Iv ... MYCOTAXON 129(2)

MY COTAXON

VOLUME ONE HUNDRED TWENTY-NINE — TABLE OF CONTENTS

COVER SECTION 129(2)

1 By Ake 1c eRe ae Oh, PreK EE Le Ae TR i Aa NN, | Seat r o Se a, Pre vi

ROVICWCTS ENE rch UE eee RE Sep eek Ett Ay he ote Pats My Ake ete es vii

SO IISSIO MD TOCCHUMES 3 xis vi Peasrih Yate ct hve Reg a, aa a meri Joa tctteere: Hyg I eS Viii

ERO MES EELO Mel? B Fhe ety inn Pie de Oe a an, A hey nw et ER ix

RESEARCH ARTICLES

Morphological and molecular evidence for a new species of Psilocybe

from southern China YANG K. Li, YE YUAN, & JUN E LianG 215

Powdery mildew on Bougainvillea spectabilis in Mexico with a

cryptic generic afhnity SYLVIA P. FERNANDEZ-Pavia,

GERARDO RODRIGUEZ-ALVARADO, JESUS GERONIMO-MAGANA,

Maria GRACIELA CABRERA, NURIA GOMEZ-DORANTES, & UWE BRAUN 223

Pseudocercospora styracigena sp. nov. on Styrax and Alniphyllum

from China Bao-Ju Lt, A-L1 CHal, & YING-LAN Guo 229

Neozygites species associated with aphids in Chile: current status and

new reports CRISTIAN MONTALVA, MAREK BARTA, ELADIO Rojas,

MONICA GUTIERREZ, & EDUARDO VALENZUELA 233

First report of Pseudoveronaea ellipsoidea causing sooty blotch and

flyspeck in China CHEN CHEN, WENHUAN LI, Liu Gao,

RONG ZHANG, GUANGYU SUN, & MARK L. GLEASON 247

Molecular annotation of type specimens of Russula described by

W.A. Murrill from the southeast United States BRIAN P. Looney 255

Molecular phylogeny of placodioid lichen-forming fungi reveal a

new genus, Sedelnikovaea S.Y. KonDRATYUK, M.-H. JEONG,

I.A. GALANINA, L.S. YAKOVCHENKO, A.P. YATSYNA, & J.-S Hur 269

A new species of Leptostroma on Pinus henryi from China

LAN ZHANG, D.W. MINTER, QING LI, & YING-REN LIN 283

A new Stemonitis species from Turkey

Hasan HUsEYIN DOGAN & GONUL EROGLU 293

A new species of Coccomyces with dimorphic paraphyses

X1IAO-YAN WANG, Halr-LIn Gu, QING LI, LEI-HONG WANG, & YING-REN LIN 297

Cytospora from Salix in northern China X1n-Ler FAN, CHENG-MING TIAN,

QIN YANG, YING-MEI LIANG, CHONG-JUAN YOU, & YU-BO ZHANG 303

New record of Melanoleuca cinereifolia in Himalayan moist temperate

forests of Pakistan M. SaBA & A.N. KHALID 317

OCTOBER-DECEMBER 2014 ... V

Entoloma species from New South Wales and northeastern

Queensland, Australia Davip L. LARGENT,

SARAH E. BERGEMANN, & SANDRA E. ABELL-Davis 329

Leptocorticium indicum sp. nov. from India

SAMITA, S.K. SANYAL, & G.S. DHINGRA 361

A new species of Podosphaera sect. Sphaerotheca from China

Lu-Cuao Bal & ZHI-MIN Cao 365

Rhizoglomus, a new genus of the Glomeraceae EWALD SIEVERDING,

GLADSTONE ALVES DA SILVA, REINHARD BERNDT, & FRITZ OEHL 373

First report of the pantropical species Diploschistes rampoddensis

from Europe SAMANTHA FERNANDEZ-BRIME, XAVIER LLIMONA,

NEsTOR HLADUN, & ESTER Gaya 395

New species of Graphium and Periconia from China

YuE-MING WU, JUN-JIE XU, JIN-HuA Kona, & TIAN-YU ZHANG 397

Two new species of Myrothecium from the

Qinghai-Tibet Plateau Area, China YuE-MING Wu, Yu-LAN JIANG,

Ya-NAN Ma, & TiAN-Yu ZHANG 403

Nomenclatural novelties in the Postia caesia complex VIKTOR Papp 407

The new lectotypification of Umbilicaria kisovana

(Umbilicariaceae, lichenized Ascomycota)

Evceny A. Davypov, LipIA YAKOVCHENKO, & YOSHIHITO OHMURA 415

Galerella xalapensis sp. nov. found in an urban green area

in Xalapa, Veracruz, Mexico VicTOR M. BANDALA & LETICIA Montoya 421

A new report of Uromyces ambiens on Buxus from Pakistan

SADIQULLAH, A. IsHAQ, M. Fiz, N.S. AFSHAN, & A.N. KHALID 429

Lophium elegans (Ascomycota), a rare European species

GEIR MATHIASSEN, ALFRED GRANMO, & TEPPO RAMA 433

New and interesting Laboulbeniales from southern and

southeastern Asia D. HAELEWATERS & S. YAAKOP 439

Dictydiaethalium dictyosporangium sp. nov. from China Bo ZHANG & Yu Li 455

Lophodermium quadrisporum sp. nov. (Rhytismataceae) on

Rhododendron faberi spp. prattii Dan-DaNn Lu, YAN-PING TANG,

LEI-HONG WANG, SHI-JUAN WANG, & YING-REN LIN 459

Diploschistes xinjiangensis, a new saxicolous lichen

from northwest China ABDULLA ABBAS, SHOU-YU GUO,

GULIBAHAER ABABAIKELI, ADILJIAN ABDULLA, & HUERNISA XAHIDIN 465

Uncispora hainanensis, a new species isolated from decayed leaves

JIAN- YING Li, MIN QIAO, JIE PENG, WEN-YUN QIAN,

GUANG-ZHU YANG, & ZE-FEN Yu 473

VI ... MYCOTAXON 129(2)

Lactarius vesterholtii, a new species from India

KANAD DAs & DYUTIPARNA CHAKRABORTY 477

REGIONAL MYCOBIOTAS NEW TO THE MYCOTAXON WEBSITE

A checklist of the fungi recorded from Serra da Jibdia, Bahia state, Brazil

FLAvia R. BARBOSA, MONIQUE MACHINER,

GLEYSON CRISTIANO K. BARBOSA, & Luis F. PR GusmMAo 485

BOOK REVIEWS AND NOTICES ELsE C. VELLINGA (Editor) 487

NOMENCLATURAL NOVELTIES AND TYPIFICATIONS

PROPOSED IN MYCOTAXON 129(2) 493

ERRATA FROM PREVIOUS VOLUMES

VOLUME 129(1)

p. 44, line 10 FOR: Sporidesmium-like genera, are polyphyletic (Shenoy

READ: Sporidesmium-like genera, is polyphyletic (Shenoy

p. 44, line 34 FOR: and sometimes the conidium between the

READ: and sometimes the portion of the conidium between the

p. 54, line TaBLE 2 FOR: Dai et al. 2007

READ: Dai YC, Yu CJ, Wang HC (2007) Polypores from eastern Xizang

(Tibet), western China. Ann Bot Fennici 44: 135-145.

PUBLICATION DATE FOR VOLUME ONE HUNDRED TWENTY-NINE (ONE)

MYCOTAXON for JuLy-SEPTEMBER, VOLUME 129[1] (I-x + 1-214)

was issued on November 19, 2014

OcTOBER-DECEMBER 2014 ... VII

REVIEWERS — VOLUME ONE HUNDRED TWENTY-NINE (TWO)

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 volume.

Hakan Alli

Joe Ammirati

Vladimir Antonin

A. Aptroot

N.S. Atri

José Luiz Bezerra

Uwe Braun

Ana Rosa Burgaz

Bart Buyck

Rafael E Castaheda-Ruiz

Shuang-Lin Chen

R.T.A. Cook

Danny Coyne

Bao-Kai Cui

John A. Elix

Ove E. Eriksson

Shouyu Guo

Nils Hallenberg

Liu-Fu Han

Anton Hausknecht

Cheng-Lin Hou

Lili Huang

Richard A. Humber

Yogesh Joshi

Ingeborg Klingen

Irmgard Krisai-Greilhuber

Steven Leavitt

Tai-Hui Li

VZ..Lite

Bruce McCune

Eric H.C. McKenzie

Julio Mena-Portales

Andrew S. Methven

Abdul Rehman Niazi

Lorelei L. Norvell

Jorinde Nuytinck

Omar Paino Perdomo

Shaun R. Pennycook

Liliane Petrini

Huzefa Raja

Valter Rossi

Amy Y. Rossman

Ivan Sanchez-Castro

Sergi Santamaria

Alexander Sennikov

B.M. Sharma

Danielle Karla Alves da Silva

Steven L. Stephenson

Guangyu Sun

Kazuaki Tanaka

Arne Thell

K.B. Vrinda

Yong Wang

Zheng Wang

Solomon P. Wasser

A.J.S. Whalley

Ming Ye

Xiu-Guo Zhang

Zhongyi Zhangt

Ivan V. Zmitrovich

vill ... MYCOTAXON 129(2)

FOUR STEPS TO SUCCESSFUL MYCOTAXON PUBLICATION IN 2015

Prospective MycoTaxon authors should download instructions PDF, review and

submission forms, and other helpful templates by clicking the ‘file download page’ link

on our INSTRUCTIONS TO AUTHORS page before preparing their manuscript. Below is a

summary of our ‘4-step’ publication process.

1—PEER REVIEW: Email formatted text and illustration files with a2014 MycoTAxon

Reviewer Comments Form to 2-3 experts for peer review. Authors should (i) ask

peer reviewers to return revisions and comment forms to BOTH authors and Editor-

in-Chief <editor@mycotaxon.com> and (ii) follow reviewer suggestions before

sending revised files to the Nomenclature Editor for nomenclatural review.

