hughes 1958]. Fries

(1825) also added “Aliud genus, mucedineum, representat vulgatissimum

Trichoderma tuberculatum Pers. cui accedunt multae species novae.... Sed de

his plura in S. M. II” [a reference to the forthcoming Systema Mycologicum

3]. “Huic generi Trichodermatis nomen tribuerem, nisi T: viridi jam a Cel.

Linkio affixum, quod sancte servandum videtur.”

However, instead of creating his proposed new genus (‘aliud genus’),

Fries (1829) actually divided Hyphelia into two unranked infrageneric

parts: Thelephoroideae (for H. rosea, H. spadicea, H. fusca, and H. nigrescens)

and Hyphomycetoidea (for the new species H. terrestris, described only

macroscopically and with four synonyms).

Fries (1849) reduced his concept of Hyphelia by omitting the type,

H. rosea (Pers.) Fr. and chose section names (instead of adjectives) for the

subdivisions: Xylohypha for H. nigrescens and H. fusca; and Geohypha for

H. terrestris. The omission of the type, H. rosea, made Hyphelia Fr. 1849 an

illegitimate later homonym of Hyphelia Fr. 1825, nom. sanct. The names of

the subdivisions Geohypha Fr. and Xylohypha Fr. are legitimate and available

for elevation to generic rank.

Fries (1829) described Hyphelia terrestris only macroscopically, with four

synonyms: “Trichoderma tuberculatum Pers.; T: nemorosum Pers.; T. laeve

Schum. (dubitans ipse citat T: laeve Pers.); and T. varium Ehrenb.”

Searching for the type material of these names, I have been informed

that no authentic specimen exists for Hyphelia terrestris in UPS or S, and

no specimen of Trichoderma tuberculatum remains in L. Specimens of

Trichoderma laeve and of Trichoderma nemorosum are preserved in L. and

of T. varium in B and STR.

The examination of these authenticated specimens, one of Trichoderma

laeve, (not the one labelled “Trichoderma laeve?” by Persoon), one of

T; nemorosum, and three of T. varium revealed that they represent one

species distinguished from Chromelosporium-like species by conidiogenesis.

Persoon (1796) describes Trichoderma tuberculatum, with ash grey

conidia (“pulvere cinereo”), like the grayish conidia (“clair-cendré”) of

T. nemorosum, suggesting that T: tuberculatum was similar to T: nemorosum.

708 ... Hennebert

Persoon also described the conidia of T: laeve as yellow, as Schumacher

described his T: laeve. Ehrenberg described conidia of T: varium as variable

in colour, while Fries described them as “subargillaceis”.

The other specimen, labelled “Trichoderma laeve ?’ by Persoon, contains

pieces of young conidiogenous cells of a Chromelosporium-like fungus,

differing from his other labelled ‘Trichoderma laeve,’ thus explaining

Persoon’s question mark. Persoon’s doubt affected Fries (1829), who

wrote “dubitans ipse citat T: aeve Pers: and cited instead “T: laeve Schum.

! Saell. 2 p. 236”, a later homonym, of which Fries had seen a specimen.

This illuminates the subsequent confusion between Hyphelia terrestris

and the Chromelosporium-like fungi. This indicates also that the Friesian

concept of Hyphelia terrestris differed from its traditional interpretation as a

Chromelosporium-like fungus.

The most accurate interpretation of the nomenclature is to respect Fries’s

1829 protologue of Hyphelia terrestris (a sanctioned name) and lectotypify

the species by one synonym, of which material exists— Trichoderma varium

Ehrenb.—rather than T: tuberculatum Pers. and T: laeve Schumach., which

lack extant material.

With the generic name Hyphelia 1849 being illegitimate as a later

homonym, Hyphelia terrestris therefore has status of type species

of Geophypha Fr. which can be raised in rank to a genus. A parallel

nomenclatural interpretation was made for Hyphelia nigrescens (Pers.) Fr.,

now regarded as the type species of the genus Xylohypha (Fr.) E.W. Mason

by Deighton (1960).

Geohypha (Fr.) Hennebert, stat. nov.

MB 835628

= Hyphelia sect. Geohypha Fr., Summa Veg. Scand. 2: 447, 1849.

TYPE SPECIES: Geohypha terrestris (Fr.) Hennebert

= Hyphelia [unranked] Hyphomycetoidea Fr., Syst. Mycol. 3(1): 213, 1829, nom. sanct.

