IV ... MYCOTAXON 136(3)

MY COTAXON

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

FREVICWETSS <1), Muveting con tarps titecedTt. ext: Pb anGieg HE MIE ENTE > ath ee mie neat arp oe Me epee Eee vi

Nomenclatural novelties & typifications ....... 0... c cece eee vii

CORTICCIT sce te Me ahs 0 este Se ent ct AE PRR de Praha Ae A tegen ase eet Viii

PROTA LCE ABOU Dc hoon ph ph ane,Sarme sleebi thane tea Te Fh dene MaFncten EC ieee SLE Sel 4a aday 9 ix

DOZE SUDIISSIOMPTOCC AITO, 5 maw. 2 cnn ea nth apt ES 6 VEE A ER ARIS xii

NEw TAXA

Vanakripa chinensis sp. nov. from China

and notes on the genus Kal ZHANG, WEIHUA Guo,

De-WE!I LI, RAFAEL FR CAsTANEDA-RUIZ

Cordana sinensis sp. nov. from southern China

Linc Qu, KAI ZHANG, ZHAO-HUAN Xu,

RAFAEL FE, CASTANEDA-RUIZ, JIAN MA

Pezicula endophytica sp. nov., endophytic in Dendrobium

from Thailand X1A0-YA Ma, JI-CHUAN KANG,

KEVIN D. HypDE, MINGKWAN DOILOM, PUTARAK CHOMNUNTI

Globoramichloridium delicatum and Heteroconium simile spp. nov.

from southern China LING Qiu, Xu-GEN SHI, X1u-Guo ZHANG,

RAFAEL FE, CASTANEDA-RUIZ, JI- WEN XIA, JIAN Ma

Neosporidesmium himachalense sp. nov. from India

and Neosporidesmina gen. nov. to accommodate N. micheliae

RAJNISH KUMAR VERMA, I.B. PRASHER, SUSHMA,

Ajay KUMAR GAUTAM, KUNHIRAMAN C. RAJESHKUMAR,

ASHISH KUMAR, RAFAEL F. CASTANEDA-RUIZ

Cladorrhinum yunnanense sp. nov. from China

YUE PAN, JI-WEN XIA, SHENG-TING HUANG,

CHUN-YUAN ZHU, XIU-GUO ZHANG, ZHUANG LI

Mazosia hainanensis sp. nov. from tropical China

ZONG-TING YAO, ZE-FENG JIA, SHU-HUA JIANG

Racocetra cromosomica sp. nov. from Oaxaca, Mexico

EDUARDO CHIMAL-SANCHEZ, IRMA REYES-JARAMILLO,

SARA LuciA CAMARGO-RICALDE, LUCIA VARELA,

JOSE YABIN SALMERON CASTRO, NOE MANUEL MONTANO

545

553

563

579

587

597

607

615

JULY-SEPTEMBER 2021... V

Bactrospora cozumelensis sp. nov. from MexicoYEniTzE A. GARC{A-MARTINEZ,

JORGE GUZMAN-GUILLERMO, RICARDO VALENZUELA, TANIA RAYMUNDO 627

Lembosia mimusopis sp. nov. from Thailand

Diana S. MARASINGHE, MONIKA C. DAYARATHNE,

SAJEEWA S.N. MAHARACHCHIKUMBURA, ABDALLAH M. ELGORBAN,

SINANG HONGSANAN, KEVIN D. HyDE 635

ENVIRONMENTAL RANGE VERIFICATION

Fibroporia gossypium isolated from an indoor environment

in Argentina CAROLINA A. ROBLES, MARiA BELEN PILDAIN,

FRANCISCO SAUTUA, MARCELO A. CARMONA, MARIO RAJCHENBERG 645

NEW RANGES/HOSTS

Rubellofomes cystidiatus and Spongiporus floriformis

newly recorded for India = VinyusHA NELLIPUNATH & T.K. ARUN. KuMaAR 661

Diplodia bulgarica, a new record for Turkey CAFER EKEN 669

Gerhardtia foliicola, a new record for Pakistan AIMAN IZHAR,

MUHAMMAD USMAN, MUNAZZA KIRAN, ABDUL NASIR KHALID 675

MycoBIOTA (FUNGAE) NEW TO THE MYCOTAXON WEBSITE

Checklist of the lichens of The Reserva Florestal Adolphe Ducke

in Manaus (Amazonas, Brazil) (summary)

ANDRE APTROOT, JANICE GOMES CAVALCANTE, LIDIANE ALVES DOS SANTOS,

ISAIAS OLIVEIRA JUNIOR, DAYANE DE OLIVEIRA LIMA,

MARCELA EUGENIA DA SILVA CACERES 685

First molecular-based contribution to the checklist

of Lebanon macrofungi (summary) SANDRA SLEIMAN,

JEAN-MICHEL BELLANGER, FRANCK RICHARD, JEAN STEPHAN. 687

Checklist of powdery mildew fungi (Erysiphaceae)

of Pakistan (SUMMARY) N.S. AFSHAN, I. ZAFAR, A.N. KHALID 689

Aphyllophoroid fungi (Basidiomycota) of Chile:

An annotated checklist (SUMMARY)

CRISTIAN RIQUELME & MARIO RAJCHENBERG 691

VI ... MYCOTAXON 136(3)

REVIEWERS — VOLUME ONE HUNDRED THIRTY-SIX (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.

Artur Alves

André Aptroot

Boris Assyov

Rosa Maria Arias Mota

D. Jayarama Bhat

Janusz Blaszkowski

Harold H. Burdsall, Jr.

Manuela Dal Forno

Ismail Erper

Genevieve Maria Gates

Tuba Geng Kesimci

Sergio P. Gorjon

Bruno Tomio Goto

Jorge Guzman Guillermo

Manoj Emanuel Hembrom

Gabriela Heredia Abarca

Bryce Kendrick

Junaid Khan

Seyed Akbar Khodaparast

De-Wei Li

Ting Li

Michael Loizides

Lei Lu

Jian Ma

Eric H.C. McKenzie

Armin Mesi¢

Karen K. Nakasone

Lorelei L. Norvell

Shaun R. Pennycook

Mario Rajchenberg

Andrea C. Rinaldi

Amy Y. Rossman

Israel Pérez- Vargas

Harrie J.M. Sipman

Susumu Takamatsu

Joanne Taylor

Ricardo Valenzuela

Fang Wu

Zefen Yu

Lulu Zhang

JULY-SEPTEMBER 2021...

NOMENCLATURAL NOVELTIES AND TYPIFICATIONS

PROPOSED IN MYCOTAXON 136(3)

Bactrospora cozumelensis Y. Garcia, Guzm.-Guill., R. Valenz. & Raymundo

[MB 836514], p. 630

Cladorrhinum yunnanense Y. Pan, J.W. Xia, X.G. Zhang & Z. Li

[MB 836106], p. 602

Cordana sinensis L. Qiu, K. Zhang, R.E Castafieda & Jian Ma

[MB 836854], p. 556

Globoramichloridium delicatum L. Qiu, Jian Ma, R.F. Castafieda &

X.G. Zhang

[MB 836851], p. 580

Heteroconium simile L. Qiu, Jian Ma, R.F. Castafieda & X.G. Zhang

[MB 836852], p. 582

Lembosia mimusopis Marasinghe, Daya., Maharachch., Hongsanan &

K.D. Hyde,

[IF 556875], p. 640

Mazosia hainanensis Z.T. Yao, S.H. Jiang & Z.F. Jia

[MB 836195], p. 610

Neosporidesmina R.F. Castafieda, Rajn.K. Verma, Prasher, Sushma,

A.K. Gautam & Rajeshk.

[IF 557995], p. 592

Neosporidesmina micheliae (Y.D. Zhang & X.G. Zhang) R.F. Castaneda,

Rajn.K. Verma, Prasher, Sushma, A.K. Gautam & Rajeshk.

[IF 557996], p. 592

Neosporidesmium himachalense Rajn.K. Verma, Prasher, Sushma,

A.K. Gautam, Rajeshk. & R.F. Castafieda

[IF 557994], p. 589

Pezicula endophytica X.Y. Ma, K.D. Hyde & J.C. Kang

[IF 558634], p. 570

Racocetra cromosomica Chim.-Sanch., Varela & Montafo

[IF 557326], p. 618

Vanakripa chinensis K. Zhang, W.H. Guo, R.F. Castaneda & D.W. Li

[IF 557829], p. 546

VII

VII ... MYCOTAXON 136(3)

CORRIGENDA

MYCOTAXON 136(2)

p. 343, line 14 FOR: BAsionym: R.H. Petersen & K.W. Hughes. 2014. N. Am. Fungi

9: 6.

READ: BASIONYM: Gymnopus micromphaloides R.H. Petersen & K.W.

Hughes. 2014. N. Am. Fungi 9: 6.

NOTE: Omission of the basionym name invalidated the proposed combination.

Collybiopsis micromphaloides was validly published in Petersen & Hughes, Index

Fungorum 491: 1 (2021).

pp. 359-360 Miussinc! The missing pages resulting from misnumbering p. 361 were not

noticed until after publication

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

MYCOTAXON for APRIL-JUNE 2021 (I-XII + 263-544)

was issued on July 23, 2021

JULY-SEPTEMBER 2021 ... IX

FROM THE EDITOR-IN-CHIEF

SO, YOU THINK YOU'VE FOUND A NEW SPECIES? One of the joys your MycoTAxoNn

editors share is overseeing the publication of new taxa (the primary focus of this

journal) and which generally involves shepherding authors to the realization that a

new species best withstands the scrutiny of other researchers and the test of time

if it is based on more than a single specimen. Our Nomenclature Editor shares his

thoughts on the subject:

“THE IDEAL STANDARD FOR A PROPOSAL OF A NEW TAXON: Many manuscripts

submitted to MycOTAXON propose a new taxon based on a single specimen

(the holotype). This is completely acceptable if this is the only known material,

but it has come to our attention that some of these manuscripts are failing to

report other specimens that have been collected and vouchered. Whenever

possible, the proposal of a new taxon should present: (1) a type collection

that was large enough to be divided and vouchered in two (or more) different

herbariums, one designated as the holotype and the others as isotypes; (2)

other specimens (ideally collected from different localities, or from different

hosts/substrates, or on different dates), which the manuscript should present,

with full collection and vouchering data, in an ADDITIONAL SPECIMEN(S)

EXAMINED Section inserted immediately after the description of the new

taxon. Authors should aim to achieve this ideal standard when collecting

material and are encouraged (one might even say required) to report all

conserved specimens of a new taxon in any manuscript inaugurating a new

taxon.”

Contending with only a single specimen on which to base a new species (and possibly

new genus) has long been a bane of careful workers. With the advent of sequence

analyses, many have been driven to “check the DNA’ to reassure themselves that

their morphologically and/or culturally based holotype also differs phylogenetically.

Unfortunately, several isolates obtained from a single specimen do not increase

species support when they cluster together nor do they address the variation likely to

be encountered in different specimens. Both phylogenetic and morphological species

concepts are strengthened when several specimens are studied.

OF PARATYPES AND PROTOLOGUES. Early in my taxonomic studies, I was convinced

that a collection cited by a scientist in the EXAMINED SPECIMENS sections of a new

species (technically referred to as a PARATYPE) had taxonomic and nomenclatural

significance. Not until I compared several paratype collections with one holotype

specimen to discover several cryptic but morphologically distinct species hidden

under one name did I realize that while a paratype (representing the original author’s

species concept) should always be evaluated as taxonomically important, it held no

nomenclatural significance, despite its “-type” suffix. A paratype is simply a specimen

X ... MYCOTAXON 136(3)

included in the protologue of the type description. And the PROTOLOGUE? As defined

in the DICTIONARY OF THE FuNGI (Kirk & al. 2008), it is: “everything associated

with a name on its first publication, i.e. diagnosis, description, references, synonymy,

geographical data, citation of specimens, discussion, comments, illustrations.”

The delayed 2021 July-September Mycotaxon, is unusually slim, offering 18

contributions by 79 authors (representing 14 countries) as revised by 40 expert

reviewers and the editors.

With ten titles, the NEW TAXA section proposes ONE new genus (Neosporidesmina

from India) plus 11 species new to science representing Bactrospora and Racocetra

from Mexico; Cladorrhinum, Cordana, Globoramichloridium, Heteroconium,

Mazosia, Pezicula, and Vanakripa from Curina; Lembosia from THAILAND; and

Neosporidesmium from INp1A. We also offer one new combination in Neosporidesmina

from India.

The single title in the ENVIRONMENTAL RANGE VERIFICATION section explores

the mysterious discovery of Fibroporia gossypium, a widely distributed brown-rot

polypore (described 122 years ago by Spegazzini and normally collected in conifer

(Europe, North America, Asia) or Nothofagus forests) in an abandoned basement of

a university plant pathology lab.

The NEw RANGES/HOsTs section contains three titles reporting range extensions

for [basidiomycetes] Rubellofomes and Spongiporus for INDIA and Gerhardtia for

PAKISTAN and [dothideomycetes] Diplodia for TurKEy. MycoTaxon 136(3) also

provides keys to species in Cordana, Mazosia, and Vanakripa, establishes a new

Racocetra species through culture study, and confirms the pathogenicity of Diplodia

bulgarica on apples in Turkey.

Papers providing conclusions supported by sequence analyses cover six new

species representing Cordana, Cladorrhinum, Lembosia, Mazosia, Pezicula, and

Vanakripa and five environmental or range extensions for Diplodia, Fibroporia,

Gerhardtia, Rubellofomes, and Spongiporus.

Our issue concludes with the announcement of four excellent new annotated

species lists on our MYCOBIOTA website, covering lichens in BRAzIL, macrofungi of

LEBANON, powdery mildews of PAKISTAN, and the aphyllophoroid fungi of CHILE.

Warm regards and best wishes,

Lorelei L. Norvell (Editor-in-Chief)

Shaun R. Pennycook (Nomenclature Editor)

15 October 2021

JULY-SEPTEMBER 2021 ... XI

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MY COTAXON

July-September 2021— Volume 136, pp. 545-551

https://doi.org/10.5248/136.545

Vanakripa chinensis sp. nov. from China

and notes on the genus

KAI ZHANG’”, WEIHUA Guo", DE-WEI Li?, RAFAEL F. CASTANEDA-RUuIZz?

' Institute of Ecology and Biodiversity, School of Life Sciences, Shandong University,

Qingdao, 266237, China

? Shandong Agriculture and Engineering University, Jinan, Shandong 250100, China

> The Connecticut Agricultural Experiment Station, Valley Laboratory,

153 Cook Hill Road, Windsor, CT 06095, USA

‘ Instituto de Investigaciones Fundamentales en Agricultura,

Tropical Alejandro de Humboldt’ (INIFAT), OSDE, Grupo Agricola,

Calle 1 Esq. 2, Santiago de Las Vegas, C. Habana, Cuba, C.P. 17200

* CORRESPONDENCE TO: whguo_sdu@163.com

ABSTRACT—A new species Vanakripa chinensis found on decaying branches of an

unidentified tree is described and illustrated. It is characterized by sporodochial

conidiomata composed of macronematous, cylindrical conidiophores that are sometimes

reduced to conidiogenous cells and obovoid, unicellular, brown, smooth conidia with a

black spot near the middle. Notes on conidiogenous events in the genus Vanakripa and a

key to the species are provided.

Key worps—asexual fungi, hyphomycetes, taxonomy

Introduction

Vanakripa, was introduced by Bhat & al. (Bhat & Kendrick 1993) with

V. gigaspora as type species; it is distinguished by scattered, punctiform,

pulvinate to postulate black or dark brown loosely or compact sporodochial

conidiomata. The conidiophores are macronematous, mononematous,

cylindrical below and inflated toward the apex, erect, flexuous, un- or slightly

branched, refractively septate, smooth, and hyaline. The conidiogenous cells

are monoblastic, cylindrical to narrowly or broadly clavate, truncated after

546 ... Zhang & al.

conidial secession, terminal, integrated, hyaline (sometimes with a refractive

lumen), and smooth. Conidia are solitary, acrogenous, variably shaped

(clavate, broadly pyriform, obovoid, ellipsoidal), truncate at the base and

rounded at the apex, 0-1-septate, darkly pigmented, smooth, thick walled,

usually remaining attached to the conidiogenous cells (Bhat & Kendrick 1993).

Under the predominantly subtropical climate of Yunnan province, several

novel asexual fungi have been found in the rainforest and mountains by Guo

& al. (2019), Zhang & al. (2020), and Zheng & al. (2019, 2020a,b). During a

survey of fungal diversity in Xishuangbanna, Yunnan province, an interesting

fungus was collected on decaying branches; its conidiomata, conidiogenesis,

and conidial features clearly suggest a placement in the genus Vanakripa.

Materials & methods

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

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

(2016). Mounts were prepared in lactic acid (90%) or in polyvinyl alcohol-glycerol

(8 g PVA in 100 mL of water + 5 mL of glycerol). Microscopic characters were

measured at x1000 using a Nikon Eclipse Ni-U microscope with DIC optics and a

Nikon DS-Fi2 camera. The holotype was deposited in the Herbarium of Department

of Plant Pathology, Shandong Agricultural University, China (HSAUP).

Taxonomy

Vanakripa chinensis K. Zhang, W.H. Guo, R.F. Castafieda & D.W. Li, sp. nov. Fic. 1

IF 557829

Differs from Vanakripa fasciata by its narrow obovoid, smaller conidia; and from

V. rhizophorae by its smaller, uniformly obovoid, dark spotted conidia.

Type: China. Xishuangbanna Tropical Botanical Garden, on decaying branches of

an unidentified broadleaf tree, 22 July 2019, coll. K. Zhang & W.H. Guo (Holotype,

HSAUP Y190114).

EtryMo oey: Latin, chinensis, referred to the country of collection, China.

CoLonlgs on the natural substrate effuse, black. CONIDIOMATA sporodochial,

scattered, punctiform, pulvinate or pustuliform, <250 um diam. MyceLium

superficial and immersed, composed of septate, branched, hyaline, smooth-

walled hyphae. CONIDIOPHORES macronematous, mononematous, erect,

straight or flexuous, 0-2-septate, sometimes refractive at the septa, smooth-

walled, hyaline, <35 um length, sometimes reduced to conidiogenous cells.

CONIDIOGENOUS CELLS monoblastic, cylindrical, sometimes narrowly clavate

lumen refractive or translucent, determinate, integrated, terminal, smooth,

hyaline, 15-29 x 2.5-5 um. CONIDIAL SECESSION schizolytic. ConrpI1a solitary,

Vanakripa chinensis sp. nov. (China) ... 547

Fic. 1. Vanakripa chinensis (holotype, HSAUP Y190114). A. Conidia after schizolytic secession;

B. Detached and attached conidia and conidiogenous cells; C. Conidiogenous cell with attached

conidium; D. Conidium after schizolytic secession; E. Sporodochium. Scale bars: B-E = 5 um.

acrogenous, obovoid to broad obovoid, truncate at the base, unicellular, thick-

walled, guttulate, 19.8-22.8 x 12.7-14.8 um, smooth, brown to black, with a

conspicuous or inconspicuous black spot near the middle.

Notes: Vanakripa now comprises nine accepted species (Index Fungorum

2020; this paper): V. chinensis, V. fasciata R.F. Castafieda & al., V. gigaspora

Bhat & al., V. inexpectata S.M. Leao & Gusmao, V. inflata (Hol.-Jech.) Melnik,

V. menglensis D.M. Hu & al., V. minutiellipsoidea Pinnoi, V. parva Bhat & al.,

548 ... Zhang & al.

and V. rhizophorae R.M. Arias & al. €Arias-Mota & al. 2008, Bhat & Kendrick

1993, Castaneda-Ruiz & al. 2005, Hu & al. 2010, Leao-Ferreira & al. 2013,

Melnik 2011, Pinone & al. 2003). Additionally there is an invalid species,

“V. ellipsoidea” K.M. Tsui & al. (Tsui & al. 2003), which lacks a designated

holotype. Vanakripa chinensis superficially resembles V. fasciata in the dark

spotted conidia, but V. fasciata has bigger (24-34 x 16-23 um) ellipsoidal

conidia that are dark brown with a black transverse band at the centre.

Also somewhat similar to V. sinensis, V. rhizophorae is distinguished by its

brown to black, variably shaped (globose, ovoid to ellipsoidal; 16.6-22.3 x

6.3-14.7 um) conidia.

Key to Vanakripa species

| onidia-che celled:ay.s, ams. samy. ac ea ae etal Se i Se Ee 3

1. Conidia with a submedian septum ............. 00. cee eee eee eee eee eens 2

2. Conidia ellipsoidal or obovoid, 17-27 x 7-12 um, smooth, asymmetric, apical cells

large, brown to dark brown, basal cells small, pale brown......... V. inexpectata

2. Conidia clavate, obovoid to obpyriform, 32-43 x 18-22 um, smooth, asymmetric,

apical cells large, 30-38 um long, dark brown to black, basal cell small, 3-5 um

lone pppale Drowns sep wane eine at een vate oe veda y tee V. gigaspora

3. Conidia mostly ovoid, obovoid, or fusiform, truncate at the base................ 4

3. Conidia not primarily ovoid, obovoid, or fusiform ............. 0... e eee eee eee 5

4. Conidia obovoid, larger (19.8-22.8 x 12.7-14.8 um), guttulate,

brown to black, with a conspicuous or inconspicuous black spot

néar the middlessmeoth: ayy Poy we cae ess ees WER ee eh V. chinensis

4. Conidia ovoid or fusiform, smaller (7-10 x 4-6 um),

browirtocdaris Drowal Meade Seas hake Rekha Bh tm ha ay Bee Be he ee V. parva

Se Conidia-mostly ell ps@idaltss .. ss so heey she wise kaorei, «oprah Apts chalet bp acta a 6

5. Conidia mostly clavate or globose ........ 0. eee cee eee eens 8

6. Conidia fasciate, ellipsoidal, 24-34 x 16-23 um, brown to dark brown,

with a black transverse band at the centre, smooth-walled .......... V. fasciata

Gs Conidis NObTASC Ate 4, tle. Prete perks Mester «Renters «Foren « fives ePaiew eevee aehioren she 7

7. Conidia ellipsoidal, 25-33 x 16-23 um, with rounded apex and base,

dark brown to black, smooth-walled .................... V. minutiellipsoidea

. Conidia broadly ellipsoidal to subspherical, 14.5-24 x 9.5-19.5 um,

truncate at the base, dark brown, smooth-walled;

conidiogenous cell 27-48 x 8-14 Um... eee ee eee V. inflata

8. Conidia clavate to obpyriform, truncate at the base, (8-)17-23 x 8-13 um, brown to

dark brown, smooth; conidiogenous cells 20-40 x 4-6 um.......... V. menglensis

8. Conidia globose, obovoid to ellipsoidal, 17-22 x 6.3-14.7 um, dark brown

to black, smooth; conidiogenous cells 29.4-54 x 4-9.3 um........ V. rhizophorae

Vanakripa chinensis sp. nov. (China) ... 549

Discussion

Bhat & Kendrick (1993) described Vanakripa as having “conidiogenous

cells [that are] holoblastic, obpyriform, clavate, colorless, often curved,

narrowed toward the base, truncate after conidial secession” and “conidia

[that are] clavate to obpyriform, rounded at the tip, truncate at the base,

darkly pigmented, smooth, 0-1-septate-” Obviously, the conidial secession of

Vanakripa is schizolytic, and only after this event, both conidiogenous cells

and conidia have flat or truncate scars at the cell apex and conidial base.

A conidiogenous cell is defined as any cell from or within which a conidium is

directly produced, and a locus is the place on the conidiogenous cell where a

conidium arises (Kendrick 1971, Kirk & al. 2008). In Vanakripa mature conidia

usually remain attached on the conidiogenous loci, and after mechanical

disturbance the conidiogenous cells and conidia separate by rhexolytic

secession from the conidiophores. However, conidia that have been captured

from the surface of substrate or released from the conidiogenous cells in a

drop of KOH 4% show a flat, truncate base without any portion or frill of the

conidiogenous cells (Fic. 1A). Bhat & Kendrick (1993), Holubova-Jechova

(1983), Ledo-Ferreira & al. (2013), Hu & al. (2010), and Seifert & al. (2011),

have illustrated similar flat, basal truncated conidia. Hughes (1951) discussed

Penzig’s illustration of Beltrania rhombica Penz., with special reference to the

attached intermediate cell found between conidiogenous cell and conidia that

Penzig called “stermmi” to be later renamed a “separating cell” by Hughes

(1951) and also adopted by Pirozynski (1963) in his taxonomical studies

of Beltrania, Beltraniella, Beltraniopsis, and Ellisiopsis. In the strict sense, a

separating cell is a cell between a conidium and a conidiogenous cell (Kirk &

al. 2008); but the term is also applicable to the conidium, at first attached to

the conidiogenous cell, then separating by schizolytic or rhexolytic secession.

In Vanakripa the conidiogenous cells are monoblastic, terminal, integrated,

cylindrical, vermiform, long clavate, usually slightly or strongly inflated near

the conidiogenous locus and the conidia are produced on the apical locus,

but not separating cell, which exists between the conidiogenous cells and

conidia (Bhat & Kendrick 1993). However, separating cells associated with

rhexolytic and schizolytic conidial secession have also been reported and

illustrated in Pseudotrichoconis, Rhexodenticula, and Dictyoaquaphila (Baker

& al. 2001, Li & al. 2011, Arias-Mota & al. 2016, Monteiro & al. 2016).

Acknowledgments

The National Natural Science Foundation Program of PR China (31870016)

financed this work. We are indebted to Prof. Dr. Bryce Kendrick (8727 Lochside

550 ... Zhang & al.

Drive Sidney, BC, Canada), Dr. Rosa Maria Arias Mota (Instituto Tecnoldgico

Superior de Xalapa, Mexico) and Dr. Gabriela Heredia Abarca (Instituto de

Ecologia A.C.) for their critical reviews. We acknowledge the websites provided

by Dr. P.M. Kirk (Index Fungorum) and Dr. K. Bensch (MycoBank). Dr. Lorelei

L. Norvell’s editorial review and Dr. Shaun Pennycook’s nomenclature review are

greatly appreciated.

Literature cited

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and Vanakripa and the other microfungi recorded from mangrove in Veracruz, Mexico.

Mycotaxon 106: 29-40.

Arias-Mota RM, Heredia G, Castafieda-Ruiz RF. 2016. Two new species of Bactrodesmium and

Dictyoaquaphila from Mexico. Mycotaxon 131: 291-295. https://doi.org/10.5248/131.291

Baker WA, Partridge EC, Morgan-Jones G. 2001. Notes on Hyphomycetes. LXXXIV.

Pseudotrichoconis and Rhexodenticula, two new monotypic genera with rhexolytically

disarticulating conidial separating cells. Mycotaxon 79: 361-373.

Bhat DJ, Kendrick B. 1993. Twenty-five new conidial fungi from the Western Ghats and the

Andaman Islands (India). Mycotaxon 49: 19-90.

Castafieda-Ruiz RF, Stadler M, Saikawa M, Iturriaga T, Decock C, Heredia G. 2005. Microfungi

from submerged plant material: Zelotriadelphia amoena gen. et sp. nov. and Vanakripa fasciata

sp. nov. Mycotaxon 91: 339-345.

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America. 197-217, in: DW Li (ed.). Biology of Microfungi. Springer International Publishing.

https://doi.org/10.1007/978-3-319-29137-6_9

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new members of Wiesneriomycetaceae. International Journal of Systematic and Evolutionary

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Hu DM, Cai L, Chen H, Bahkali AH, Hyde KD. 2010. Four new freshwater fungi associated with

submerged wood from Southwest Asia. Sydowia 62: 191-203.

Hughes S. 1951. Studies on micro-fungi. Beltrania, Ceratocladium, Diplorhinotrichum and

Hansfordiella (gen. nov.). Mycological Papers 47, 1-15.

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[Accessed 26 July 2020].

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Imperfecti. Edited by W. B. Kendrick. University of Toronto Press, Toronto.

https://doi.org/10.3138/9781487589165-018

Kirk PM, Cannon PF, Minter DW, Stalpers JA. 2008. Ainsworth & Bisby’s dictionary of the fungi.

10th edition. CAB International Wallingford, U.K. https://doi.org/10.1079/9780851998268.0000

Leao-Ferreira SM, Gusmao LFP, Castafieda-Ruiz RE 2013. Conidial fungi from the semi-arid

Caatinga biome of Brazil. Three new species and new records. Nova Hedwigia 96: 479-494.

https://doi.org/10.1127/0029-5035/2013/0084

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Mycotaxon 117: 287-290. https://doi.org/10.5248/117.287

Mel'nik VA. 2011. Materialy k izucheniyu anamorfnykh gribov Vetnama [Contribution to the

examination of anamorphic fungi of Vietnam]. Mikologiya i Fitopatologiya. 45: 323-331.

Vanakripa chinensis sp. nov. (China) ... 551

Monteiro JS, Conceigao LB, Marques MFO, Gusmao LFP, Castafieda-Ruiz RF. 2016. Dictyoaquaphila

appendiculata gen. & sp. nov. from submerged wood from Brazil. Mycotaxon 131: 177-183.

https://doi.org/10.5248/131.177

Pinnoi A, McKenzie EHC, Jones EBG, Hyde K.D. 2003. Palm fungi from Thailand: Custingophora

undulatistipes sp nov. and Vanakripa minutiellipsoidea sp. nov. Nova Hedwigia 77: 213-219.

https://doi.org/10.1127/0029-5035/2003/0077-0213

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V. ellipsoidea sp. nov. Mycologia 95: 124-127. https://doi.org/10.1080/15572536.2004. 11833140

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characterization of two new species of Orbilia (Orbiliomycetes) from China.

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and a key to species of the genus. International Journal of Systematic and Evolutionary

Microbiology, 69: 3161-3169. https://doi.org/10.1099/ijsem.0.003605

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from China and a key to species of the genus. International Journal of Systematic and

Evolutionary Microbiology 70: 1178-1185. https://doi.org/10.1099/ijsem.0.003897

Zheng H, Yu ZF, Xu JP, Castafeda-Ruiz RF, Qiao M. 2020b. Ramichloridium endophyticum

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China. International Journal of Systematic and Evolutionary Microbiology 70: 433-439.

https://doi.org/10.1099/ijsem.0.004190

MY COTAXON

July-September 2021— Volume 136, pp. 553-562

https://doi.org/10.5248/136.553

Cordana sinensis sp. nov. from southern China

LING Qu’, Kal ZHANG’, ZHAO-HUAN XU’,

RAFAEL FE, CASTANEDA-RUuiIz?}, JIAN Ma***

' College of Agronomy, Jiangxi Agricultural University, Nanchang, Jiangxi 330045, China

? College of Forestry Engineering, Shandong Agriculture and Engineering University,

Jinan, Shandong 250100, 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

‘Jiangxi Key Laboratory for Conservation and Utilization of Fungal Resources,

Jiangxi Agricultural University, Nanchang, Jiangxi 330045, China

* CORRESPONDENCE TO: majian821210@163.com; jxaumj@126.com

ABSTRACT—A new species, Cordana sinensis, is described and illustrated from a specimen

collected on dead branches of an unidentified broadleaf tree in Jiangxi Province, China.

It is characterized by its integrated, polyblastic conidiogenous cells that produce ovoid to

obclavate, medially 1-2-septate pale brown, smooth conidia with a slightly prominent hilum.

A dichotomous key and a synoptic table to Cordana species, are provided.