2—NOMENCLATURAL REVIEW: Email all text-based files (with phylotrees but No

photos/drawings) to the Nomenclature Editor <PennycookS@LandcareResearch.co.nz>

for accession and pre-submission review. Place ‘“MycoTaxon’ + first author

surname on the subject line; list all peer reviewer names+Email addresses in the

message. The Nomenclature Editor will assign accession numbers and return

annotated files with a list of needed corrections to the authors and Editor-in-Chief.

3—FINAL SUBMISSION: Consult experts, revise and thoroughly proof manuscripts,

and prepare error-free text and image files ready for immediate publication.

Then send the (i) completed 2015 Mycotaxon submission form; (ii) separate

text files for main text, tables, and legends; and (iii) art files to the Editor-in-Chief

<editor@mycotaxon.com>. The Editor-in-Chief usually Emails all coauthors and

expert reviewers within two weeks of final submission, but please wait at least

14 days before sending a follow-up query (without attachments); this helps us

keep Email traffic to a minimum during Mycotaxon publication deadlines or

temporary closures of the editorial office.

4—FINAL EDITORIAL REVIEW & PRESS PREPARATION: Files not ready for publication

will be rejected or returned to authors for further revision; the Editor-in-Chief

gives tentatvely approved manuscripts a final grammatical and scientific review

before converting all files into publishable format. The PDF proof, bibliographic

citation, and nomenclatural entries are sent to all coauthors for final inspection

prior to publication. Thereafter, the Editor-in-Chief corrects ONLY processing or

editorial errors prior to publication but will list corrections of author errors in the

ERRATA of a subsequent volume for no charge. Authors are expected to arrange

payment of page charges and optional open access fees with the Business Manager

<subscriptions@mycotaxon.com> at this time.

MyYcOTAXON LTD— www.mycotaxon.com

The Mycotaxon Webmaster <mycotaxon@gmail.com> posts general and

subscription information, important announcements, and author forms and templates

on the official MycoTaxon site. The server also hosts the regional mycobiota webpage

for free download of distributional annotated species lists.

MYCOTAXON ONLINE— www.ingentaconnect.com/content/mtax/mt

Mycotaxon publishes quarterly. Both open access and subscription articles are

offered.

OcCTOBER-DECEMBER 2014 ... IX

FROM THE EDITOR-IN-CHIEF

A FOND AU REVOIR TO ELSE — Responding to a series of wheedling editorial Emails

during late 2010, the esteemed Dr. Vellinga agreed to assume the mantle of Book

Review Editor from David Hawksworth—but with only (alas) a one to two year

commitment. Her fourth year now concluding and with the California drought

finally promising to break, Else wishes to relinquish her post to make time for

intensive field and lab research. We are sad to lose her but hope to publish some of

her research (and perhaps an occasional book review?) in the not too distant future.

MyYcOTAXON 129(2) contains 30 research papers by 115 authors (representing 23

countries) that have been previously revised by 115 expert reviewers.

Within its pages are two new genera (Rhizoglomus for arbuscular mycorrhizal

fungi previously placed in Rhizophagus and Sedelnikovaea for a distant relative of

Protoparmeliopsis), one new subgenus (Postia subg. Cyanosporus), and 28 species

new to science representing Coccomyces, Cytospora, Dictydiaethalium, Diploschistes,

Graphium, Leptostroma, Myrothecium, Periconia, Podosphaera, Pseudocercospora,

Psilocybe, and Uncispora from China; Entoloma from Australia; Galerella from

Mexico; Laboulbenia from The Philippines; Lactarius and Leptocorticium from India;

and Stemonitis from Turkey.

In addition to range extensions and/or new hosts for previously named taxa, we

also offer 18 new combinations in Postia, Rhizoglomus, and Sedelnikovaea, a new

typification for Umbilicaria kisovana, an examination of the potential of Neozygites

for aphid pest control in Chile, and an excellent molecular analysis of Murrill’s

southeastern U.S.A. Russula type specimens.

ONWARD TO 2015 — MycorTaxon is pleased to close 2014 having successfully

published a total of five issues (1267 pages), 140 research papers, and 144 new taxa.

Editorial health has improved and is holding, making attainable our new year’s goal

of publishing each quarterly well wITHIN the dates emblazoned on its spine (instead

of far too many weeks afterwards). We take this opportunity to wish all our readers,

authors, and reviewers the happiest, healthiest, and most productive New Year.

Warm regards,

Lorelei L. Norvell (Editor-in-Chief)

30 December 2014

MY COTAXON

http://dx.doi.org/10.5248/129.215

Volume 129(2), pp. 215-222 October-December 2014

Morphological and molecular evidence for a new species of

Psilocybe from southern China

YANG-KuN LI, YE YUAN, & JUN-FENG LIANG*

Research Institute of Tropical Forestry, Chinese Academy of Forestry

Guangzhou, 510520, P. R. China

*CORRESPONDENCE TO: jfliang2000@163.com

AxBstRAcT—Psilocybe cinnamomea sp. nov. is reported from China. It is characterized by

its campanulate to plano-convex pileus, ellipsoid spores with a distinctly broad germ pore,

clavate basidia, polymorphic cheilocystidia, and clavate to fusiform pleurocystidia with a

short unbranched neck. Phylogenetic relationships among the new species and allied species

in the genus were inferred from the internal transcribed spacer (ITS) region.

Key worps—Agaricales, Strophariaceae, taxonomy, phylogeny

Introduction

Psilocybe (Fr.) P. Kumm. was formerly a broadly circumscribed genus,

including bluing, hallucinogenic species and non-bluing, non-hallucinogenic

species (Guzman 1983, Allen et al. 1992, Stamets 1996, Guzman 2005,

Borovicka et al. 2011, Guzman 2012). Recently, this broad polyphyletic genus

was divided: Psilocybe sensu stricto (with P. semilanceata as its conserved type)

was restricted to the bluing species, while the non-bluing species were assigned

to Deconica (W.G. Sm.) P. Karst. (Redhead et al. 2007, Norvell 2010).

Psilocybe s. s. contains more than 150 species worldwide (Guzman 2005,

2009). However, only 11 species have been reported from China, of which

only three were originally described from the country (Bau & Sarentoya 2009,

Guzman & Zhu L. Yang 2010, Ma et al. 2014). In the present study, a new

species of Psilocybe is described from southern China. In order to confirm the

phylogenetic uniqueness of this species, sequences of its internal transcribed

spacer (ITS) were generated and compared with sequences of other Psilocybe

species.

Material & methods

Field collections were conducted in 2010. Notes and photographs were taken on

macro-morphological features, and specimens were dried using a Dérrex dehydrator at

50°C. Specimens were deposited in the Research Institute of Tropical Forestry (RITF),

216 .. Li, Yuan, & Liang

Chinese Academy of Forestry. Macroscopic descriptions were based on the photos and

notes made in the field. Descriptive terminology follows Vellinga (1988), and colours

are designated according to Kornerup & Wanscher (1981).

For microscopic observations, sections of specimens were cut by hand and mounted

in 5% aqueous KOH solution or 1% aqueous Congo red solution. Basidia, basidiospores,

cheilocystidia, pleurocystidia, caulocystidia, and pileocystidia were measured using the

MShot Digital Imaging System (n=25 for each character). The abbreviation [n/m/p]

indicates that measurements were made on n basidiospores in m basidiomata from p

collections. Basidiospore dimensions follow the form (a—)b-c(-d) with b-c representing

95% of the measured values and extreme values shown in parentheses.

DNA was extracted from herbarium materials (Zhou & Liang 2011). The internal

transcribed spacer (ITS) regions were amplified by polymerase chain reaction (PCR)

with the primers ITS1 and ITS4 (White et al. 1990). Both strands were sequenced with an

ABI 3730 DNA analyzer and an ABI BigDye 3.1 terminator cycle sequencing kit (Beijing

AuGCT DNA-SYN Biotechnology Co., Ltd, Beijing). The newly generated sequences

were submitted to GenBank (accession numbers KJ433483 and KJ433484). DNA

sequences were edited and aligned using SeqMan (DNA STAR Package) and Clustal

X (Thompson et al. 1997) and manually checked and adjusted. Ambiguous positions

were excluded from the matrix. The dataset was analyzed with the RAxML BlackBox

online server (Stamatakis et al. 2008) for the maximum likelihood and MrBayes 3.1 for

bayesian inference (Huelsenbeck & Ronquist 2005).

Taxonomy

Psilocybe cinnamomea J.F. Liang, Yang K. Li & Ye Yuan, sp. nov. Fig. 1

MycoBAank 807941

Differs from Psilocybe zapotecorum by its smaller basidiomata, its greenish white to pale

green lamellae, and its smaller basidia.

Type: China, Guangdong Province, Lechang County, Yangdongshan Shierdushui Nature

Reserve, 4 October 2010, Wang 189 (Holotype, RITF 746; GenBank KJ433484).

Erymo oey: The Latin word “cinnamomea” refers to the cinnamon-like color of young

basidiomata.

BASIDIOMATA small. PiLEus 20-40 mm in diameter, campanulate when young,

expanding to subconvex or plano-convex, slightly papillate at the apex, glabrous,

hygrophanous, cinnamon (5B3-5C6) when young, surface pale (2A2) to cream

(4A2-4), light orange (5A4-5B7) at the disk when mature, yellowish brown

(5C4-8) in dry specimens, blackish blue with age or when bruising, margin

with white floccose remnants from the veil. LAMELLAE subadnate or sinuate,

crowded, unequal, greenish white to pale green (28A2-3). STIPE 20-80 x 2-5

mm, uniform or slightly thickened at the base, somewhat flexuous, hollow,

whitish to pale green (5B3) at the upper, grayish orange (5B3-5) to brownish

yellow (5C6-8) at the lower part, covered with milk white (1A2) fibrillose,

bluing, blackish when dried. CoNTExT thin, whitish. ODor or TASTE not

recorded. ANNULUS absent.

Psilocybe cinnamomea sp. nov. (China) ... 217

; um

agirr00 |}

PS Ee

en) ke

| 10um

IF ik

Fic. 1 Psilocybe cinnamomea. A. Basidiomata. B. Basidiospores. C. Basidia.

D&H. Cheilocystidia. E. Pleurocystidia. F. Pileipellis.