Ascomycota, ascomata unknown.

Hypuae septate, hyaline, intricate, irregular, branched.

CONIDIOPHORES short as lateral outgrowths from vegetatve hyphae

bearing one conidium or long narrow sinuous and self-ramified branch,

sparsely septate, bearing a conidium on each of the many lateral outgrowths

and terminal ends.

CONIDIOGENESIS thallic, solitary on hyphal outgrowths, secession

schizolytic.

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 709

Conrpi1A one-celled, globose or subglobose, sometimes napiform, hyaline,

smooth to verrucose.

Hasitat on soil.

Geohypha terrestris (Fr.) Hennebert, comb. nov. Figs 22, 23

MB 835629

= Hyphelia terrestris Fr., Syst. Mycol. 3(1): 213, 1829, nom. sanct.

= Ostracoderma terrestre (Fr.) Nannf., Fungi. Exsicc.

Suec., Fasc. 53-54, Schedae: 40, 1959.

= Chromelosporium terrestre (Fr.) M.B. Ellis, More Demat. Hyphom.: 154, 1976.

= Trichoderma tuberculatum Pers., Ann. Bot. (Usteri) 15: 12, 1795 [as “tuberculata”].

= Chromelosporium tuberculatum (Pers.) Hennebert, Persoonia 7: 198, 1973.

= Trichoderma laeve Pers., Observ. Mycol. 1: 12, 1796.

= Trichoderma varium Ehrenb., Sylv. Mycol. Berol.: 22, 1803.

= Trichoderma nemorosum Pers.,Traité Champ. Comest.: 131, 1818.

= Botrytis ceratioides Peck, Annual Rep.New York State Mus. Nat. Hist. 35: 139, 1884.

= Sporotrichum fossarum Fautrey, Rev. Mycol. (Toulouse) 17: 71, 1893.

= Ostracoderma fossarum (Fautrey) S. Hughes, Canad. J. Bot. 36: 792, 1958.

Type: Trichoderma taeve-Pers- [laeve Pers.? scratched off] varium m. [mei] ad Berol.

in terra umbrosa humida, Thg. [Thiergarten] Berlin, 7/8 [Aug. 7] [scr. Erhenberg].

Herb. Ehrenberg (B, MBT 392413, here designated as lectotype; isolectotypes:

[DAOM 83392, DAOM 83393, MUCL 2387, MUCL 2388}).

CoLoniegs in small cushions, around 5 mm across, linked by a web-like

mycelium, greyish yellow when fresh, brown when dried.

HyPHAE narrow, irregular in diameter, 2-4 um, sometimes inflated to 6 um

before septa and constricted at septa, hyaline, thin-walled, with abundant

lateral narrower branchlets, fertile.

CONIDIOGENUS CELLS either reduced to a conical to cylindrical lateral

outgrowth of the hypha, 2-8 x 1-1.5 um or (most often) developed into a long

and narrow, sinuous, irregular, hypha, 8-50 x 1-1.5 um, possibly ramified,

producing <15-20 thallic conidia on lateral outgrowths and terminal ends.

Conrp1A thallic, solitary, borne on each hyphal outgrowth, seceding

schizolytically through a 1-1.5 um wide septum, one-celled, globose or

subglobose, 4.5-6(-6.5) um, hyaline, with a thick wall, at first smooth, soon

becoming verrucose, with 12-15 warts in median view.

HasitaT: on bare soil after rain in forest.

COMMENTS—'he type and authentic specimens of the species cited in the

nomenclator are microscopically characterized as having narrow, sinuous

conidiogenous hyphae and verrucose conidia mixed with young smooth

conidia, the relative abundance depending on maturity of the fungus.

710 ... Hennebert

Fig. 22. Geohypha terrestris. A. Trichoderma laeve (MUCL 2462) conidiogenous cell with young

conidia. B. Trichoderma varium (lectotype, MUCL 2387) with mature conidia. C. Sporotrichum

fossarum (holotype, MUCL 2486b and isotype, MUCL 2394), conidia borne either solitary on

hyphae, or numerous on tortuous conidiogenous hyphae. D. Botrytis ceratioides (holotype,

MUCL 2467) conidiogenous cell, after release of some conidia. E. Mature conidia from these

four specimens. Scale bar = 10 um.

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 711

Fig. 23. A Trichoderma laeve Pers. (MUCL 2387). B. Trichoderma varium Ehrenb. (MUCL 2387)

(Photos made by J.W. Carmichael, University of Alberta Mold Herbarium in 1961).