KEY worps—asexual Ascomycota, Cordanaceae, hyphomycetes, saprobes, taxonomy

Introduction

Cordana Preuss was established with C. pauciseptata Preuss as the

type species (Preuss 1851) and characterized mainly by macronematous,

mononematous, unbranched, cylindrical, nodose or subnodose

conidiophores and polyblastic, integrated, terminal or intercalary, sympodial

conidiogenous cells, usually denticulate with tiny or conspicuous cylindrical

denticles. The conidia are solitary, acropleurogenous, variably shaped (ovoid,

obovoid, ellipsoidal, pyriform, or obpyriform), 0-1-euseptate and seceding

schizolytically (Preuss 1851, Hughes 1955, Ellis 1971, Markovskaja 2003,

554 ... Qiu & al.

Seifert & al. 2011). Hernandez-Restrepo & al. (2014), who expanded the

generic concept to include integrate and discrete polyblastic conidiogenous

cells that are intercalary nodose and swollen or umbellate at the conidiophore

apices, provided a key for 19 accepted species. Thereafter, C. johnstonii

M.B. Ellis, C. musae (Zimm.) Hohn., and C. versicolor D.J. Soares &

R.W. Barreto were transferred to Neocordana Hern.-Rest. & Crous based

on a phylogenetic analysis (Hernandez-Restrepo & al. 2015). After the

addition of four Cordana species by Ai & al. (2019) and Luo & al. (2019),

Cordana currently contains 20 recognized species, of which only ten have

been reported from China: C. abramovii Seman & Davydkina, C. aquatica

Z.L. Luo & al., C. ellipsoidea de Hoog, C. lignicola Z.L. Luo & al., C. lithuanica

Markovsk., C. lushanensis C.C. Ai & al., C. meilingensis C.C. Ai & al.,

C. pauciseptata, C. uniseptata L. Cai & al., and C. terrestris (Timonin)

Hern.-Restr. & al. (Lu & al. 2000, Cai & al. 2004, Zhang 2018, Ai & al. 2019,

Luo & al. 2019).

In the past, Cordana was associated with Trichosphaeriaceae (Miller

& Samuels 1982) and Chaetosphaeriaceae (Réblova & al. 1999) based on

ascomatal morphology.

Saprobic hyphomycetes are highly diverse on dead branches, and much

taxonomic information has been published in China (e.g., Wu & Zhuang

2005; Ma & al. 2014, 2015, 2016; Ma & Zhang 2015; Xia & al. 2015, 2016;

Ma 2016a,b; Xu & al. 2017, 2019; Zhang 2018; Qiu & al. 2020). In our study

of these fungi from Jinggangshan Mountain, Jiangxi Province, an interesting

hyphomycete with the morphological features typical of Cordana was

collected on dead branches. It shows significant differences from previously

described Cordana species and therefore is proposed as new.

Materials & methods

Dead branches were collected from humid environments and watersides in

the forest ecosystems of southern China and placed in Ziploc™ plastic bags for

transport to the laboratory, where they were processed, and examined as described by

Ma & al. (2011). Microphotographs were prepared using a Nikon Eclipse E200 and

SmartV550Dc digital camera, with a 100 x (oil immersion) objective at the same

background and scale. Conidia were measured at their widest point. The minimum

and maximum values for microscopic measurements are provided, with outliers in

parentheses. Adobe Photoshop 7.0 was used to assemble photographs into plates. The

specimens are deposited in the Herbarium of Jiangxi Agricultural University, Plant

Pathology, Nanchang, Jiangxi, China (HJAUP).

Cordana sinensis sp. nov. (China) ... 555

s A ; B

g 4

Fic. 1. Cordana sinensis (holotype, HJAUP M0172).

A-C. Conidiophores and conidia; D. Conidia.

556 ... Qiu & al.

Taxonomy

Cordana sinensis L. Qiu, K. Zhang, R.F. Castafieda & Jian Ma, sp. nov. FIG. 1

MB 836854

Differs from Cordana crassa and C. lithuanica by its ovoid to obclavate, medially

1-septate or with 2-septate, narrower conidia.

Type: China, Jiangxi Province: Jinggangshan Mountain, Jingzhushan scenic spot,

26°32’28’”N 114°06’57”E, on dead branches of an unidentified broadleaf tree, 6 Nov.

2014, J. Ma (holotype, HJAUP M0172).

EryMo_oey: refers to China, where the fungus was collected.

COLONIES on the natural substratum effuse, brown to dark brown. Mycelium

partly superficial, partly immersed, composed of branched, septate, smooth,

subhyaline to pale brown hyphae. CoNIDIOPHORES macronematous,

mononematous, simple or branched, erect, straight to flexuous, smooth,

septate, brown to pale brown, cylindrical, sub-nodose, <400 um long,

4-6.5 um wide, with terminal and intercalary nodes, 5-7.5 um wide.

CONIDIOGENOUS CELLS integrated, polyblastic, terminal and intercalary,

cylindrical-subnodose, with tiny prominent denticles at the nodes, percurrent

elongated, 9.5-32 x 4-6.5 um. Conidial secession schizolytic. CONIDIA

solitary, acropleurogenous, ovoid to obclavate, 1-2-septate, pale brown,

smooth, thick-walled, 13-20(-26.5) x 5-7.5 um, dry, apical cell rounded,

basal cell with a tiny prominent hilum.

COMMENTS —Among the known species of Cordana, C. sinensis most closely

resembles C. crassa Toth and C. lithuanica Markovsk. in conidial shape.

However, C. crassa and C. lithuanica produce asymmetrically 1-septate

conidia while those of C. sinensis are medially 1-2-septate. In addition,

C. crassa produces larger (20-24 um long), ovoid conidia (Toth 1975), and

C. lithuanica produces shorter and wider (13-17.5 x 8-10 um) obpyriform

conidia (Markovskaja 2003).

With the addition of C. sinensis, Cordana is represented by 21 species

(TaBLE 1). All these species have integrated, polyblastic, terminal or

intercalary, sympodial conidiogenous cells, except C. bisbyi (Timonin)

Hern.-Restr. & al. and C. terrestris (Timonin) Hern.-Restr. & al., which bear

discrete, polyblastic conidiogenous cells umbellately arranged at the apex of

the conidiophores (Hernandez-Restrepo & al. 2014). Cordana conidia are

mostly smooth-walled and concolorous, but C. indica Subramon. & V.G.

Rao and C. verruculosa Hern.-Restr. & al. produce echinulate or verruculose

conidia. Cordana inaequalis S. Hughes, C. mercadoana Hern.-Restr. & al.,

557

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C. parasitica Togashi & Onuma, and C. uniseptata L. Cai & al. produce

versicolored conidia. Only five species produce nonseptate conidia: C. bisbyi,

C. lushanensis C.C. Ai & al., C. semaniae Davydkina & al., C. solitaria V. Rao

& de Hoog, and C. verruculosa, while most form 1-septate conidia, although

in C. lignicola Z.L. Luo & al. and C. sinensis, 2-septate conidia are occasionally

present. All 21 species except C. parasitica are usually found on rotten wood

or dead branches, or in soil.

Key to Cordana accepted species

1. Conidiogenous cells discrete, polyblastic, umbellately arranged over the swollen

apex of the conidiophores ......... 00... cece eee eee ene eee eee eee ee 2

1. Conidiogenous cells integrated, polyblastic,

intercalary or terminal on the conidiophores ................... ec eee eens 3

2. Conidia 0-septate, cylindrical-oval, 5-7.2 x 2.5-3.6 um .............004. C. bisbyi

2. Conidia 1-septate, oblong to ovate, 6-11 x 3-4um ..... ee... C. terrestris

B COMIGTa DOMSCD ALG nang on tare tet ated tober tan ta eed Cece tee Gaetan Cae eG cep et wee 4

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4. Conidia verruculose, 3-5.5 X 2-3.5 UM ....... eee eee eee eee eee C. verruculosa

4Conidiasmocthy:25:5-UnPlOng.. 64 vox nebo eens toad het a hearse tye Pema oon 5

5. Conidia obovoid, guttulate, 21-27 x 9-15um....... eee C. semaniae

SRC Orig 9 25K Py MT Se Yoon tas a Viner cs inate Sev Gee Pung cee Meceg eee deere Me tre Seelgeen Mule es 6

6. Conidia ellipsoidal to obovoid, 5.5-8 x 2.5-4um...... 22. eee eee C. lushanensis

6. Conidia obovoidal, 6.5-9.5 X 4.5-6.5 UM ....... ee eee eee eee ee eee C. solitaria

7. Conidia oval to oblong, echinulate, 11-20 x 4.5-7um ..............0.. C. indica

7.Conidia‘smooth,-variably.sized*-. Stn; ces \ bas8 seas Gases Paes Cake ees we 8

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8. GOMIdaCONCOLOTGUS «22g 8 ash ae apn ee daile ce dott nee dah ae dah aae dosh ote dash he dphens 12

9. -Gonidia ellipsoidal or Gvoid.:. i158 sesh ekak de Loh bn Vo de Peed OS SE Oh oe Es 10

9. Conidia oblong, obovoid, or cylindrical ....... 0... eee eee eee 11

10. Conidia mostly broadly ellipsoidal, 13.5-23 x 8.5-11.5 um ......... C. uniseptata

10. Conidia narrowly ellipsoidal to ovoid, 10-12.5 x 4-5 um ........... C. inaequalis

11. Conidia oblong, only 1-septate, 6-10 x 3-5 um .... eee eee. C. parasitica

11. Conidia oblong, obovoid or cylindrical, 0-1-septate,

GAO: xe SeA IVI a cPak and acon deh shod ts poe duh FONE Sy Pace Sh Gael ak eS C. mercadoana

12-Conidia-0-2-or 12) =septate A. Saks, Sie ree tory trees trues eres ree 13

12Conidia-only Taséptatert 108.0 sable jcte x gun tints bntned Hele bslnitt baie pee oe 14

13. Conidia mostly oblong, sometimes ellipsoid, 0-2-septate,

OV XB, 54 Salih, § BY, vB GNM ue Oe A ENA A i RA BN 1, BI, 27. BN, C. lignicola

. Conidia ovoid to obclavate, 1-2-septate, 13-20 x 5-7.5um ........... C. sinensis

—

ies)

560 ... Qiu & al.

14. Conidia of both cells more or less equally wide ............ 0... e eee eee eee 15

14. Conidia of both cells unequally wide ......... 0... cece eee eee eee ee 18

15. Conidia ovoid, with a pigmented septal pore,

VSS EZ S PUM aa dt gala Bittlgel gs dik gee a dog sce ge 8 aytee-o 8 Atse wae! C. abramovii

15. Conidia oblong, cylindrical, ellipsoid to oblong or ellipsoid to ovoid,

lacking pigmented septal pore, <13 X 7 UM ...... eee eee eee 16

16. Conidia ellipsoid to oblong, 8.7-11.5 x 4.3-4.7 um .............04. C. ellipsoidea

16-CWonidia SS lin widesss, \cskeie tek Apa ees REN do PN Ay 2B son Lobe es 17

17. Conidia oblong or cylindrical, 10-13 x 5.5-7 um................ C. meilingensis

17. Conidia broadly ellipsoid to ovoid, 8-12 x 5-7 um .............. C. pauciseptata

18. Conidia ovoid, 20-24 x 13-15 Um 2... eee eee eens C. crassa

18. Conidia obpyriform, obovoid, ellipsoid or oblong, <19 x 13 um .............. 19

19. Conidia mostly oblong, sometimes ellipsoid, 8-10 x 3-5 um......... C. aquatica

19F Conidia 213 28 [Mh 5... eee y cep iee we sohad te wheel orcad vhs iclyadl apie Galea es 20

20. Conidia obovoid to ellipsoid, 17-19 x 10-13 um ........... C. andinopatagonica

20. Conidia obpyriform, 13-17.5 x 8-10 um ....... eee eee C. lithuanica

Acknowledgments

The National Natural Science Foundation of China (Nos. 31970018, 31360011)

supported this project. The authors express gratitude to Dr. Rosa Maria Arias

Mota (Instituto Tecnolégico Superior de Xalapa, Mexico) and Dr. De-Wei Li (The

Connecticut Agricultural Experiment Station, USA) for serving as pre-submission

reviewers and to Dr. Lorelei L. Norvell for final editorial review and Dr. Shaun

Pennycook for nomenclatural review.

Literature cited

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spp. nov. from Jiangxi, China. Mycotaxon 134: 329-334. https://doi-org/10.5248/134.329

Cai L, McKenzie EHC, Hyde KD. 2004. New species of Cordana and Spadicoides from decaying

bamboo culms in China. Sydowia 56: 222-228.

Davydkina TA, Mel'nik VA. 1989. Two new hyphomycetes from the genera Cordana and

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MY COTAXON

July-September 2021— Volume 136, pp. 563-577

https://doi.org/10.5248/136.563

Pezicula endophytica sp. nov.,

endophytic in Dendrobium in Thailand

X1A0-YA MA**3, JI-CHUAN KANG", KEvIN D. HyDE”?*,

MINGKWAN DOILOM?*>, PUTARAK CHOMNUNTI”?

"Engineering & Research Center for Southwest Biopharmaceutical Resource of

National Education Ministry of China, Gui Zhou University,

Gui Yang 550025, Gui Zhou Province, Peoples Republic of China

*Center of Excellence in Fungal Research, Mae Fah Luang University,

Chiang Rai Province, 57100, Thailand

*School of Science, Mae Fah Luang University,

Chiang Rai Province, 57100, Thailand

‘Institute of Plant Health, Zhong Kai University of Agriculture & Engineering,

Haizhu District, Guang Zhou Province, 510225, P.R. China

°Key Laboratory for Plant Diversity & Biogeography of East Asia,

Kun Ming Institute of Botany, Chinese Academy of Science,

Kun Ming City, 650201, Yun Nan Province, Peoples Republic of China

* CORRESPONDENCE TO: jckang@gzu.edu.cn

ABSTRACT—A new species, Pezicula endophytica, was isolated from roots and stems of two

Dendrobium species in northern Thailand. Evidence to support the new species is based

on morphology and phylogenetic analysis of the combined ITS, LSU, and RPB2 DNA

sequence dataset. Pezicula endophytica, which constituted a clade independent from other

Pezicula species, has 4% distinct base pair differences in all genes. Pezicula endophytica

has larger macroconidia and longer conidiophores compared with phylogenetically

neighboring species. This is the first report of an endophytic Pezicula species from

Dendrobium in Thailand.

KEY wWoRDS— ascomycetes, Cryptosporiopsis, Dermateaceae, multi-loci, Orchidaceae

Introduction

Fungal endophytes in orchids may be symbionts that help plant growth

and are possible alternative sources for the bioactive compounds produced

564 ... Ma & al.

in plants (Bungtongdee & al. 2019, Ma & al. 2015, Vujanovic & al. 2000).

They also play a role as mycorrhizae in orchids and transfer water and

nutrition to the host (Bungtongdee & al. 2019, Ma & al. 2015, Smith & Read

2010). Pezicula Tul. & C. Tul. (Dermateaceae; anamorph: Cryptosporiopsis

Bubak & Kabat) is apothecial (Ekanayaka & al. 2017, 2019). The name

Pezicula has been proposed as the accepted name because Cryptosporiopsis

species are the asexual morphs of both Pezicula and Neofabraea H.S. Jacks.

(Chen & al. 2016; Johnston & al. 2014; Verkley 1999; Verkley & al. 2003;

Wijayawardene & al. 2017a,b). Pezicula is characterized by brightly colored,

short-stalked apothecia with circular discs that are initially concave and

surrounded by slightly elevated margins but later become convex. They

occur on the bark of woody plants and are involved in wood degradation

(Kowalski 1996). Asci are inoperculate and mostly clavate or cylindric-

clavate, ascospores vary in shape from broadly ellipsoid to elongate-

ellipsoid to allantoid or fusoid (Ekanayaka & al. 2016, Ooki & al. 2003,

Verkley 1999, Verkley & al. 2003). The asexual morph of Pezicula species

produces sporodochial conidiomata, with macro- and microconidia

formed on different conidiophores (Ekanayaka & al. 2016, Verkley 1999).

Pezicula species are recorded as endophytes, plant pathogens, and saprobes

from a wide range of hosts (Chen & al. 2016, Wijayawardene & al. 2017a).

They have been isolated as endophytes from Dendrobium nobile Lindl. and

Holcoglossum wangii Christenson (Orchidaceae) in China (Chen & al. 2011,

Tan & al. 2012).

Dendrobium Sw. is one of the largest genera in Orchidaceae (https://www.

kew.org/), and many species are important medicinal and ornamental plants

(Ma & al. 2015). However, few reports concern endophytic Pezicula species in

Dendrobium (Chen & al. 2011, Tan & al. 2012).

In this study, we isolated four endophytic fungal strains from two

different, undetermined Dendrobium species sampled in northern

Thailand. The strains were identified as representing a new species, Pezicula

endophytica, based on morphological and phylogenetic analyses. ‘This is the

first report of an endophytic Pezicula in Dendrobium in Thailand.

Materials & methods

Sample collection

Healthy roots and stems of two different but undetermined Dendrobium species

(sp.1 and sp.2) were collected from outside the temple of Wat Phra That Doi Tung

(Temple of Doi Tung Pagoda), Mae Fah Luang district, Chiang Rai Province,

Pezicula endophytica sp. nov. (Thailand) ... 565

Thailand. Fresh material was kept fresh by placing into Zip-lock bags or tubes

containing silica gel on ice.

Fungal isolation & cultivation

The material was processed within 48 hours. Surface sterilization followed

Nontachaiyapoom & al. (2010) with minor modifications. Healthy roots and stems

were washed in tap water and then immersed for 5 minutes in a solution containing

3% (v/v) H,O, and 70% (v/v) ethanol, and then rinsed with three changes of sterile

distilled water. The sterilized materials were cut into 2 mm?’ pieces and placed

on potato dextrose agar (PDA, Difco, BBL/USA’ #213400) containing 50 ug/ml

oxytetracycline, 50 ug/ml penicillin, and 50 ug/ml streptomycin (Otero & al. 2002).

The surface sterilization method was checked, as described by Petrini (1991). Petri

dishes were incubated at 28 °C under natural light and observed daily; mycelia on

the edge of developing fungal colonies were transferred to fresh PDA to obtain pure

cultures. The pure cultures were deposited in Mae Fah Luang University Culture

Collection, Chiang Rai, Thailand (MFLUCC). Dry cultures of the new species were

deposited in the herbarium of Mae Fah Luang University, Chiang Rai, Thailand

(MFLU).

Morphological analysis

Pure cultures were cultivated on both PDA and water agar (WA) media in a dark

cabinet at room temperature (28 °C) and observed every week. Sterilized toothpicks

were placed on the WA surface to encourage fungal sporulation. The growth rate was

evaluated after mycelia had nearly covered the whole agar surface. Any observable

structures were examined using a Zeiss SteREO Discovery.V8 stereomicroscope.

The colonies were examined for morphological characteristics and structures were

removed and mounted in water. The mounted slides were observed using a Nikon

EOS 700D compound microscope. All observed structures were measured with

Image Frame Work v.0.9.7 and the photo plates were assembled using Photoshop

CS 6.0.

DNA extraction & amplification

Genomic DNA was extracted from mycelium of endophytic fungi using an

EZgene Fungal gDNA Kit following the manufacturer's instructions. The selected

genes for amplification were partial nuclear rDNA internal transcribed spacer (ITS),

partial large subunit (LSU), and partial RNA polymerase II large subunit 2 (RPB2).

Each 25 ul amplification reaction contained 12.5 ul of 2*Bench TopTM Taq Master

Mix (0.05 units/ul Taq DNA polymerase, 0.4 mM dNTPs and 4mM MgCl); 2 ul

forward and reverse primers; 1 ul of DNA template and 9.5 ul of threefold-distilled

water. The primers and PCR protocol followed Chen & al. (2016). PCR products

were visualized on 1% agarose gel stained with Realtimes Biotech Goldview and then

sent to Sangon Biotech for purification and sequencing. All DNA sequences from

this study were submitted to GenBank.

566 ... Ma &al.

TABLE. 1. Fungal strains and sequences of Pezicula and related taxa sourced from the

present study, BLAST searches in GenBank, or Chen & al. (2016).

GENBANK ACCESSION No.

eee aan ITS LSU RPB2

Neofabraea. inaequalis CBS 326.75 KR859081 KR858872 KR859321

N. krawtzewii CBS 102867 KR859084 KR858875 KR859324

N. malicorticis CBS 102863 KR859085 KR858876 KR859325

CBS 122030 KR859086 KR858877 KR859326

Parafabraea caliginosa CBS 124806 KR859090 KR858881 KR859330

Pa. eucalypti CBS 124810 KR859091 KR858882 KR859331

Pezicula acericola CBS 239.97 KR859093 KR858884 KF376214

CBS 245.97 KR859098 KR858889 KF376213

P. aurantiaca CBS 201.46 KR859102 KR858893 KF376210

P. brunnea CBS 120291 KR859103 KR858894 _—

P. californiae CBS 124805 KR859104 KR858895 KR859332

P. carpinea CBS 923.96 KR859108 KR858899 KF376158

CBS 921.96 KR859107 KR858898 KF376159

P. chiangraiensis MFLUCC 15-0170 KU310621 KU310622 KU310623

P. cinnamomea CBS 626.96 KR859152 KR858944 KF376162

CBS 239.96 KR859124 KR858915 KF376165

P. cornina CBS 285.39 KR859163 KR858955 KR859333

P. corticola CBS 259.31 KR859164 KR858956 _—

CBS 260.31 KR859165 KR858957 _

P. corylina CBS 140.22 KR859166 KR858958 KR859334

CBS 249.97 KR859168 KR858960 KF376161

P. diversispora CBS 185.50 KR859170 KR858962 _

CBS 282.47 KR859171 KR858963 _—

P. endophytica MFLUCC 14-0116 MN908669 MN908642 MT371880

MFLUCC 14-0118 MN908666 MN908640 _—

MFLUCC 14-0140 MN908667 MN908641 _—

MFLUCC 14-0144 MN908668 = =

P ericae CBS 120290 KR859173 KR858965 _—

CBS 120292 KR859174 KR858966 _

P. eucrita CBS 259.97 KR859179 KR858971 KF376205

CBS 656.96 KR859185 KR858977 KF376208

P. fagacearum CBS 112400 KR859201 KR858993 KR859335

CBS 112402 KR859203 KR858995 KR859336

P._ frangulae subsp. frangulae CBS 100244 KR859204 KR858996 KF376211

CBS 778.96 KR859209 KR859001 KF376212

P. heterochroma CBS 199.46 KR859210 KR859002 —

P. melanigena CBS 898.97 KR859211 KR859003 _

P. microspora CBS 124641 KR859212 KR859004 KR859337

P. neocinnamomea CBS 100248 KR859213 KR859005 KF376209

P. neoheterochroma CBS 127388 KR859221 KR859013 KR859338

Pezicula endophytica sp. nov. (Thailand) ... 567

GENBANK ACCESSION No.

SPECIES STRAIN

ITS LSU RPB2

P. neosporulosa CBS 101.96 KR859223 KR859015 KF376193

CBS 102.96 KR859224 KR859016 KF376181

P. ocellata CBS 268.39 KR859232 KR859024 KR859339

CBS 949.97 KR859233 KR859025 KF376215

P. pruinosa CBS 292.39 KR859234 KR859026 —

P. pseudocinnamomea CBS 101000 KR859235 KR859027 KR859340

P. querciphila CBS 134525 JX144750 — —

P. radicicola CBS 640.94 KR859236 KR859028 —

CBS 681.83 KR859237 KR859029 —

P rubi CBS 253.97 KR859250 KR859042 KF376204

CBS 593.96 KR859253 KR859045 KF376203

P. sporulosa CBS 224.96 KR859261 KR859053 KF376201

CBS 225.96 KR859262 KR859054 KF376202

Phlyctema vagabunda CBS 109875 KR859275 KR859069 KR859346

CBS 304. 62 KR859276 KR859070 KR859347

Rhizodermea veluwensis CBS 110605 KR859282 KR859076 KR859353

CBS 110615 KR859283 KR859077 KR859354

*Ex-type, ex-epitype or other type strains are in bold

Sequence analysis

The sequencing results were assembled using DNASTAR SeqMan v.7.1.0. The

assembled sequences were subjected to nucleotide BLAST searches in GenBank

(www.ncbi.nlm.nih.gov/BLAST) for establishing highly similar strains (Altschul & al.

1997). Supplemental strains of Pezicula and related genera were selected from Chen

& al. 2016, especially ex-type and ex-epitype sequences where available. Sequence

alignment was carried out by MAFFT v.7.0 (https://mafit.cbrc.jp/alignment/server/).

Misaligned sequences were manually edited using AliView v.1.26 (Larsson 2014).

The three loci were combined into a concatenated dataset with SequenceMatrixa

1.7.8 (Vaidya & al. 2011).

The best fitting nucleotide substitution models for each gene were estimated by

IQ-TREE (Nguyen & al. 2014) with ultrafast bootstrap (Hoang & al. 2017). Gaps

were regarded as missing data. Both maximum likelihood (ML) and Bayesian (BI)

trees were constructed on The CIPRES Science Gateway V. 3.3 (http://www.phylo.

org/index.php/). The ML tree was constructed using RAxML-HPC2 on XSEDE.

Maximum likelihood bootstrap values (MLSB) were calculated with 1000 iterations.

The generalized time-reversible (GTR) was set with gamma distribution. The BI

tree was implemented by MrBayes on XSEDE (3.2.6). Two sets of four simultaneous

independent chains of Markov chains Monte Carlo (MCMC) simulations were run

for 5,000,000 generations; 25% of trees were discarded and the remaining trees were

used to calculate the Bayesian posterior probabilities (PP). Convergence was assumed

568 ... Ma &al.

when the standard deviation of split sequences was less than 0.01. The generated

trees were submitted to TreeBase (http://purl.org/phylo/treebase/phylows/study/

TB2:S25576). The resulting trees were viewed with Figtree v.1.4.0. The layout was

made with Adobe Illustrator CS 6.

Results

Fungal isolation & sporulation

Four endophytic Pezicula strains were isolated from healthy roots and

stems of Dendrobium species. Isolate MFLUCC 14-0116 sporulated on PDA

after one month and on WA with toothpicks after two months. The conidia,

conidiomata, conidiophores and mycelia were recorded from cultures.

Phylogenetic results

Sequences of the isolated Pezicula and reference sequences of related taxa

are listed in TABLE 1. Combined data for the phylogenetic tree comprised 57

sequences containing 2478 sites (including 582 ITS sites, 843 LSU sites, and

1053 RPB2 sites). The selected nucleotide substitutional model for ITS and

RPB2 was a symmetrical model (SYM) with gamma distribution, and for LSU

was Kimura's two-parameter substitution model with invgamma distribution.

The phylogenetic tree (Fic. 1) was rooted with two selected outgroup

species Parafabraea caliginosa (CBS 124806) and P. eucalypti (CBS 124810)

with robust bootstrap support (ML = 100%, PP = 1). In the phylogenetic tree

(RAxML), Pezicula endophytica strains clustered and formed an independent

clade. Strains MFLUCC 14-0140, MFLUCC 14-0144 and MFLUCC 14-0116

clustered with MFLUCC 14-0118 (ML = 100%, PP = 1). Pezicula aurantiaca,

P. cornina and P. pseudocinnamomea formed a clade close to P. endophytica.

The neighboring genera Neofabraea H.S. Jacks., Phlyctema Desm. and

Rhizodermea Verkley & Zijlstra were well separated from Pezicula

(ML = 94%, PP = 1).

Pezicula endophytica (MFLUCC 14-0116, ex-type) differs from Pezicula

diversispora (CBS 185.50, ex-type) in 4% of base pairs [2.9% in ITS (17/582bp),

1.1% in LSU (9/843bp)] and from Pezicula cornina (CBS 285.39) in 8.6% of

base pairs [3.3% in ITS (19/582bp); 1.5% in LSU (13/843bp); 3.8% in RPB2

(40/1053bp)].

Fic. 1. The consensus phylogram resulting from a RAxML analysis of the combined three loci

(ITS-LSU-RPB2) of Pezicula and related taxa. Parafabraea caliginosa and P. eucalypti were

selected as outgroup taxa. Isolates generated in this study are labelled with *. Holotype and

ex-type isolates are in bold. Support values from maximum likelihood (MLBS) and Bayesian

posterior probabilities (PP) are labelled at the end of nodes (50% majority rule). Dashes indicate

values <50%. Scale bar represents 0.04 substitutions per site.

Pezicula endophytica sp. nov. (Thailand) ...

9s/lnj Pezicula sporulosa CBS 224.96

Pezicula sporulosa CBS 225.96

Pezicula querciphila CBS 134525

|) Pezicula neosporulosa CBS 101.96

Pezicula neosporulosa CBS 102.96

100/K.; Pezicula corticola CBS 259.31

“een Pezicula corticola CBS 260.31

“A iN Pezicula cinnamomea CBS 626.96

Pezicula cinnamomea CBS 239.96

| Pezicula chiangraiensis MFLUCC 15-0170

ae ! | Pezicula eucrita CBS 259.97

een \ Pezicula eucrita CBS 656.96

“0.7 (| Iq Pezicula rubi CBS 593.96

Pezicula rubi CBS 253.97

Pezicula neocinnamomea CBS 100248

- " Ky Pezicula diversispora CBS 185.50

7, Pezicula diversispora CBS 282.47

86/0.94e Pezicula endophytica MFLUCC 14-0140*

eye [Ny Pezicula endophytica MFLUCC 14-0144*

“ACS! Pezicula endophytica MFLUCC 14-0116*

Pezicula endophytica MFLUCC 14-0118*

ore i Pezicula cornina CBS 285.39

Pezicula aurantiaca CBS 201.46

Pezicula pseudocinnamomea CBS 101000

10.87 Pezicula microspora CBS 124641

Pezicula radicicola CBS 640.94

| Pezicula radicicola CBS 681.83

Pezicula melanigena CBS 898.97

pice’ | Pezicula ericae CBS 120292

“ '\’ Pezicula ericae CBS 120290

Pezicula acericola CBS 245.97

Se Pezicula acericola CBS 239.97

Pezicula pruinosa CBS 292.39

_ Iq Pezicula ocellata CBS 268.39

roaict Pezicula ocellata CBS 949.97

Pat [s Pezicula frangulae subsp. frangulae CBS 100244

N ' Pezicula frangulae subsp. frangulae CBS 778.96

Pezicula neoheterochroma CBS 127388

Pezicula brunnea CBS 120291

34/0.94,|] 12°.) Pezicula corylina CBS 249.97

1094. | Pezicula corylina CBS 140.22

Rj LXy Pezicula fagacearum CBS 112400

98/1

65/0.99

100/1 =

54/0.61

99/1

80/1 A Pezicula fagacearum CBS 112402

ioo,| S— Pezicula heterochroma CBS 199.46

Xa PQ Pezicula carpinea CBS 921.96

Pezicula carpinea CBS 923.96

Pezicula californiae CBS 124805

00/1, | Rhizodermea veluwensis CBS 110615

ae Rhizodermea veluwensis CBS 110605

100/In) Phlyctema vagabunda CBS 304.62

a Phlyctema vagabunda CBS 109875

10/1 Neofabraea krawtzewii CBS 102867

1001+; Neofabraea malicorticis CBS 122030

Toso |f Neofabraea malicorticis CBS 102863

Neofabraea inaequalis CBS 326.75

10/1, ¢ Parafabraea eucalypti CBS 124810 Outgroup

Parafabraea caliginosa CBS 124806

0.04

569

570 ... Ma & al.

Taxonomy

Pezicula endophytica X.Y. Ma, K.D. Hyde & J.C. Kang, sp. nov. Fic. 2

IF 558634

Differs from Pezicula diversispora by its longer conidiophores and narrower

macroconidia.