G. Caulocystidia. (A-G from holotype, RITF 746; H from RITF 726).

218 .. Li, Yuan, & Liang

BASIDIOSPORES (5.5—)6.5—7.5(-8) x (3-)4-4.5(-5) um, ellipsoid to subovoid

in face view, subellipsoid in side view, sometimes subventricose, smooth, thin-

walled, wall about 0.3-0.5 um thick, with a distinctly broad germ pore and

a short apical appendage, yellowish-brown pigment in KOH. Basip1a 7-13.5

x 3.0-6.0 um, 4-spored, rarely 2-spored, hyaline, clavate, sometimes with a

median constriction. CHEILOCYSTIDIA common, (17.5-)20-30 x 6.5-10 um,

unbranched, polymorphic (subcylindrical, clavate-mucronate, utriform,

lageniform, fusiform), frequently with a median constriction and with a short or

long neck, hyaline in 5% KOH. PLEurocystip1a abundant, 17.5-32 x 5.5-10.0

um, clavate, narrowly fusiform to fusiform, frequently with a short neck, the

neck <5 um long, unbranched, hyaline, walls smooth. PILEIPELLIs a subcutis,

not subgelatinized, 9-13 um thick, hyaline, hyphae 3-5 um diam., rarely with

surface subcylindric or fusiform cystidioid elements 7.5-17.5 x 4.5-7.5 um.

CAULOCYSTIDIA rare, 9.5-85 x 5-11 um, variable in shape (subcylindrical or

attenuate). CLAMP connections common.

ADDITIONAL SPECIMEN EXAMINED: CHINA: Guangdong Province, Lechang County,

Yangdongshan Shierdushui Nature Reserve, 4 October 2010, Wang 234 (RITF 726;

GenBank KJ433483).

Hasitat & DISTRIBUTION - Gregarious to caespitose on rotten wood or

nutrient rich soils in a subtropical evergreen broad-leaved forest. Known only

from the type locality.

Phylogenetic analyses

A dataset of ITS sequences with 700 nucleotide sites was analyzed for 42 taxa

(including 41 from GenBank). The dataset contained available sequences of

Psilocybe species; Stropharia rugosoannulata Farl. ex Murrill and S. hornemannii

(Fr.) S. Lundell & Nannf. were chosen as outgroup.

Bayesian and RAxML phylogenetic analyses (Fic. 2) clustered together

two P. cinnamomea sequences on a well-supported branch (bootstrap = 100%;

posterior probability = 1.00) and placed P cinnamomea with P. antioquiensis

Guzman et al., P zapotecoantillarum Guzman et al., P zapotecorum R. Heim

emend., P. thaizapoteca Guzman et al., and P argentipes K. Yokoy. in the same

subclade /zapotecorum (bootstrap = 87%; posterior probability = 1.00).

Discussion

Psilocybe cinnamomea is easily recognized by its campanulate to plano-

convex pileus, ellipsoid spores with a distinctly broad germ pore, clavate basidia,

polymorphic cheilocystidia, and clavate to fusiform pleurocystidia with a short

unbranched neck. Its ellipsoid thin-walled spores and bluing discoloration

suggest placement of P cinnamomea in P. sect. Zapotecorum (Guzman 1983).

Psilocybe cinnamomea sp. nov. (China) ... 219

. subaeruginosa KC669296

Ie endeioetie GUS565173

93/1.00|! p, cyanescens GU565176

P. allenii HE994443

. thaiaerugineomaculans KC669298

. thaiduplicatocystidiata KC669299

P. ovoideocystidiata HE994451

P. mescaleroensis KC669290

. cubensis HM035075

‘P. subcubensis KC669297

P. serbica GU565177

75/1.00

91/1.00

P. moravica var. moravica GU565182

P. quebecensis AY 129373

95/1.00/-P. mexicana HM035083

P. tampanensis AF548391 :

P. samuiensis AB257586 |MexXIcanae

P. caerulescens HM035072

00/1.007P: vungensis KC669301

P. fagicola KC669288

P. antioquiensis JN113590

‘P. thaizapoteca KC669300

/1.00/P: cinnamomea KJ433484

'P. cinnamomea KJ433483 Zapotecorum

Q\P. zapotecorum KC669303

P. argentipes AB092792

P. zapotecoantillarum KC669302

P. semilanceata AY 129353

P. pelliculosa AY 129354

P. stuntzii KC669295

P. hispanica KC669289

P. silvatica AY 129362

P. caerulipes KC669282

93 P. rhombispora FJ596920

100/1.00 Lp crobula AY 129358

100/1.00 | P. australiana AY 129366

100/0.99P. merdaria AY995175

P. fasciata DQ001401

85/1.00

cordisporae

Clade I

87/1.

98/1.00

95/0.99

P. magnivelaris GU234140

P. calongei AJ519794

on nara hornemannii JNO21095 |

Stropharia rugosoannulata FJ810166 Outgroup

0.05

Fic. 2 One of 100 RAxML likelihood trees (-In L 5242.090489) based on the Psilocybe ITS dataset.

Support values in bold type are RAxML likelihood bootstrap (270%). Values in normal type are

Bayesian posterior probabilities (20.95).

220 .. Li, Yuan, & Liang

Observations of both collections suggest that P cinnamomea shows

considerable variation in shape and size of cheilocystidia. In young specimens

(Fic. 1H), the cheilocystidia are utriform to lageniform (17.5-22.5 x

3.5-5.5 um) with a long neck (<6.5 um long), while those in mature specimens

(Fic. 1D) are subcylindrical to clavate-mucronate with a short neck. Although

there are differences between young and mature specimens, they are confirmed

as representing the same species based on their same habitat and an ITS

sequence similarity between young and mature specimens of 99%.

Bayesian and RAxML phylogenetic analyses (Fic. 2) supported P. cinnamomea

in Clade I, representing hallucinogenic and mainly tropical and subtropical

taxa (Guzman et al. 2012) that were divided into three subclades (/cordisporae,

/mexicanae, /zapotecorum) that grouped together with 93% bootstrap support

and 0.98 posterior probability. Within /zapotecorum, P antioquiensis, which

has slightly thick-walled and angular spores, has been placed in P sect.

Mexicanae (Guzman et al. 1994), while the other species, which share ellipsoid,

thin-walled, non-angular spores, have been assigned to P. sect. Zapotecorum

(Guzman et al. 2003, 2012).

As P. cinnamomea bruises or dries blackish blue, it might also be

hallucinogenic.

Closely related to P. cinnamomea are P. subcaerulipes Hongo (from Japan),

which differs by its distinctly subumbonate brown to orangish brown pileus,

gray-brown to dark violaceous brown lamellae, longer basidia (15-25 x

4.5-6 um), and irregularly branched pleurocystidia (Guzman et al. 2013);

P. zapotecorum (from Mexico), which has larger basidiomata, whitish-

brown to dark violaceous lamellae, and larger basidia (Guzman 2012); and

P. zapotecoantillarum (from Puerto Rico), which is easily distinguished by light

brown to dark reddish-brown adnexed lamellae, longer basidia, and smaller

ventricose pleurocystidia (Guzman et al. 2003).

Psilocybe antioquiensis, described from Colombia, is close to P. cinnamomea

in phylogenetic analyses but is distinguished by its reddish brown to fulvous

pileus, gray brownish or violaceous brown lamellae, bigger angular spores

(6-10.6 x 4.5-7 um), and longer basidia (20-30 x 6-8 um; Guzman et al. 1994).

Morphologically, the new species is similar to the Caribbean species

Psilocybe guilartensis Guzman et al. and the Mexican species P moseri Guzman.

However, P. guilartensis (placed in P. sect. Brunneocystidiatae) has thick-walled

rhomboid spores and longer basidia (20-32 x 4-7 um; Guzman et al. 2003),

while P moseri has a dark buff to brownish pileus, brownish lamellae, and

smaller spores (4-6.5 x 3-3.5 um) and pleurocystidia (12-17 x 4-5.5 um;

Guzman 1995).

Psilocybe cinnamomea sp. nov. (China) ... 221

Acknowledgments

We are grateful to Dr. Dewei Li for improving the manuscript and checking the new

name. Thanks are also given to Keliang Zhang and Lin Li for the language polishing

and Duo Wang for collecting specimens. We are greatly indebted to Dr. Taihui Li and

Dr. Baokai Cui for their critical reviews of the manuscript. This study was supported

by the National Nonprofit Institute Research Grant of CAF (CAFYBB2011004) and the

National Natural Science Foundation of China (No. 31070014).

Literature cited

Allen JW, Gartz J, Guzman G. 1992. Index to the botanical identification and chemical analysis of

the known species of the hallucinogenic fungi. Integration 2: 91-97.

Bau T, Sarentoya. 2009. Strophariaceae of China (IV) Psilocybe. Journal of Fungal Research 1: 14-36.

Borovicka J, Noordeloos ME, Gryndler M, Obornik M. 2011. Molecular phylogeny of Psilocybe

cyanescens complex in Europe, with reference to the position of the secotioid Weraroa novae-

zelandiae. Mycological Progress 10: 149-155. http://dx.doi.org/10.1007/s11557-010-0684-3

Guzman G. 1983. The genus Psilocybe. Beihefte Nova Hedwigia 74. Cramer, Vaduz.

Guzman G. 1995. Supplement to the monograph of the genus Psilocybe. Petrini O, E Horak,

Taxonomic monographs of Agaricales. Bibliotheca Mycologica 159: 91-141.

Guzman G. 2005. Species diversity of the genus Psilocybe in the world mycobiota, with special

attention to hallucinogenic properties. International Journal of Medicinal Mushrooms 7:

305-331. http://dx.doi.org/10.1615/IntJMedMushr.v7.i12.280

Guzman G. 2009. The hallucinogenic mushrooms: diversity, traditions, use and abuse with special

reference to the genus Psilocybe. 256-277, in: Jk Misra, SK Deshmukh (eds). Fungi from

different environments, Science Publishers, Enfield.

Guzman G. 2012. New taxonomical and ethnomycological observations on Psilocybe s.s. (Fungi,

Basidiomycota, Agaricomycetidae, Agaricales, Strophariaceae) from Mexico, Africa and Spain.

Acta Botanica Mexicana 100: 79-106.