The size of the conidia obtained from each specimen drawn in Fig. 21 are

T. laeve 4.5-5.5 (av. 5) um, T: varium 4-6 (av. 5) um, B. ceratioides 5-6(-6.8)

(av. 5.7) um, and S. fossarum 4-6.5 (av. 5) um.

In 1961, I sent slides of Trichoderma laeve and T. varium from Persoon’s

herbarium to J.W. Carmichael for comparison with Chrysosporium species.

He responded with photographs of the slides, and the opinion that the

species were not congeneric with Chrysosporium.

The type of Sporotrichum fossarum contains two packets, one wrapped in

paper contains soil in powder form ona piece of blotting paper (MUCL 2486a)

and the other, in an aluminium sheet, contains the same soil kept compact

and bearing the fungus (MUCL 2486b). The same fungus was distributed in

G. Roumeguere’s Fungi Selecti Exsiccati 6790 (MUCL 2394). The fungus has

abundant vegetative hyphae that are often inflated before the septa and, when

mature, verrucose conidia. Seeing the similarities with an Ostracoderma sp.

sensu Juel, Hughes (1958) classified the species as Ostracoderma fossarum.

Hennebert (1973) made it a synonym of Chromelosporium tuberculatum, an

opinion later reported by Stalpers (1984). Revisiting the collection, I now

assert that Sporotrichum fossarum is neither an Ostracoderma nor a

Chromelosporium species but what is identified here as Geohypha terrestris.

712 ... Hennebert

SPECIMENS EXAMINED

TYPE & AUTHENTIC SPECIMENS: EUROPE: (3) Trichoderma laeve [scr. Persoon]

Herb. Persoon (L 910.264.459) [MUCL 2462] (not L 910.264. 31, Trichoderma laeve?

as written by Persoon). (4) Trichoderma nemorosum [scr. Persoon] Herb. Persoon

(L 910.264-) [DAOM 83900, MUCL 2463]. (5) Trichoderma varium Ehbg. orig.

[on soil] [scr. Ehrenberg] Herb. Schwaegrichen [stamped]” (STR) [DAOM 83898,

MUCL 2441]. (6) “Trichoderma varium Ehrenb. [on soil, no locality, no date] [scr.

Ehrenberg] (B) (DAOM 83394, MUCL 2389). FRANCE (7). Sporotrichum fossarum

sp.n. Fautrey, sur la terre déjetée des fossés dans les bois humides. La Forét de

Clamecy. Eté 1894, Fautrey 212 (holotype) (UPS) [DAOM 83912, MUCL 2486-

a, 2486-b]. (8) Sporotrichum fossarum n. sp. Fautrey, sur la terre rejetée des fossés

dans les bois humides, été 1894. F. Fautrey (isotype). G. Roumeguere Fungi Selecti

Exsiccati 6790 (Rev. Myc. 1895, p. 71, n.49) (G, NY) [MUCL 2394]. GERMANY (9)

Botrytis epigaea Link, [var. alba] ad terram humidam, non raro, Aestate. Fuckel. Fungi

Rhenani Fasc. II, 1863, n° 147 (BX 47111, GRO, S, K, FH) [MUCL 1798]. (Jahrb.

Nassauischen Vereins. Naturk. 23: 363. 1870, p. 363 as, Hyphelia terrestris Fr. var. alba,

nom. inval.). USA: NEw york (10) Botrytis ceratioides Peck, on decaying wood of

Tsuga canadensis, Albany, June, leg. C.H. Peck. (holotype). See 35th Report p.139,

1884 (NYS) [MUCL 2467].

OTHER SPECIMENS: EUROPE: ITaty (11) Botrytis epigaea Lk. f. cinerea, ad terram

argillosam udam, Bosco Montello (Treviso), Sept. 1875. Saccardo Mycotheca Veneta

58 (K-M) [MUCL 2295]. FRANCE (12) Trichoderma nemorosum Pers. Autumno,

ad terram, St Cloud, near Paris. Herb. de Candolle (G 005431) [DAOM 83992,

MUCL 2392]. (13) Trichoderma nemorosum Pers. [on soil] Meudon, Augusto [no

year] Herbier Léveillé in Herb. de Candolle (G 005432) [DAOM 83899, MUCL

2393]. SWEDEN (14) Hyphelia, on half buried birch branches in Sphagnum cushion,

Gastrikland, NNW of Tolffors Swampy wood, Aug. 14 1950, J.A. Nannfeldt. Flora

Suecica 11057 (UPS) [MUCL 2479a]. NETHERLANDS (15) Hyphelia terrestris Fr. in

terra argillacea, Lugd. Batava, 1844, Oudemans (GRO) [MUCL 2775]. DENMARK

(16) Hyphelia terrestris Fr. on bare soil, Saelland, Tisvilde Hegn., Oct. 4 1955, J.A.