Type— Thailand, Chiang Rai Province, Mae Fah Luang district, outside Temple of

Doi Tung Pagoda, endophytic in the roots of Dendrobium sp. 1, 19 December 2013,

Nontachaiyapoom S, Aewsakul N & Ma XY (holotype, MFLU 17-2789; ex-type living

culture, MFLUCC 14-0116; isotype, MFLU 20-0442; ex-type living culture, MFLUCC

14-0118.

EryMOLoGyY—in reference to its endophytic habit.

SEXUAL MORPH undetermined. ASEXUAL MORPH observed on both PDA and

(with toothpicks) WA.

Cotoniges on WA superficial, white with light brownish grey mycelia,

sparse, irregular edge, fluffy, reverse light brown, growth rate 0.8 mm/day.

Mycettium hyaline to light brown, smooth, 2.5-4.4 um diam. CONIDIOMATA

on toothpicks on WA aggregated or scattered, with aerial and fluffy hyphae,

yellow-grey, globose.

CoLoniges on PDA medium superficial, white and brown in the center,

lobate edge, dense, fluffy, concentric ring close to the edge, reverse brown,

growth rate: 1.2 mm/day. CoNIDIOMATA erumpent on surface, aggregated

or scattered, with irregular sporodochia, with brown to milky white conidial

masses, globose to irregular. MACROCONIDIOPHORES (16-)22-30(-40) x

(2-)3-4(-5.5) um (x = 26 x 3.5 um, n=5), arising from the inner wall layers

of conidiomata, hyaline, cylindrical, straight to flexuous, septate, unbranched

or branched, smooth-walled. MacrocoNIDIOGENOUS CELLS hyaline,

enteroblastic, phialidic, determinate, proliferating percurrently, sometimes

with a minute collarette, cylindrical, integrated, determinate, smooth-walled.

MACcROCONIDIA (30-)31-43(-44) x (5-)7-9 (-10) um (x = 37 x 8 um, n=16),

L/W =4.4, hyaline, elongated ellipsoid, rounded at the apex, narrow and slightly

truncate at the base, sometimes attenuated and with a protruding scar at the

base, septate when mature, straight, smooth-walled, with guttules.

ADDITIONAL SPECIMENS EXAMINED— THAILAND, CHIANG RAI PROVINCE, Mae Fah

Luang district, outside Temple of Doi Tung Pagoda, endophytic in roots and stems of

Dendrobium sp. 2, 19 December 2013, Nontachaiyapoom S, Aewsakul N & Ma XY.

(MFLUCC 14-0140, GenBank MN908667, MN908641; MFLUCC 14-0144, GenBank

MN908668).

CoMMENTS—Pezicula endophytica formed an independent clade differing

from other species in the phylogenetic tree, and is adjacent to P. aurantiaca,

Pezicula endophytica sp. nov. (Thailand) ... 571

Fic. 2. Pezicula endophytica (ex-holotype MFLUCC 14-0116). a, b. Colony on PDA (a. top;

b. reverse). c. Colony on WA with toothpicks. d, e. Conidiomata on toothpicks on WA.

f-h. Conidiomata with conidial masses on PDA. i-n. Macroconidiophores, macroconidiogenous

cells and macroconidia. o-r. Macroconidia. Norte: i-r from PDA. Scale bars: d~h = 500 um;

i-n, p, q = 10 um; 0, r= 5 um.

572... Ma &al.

P. cornina, P. diversispora, and P. pseudocinnamomea. Pezicula endophytica

is distinguished from those species by its longer macroconidiophores

and narrower macroconidia with septa (P aurantiaca and P. cornina are

aseptate; Chen & al. 2016, Robak 1950, Verkley 1999). Pezicula endophytica

(ex-type MFLUCC 14-0116) has >4% different gene sequences than

P. diversispora (ex-type CBS 185.50). Compared with their cultures on MEA,

Pezicula endophytica produces no diffusing pigment on WA (Verkley 1999).

Pezicula aurantiaca, P. cornina, P. diversispora, and P. pseudocinnamomea

were isolated from dead branches, bark, and twigs of dicotyledonous woody

plants from European countries (Chen & al. 2016, Robak 1950, Verkley 1999),

whereas P. endophytica is an endophytic fungus from monocotyledonous

Dendrobium orchids.

Discussion

The taxonomy and identification of Pezicula is complicated because

many species have been introduced based on single-morph taxa, some

lack molecular data, and their morphology may vary depending on the

growing medium (Verkley 1999, Verkley & al. 2003). Of the 136 species

listed in Pezicula (http://www.speciesfungorum.org), only 29 (including

P. endophytica) have DNA sequence data available in the 2021 NCBI

database (Yuan & al. 2015, Ekanayaka & al. 2016; http://www.ncbi.nlm.nih.

gov/). A revision of the genus based on comprehensive morphological and

phylogenetic data is necessary. The asexual morphs of most Pezicula species

produce both macro- and microconidia on the natural substrata and in

cultures (Verkley 1999). In this study, only macroconidia were observed

in cultures. Macroconidia of P endophytica resemble P. diversispora, which

differs by its longer conidiophores, narrower macroconidia, and a 4% gene

sequence difference. Pezicula endophytica, which only produced its asexual

morph in cultures, differs from P carpinea (the type species) by lacking

conidia along the conidiophore sides and large gene sequence differences.

Presumably P. endophytica microconidia will not be produced on WA after

two sporulation months.

Chen & al. (2011) first reported an endophytic Pezicula sp. from

Dendrobium orchids in China. Pezicula endophytica was the first Pezicula

endophyte identified in Thailand. Pezicula species are frequently isolated

endophytes in angiosperms, particularly in roots of Abies, Calluna, Erica,

Larix, Pleurothallis, Pseudorchis, Stelis, and Vaccinium (Chen & al. 2016,

Devi & Joshi 2014, Herrera & al. 2010, Kohout & al. 2013, Noble & al. 1991,

Pezicula endophytica sp. nov. (Thailand) ... 573

Schulz & al. 2002, Strobel & al. 1999, Talontsi & al. 2012, Verkley & al. 2003,

Wang & al. 2014, Yuan & Verkley 2015, Zilla & al. 2013). They have also

been found in peridermal bark of living branches of temperate trees such as

Abies, Acer, Alnus, Betula, Carpinus, Castanea, Fagus, Fraxinus, Picea, Pinus,

Quercus, and Sequoia (Espinosa-Garcia & Langenheim 1990, Kowalski &

Kehr 1992, Matsumura & al. 2013, Petrini & Miller 1979, Schulz & al. 1995,

Sieber 1988, Sieber & Dorworth 1994, Verkley 1999).

Although many Pezicula species are reported from temperate and

boreal forests of the northern hemisphere, such as Canada, Germany, and

the Netherlands (Kowalski & Kehr 1992, Chen & al. 2016, Verkley 1999),

few geographic and climatic limitations have been observed. Saprobic

and pathogenic Pezicula species have been recorded from temperate to

tropical and Mediterranean areas (Bergero & al. 2003, Cheewangkoon &

al. 2010, Old & al. 2002). Pezicula eucalyptigena was isolated from leaves

of Eucalyptus sp. in the mediterranean Western Cape, South Africa, which

has smaller conidia ((23—)24—27(-30) x (6-)7(-8) um; Crous & al. 2019)

than those of P endophytica. Ekanayaka & al. (2016) described a sexual

morph from Thailand, P chiangraiensis, saprobic on the bark of decaying

wood. Pezicula ericae was found as ericoid mycorrhizal fungi in the roots

of velvetleaf blueberry (Vaccinium myrtilloides) seedlings in Canada,

which could enhance development of the host plant (Mu 2020). For most

endophytic Pezicula species, their roles remain unknown.

Acknowledgments

We sincerely thank the National Natural Science Foundation of China (NSFC

grants 31670027 & 31460011) and the Open Research Foundation of the Key

Laboratory (Engineering Center) of National Education Ministry at Gui Zhou

University (grant GZUKEY 20160705). We also thank Thailand Research Funds for

the grant “Future of specialist fungi in a changing climate: baseline data for generalist

and specialist fungi associated with ants, Rhododendron species and Dracaena species

(grant DBG6080013)” for partly supporting this research. Mingkwan Doilom thanks

the 64th batch of China Postdoctoral Science Foundation (grant Y913082271) and

the 5th batch of Postdoctoral Orientation Training Personnel in Yun Nan Province

(grant Y934283261). We sincerely acknowledge Dr Sureeporn Nontachaiyapoom for

her guidance in collecting Dendrobium samples, identification, and endophytic fungal

isolation. We thank Dr. Eric H.C. McKenzie (Manaaki Whenua Landcare Research,

Auckland, New Zealand) and Dr. Joanne Taylor (Royal Botanic Garden Edinburgh,

Scotland, UK) for presubmission reviews. We acknowledge Shaun Pennycook for

his Latin name correction and Paul Kirk for his help in providing original papers

containing Pezicula species descriptions.

57A ... Ma &al.

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MY COTAXON

July-September 2021— Volume 136, pp. 579-586

https://doi.org/10.5248/136.579

Globoramichloridium delicatum and

Heteroconium simile spp. nov. from southern China

LING Qiu’, XU-GEN SHI’, XIU-GUO ZHANG’,

RAFAEL FE, CASTANEDA-RUuiIz?}, JI-WEN XIA’, JIAN Ma*4*

‘College of Agronomy, Jiangxi Agricultural University, Nanchang, Jiangxi 330045, China

? Department of Plant Pathology, 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

‘ Jiangxi Key Laboratory for Conservation and Utilization of Fungal Resources,

Jiangxi Agricultural University, Nanchang, Jiangxi 330045, China

*CORRESPONDENCE TO: majian821210@163.com jxaumj@126.com

ABSTRACT — Two new anamorphic fungi, Globoramichloridium delicatum and Heteroconium

simile, collected from dead branches of unidentified broadleaf trees in Jiangxi Province,

China, are described and illustrated. Globoramichloridium delicatum is characterized by

unicellular, subglobose to narrow subnapiform, yellowish-green conidia. Heteroconium simile

is distinguished by its monoblastic, integrated, terminal, determinate conidiogenous cells

that produce acrogenous, blastocatenate, dimorphic, 0-1-septate, smooth conidia.

KEY worps—Antennulariellaceae, Dissoconiaceae, Dothideomycetes, hyphomycetes,

taxonomy

Introduction

Saprobic dematiaceous hyphomycetes are highly diverse on rotting

woody plant materials (Xia & al. 2014, 2016). As essential components in

forest ecosystems, they play important roles in nutrient recycling, but their

geographical distribution and alpha taxonomy are poorly known. In order to

enrich the inventory of saprobic dematiaceous hyphomycetes in China, several

580 ... Qiu & al.

studies on fungi occurring on dead branches have been conducted during

the last 15 years (Zhang & Shi 2005; Ma & Zhang 2007, 2015; Ma & al. 2008,

2014, 2016; Zhang & al. 2009, 2014; Xia & al. 2012, 2015; Xu & al. 2017, 2019;

Qiu & al. 2020).

Jinggangshan Mountain is a small range in southeast Jiangxi, China. It lies

in the center part of Luoxiao Mountains, which border both Hunan and Jiangxi

provinces. Its scenic area covers c. 260 km? and exhibits complex geographical

and topographical conditions with an 81.2% forest coverage rate and a 14.2

°C annual average temperature. It is considered one of the world’s most

abundant biological areas within this latitudinal zone. A field trip was made to

Jinggangshan Mountain to collect saprobic hyphomycetes in November 2014.

Two new species with morphologic features typical of Globoramichloridium

Y. Marin & Crous (Marin-Felix & al. 2019; Dissoconiaceae) and Heteroconium

Petr. (Petrak 1949; Antennulariellaceae) were collected on dead branches.

The two new fungal species are described and illustrated below.

Materials & methods

Dead branches were collected from humid environments and watersides in the

forest ecosystems of Jinggangshan Mountain and placed in Ziploc™ plastic bags for

transport to the laboratory, where they were processed, and examined as described

by Ma & al. (2011). Microphotographs were prepared using a Nikon Eclipse E200

microscope equipped with a SmartV550Dc digital camera, with a 100x (oil

immersion) objective at the same background and scale. Conidia were measured at

their widest point. The range between minimum and maximum values for microscopic

measurements is given with outliers in parentheses. Adobe Photoshop 7.0 was used to

assemble photographs into plates. The specimens are deposited in the Herbarium of

Jiangxi Agricultural University, Plant Pathology, Nanchang, Jiangxi, China (HJAUP).

Taxonomy

Globoramichloridium delicatum L. Qiu, Jian Ma, R.F. Castafieda &

X.G. Zhang, sp. nov. FIG. 1

MB 836851

Differs from Globoramichloridium indicum by its larger unicellular subglobose to

narrow subnapiform yellowish green conidia.

Type: China, Jiangxi Province: Jinggangshan Mountain, Shuikou scenic spot, 26°33’03”N

114°07’55”E, on dead branches of an unidentified broadleaf tree, 6 Nov. 2014, J. Ma

(Holotype, HJAUP M0217).

Erymo_oey: Latin, delicatum, meaning “delicate”, referring to the dainty conidia.

CoLonigs on the natural substratum, effuse, brown. Mycelium superficial

and immersed composed of branched, septate, pale brown or brown,

Globoramichloridium delicatum & Heteroconium simile spp. nov. (China) ... 581

C D

un oP

Fic. 1. Globoramichloridium delicatum (holotype, HJAUP M0217).

A-C. Conidiophores, conidiogenous cells, and conidia; D. Conidiophore; E. Conidia.

smooth-walled hyphae. CONIDIOPHORES macronematous, mononematous,

unbranched, erect, straight or flexuous, cylindrical, 4-9-septate, smooth,

brown to dark brown, paler towards the apex, <310 um long, 6-8 um wide.

CONIDIOGENOUS CELLS polyblastic, integrated, terminal, sympodial extended,

with crowded, prominent, thickened, slightly melanized scars. CONIDIA

solitary, acropleurogenous, subglobose to narrow subnapiform, unicellular,

yellowish green, verrucose or smooth, dry, 11-14 x 9.5-13 um, truncate at the

base, 1-2 um wide, with a conspicuous 1 tm long hilum.

ComMEnts -—The monotypic genus Globoramichloridium was erected based

on molecular phylogenetic analysis, with Chloridium indicum Subram.

(Subramaniam 1955) [= Globoramichloridium indicum (Subram.) Y. Marin

& Crous] as its type (Marin-Felix & al. 2019). Globoramichloridium is

582 ... Qiu & al.

characterized by broadly ellipsoidal to globose, (0—)1-septate, smooth-walled or

coarsely verrucose conidia seceding schizolytically from polyblastic, integrated,

terminal, sympodial, geniculate or nodose conidiogenous cells with prominent,

thickened and darkened scars (Marin-Felix & al. 2019). These morphological

characteristics differ from the similar genus Ramichloridium Stahel ex de Hoog

(de Hoog & Hermanides-Nijhof 1977), which has conspicuous conidiogenous

loci with crowded or scattered, unpigmented to faintly pigmented, or slightly

prominent denticles, thin scars, and clavate or oblong to ellipsoid, or obovate

to obconical, aseptate, smooth-walled to finely verrucose conidia (Arzanlou &

al. 2007, Seifert & al. 2011).

Globoramichloridium was previously known only from the type species,

G. indicum [= Chloridium indicum; = Veronaea indica (Subram.) M.B. Ellis;

= Ramichloridium indicum (Subram.) de Hoog], which has been reported

from Australia, Cuba, Hong Kong, Malaysia, and Brazil (Castafteda-Ruiz 1986,

GBIF 2020). Globoramichloridium delicatum, the second species of the genus,

differs from G. indicum, which has smaller [(5-)7—8(-10) x (4-)6-6.5(-9) um],

broadly ellipsoidal to globose, (0—)1-septate, subhyaline to pale brown conidia

(Arzanlou & al. 2007).

Heteroconium simile L. Qiu, Jian Ma, R.E Castafieda &

X.G. Zhang, sp. nov. Fic. 2

MB 836852

Differs from Heteroconium ponapense and H. indicum by its shorter or smaller,

0-1-septate, dimorphic conidia that are obviously constricted at the septum.

Type: China, Jiangxi Province: Jinggangshan Mountain, Shuikou scenic spot,

26°33'03”N_ 114°07'55”E, on dead branches of an unidentified broadleaf tree,

6 Nov. 2014, J. Ma (holotype, HJAUP M0218).

EtrymMo oey: Latin, simile, referring to its similarity to Heteroconium ponapense and

H. indicum.

Co.Lontgs on the natural substratum effuse, dark brown. Mycelium partly

superficial, partly immersed, composed of branched, septate, pale brown,

smooth hyphae. CONIDIOPHORES macronematous, mononematous, simple

or branched, erect, straight or flexuous, cylindrical, smooth, brown to dark

brown at the base, and paler towards the apex, 2-7-septate, 28-72 x 3-5 um.

CONIDIOGENOUS CELLS monoblastic, integrated, terminal, cylindrical,

brown, smooth, 10-14 x 3.5-4.5 um. Conidial secession schizolytic. CONIDIA

acrogenous, blastocatenate, 11-15 x 4-5 um, smooth, 0-1-septate, dimorphic,

i) unicellular, ellipsoidal, fusiform, or ovoid, truncate at both ends or truncate

at the base and rounded at the apex, concolorous, pale brown to dark brown,

Globoramichloridium delicatum & Heteroconium simile spp. nov. (China) ... 583

B C

wiz

winigz

Fic. 2. Heteroconium simile (holotype, HJAUP M0218).

A-C. Conidiophores, conidiogenous cells, and conidia.

ii) bicellular, constricted at the septum, obcalceiform, ellipsoidal to obclavate,

mostly bicolorous, basal cell brown or dark brown basal, apical cell pale brown

or brown.

ComMENTS —Heteroconium was erected by Petrak (1949) with H. citharexyli

Petr. as type species. It is characterized by macronematous, mononematous,

usually unbranched conidiophores, sometimes with a secondary branch

originating after conidial secession or near percurrent regeneration. The

conidiogenous cells are monoblastic, integrated, terminal, determinate or

sometimes percurrent extended and produce acrogenous, blastocatenate,

euseptate conidia that secede schizolytically (Petrak 1949; Ellis 1971; Castaneda

& al. 1999, 2008; Taylor & al. 2001; Ma & al. 2012b).

Twenty-five Heteroconium epithets were listed in Index Fungorum (2020).

However, H. tetracoilum (Corda) M.B. Ellis was transferred to Lylea Morgan-

Jones due to its distoseptate conidia (Ellis 1976, Holubova-Jechova 1978).

Heteroconium solaninum (Sacc. & P. Syd.) M.B. Ellis is not congeneric with

H. citharexyli; it has an obligate association with asterinaceous fungi and

584 ... Qiu & al.

different conidial septation sequence and was designated as the type species of

Pirozynskiella S. Hughes (Ellis 1976, Hughes 2007). Heteroconium chaetospira

(Grove) M.B. Ellis, H. eucalypti Crous & M.J. Wingf., and H. kleinzeense Crous

& Z.A. Pretorius were transferred to other genera (Cladophialophora Borelli;

Thyrinula Petr. & Syd.; Alysidiella Crous) based on molecular phylogenetic

analyses (Ellis 1976; Crous & al. 2006, 2007a,b, 2019; Cheewangkoon & al.

2012). Although H. queenslandicum Matsush. is a currently accepted name, this

species exhibits a similar conidial ontogeny with Parapleurotheciopsis P.M. Kirk

in its holoblastic conidia with an acropetal branched chain through an apical

ramoconidium that secedes schizolytically (Matsushima 1989, MycoBank

2020); these characters were remarked by Castafeda-Ruiz & al. (2008). Thus,

Heteroconium currently contains 19 species, among which 11 species have

been recorded from China: H. annesleae S.C. Ren & X.G. Zhang, H. arundicum

Chowdhry, H. bannaense J.W. Xia & X.G. Zhang, H. decorosum R.F. Castaneda

& al., H. fici L.G. Ma & X.G. Zhang, H. indicum, H. neolitseae S.C. Ren & X.G.

Zhang, H. phellodendri Jian Ma & X.G. Zhang, H. ponapense, H. schimae Y.D.

Zhang & X.G. Zhang, H. tsoongiodendronis L.G. Ma & X.G. Zhang (Zhang & al.

2010; Ma & al. 2012a,b; Ren & al. 2012; Xia & al. 2012; Zhang 2018).

Among the 19 Heteroconium species, only H. ponapense Matsush. and

H. indicum Varghese & V.G. Rao are resemble H. simile in having predominately

1-septate conidia. However, H. ponapense differs by its obclavate to ellipsoid,

brown, longer (12-24 um) conidia (Matsushima 1983), while H. indicum differs

by its larger (23-42.5 x 8.5-10.5 um), fusiform to cylindrical, pale brown to

reddish brown conidia (Varghese & Rao 1980).

Acknowledgments

The authors express gratitude to Dr. Rosa Maria Arias Mota (Instituto Tecnoldgico

Superior de Xalapa, Mexico) and Dr. De-Wei Li (The Connecticut Agricultural

Experiment Station, USA) for serving as pre-submission reviewers and to Dr. Lorelei L.

Norvell for final editorial review and Dr. Shaun Pennycook for nomenclatural review.

We thank to the National Natural Science Foundation of China (Nos. 31970018,

31360011) for supported this project.

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MY COTAXON

July-September 2021— Volume 136, pp. 587-595

https://doi.org/10.5248/136.587

Neosporidesmium himachalense sp. nov. from India and

Neosporidesmina gen. nov. to accommodate N. micheliae

RAJNISH KUMAR VERMA’, I.B. PRASHER’, SUSHMA3,

AjAY KUMAR GAUTAM‘, KUNHIRAMAN C, RAJESHKUMAR),

ASHISH KUMAR®, RAFAEL F. CASTANEDA-RUIZ7"*

' Department of Plant Pathology, Punjab Agricultural University,

Ludhiana, Punjab, 141004, India

*Department of Botany, Mycology and Plant Pathology Laboratory, Panjab University,

Chandigarh, 160014, India

> Department of Biosciences, Chandigarh University Gharuan, Punjab, India

‘School of Agriculture, Abhilashi University, Mandi, Himachal Pradesh, 175028, India

° National Fungal Culture Collection of India (NFCCI),

Biodiversity and Palaeobiology (Fungi) Gr. MACS, Agharkar Research Institute,

G.G. Agarkar Road, Pune, 411 004, Maharashtra, India

° Department of Geography, Panjab University Chandigarh, 160014, India

” Instituto de Investigaciones Fundamentales en Agricultura Tropical Alejandro de Humboldt

(INIFAT), OSDE, Grupo Agricola,

Calle 1 Esq. 2, Santiago de Las Vegas, C. Habana, Cuba, C.P. 17200

*CORRESPONDENCE TO: “vermarajnish1985@gmail.com * rfcastanedaruiz@gmail.com

ABSTRACT—A new species, Neosporidesmium himachalense, collected from a decaying twig

of an unidentified angiosperm in Mandi Himachal Pradesh, India, is described and illustrated.

It is characterized by macronematous, laterally and terminally diverging conidiophores,

terminal, doliiform, monoblastic conidiogenous cells, and obclavate 7—18-distoseptate, pale

brown conidia. A new genus Neosporidesmina is proposed for Neosporidesmium micheliae,

which has euseptate conidia.

KEY worpDs—anamorphic fungi, hyphomycetes, taxonomy

Introduction

Recent works on fungal biodiversity in India have prompted an

increase in the mycobiota of the country with several new species and

588 ... Verma & al.

new records (Adam¢ik & al. 2015; Buyck & al. 2017; Gautam &. al. 2020;

Sushma & al. 2020; Verma & al. 2019, 2020). During continuing surveys of

microfungi associated with plant materials in Himachal Pradesh, India, a

synnematous fungus was collected on an unidentified angiosperm decaying

twig. Its conidiogenesis and conidial morphology placed this fungus in

Neosporidesmium Mercado & J. Mena. Fifteen Neosporidesmium species

have already been reported, from four countries: China (8 spp.), Cuba

(1 sp.), India (4 spp.), and Vietnam (2 spp.). Distribution is depicted in the

map (Fic. 1) constructed using the geographical coordinates mentioned in

the protologues of these taxa. The original Neosporidesmium generic concept

was expanded by Zhang & al. (2011), after the description of N. micheliae,

which is characterized by having euseptate conidia. This species is clearly

separated from the distoseptate Neosporidesmium generic concept, and we

propose a new genus, Neosporidesmina.

20°0'0"N

Es 120°0'0"W 100°O'O"W SO*O'O"W 60°0'O"W 40°00" WW 20h0'0' 71

Chandigarh.

J i bs Nv ion .

somes ep face

N. diaoluoshanense Hainan, China N. sinense

6 | Garo His,

—— as

= ra i | India

4 Location of Species | a, nda

¥ ] N. . : Chandigarh, 13 N. wevishaner

—— International Boundaries N. macrosporum India N. wiqvishanense

N maestrense

1,520

Miles eX

60°0'0"E 70°0'0"E. $0°0'0"E 90°0'0"E 100°0'0"E 110°00"E 120°0'0"E

10°0'0"S_ = :10°0'0"N_-30°0'0"-N ape

0°0'0'

10°0'0"S = 10°O'0"N_30°0'0"N-50°0'0"N

10°0'0"S

Fic. 1. Map showing the world distribution of Neosporidesmium spp.

Neosporidesmium sp. nov. & Neosporidesmina gen. & sp. nov. (India) ... 589

Materials & methods

Decaying culms, leaves, twigs, bark, and dead wood were collected in separate

zip lock plastic bags and brought to the laboratory. The specimens were mounted on

glass slides in either 4% KOH, 0.01% cotton blue or lactophenol (Kirk & al. 2008),

The specimens were studied microscopically under a matrix stereo trinocular

microscope (VL-Z60) and a compound microscope (VRS-2f) for macroscopic

and microscopic characters. All the measurements were taken assisted by Pro

MED software and represented as the extreme values without statistical treatment.

The specimens were deposited in herbarium of Panjab University, Chandigarh,

India (PAN).

Repeated attempts to culture the fungus failed on various media (i.e., 2% agar

broth, cornmeal agar (CMA), potato dextrose agar (PDA), oatmeal agar, malt

extract agar (MEA), and solidified glucose peptone medium comprising: glucose

10 g/l, peptone 2 g/l, KH,PO, 1 g/l, MgSO,.7H,O 0.5 g/l, agar 20 g/l in 1000 ml

H,O with chips of host) with the addition of 50 ug/mL of two antibiotics (penicillin

and streptomycin)—according to Choi & al. (1999) and Kirk & al. (2008).

The distribution map was constructed by using ArcGIS software (Beyer 2004).

Taxonomy

Neosporidesmium himachalense Rajn.K. Verma, Prasher, Sushma,

A.K. Gautam, Rajeshk. & R.F. Castaneda, sp. nov. Fics 2, 3

IF557994

Differs from Neosporidesmium macrosporum by its smaller conidia with fewer

distosepta.

Type: India. Himachal Pradesh: Bilaspur (Jolplakhin), on dead and decaying twigs of

angiosperm plant, 23 Nov 2015, R. K. Verma (Holotype, PAN 32815).

Erymo coey: Latin, himachalense, refers to the Himachal, where the type was collected.

CoLonigs on the natural substrate effuse, hairy, dark brown. Mycelium

superficial, partly immersed in the substratum. CONIDIOMATA synnematal,

erect, cylindrical, scattered, dark brown to black, tapering toward the apex,

up to 2400 um high and 90-150 um wide at the base. CONIDIOPHORES

macronematous, unbranched, septate, smooth, brown to dark brown,

diverging laterally and terminally. CoNIDIOGENOUS CELLS monoblastic,

doliiform, integrated, terminal, smooth, brown to dark brown, having

2-4 enteroblastic extensions (11.5—21 x 6.5—8.7 um). Conidial secession

schizolytic. Conrp1a 72.5-151.5 x 10.5-15.7 um, solitary, acrogenous,

obclavate, 7—18-distoseptate, smooth, pale brown and paler at the apex.

590 ... Verma & al.

Fic. 2. Neosporidesmium himachalense (holotype, PAN 32815): A. Synnemata attached to natural

substratum; B. Synnemata; C. Conidia attached to conidiogenous cells; D, E. Conidia. Scale bars:

A = 100 mm; B = 200 um; C-E = 20 um.

Neosporidesmium sp. nov. & Neosporidesmina gen. & sp. nov. (India) ... 591

Fic. 3. Neosporidesmium himachalense (holotype, PAN 32815): Developing conidia and

conidiogenous cells. Scale bars: A, B = 20 um; C, D = 10 um.

592. ... Verma & al.

Neosporidesmina R.F. Castafieda, Rajn.K. Verma, Prasher, Sushma,

A.K. Gautam & Rajeshk., gen. nov.

IF 557995

Differs from Sporidesmina by its invariably euseptate conidia.

TYPE SPECIES: Neosporidesmium micheliae Y.D. Zhang & X.G. Zhang [=Neosporidesmina

micheliae (Y.D. Zhang & X.G. Zhang) R.F. Castaneda & al.]

Erymo .oey: Latin, Neo-, meaning, new + Latin, sporidesmina, referring to the genus

Sporidesmina.

Cotonigs on the natural substrate effuse, hairy, dark brown. Mycelium

superficial, partly immersed in the substratum. CONIDIOMATA synnematal,

scattered, erect, cylindrical, dark brown to black. Mycelium superficial and

immersed. CONIDIOPHORES macronematous, unbranched, diverging laterally

and terminally, septate, smooth, brown to dark brown. CONIDIOGENOUS CELLS

monoblastic, cuneiform, cylindrical or subulate, integrated, determinate,

terminal, smooth, brown to dark brown. Conidial secession schizolytic.

ConipiA solitary, acrogenous, obclavate or cylindrical, euseptate, smooth,

brown.

Neosporidesmina micheliae (Y.D. Zhang & X.G. Zhang) R.F. Castafieda,

Rajn.K. Verma, Prasher, Sushma, A.K. Gautam & Rajeshk., comb. nov.

IF 557996

= Neosporidesmium micheliae Y.D. Zhang & X.G. Zhang, Sydowia 63(1): 128 (2011).

Discussion

Neosporidesmium was erected by Mercado Sierra & Mena Portales (1988)

for a single species, N. maestrense. The genus is characterized by effuse,

dark brown to black, hairy colonies, unbranched conidiophores aggregating

into erect, cylindrical, dark brown to black synnemata and integrated,

terminal, monoblastic, lageniform or doliiform, determinate or percurrent

extending conidiogenous cells. The conidia are solitary, obclavate or

cylindrical, distoseptate, pale brown to brown, smooth. Subsequently,

14 other species were described: N. antidesmatis, N. appendiculatum,

N. diaoluoshanense, N. garoense, N. khasianum, N. macrosporum, N. malloti,

N. micheliae, N. microsporum, N. sinense, N. subramanianii, N. vietnamense,

N. wuyishanense, and N. xanthophylli (Wu & Zhuang 2005; Ma & al. 2011;

Zhang & al. 2011; Melnik & Braun 2013; Prasher & Verma 2015a,b; Li & al.

2015, 2017; Mel'nik & al. 2016, Pratibha & al. 2018; Index Fungorum 2020).

The criteria used for species delimitation are based primarily on

conidiomata size, the enteroblastic extensions of conidiogenous cells, and

Neosporidesmium sp. nov. & Neosporidesmina gen. & sp. nov. (India) ... 593

conidial morphology (Mercado Sierra & Mena Portales 1988, Wu & Zhuang

2005). Additionally, Prasher & Verma (2015b) described N. appendiculatum

Prasher & Rajn.K. Verma as having conidial appendages. Neosporidesmium

macrosporum Prasher & Rajn.K. Verma is comparable with N. himachalense

but differs by its <1100 um tall conidiomata and larger (108-250 x

12—16.5 um) 9-20-distoseptate conidia. Neosporidesmium himachalense

also differs from other species in size of synnemata and number of conidial

distosepta and sizes (TABLE 1).