Guzman G, Yang ZL. 2010. A new species of a bluing Psilocybe from Asia (Basidiomycota, Agaricales,

Strophariaceae). Sydowia 62: 185-189.

Guzman G, Saldarriaga Y, Pineda F, Garcia G, Velazquez LE. 1994. New species of Psilocybe from

Colombia and discussion on the known species. Mycotaxon 51: 225-235.

Guzman G, Tapia F, Ramirez-Guillén FE, Baroni TJ, Lodge DJ, Cantrell SA, Nieves-Rivera AM. 2003.

New species of Psilocybe in the Caribbean, with an emendation of P guilartensis. Mycologia

95: 1171-1180. http://dx.doi.org/10.2307/3761918

Guzman G, Ramirez-Guillén F, Hyde KD, Karunarathna SC. 2012. Psilocybe s. s. in Thailand:

four new species and a review of previously recorded species. Mycotaxon 119: 65-81.

http://dx.doi.org/10.5248/119.65

Guzman G, Cortes-Perez A, Ramirez-Guillén F. 2013. The Japanese hallucinogenic mushrooms

Psilocybe and a new synonym of P. subcaerulipes with three Asiatic species belong to section

Zapotecorum (higher basidiomycetes). International Journal of Medicinal Mushrooms

15: 607-615. http://dx.doi.org/10.1615/IntJ MedMushr.v15.i6.90

Huelsenbeck JP, Ronquist F. 2005. Bayesian analysis of molecular evolution using MrBayes. In:

27 Nielsen R (ed) Statistical methods in molecular evolution. Springer, New York, 183-232.

Kornerup A, Wanscher JH. 1981. Taschenlexikon der Farben. 3. Aufl. Muster-Schmidt Verlag,

Gottingen.

222 .. Li, Yuan, & Liang

Ma T, Feng Y, Lin XE Karunarathna SC, Ding WE, Hyde KD. 2014. Psilocybe chuxiongensis, a new

bluing species from subtropical China. Phytotaxa, 156: 211-220.

http://dx.doi.org/10.11646/phytotaxa.156.4.3

Norvell LL. 2010. Report of nomenclature committee for Fungi 15. Taxon 59: 291-293.

Redhead SA, Moncalvo JM, Vilgalys RE, Matheny PB, Guzman-Davalos L, Guzman G. 2007.

Proposal to conserve to the name Psilocybe (Basidiomycota) with a conserved type (1757).

Taxon 56: 255-257.

Stamatakis A, Hoover P, Rougemont J. 2008. A rapid bootstrap algorithm for the RAxML Web

servers. Systematic biology, 57(5): 758-771. http://dx.doi.org/10.1080/10635 150802429642

Stamets P. 1996. Psilocybin mushrooms of the world: an identification guide. Berkeley.

Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG. 1997. The CLUSTAL_X windows

interface: flexible strategies for multiple sequence alignment aided by quality analysis tools.

Nucleic Acids Research 25: 4876-4882. http://dx.doi.org/10.1093/nar/25.24.4876

Vellinga E. 1988. Glossary. 54-64, in: C Bas et al. (eds). Flora Agaricina Neerlandica. Vol. 1. A.A.

Balkema, Rotterdam.

White TJ, Bruns T, Lee S, Taylor J. 1990. Amplification and direct sequencing of fungal ribosomal

RNA genes for phylogenetics. 315-322, in: MA Innis et al. (eds). PCR protocols: a guide to

methods and applications. Academic Press, New York.

Zhou LL, Liang JF. 2011. An improved protocol for extraction of DNA from macrofungi.

Guangdong Forestry Science and Technology 27: 13-16.

MY COTAXON

http://dx.doi.org/10.5248/129.223

Volume 129(2), pp. 223-228 October-December 2014

Powdery mildew on Bougainvillea spectabilis in Mexico

with a cryptic generic affinity

SYLVIA P. FERNANDEZ-PAViA!, GERARDO RODRIGUEZ-ALVARADO!,

JESUS GERONIMO-MAGANA’, MARIA GRACIELA CABRERA’,

NuRIA GOMEZ-DORANTES’, & UWE BRAUN?

‘Instituto de Investigaciones Agropecuarias y Forestales, Universidad Michoacana de San Nicolas

de Hidalgo, Km. 9.5 Carr. Morelia-Zinapécuaro, Tarimbaro, Michoacan 58880, México

*Universidad Nacional del Nordeste, Facultad de Ciencias Agrarias, Catedra de Fitopatologia,

Sargento Cabral N° 2131, Corrientes 3400, Argentina

3Martin-Luther-Universitdt, Institut fiir Biologie, Bereich Gebotanik und Botanischer Garten,

Herbarium, Neuwerk 21, 06099 Halle (Saale), Germany

“CORRESPONDENCE TO: fpavia@umich.mx, fernandezpavia@hotmail.com

ABSTRACT —An asexual powdery mildew on Bougainvillea spectabilis resembles Ovulariopsis

(anamorphic Phyllactinia and Pleochaeta), while its combination of sinuous to coiled

conidiophores forming dimorphic conidia singly suggests a Pleochaeta anamorph. However,

this severely contradicts the rDNA ITS molecular data. The Bougainvillea powdery mildew

shows the closest genetic similarities (80-83% sequence homologies) with various Leveillula

and Phyllactinia species (Phyllactinieae), whereas Pleochaeta species were not genetically

close. These results suggest that the Bougainvillea powdery mildew has an affinity with

tribe Phyllactinieae of the Erysiphaceae, but the percentage of genetic similarity is below the

scope of generic limits in powdery mildew ITS data. Although a new genus may be involved,

we cannot formally propose it based on a single sequence, an anamorph morphologically

indistinguishable from Ovulariopsis, and an unknown teleomorph.

Key worps— Erysiphales, ornamental plant, new species, Michoacan

Introduction

Bougainvillea is a widely used ornamental plant in Mexico, with Morelos

state one of the main producers in the country. In Michoacan it is the most

common ornamental used in gardens, avenues, and parks. The flowers are

also used for medicinal purposes, mainly as remedy for respiratory illnesses.

The powdery mildew Leveillula taurica (Lév.) G. Arnaud has been associated

with Bougainvillea in Mexico (Ramirez-Rojas et al. 2010), but its identification

224 ... Fernandez-Pavia & al.

has not yet been confirmed by molecular methods. The objective of this study

was to determine the identity of the pathogen causing powdery mildew on

100-250 cm tall flowering Bougainvillea spectabilis plants collected during

July and September 2009 in two urban municipalities in Michoacan, México.

Some Bougainvillea plants showed chlorotic lesions on both sides of the leaves.

Lesions were small (3-6 mm diameter), irregularly shaped, and with a central

necrotic area. Most mature leaves of the affected plants showed lesions on

the upper side of the leaves but lacked mycelia and other fungal reproductive

structures. Lesions on the undersides were colonized by sparingly developed

mycelia with few conidiophores and conidia.

Materials & methods

Two hundred forty-six Bougainvillea spectabilis plants were examined for powdery

mildew in Morelia and Tarimbaro, municipalities in Michoacan state. Powdery mildew

colonies on infected plants were prepared by means of a dissecting microscope and

examined by fresh mounts in distilled water (with or without aniline blue) or dry mounts

in 85% lactic acid, with staining and gentle heating (modified from Shin & La 1995

according to technique 5 in Liberato et al. 2005) and using standard light microscopy

with oil immersion (bright field and phase contrast, Olympus BX 50, Hamburg,

Germany). 30 measurements (1000x magnification) of conidia and other structures

were made. Voucher specimens were conserved in the herbariums at Universidad

Michoacana de San Nicolas de Hidalgo, Morelia, Michoacan, Mexico (EBUM) and

Martin-Luther- Universitat, Halle, Germany (HAL).

Genomic DNA was obtained from powdery mildew infected leaves from samples

collected in gardens. Standard protocols followed instructions of the manufacturer

(Wizard Genomic DNA Purification Kit, Promega Corp., Madison, Wisconsin). The

internal transcribed spacer (ITS) region of nuclear ribosomal DNA, including 5.8S

rDNA, the 3’ end of 18S rDNA and the 5’ end of 28S rDNA, was amplified from

powdery mildew according to Cunnington et al. (2003). PCR reactions were carried

out using HotStarTaq DNA polymerase (HotStarTaq Master Mix Kit, Qiagen, Valencia,

CA) in a thermal cycler (Mastercycler Gradient, Eppendorf). The PCR DNA products

were cleaned using a commercial kit following the manufacturer's protocol (Wizard SV

Gel and PCR Clean-up System (Promega, Corp., Madison, Wisconsin). PCR products

were visualized by electrophoresis in 1.5% agarose gels in TAE buffer. Direct sequencing

of the PCR products was carried out by an external company (Macrogen Inc. Seoul,

Korea). Programs Pregap and Gap in the Staden Package (http://staden.sourceforge.

net) were utilized to obtain consensus sequences for the amplified region from each

isolate. Sequences were deposited in GenBank, 626 bp as Ovulariopsis sp. GRA-2013

(Accession no. KC556804). The sequences concerned were used in the BLASTN 2.2.25+

program (NCBI http://www.ncbi.nlm.nih.gov/) to determine the identity or at least

the affinity based on sequence similarities expressed as percent sequence identity.

A restricted phylogenetic tree was prepared, using the program Mega5 by neighbor-

joining method with Tajima-Nei distance calculation to demonstrate relationships to

the closest relatives.

Cryptic powdery mildew on Bougainvillea (Mexico) ... 225

Figure 1. Bougainvillea powdery mildew.

A: Hypha with appressorium; B: Conidiophores; C: Conidium; D: Conidium with germ tube.

Based on type material [HAL 2588 F]. Bar = 10 um. U. Braun del.