Nannfeldt. Flora Danica 14249 (UPS) [DAOM 84938a, MUCL 2495]. BELGruM (17)

Trichoderma laeve on bare soil, Lauzelle forest, Ottignies-LLN, Oct. 2017, G.L.H.

(MUCL 56656).

NORTH AMERICA: USA: NEBRASKA (18) Botrytis ceratioides Peck [on leaves

and mosses] Lincoln, 1200 ft, July 22 1890, T.A. Williams. Nebraska Flora 312

(BP) [MUCL 2423]. CANADA: ONTARIO (19) Rhinotrichum carneum Ell. & Ev. on

dead wood, Oct. 3 1896, Macoun, Ellis collection 185. (NY) [MUCL 2838]. (20)

Trichoderma laeve Pers. on soil, Queen’s University Biological Station, near Chafey’s

Lake, Ontario, July 12 1961, det. G.L. Hennebert (MUCL 2537).

Plicaria endocarpoides

Hennebert (1973) suggested that Plicaria endocarpoides could have a

Chromelosporium-like asexual morph in culture based on a strain from J.W.

Paden. The strain DAOMC 199565 produced on malt agar a conidial state

having no similarity with Chromelosporium as illustrated here.

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 713

al? ae

Fig. 24. Plicaria endocarpoides (DAOMC 199565 = MUCL 57202).

Conidial morph on malt-agar.

Fig. 25. Plicaria endocarpoides (DAOMC 199565 = MUCL 57202).

A. Conidiophores and conidia on malt agar. Scale bar =10 um.

714 ... Hennebert

Plicaria endocarpoides (Berk.) Rifai, Verh. Kon. Ned. Akad. Wetensch., Afd.

Natuurk., Sect. 2, 57(3): 255, 1968. Fics 24, 25

= Peziza endocarpoides Berk., in Hooker, Fl. Nov.-Zel. 2: 199, 1855.

Myce.ivum hyaline prostrate on malt agar.

CONIDIOPHORES erect from creeping hyphae, one or pluri-celled, 10-110

um high, the basal cell enlarged, triangular, the stipe often furcate.

CONIDIOGENUS CELLS intercalary or terminal of the conidiophore,

cylindrical, fusiform, or triangular, 5-8 um wide, producing acropetal solitary

or branched chains of conidia, leaving unthickened scars at schizolytic

secession.

Conrp!1A hyaline, smooth, one-celled, some 2-3-celled, variable in shape,

from globose (3-5 um), to ovate, pyriform, or citriform (5-23 x 3-8 um),

most one celled, some septate, ramoconidia bearing one to three scars of

attachment of simple or branched chains of conidia.

SPECIMENS EXAMINED: CANADA: BritTisH CoLumBiA. Plicaria endocarpoides

(Berk.) Rifai, on burnt wood, slash and ash in coniferous forest, Lightning Lakes trail,

Manning Provincial Park, British Columbia, June 9 1985, K.N. Egger 2044 in living

culture (CCF 6892 = DAOMC 199565 = MUCL 57202).

Conclusion

In light of certain mononematous Chromelosporium asexual morphs

actually representing Peziza or Plicaria, and certain synnematous

Chromelosporiopsis asexual morphs representing Pachyphlodes, it was logical

to emphasize a morphological character to distinguish the asexual morphs of

these Pezizaceae. The chosen criterion of conidiophore fasciculation allows

segregating similar conidial fungi not known to have sexual morphs into two

genera—Chromelosporium around its mononematous type and the new genus

Chromelosporiopsis for the synnematous species. Chromelosporium includes

here five taxa with distinct morphologies. Chromelosporiopsis comprises so

far two named taxa. In addition to the excluded taxa, there remain some

doubtful taxa showing some (but insufficient) Chromelosporiopsis and a set

of unidentified species detected among the herbarium specimens that further

morphological investigation of might allow segregation into species. It is

recommended that any new fresh conidial samples be not only genetically

analysed but accurately described in their finest morphological details

(including electron microscopy) to allow characterization and identification

of these conidial fungi, many of which are likely to be asexual morphs of

some known Pezizaceae.