Neosporidesmina is close to Sporidesmina Subram. & Bhat, typified by

S. malabarica, which is characterized by erect, carbonaceous, compact,

unbranched or mostly branched, divergent, synnematal conidiomata

formed by indistinguishable, unbranched, tiny packed, septate, pale to dark

TABLE 1. Morphological comparison of accepted Neosporidesmium species.

SPECIES

N. antidesmatis

N. appendiculatum

N. diaoluoshanense

N. garoense

N. himachalense

N. khasianum

N. macrosporum

N. maestrense

N. malloti

N. microsporum

N. sinense

N. subramanianii

N. vietnamense

N. wuyishanense

N. xanthophylli

SYNNEMATA

[um]

950 x

30-50

<4700

long

605 x

40-60

700-1620 x

50-65

2400 x

90-150

855-2750 x

50-60

1100 x

80-140

2500 x

60-250

380 x

20-30

1200 x

20-40

1000 x

80-110

600-1200 x

45-80

500-1430

long

<1290 x

120

1000 x

40-55

CONIDIAL

SEPTA

11-15

CONIDIAL SIZE

[um]

48-75 x

7.5-9

82-148 x

17-19

105-135 x

10.5-13.5

75-100 x

8-11

72-151 x

10.5-15.7

103-135 x

10-18

108-250 x

12-16.5

55-100 x

15-19

22.5-33.5 x

7-9

45-67 x

6-7

120-150 x

12-15

80-120 x

10-14

75-110 x

11-13.5

30-60 x

5-9

33.5-51.5 x

12-14

REFERENCE

Ma &al. 2011

Prasher & Verma 2015b

Li & al. 2015

Pratibha & al. 2018

This paper

Pratibha & al. 2018

Prasher & Verma 2015a

Mercado Sierra &

Mena Portales 1988

Ma & al. 2011

Wu & Zhuang 2005

MePnik & al. 2016

MePnik & Braun 2013

Li & al. 2017

Ma & al. 2011

594 ... Verma & al.

brown conidiophores. The conidiogenous cells are monoblastic, cylindrical

integrated, terminal, and producing solitary acrogenous elongated-fusiform

brown conidia, that are 8-12-euseptate from the base toward near the end,

but abruptly attenuate, sub-rostrate or subulate, hyaline, 1-2-distoseptate

just below the apex (Subramanian & Bhat 1989). The two types of septa

separate Sporidesmina from Neosporidesmina, which has only euseptate

conidia.

Acknowledgments

We are indebted to Dr. De-Wei Li (The Connecticut Agricultural Experiment

Station, USA) and Dr. Rosa Maria Arias Mota (Instituto Tecnoldgico Superior de

Xalapa, Mexico) for serving as pre-submission reviewers and to Dr. Lorelei L. Norvell

for final editorial review and Dr. Shaun Pennycook for nomenclatural review. The

IBP is thankful to the Ministry of Environment and Forests, Government of India,

for financial assistance (vide letter 14/26/2008—ERS/RE dt. 06.07.2010), and to UGC

(SAP, DRSIII). The authors (RKV, IBP and Sushma) are thankful to Chairperson

Department of Botany Panjab University for providing infrastructural and laboratory

facilities. RFCR is grateful to Cuban Ministry of Agriculture for facilities.

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MY COTAXON

July-September 2021— Volume 136, pp. 597-605

https://doi.org/10.5248/136.597

Cladorrhinum yunnanense sp. nov. from China

YUE PAN™, JI-WEN XIA* 4, SHENG-TING HUANG?,

CHUN-YUAN ZHU?, XIU-GUO ZHANG!, ZHUANG LI™?

' Shandong Provincial Key Laboratory for Biology of Vegetable Diseases and Insect Pests,

College of Plant Protection, Shandong Agricultural University,

Taian, Shandong 271018, China

? College of Life Sciences, Yangtze University,

Jingzhou, Hubei 434023, China

* College of Life Sciences, Shandong Agricultural University,

Taian, Shandong, 271018, China

“CORRESPONDENCE TO: “xiajiwen1@126.com, *liz552@126.com

ABSTRACT—A new asexual fungus, Cladorrhinum yunnanense, isolated from unidentified

fallen leaves of a broadleaf tree in Yunnan, China, is described by morphological characters

and phylogenetic analyses of DNA sequence data from three loci (ITS, LSU and TUB).

Key worps—Lasiosphaeriaceae, mitosporic fungi, Sordariales, taxonomy

Introduction

Cladorrhinum Sacc. & Marchal was established in 1885 with

C. foecundissimum Sacc. & Marchal as the type species (Marchal 1885).

The genus is characterized by fast-growing colonies with pale to dark

greenish, greyish, or brownish pigmentation, and conidiophores typically

comprising intercalary phialides with lateral openings (pleurophialides),

usually with a flaring collarette. The conidia are one-celled, hyaline or

subhyaline, smooth-walled, commonly globose to dacryoid, and arranged

in slimy masses. Bahupaathra Subram. & Lodha is synonymised with the

earlier described Cladorrhinum (Mouchacca & Gams 1993).

* These authors contributed equally to this work.

598 ... Pan, Xia & al.

Mouchacca & Gams (1993), who revised Cladorrhinum, accepted five

species: C. foecundissimum, C. brunnescens W. Gams, C. bulbillosum W.

Gams & Mouch., C. phialophoroides Mouch. & W. Gams, and C. samala

(Subram. & Lodha) W. Gams & Mouch. Subsequently added to the genus

were C. flexuosum Madrid & al., C. microsclerotigenum Madrid & al.,

C. australe Gasoni, C. globisporum Z.F. Zhang & L. Cai, C. hyalocarpum

(Arx) X. Wei Wang & Houbraken, and C. intermedium (Stchigel & Guarro)

X. Wei Wang & Houbraken (Madrid & al. 2011, Carmaran & al. 2015,

Zhang & al. 2017, Wang & al. 2019). Wang & al. (2019) recently transferred

C. bulbillosum to Podospora Ces. and C. phialophoroides to Triangularia

Boedijn.

Fungal diversity in China is high but still mostly unexplored. Several

new species from Yunnan Province have been previously described (Xia &

al. 2012, 2014, 2016; Ma & al. 2014 a,b,c; Zhang & al. 2016; Guo & al. 2019;

Zheng & al. 2019, 2020). During our ongoing survey of anamorphic fungi

associated with plant leaves in tropical and subtropical forests of southern

China, we collected a new species representing Cladorrhinum.

Materials & methods

Isolates and Morphological analysis

Diseased leaves were collected from an unidentified plant in Xishuangbanna,

Yunnan Province, China. Tissue pieces (5 x 5 mm) were taken from the margin

of leaf lesions and surface-sterilized by immersing in 75% ethanol solution for

1 min, 5% sodium hypochlorite solution for 30 s, and then rinsing three times in

sterile distilled water for 1 min. The pieces were dried with sterilized paper towels

and then placed on potato dextrose agar (PDA) (Cai & al. 2009). All the PDA

plates were incubated at room temperature 25 °C for 2-4 d, and from which the

peripheries of the colonies were picked out and inoculated onto new PDA plates.

Colonies were photographed after 7 d and 15 d with a Powershot G7X mark II

digital camera. Micromorphological characters were observed using an Olympus

SZX10 stereomicroscope and Olympus BX53 microscope, both fitted with Olympus

DP80 high definition colour digital cameras. All fungal strains were stored in 10%

sterilized glycerin at 4 °C for further studies. The specimens are deposited in the

Herbarium of Plant Pathology, Shandong Agricultural University, Taian, China

(HSAUP). Ex-type cultures are deposited in the Shandong Agricultural University

Culture Collection, Taian, China (SAUCC).

DNA extraction, PCR amplification and sequencing

Genomic DNA was extracted from colonies grown on PDA, using the CTAB

method (Doyle & Doyle 1990). The primer pair ITS1/ITS4 (White & al. 1990) was

Cladorrhinum yunnanense sp. nov. (China) ... 599

TABLE 1. Cladorrhinum and related strains included in the phylogenetic analyses.

SPECIES NAME

Cercophora ambigua

Cercophora appalachianensis

Cercophora areolata

Cercophora caudata

Cercophora grandiuscula

Cercophora samala

Cercophora sulphurella

Cladorrhinum australe

Cladorrhinum brunnescens

Cladorrhinum bulbillosum

Cladorrhinum flexuosum

Cladorrhinum foecundissimum

Cladorrhinum globisporum

Cladorrhinum hyalocarpum

Cladorrhinum intermedium

Cladorrhinum microsclerotigenum

Cladorrhinum phialophoroides

Cladorrhinum samala

Cladorrhinum yunnanense

Phialophora brevicollaris

Podospora cochleariformis

Podospora cupiformis

Podospora curvicolla

Podospora decipiens

Podospora didyma

Podospora fibrinocaudata

Podospora fimiseda

Podospora intestinacea

CULTURE ACC. NO.

CBS 215.60

HKUCC3711

UAMH7495

CBS 606.72

CBS 1200137

CBS 109.937

SMH2531

INTA-AR 108

INTA-AR 110

CBS 643.757

INTA-AR 102

CBS 304.907

FMR 10415

BCCM 6980

CBS 180.667

CGMCC3.179217

CGMCC3.17922

CBS 322.707

CBS 102198

CBS 433.967

CBS 100257

CBS 290.757

CBS 301.907

CBS 302.90

INTA-AR 168

SAUCC 0525 7

SAUCC 0791

CBS 126.747

CBS 249.71

CBS 246.717

IFO 8548

CBS 258.69

CBS 232.78

CBS 315.917

CBS 990.96

CBS 113106

GENBANK ACCESSION NO.

ITS

AY999137

AF177155

AY587911

AY999135

GQ922544

AY999134

AY587913

KT321073

KT321074

NR137152

KT321068

NRO77199

NR154757

KT321080

MH858767

KU746680

KU746679

MK926857

MK926858

MK926859

MK926860

NR154758

NRO77198

KT321079

KT321078

MN956794

MN956795

NR121471

MH860099

MH860098

AY999122

KX171946

AY999127

MH862255

KX348039

AY999121

LSU

AY999114

AY587936

MH872289

KF557693

AY999111

AY587938

KT312986

KT312987

FR692346

KT312982

FR692339

NG058773

KT312993

FR692343

KU746726

KU746725

MK926857

MK926858

MK926859

MK926860

FR692342

FR692344

KT312992

KT312991

MN956785

MN956786

AY999098

AY999102

AY999099

AY780073

AY999100

MK926844

AY346296

AY999104

TUB

AY999147

AY600252

AY999151

AY999140

AY600254

KT291711

KT291712

KT291706

KT291720

KT291721

KT291717

KU746771

KU746772

MK926957

MK926958

MK926959

MK926960

KT291718

KT291719

KT291716

MT376683

GU727570

AY999145

AY999149

AY999148

AY780130

AY999142

MK926944

AY780133

AY999152

600 ... Pan, Xia & al.

used to amplify the rDNA internal transcribed spacer (ITS) region following White

& al. (1990). The primer pair LROR/LRS (Vilgalys & Hester 1990, Rehner & Samuels

1994) was used to amplify a partial fragment of the large subunit (LSU) gene. The

primer pair Bt2a/Bt2b (Glass & Donaldson 1995) was used to amplify the beta-

tubulin (TUB) gene.

The DNA was amplified using an Eppendorf Master Thermocycler in 25 uL

reaction volumes containing 12.5 uL Vazyme Green Taq Mix, 1 uL of each Biosune

forward and reverse primer (10 uM), and 1 uL template genomic DNA adjusted with

distilled deionized water to a total 25 uL volume. PCR parameters were 95 °C for 5

min, followed by 35 cycles of denaturation at 95 °C for 30 s, annealing at 55 °C (ITS,

LSU) or 60 °C (TUB) for 30 s, extension at 72 °C for 1 min, and a final elongation

step at 72 °C for 10 min. PCR products were estimated visually by staining with

GelRed after 1% agarose gel electrophoresis. Sequencing was done bi-directionally,

conducted by the Biosune Company Limited. Consensus sequences were obtained

using MEGA v. 7 (Kumar & al. 2016). All sequences generated in this study were

deposited in GenBank (TABLE 1).

Sequence alignment and phylogenetic analysis

The quality of our amplified nucleotide sequences was checked and combined

by MEGA v. 7 (Kumar & al. 2016), and reference sequences were retrieved from

the National Center for Biotechnology Information (NCBI), based on recent

publications on Cladorrhinum (Madrid & al. 2011, Carmaran & al. 2015, Zhang

& al. 2017). Sequences were aligned using MAFFT v. 7.310 (Katoh & al. 2019;

http://mafft.cbrc.jp/alignment/server/index.html) and manually corrected using

MEGA v. 7.

The combined gene regions were phylogenetically analysed using Maximum-

Likelihood (ML) and Bayesian Inference (BI) methods. RaxML (v. 8.2.9) and

Bayesian analyses (MrBayes v. 3.2.6) were run on the CIPRES Science Gateway

portal (Miller & al. 2012). Evolutionary models were calculated using MrModelTest

v. 2.3 (Nylander 2004) selecting the best-fit model for each data partition according

to the Akaike criterion. For ML analyses default parameters were used and bootstrap

support (BS) used the rapid bootstrapping algorithm with the automatic halt option.

Bayesian analyses included two parallel runs of 5,000,000 generations, with the

stop rule option and a sampling frequency set to each 1000 generations. The 50%

majority rule consensus trees and posterior probability (PP) values were calculated

after discarding the first 25% of the samples as burn-in. The resulting trees were

plotted using FigTree v. 1.4.2 (http://tree.bio.ed.ac.uk/software/figtree) and edited

with Adobe Illustrator CS v. 5. The individual gene datasets were assessed for

incongruence before being concatenated by checking their individual phylogenies

for conflicts between clades with significant ML and BI support (Mason-Gamer &

Kellogg 1996, Wiens 1998).

Cladorrhinum yunnanense sp. nov. (China) ... 601

- Cladorrhinum yu

—L Cladorrhinum yun sé

Cladorrhinum samala CBS 302.90

Cladorrhinum samala INTA-AR 168

Cladorrhinum flexuosum FMR 10415

4100/1 Cladorrhinum australe INTA-AR 110

Cladorrhinum australe INTA-AR 108

4100/1 Cladorrhinum bulbillosum \NTA-AR 102

Cladorrhinum bulbillosum CBS 304.90

99/1 Cercophora samala CBS 109.93

97/1

Cladorrhinum microsclerotigenum CBS 290.75

Triangularia phialophoroides CBS 301.90

Cercophora grandiuscula CBS 120013

Cladorrhinum foecundissimum BCCM 6980

Cladorrhinum foecundissimum CBS 180.66

"00/1 Cladorrhinum hyalocarpum CBS 102198

Cladorrhinum hyalocarpum CBS 322.70

Cladorrhinum globisporum LC5370

Cladorrhinum globisporum LC5415

100/1 | Cladorrhinum intermedium CBS 100257

Cladorrhinum intermedium CBS 433.96

Cladorrhinum brunnescens CBS 643.75

Podospora decipiens CBS 258.69

Podospora fimiseda CBS 990.96

baal Cercophora areolata UAMH7495

Cercophora sulphurella SMH2531

Podospora cupiformis CBS 246.71

Podospora curvicolla IFO 8548

100/1

96/0.99 Cercophora caudata CBS 606.72

Podospora intestinacea CBS 113106

Cercophora appalachianensis HKUCC3711

Podospora cochleariformis CBS 249.71

Cercophora ambigua CBS 215.60

Podospora didyma CBS 232.78

Podospora fibrinocaudata CBS 315.91

Hyphodiscus brevicollaris CBS 126.74

Fic. 1. The ML consensus tree inferred from the combined ITS, LSU and TUB sequence

alignment. Branches are labeled with Maximum-Likelihood bootstrap support 275%

and Bayesian posterior probabilities 20.95. The tree is rooted to Hyphodiscus brevicollaris

(CBS 126.74). The scale bar indicates 0.1 expected changes per site.

Phylogenetic results

The dataset comprised 36 taxa representing 28 species including

Hyphodiscus brevicollaris (CBS 126.74) as the outgroup. The final alignment

comprised a total of 2101 characters of the combined ITS, LSU, and TUB

including gaps: ITS 1-545; LSU, 546-1357; and TUB, 1358-2101. Of these

characters, 1315 were constant, 221 parsimony-uninformative, and 565

parsimony-informative. For ML and BI analyses, GTR+I+G for ITS and LSU,

and GTR+G for TUB were selected and incorporated into the analyses. As

the topology of ML tree confirmed the tree topologies obtained from the

BI analyses, only the ML tree is presented (Fic. 1). In this tree, our strains

formed a distinct clade and grouped together with Cladorrhinum samala and

C. flexuosum with good support (BS = 95%, PP = 1). Therefore, we determined

that our strains belonged to a novel species of Cladorrhinum.

602 ... Pan, Xia & al.

Taxonomy

Cladorrhinum yunnanense Y. Pan, J.W. Xia, X.G. Zhang & Z. Li, sp. nov. FIG. 2

MB 836106

Differs from all other Cladorrhinum spp. by its blastic conidiogenous cell.

Type: China, Yunnan Province, Xishuangbanna, on unidentified fallen leaf of a

broadleaf tree, 16 Apr. 2019, Y. Pan (Holotype, HSAUP 0525; ex-type living culture

SAUCC 0525).

EryMo_oey: in reference to the province containing the type locality.

CoLonigs on PDA pale grey to dark grey, cottony to fluffy, flat to slightly

plicate, margin entire. Reverse pale brown to brown. Culture on OA white

to grayish, margin entire, aerial mycelia sparse. Reverse white. Culture on

MEA white to yellowish-brown, cottony to fluffy, margin filiform. Reverse

white to yellowish. VEGETATIVE HYPHAE hyaline, branched, septate, thin-

walled, slightly rough, 1.5-4 um diam. MIcROSCLEROTIA not observed.

SPORODOCHIUM forming on PDA within 30 d or longer, margin round, black,

scattered over entire colony. CONIDIOPHORES single, unbranched, erect,

straight or slightly flexuous, cylindrical, smooth, thick-walled, hyaline at the

base, pale brown to brown in the upper part, 2—4-septate, 40-60 x 2-4 um.

CONIDIOGENOUS CELL polyblastic, integrated, terminal, cylindrical, pale

brown to brown, swollen at the apex, 15-25 x 2-4 um. Conip1A pale brown,

smooth, thick-walled, oval to ellipsoid, aseptate, 6-10 x 2-4 um, guttulate,

aggregated in slimy head.

ADDITIONAL SPECIMEN EXAMINED: CHINA, YUNNAN PROVINCE, Xishuangbanna,

on unidentified fallen leaf of a broadleaf tree, 16 Apr. 2019, Y. Pan (HSAUP 0791;

living culture SAUCC 0791).

CoMMENTS—According to Madrid & al. (2011) regarding convergent

morphology in Cladorrhinum, characterization of any potentially new

species within this genus should be included in the phylogenetic analyses.

Based on a megablast search of GenBank nucleotide sequence database,

the closest hit using the ITS sequence was Cladorrhinum sp. (strain RS_94,

GenBank MK379585.1; Identities = 481/489 (98%), 3 gaps (0%)); the closest

hit using the LSU sequence was Cercophora costaricensis (strain SMH4021,

GenBank AY780059.1; Identities = 827/833 (99%), 1 gap (0%)), and the

closest hit using the TUB sequence was Cladorrhinum foecundissimum

(strain CBS 391.42, GenBank HQ877462.1; Identities = 282/300 (94%),

3 gaps (1%)).

Cladorrhinum yunnanense formed a well-supported sister clade

(BS = 99%, PP = 1.0) to C. flexuosum and C. samala. Based on conidial

Cladorrhinum yunnanense sp. nov. (China) ... 603

Fic. 2. Cladorrhinum yunnanense (holotype, HSAUP 0525). a. Diseased leaf; b,c. Colony on PDA;

d,e. Colony on OA; f,g. Colony on MEA; h. Conidiomata; i-k. Conidiophores and conidia; l-m.

Conidia. Scale bars: i-m = 10 um

morphology, C. yunnanense (oval to ellipsoid, 6-10 x 2-4 um) can be

distinguished from C. flexuosum (mostly globose to dacryoid, 2-4 x 2-3 um)

and C. samala (dacryoid, 2-4 x 1.5-3 um). Cladorrhinum yunnanense also

differs from the previously described Cladorrhinum species by its blastic

conidiogenous cell.

604 ... Pan, Xia & al.

Acknowledgments

The authors express gratitude to Dr. Jian Ma (College of Agronomy, Jiangxi

Agricultural University, Nanchang, China) and Dr. Zefen Yu (School of Life Science,

Yunnan University, Kunming, China) for serving as pre-submission reviewers and

to Dr. Shaun Pennycook for nomenclatural review and Dr. Lorelei L. Norvell for

editorial review. This work was jointly supported by the National Natural Science

Foundation of China (Nos. 31900014, 31770016, 31750001) and the China

Postdoctoral Science Foundation (No. 2018M632699).

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MY COTAXON

July-September 2021— Volume 136, pp. 607-613

https://doi.org/10.5248/136.607

Mazosia hainanensis sp. nov. from tropical China

ZONG-TING YAO”?, ZE-FENG JIA’ “, SHU-HUA JIANG? ®

' College of Life Sciences, Liaocheng University, Liaocheng, P. R. China

’ State Key Laboratory of Mycology, Institute of Microbiology, Chinese Academy of Sciences,

Beijing 100101, PB. R. China

" CORRESPONDENCE TO: “zfjia2008@163.com "jiangsh@im.ac.cn

ABSTRACT—A new species, Mazosia hainanensis, is described from Hainan, China. It is

most similar to M. pseudobambusae but differs by its yellowish green thallus and smaller

ascospores (14-18.5 x 3-4 um). An analysis of its relationships based on molecular

phylogeny is given. A key to the Mazosia species recorded in China is also presented.

Key worps—Arthoniales, lichenized fungi, Roccellaceae, taxonomy

Introduction

Mazosia A. Massal. is a genus of lichenized fungi typified by M.

rotula (Mont.) A. Massal. (Massalongo 1854), belonging to Roccellaceae,

Arthoniales, Arthoniomycetes (Licking 2008, Aptroot & al. 2014, Liicking &

al. 2016). The status of this genus was in disarray until 1996, when Licking

& Matzer determined its position in Roccellaceae (Licking & Matzer

1996). Mazosia is characterized by the rounded ascomata with three layers:

excipulum layer with dark brown hyphae, a layer of crystals, and a thin

thallus layer dominated by the photobiont. There are 8 ascospores per ascus

that are hyaline, fusiform, and transversely septate. Conidia of two types

are sometimes present (Santesson 1952, Licking 2008, Liicking & al. 2016).

According to Index Fungorum, Mazosia includes c. 34 species, most of which

occur on leaves, except for four found on bark (Sparrius 2004, Aptroot & al.

2014).

608 ... Yao, Jia, Jiang

Only six Mazosia species have been found in China, distributed in

Guangdong, Hongkong, Sichuan, Taiwan, and Yunnan (Zahlbriickner 1930,

Santesson 1952, Thrower 1988, Wei & Jiang 1991, Aptroot & Seaward 1999,

Aptroot & Sipman 2001, Aptroot & al. 2003, Aptroot & Sparrius 2003).

TABLE 1. Specimens of Mazosia and related species and their LSU sequences

used in the phylogenetic analysis.

SPECIES VOUCHER SPECIMEN GENBANK No.

Dendrographa alectoroides Ertz 12418 (BR) HQ454527

Robertson 8970 (S-L63276) EF081384

Dendrographa decolorans DUKE:0047570 NG027622

UPS: Frisch 11/Se28 KJ851054

Mazosia bambusae UPS: Joensson Guyana 3c KJ851057

Mazosia carnea Ertz 15686 (BR) KJ524309

Ertz 15684 (BR) KJ524308

Mazosia melanophthalma UPS: Joensson Guyana 3b2 KJ851063

Mazosia paupercula Ertz 9264 (BR) KJ524310

Roccella fuciformis S-F206191 KF036035

Tehler 8171 (L12540) FJ638979

Tylophoron hibernicum Diederich 16335 JF295084

Ertz 11546 (BR) JF295083

Mazosia hainanensis HMAS-L 0145281 [T] MT683119

HMAS-L 0145282 MT683120

Note: Newly generated sequences are set in bold font.

Materials & methods

SPECIMENS. All materials were collected in Hainan Province, China, and are deposited

in the Fungarium-Lichenes of the Institute of Microbiology, Chinese Academy of

Sciences (HMAS-L). Morphological characters were examined and photographed

under a LEICA M125 dissecting microscope. The anatomical characters were

examined and photographed under Zeiss Axioscope2 compound microscope with a

Zeiss Axio Imager A2 and an AxioCam MRc5 camera. Ascospores were tested using

I (1% iodine solution). The lichen substances were detected using standardized thin

layer chromatography (TLC) using solvent systems C and A (Orange & al. 2010).

DNA EXTRACTION, AMPLIFICATION, & SEQUENCING. Genomic DNA was extracted

from ascomata of the specimens followed a modified CTAB method (Rogers &

Bendich 1988). The nuclear ribosomal RNA gene region, large subunit rDNA (LSU)

was amplified using the LIC24R with LR7 primers (Frisch & al. 2014, Ertz & al.

2009). Reactions were carried out in 25 uL reaction system containing 1 uL each

primer solution (10 uM), 2 uL genomic DNA, 8 uL ddH,O, and 13 uwL 2xTaq PCR

Mazosia hainanensis sp. nov. (China) ... 609

MasterMix®. Thermocycling conditions comprised: initial denaturation for 15 min

at 95 °C; followed by 45 cycles of 45 s at 95 °C, 45 s at 53 °C, 1 min at 72 °C, and

a final extension for 7 min at 72 °C. The target product of PCR was afhrmed by

electrophoresis on 1% agarose gels and sequenced by Majorbio (Shanghai).

PHYLOGENETIC ANALYSIS. Due to lack of Mazosia ITS sequences, we chose LSU

for sequence analysis. Our two newly generated LSU sequences were submitted

to GenBank and aligned using MEGA (Kumar & al. 2016) with seventeen other

sequences representing similar taxa in Arthoniales (TABLE 1), selected based on

the results of LSU sequence Blast searches. Tylophoron hibernicum (D. Hawksw. &

al.) Ertz & al., was chosen as outgroup (Urbanavichus & Urbanavichene 2017). The

ML tree involving 1000 pseudoreplicates was generated by MEGA (Kumar & al.

2016), and the evolutionary history was inferred by using the Maximum Likelihood

method based on the Tamura-Nei model (Tamura & Nei 1993). The alignment

was also subjected to a Neighbor-Joining analysis involving 1000 replicates with

MEGA7 (Kumar & al. 2016), and the evolutionary distances were computed using

the Maximum Composite Likelihood method (Tamura & al. 2004).

0 ).0 I

100/100 r Mazosia hainanensis MT683119 [T]

Mazosia hainanensis MT683120

Mazosia bambusae KJ851057

Mazosia paupercula KJ524310

Mazosia carnea KJ524308

99/100

Mazosia carnea KJ524309

Mazosia melanophthalma KJ851063

99/100 Dendrographa alectoroides EF081384

Dendrographa decolorans KJ851054

100/100

Dendrographa decolorans NGO27622

Roccella fuciformis FJ638979

100/100! Roccella fuciformis KF036035

100/100 Lyephoron hibernicum JF295083

Tylophoron hibernicum JF295084

Fic. 1. Phylogenetic tree constructed from ML based on nrLSU sequences of Mazosia and

related species. Maximum likelihood and Neighbor-Joining bootstrap support >50% are shown

at nodes (ML/NJ). New sequences are set in bold font; other sequences were downloaded from

GenBank. Genetic distance scale = 0.01 changes per site.

610 ... Yao, Jia, Jiang

Phylogeny

The phylogenetic tree constructed from ML based on the LSU sequences

(1200 bp) is shown in Fic. 1. Other genera in Roccellaceae—Dendrographa

Darb. and Roccella DC.—were included in the phylogenetic analysis. Within

the well-supported (93/95) monophyletic five-species clade of Mazosia,

the new species M. hainanensis formed an independent clade. Mazosia

bambusae (Vain.) R. Sant. is placed as sister to M. paupercula (Mill. Arg.)

R. Sant. with M. carnea (Eckfeldt) Aptroot & M. Caceres shown as sister to

M. melanophthalma (Mill. Arg.) R. Sant.

Taxonomy

Mazosia hainanensis Z.T. Yao, S.H. Jiang & Z.F Jia, sp. nov. Fic. 2

MB 836195

Differs from Mazosia pseudobambusae by its yellowish green thallus and smaller

ascospores.

Type: China, Hainan, Baoting County, Mount Qixian, on unidentified angiosperm

leaf, 7 Sept. 2017, S.H. Jiang HN20171277 (Holotype, HMAS-L 0145281; GenBank

MT683119).

Erymo.oecy: The epithet refers to the type locality province.

THALLUS yellowish green, continuous, smooth, slightly shiny, often irregular

in outline, finely verrucose, verrucae 20-30 um tall. Photobiont cells

rectangular, 6-15 x 2-4 um, in radiate plates or slightly irregular. Ascomata

rounded, 50-170 um wide and 59-76 um tall; disc black, translucent

when moistened; margin slightly higher than disc and the disc is slightly

lower than or as high as the thallus. Excipulum brown, 8-16 um thick.

Hypothecium 5-15 um high, pale brown. Hymenium 42-50 um high,

colorless. Asci clavate, 34-45 x 10-12.5 um. Ascospores fusiform, 3-septate,

14-18.5 x 3-4 um, the second cell is slightly enlarged. Pycnidia low conical,

10-50 um, Microconidia fusiform-ellipsoid, non-septate, 2-3.5 x 1-1.5 um.

Macroconidia not seen.

CHEMISTRY: unidentified red fluorescent substance in the fourth and sixth

zones in solvent system C and in the fourth and seventh zones in solvent

system A.

ECOLOGY & DISTRIBUTION: On unidentified angiosperm leaves in tropics;

known only from the type locality, Hainan.

ADDITIONAL SPECIMEN EXAMINED: CHINA, HAINAN, Baoting County, Mount

Qixian, on unidentified angiosperm leaf, 7 Sept. 2017, S.H. Jiang HN20171184

(HMAS-L 0145282; GenBank MT683120).

Mazosia hainanensis sp. nov. (China) ... 611

Fic. 2. Mazosia hainanensis (holotype, HMAS-L 0145281). A. Thallus with ascomata;

B. Apothecium section; C. Photobiont; D. Ascus with ascospores; E. Ascospores; F. Microconidia.

Scale bars: A = 0.2 mm; B = 50 um; C-F = 5 um.

REMARKS: This species is characterized by its yellowish green, finely verrucose

thallus, verrucae 20-30 um, transversely septate ascospores 14-18.5 x 3-4

um. Mazosia pseudobambusae Kalb & Vézda differs from the new species

by its very pale brownish grey thallus, dark brown prothallus, and larger

(20-28 x 4-5 um) ascospores (Liicking 2008). The new species is also similar

to M. melanophthalma, which differs by its larger (50-100 um) verrucae,

longer (15-22 um) ascospores, and an ascomatal margin that always slopes

outwards (Liicking 2008; Conran & Rogers 1983). Another related species

is M. dispersa (J. Hedrick) R. Sant., distinguished by its less finely verrucose

thallus and longer (26-39 um) ascospores (Lticking 2008).