Taxonomy

Bougainvillea powdery mildew Fic. 1

Mycelium on leaves, hemiendophytic, superficial mycelium mostly

hypophyllous, with hyphae emerging through stomata. Hyphae hyaline,

septate, thin-walled, 2-6 um wide, frequently with approximately right angled

226 ... Fernandez-Pavia & al.

branching. Hyphal appressoria numerous, distinct, oblong, hooked, sinuous

and/or coral-like. Conidiophores consistently arising from the upper surfaces

of superficial hyphae (conidiophores emerging through stomata not observed),

and more or less midway between two septa, cylindrical, straight, unbranched,

140-252.5 (average 180) um long and 7.5-9 um wide, hyaline, thin-walled,

smooth; foot-cells very long, cylindrical, but sinuous to somewhat coiled at the

very base just above the basal septum that is slightly elevated above the junction

with the mother cell, foot-cells followed by 1-2 much shorter cells. Conidia

formed singly, one-celled, dimorphic, primary conidia lanceolate to ovoid-

lanceolate, distinctly attenuated towards the apex, 62.5-87.5 (average 77.65) x

12.5-27.5 (average 20.85) um, secondary conidia ellipsoid-subcylindrical, ends

subtruncate, 55-77.5 x 7.5-15 um, hyaline, with numerous oil-droplets, germ

tubes arising near base or apex, single, short, producing hooked and/or coral-

like appressoria.

MATERIAL EXAMINED: MEXICO, MicHoacan, Morelia municipality, urban

garden, on leaves of Bougainvillea spectabilis Willd. (Nyctaginaceae), September 2009,

S. Fernandez-Pavia (HAL 2588 F); September 2009, S. Fernandez-Pavia (EBUM 23687,

GenBank KC556804); Tarimbaro municipality, urban garden, on leaves of B. spectabilis,

July 2009, S. Fernandez-Pavia (EBUM 24362).

Discussion

The incidence of powdery mildew on Bougainvillea was very low: only four

plants, or 1.6%, were infected. This is an atypical powdery mildew because all

structures are only sparingly developed and rather inconspicuous, perhaps one

reason that this fungus has been overlooked hitherto.

A blast search based on the sequence retrieved from our Bougainvillea

powdery mildew (KC556804) did not yield any hits and showed only a distant

similarity (80-83%) with other sequences representing Leveillula G. Arnaud

and Phyllactinia Lév. These homologies are listed below:

83%: Leveillula chrozophorae U. Braun (AB045147, Chrozophora tinctoria, Iran),

L. duriaei (Lév.) U. Braun (AB667859, Phlomis persica, Iran), and L. taurica s. lat.

(AB667864, Carthamus sp., Iran).

82%: L. duriaei (AB044373, Salvia nemorosa, Iran), L. elaeagni (Jacz.) Simonyan

& Heluta (AB042642, Elaeagnus orientalis, Turkmenistan), L. lactucarum

Durrieu & Rostam (AB044375, Lactuca serriola, Iran), L. lanuginosa (Fuckel)

Golovin (AB044376, AB045153, Echinophora sibthorpiana and Heracleum

persicum, Iran), L. loranthi Haajian et al. (AB044377, Loranthus europaeus,

Iran), L. thevenotiae (Jacz.) Golovin (AB044383, Thevenotia persica, Iran) as

well as several sequences deposited as L. taurica s. lat. (AB045108, 667868,

667874, 667882, diverse hosts, Iran; AY912077, Triglochin maritima, origin

unclear; GQ860947, Mentzelia laevicaulis, origin unclear; JQ885445, Capsicum

sp., South Korea).

Cryptic powdery mildew on Bougainvillea (Mexico) ... 227

gg , AB022403 Pleochaeta shiraiana

AB243757 Pleochaeta indica

AB218773 Pleochaeta turbinata

AB022387 Leveillula taurica

AB022401 Phyllactinia moricola

AB022372 Phyllactinia kakicola

KC556804 GRA-2013 (Bougainvillea)

Sal

0.1

FIGURE 2. Bougainvillea powdery mildew. Restricted phylogenetic neighbor-joining tree, prepared

by using the program Mega5 with Tajima-Nei distance calculation to demonstrate relationships of

this powdery mildew to its closest relatives.

81%: Phyllactinia fraxini (DC.) Fuss (AB080513, Fraxinus longicuspis, Japan;

AB080553, F. excelsior, Lithuania; AB080543, Syringa vulgaris, Germany).

80%: P. roboris (Gachet) S. Blumer (AB080516, Castanea crenata, Japan),

Phyllactinia sp. (AB080546, Castanopsis sp., China; AB080564, Chionanthus

virginicus, Germany).

No sequence agreeing with that of our Bougainvillea powdery mildew has yet

been deposited in GenBank; our comparisons indicate that within powdery

mildews the closest affinity is with species of the tribe Phyllactinieae R.T.A.

Cook et al. Based on its most significant morphological characters —

conidiophores that arise exclusively from superficial hyphae and with sinuous

to somewhat coiled foot-cells — this fungus might be assigned to Ovulariopsis

Pat. & Har., the anamorph of Phyllactinia and Pleochaeta Sacc. & Speg., as

currently circumscribed by Braun & Cook (2012). The formation of dimorphic

conidia rather suggest an asexual morph of Pleochaeta previously referred to

as Streptopodium R.Y. Zheng & G.Q Chen, a genus originally introduced for

Ovulariopsis-like powdery mildew anamorphs characterized by conidiophores

with twisted foot-cells and associated with sexual morphs belonging to

Phyllactinia and Pleochaeta.

Braun & Cook (2012) discussed asexual morphs of Phyllactinia and Pleochaeta

in detail. Combinations of conidiophores with coiled foot-cells and formation

of dimorphic conidia are not confined to Pleochaeta anamorphs, but are also

known in various asexual morphs of Phyllactinia species, e.g., P. bauhiniae

Y.S. Paul, P. cassiae G.J.M. Gorter & Eicker, and P. chorisiae Viégas. There is no

real morphological basis for maintaining Streptopodium as a separate genus,

and Braun & Cook (2012) reduced it to synonymy with Ovulariopsis, emending

the circumscription of that genus. In addition, various sequences retrieved from

228 ... Fernandez-Pavia & al.

Pleochaeta species were not among those with highest homology (Fic. 2), in

that Pleochaeta is more distantly related to the Bougainvillea powdery mildew

than are Phyllactinia and Leveillula. The formation of dimorphic conidia and

the conidial shape agrees well with species of Oidiopsis Scalia (anamorphs of

Leveillula) but in Leveillula conidiophores emerge exclusively (or at least partly)

through stomata and twisted conidiophore foot-cells are unknown.

In conclusion, we believe that the Bougainvillea powdery mildew

undoubtedly represents an undescribed species related to genera of tribe

Phyllactinieae. This powdery mildew is morphologically indistinguishable from

the Ovulariopsis anamorphs of Phyllactinia and Pleochaeta, but it cannot be

assigned to Ovulariopsis due to a conflicting molecular sequence analysis. This

analysis confirms a distant relationship to Leveillula, Phyllactinia, Pleochaeta,

and Queirozia Viégas & Cardoso but excludes it from being congeneric with

any of these genera. The introduction of a new anamorph genus, based on

molecular data of a single taxon and indistinguishable morphologically from

Ovulariopsis, is not justified, as it would be of little help in clarifying the

taxonomy of this powdery mildew. In the absence of an appropriate genus, it

is currently not possible to provide a formal description of the Bougainvillea

powdery mildew species. The discovery of the teleomorph of this mildew

or additional genetically closely allied taxa would be useful, and so further

research is necessary.

Acknowledgments

We are much obliged to Professor Xavier Madrigal Sanchez of Universidad

Michoacana de San Nicolas de Hidalgo for support in the identification of the host

plant, and we are thankful to the two reviewers, R.T.A. Cook (U.K.) and T.Z. Liu (Inner

Mongolia, China), for critical comment.

Literature cited

Braun U, Cook RTA. 2012. Taxonomic manual of the Erysiphales (powdery mildews). CBS

Biodiversity Series 11. 707 p.

Cunnington HJ, Takamatsu S, Lawrie CA, Pascoe IG. 2003. Molecular identification of

anamorphic powdery mildews (Erysiphales). Australasian Plant Pathology 32: 21-428.

http://dx.doi.org/10.1071/AP03038

Liberato JR, Barreto RW, Shivas RG. 2005. Leaf clearing and staining techniques for the observation

of conidiophores in the Phyllactinoideae (Erysiphaceae). Australasian Plant Pathology

34: 401-404.

Ramirez-Rojas S, Garcia-Pérez F, Osuna-Canizalez F. 2010. Patégenos asociados al cultivo de

bugambilia (Bouganvillea spp. Willd.) en el estado de Morelos. XII Congreso Internacional/

XXXVII Congreso Nacional de la Sociedad Mexicana de Fitopatologia, Mérida, Yucatan,

México, July 4-8, 2010.

Shin H-D, La YJ, 1993. Morphology of edge lines of chained immature conidia on conidiophores in

powdery mildew fungi and their taxonomic significance. Mycotaxon 46; 445-451.

MY COTAXON

http://dx.doi.org/10.5248/129.229

Volume 129(2), pp. 229-232 October-December 2014

Pseudocercospora styracigena sp. nov.

on Styrax and Alniphyllum from China

Bao-Ju Li’, A-L1 CHAI’ & YING-LAN GUO”**

‘Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences,

Beijing 100081, P.R. China.

*State Key Laboratory of Mycology, Institute of Microbiology, Chinese Academy of Sciences,

Beijing 100101, PR. China

* CORRESPONDENCE TO: guoyl@im.ac.cn

ABSTRACT — A new species, Pseudocercospora styracigena, is described, illustrated, and

discussed. The type specimen is deposited in HMAS.

KEY worps — cercosporoid fungi, plant pathogen, Styrax tonkinensis

Introduction

Pseudocercospora was established in 1910 by Spegazzini. With more than 1500

described species, it is the largest genus of cercosporoid fungi and is distributed

predominantly in tropical and subtropical regions. Pseudocercospora species

are plant pathogens, causing leaf and fruit spot diseases, blights, and fruit rots

(Chupp 1954, Deighton 1976, von Arx 1983, Pons & Sutton 1988), and some

species are used for biological control purposes against weeds (Den Breeyen et

al. 2006). Therefore, Pseudocercospora species are an economically significant

fungal group. Pseudocercospora styracigena, a new species on leaves of Styrax

tonkinensis collected from Yunnan Province, China, is described, illustrated,

and compared with other cercosporoid species on Styracaceae.