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 715

Examination of many herbarium specimens revealed a long history

of misapplication of the name Hyphelia terrestris, irrespective of Fries

protologue of the species. A neotype of the species designated among the

synonyms given by Fries allowed a new description of the species renamed

Geohypha terrestris.

Acknowledgments

I am thankful to my colleagues, Dr. Keith Seifert (Adjunct Professor, Carleton

University, Ottawa and formerly of Ottawa Research and Development Center,

Agriculture and Agri-Food Canada) and Dr. David L. Hawksworth (Royal Botanic

Gardens Kew), who thoroughly and accurately revised the manuscript in agreement

with the present rules of nomenclature. I thank Dr Scott Redhead (Curator, National

Mycological Herbarium (DAOM), Agriculture and Agri-Food Canada) for his

appreciated nomenclatural advice and Dr Konstanze Bensch, (MycoBank curator,

Westerdijk Fungal Biodiversity Institute, Utrecht) for the registration of names

and types in MycoBank and her meticulous revision of all name citations. I thank

Dr M. Scherrenberg and Dr J. Nuytinck of Naturalis Biodiversity Center in Leiden for

the photographs and examination of the precious Persoon’s material of Isaria carnea

and Mr Jonathan Mack from Quebec for the photographs of Chromelosporium

coerulescens I thank also Dr Shaun Pennycook and Dr Lorelei Norvell, editors of

Mycotaxon, for editing the paper through very friendly exchanges. Also I remain

grateful to the late Dr R.P. Korf of Cornell University, Ithaca, for his profound

friendship and the multiple scientific exchanges and support for this research,

particularly during 1973.

A personal note of gratitude to late Dr. Stanley J. Hughes

I am overall deeply grateful to late Dr. Stanley J. Hughes, a great mycologist, who

was a great teacher and a loyal friend. After sending him some pages of my doctoral

thesis on the genus Botrytis, inspired by some of his major publications, I received

an invitation to apply for a National Research Council of Canada postdoctoral

fellowship in the Mycology Section of the then Plant Research Institute of the

Department of Agriculture Canada, in Ottawa, from October 1960.

My wife Lidwina and I arrived in Canada at the end of September 1960 and were

received by Stanley and Lyndell Hughes. Once we were settled in the apartment they

had carefully prepared for us, Stan introduced us to his colleagues of the Mycology

Section in the Neatby Building at the Central Experimental Farm. The following

day, Stan immediately began my training with a collecting trip to Gatineau Park

near Ottawa (Fig. 26).

In the laboratory, showing great patience and efficiency with the student I then

was (speaking only a few words of English accented French), Stan tested my skills

with a simple project, describing Balanium from the DAOM herbarium. Then

followed the description of Arachnophora fagicola, a new genus and species collected

716 ... Hennebert

in Belgium. As many as seven times, he asked me to improve the manuscripts I

submitted to him, each word having to be useful and necessary.

To introduce me to ancient mycological literature, Stan asked me to clarify the

complex history of Oedemium didymum. To describe the apothecial stage of certain

Botrytis species reported in my doctoral thesis, Dr. J.W. Groves, in charge of the

Mycology Section, with Dr. Elliot, introduced me to the in vitro production of the

apothecia of Botrytis from living strains.

During the second year of the award, Stan (Fic. 27A) urged me to develop my

research on the Botrytis-like fungi begun in my PhD thesis and which he classified

in Ostracoderma. In 1973, incited by Dr Korf to publish, I revised some Botrytis-

like genera and renamed Ostracoderma species in Chromelosporium (also described

and illustrated in the present paper).

Chromelosporiopsis gen. nov. & Geohypha stat. nov. ... 717

Those two years of mycological training with Dr. Hughes and Dr. Groves not

only were essential to my career, but also provided the opportunity for creating an

indestructible bond of respect and friendship. Later, Stan Hughes did to me the

honour of visiting my laboratory at the UCL in Herverlee-Leuven in May 1973

(Fic. 27B) and in Louvain-la-Neuve in June 1994 (Fic. 27C) at the celebration

of the Centenary of the MUCL fungus collection (Hennebert 2010). I remain

very grateful to the late Stanley J. Hughes for the quality of my training and his

friendship.

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