Key to species of Mazosia known from China

Ire LiialltistelabrOuss.. cautunteatHenteak He cheats Se ptide Monk a Re kre Meh tee t ate hehe nk Has 2

1. Thallus with radiate ridges or finely to coarsely verrucose ...............0 00000. 3

2 PASeOSpoles: S25 IN 1ONS Boe esi ae BE ctr itn wR ita atta ae M. phyllosema

2. Ascospores 25-35 um long, irregularly 3-7-septate,................ M. paupercula

612 ... Yao, Jia, Jiang

3. Thallus with radiate ridges, ascospores 3-septate, 14-22 x 4-6 um ....... M. rotula

3. Thallus lacking radiate ridges, thallus verrucose ............ 0. cece eee eee eee eee 4

4. Ascospores 5(—7)-septate, 26-39 um long ............. 0. cece eee eee M. dispersa

4, ASCOSPOFes. 3-SEptate Viner es hee cee ee lates Helicase Hecate alpen patos Spares ee 5

5. Verrucae 50-100 um tall, ascospores 15-22 x 3-4um......... M. melanophthalma

5. Verrucae 20-30 um tall, ascospores 14-18.5 x 3-4 um ............ M. hainanensis

Conclusion

The morphological and molecular data demonstrate that the material

from Hainan province represents a previously unknown foliicolous lichen

that merits the status of a new species in Mazosia. The preliminary key is

provided to the species of Mazosia currently known from China.

Acknowledgments

This research was funded by the National Natural Science Foundation of China

(Project No. 31800010, 31750001). The authors are grateful to the Dr. L.L. Zhang

(Institute of Environment and Ecology, Shandong Normal University, China) and

Dr. L. Lii (College of Food Science and Engineering, Qilu University of Technology,

Jinan, China), reviewers for the manuscript.

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MY COTAXON

July-September 2021— Volume 136, pp. 615-626

https://doi.org/10.5248/136.615

Racocetra cromosomica sp. nov. from Oaxaca, Mexico

EDUARDO CHIMAL-SANCHEZ’*”, IRMA REYES-JARAMILLO’,

SARA LuciA CAMARGO-RICALDE’, LUCIA VARELA’,

JOSE YABIN SALMERON CASTRO’, NOE MANUEL MONTANO'™*

"Departamento de Biologia, Divisién de Ciencias Bioldgicas y de la Salud,

Universidad Auténoma Metropolitana-Iztapalapa,

Av. San Rafael Atlixco 186, Col. Vicentina, 09340, Ciudad de México, México

’ Facultad de Estudios Superiores-Zaragoza, Universidad Nacional Autonoma de México,

Batalla del 5 de Mayo s/n, Col. Ejército de Oriente, 09230, Ciudad de México, México

* CORRESPONDENCE TO: nmma@xanum.uam.mx

AsstTRAcT—Analysis of soil samples collected in the rhizosphere of Agave karwinskii

in the Central Valleys of Oaxaca, Mexico, revealed an undescribed species of Racocetra

that possesses ornamented spores. The ornamentation on the outer layer of the external

spore wall consisted of rounded and elongated processes, which are organized in such a

way that they look like chromosomes. Therefore, the new species was named Racocetra

cromosomica. It is the first species of Racocetraceae (Gigasporales) described from Oaxaca,

one of the Mexican states having the greatest floristic diversity.

Key worps—arbuscular mycorrhizal fungi, Glomeromycota, mezcal, semiarid ecosystems,

taxonomy

Introduction

Mexico has a wide variety of ecosystems, and it is the center of origin,

diversification and domestication of several plant species (Casas 2019).

Oaxaca is one of the Mexican states characterized by the highest floristic

diversity, comprising 8431 plant species (Garcia-Mendoza & al. 2004). The

diversity in Oaxaca arises from the convergence of five biogeographical

provinces varying in climates, soils, and vegetation types (Morrone 2019),

such as xerophilous scrublands and tropical dry forests, containing plants

616 ... Chimal-Sanchez & al.

of Agavaceae, a family endemic to America and one of ten monocot families

with the highest species richness. In Oaxaca, the Agave genus is highly

diverse, containing 32 of the 159 taxa distributed in Mexico (Garcia-Mendoza

& Galvan 1995, Garcia-Mendoza & al. 2004), all with great ecological,

economic, and cultural importance for the inhabitants (Torres-Garcia

& al. 2019). Several Agave species are used to prepare mezcal, and people

identify both agaves and mezcal as symbols of cultural identity. In preparing

mezcal in the Central Valleys and the Sierra Sur of Oaxaca, people use Agave

vivipara L. [= A. angustifolia Haw.], known in Mexico as agave espadin, and

some wild species, as Agave karwinskii Zucc. (Ordohez & Rodriguez 2008,

Garcia-Mendoza & al. 2004, 2017). Several Agave species are listed in the

IUCN-Red List of Threatened species, including A. karwinskii, due to its

limited distribution range (14,492 km?) and the rapidly increasing demand

for mature individuals for mezcal production (Garcia-Mendoza & al. 2019).

Arbuscular mycorrhizal fungi (AMF) species (Glomeromycota) form

mutualist symbiosis with almost 85% of land plant families, including

Agavaceae (Carballar-Hernandez & al. 2013, Goto & al. 2013; Chimal-Sanchez

& al. 2018), and they promote the growth and nutrition for most plants

(Smith & Read 2008), including A. tequilana EA.C. Weber ‘Azul’ (Montoya-

Martinez & al. 2019). Thus, in Mexico, AMF could be very important not

only for conservation of floristic diversity but also for production of agave

plants. In Mexico, currently 160 species of AMF are registered; however, the

morphology of many AMF species found in the country suggests that they

are undescribed (Varela & al. 2019, Polo-Marcial & al. 2021).

During a study on AMF associated with rhizosphere soil of mezcal agaves

(A. vivipara, A. karwinskii) in the Central Valleys and Sierra Sur of Oaxaca,

Mexico (the mezcal range), four new records and 48 morphospecies

were registered (Chimal-Sanchez & al. 2018, Reyes-Jaramillo & al. 2019);

however, ca. 30% of the AMF listed exhibit a morphology that does not fit

those described species. Among them is a member of Racocetraceae (for this

family see: Silva & al. 2012, Goto & al. 2012, Souza & al. 2018, Wijayawardene

& al. 2020) that was registered in the rhizosphere of A. karwinskii and

reported as Racocetra sp. 1 by Reyes-Jaramillo & al. (2019). Its subcellular

spore structure, germination shield morphology, and histochemical

properties indicates that it does belong in Racocetra (Gigasporales). Most

importantly, spores of this fungus have a unique ornamentation consisting

of chromosome-like outgrowths. Therefore, the fungus is here described as

Racocetra cromosomica.

Racocetra cromosomica sp. nov. (Mexico) ... 617

Materials & methods

The Central Valleys (CV) and Sierra Sur of Oaxaca (SSO) are two of the eight

economic regions of the State of Oaxaca. The CV and SSO belong to the “Sierra

Madre del Sur” biogeographic province (Morrone 2019). The climate of the region

is semi-dry, warm to semi-warm, with rains in summer. The soils according to

World Reference Base for Soil Resources (WRB 2014) are Regosols and Leptosols,

although Calcisols and Phaeozems are also present. The vegetation corresponds

to tropical dry forests and xerophytic scrubland that have been strongly altered by

agricultural activities, so that there are only fragments of this vegetation, with wild

populations of A. karwinskii and cultivated fields with A. vivipara (INEGI 2010,

Reyes-Jaramillo & al. 2019).

In the Central Valleys and Sierra Sur of Oaxaca, seven study sites (S) with wild

populations of A. karwinskii were established in the municipalities of San Pedro

Totolapan (S1), San Agustin Amatengo (S3, S4), Ejutla (S5), and San Luis Amatlan

(S6, S9), while only two sites were established in fields cultivated with Agave vivipara

in Ejutla (S2) and San Luis Amatlan (S7). The soils have a texture of Loamy sand

(S1, S3-S9) or Sandy loam (S2), and pH ranging from neutral (S1, pH = 7.5; S2,

pH = 7.19, and S3, pH = 7.28) to lightly alkaline (S4, pH = 7.86; S5, pH = 8.06; S6,

pH = 8.02; S7, pH = 8.04; S8, pH = 7.96, and S9, pH = 8.0). On average, they are

rich in organic matter (OM = 2.65% from S1-S4; OM = 4.9% from S5-S9), poor

in available phosphorus (PO,- Olsen = 1.6 mg kg"), and rich in extractable bases

of calcium and magnesium, but low in potassium and sodium (Reyes-Jaramillo &

al. 2019). However, our new Racocetra species was registered only in San Agustin

Amatengo (S3) in soil with the highest percentage of sand (79%). Our new Racocetra

coexisted with other AMF species, the most frequent of were Acaulospora rehmii

Sieverd. & S. Toro, A. scrobiculata Trappe, Cetraspora pellucida (T.H. Nicolson &

N.C. Schenck) Oehl & al., Gigaspora gigantea (T.H. Nicolson & Gerd.) Gerd. &

Trappe, Funneliformis halonatus (S.L. Rose & Trappe) Oehl & al., F. mosseae (T.H.

Nicolson & Gerd.) C. Walker & A. Schiifler, and Racocetra fulgida (Koske & C.

Walker) Oehl & al. (Reyes-Jaramillo & al. 2019).

Pot trap cultures were established with native soil samples (600g per pot) and

grown without fertilization in glasshouse conditions (Stutz & Morton 1996) using

corn (Zea mays L.) and “guaje” (Leucaena sp., Leguminosae) as host plants. Host

plants were grown for six months and watered every third day with distilled water,

and then left to dry for two weeks to favor formation of AMF spores. Due to the

small number of Racocetra spores obtained (1 spore per 300 g dry soil), only one

single-species culture was set using two spores as inoculum and the same host plants.

Unfortunately, this attempt failed, and no newly produced spores and mycorrhizal

structures were found. Consequently, the new Racocetra species is here described

based on field-collected spores and those extracted from trap cultures.

Racocetra spores were isolated by wet sieving and decanting (Gerdemann &

Nicolson 1963), followed by centrifugation in 60% sucrose solution to eliminate

organic matter and mineral remains. Several spores were then placed in Petri

618 ... Chimal-Sanchez & al.

dishes with distilled water, and their color was determined under a dissection

microscope using the web color chart hosted by the International Culture

Collection of (Vesicular) Arbuscular Mycorrhizal Fungi (INVAM 2018). Ten spores

were mounted on microscopic slides in polyvinyl alcohol lacto-glycerol (PVLG)

and in PVLG mixed with Melzer’s reagent 1:1 (v/v). Terminology for the species

description, spore characteristics, and spore wall nomenclature are those adopted

from Walker (1983) and Oehl & al. (2008). Likewise, we used an updated taxonomic

key for Racocetra species proposed by Souza & al. (2018). Voucher specimens were

deposited in the Herbarium of Instituto Politécnico Nacional, Mexico City, Mexico

(ENCB), the Herbarium of Universidad Aut6noma de Tlaxcala, Tlaxcala state,

Mexico (TLXM), and the Laboratory of Legume Biosystematics, Department of

Biology, Universidad Auténoma Metropolitana-Iztapalapa, Mexico City, Mexico

(UAMI).

Taxonomy

Racocetra cromosomica Chim.-Sanch., Varela & Montano, sp. nov. FIGS 1, 2

IF 557326

Differs from all other Racocetra and gigasporalean species by its unique ornamentation

formed on the outer spore wall, consisting of rounded and elongated processes

resembling condensed chromosomes.

TypE—Mexico, Oaxaca, Central Valleys in San Agustin Amatengo, 16°31’06”N

96°47'08”W; isolated from the rhizospheric soil of Agave karwinskii growing within a

xerophytic scrubland (SAASM) and coming from a propagation pot (SAASM-Prop)

with Zea mays and Leucaena sp. (guaje) as trap plants; isolated June 2017, E. Chimal-

Sanchez (Holotype, ENCB121802 [SAASM-Prop-slide-001]; isotypes, TLXM-5034

[SAASM-Prop-slide-002], VAMI-002 [SAASM-Prop-slide-003]).

ErymMoLocy—referring to the distinctive ornamentation (rounded and elongated

processes) on the outer spore wall that resembles chromosomes.

SPOROCARP formation is unknown. Spores are formed singly in the soil and

terminally on a bulbous sporogenous cell. Spores are red-brown (20-80-60-0)

to dark red-brown (60-80-100-0); globose, 368.6-445.3 um.

Fic. 1. Racocetra cromosomica (holotype, ENCB121802): a, b. Spore (S) in water (a) and PVLG

(b) with bulbous sporogenous cell (sc) characteristic of the Gigasporales; c. The spore surface of

an intact spore ornamented with elongated processes seen in plan view; d. Outer spore wall (OW)

separated from inner spore wall (IW) of a crushed spore; the unique chromosome-like processes

ornamenting the upper surface of the outer spore wall are visible; e. Outer (OW) and inner (IW)

spore wall and bulbous sporogenous cell with no reaction in Melzer’s reagent; f. Outer spore wall

with three permanent layers (OWL1-3), of which OWLI is ornamented, and inner spore wall

(IW); g, h. The upper surface of layer 1 of the outer spore wall ornamented with rounded (RP) and

elongated (EP) processes; i. The distribution and arrangement of rounded (RP) and elongated (EP)

processes (chromosomes-like) on the spore surface.

Racocetra cromosomica sp. nov. (Mexico) ... 619

620 ... Chimal-Sanchez & al.

SPORE WALL comprising two walls: an outer wall (OW) and an inner wall

(IW). OW has three closely adherent layers (OWL1, OWL2, and OWL3) of

a total thickness of (15.4-)16.6(-17.6) um. OWL] is a permanent rigid layer,

yellow-brown (0-10-40-0) to dark red-brown (60-80-100-0). The surface of

this layer has double ornamentation resembling condensed chromosomes

when observed in plan view. The first ornamentation type comprises

rounded processes, (1.9-)3.7(-5.4) um high and (3.8-)4.8(-5.7) um wide at

their base. Regularly, these processes are arranged in groups of 2, 4, 6, 8, or

more, and when grouped they have a width of (11.1-)11.9(-12.7) um and a

height of (4.9-)5.1(-5.4) um. The second type of ornamentation consists of

elongated outgrowths, which are fused to the rounded ones and 1, 2, 3 or 4

elongated processes may be attached. In addition, the elongated processes

may be connected to another group of rounded processes forming an

irregular reticulum when the spores are observed in plan view. The elongated

processes are claviform, (10.5-)16.4(-21.2) um long, (4.4-)5.2(-6.5) um

wide, and (2.8-)3.4(-4.3) um high. Both types of ornamentations are yellow-

brown (0-10-40-0) to dark red-brown (60-80-100-0). The second layer of

the spore wall (IWL2) is laminated, (9.1-)10.3(-12.4) um thick, smooth, and

hyaline (20-0-20-0). OWL3 is hyaline, often difficult to observe as it closely

adheres to OWL2, <1 um thick. None of the spore wall layers stained in

Melzer’s reagent.

INNER WALL has two adherent layers (IWL1 and IWL2) that are semi-

flexible, hyaline (30-0-20-0), jointly <5 um thick, and do not stain in

Melzer’s reagent.

GERMINATION SHIELD formed on the outer surface of IW. It is hyaline

(20-0-20-0), oval, (149.7-)165.5(-181.7) um long x (102.4-)126.5(-140.9)

Fic. 2. Racocetra cromosomica (isotype, TLXM-5034): a, b. The arrangement and shapes of

rounded (RP) and elongated (EP) processes (chromosomes-like) on layer 1 of the outer spore

wall from spores crushed in Melzer’s reagent; c. Ornamented, dark-colored layer 1 (OWL1)

and smooth, colorless layer 2 (OWL2) of the outer spore wall; d. Three-layered outer spore wall

(OWL1-3) and two-layered inner spore wall (IWL1-2); e-g. Spores with germination shields

(gs) composed by multiple small compartments (6-8), each with generally one germ tube

initiations (gti); h. Germination from gti; note the thick and reddish brown color of the hyphae

(H); i, j. Subglobose to ovoid sporogenous cell (sc) with a septum (S) at its base; note sc has no

ornamentations and its wall consists of only two layers (scwl2 and scwl3) continuous with outer

spore wall layers 2 and 3.

Racocetra cromosomica sp. nov. (Mexico) ... 621

622 ... Chimal-Sanchez & al.

um wide, multilobed (6-8), each lobe generally with one germ tube initial

(gti) from where germination tubes emerge.

BULBOUS SPOROGENOUS CELL formed terminally from an extraradical

mycorrhizal hypha, yellow brown (20-40-100-0), sub-globose to ovoid,

(56.1-)61.3(-63.9) um wide and (68.9-)74.6(-77) um long in the region

from the spore base to the subtending hypha septum. Sporogenous cell wall

smooth with two layers (scwl2, scwl3) continuous with OWL2 and OWL3.

Scwll is absent since the bulb does not have the ornate layer of the spores

(OWL}1), scwl2 is yellow-brown (20-40-100-0), 2-3 um thick, and scwl3 is

yellow (0-30-80-0), 1.1-1.4 um thick.

AUXILIARY CELLS were not observed.

DISTRIBUTION & ECOLOGY — Racocetra cromosomica is known only

from Central Valleys, San Agustin Amatengo, Oaxaca, Mexico. The spores

of R. cromosomica have been recorded only from soil associated with the

rhizosphere of A. karwinskii growing in a xerophilous scrubland with

remnants of other A. karwinskii wild populations. Soil characteristics are

= 7.2, soil organic matter = 3.2%, available

« Kg’ Mg = 7.5

cmoles,,, kg", K = 0.29 cmoles,,, kg", and Na = 0.14 cmoles,,) kg* (Reyes-

Jaramillo & al. 2019). Racocetra cromosomica probably has preference for

sandy soils like other Gigasporales (Lekberg & al. 2007; Vieira & al. 2020).

MYCORRHIZAL ASSOCIATIONS unknown. In the AMF multi-species

loamy sandy (79% sand), pH,,,.,

phosphorus (PO,-Olsen) = 0.7 mg kg’, Ca = 15 cmoles

propagation pot, R. cromosomica was propagated with maize (Zea mays)

and “guaje” (Leucaena sp.) as host plants, but its sporulation was very low

(one spore per 300 g dry soil). However, the presence of spores in these

cultures suggested that R. cromosomica formed arbuscular mycorrhiza, as

do other Racocetra species grown in single-species cultures.

Discussion

The formation of spores from a bulbous sporogenous cell, the subcellular

structure of spores containing two spore walls, the features of the spore

germination shield, and the lack of staining reaction of the inner spore wall

in Melzer’s reagent unambiguously support the fungus collected by us in

Mexico and here described as a new species of Racocetra.

The ornamentation of the upper surface of Racocetra cromosomica spores,

consisting of rounded and elongated processes that are organized as if they

were chromosomes, is a unique and very important diagnostic character that

easily distinguishes this species from other Racocetra species and species

Racocetra cromosomica sp. nov. (Mexico) ... 623

of other gigasporalean genera producing ornamented spores. Spores of

R. undulata T.C. Lin & C.H. Yen are much smaller (195-225 um vs.

368.6-445.3 um) and are ornamented with pit undulations (Lin & Yen 2011),

and the spore ornamentation in R. persica (Koske & C. Walker) Oehl & al.,

consists of fine spines (Oehl & al. 2008, Souza & al. 2018). Racocetra minuta

(Ferrer & R.A. Herrera) Oehl & al., has a double ornamentation on the outer

spore wall, but this ornamentation is composed of projections with central

depressions (Oehl & al. 2008, Souza & al. 2018). Intraornatospora intraornata

Goto & al., (= R. intraornata B.T. Goto & Oehl) produces ornamented

gigasporoid spores, but the ornamentation consists of densely packed tubes

that, importantly, occur on the lower surface of a laminate layer of the outer

spore wall (Goto & al. 2009, 2012).

The only Racocetra species that have spores ornamented with processes

(outgrowths resembling warts) are R. beninensis Oehl & al., R. verrucosa

(Koske & C. Walker) Oehl & al., R. coralloidea (Trappe & al.) Oehl & al.,

R. gregaria (N.C. Schenck & T.H. Nicolson) Oehl & al., and R. crispa RA. Souza

& al. However, the spores of both R. beninensis and R. verrucosa are lighter

in color and ornamented with processes that are smaller (0.9-2.1(-2.8) um

long, 0.9-3.8 um wide in R. beninensis; 0.5-1.5 um long, 0.5-1.5 um wide in

R. verrucosa) and simpler in construction (Koske & Walker 1985, Tchabi & al.

2009). Racocetra coralloidea spores are ornamented with flattened warts with

angular margins (most 2-12 um wide x 1-3 um high) and those of R. gregaria

with rounded warts (most 11-26.7 um wide x 3-7 um high) that differ in size

and are clearly separated from each other, never occurring in groups as in

R. cromosomica (Oehl & al. 2008, Souza & al. 2018). The ornamentation of

R. crispa clearly differs from that of R. cromosomica by displaying multiple

cloud-like projections with a basal diameter of (5-)15-41 um and a height of

(5.2-)9.5-18 um (Souza & al. 2018).

Other species of the Gigasporales that can be morphologically confused

with R. cromosomica due to the formation of similarly colored ornamented

spores are Scutellospora dipapillosa (C. Walker & Koske) C. Walker & FE.

Sanders, Dentiscutata colliculosa B.T. Goto & Oehl, D. nigerita Khade,

D. nigra (J.E. Redhead) Sieverd. & al., and D. reticulata (Koske & al.) Sieverd.

& al. Apart from morphological differences in the spore ornamentation,

these species also differ from R. cromosomica by possessing three spore

walls instead of two (Koske & Walker 1985, Goto & al. 2010, Khade 2010,

Oehl & al. 2008).

Although R. cromosomica sporulated in multi-species trap cultures, the

low numbers of spores obtained did not allow us to conduct molecular

and phylogenetic analyses to determine the position of this species among

sequenced Racocetra spp. Nonetheless its spore morphology both clearly

624 ... Chimal-Sanchez & al.

supports its novelty and easily distinguishes R. cromosomica from other

ornate species in the Gigasporales.

We are aware that R. cromosomica should be characterized molecularly

in the future to track occurrence elsewhere based on environmental studies.

Finally, the fact that R. cromosomica has so far been found only in the

rhizosphere of A. karwinskii, a highly vulnerable species endemic to Mexico,

suggests that A. karwinskiiis critical for in situ conservation of this mycorrhizal

fungus and that this new Glomeromycota species could be essential in the

recovery of A. karwinskii populations within mezcal producing regions in

Oaxaca, Mexico, particularly in the town of San Agustin Amatengo.

Acknowledgements

We thank Genaro Sierra for his support in the field collection in the town of

Ejutla, Oaxaca as well as Rosalva Garcia Sanchez, Alejandro Alarcon, Ronald Ferrera

Cerrato, Sandra Cortés; Maria Eugenia Fraile, and Rosaura Grether for their support

with the microphotographs. L. Varela was supported by the professorship “Ramon

Riba y Nava Esparza” at the Universidad Autonoma Metropolitana-Iztapalapa. We

thank Janusz Blaszkowski (West Pomeranian University of Technology, Szczecin

Poland) and Bruno Tomio Goto (Universidade Federal do Rio Grande do Norte,

Brazil) for reviewing our manuscript and their helpful, constructive comments

and suggestions that improved this paper. We are also grateful to Nomenclature

Editor Shaun Pennycook and Editor-in-Chief Lorelei L. Norvell for their excellent

reviews.

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https://doi.org/10.17129/botsci.2202

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ed. electrénica, UNAM, México. 1487 p.

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arbuscular mycorrhizalfungicommunityalonganenvironmental gradient. Plants 9(52):[16p.].

https://doi.org/10.3390/plants9010052

Walker C. 1983. Taxonomic concepts in the Endogonaceae: spore wall characteristics in species

descriptions. Mycotaxon 18: 443-455.

Wijayawardene NN, Hyde DK, Al-Ani LKT, Tedersoo L, Haelewaters D, Rajeshkumar

KC & al. 2020. Outline of fungi and fungi-like taxa. Mycosphere 11(1): 1060-1456.

https://doi.org/10.5943/mycosphere/11/1/8

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maps. World Soil Resources Reports No. 106. FAO, Rome. 186 p.

MY COTAXON

July-September 2021— Volume 136, pp. 627-634

https://doi.org/10.5248/136.627

Bactrospora cozumelensis sp. nov. from Mexico

YENITZE A. GARCIA-MARTINEZ’, JORGE GUZMAN-GUILLERMO’,

RICARDO VALENZUELA’, TANIA RAYMUNDO*

' Instituto Politécnico Nacional, Escuela Nacional de Ciencias Biol6gicas,

Plan de Ayala y Carpio s/n Col. Santo Tomas, México, D.F. 11340, México

? Universidad Veracruzana, Facultad de Biologia,

Circuito Gonzalo Aguirre Beltran s/n, Zona Universitaria,

Xalapa-Enriquez, 91090, México.

*CORRESPONDENCE TO: traymundoo@ipn.mx

ABSTRACT—Bactrospora cozumelensis is described as a new species associated with mangrove

forests in Cozumel Island, Mexico. The specimens were collected on Rhizophora mangle bark

in Punta Sur Ecological Park, Cozumel Island Biosphere Reserve, Quintana Roo, Mexico.

This species is characterized by an erumpent to sessile apothecium, laterally carbonized

excipulum, and 3-septate ascospores that are slightly constricted in the middle.

Key worps—Arthoniales, lichens, Roccellaceae, taxonomy

Introduction

Bactrospora A. Massal. comprises 37 species of crustose lichenized fungi

that grow mainly on the bark of trees (Wijayawardene & al. 2018, Van

den Boom & al. 2018, Herrera-Campos & al. 2019). The genus is widely

distributed in both tropical and temperate areas but often goes unnoticed

(Sobreira & al. 2015). It is characterized by a thallus that is poorly developed,

crustose, and associated with photobionts of Trentepohliaceae, bitunicate

asci with an apical KI+ blue apparatus, and ascospores that are cylindrical to

acicular and multiseptate (Egea & Torrente 1993). Bactrospora differs from

similar genera such as Lecanactis or Opegrapha by its cylindrical to acicular

and multiseptate ascospores, its apical ascal apparatus with a conspicuous

KI+ blue ring, its pruinose thallus or disk, and its ascomatal shape (Egea

628 ... Garcia-Martinez & al.

& Torrente 1993). Bactrospora displays four ascospore types (dryina-type,

patellarioides-type, homalotropa-type, and jenikii-type) and its asexual form

produces pycnidia that are immersed or subimmersed in the thallus or

substratum and produce variably shaped conidia (Egea & Torrente 1993).

The genus was initially placed in the Bactrosporaceae by Rabenhorst

(1869: 37, 60, as “Bactrosporeae”) and this was accepted by Eriksson (1981).

Subsequently, it was regarded as a synonym of the Opegraphaceae (Cannon &

al. 1985, Eriksson & Hawksworth 1991), and later Hawksworth & al. (1995)

and Grube (1998) included it in Roccellaceae (Arthoniales) based on macro-

and micromorphological and chemical features. Recent phylogenetic studies

treat Bactrospora as a genus incertae sedis within Arthoniales (Licking & al.

2016, Wijayawardene & al. 2018).

In the Americas, 17 Bactrospora species have been reported, mostly

from coastal ecosystems (Egea & Torrente 1993, Egea & al. 2004). Although

Mexico is a megadiverse country, the lichen biota is insufficiently studied,

and only two Bactrospora species have been reported: B. jenikii (Vézda)

Egea & Torrente from Quintana Roo (Guzman-Guillermo & al. 2019) and

B. lecanorina Herrera-Camp. & al. from Jalisco (Herrera-Campos & al.

2019); both species were discovered in tropical forests.

During a mycological survey of mangrove on Cozumel Island, we

collected specimens belonging to this genus that could not be assigned to

any published species and so propose a new species here.

Materials & methods

The mangrove forests lie within Cozumel Island Biosphere Reserve, in the Punta

Sur Ecological Park, 20°16’41”-18’27”N 86°58’05”-87°00'41’W, covering 1114 haat

0-7 maz.s.l. The climate is tropical with an average temperature of 25°C and a mean

annual precipitation of 1570 mm (annual maximum of 250 mm occurring in October

and minimum of 40 mm occurring in April; INEGI 2013). The mangrove forests

include Rhizophora mangle L., Laguncularia racemosa (L.) C.F. Gaertn., Conocarpus

erectus L., and Avicennia germinans (L.) L. (Téllez- Valdez & al. 1989). The specimens

were collected in Punta Sur Ecological Park on 18 May 2019 on R. mangle bark and

are deposited in fungal collection “Dr. Gast6n Guzman Huerta” at the Herbarium

Escuela Nacional de Ciencias Biolégicas of Instituto Politécnico Nacional, Mexico

City, Mexico (ENCB) and the Herbarium in Universidad Veracruzana, Xalapa,

Veracruz, Mexico (XALU). Latitude/longitude coordinates were obtained with

GPS eTrex (Garmin). Morphological examinations were conducted as outlined by

Brodo & al. (2001). The specimens were morphologically examined using a Carl

Zeiss Primo Star microscope, and hand-cut sections were examined under a Carl

Zeiss model Stemi Dv4 stereoscope. Ascospore measurements were made in water

Bactrospora cozumelensis sp. nov. (Mexico) ... 629

Fics 1-6: Bactrospora cozumelensis (holotype, ENCB—Garcia & Bravo 31a). 1. Gregarious

young and mature apothecia on dead wood; 2. Apothecia mature, discoid; 3. Apothecium in

transverse section; 4. Excipulum in lateral section; 5. Under side of subhymenium; 6. Excipulum

after KI.

630 ... Garcia-Martinez & al.

at 1000x magnification and only well-developed ascospores lying outside the asci

were measured. Amyloidy was tested with Lugol’s solution before and after pre-

treatment with KOH 5%. Spot tests were made using standard methods (Brodo

& al. 2001). A thin-layer chromatography (TLC) was performed with solvent C

(Elix 2014). Measurements of micromorphological characters were taken from

rehydrated tissues in water. Terminology is based on Egea & Torrente (1993).

Taxonomy

Bactrospora cozumelensis Y. Garcia, Guzm.-Guill., R. Valenz. &

Raymundo, sp. nov. Figs 1-11

MB 836514

Differs from Bactrospora incana by its smaller ascospores, its epruinose apothecia,

and its corticolous habit.

Type: México, Quintana Roo, municipality of Cozumel, Punta Sur Ecological Park,

20°18’00’N 87°00'36”W, 0 m a.s.l., mangrove forest in a coastal lagoon, on bark of

Rhizophora mangle (Rhizophoracaeae), 18 May 2020, Y.A. Garcia & M.A. Bravo 31a

(Holotype, ENCB).

ETyMOLOoGy: cozumelensis refers to the type locality.

THALLUS crustose, continuous, effuse, thin, whitish, slightly shiny,

surrounded by a black prothallus line; I-, K/I-, and Cl-; trentepohlioid

ellipsoid cells inside of the thallus; ApoTHEcrIA 0.1-0.3 mm diameter,

roundish, dark, with thin margin to immarginate distinguishable for being

darker, epruinose, initially erumpent on the bark, turning sessile at maturity,

numerous, scattered in the thallus; ExcipuLum 25-30 um at the top and

40-60 um at the base, laterally carbonized, open below the subhymenium,

I-, K/I+ deep blue; HymEentum (85-)130-200(-230) um, not inspersed, I+

reddish, K/I-; SuBHYMENIUM 100-130 um, hyaline I-, K/I+ blue. Paraphyses,

long, branched, forming a reticulate, light brown pseudoepithecium without

pruinose granules, K+ dark olive, I- and KI-; Asc (40—)45-52(-55) x 8-10

um (n = 20), cylindrical, hyaline, bitunicate, stipulated; AscosPorEs eight

per asci, (15-)17-20(-23) x (2-)3-4 um (n = 35), of jenikii-type, hyaline,

with 3(—4) transverse septa, slightly constricted in the middle.