Taxonomy

Pseudocercospora styracigena Y.L. Guo & B,J. Li, sp. nov. FIG. 1

MycoBank MB 808331

Differs from Pseudocercospora fukuokaensis by its longer and wider conidiophores,

wider conidia, and developed external hyphae.

Type — China, Yunnan Province, Jinghong, on living leaves of Styrax tonkinensis

(Pierre) Craib ex Hartwich (Styracaceae), August 1973, coll. Liu Xi-jin (HMAS 62748,

holotype).

230 ... Li, Chai, & Guo

FiGuRE 1. Pseudocercospora styracigena (HMAS 62748, holotype).

a. Conidia; b. Conidiophores; c. Stroma; d. External hypha. Bar = 40 um.

Erymo oey: Derived from the host genus Styrax.

LEAF spots amphigenous, circular to irregular, 0.5-8 mm diam., often confluent,

at first gray to grayish brown, later center olivaceous-brown to dark brown,

surrounded by a brown to dark brown line on the upper surface, grayish to pale

brown on the lower surface. CAESPITULI amphigenous. PRIMARY mycelium

internal; secondary mycelium external, hyphae pale olivaceous to olivaceous,

branched, septate, smooth, 2-3.5 um wide. Srromata substomatal, globose,

brown to dark brown, 20-80 um diam. CoNIDIOPHORES densely fasciculate

or solitary, arising from the external hyphae as lateral branches, olivaceous

to moderate olivaceous-brown, uniform in color, irregular in width, straight

to curved, branched, geniculate, denticulate, rounded to conical at the

Pseudocercospora styracigena sp. nov. (China) ... 231

apex, 0-6-septate, sometimes constricted at the septa, 10-110 x 3-5.3 um.

CONIDIOGENOUS LOCI inconspicuous. Conip1A cylindrical to cylindrical-

obclavate, pale olivaceous to olivaceous, straight to curved, obtuse at the apex,

obconically truncate at the base, 2.4—4 um wide, 3-9-septate, 30-95 x 2.5-5um.

ADDITIONAL SPECIMENS EXAMINED — CHINA, HUNAN PROVINCE, Zhangjiajie, on

living leaves of Alniphyllum sp. (Styracaceae), 14 September 1987, coll. Zhao Wen-xia,

no. 51 (HMAS 62127, as Pseudocercospora styracis); YUNNAN PROVINCE, Jinghong, on

living leaves of Styrax tonkinensis, 16 July 1976, coll. Han Shu-jin, Zheng Ru-yong, no.

331 (HMAS 62747).

Discussion

Zhao & Guo (1993) previously determined the specimen of Pseudocercospora

styracigena on Alniphyllum sp. from Hunan (HMAS 62127) as Pseudocercospora

styracis (Chupp) Y.L. Guo & W.X. Zhao [=Passalora styracis], but Crous &

Braun (2003: 392) stated that this was a misdetermination of an undescribed

species.

Four cercosporoid species have been reported previously on Styracaceae:

Cercospora styracicola X.J. Liu & Y.L. Guo on Styrax faberi Perkins (Liu &

Guo 1984); Passalora halesiicola U. Braun et al. [=“P. halesiae” nom. inval.] on

Halesia spp. (Braun et al. 2009, Crous & Braun 2003, Dearness 1941); Passalora

styracis (Chupp) U. Braun & Crous on Styrax americana (Chupp 1954, Crous &

Braun 2003); and Pseudocercospora fukuokaensis (Chupp) X.J. Liu & Y.L. Guo

on Styrax spp. (Chupp 1954, Crous & Braun 2003, Guo & Liu 1992, Togashi &

Katsuki 1952). Cercospora styracicola, Passalora halesiicola, and P. styracis differ

from Pseudocercospora styracigena by having different generic characteristics.

Pseudocercospora fukuokaensis differs from P. styracigena by its shorter and

narrower conidiophores (5-30 x 2.5-4 um) and narrower conidia (30-70 x

2-3.5um) as well as in lacking external hyphae (Togashi & Katsuki 1952, Chupp

1954).

Acknowledgments

This work was supported by the Ministry of Science and Technology of China

(No.2013FY110400), CAAS Agricultural Science and Technology Innovation Program

and funded by Key Laboratory of Biology and Genetic Improvement of Horticultural

Crops, Ministry of Agriculture, China. We are grateful to U. Braun and S.Y. Guo for

reviewing the manuscript.

Literature cited

Arx JA von 1983. Mycosphaerella and its anamorphs. Proceedings of the Koninklijke Nederlandse

Akademie van Wetenschappen, Series C, 86: 15-54.

Braun U, Farr DF, Minnis AM. 2009. Nomenclatural notes on some cercosporoid hyphomycetes.

Schlechtendalia 19: 81-84.

Chupp C. 1954. A monograph of the fungus genus Cercospora. Ithaca, New York

232 ... Li, Chai, & Guo

Crous PW, Braun U. 2003. Mycosphaerella and its anamorphs: 1. Names published in Cercospora

and Passalora. CBS Biodiversity Series 1. 571 p.

Dearness J. 1941. New species of Tennessee fungi. Mycologia 33: 360-366.

http://dx.doi.org/10.2307/3754889

Deighton FC. 1976. Studies in Cercospora and allied genera VI. Pseudocercospora Speg., Pantospora

Cif. and Cercoseptoria Petr. Mycological Papers 140. 168 p.

Den Breeyen A, Groenewald JZ, Verkley GJM. 2006. Morphological and molecular characterisation

of Mycosphaerellaceae with the invasive weed, Chromolaena odorata. Fungal Diversity 23:

89-110.

Guo YL, Liu XJ. 1992. Studies on the genus Pseudocercospora in China VI. Mycosystema 5: 99-108.

Liu XJ, Guo YL. 1984. Five new species of Cercospora. Acta Mycological Sinica 3(2): 102-108.

Pons N, Sutton BC. 1988. Cercospora and similar fungi on yams (Dioscorea species) Mycological

Papers 160. 78 p.

Togashi K, Katsuki S. 1952. New or noteworthy Cercospora from Japan. Botanical Magazine Tokyo

65: 18-26. http://dx.doi.org/10.15281/jplantres1887.65.18

Zhao WX, Guo YL. 1993. Studies on hyphomycetes of Zhangjiajie in Hunan II. Cercospora and

Pseudocercospora. Acta Mycologica Sinica 12(3): 193-199.

MY COTAXON

http://dx.doi.org/10.5248/129.233

Volume 129(2), pp. 233-245 October-December 2014

Neozygites species associated with aphids in Chile:

current status and new reports

CRISTIAN MONTALVA »?*, MAREK BARTA 3, ELADIO ROJAS 4,

MONICA GUTIERREZ *, & EDUARDO VALENZUELA °

‘Facultad de Ciencias Forestales y Recursos Naturales, Escuela de Graduados,

Casilla 567, Universidad Austral de Chile, Valdivia, Chile

?Instituto de Patologia Tropical e Saude Publica, Universidade Federal de Goias,

Goidania, Brazil

°Mlynany Arboretum, Institute of Forest Ecology SAS,

Vieska nad Zitavou 178, Slepéany 95152, Slovakia

‘Laboratorio Regional, Servicio Agricola y Ganadero,

Ruta a Puerto Octay U-55-V, Calle de Servicio, Osorno, Chile

°Facultad de Ciencias, Instituto de Bioquimica y Microbiologia,

Casilla 167, Universidad Austral de Chile, Valdivia, Chile

* CORRESPONDENCE TO: montalva.cristian@gmail.com

AxBstTRAcT —Three species of Neozygites were recorded during a 2007-13 survey of the

occurrence of the genus on aphids in Chile. Neozygites osornensis is known from recent

studies, and N. fresenii and N. cinarae are reported as new records for Chile. Neozygites

lageniformis, which was not found during the survey, had been recorded previously in Chile.

Morphological descriptions, symptoms on infected insects, host spectrum, and geographical

distribution of all four species are presented, and a key to Neozygites species associated with

aphids in Chile is included. These fungi, which are important natural enemies of aphids, may

be considered for future aphid pest control.

KEY worpDs — entomopathogenic fungi, aphid pathogens, Entomophthoromycota

Introduction

Insect-pathogenic fungi represent a phylogenetically heterogeneous group

of microorganisms. However, a majority of the economically important species

belongs to the phyla Ascomycota (order Hypocreales) and Entomophthoromycota.

In general, these fungi are considered excellent candidates for the biological

control of various insect pests (Latgé & Papierok 1988, Charnley & Collins 2007,

Roy et al. 2010). The hypocrealean species are well studied and some of them are

234 ... Montalva & al.

commercially used with success in various strategies of biological control (Shah

& Pell 2003, Sandhu et al. 2012). While species in the Entomophthoromycota have

been used successfully in the laboratory, they have not yet been commercially

formulated into an applicable mycoinsecticide (Barta & Cagan 2006a), although

they have been introduced and spread successfully as an inoculum (Milner et

al. 1982, Elkinton et al. 1991). Conservation biological control has been used

for other fungi in Entomophthoromycota, and there have been attempts in the

United States of America to suppress mite populations in soybean crops with

fungal pathogens rather than insecticide treatments (Klubertanz et al. 1991).

Neozygites (Neozygitales, Neozygitaceae) (Humber 2012) comprises a

relatively homogenous group of 18 species exclusively pathogenic to arthropods

(Acari, Hemiptera, and Thysanoptera) (Delalibera et al. 1992, Montserrat et al.

1998, Shah & Pell 2003, Keller & Petrini 2005, Keller 2007, Wekesa et al. 2010).

Neozygites species are important natural enemies that may control arthropod

populations under microclimatic conditions favorable for the fungus (Latgé

& Papierok 1988; Barta & Cagan 2003, 2006a; Klingen & Westrum 2007;

Castro et al. 2013). Generally, most Neozygites species can be divided into two

subgroups (Neozygites fresenii-type or Neozygites floridana-type) based on

their morphologies and life cycles (Keller 1997, 2006; Delalibera et al. 2004).

Their narrow host specificity makes them valuable control agents that present a

negligible threat to non-target organisms (Barta 2004). Although the genus has

a worldwide distribution, some Neozygites species are known only from their

type localities. As there is limited literature about the genus in Chile (Aruta et

al. 1984; Aruta & Carrillo 1989; Montalva et al. 2010, 2013), our goal was to

review and update knowledge on Neozygites species associated with aphids in

Chile.

Materials & methods

The literature was searched for publications on Neozygites associated with

aphids in South America with an emphasis on Chile. During 2007-13, aphid

populations were observed for Neozygites in 25 localities of southern Chile

(36°49’41.5”-53°10’00’S 70°56’00”-73°03'04.93”W).

Dead aphids were collected in the field. Black cadavers filled with internally

produced resting spores were either examined immediately after being taken

to the laboratory or stored at 4 °C until examination. Brown cadavers filled

with vegetative cells and hyphal bodies were incubated at 25 °C in moist

chambers (plastic containers and moistened filter paper) to induce external

conidiogenesis. Neozygites species were identified based on morphology

according to Keller (1991, 1997).

Mounts were prepared in lactophenol-aceto-orcein (LPAO) as described

by Keller (1987). Fungal structures were examined with a Nikon Eclipse

Neozygites species associated with aphids (Chile) ... 235

E600 microscope, photographed with a Nikon DS-Fil digital camera at a

magnification of 1500x, and measured with Motic Images Plus 2.0 software.

At least four series of measurements were taken for taxonomically important

microstructures. Each series represented 50 measurements, and for each series

a mean value was calculated. A final value of each dimension consisted of two

ranges: the range of mean values of all series and the range of extreme values of

all measurements (in parentheses).

Vouchers of the newly recorded species were conserved in Laboratorio

Regional, Servicio Agricola y Ganadero, Osorno, Chile (LA-N).

Results

Current status of aphid-pathogenic Neozygites species in Chile

Neozygites lageniformis was not found during this survey but has been

recorded previously in Chile. Two of the three aphid-pathogenic Neozygites

species observed during the survey are presented as first records from this

country. TABLE 1 provides morphometric characteristics of all Neozygites

species recorded in Chile. Detailed data on each species are presented below.

Taxonomic descriptions

Neozygites cinarae S. Keller, Sydowia 49: 137. 1997. PLATE 1

Hyphal bodies not observed. Primary conidia ovoid to pyriform, formed

apically on unbranched conidiophores, with papilla distinct, truncate or

slightly rounded, 25.7-26.8 x 18.4-19.5 um. Forcibly discharged secondary

conidia were only rarely observed; capilliconidia are short almond-shaped with

a terminal mucoid droplet, brownish, produced apically on slender capillary,

26.0-30.3 x 11.5-12.8 um. Zygospores ellipsoidal, black, with smooth surface,

33.8-35.3 x 24.3-24.7 um.

SPECIMEN EXAMINED: on Cinara cedri Mimeur: CHILE, Valdivia, 39°48’18"S

73°15'10”W, March 2013, by C. Montalva (LA-N15032013).

ECOLOGY & OCCURRENCE: Infected aphids were found fixed by their probosces

on the underside of branches of Cedrus atlantica (Endl.) Manetti ex Carriére.

Almost all aphids (> 90%) sampled from March to May 2013 were infected and

brown or black when filled with resting spores. This is the first record for Chile.

Neozygites fresenii (Nowak.) Remaud. & S. Keller, Mycotaxon 11: 332. 1980. PLATE 2

Hyphal bodies spherical to subspherical, 13.0-13.9 um in diameter.

Conidiophores unbranched. Primary conidia spherical, 21.0-21.7 x 15.9-16.3

um; papilla distinct, cylindrical, blunt to slightly rounded. Secondary conidia

(if discharged) like primary conidia on short unbranched conidiophores;

capilliconidia passively dispersed, almond-shaped, apical on slender

236 ... Montalva & al.

PLATE 1. Neozygites cinarae on Cinara cedri. a. Conidiophore forming primary conidium.

b. Primary conidium. c. Formation of secondary conidium. d. Capilliconidium. e. Resting spore.

f. Cinara cedri infected with Neozygites cinarae. Bars = 25 um.

ws — Sk

y and

PLATE 2. Neozygites fresenii on Myzocallis coryli. a. Hyphal body with visible nuclei. b. Unbranched

conidiophores. c. Primary conidium. d. Capilliconidium. e. Formation of capilliconidia. f. Resting

spore. g. Myzocallis coryli infected with Neozygites fresenii. Bars = 20 um.

Neozygites species associated with aphids (Chile) ... 237

PLATE 3. Neozygites osornensis on Cinara cupressi. a. Subspherical hyphal bodies with visible nuclei.

b. Conidiophore forming primary conidium. c. Primary conidia attached to aphid leg. d. Primary

conidium. e. Crescent-shaped capilliconidium with visible nuclei. f. Zygospore with visible nuclei

developing by conjugation of two hyphal bodies containing degenerating nuclei. g. Immature

zygospore. h. Fully developed zygospore. i. Young germ capilliconidium produced by zygospore on

long capillary germ conidiophore. Bars = 20 um.

capilliconidiophores, 22.8-24.3 x 12.8-13.2 um. Zygospores broadly ellipsoidal,

thick-walled, episporium black, smooth, 26.1-28.4 x 18.7-20.4 um.

SPECIMENS EXAMINED: on Myzocallis coryli (Goetze): CHILE, Puyehue, 40°38’19"S

72°40'55"W, 2012, by C. Montalva (LA-N25032012).

ECOLOGY & OCCURRENCE: Infected aphids fixed by their probosces and/or

legs were found on the underside of branches of the common hazel (Corylus

avellana L.); grayish brown to brick or dark brown when conidia were formed,

and black when resting spores were present. About 30% of the aphids sampled

from February to April 2012 were infected. This is the first record for Chile.

Neozygites osornensis Montalva & Barta, Mycologia 105: 663. 2013. PLATE 3

Hyphal bodies spherical or subspherical, 16.4-16.9 um in diameter. Primary

conidia form apically on unbranched conidiophores, ovoid to pyriform, with

papilla truncate or slightly rounded, 20.8-23.8 x 13.6-15.9 um. capilliconidia

apical on long slender capillary conidiophores, elongate, crescent-shaped,

28.0-28.5 x 8.5-8.8 um. Zygospores ellipsoidal, with episporium smooth, dark-

brown to black, 31.8-40.8 x 21.5-22.7 um; germinating by forming capillary

238 ... Montalva & al.

€LOT Te 39 ATLIUOYY

(Salas F) SS I-ZLF'I

(salqas F) L°77-S'1Z

(saltas F) 8°OF-8'TE

(Salqas F) 9€°€-€7'E

(S9T19S 7) 8°8-S'8

(Saltas #) S°8Z-0'87

(salsas ) 7S I-8P'T

(salJas ) 6'S1-9'€T

(salqas F) 8°€7-8°07

(solJas F) 6°91-F'91

SISUAUAOSO "NT

6861 O[[MTeD 29 eINIV

vyep ON

9'Ee-0'0E

vyep ON

STC-€LT

CCE-V'8T

vyep ON

SIULOfIUasv] ‘N

*‘SJUsMMoINseotw JO Joquinu = U,,

*SOTIOS JO SON[RA IUIII}X9 UPIUT FO S}STSUOD asuel oy} ‘SJUDUIIINSVIUI (SG sjuasaida1 SOTI9S 9UO = Soles ,

Joded styy,

(Salles F) 6€'1

(saltas F) F'07-L’8T

(salqas F) $87-1'97

(Sal1as F) S8'1-6Z'T

(salqas F) Z'€1-8°Z1

(salqas F) €'F7-8°T7

(saldas 7) 9€ I-€€'T

(salqas F) €°91-6'°ST

(saldas F) Z'1Z-0'°1Z

(soltas F) 6€T-O'ET

iuasalf ‘N

Joded styy, QOUIIIZOY

(salqas F) EF I-LE'1

(sallas f) L7-€'b7 (q) sa}0uIeIG

(sal1as 7) €°GE-8'€€ (1) y8ueT

(um) sazods 8unsoy

(solias #) LE°T-€7'T

(soldas F) 8°7I-S'TT (q) Jo}oWIeIG

(saltas F) €°0€-0°9Z (1) y8ueT

(tun) erpruosyidep

(sal1as F) I-F'1

(Salas $) S'61-F'81 (qd) JajoureIG

(,S9Has 7) 8°97-L'ST (J) ysuaT

(um) erpruos Areuntig

eyep ON (um Surerp) satpoq teydAY

IDADUII *NT SHYNLONGLS TVONNA

‘gTIYD ut spryde uo sateds sapyisdzoan JUIIIYIP INoJ Jo somnjons}s Jue}odurt AT[esTuIOUOXe} Jo sisATeuR sIJOUIOYdIOPY ‘Tt ATAV I],

Neozygites species associated with aphids (Chile) ... 239

conidiophores, averaging 72.4 um long (n: 43), with apical germ capilliconidia,

28.0 x 12.4 um (n: 45) (Montalva et al. 2013).

SPECIMENS EXAMINED: on Cinara cupressi: CHILE, Osorno, 40°35’14"S 73°05’38"W,

4 April 2007, by C. Montalva (SGO 161 401, holotype).

ECOLOGY & OCCURRENCE: Infected aphids can be easily recognized hanging on

twigs and by an obvious change of their color during sporulation (turning from

yellowish-brown to reddish-brown). During the infection progress, aphids

gradually became swollen and turgid. Cadavers that were reddish-brown during

the formation of external conidia became black when internal zygospores

developed. Cadavers usually hung and remained with their probosces inserted

in the branches. After certain time the cadavers either dried up and fell down to

the soil or liquefied and disintegrated during zygosporogenesis, thus dispersing

the resting spores on the bark of twigs. More than 80% of aphids sampled from

November to December 2013 in Osorno were infected (Montalva et al. 2013).