PYCNIDIA not seen.

CHEMISTRY: K-, C-, UV-. TLC: no substances detected.

ADDITIONAL SPECIMEN EXAMINED: MEXICO, QuINTANA Roo, Municipality of

Cozumel, Punta Sur Ecological Park, 20°18’00”N 87°0’36”W, 0 m a.s.l., mangrove

forest in a coastal lagoon, on bark of Rhizophora mangle, 18 May 2020, Y.A. Garcia &

M.A. Bravo 32 (XALU).

ECOLOGY & DISTRIBUTION: Growing on primary branch bark of Rhizophora

mangle in a mangrove forest situated in a coastal lagoon; known only from

Bactrospora cozumelensis sp. nov. (Mexico) ... 631

LPHA |

be UY) Sum

Saree

Fics 7-12: Bactrospora cozumelensis (holotype, ENCB—Garcia & Bravo 31a). 7. Hymenium in

water; 8. Paraphyses; 9. Asci in water; 10. Asci after KI; 11, 12. Ascospores in 5% KOH.

the type locality. Bactrospora cozumelensis was found growing together with

other lichens including Pyrenula cerina Eschw., Coniocarpon cinnabarinum

DC., and Arthonia antillarum (Fée) Nyl.

632 ... Garcia-Martinez & al.

COMMENTS—Bactrospora cozumelensis is diagnosed by apothecia that are

subimmersed when immature and sessile when mature and its 3(—4)-septate

ascospores that are slightly constricted in the middle. The similar species

Bactrospora brevispora R.C. Harris is distinguished by its sessile apothecia,

an excipulum closed below the subhymenium, and larger (20-32 x

3-3.5 um) ascospores (Harris 1990; Egea & Torrente 1993). Bactrospora

incana Egea & Torrente, which also resembles B. cozumelensis, differs by

its larger (28-42 x 4.5-6.5) ascospores, densely hairy young ascomata, and

epilithic habitus (Egea & Torrente 1993) (TABLE 1). Neither B. brevispora

nor B. incana have been reported in mangrove forests.

TABLE 1. Comparisons between Bactrospora cozumelensis and related species.

CHARACTER B. BREVISPORA B. COZUMELENSIS B. INCANA

Unicellular hairs Absent Absent Present

on thallus

Ascomata size 0.3-0.8 0.1-0.3 0.2-0.4

(mm)

Excipulum opening Closed Open Open or

below subhymenium reduced to a thin band

Ascus size (45-)50-70 (40-)45-52(-55) 60-75

(um) x 10-12 x 8-10 x 18-21

Ascospore type jenikii-type jenikii-type jenikii-type

Ascospore size 20-32(-35) (15-)17-20(-23) 28-42

(um) x 3-3.5(-4) x (2-)3-4 x 4.5-6.5

Number o 4-9 asi 3-7

ascospore septa

Habitus Epiphyte Mangrove epiphyte Epilithic

Distribution USA, Jamaica Mexico Venezuela

(Egea & Torrente (this publication) (Egea & Torrente

1993) 1993)

In the Americas, only three other species of Bactrospora show the jenikii-

type ascospores produced by B. cozumelensis: B. brevispora, B. incana, and

B. jenikii; of these, B. jenikii with 8-13 septa is the most distinct (Sobreira

& al. 2015) while B. cozumelensis has the smallest jenikii-type spores. Of the

three Bactrospora species reported in Mexico, only B. cozumelensis is found

in mangrove forest; B. lecanorina, found in a dry tropical forest in Jalisco,

has patellarioides-type ascospores with more septa (Herrera-Campos & al

2019), and B. jenikii, found in sub-deciduous forest in Bacalar lagoon, has

larger ascospores (Guzman-Guillermo & al. 2019).

Bactrospora cozumelensis sp. nov. (Mexico) ... 633

Acknowledgments

We wish to express our gratitude to Dr. André Aptroot (Lb. Botanico, UFMS,

Campo Grande, Brazil) and Dr. Israel Pérez Vargas (Dpt. Botanica, Ecologia y

Fisiologia Vegetal, Universidad de La Laguna, Canary Islands, Spain) for reviewing

the manuscript and their useful comments. We want to thank Lic. Emilio Villanueva

(Head, Research of the Parks and Museums Foundation of Cozumel) for the facilities

granted during specimen collection and Lic. José de Jesus Benavides Andrade

(Director for Punta Sur Eco Beach Park). This study was supported by CONACYT

projects 252934 and by the Instituto Politécnico Nacional (IPN) through the

Secretaria de Investigacién y Posgrado (SIP), projects 20210315 and 20210661.

RV thanks the Comisidn de Operacién y Fomento de las Actividades Académicas,

IPN (COFAA). RV and TR thank Secretaria de Investigacion y Posgrado, IPN and

Sistema Nacional de Investigadores (SNI, CONACYT) for the support received

through scholarships. YG thanks the CONACYT for the scholarship awarded to

carry out their master’s degree in Posgrado en Biociencias of the Escuela Nacional

de Ciencias Biolégicas and program BEIFI of the SIP.

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de Quintana Roo, México. Boletin Micoldgico 34 (2): 25-32.

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Garden, USA.

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the Fungi. 8th ed. CAB International, Wallingford, UK. 616 p.

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estudios/conociendo/QUINTANA_ROO.pdf

Licking R., Hodkinson BP, Leavitt SD. 2016. Classification of lichenized fungi in the Ascomycota

and Basidiomycota - approaching one thousand genera. Bryologist 119: 361-416.

https://doi.org/10.1639/0007-2745-119.4.361

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167-236. https://doi.org/10.1007/s13225-018-0394-8

MY COTAXON

July-September 2021— Volume 136, pp. 635-644

https://doi.org/10.5248/136.635

Lembosia mimusopis sp. nov. from Thailand

DIANA S. MARASINGHE””, MONIKA C. DAYARATHNE’?”?,

SAJEEWA S.N. MAHARACHCHIKUMBURA‘, ABDALLAH M. ELGORBAN®*”,

SINANG HONGSANAN’, KEVIN D. HyDE??*

‘Center of Excellence in Fungal Research, Mae Fah Luang University,

Chiang Rai 57100, Thailand

*The Faculty of Science, Mae Fah Luang University, Chiang Rai, 57100, Thailand

°Key Laboratory for Plant Biodiversity and Biogeography of East Asia (KLPB),

Kunming Institute of Botany, Chinese Academy of Science,

Kunming, Yunnan 650201, China

‘School of Life Science and Technology,

University of Electronic Science and Technology of China (UESTC),

No. 2006, Xi- Yuan Avenue, High-Tech West Zone, Chengdu 611731, PR. China

°Center of Excellence in Biotechnology Research, King Saud University,

PO. Box. 2455, Riyadh 11451, Saudi Arabia

°Department of Botany and Microbiology, College of Sciences, King Saud University,

PO. Box. 2455, Saudi Arabia, Riyadh 11451, Saudi Arabia

7Shenzhen Key Laboratory of Microbial Genetic Engineering,

College of Life Sciences and Oceanography, Shenzhen University,

Shenzhen, Guangdong 518060, PR China

“ CORRESPONDENCE TO: kdhyde3@gmail.com

ABSTRACT—A novel species Lembosia mimusopis is introduced with evidence from

morpho-molecular characterization. It was collected from the leaves of Mimusops elengi in

Chiang Rai Province, Thailand. The new species is unique in having a mucilaginous sheath

surrounding its immature ascospores. LSU sequence analyses phylogenetically support

separation of this species from other Lembosia species.

Keyworps—Asterinaceae, Asterinales, Dothideomycetes, Lembosiaceae, Sapotaceae

Introduction

Léveillé (1845) established Lembosia with four species: Lembosia tenella

Lév. (type species), L. dendrochili Lév., L. drimydis Lév., and L. macula Lév.

636 ... Marasinghe & al.

Hansford (1946) included Lembosia in his new family Asterinaceae Hansf.

Based on morphology, Hosagoudar & al. (2001) and Hosagoudar (2012)

excluded Lembosia from Asterinaceae and treated the genus as the type of

Lembosiaceae Hosag However, Hongsanan & al. (2014) re-instated Lembosia

within the Asterinaceae based on molecular data derived from L. albersii

Henn. Dai & al. (2018) revealed that two species, L. albersii and L. xyliae

Zeng & al., are phylogenetically separate from Asterinaceae but retained

them in the same order, Asterinales. Three other species (L. syzygii Sivan. &

R.G. Shivas, L. abaxialis Firmino & R.W. Barreto, L. brigadeirensis Firmino

& al. clustered with Asterina species in a distinct clade and Dai & al. (2018)

treated them in Asterinales sensu lato. There are about 200 Lembosia species

epithets listed in Index Fungorum (2019). The type species L. tenella has not

yet been sequenced, and only five Lembosia species have been sequenced.

Lembosia species are considered as host-specific on plants in tropical

and sub-tropical regions and are usually identified based on association

with their host family or genus (Miller & von Arx 1962, Sivanesan & Shivas

2002). For instance, Sivanesan & Shivas (2002) introduced L. syzygii from

the leaves of Syzygium suborbiculare (Benth.) T.G. Hartley & L.M. Perry

and Araucaria heterophylla (Salisb.) Franco. The inherent morphological

characters of Lembosia are oval, elongated thyriothecia with an X- or

Y-shaped or longitudinal dehiscence, lateral appressoria on the hyphae,

and ovate to globose asci containing 4-8 brown uniseptate ascospores

(Hosagoudar & Goos 1991, Hosagoudar 2012). The asexual morph of this

genus is undetermined.

In this study, we introduce a novel species, Lembosia mimusopis, which

was collected from the leaves of Mimusops elengi in Thailand. Morphological

characters of the new taxon and its taxonomic placement supported by

phylogenetic analysis are provided.

Materials & methods

Sample collection, morphological studies, specimen deposition

Fresh leaf specimens were collected from Mae Fah Luang Botanical Gardens,

Chiang Rai, Thailand, in December 2018. Specimens were examined using a Motic

SMZ 168 Series microscope. Hand sections of the fruiting structures were mounted

in water and 5% KOH for microscopic studies and photomicrography. Blue ink

was used to stain the mucilaginous sheath around the ascospores. The micro-

morphology was examined using a Nikon ECLIPSE 80i compound microscope

and photographed using a Canon 750D digital camera fitted to the microscope.

Lembosia mimusopis sp. nov. (Thailand) ... 637

Measurements were made with the Tarosoft (R) Image Frame Work program and

figures were processed with Adobe Photoshop CS3 Extended version 10.0 software.

Specimens were deposited in the Mae Fah Luang University Herbarium, Chiang

Rai, Thailand (MFLU). The new taxon was linked with Facesoffungi and Index

Fungorum databases as explained in Jayasiri & al. (2015) and Index Fungorum

(2019).

TABLE 1. Taxa used in the phylogenetic analyses and their GenBank accession numbers

and voucher numbers

SPECIES

Asterina cestricola

Asterina chrysophylli

Asterina cynometrae

Asterina fuchsiae

Asterina magnoliae

Asterina melastomatis

Asterina phenacis

Asterina siphocampyli

Asterina weinmanniae

Asterina zanthoxyli

Asterotexis cucurbitacearum

Batistinula gallesiae

Buelliella physciicola

Buelliella poetschii

Hemigrapha atlantica

Inocyclus angularis

Karschia cezannei

Karschia talcophila

Labrocarpon canariense

Lembosia abaxialis

Lembosia albersii

Lembosia brigadeirensis

Lembosia mimusopis

Lembosia syzygii

Lembosia xyliae

Melaspileella proximella

Melaspileopsis diplasiospora

Parmularia styracis

Prillieuxina baccharidincola

Stictographa lentiginosa

VOUCHER

TH 591

VIC 42823

MFLU 13-0373 [T]

TH 590

MELU 16-0072

VIC 42822 [T]

TH 589

PPMP 1324

TH 592

TH 561

PMA M-0141224 [T]

VIC 42514

Ertz 18113

Ertz 18115

Ertz 14014

VIC 39748

Ertz 19186 [T]

Ertz 16749

Ertz 16907

VIC 42825 [T]

MELU 13-0377

VIC 44208

MELU 19-1225 [T]

MFLU 13-0633

MEFLU 14-0004 [T]

G.M. 2015-04-29

Ertz 16624

VIC 42447 [T]

VIC 42817

Ertz 17570

GENBANK (LSU)

GU586215

KP143738

NG057120

GU586216

MG844186

NG057055

GU586217

HQ701140

GU586218

GU586219

HQ610510

KP 143736

KP456147

KP456149

KP456151

KP143732

NG060323

KP456155

KP456158

NG060317

KM386982

MF664531

MN563123

MG844185

NG059589

KY654747

KP456165

NG060137

KP143735

KP456170

[T] = Ex-types and authentic strains. Newly generated sequences are indicated in bold.

638 ... Marasinghe & al.

DNA extraction, PCR amplification, sequencing

Genomic DNA was extracted directly from fresh thyriothecia using a Forensic

Genomic DNA Extraction Kit.

The total volume of PCR mixture (25 uL) contained double distilled water

(ddH,O) (11 uL), 2x QinKe PCR Master Mix (11 uL), DNA template (1 uL) and

10 um of each primer (1 pL). The complete 28S large subunit rDNA (LSU) gene was

amplified using LROR/LRS5 primers (Vilgalys & Hester 1990). PCR amplification

conditions consisted of an initial denaturation step of 5 min at 94 °C and final

extension step of 10 minutes at 72 °C. For the LSU amplification, the 37 cycles

consisted of denaturation at 94 °C for 1 minute, annealing at 54 °C for 50 seconds

and elongation at 72 °C for 1 minute. PCR products were viewed on 1% agarose

electrophoresis gels, stained with ethidium bromide (Thambugala & al. 2015). The

amplified PCR products were sent to Qinke, a commercial sequencing provider in

Kunming, China.

Sequence alignment & phylogenetic analyses

Newly generated sequences were assembled using Seqman and subjected to

the standard BLAST search (https://blast.ncbi.nlm.nih.gov) to identify the closest

matches in GenBank. The accession numbers of taxa used in our analyses are shown

in TABLE 1. Sequences (LSU) were aligned using MAFFT v. 6.864b (Katoh & al.

2017) and manually adjusted in BioEdit v. 7.0 (Hall 2004).

Maximum likelihood analysis was performed using RAxML on XSEDE in

CIPRES (Miller & al. 2010, Silvestro & Michalak 2012). The optimal ML tree

was obtained with 1000 separate runs under the GTR+GAMMA substitution

model resulted from model tests. Bayesian inference (Larget & Simon 1999) was

performed using the MrBayes 3.2.2 on XSEDE tool in CIPRES (Ronquist & al.

2012). Posterior probabilities (PP) were obtained from Markov chain Monte Carlo

sampling (MCMC) (Rannala & Yang 1996, Ronquist & al. 2012) when the average

standard deviation of split frequencies fell below 0.01. MCMC chains were run from

random trees for 1,000,000 generations and sampled every 100 generations with the

burn-in value of 25%. The remaining trees were used to calculate PP values. All trees

were visualized in FigTree v1.4.0 (Rambaut 2012) and the final layout was done with

Microsoft PowerPoint. The final alignment and trees were registered in TreeBASE

(2019) under the submission ID: 25248

Phylogenetic conclusions

LSU sequence data of representative families comprised 39 strains,

including two outgroup strains of Parmularia styracis Lév. (V1C42447,

V1C42450). The alignment contained 32,916 characters (LSU: 1-844)

including alignment gaps. The best scoring RAxML tree was selected

to represent the relationships among taxa with a final likelihood value

of -5964.8722. The matrix had distinct alignment patterns with 12.01%

undetermined characters or gaps. Estimated base frequencies were:

Lembosia mimusopis sp. nov. (Thailand) ... 639

* /9.73 | Asterina cestricola TH 591

61/* Asterina cynometrae MFLU 13-0373

Asterina siphocampyli PPMP

Asterina phenacis TH 589

goog Asterina zanthoxyli TH 561 Asterinaceae sensu stricto

97/1.00 Ni Asterina weinmanniae TH 592

Asterina fuchsiae TH 590

Asterina magnoliae MFLU 16-0072

66/0.96

100/1.00 | Asterotexis cucurbitacearum VIC 42814

Asterotexaceae

Asterotexis cucurbitacearum PMA M-0141224

93/100 100/1.00

100/1.00 Lembosiaceae

Lembosia albersii MFLU 13-0377

100/1.00

Lembosia xyliae MFLU 14-0004

82/0.97 Inocyclus angularis VIC 39747 ererinales

A DOTEEOD Inocyclus angularis VIC 39749 incertae cedis

Inocyclus angularis VIC 39748

100/1.00 m= . a

asterinales incertae

cedis

Hemigraphaceae

Melaspileella proximella G.M. 2014-05-25.1 Melaspileellaceae

Stictographa lentiginosa Ertz 17570

Stictographa lentiginosa Ertz 17447

Buelliella physciicola Ertz 18113

Karschia cezannei BR Ertz 19186 ‘

Stictographaceae

Karschia talcophila Ertz 16749

Labrocarpon canariense Ertz 16308

10 Labrocarpon canariense Ertz 16907

Melaspileopsis cf diplasiospora Ertz 16247 BR

IDET D Melaspileopsis cf diplasiospora Ertz 16624 BR

100/1.00__[ Asterina melastomatis VIC 42822

Lembosia abaxialis VIC 42825

85/1.00 Asterina chrysophylli VIC 42823 Asterinaceae sensu lato

anes Prillieuxina baccharidincola VIC 42817

Batistinula gallesiae VIC 42514

sata Lembosia syzygii MFLU 13-0633

Parmularia styracis VIC 42450

Parmularia styracis VIC 42447 Parmulariales (out group)

0.06

FiGuRE 1. Phylogram of Lembosia and related genera, generated from maximum likelihood

(RAxML) based on LSU sequence data. ML bootstrap support (260%) and Bayesian posterior

probability (= 0.80) are indicated above the branches as ML/PP. The tree is rooted in Parmularia

styracis (VIC 42447, VIC 42450). Ex-type strains are in bold and the newly generated sequences

are in red.

640 ... Marasinghe & al.

A = 0.234839, C = 0.253073, G = 0.326282, T = 0.185806; substitution rates

AC = 0.902460, AG = 2.658802, AT = 0.629941, CG = 1.176560, CT = 8.729825,

GT = 1.000000; gamma distribution shape parameter a = 0.3196 (Fie. 1).

Lembosia albersii, L. xyliae, and our new taxon clustered with high

statistical support (100% ML, 1 BYPP), forming a sister clade to three strains

of Inocyclus angularis Guatim. & R.W. Barreto (VIC 39747, VIC 39748, VIC

39749). The new taxon is clearly distinct from the two other species in the

clade with high statistical support. However, L. abaxialis, L. brigadeirensis,

and L. syzygii clustered with species in Asterina sensu lato.

Taxonomy

Lembosia mimusopis Marasinghe, Daya., Maharachch., Hongsanan &

K.D. Hyde, sp. nov. FIG 2

IF 556875

Differs from Lembosia bomjardinensis in having ascospores that are sheathed when

immature.

Type: Thailand, Chiang Rai, MFU Botanical Gardens, on living leaves of Mimusops

elengi L. (Sapotaceae), 19 December 2018, M.W.D. Sandamali BSM-2019a (Holotype,

MFLU_ 19-1225; GenBank MN563123. Isotype, MFLU 19-0724; GenBank

MN565982).

EryMo.oecy: referring to the host genus Mimusops from which the fungus was

collected.

CoLonies hypophyllous on Mimusops elengi leaves forming blackened

circular to irregular areas, single to confluent. HyPpHAE 3.2-5 um diam.

(x = 4.5 um, n = 10), superficial, brown, straight to undulate, branching

opposite to irregular at acute to wide angles, reticulate. APPRESSORIA

4—5.5 um diam. (x = 4.5 um, n = 10), unicellular, brown, irregular, opposite

to alternate.

SEXUAL MORPH: THYRIOTHECIA 350-500 x 350-450 um (x = 451 x

393 um, n = 6), scattered, sub-dense, initially rounded, elongated at maturity,

longitudinally dehisced at the centre, margin crenate to fimbriate, borne on

the surface of mycelium. PSEUDOPARAPHYSES not observed. Asci 55-65 x

45-55 um (x = 60 x 48 um, n = 10), 8-spored, bitunicate, sub-globose to

globose, apical region of asci usually thick, opaque, ocular chamber not

observed, not staining blue in IKI. Ascospores 25-30 x 8-11 um (x = 28 x

9 um, n = 10), overlapping 4-5-seriate, oblong to obovoid, with broad to

narrowly rounded ends, lower cell slightly longer and narrower, hyaline,

brown when mature, covered with mucilaginous sheath when immature,

1-septate, constricted at the septum, verruculose.

Lembosia mimusopis sp. nov. (Thailand) ... 641

FIGURE 2. Lembosia mimusopis (holotype, MFLU 19-1225): a. Leaf sample; b, c. Thyriothecia

on host substrate; d. Squash mount of thyriothecium showing longitudinal fissure (red arrow);

e. Upper wall of thyriothecium; f. Hyphae with appressoria; g-j. Immature asci; k. Mature

ascus; |. Ascospores with mucilaginous sheath stained with blue ink; m-s. Immature to mature

ascospores. Scale bars: d—f = 100 um; g—k = 20 um; |-s = 10 um.

ASEXUAL MORPH: Undetermined.

642 ... Marasinghe & al.

COMMENTS: Lembosia mimusopis (FoF06234) is morphologically similar

to L. bomjardinensis (Bat.) Hongsanan & K.D. Hyde [= Yamamotoa

bomjardinensis Bat.] in its globose to subglobose, apedicellate asci with

a thick hyaline apical region and oblong to obovoid, 1-septate, hyaline

to brown ascospores with a verruculose wall. However, L. mimusopis is

distinguished by having sheathed ascospores when immature; and in its

sapotaceous host in contrast to the proteaceous host of L. bomjardinensis.

Attempts to isolate L. mimusopis onto potato dextrose agar and malt extract

agar were unsuccessful so that the DNA was extracted directly from fruiting

bodies.

Discussion

Lembosia mimusopis represents the first record of a Lembosia species from

Mimusops (Sapotaceae). However, L. sapotacearum Mibey was recorded from

Sapotaceae in East Africa (Mibey & Hawksworth 1997). Lembosia species

are also recorded from other host families (Araucariaceae, Asparagaceae,

Dipterocarpaceae, Ebenaceae, Lauraceae, Myrtaceae, Proteaceae, Orchidaceae;

Index Fungorum 2019). A comprehensive comparison of morphology within

this genus is complicated by the lack of good illustrations and descriptions of

previously recorded species. Host-specificity of Lembosia species has not been

phylogenetically studied (Hongsanan & al. 2014). Hongsanan & al. (2014)

synonymized Trichamelia Bat., Viegasia Bat., and Yamamotoa Bat. under

Lembosia. Dai & al. (2018) pointed out the necessity of recollecting the type

species, L. tenella, to obtain molecular data to establish its placement within

Asterinales. In agreement with the results reported by Dai & al. (2018), our

study also shows that four Lembosia species form a sister clade with Inocyclus

in Asterinales.

Acknowledgments

The authors extend their appreciation to the Researchers supporting project

number (RSP-2019/56) King Saud University, Riyadh, Saudi Arabia. We would

like to thank the National Natural Science Foundation of China for supporting

the project Biodiversity, Taxonomy, Phylogeny, Evolution and Phytogeography

of Phytopathogens in Dothideomycetes from Southern China (Project number:

31950410548) for funding this research. The Thailand Research fund is thanked

for the grant “Impact of climate change on fungal diversity and biogeography in

the Greater Mekong Subregion grant number: RDG6130001” and Thailand Science

Research and Innovation (TSRI) grant “Macrofungi diversity research from the

Lancang-Mekong Watershed and Surrounding areas (grant no. DBG6280009). Prof.

K.D. Hyde thanks Chiang Mai University for the award of Visiting Professorship.

Lembosia mimusopis sp. nov. (Thailand) ... 643

The authors thank Prof. D. Jayarama Bhat (Curca, Goa Velha-403108, India) and Dr.

Eric H.C. McKenzie (Manaaki Whenua Landcare Research, Auckland, New Zealand)

for presubmission review. Prof. Mibey and Dr. Shaun Pennycook are thanked for

valuable suggestions and for nomenclatural advice.

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MY COTAXON

July-September 2021— Volume 136, pp. 645-660

https://doi.org/10.5248/136.645

Fibroporia gossypium isolated from an indoor environment

in Argentina

CAROLINA A. ROBLES??’, MARIA BELEN PILDAIN??*+, FRANCISCO SAUTUA5,

MARCELO A. CARMONA®, MARIO RAJCHENBERG”?°

' Departamento de Biodiversidad y Biologia Experimental, Universidad de Buenos Aires,

Intendente Guiraldes 2160, C.A.B.A, CP 1428, Argentina.

? Consejo Nacional de Investigaciones Cientificas y Técnicas, Argentina (CONICET)

* Centro de Investigacion y Extension Forestal Andino Patagonico (CIEFAP),

C.C. 14, Ruta 259 km 16.24, Esquel, Chubut, 9200, Argentina

‘ Facultad de Ciencias Naturales, Universidad Nacional de la Patagonia S.J. Bosco,

Sede Esquel, Ruta 259 km 16.24, Esquel, Chubut, 9200, Argentina.

° Catedra de Fitopatologia, Facultad de Agronomia, Universidad de Buenos Aires,

Av. San Martin 4453, C.A.B.A, C1417DSE, Argentina

° Facultad de Ingenieria, Universidad Nacional de la Patagonia S.J. Bosco,

Sede Esquel, Ruta 259 km 16.41, Esquel, Chubut, 9200, Argentina.

* CORRESPONDENCE TO: caroanarobles@gmail.com, carorobles@bg.fcen.uba.ar

ABSTRACT—Using culture studies and molecular techniques, the brown rot polypore

Fibroporia gossypium was identified from a cellar in an old building of Buenos Aires city, the

second record of the species in an urban environment in Argentina. The fungus formed large

mycelial masses hanging from a wooden support and growing on the walls but did not form

basidiomes. The conspecificity of specimens from distant regions is considered and the origin

of the present finding is discussed.

Key worps—multigene analysis, South America, taxonomy, wood-rotting fungus

Introduction

Fibroporia gossypium |= Antrodia gossypium| is a widely distributed brown

rot wood-decay basidiomycete occurring in coniferous stands in Europe

(Spirin 2007, Bernicchia & al. 2012, Ryvarden & Melo 2014, Zétciak & al.

2016), North America (Lombard 1990), and Asia (Yuan & Dai 2008). It is

646 ... Robles & al.

also found on Nothofagus spp. in Patagonia, Argentina (Rajchenberg 2006,

Rajchenberg & al. 2011).

The species was originally described as Poria gossypium (holotype LPS

25495) from a specimen isolated from vine plant borders in La Plata, Buenos

Aires Province, 122 years ago (Spegazzini 1898, Rajchenberg 1984, Rajchenberg

& Wright 1987). After that, the species was not recorded in Argentina until

studies in the southern region of the country (Rajchenberg 2006).

During summer of 2018, an extensive mycelium of a wood-decay

basidiomycete was found in a basement located in the plant pathology

laboratory of the School of Agriculture of the University of Buenos Aires.

There was mycelium hanging from a door frame (Fic. 1A) and typical mycelial

cords (rhizomorphs) were observed on the walls (Fic. 1B). The fungus was

also covering the furniture (Fic. 1C) and had decayed a wooden door, made

from an unidentified gymnosperm, extremely decayed, and found fallen to

the floor (Fic. 1D). The basement had been formerly used as a cheese ripening

chamber with an environment of high relative humidity, cool temperature,

and darkness. More recently, the basement had been used as a depository for

books and materials used in the dairy industry and had been abandoned for

at least 15 years until the mycelium was found covering the walls, wooden

shelves, door, and door-frame.

The aim of this work was to identify and to discuss the presence of

FE. gossypium growing in an indoor environment in Buenos Aires city,

Argentina. This is the second urban record of the species, originally described

by Spegazzini (1898) from La Plata city (ca. 60 km from Buenos Aires).

Materials & methods

Isolation and cultural studies

The fungus was isolated directly from the mycelium hanging from the door frame

and cultured in 2% MEA, 2% PDA, yeast media and 2% water agar medium and

incubated at both dark and natural light at 24 °C. The micro and morphological

characteristics were observed regularly under the microscope.

The strain, named ARG_2018_Agro, was cultured on 1.25% MEA (Difco Lab.)

in the dark at 25 °C (Nobles 1965), and macro- and micromorphological characters

were recorded weekly for 6 weeks. The species was identified by means of keys based

on mycelial characters (Nobles 1965, Stalpers 1978) and codified according to Nobles

(1965). Oxidase reactions were performed using gallic (GAA) and tannic (TAA) acid

agar media (Davidson & al. 1938) and tyrosine, p-cresol and guaiacol agar media

(0.2%) according to Boidin (1954). The relative intensity of the reaction, recorded one

week after incubation in the dark at 25 °C, was indicated with plus or minus signs.

Fibroporia gossypium indoors (Argentina) ... 647

Fic. 1. Fibroporia gossypium (ARG_2018 Agro), sampling site, with mycelium covering the

walls, furniture, and wooden door frame. A. Conspicuous mycelial flaps hanging from the

wooden frame; B, C. Mycelium and rhizomorphs covering furniture and walls; D. Wooden door

rotted by the fungus.

Strain ARG_2018_ Agro is conserved in the Department of Plant Pathology, School of

Agriculture, University of Buenos Aires, Buenos Aires City (DPP FAUBA).

Molecular studies

The genomic DNA was extracted from 7-day-old fungal cultures grown in 2%

MEA culture plates using the PuriPrep T Kit, according to the manufacturer's

648 ... Robles & al.

instructions. Five genomic regions were included in the analyses, following Chen

& al. (2017). DNA was amplified in 50 ul volumes containing 10 mM PCR buffer

supplied by the manufacturer, 50 mM MgCl, 1 mM of each dNTP, 10 mM of each

primer, 1 U Recombinant Taq DNA polymerase, and 5 ul fungal genomic DNA.

The internal transcribed spacer (ITS) region of the isolates was amplified using the

universal primers ITS1 and ITS4 under the following PCR conditions: denaturation

at 94 °C for 3 min, 50 cycles of denaturation at 94 °C for 45 s, annealing at 57 °C for

45 s, extension at 72 °C for 1 min and a final extension at 72 °C for 7 min (Schoch

& al. 2012). The nLSU regions were amplified with primer pairs LROR and LR7

(https://sites.duke.edu/vilgalyslab/rdna_primers_for_fungi/): the nSSU regions with

the primer pair NS1 and NS4 (White & al. 1990); the mtSSU regions with the primer

pair MS1 and MS2 (White & al. 1990); the tefl with the primer pair EF1-983F and

EF1-1567R (Rehner & Buckley 2005); and rpb2 with the primer pair bRPB2- 6F and

bRPB2-7R (Matheny 2005). The PCR conditions for nSSU, mtSSU, tefl, and rpb2

are described in Chen & al. (2017). The PCR products were purified and sequenced

at sequencing service of Facultad de Ciencias Exactas y Naturales, Universidad de

Buenos Aires (Argentina) and at Macrogen Sequencing Service (Macrogen, Korea),

with the same primers.

Phylogenetic analysis

BLAST queries were performed in GenBank. As a framework for taxon

selection, we used sequences of Fibroporia and its related genera in the antrodia

clade obtained by Chen & al. (2017) (TaBLE 1). Two data sets were analyzed for

this study: concatenated analyses of the ITS, nLSU, nSSU, mtSSU, tefl, and rpb2

gene regions and the ITS region to ascertain the phylogenetic structure within the

F. gossypium-pseudorennyi clade. Nucleotide sequences for the ITS, nLSU, nSSU,

mtSSU, tefl, and rpb2 gene regions were initially edited using BIOEDIT 7.0.9.0

(Hall 1999) then aligned using L-INS-i strategy as implemented in MAFFT v 7.0

(Katoh & Standley 2013) and manually adjusted using MEGA version X (Kumar &

al. 2018). The final ITS data set comprised 45 sequences and 668 characters including

gaps, the nLSU data set comprised 45 sequences and 1319 characters including gaps,

while the mtSSU, tefl, and rpb2 data sets comprised 40 each and 1015 (mtSSU), 487

(tefl), and 651 (rpb2) characters including gaps. The datasets were combined for

concatenated analyses using Mesquite 3.40 (Maddison & Maddison 2018). ITS data

set for the extended Fibroporia species analysis comprised 29 sequences and 623

characters including gaps. The best nucleotide substitution model was estimated by

using the Akaike Information Criterion (AIC) (Akaike 1974) using JMODELTEST

2 2.1.1 (Guindon & Gascuel 2003, Darriba & al. 2012) and were GTR+I+G for ITS,

nLSU, nSSU, and rpb2; GTR+G for mtSSU; and TIMef+I+G for tefl. Maximum

likelihood (ML) phylogenetic trees for the combined data was estimated under

these models in RAxML 7.2.8 (Stamatakis 2014), using the default parameters,

executed on the CIPRES (Cyberinfrastructure for Phylogenetic Research) Science

Gateway V.3.1 (http://www.phylo.org/sub_sections/portal/, Miller & al. 2010) with

bootstrap support values (MLB) calculated with 1000 repetitions, and Bayesian

649

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inferences (BI) of phylogenies in MrBayes v.2.2 (Ronquist & al. 2012). Four

independent chains were run for 8 million generations; trees were sampled every

1000 generations. Log files for each run were viewed in Tracer v1.6.0 (http://evolve.

z00.0x.ac.uk/software.html/tracer/) to determine convergence. Trees generated

prior to stationarity were discarded, and the remainder of the trees was summarized

in a majority rule consensus tree from the four independent runs. Branch support

was assessed using posterior probabilities (BPP) calculated from the posterior set

of trees after stationarity was reached. Sequences of Trametes suaveolens (L.) Fr.

(Cui 11586) and Trametes polyzona (Pers.) Justo (Cui 1104042) were used as

outgroup in the concatenated analyses of the ITS, nLSU, nSSU, mtSSU, tefl and rpb2

gene regions and sequences of Pseudofibroporia citronella Yuan Y. Chen & B.K. Cui

(He 20120721-15 and Yuan 6181) were used as outgroup in the ITS region analyses.

The final alignments were deposited in TreeBASE (http://www.treebase.org), under

accession [D25872.

Taxonomy

Fibroporia gossypium (Speg.) Parmasto,

Consp. System. Corticiac.: 207. 1968. Figs 1, 2

= Poria gossypium Speg., Anales Mus. Nac. Buenos Aires 6: 169. 1898.

= Antrodia gossypium (Speg.) Ryvarden, Norweg. J. Bot. 20: 8. 1973.

Nobles culture code: 1.3.7.34.35.36.38.43.51.55.

GROWTH CHARACTERISTICS: Mats white, cottony, loose at first, appressed

during the second week, powdery in the following weeks; margins distinct,

even, or finely fimbriate; odorless; reverse unchanged; the colony reaching a

radius of 17 mm after 7 da on 1.25% MEA, covering the plate in three weeks;

oxidase reactions negative, GAA at 7 da 25-27 mm in diam, no growth on

TAA.

HyYPHAL CHARACTERISTICS: all hyphae staining in phloxine, thin-walled,

mostly clamped with single clamps, rarely simple-septate, (2.5-)3.5-5(-6)

um diam. Crystals present from the first week, 3-4 x 2-3 um. From the

second week terminal chlamydospores 5-7(-11) x 4-6 um and arthrospores

(6-)9-15(-28) x 2-4 um, very abundant at the sixth week.

SPECIMEN EXAMINED: ARGENTINA, BUENOs AIRES CiTy, University of Buenos

Aires, School of Agriculture, 34°35’30”S 58°28’46”W, basement wood-decay,

December 2018, F. J. Sautua (DPP FAUBA ARG_2018_Agro).

CoMMENTs: The description does not completely agree with Lombard (1990;

as Antrodia “gossypia”) and the southern Argentina specimens studied by

Rajchenberg (2006), because those strains showed few clamped hyphae,

there was no mention of the presence of chlamydospores or arthrospores,

Fibroporia gossypium indoors (Argentina) ... 653

Fig. 2. Fibroporia gossypium (ARG_2018 Agro), cultural characteristics. A. Six-week-old

culture; B. GAA medium; C. TAA medium; D. Clamp connections; E. Crystals; F. Arthrospores;

G. Chlamydospores. Scale bars = 10 um.

and growth was very slow, covering the plates after only after 7 weeks of

incubation. However, Spirin (2007) noted that one specimen of FE gossypium

(LE 210088) had thick-walled chlamydospores in the subiculum.

After a four-month incubation in 2% MEA, 2% PDA, yeast media, and

2% water agar medium, whether cultured in the dark or in natural light,

there was no production of basidiomes.

Phylogeny

Phylogenetic analyses based on the ITS and the combined concatenated

data set ITS + nLSU + nSSU + mtSSU + tefl + rpb2 for the antrodia clade

(Fics. 3, 4) clustered strain ARG_2018_Agro and two Argentinian specimens

[CIEFAP 131 from Chubut and CIEFAP M. Rajchenberg 11443 from Tierra

del Fuego (Ushuaia, Tierra Mayor, in pure Nothofagus pumilio forests, on

fallen trunk, 22 March 1998; duplicate as LY-BR 3994, cf. Chen & al. 2017]

within the monophyletic group F. gossypium that includes E gossypium and

F. pseudorennyi (BPP = 1.0, MLB = 0.99, Fic. 3; BPP = 1.0, MLB = 0.95, Fie. 4).

654 ... Robles & al.

Discussion

Molecular analysis led to the identification of the ARG_2018 Agro

strain found in the humid basement as the decay basidiomycete Fibroporia

gossypium. The cultural characters showed, on the other hand, differences

from previous descriptions of this species, in particular an absence of

chlamydospores or arthrospores, fewer clamped hyphae, and slower growth

(Lombard 1990, Rajchenberg 2006). Cultural features have facilitated

separation and identification of different basidiomycete groups (Nobles 1965,

Stalpers 1978); however, conflicting characters have also been noticed in such

cultural studies.

Strain ARG_2018_Agro produced a brown rot decay, as indicated by the

substrate from which it was isolated and the tannic acid agar media results.

Fibroporia gossypium has already been associated with gymnosperms in the

Northern Hemisphere and on Nothofagus sp. in the Patagonian Andes forests

of Argentina (Rajchenberg 2006).

Spegazzini described the original material of Poria gossypium from border

plants of (cultivated) vines near La Plata city: “Hab. In ollis vinariis ad plantas

colendas adhibitis, in La Plata, aest. 1890 (leg. Cl. Fl. Ameghino)” (Spegazzini

1898), although he wrote on holotype envelope: “In limis plantarum, FI.

Ameghino, Aest. 1890” (Rajchenberg & Wright 1987). This exotic environment

and host present the unusual situation of a taxon that was first described from

a strange habit while it was naturally growing in other situations. While the

species was widely recorded in the Northern Hemisphere (e.g., Lowe 1966,

Nunez & Ryvarden 2001, Dai & Niemela 2002, Ryvarden & Melo 2014), it was

only recorded from the Southern Hemisphere from Patagonia, Argentina,

many years later, there growing exclusively on fallen trunks and branches

of different Nothofagus spp. (Rajchenberg 2006), contrasting with other

geographical areas where it is mainly recorded on coniferous wood. The taxon

is unknown from other austral continents.

How did this taxon arrive and profusely develop its mycelium in a cellar-

like, indoor environment? We have no answer to that. An exotic origin is a

possible explanation. Certainly, its dissemination and arrival seem to be an

anomaly, as the species is not present as such in Buenos Aires city and is not

associated with the introduced tree species planted in the city’s groves (Robles

& al. 2011, 2015). Its establishment and profuse growth has most possibly

been encouraged by the still environment and optimum temperatures present

in the abandoned room where it developed.

Fibroporia gossypium indoors (Argentina) ...655

Fibroporia gossypium 3994

Fibroporia gossypium Cui 9472

1.0/ 100 Fibroporia gossypium LY BR 3914

Fibroporia pseudorennyi X1377

Fibroporia pseudorennyi X1384

0/ 100

ARG_2018_Agro

1.0/ 96 1.0/ 100] Fibroporia norrlandica 4115

Fibroporia norrlandica 4122

Fibroporia ceracea Dai 13013

Fibroporia radiculosa Cui 11404

1.0/ oof Fibroporia radiculosa Cui 2796

) Fibroporia radiculosa Dai 6473

Fibroporia radiculosa Dai 13041

Fibroporia bambusae Dai 16210

1.0/ 100]Fibroporia bambusae Dai 16211

Fibroporia bambusae Dai 16212

1.0/ 75 Fibroporia bohemica BJFC 12750

4.0/1 190] Fibroporia vaillantii FP 90877 R

Fibroporia vaillantii RWD 63 263

Fibroporia albicans Cui 9464

Fibroporia albicans Cui 9495

Fibroporia albicans Cui 9504

1.07 a] Fibroporia albicans Dai 10595

Fibroporia citrina Cui 11604

Fibroporia citrina Cui 9683

of 1007 PSeudofibroporia citrinella Yuan 6181

Pseudofibroporia citrinella He 20120721-15

4.01 100 7 La@etiporus sulphureus Dai 12154

_/84 Laetiporus sulphureus Dai 12826

91 100 | -iptoporus soloniensis Cui 11390

Piptoporus soloniensis Dai 11872

Amyloporia xantha Cui 11544

4.0! 100 Antrodia heteromorpha Dai 12755

0/ 100 Antrodia serpens Dai 7465

Antrodia tanakae Cui 9743

-0/ 100

1.0/ 100 1.0/ 100)

1.0/-

1.0/ -

O/ 100 4.0/ 100 ¢ Antrodia serialis Cui 10519

1.0/ 100 Antrodia serialis Cui 9706

of 100 [ Daedalea dickinsii Yuan 1090

10 100 Daedalea quercina Dai 12659

Piptoporus betulinus Cui 10756

1.0/ 100 Fomitopsis pinicola Cui 10312

4.0/ 100 Postia duplicata Cui 10366

Oligoporus lacteus Cui 12206

Trametes polyzona Cui 11040

Trametes suaveolens Cui 11586

0.1

Fic. 3. ML tree based on the combined concatenated data of ITS + nLSU + nSSU + mtSSU + tefl +

rpb2 including 45 sequences. BPP >0.95 and MLB >50% are marked above the nodes.

656 ... Robles & al.

Fibroporia gossypium CIEFAP131 Argentina

Fibroporia gossypium 3994 Argentina

ARG_2018_Agro Argentina

1.0/ 95 Fibroporia gossypium Cui 9472 China

Fibroporia gossypium FG2011 Poland

Fibroporia gossypium HUBO7724 ttaly

Fibroporia gossypium LY BR3914 France

186 Fibroporia pseudorennyi X1384 Russia

Fibroporia gossypium X1403 Finland

Fibroporia pseudorennyi X1377 France

Fibroporia norriandice 4115 Finland

bet Fibroporia norriandice 4122 Finland

Fibroporia norriandica 4151 Finland

T.0/

100 | Fibroporia norriandica UC2022910 USA

10/98 Fibroporia albicans Cui 9464 China

1.0/ 100 4.0199 Fibroporia albicansDai10595 China

Fibroporia citrine Cui11604 China

1.0/ 99 | 0.994 Fibroporia citrina HUBO7715 ttaly

e Fibroporia vailiantiiFP 90877 USA

Fibroporia vaillantiiRWD 63 263 USA

Fibroporia bambusae Dai 16210 China

1.0/ 100

1.0/ Fibroporia bambusae Dai 16212 China

100) Fibroporia radiculosa Cui2796 China

iN Fibroporia radiculosa Dai 13041 China

+84 Fibroporia bohemica BJFC12750 Czech Republic

Fibroporia ceracea CLZhao 2308 China

1.0/ 100

Fibroporia ceracea Dai 13013 China

Pseudofibroporia citinella He 20120721-15 China

Pseudofibroporia citinela Yuan 6181 China

Fic. 4. The 50% majority-rule consensus tree from Bayesian inference of the ITS region including

29 sequences. BPP >0.95 and MLB >75% are marked above the nodes.

Fibroporia gossypium indoors (Argentina) ... 657

Leptoporus destructor var. resupinatus Bourdot & Galzin (1928: 547) was

accepted as a form of Fibroporia gossypium by Lowe (1966, as Poria gossypium),

Parmasto (1968, as E gossypium), Domanski (1972, as FE gossypium) and

Ryvarden (1976, as Antrodia gossypium), growing on mine poles where

environmental conditions are constant. The type species of Fibroporia,

FE. vaillantii (DC.) Parmasto, another “mine” fungus, is also associated with

mine timber and affected wood in deteriorated buildings (Wainwright 1999,

Cojocariu & al. 2007).

Spirin described Fibroporia pseudorennyi (Spirin) Spirin [= Oligoporus

pseudorennyi Spirin] as phylogenetically close to FE gossypium but with smaller

spores and lacking cystidia (Spirin 2004, 2007). Fibroporia pseudorennyi also

produced chlamydospores ((6-)8-10 x 6-7.5 um) in the basidiome margin;

currently there are no cultural descriptions of F pseudorennyi. Given the

phylogenetic results of the present study and those of Chen & al. (2017) based

on five genomic regions, the synonymy of F. gossypium and E pseudorennyi is

maintained (cf. Ryvarden & Melo 2014).

Acknowledgments

The research was supported by the National Council for Scientific and

Technological Research (CONICET, Argentina) PIP 0956 and by University of

Buenos Aires (UBACYT 20020150100067BA and UBACYT 20020170100147BA).

We acknowledge Dr. O. Troncoso (Facultad de Ingenieria, U.N. de la Patagonia S.J.

Bosco, Argentina) for his inputs on the nature of the wooden door decayed by the

strain studied. We thank Harold H. Burdsall (Fungal & Decay Diagnostics, Black

Earth WI, USA) and Sergio P. Gorjén (Department of Botany and Plant Physiology

University of Salamanca, Spain) for their expert reviews.

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MY COTAXON

July-September 2021— Volume 136, pp. 661-667

https://doi.org/10.5248/136.661

Rubellofomes cystidiatus and Spongiporus floriformis

newly recorded for India

VINJUSHA NELLIPUNATH & T.K. ARUN KUMAR™

The Zamorins Guruvayurappan College, Kerala 673014, India

“CORRESPONDENCE TO: tkakumar@gmail.com

ABSTRACT-In an effort to document the macrofungi of Kerala state, India, we collected and

identified two brown rot polypores, Rubellofomes cystidiatus and Spongiporus floriformis,

which represent new records for India. Here we present taxonomic accounts of the two

species.

Key worps-biodiversity, brown rot, Dacryobolaceae, Fomitopsidaceae, Polyporales

Introduction

Rubellofomes B.K. Cui & al. (Fomitopsidaceae, Polyporales) was erected to

accommodate two brown rot polypores, Fomitopsis cystidiata B.K. Cui & M.L.

Han and F. minutispora Rajchenb. According to Han & al. (2016), these two

species were distinct from other Fomitopsis species by their purple pink to

pinkish brown context. Rubellofomes can be distinguished from the closely

related Niveoporofomes B.K. Cui & al. by its orange brown to dark brown pileal

surface, purple pink to light pinkish brown context, and cylindrical to ellipsoid

basidiospores. Niveoporofomes species produce ivory white to pale ochraceous

pileus surfaces, white to ochraceous context, and ovoid to broadly ellipsoid

basidiospores (Han & al. 2016). Ungulidaedalea B.K. Cui & al., another closely

related genus, is separated by its ungulate, fragile fruit bodies and septate

skeletal hyphae (Han & al. 2016). Rubellofomes has been reported previously

from Argentina and China.

Spongiporus Murrill (Dacryobolaceae, Polyporales) was established by

Murrill (1905), with S. leucospongia (Cooke & Harkn.) Murrill as the type

662 ... Nellipunath & Kumar

species. The genus accommodates brown rot species with a monomitic hyphal

system (David 1980). Spongiporus representatives are characterized by an

annual growth habit, effused reflexed to pileate basidiomata, a monomitic

hyphal system with clamped generative hyphae, and hyaline, smooth, thin-

walled, acyanophilous, inamyloid, allantoid to cylindrical basidiospores (Shen

& al. 2019). Spongiporus is distributed in Asia, Europe, and North America

(Murrill 1907, Ryvarden & Gilbertson 1994, Spirin & al. 2006). Spongiporus

has been considered as a synonym of other two brown rot polypore genera,

Oligoporus Bref. and Postia Fr., by different workers (Gilbertson & Ryvarden

1985, Ryvarden & Gilbertson 1994, Pildain & Rajchenberg 2013, Ryvarden

& Melo 2014). According to Shen & al. (2019), Oligoporus basidiocarps are

resupinate while Spongiporus produces effused reflexed to pileate basidiocarps.

Postia species differ from Spongiporus by a universally solitary habit and

narrower basidiospores (Shen & al. 2019). Pildain & Rajchenberg (2013)

treat Oligoporus as a synonym of Postia. Postia species produce thin walled

basidiospores and generative hyphae that are metachromatic in cresyl blue.

Oligoporus representatives produce thick-walled cyanophilous spores and

their hyphae are non-metachromatic in cresyl blue (Erkkila & Niemela 1986,

Renvall 1992, Pildain & Rajchenberg 2013). Recent phylogenetic studies treat

Oligoporus, Postia, and Spongiporus as independent genera (Ortiz-Santana &

al. 2013, Binder & al. 2013, Cui & al. 2014, Shen & al. 2019). Seven species

have been recognized in Spongiporus (He & al. 2019; www.indexfungorum.

org).

Kerala state in India has a rich mycobiota, with a documented diversity

of 148 polyporoid species (Adarsh & al. 2018). As part of studies on the

polyporoid fungi of the region, two interesting species were encountered

that were later found to represent new records for India. Here we present the

taxonomic details of the two new polypore records.

Materials and methods

Fruitbodies were collected from forest areas of the Kerala region of Western

Ghats in India during the monsoon seasons. Macroscopic characters of fresh

specimens were recorded. Microscopic observations were made on materials

stained using aqueous solutions of 3% phloxine and 1% congo red and mounted

in 5% aqueous KOH. Reaction of the basidiospores on treatment with Melzer’s

reagent and cotton blue was noted. Twenty basidiospores from each specimen were

measured for obtaining the spore dimensions, range of spore quotient (Q, length/

width ratio), and its mean value (Q_). All collections examined are deposited at

the Zamorin’s Guruvayurappan College Herbarium, Kozhikode, Kerala, India

(ZGC).

Rubellofomes cystidiatus & Spongiporus floriformis (India) ... 663

Taxonomy

Rubellofomes cystidiatus (B.K. Cui & M.L. Han) B.K. Cui, M.L. Han &

Y.C. Dai, Fungal Diversity 80: 366. 2016 FIG. 1A-C

BASIDIOMATA annual, medium sized, pileate, hard, woody, light weight,

when dry. PrLEus 30-65 mm long, 14-20 mm thick, sessile, broadly attached,

difficult to detach from the host, applanate to slightly irregular with minute

ridges and grooves, almost semicircular, glabrous, azonate, purplish brown

to cinnamon brown (browner on drying), turning brownish black in KOH.

MARGIN even, round, sterile, orange yellow. HyMENOPHORE poroid, pale

purplish brown, more brownish on drying. Porzs large, 1-3 mm wide,

narrow towards margin, angular, pore tubes 10-15 mm long, concolorous

with the hymenophore. CONTEXT 3-5 mm thick, very pale pinkish brown,

homogeneous. ODovR not distinctive. SPORE PRINT not observed.

HyPHAL SYSTEM dimitic, generative hyphae septate, with clamp

connections, slightly thick to thick walled, skeletal hyphae unbranched to

slightly branched. PILEIPELLIs an agglutinated cutis, easily crushed in sections,

pileipellis hyphae 2-3 um wide, slightly thick walled, yellowish brown. PILEAL

TRAMA interwoven, generative hyphae 2-3 um wide, septate, hyaline, mostly

thick walled, branched, clamp connections present. HYMENIAL TRAMA

interwoven, generative hyphae 2-3 um wide, septate, hyaline, slightly thick to

thick walled, branched, clamp connections present and skeletal hyphae 2-5

um wide, hyaline, simple to rarely branched, aseptate, thick walled (1 um).

Skeleto-ligative hyphae not observed. Cystip1a 31-42 x 4-6 um, clavate to

cylindrical, hyaline, thick walled (1 um), encrusted. BAstp1a 26-32 x 3-4

um, clavate, 2-sterigmate, with basal clamp connections. BASIDIOSPORES 5-8

x 2-3 um, Q = 3-4, Q_ = 3.4, cylindrical, often slightly bent, smooth, thin

walled, eguttulate, inamyloid in Melzer’s reagent.

SPECIMEN EXAMINED: INDIA. KERALA: Palakkad, Silent Valley National Park,

11.0641°N 76.5378°E, on dead wood of unidentified tree, 14 January 2018, Vinjusha N.

(ZGC VN667).

KNOWN DISTRIBUTION: China

CoMMENTs: ‘The present collection possesses slightly longer spores when

compared with the original description of the material of R. cystidiatus

((4.8-)4.9-6.2(-6.5) x 2.0-2.8(-2.9) um, Han & al. 2014). The pileus surface

of R. cystidiatus was described as concentrically zonate (Han & al. 2014), but

proper zonations or concentric rings were not observed in our collection.

According to Han & al. (2014), the pore surface of R. cystidiatus is pale

purple pink in fresh condition and turns clay buff to cinnamon brown on

664 ... Nellipunath & Kumar

drying. However, a pinkish tint was absent in our collection and its more

purplish to brownish shades may be because the specimen was collected in a

more or less dried form.

Spongiporus floriformis (Quél.) Zmitr.,

Folia Cryptogam. Petropolitana 6: 90. 2018. FIG. 1D, E

BASIDIOMATA annual, small to medium, pileate, in clusters, slightly

coriaceous and spongy when fresh, becoming hard on drying. PILEUs

15-30 mm long, 3-6 mm thick, laterally attached with a distinct region

of attachment, rarely sessile, conchate, mostly flabelliform, sometimes

semicircular, fruit bodies fused at the region of attachment, minutely

pubescent to glabrous, cream white with pale brown zones when fresh, more

brownish when mature, surface with concentric zones. MARGIN even to

slightly wavy, incurved on drying, acute, sterile, concolor with the surface

pileus. HyYMENOPHORE poroid, creamish white, pale brown on bruising.

Pores 6-8 per mm, absent near margin, angular to irregular, pore tubes 2-3

mm long, non-stratified, white when fresh, cream on bruising. CONTEXT

1-2 mm thick, white, homogenous. Opor not distinctive. SPORE PRINT not

observed.

HyYPHAL SYSTEM monomitic, generative hyphae with clamp connections,

thin to thick walled. PILEIPELLIs an irregular cutis, with scattered short erect

agglutinated hyphal patches that are hyaline that are up to 20 um long. PILEAL

TRAMA interwoven, generative hyphae 2-5 um wide, hyaline, branched, thin

to mostly thick walled (1-2 um), clamp connections frequent. HYMENIAL

TRAMA interwoven, generative hyphae 2-4 um wide, hyaline, branched, thin

to mostly thick walled (1-2 um), clamp connections frequent. CystTip1a and

other sterile structures absent. BAsID1IA 12-30 x 4—5 um, clavate, 4-sterigmate,

with basal clamp connections. Basip1osPorREs 3-4 x 2-2.5 um, Q = 1.5-1.6,

Q = 1.83, subcylindric to cylindric, hyaline, guttulate, usually with one or

two large guttules, thin walled, smooth, inamyloid in Melzer’s reagent.

SPECIMEN EXAMINED: INDIA. KERALA: Kottayam, Vagamon, Pine valley, 9.6862°N

76.9052°E, on rotten wood of Pinus species, 25 October 2015, Vinjusha N. (ZGC

VN258).

KNOWN DISTRIBUTION: Asia, northern to southern Europe, circumpolar in

the boreal conifer zone.

ComMENTs: The Indian specimen resembles Postia tephroleuca (Fr.)

Jiilich, a species already reported from India (Ranadive 2013). However,

Rubellofomes cystidiatus & Spongiporus floriformis (India) ... 665

Fic. 1: Rubellofomes cystidiatus (ZGC VN667): A. Basidiomata; B. Encrusted cystidia;

C. basidiospore. Spongiporus floriformis (ZGC VN258): D. Basidiomata; E. Basidia with attached

basidiospores. Scale bars: A = 25 mm, B = 15 um, C = 10 um, D = 10 mm, E= 6 um.

P. tephroleuca fruitbodies are always sessile whereas fruit bodies of

our collection have a distinct point of attachment. Also, P tephroleuca

basidiospores are longer and narrower (4.5-6 x 1-1.5 um) than those of

our collection (3-4 x 2-2.5 um). According to the description from the

isotype by Lowe (1975), S. floriformis produces conchate to often spathulate

pileus with a central stem-like point of attachment. While Ryvarden & Melo

(2014) describe the same collection as petal-like, sessile, or effused reflexed.

According to Lowe (1975), the fruitbody trama contains some gloeoplerous

hyphae; however, we did not observe any hyphae with gloeoplerous

contents.

666 ... Nellipunath & Kumar

Discussion

Rubellofomes comprises two species, R. cystidiatus and R. minutisporus

(Rajchenb.) B.K. Cui & al. The presence of cystidia and larger spores

distinguish R. cystidiatus from R. minutisporus. Except for the absence of

distinct concentric zonations on the pileus and small variations in the pore

surface colour and size of basidiospores, characters of our collection fit well

with those in the original description of R. cystidiatus from South China (Han

& al. 2014, as Fomitopsis cystidiata). Since then, the species has never been

reported from elsewhere, and this is the first report of the species outside its

type locality.

The Kerala collection of Spongiporus floriformis resembles Postia

tephroleuca, except that P. tephroleuca has sessile fruit bodies and longer

and narrower basidiospores. Possession of leptocystidia and amyloid tramal

hyphae help separate S. rhodophilus Spirin & Zmitr. from our Spongiporus

collection. Spirin & al. (2006) reported conidial cells on the pileus of

S. floriformis, but we did not find any conidia in our collections. Reported

hosts of S. floriformis are Abies, Cedrus, Crataegus, Cupressus, Larix, Picea,

and Pinus (Ryvarden & Melo 2014). The collection from Kerala was obtained

from dead wood of Pinus.

Rubellofomes cystidiatus and Spongiporus floriformis are new records for

the mycobiota in India.

Acknowledgments

The authors thank Dr. Ricardo Valenzuela Garza (Escuela Nacional de Ciencias

Bioldgicas Instituto Politécnico Nacional, Mexico) and Dr. Manoj Emanuel Hembrom

(Botanical Survey of India, India) for critically reviewing, and recommending our

manuscript. The authors also thank Dr. Mario Rajchenberg (National and Technical

Research Council, Argentina) and Dr. Fang Wu (Beijing Forestry University, China)

for reviewing an earlier version. Vinjusha Nellipunath acknowledges support from

the Kerala State Council for Science, Technology and Environment (KSCSTE) in

the form of a research fellowship. The authors also thank the Chief Conservator of

Forests & Chief Wildlife Warden, Kerala, for permission for field work in the forest

areas of Kerala.

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MY COTAXON

July-September 2021— Volume 136, pp. 669-673

https://doi.org/10.5248/136.669

Diplodia bulgarica, a new record for Turkey

CAFER EKEN

Department of Agricultural Biotechnology, Faculty of Agriculture,

Aydin Adnan Menderes University, Aydin 09970 Turkey

CORRESPONDENCE TO: cafereken@hotmail.com

ABSTRACT—Diplodia bulgarica on Malus domestica, a new species for the Turkish mycobiota,

is described briefly and illustrated. A pathogenicity test showed that D. bulgarica was highly

aggressive to apple plants.

Key worps—canker, Botryosphaeriaceae, Dothideomycetes

Introduction

Apple (Malus domestica Borkh.) is an economically important horticultural

crop in Turkey; more than 3,500,000 t of apples are produced in Turkey every

year, making the country the fourth largest producer worldwide (FAO 2018).

Fungal pathogens are major biotic stresses affecting apple tree growth and

fruit production. Botryosphaeriaceae encompasses a range of morphologically

diverse fungi that are either pathogens, endophytes, or saprophytes, occurring

mainly on woody plants such as apple (Slippers & Wingfield 2007; Phillips

& al. 2012, 2013). Among the 23 genera recognized in this family is Diplodia

(Dissanayake & al. 2016). Diplodia is a large genus currently comprising more

than 1000 species (Phillips & al. 2012). Species of Diplodia can cause canker,

dieback, gummosis, fruit rot, and twig blight diseases of numerous woody hosts

(Crous & al. 2006, Slippers & Wingfield 2007, Lazzizera & al. 2008, Phillips &

al. 2012, Abdollahzadeh 2015, Nabi & al. 2020, Hinrichs-Berger & al. 2021).

Recently, a botryosphaeriaceous species associated with canker symptoms of

apple trees was observed in Egirdir district of Isparta province, Turkey. The

present paper reports the etiology of the disease.

670 ... Eken

Materials & methods

In 2019, extensive canker symptoms were observed on apple trees in Egirdir

district of Isparta province, Turkey. Samples were collected from symptomatic

trees showing cankers in branches and transported in a cooler to the laboratory.

Fungi were isolated from canker margins after surface sterilization of samples in

2% sodium hypochlorite solution and rinsing in two changes of sterile distilled

water and transferred to Petri plates (9 cm diam.) containing Potato Dextrose Agar

(PDA). Plates were incubated 5-7 days at 25 °C, and the fungal isolate cultures were

subculture using hyphal tips cut from the colonies under a dissecting microscope

and placed, one colony per plate, onto PDA. Identification of isolates to species

level was based on colony morphology and conidial characteristics, as described

by Phillips & al. (2012). Isolates are maintained at the Department of Agricultural

Biotechnology, Faculty of Agriculture, Aydin Adnan Menderes University, Aydin,

Turkey (ADUAB).

The identity of isolates was confirmed by analysis of the internal transcribed

spacer (ITS) region of ribosomal DNA (rDNA). Genomic DNA was extracted from

the mycelium from the surface of the PDA cultures using an Omega Bio-Tek SP-

Fungal DNA Kit according to the manufacturer's instructions. After DNA extraction

of two isolates (CEE-22, CEE-211), the rDNA ITS region was amplified using

primers ITS1/ITS4 (White & al. 1990). Amplicons were purified and sequenced

by RefGen (Ankara, Turkey). The nucleotide sequence was compared with the

GenBank nucleotide collection database through nucleotide BLAST search.

Preliminary pathogenicity tests were carried out on potted 2-year-old apple

seedlings (cv. Scarlet Spur) using the representative isolates from cankers; the

most virulent isolate (CEE-22) was used for greenhouse test. The outer bark at the

inoculation area was disinfected by spraying with 70% ethanol, and the plants were

wounded with a scalpel through the full thickness of the bark along a 1-cm long

slit and inoculated with a 5 mm plug from a 7-day-old culture grown on PDA and

covered with parafilm. Control seedlings were similarly wounded and plugs of sterile

PDA applied. The bark of the inoculated area was removed, and the lesion lengths

recorded after four weeks. Re-isolation was done from the margins of infected tissues

as described above and re-isolated fungi were identified by comparing colony and

conidial morphological characteristics to the original inoculated isolate (CEE-22).

Results

A total of 12 isolates were obtained by isolation from 12 cankered branches

of apple trees collected from different localities of Egirdir district of Isparta

province, Turkey. Based on morphological and cultural characteristics, these

isolates were tentatively identified as Diplodia bulgarica. The identity of

two representative isolates (CEE-22 and CEE-211) was confirmed by DNA

sequencing of the rDNA ITS region (GenBank accession no. MT239082 and

MZ442352). The description and illustrations of D. bulgarica given below

Diplodia bulgarica new for Turkey ... 671

Fic. 1. Diplodia bulgarica (ADUAB - CEE-22): a. Colony on PDA;

b. Conidia developing on conidiogenous cells; c. Septate and aseptate conidia.

are based on the Turkish collections of the material. This is the first report of

D. bulgarica from Turkey.

Taxonomy

Diplodia bulgarica A.J.L. Phillips, J. Lopes & S.G. Bobev,

Persoonia 29: 33. 2012. Fic. 1

CoLony growth at 25 °C on PDA reaches 47 mm in diameter after 3 days.

The colony was at first white, turned to gray to dark gray with irregular margin

having dark centre and formed pycnidia after 20-25 days were immersed in

culture. Pycnrip1A were globose to ovoid, dark brown to black. CONIDIA were

aseptate, smooth, thick walled, oblong to ovoid, both ends broadly rounded,

19.8-29.0 x 10.4-15.1 um, initially hyaline and later becoming pale brown

and sometimes 1-septate at maturity.

SPECIMENS EXAMINED: TURKEY, ISPARTA PROVINCE, Egirdir district, Serpil village,

from cankered branches of Malus domestica, 2 Nov. 2019, C. Eken CEE-22 (ADUAB;

GenBank MT239082); 9 Nov. 2019, CEE-46 (ADUAB)); Tepeli village, from cankered

branches of Malus domestica, 8 Jan. 2019, C. Eken CEE-211 (ADUAB; GenBank

MZ442352); 27 May 2019, CEE-212, CEE-223 (ADUAB); Balkiri village from

cankered branches of Malus domestica, 7 Dec. 2019, C. Eken CEE-141 (ADUAB);

Barla village from cankered branches of Malus domestica, 27 July 2019, C. Eken

CEE-178, CEE-179 (ADUAB); Mahmatlar village from cankered branches of Malus

domestica, 5 Oct. 2019, C. Eken CEE-186, CEE-191 (ADUAB); Sevingbey village from

cankered branches of Malus domestica, 24 Aug. 2019, C. Eken CEE-123, CEE-124

(ADUAB).

Symptoms of canker were observed on the stem of the inoculated apple

seedlings. Canker lesions expanded to 52 + 0.4 mm four weeks after

inoculation and extended both upwards and downwards from the inoculation

672 ... Eken

sites. The pathogen was successfully re-isolated from all inoculated stems and

isolate identity was confirmed based on colony and conidial morphological

characteristics. In control treatments, no symptoms developed.

Discussion

Diplodia bulgarica, originally isolated from dead twigs of Malus sylvestris

in 2005, was described by Phillips & al. (2012). In our study, Diplodia

bulgarica was isolated from apple trees (M. domestica) in the Egirdir district

of Isparta province of Turkey, showing canker symptoms consistent with

previous reports (Abdollahzadeh 2015, Hanifeh & al. 2017, Bari & al. 2019,

Nabi & al. 2020, Hinrichs-Berger & al. 2021).

The size of conidia of the Turkish isolates is within the ranges observed by

Phillips & al. (2012; 22.5-28.0 x 14.5-18.5 um), Hanifeh & al. (2017; 19-29 x

12.5-17.5 um), and Nabi & al. (2020; 20.0-28.0 x 9.0-14.5 um). The identity

of isolates was confirmed by rDNA ITS sequence analysis. BLAST searches

revealed that ITS sequences of Turkish isolates matched 99.82% (CEE-22)

and 100% (CEE-211) with the Bulgarian ex-holotype strain of D. bulgarica

(CBS 124136; GenBank GQ923854).

The pathogenicity of D. bulgarica strain CEE-22 obtained in this study was

confirmed through an artificial inoculation experiment in the greenhouse.

Diplodia bulgarica strain CEE-22 did cause canker lesions, and the strain

proved to be highly aggressive. Diplodia bulgarica has previously been

reported as an aggressive pathogen on M. domestica in Iran (Abdollahzadeh

2015, Hanifeh & al. 2017, Bari & al. 2019), India (Nabi & al. 2020), and

Germany (Hinrichs-Berger & al. 2021). This is the first report of D. bulgarica

associated with a disease of M. domestica from Turkey. This pathogen is

considered to be an important agent causing canker on apple trees in Isparta

province of Turkey.

Acknowledgments

The author is indebted to Prof. Ismail Erper (Kyrgyz-Turkish Manas University,

Bishkek, Kyrgyzstan), Asst. Prof. Tuba Geng Kesimci (Igdir University, Igdir, Turkey)

and Asst. Prof. Artur Alves (University of Aveiro, Aveiro, Portugal) for reviewing

the manuscript; also I thank Dr. Lorelei L. Norvell, Editor-in-Chief, and Dr. Shaun

Pennycook, Nomenclature Editor, Mycotaxon for their efforts.

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industry in Iran. Phytopathologia Mediterranea 54: 128-132.

https://doi.org/10.14601/Phytopathol_Mediterr- 14686

Diplodia bulgarica new for Turkey ... 673

Bari ZB, Abrinbana M, Ghoosta Y. 2019. Distribution of Diplodia bulgarica, the causal agent of

apple diplodia canker, in central regions of West Azarbaijan province. In: 2nd International

and 6th National Iranian Congress on Organic vs. Conventional Agriculture (25-26 August

2019), Ardabil, Iran.

Crous PW, Slippers B, Wingfield MJ, Rheeder J, Marasas WFO, Phillips AJL, Alves A, Burgess

'T, Barber P, Groenewald JZ. 2006. Phylogenetic lineages in the Botryosphaeriaceae. Studies

in Mycology 55: 235-253. https://doi.org/10.3114/sim.55.1.235

Dissanayake AJ, Phillips AJL, Li XH, Hyde KD. 2016. Botryosphaeriaceae: current status of

genera and species. Mycosphere 7: 1001-1073. https://doi.org/10.5943/mycosphere/si/1b/13

FAO. 2018. Food and Agriculture Organization of the United Nations, statistical database.

http://faostat.fao.org (accessed March 23, 2020).

Hanifeh S, Zafari D, Soleimani MJ. 2017. Reaction of some apple cultivars to Diplodia bulgarica

in Iran. Mycosphere 8: 1253-1260. https://doi.org/10.5943/mycosphere/8/2/9

Hinrichs-Berger J, Zegermacher K, Zgraja G. 2021. First report of Diplodia bulgarica causing

black canker on apple (Malus domestica) and pear (Pyrus communis) in Germany. New

Disease Reports 43(e12004): [3 p.].

Lazzizera C, Frisullo S, Alves A, Phillips AJL. 2008. Morphology, phylogeny and pathogenicity

of Botryosphaeria and Neofusicoccum species associated with drupe rot of olives in Southern

Italy. Plant Pathology 57: 948-956. https://doi.org/10.1111/j.1365-3059.2008.01842.x

Nabi SU, Raja WH, Mir JI, Sharma OC, Singh DB, Sheikh MA, Yousuf N, Kamil D.

2020. First report of Diplodia bulgarica a new species causing canker disease of

apple (Malus domestica Borkh.) in India. Journal of Plant Pathology 102: 555-556.

https://doi.org/10.1007/s42161-019-00445-w

Phillips AJL, Lopes J, Abdollahzadeh J, Bobev S, Alves A. 2012. Resolving the

Diplodia complex on apple and other Rosaceae hosts. Persoonia 29: 29-38.

https://doi.org/10.3767/003158512X658899

Phillips AJL, Alves A, Abdollahzadeh J, Slippers B, Wingfield MJ, Groenewald JZ, Crous

PW. 2013. The Botryosphaeriaceae: genera and species known from culture. Studies in

Mycology 76: 51-167. https://doi.org/10.3114/sim0021

Slippers B, Wingfield MJ. 2007. Botryosphaeriaceae as endophytes and latent pathogens

of woody plants: diversity, ecology and impact. Fungal Biology Reviews 21: 90-106.

https://doi.org/10.1016/j.fbr.2007.06.002

White TJ, Bruns T, Lee S, Taylor J. 1990. Amplification and direct sequencing of

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https://doi.org/10.1016/B978-0-12-372180-8.50042-1

MY COTAXON

July-September 2021— Volume 136, pp. 675-684

https://doi.org/10.5248/136.675

Gerhardtia foliicola, a new record for Pakistan

AIMAN IZHAR*, MUHAMMAD USMAN,

MUNAZZA KIRAN, ABDUL NASIR KHALID

Fungal Biology and Systematics Research Laboratory, Institute of Botany,

University of the Punjab, Quaid-e-Azam Campus-54590, Lahore, Pakistan

* CORRESPONDENCE TO: aimanizhar25@gmail.com

ABSTRACT—Gerhardtia foliicola is reported as a new record for Pakistan. The species is

characterized by a reddish brown and hygrophanous pileus, densely crowded lamellae, small

sized ellipsoid basidiospores with irregular outline and occurrence on leaf litter and humus in

broadleaf forests. Our phylogenetic analysis based on nuclear ribosomal internal transcribed

spacer (nrITS) region clustered the Pakistani collections in the same clade with G. foliicola

sequences from Japan. Detailed description of macro- and micro-characters, habitat, general

distribution, and diagnostic features important for delimiting G. foliicola from closely related

taxa are presented.

Key worps—Agaricales, biodiversity, molecular phylogeny, Lyophyllaceae, taxonomy

Introduction

Gerhardt (1982) published Lyophyllum subg. Lyophyllopsis Ew.

Gerhardt [non Lyophyllopsis Sathe & J.T. Daniel (Sathe & al. 1981)], with

L. incarnatobrunneum Ew. Bernhardt as its only species. Bon (1994)

raised this subgenus to generic rank as Bernhardtia Bon stat. & nom. nov.

(Agaricales, Lyophyllaceae). The type species, G. incarnatobrunnea (Ew.

Bernhardt) Bon, is a more recent heterotypic synonym of G. borealis (Fr.)

Contu & A. Ortega (Contu & Ortega 2002). Distinguishing characters

of the genus include siderophilous granulation of basidia, minutely

verrucose ornamented basidiospores, pileipellis in form of a cutis or a

trichoderm, and the absence of clamp connections (Bon 1994). Vizzini &

676 ... Izhar & al.

al. (2015) emended the genus recently to emphasize smooth or verruculose

basidiospores, an absence of clamp connections, and a pileipellis in form of

a cutis, trichoderm, or hymeniderm.

Gerhardtia is a small genus (He & al. 2019) with c. seven accepted

species which are distributed in Europe, New Zealand, North and South

America, and China (Baroni 1981; Bon 1994; Contu & Ortega 2002; Contu

& Consiglio 2004; Kalamees 2008; Mesic & Tkaléec 2009; Cooper 2014;

Vizzini & al. 2015, 2017; Li & al. 2017; Matheny & al. 2017).

Gerhardtia foliicola, originally described from Japan as Tricholoma

foliicola Har. Takah. (Takahashi 2001), is characterized by a saprotrophic

habit and smooth basidiospores with an irregular outline (Endo & al. 2019).

Species of Tricholoma (Fr.) Staude are characterized by tricholomatoid

stature, smooth, white, and sub-globose to oblong basidiospores, and

an ectomycorrhizal association with woody plants (Singer 1986), which

conflict with the key characteristics of Gerhardtia.

Knowledge about Gerhardtia is limited, and previously no species of

the genus has been reported from Pakistan. During various mycological

expeditions, basidiomata of G. foliicola were collected that are presented

here as a new species record for Pakistan mycobiota.

Materials & methods

Collection site

Specimens were collected from Kumrat valley which lies at the end of Dir Upper

district, Khyber Pakhtunkhwa, Pakistan. The area falls in the alpine zone of the

foothills of Hindukush where elevation ranges from 2439 to 3048 m above sea level

and the annual rainfall is 52.43 cm (Stucki & Khan 1999, Siddiqui & al. 2013). The

area has a dry temperate climate and is dominated by Pinus wallichiana A.B. Jacks.

and Cedrus deodara (Roxb. ex D. Don) G. Don (Gillett 2001, Ahmad & Nizami 2015).

Morphological examination

Basidiomata were collected during July-August 2019 and photographed and

morphologically characterized in the field. Munsell (1975) is followed for color

designations. For long term preservation, specimens were dried using an electric

fan heater. Basidiomata were hand-sectioned and mounted in 5% KOH and in

Congo red for contrast purposes. Melzer’s reagent was used to check basidiospore

amyloidity. Specimens were examined microscopically using a Meiji MX4300H

compound microscope. Basidiospore, basidial, basidiole, and pileus and stipe hyphae

measurements were determined from atleast 20 observations at 1000 x magnification.

Gerhardtia foliicola new to Pakistan ... 677

The abbreviation [n/l/p] refers to n (number of basidiospores) measured from |

(number of basidiomata) and p (number of collections). Basidiospore dimensions

are given as (a—)b-c(-d). The abbreviations used are Q = length/width ratio of spores

and avQ = average Q from all basidiospores + standard deviation (Ge & al. 2010). All

specimens were deposited in the Herbarium, Institute of Botany, University of the

Punjab, Lahore, Pakistan (LAH).

DNA extraction, PCR and DNA sequencing

Genomic DNA was extracted from 5-20 mg of dried specimens using a modified

CTAB method (Bruns 1995). The ITS1 +5.8S ITS2 regions were amplified in a

polymerase chain reaction nuclear ribosomal DNA using primers ITSIF and ITS4

(White & al. 1990, Gardes & Bruns 1993). Amplified products were sent to TsingKe

China for further purification and bidirectional sequencing.

Phylogenetic analyses

Both forward and reverse primer reads were manually edited and assembled to

generate a consensus sequence in BioEdit 7.2.5. (Hall 1999). Homology searches

were carried out at NCBI webpage using the BLAST tool, and sequences with closest

identity were downloaded. Closest relatives of the species and some other sequences

representing the Lyophyllaceae were selected following phylogenetic studies

(Moncalvo & al. 2000; Hofstetter & al. 2002; Cooper 2014; Bellanger & al. 2015;

Vizzini & al. 2015, 2017; Latha & al. 2016; Li & al. 2017; Matheny & al. 2017; Yang &

al. 2018; Endo & al. 2019). All ambiguous portions with missing data and gaps were

omitted from the final matrix. Online MUSCLE tool by EMBL-EBI, (http://www.

ebi.ac.uk/Tools/msa/muscle/) was used for multiple sequences alignment (Edgar

2004). The maximum likelihood tree was generated using CIPRES Science Gateway

employing RaxML -HPC 2 on XSEDE with 1000 bootstrap pseudoreplicates (Miller

& al. 2010). Rapid bootstrap search was performed and GTRGAMMA model was

chosen. The generated tree was rooted with Myochromella boudieri (Kihner &

Romagn.) V. Hofst. & al. (GenBank ITS: AF357047) from Switzerland as outgroup

(Matheny & al. 2017). Bootstrap values >50% were cited on tree as significant.

Phylogenetic results

Our sequences showed 99% identity with Gerhardtia borealis (KR673544)

from Korea and 99% similarity with G. foliicola (LC458831, LC458827,

LC458828, LC458834, LC458829, LC458833) from Japan (Fic. 1). A total of

39 ITS sequences were assembled for phylogenetic reconstruction. The final

alignment was 732 bp, of which 317 sites were conserved, 372 were variable,

261 were parsimony informative, and 102 were singletons. Maximum

likelihood analysis clustered the sequences from our Pakistani collection

with Japanese G. foliicola with a strong bootstrap support (94%).

678 ... Izhar & al.

81| MT476983 Ge

MT476982

LC458827

LC458832

LC458833 G

LC458831 ¢

KR673544

9° KP19253

66] KP858004

KP208750 Ger

100 KP012786 Ger. aratic

100K 3461890 Lyophyllum moncalvo

100]! KJ461906 Lyophyllum moncalvoanu

98 KJ461904 Lyophyllum moncalv

5? HM572525 Lyophyllum shimeji

93 100 ___ HM572526 Lyophyllum shimeji

KY684031 Lyophyllum shimeji

97 AF357039 Sagarnella

AF357041 Sagarnella gi

AF357047 Myochromella boudieri

0.06

Fic. 1. Maximum Likelihood analyses of Gerhardtia foliicola and related taxa based on ITS

sequences. The bootstrap values =50% given above branches. Species collected from Pakistan are

highlighted in bold. The analysis included 37 nucleotide sequences.

Taxonomy

Gerhardtia foliicola (Har. Takah.) N. Endo, S. Ushijima, Nagas.,

Sotome, Nakagiri, & N. Maek., Mycoscience 60(6): 324. 2019. Fics 2, 3

Basip1omata thick, dry, clustered. PILEus 35-75 mm, hemispherical

with rolled margins at first, later becoming convex to applanate, surface

Gerhardtia foliicola new to Pakistan ... 679

-_—

re.

SAN}

Fic. 2. Gerhardtia foliicola: basidiomata.

A, B = LAH 36419: C, D = LAH 36420. Scale bars = 2 cm.

glabrous, shiny when young, later dry, hygrophanous, bright reddish brown

(5YR5/6), dull reddish brown (5YR5/4) to dull orange (7.5YR7/4), somewhat

paler towards the margin. LAMELLAE white (N10), adnate to emarginate,

thin, dense, lamellulae abundant. Stipe 40-120 x 10-30 mm, equal to

clavate, sometimes with a bulbous base, slightly fibrillose, base covered with

white tomentum, thick, solid, white (N10), light grey (5Y8/2) near the apex.

ANNULUS and votva absent. Opor and TASTE not recorded.

Basip1ospores [40/2/1], (4-)4.7-5.2(-5.5) x (1.7—)1.9-2.5(-2.6) um, avl

x avw = 4.6 x 2 um, Q = 2.1-2.3, avQ = 2.2 um, oblong to ellipsoid, smooth,

sometimes with tubercles of irregular outline, thin-walled, hyaline in KOH,

inamyloid. Basrp1a (19-)20-25(-27) x (4-)4.6-6.2(-7.6) um, avl x avw =

680 ... Izhar & al.

D f

Fic. 3. Gerhardtia foliicola (LAH 36419, LAH 36420).

A. Basidia; B. Basidiospores; C. Basidioles; D. Pileus hyphae; E: Stipe hyphae.

Scale bars = 5 um. Line drawings by: Aiman Izhar.

Gerhardtia foliicola new to Pakistan ... 681

22.6 x 5.3 um, clavate, 4-spored, hyaline in KOH, clamp connections absent.

BASIDIOLES (10.6—)13-23(-28) x (2.8—)2.9-4.7(-5) um, avl x avw = 17.7 x

4.2 um, clavate, basal cells septate, hyaline in KOH, clamp connections

absent. CHEILOCYSTIDIA and PLEUROCYSTIDIA absent. PILEIPELLIS a cutis,

hyphae 2-6.5 um (avw = 4 um) diam., thin-walled, septate, branched, ends

clavate, cylindrical, hyaline in KOH. ST1IPITIPELLis a cutis of parallel hyphae,

4-11 um (avw = 7.7 um) diam., hyaline in KOH, frequently septate, clamp

connections absent.

HaBiTaT & DISTRIBUTION— Growing on leaf litter under cedars and

pines. This species was originally reported from Japan and now for the first

time from Khyber Pakhtunkhwa province, Pakistan.

SPECIMENS EXAMINED— PAKISTAN. KHYBER PAKHTUNKHWaA: Dir (Upper) district,

Kumrat valley, 35°34’56”N 72°10’10’E, elev. 2439 m, found among pine needles, 25

July 2019, Abdul Nasir Khalid K-204 (LAH 36419; GenBank: MT476982; found on

forest floor under Cedrus deodara and Pinus wallichiana, 15 August 2019, Abdul

Nasir Khalid KU-19 (LAH36420; GenBank: MT476983).

Discussion

Gerhardtia foliicola, originally described from Japan as Tricholoma foliicola,

is characterized by its habit on leaf litter, a dry, glabrous and hygrophanous,

reddish brown pileus, adnate and very crowded lamellae, small ellipsoid

basidiospores, and hyphae lacking clamp connections (Takahashi 2001). After

taxonomic reconsideration, Reschke & al. (2018) recommended transfer

of T. foliicola to Gerhardtia, a recombination later supported by molecular

phylogenetic analyses (Endo & al. 2019). Takahashi (2001) reported this

species from a broadleaf forest dominated by Quercus myrsinifolia and

Q. serrata; Endo & al. (2019) reported it as growing on humus or on leaf or bark

litter of coniferous or broadleaf trees. Our findings confirm its occurrence in

coniferous forests.

Our molecular phylogenetic analysis clustered the Pakistani collections in

the same clade with Japanese collections of Gerhardtia foliicola and with a

specimen from South Korea (Ulleungdo) labeled as G. cf. borealis (KR673544),

which probably also represents G. foliicola. Phylogenetically our nrITS

sequences have a single nucleotide difference at the 395 bp position when

compared with the Japanese sequences. The Pakistani G. foliicola specimens

possess a larger pileus, longer stipe, oblong to ellipsoid basidiospores, and

thinner pileus hyphae contrasting slightly with the Japanese collections

(characterized by a smaller pileus, shorter stipe, sub-cylindrical, ellipsoid or

subfusiform basidiospores, and broader pileipellis hyphae). Geographically,

682 ... Izhar & al.

Japanese collections are recorded from temperate region and on humus and

litter under conifers (Pinus thunbergii Parl., P. densiflora Siebold & Zucc.,

P. parviflora Siebold & Zucc., Tsuga diversifolia (Maxim.) Mast., and broadleaf

trees (Quercus spp.), (Endo & al. 2019). The Pakistani specimens were recorded

from a dry temperate climate under Cedrus deodara and Pinus wallichiana.

Gerhardtia borealis, a sister species to G. foliicola is morphologically the

closest counterpart. However, it differs from G. foliicola by its quite distant

lamellae, larger (6—8.5 x 4—5 um) basidiospores, and growth on clay or sandy

soil in coniferous forests (Contu & Consiglio 2004). This species is also

phylogenetically well separated, forming a monophyletic clade that is sister to

G. foliicola (Gerhardt 1982, Bigelow 1985, Contu & Consiglio 2004).

Acknowledgements

Higher Education Commission, Pakistan is thanked for financial assistance under

National Research Program for Universities, project no. 20-3383/NRPU/R&D/

HEC/14/184. Authors are grateful to Dr. Junaid Khan (Center for Plant Sciences

and Biodiversity, University of Swat, Pakistan), Dr. Ting Li (Guangdong Institute of

Microbiology, Guangzhou, China), and Dr. Armin MeSi¢ (Ruder BoSkovic Institute,

Zagreb, Croatia) for their peer reviews and helpful comments to improve this

manuscript.

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South-west India - Part I: Agaricales (mushrooms) of Maharashtra State. Maharashtra

Association for the Cultivation of Science, Monograph 1. 114 p.

Siddiqui MF, Shaukat SS, Ahmed M, Khan N, Khan IA. 2013. Vegetation environment

relationship of conifer dominating forests of moist temperate belt of Himalayan and

Hindukush regions of Pakistan. Pakistan Journal of Botany 45: 577-592.

Singer R. 1986. The Agaricales in modern taxonomy (4th ed.). Koenigstein: Koeltz Scientific

Books.

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Takahashi H. 2001. Notes on new Agaricales of Japan 3. Mycoscience 42: 355-360.

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Vizzini A, Consiglio G, Setti L, Ercole E. 2015. Calocybella, a new genus for Rugosomyces pudicus

(Agaricales, Lyophyllaceae) and emendation of the genus Gerhardtia. IMA Fungus 6: 1-11.

https://doi.org/10.5598/imafungus.2015.06.01.01

Vizzini A, Angelini C, Ercole E. 2017. Is the species diversity in the lyophylloid genera Calocybella

and Gerhardtia (Agaricales, Basidiomycota) underestimated? Two new species from the

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fungal ribosomal RNA genes for phylogenetics. 315-322, in: MA Innis & al. (eds).

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Yang SD, Huang HY, Zhao J, Zeng NK, Tang LP. 2018. Ossicaulis yunnanensis sp. nov.

(Lyophyllaceae, Agaricales) from southwestern China. Mycoscience 59: 33-37.

https://doi.org/10.1016/j.myc.2017.07.008

MYCOTAXON

July-September 2021— Volume 136, p. 685

https://doi.org/10.5248/136.685

REGIONAL ANNOTATED MYCOBIOTA NEW TO THE MYCOTAXON WEBSITE:

ABSTRACT—MycotTaxon is pleased to add a new annotated species distribution list to

our 146 previously posted free-access fungae. The 39-page “Checklist of the lichens of

The Reserva Florestal Adolphe Ducke in Manaus” by Aptroot, Cavalcante, Santos, Junior,

Lima, and Caceres may be downloaded from our website via

http://www.mycotaxon.com/mycobiota/index.html

SOUTH AMERICA

Brazil

ANDRE APTROOT, JANICE GOMES CAVALCANTE, LIDIANE ALVES DOS SANTOS,

ISAIAS OLIVEIRA JUNIOR, DAYANE DE OLIVEIRA LIMA, MARCELA EUGENIA

DA SILVA CACERES. Checklist of the lichens of The Reserva Florestal

Adolphe Ducke in Manaus (Amazonas, Brazil). 39 p.

ABSTRACT—A checklist is presented of the lichenized fungi of The Reserva

Florestal Adolphe Ducke in Manaus (Amazonas, Brazil), which is one of the

best studied and most accessible forest reserves in the Amazon. In total, 295

species are reported, 116 of which are newly reported here based on new

sampling by the authors, 10 of which are first reports for Brazil, and 50 more

are new reports to the Amazonas state. Among these, 214 are corticolous

species, 81 are foliicolous and five are terricolous or on termitaria, with five

species occurring both on bark and living leaves.

KEY worps—Amazon Forest, living leaves, diversity, termite nest, tree bark

MYCOTAXON

July-September 2021— Volume 136, p. 687

https://doi.org/10.5248/136.687

REGIONAL ANNOTATED MYCOBIOTA NEW TO THE MYCOTAXON WEBSITE:

AxBsTRACT—The 13-page “First molecular-based contribution to the checklist

of Lebanon macrofungi” by Sleiman, Bellanger, Richard, and Stephan may now

be downloaded from Mycotaxon’s mycobiota webpage. This well-documented

annotated species list brings to 148 the number of free-access fungae now available

on our website: http://www.mycotaxon.com/mycobiota/index.html

MID-EAST

Lebanon

SANDRA SLEIMAN, JEAN-MICHEL BELLANGER, FRANCK RICHARD, JEAN STEPHAN.

First molecular-based contribution to the checklist of Lebanon macrofungi.

13 p.

AxsstRacT—Across the Mediterranean biodiversity hotspot, our knowledge

of fungal diversity is still fragmentary, and the eastern part of the basin remains

poorly explored. This is particularly the case in Lebanon, where macrofungal

diversity has never been subject to molecular-based investigation. A first checklist

was published in 1957 and updated in 1996, based on morphological identification

of specimens. In an effort to narrow this gap of knowledge, we designed a field

survey in Akkar, north Lebanon in Abies cilicica, Quercus calliprinos and Quercus

cerris old-growth stands, carried out between 2018 and 2019. Three sampling plots

were monitored in each ecosystem. By combining morphological identification and

molecular analysis of 105 collected samples, 79 species were identified, including

62 (i.e., 78.5%) new records for Lebanon, and eight putative undescribed species.

Species diversity was the highest under Quercus calliprinos and lowest under Abies

cilicica. Most species were encountered in just one ecosystem type, and only 7 were

recorded in two ecosystems. These results highlight the overlooked fungal diversity

in this part of the eastern Mediterranean hotspot, and the importance of Lebanese

forests for biodiversity conservation. Our results also call for further investigation,

in particular under endemic tree hosts, to increment the still incomplete national

checklist of macrofungi in Lebanon.

Key worps—basidiomycetes, forest ecosystems, mycology, Mediterranean forests,

fungal diversity

MYCOTAXON

July-September 2021— Volume 136, p. 689

https://doi.org/10.5248/136.689

REGIONAL ANNOTATED MYCOBIOTA NEW TO THE MYCOTAXON WEBSITE:

ABsTRACT—The 10-page “Checklist of powdery mildew fungi (Erysiphaceae) in

Pakistan” by Afshan, Zafar, and Khalid may now be downloaded from Mycoraxon’s

mycobiota webpage. This annotated species list brings to 149 the number of free-

access fungae now available on our website:

http://www.mycotaxon.com/mycobiota/index.html

ASIA

Pakistan

N.S. AFSHAN, I. ZAFAR, A.N. Kuatip. Checklist of powdery mildew fungi

(Erysiphaceae) in Pakistan. 10 p.

ABSTRACT—This paper presents a checklist of powdery mildews (Erysiphaceae) in

Pakistan together with their known host plants. A total of 42 taxa belonging to 10

genera are included here. A host index is also provided..

KEY worDS—angiosperms, Ascomycota, biotrophic parasites, Erysiphe

MYCOTAXON

July-September 2021— Volume 136, p. 691

https://doi.org/10.5248/136.691

REGIONAL ANNOTATED MYCOBIOTA NEW TO THE MYCOTAXON WEBSITE:

ABSTRACT—MycotTaxons pleased to add our 150th annotated species

distribution list to our previously posted free-access fungae. The extraordinarily

well-documented 57-page “Aphyllophoroid fungi (Basidiomycota) of Chile: An

annotated checklist” by Riquelme & Rajchenberg may be downloaded from our

website:

via http://www.mycotaxon.com/mycobiota/index.html

SOUTH AMERICA

Chile

CRISTIAN RIQUELME & MAarIO RAJCHENBERG. Aphyllophoroid fungi

(Basidiomycota) of Chile: An annotated checklist. 57 p.

ABSTRACT—We critically reviewed the aphyllophoroid fungal species recorded

from Chile on the basis of literature reviews from 1828 to 2020. A total of 345

species names were distributed in the following categories: accepted taxa (236),

uncertain taxa (83), taxa inferred by molecular analysis only (12), and excluded

taxa (14). For each accepted species, information on the species name, its first

record from Chile, the typification data, substrate, distribution, and remarks

are presented. The distribution of accepted taxa shows a major recording from

the Valdivian Forest province. We found that 61 (25.85%) of the accepted taxa

were originally described from Chile. Nothofagus was the genus that hosted the

highest number of fungal species.

Kry worps—aphyllophorales, coralloid fungi, corticioid fungi, polypores