Key to aphid-pathogenic Neozygites species from Chile

LoObservedsoneMyzocallidinae or A phidinae oes. 634 ed et esd oe led ed 2

1. Observed on Lachninae or Pterocommatinde .......... ccc cece cence cece neues 3

2. Primary conidia average length <22 um; capilliconidia averaging < 25 um,

iat lye LADD eB tae es tiny ev etna e tillags a> thean Te tte Te thea Te patieas eH een Neozygites fresenii

2. Primary conidia average length =29 um; capilliconidia averaging = 30 um,

Wt lhe LADD 0s ota ic «esac» Wigs s ope es Maga le sagas s Nagas Was Neozygites lageniformis

3. Primary conidia average length 26 um; capilliconidia almond-shaped,

AVEFASeTCTAMIe Cer sll MTL. gto eae hace tee a Or Pe oO at Neozygites cinarae

3. Primary conidia average length 23 um; capilliconidia crescent-shaped,

AVE FASE TAMeter OUT ton ee chan a hee lee doe hae Neozygites osornensis

Discussion

Dead insects and mites killed by different Neozygites species have been

reported from several South American countries (Humber et al. 1981,

Agudelo-Silva 1986, Lohr et al. 1990, Gomez de Picho 1991, Delalibera et al.

1997, 2000, Van der Geest et al. 2002, Scorsetti & Lopez-Lastra 2007, Roggia

et al. 2009). Previously, however, Neozygites species pathogenic to aphids have

been recorded only in Argentina and Chile (Scorsetti et al. 2007, Sosa-Gomez

et al. 2010, Montalva et al. 2013). The first reports from Chile of any Neozygites

species were made in the 1980s, when N. parvispora (D.M. MacLeod & K.P. Carl)

Remaud. & S. Keller (on thrips) and N. lageniformis (on aphids) were observed

infecting natural arthropod populations (Aruta et al. 1984, Aruta & Carrillo

1989). Neozygites parvispora is a widespread fungal parasite of various thrips

species with a primarily European distribution (Balazy 1993, Keller 1997).

Neozygites lageniformis is rare and incompletely described species from the

240 ... Montalva & al.

New World (Thaxter 1888, Aruta & Carrillo 1989), previously observed only

twice (in aphids in United States of America and Chile). During the survey we

did not find this fungus in Myzocallis coryli colonies. Although there are no

complete data available on its morphology, N. lageniformis is still regarded as a

separate species (Balazy 1993). However, until additional records and a more

accurate description become available, its taxonomic status is uncertain.

Neozygites cinarae is a fungal pathogen of aphids in the genus Cinara.

Previously reported from Europe on C. pilicornis (Hartig) and C. piceae

(Panzer) (Keller 1997, Barta et al. 2005, Barta & Cagan 2006b, Montalva et al.

2013), ours is first report for South America. In Chile, N. cinarae was found

only in C. cedri colonies on the host tree, Cedrus atlantica. No data are available

about its use as a biological control in laboratory or field conditions. Its high

incidence during our survey suggests that during autumn N. cinarae could be a

good natural regulator of C. cedri populations along with other natural enemies

such as coccinellid beetles and syrphid flies.

Neozygites fresenii is a widely occurring fungal pathogen of aphids. It has

been reported from Europe (Keller 1991, Batazy 1993, Barta & Cagan 2002),

North and South America (Steinkraus et al. 1995, Scorsetti et al. 2007), South

Africa (Hatting et al. 1999), the South Pacific (Keller 1997), and Australia

(Milner & Holdom 1986). Although the fungus is reported to have a broader

host spectrum, in Chile N. fresenii was found only in Myzocallis coryli colonies.

In Argentina, N. fresenii has also been identified from Brevicoryne brassicae

L., Myzus sp., and Aphis fabae Scop. in Buenos Aires province (Scorsetti et

al. 2007). Neozygites fresenii often causes widespread epizootics in aphid

populations (Steinkraus et al. 1995, Keller 1997, Barta 2004). While this fungus

is considered to be best adapted to hot, humid, and even tropical conditions

(Remaudieére 1977, Keller 1997), epizootics have also been reported in Iceland

(Austara et al. 1997, Nielsen et al. 2001). The epizootiology of N. fresenii in

annual crop ecosystems has been well studied (Steinkraus et al. 1995, 1996,

Barta & Cagan 2002), but its commercial exploitation is limited due to absence

of in vitro isolates. In Arkansas (USA), a method for mass-harvesting N.

fresenii from natural epizootics in Aphis gossypii populations in a commercial

cotton field was developed with the intention of using the collected inoculum

for targeted aphid control (Steinkraus & Boys 2005).

Neozygites osornensis, first described during a survey of natural enemies of

invasive Cinara cupressi colonies in Chile (Montalva et al. 2013), was also found

in five localities in southern Chile on C. cupressi and C. tujafilina, which usually

co-occurs with C. cupressi. The infection usually occurred between November

(end of spring) and March (end of summer). We suggest that N. osornensis is

a natural enemy that could serve as a biological control of C. cupressi in Chile.

Neozygites osornensis might also be a candidate for conservation biological

Neozygites species associated with aphids (Chile) ... 241

control of C. tujafilina, an aphid attacking a native host tree, Austrocedrus

chilensis (D. Don) Pic. Serm. & Bizzarri, whose populations are now defined as

vulnerable (Hechenleitner et al. 2005). The application of chemical insecticides

to control C. tujafilina is forbidden in national parks (Corporacion Nacional

Forestal, CONAF) without permission (CONAF 2006).

All Neozygites species that we found in Chile produce resting spores that allow

the fungi to persist during unfavorable conditions for several years (Hajek 1997)

and, thereby, to serve as a reservoir of infection in the ecosystem. For Neozygites

collected during the surveys, we usually found capilliconidia on aphid cadavers

diseased by N. cinarae and N. fresenii during March (end of summer). On the

other hand, N. osornensis capilliconidia were less frequently observed and

were obtained only in the laboratory in a humid chamber on infected Cinara

cupressi collected in Osorno during November (end of spring). Capilliconidia

of Neozygites spp., which are usually produced during the vegetation period, are

responsible for disseminating infection among host populations (Keller 1997).

Contrary to what Keller reported (1997), we observed forcibly discharged

secondary conidia of N. cinarae in Valdivia, although this conidial type was

found only rarely. The absence of rhizoids and pseudocystidia for all species

discussed here agrees with Keller (1991, 1997) and Balazy (1993). Very few

studies have been published for any location treating natural occurrences of

entomophthoroid fungi infecting aphids feeding on woody plants. However, it

appears that N. fresenii, Entomophthora planchoniana Cornu, and Zoophthora

aphidis (Hoffm.) A. Batko are the most frequent fungal species associated

with tree-dwelling aphids (Nielsen et al. 2007, Barta 2009); these findings are

reinforced by our findings about N. fresenii.

The identification of these fungi is an important step for a better knowledge

of natural fungal enemies of aphids in Chile and throughout the world.

However, more studies are on virulence, culturing, strain characterization, and

molecular identification are needed to bring greater and much needed clarity

to our understanding of these relatively little known fungi.

Acknowledgments

The authors thank Servicio Agricola y Ganadero for support the development of this

study in its facilities and DID of the Universidad Austral of Chile project D-2011-11 for

financial support. The authors also thank Richard A. Humber (Cornell University, Ithaca

NY, U.S.A.), Ingeborg Klingen (Norwegian Institute for Agricultural and Environmental

Research, As, Norway), and Christian Luz (Instituto de Patologia Tropical e Satide

Publica, Universidade Federal de Goias, Brasil) for critical review of the manuscript.

Literature cited

Agudelo-Silva P. 1986. A species of Triplosporium (Zygomycetes: Entomophthorales) infecting

Mononychelus progressivus (Acari: Tetranychidae) in Venezuela. Fla. Entomol. 69: 444-446.

242 ... Montalva & al.

Aruta C, Carrillo R. 1989. Identificacién de hongos del orden Entomophthorales en Chile. III. Agro

Sur 17: 10-18.

Aruta C, Carrillo R, Montealegre J. 1984. Determinacién para Chile de hongos del orden

Entomophthorales (Zygomycetes). Agro Sur 12: 36-42.

Austara ©, Carter C, Eilenberg J, Halldorsson G, Harding S. 1997. Natural enemies of the green

spruce aphid in spruce plantations in maritime north-west Europe. Icel. Agric. Sci. 11: 113-124.

Balazy S. 1993. Flora of Poland (Flora Polska). Fungi (Mycota), Volumen 24: Entomophthorales.

Polish Academy of Science, W. Szafer Institute Botany of Botany.

Barta M. 2004. Fungi of the order Entomophthorales infecting aphids in Slovakia, Ph.D. thesis,

Slovak Agricultural University, Dept. of Plant Protection.

Barta M. 2009. Entomophthoralean fungi associated with aphids in woody plants in the Arboretum

Mlynany SAS. Folia Oecologica 36: 1-7.

Barta M, Cagan L. 2002. Prevalence of natural fungal mortality of black bean aphid, Aphis fabae

Scopoli, on primary host and two secondary hosts. Acta Fytotechnica et Zootechnica 5: 57-64.

Barta M, Cagan L. 2003. Entomophthoralean fungi associated with common nettle aphid

(Microlophium carnosum Buckton) and a potential role of nettle patches as reservoirs for the

pathogens in landscape. Anzeiger ftir Schadlingskunde 76: 6-13.

http://dx.doi.org/10.1046/j.1439-0280.2003.03004.x

Barta M, Cagan L. 2006a. Aphid-pathogenic Entomophthorales (their taxonomy, biology and

ecology). Biologia (Section Zoology) 61: 543-616.

http://dx.doi.org/10.2478/s11756-007-0100-x

Barta M, Cagan L. 2006b. Observations on the occurrence of Entomophthorales infecting aphids

(Aphidoidea) in Slovakia. BioControl 51: 795-808.

http://dx.doi.org/10.1007/s10526-006-9007-7

Barta M, Keller S, Cate P, Wegensteiner R. 2005. Observations on the occurrence of Entomophthorales

in Austria. IOBC/wprs Bull 28: 49-53.

Castro T, Wekesa V, Moral R, Demétrio C, Delalibera I, Klingen I. 2013. The effects of photoperiod

and light intensity on the sporulation of Brazilian and Norwegian isolates of Neozygites

floridana. J. Inverteb. Pathol. 114: 230-233. http://dx.doi.org/10.1016/j.jip.2013.08.004

Charnley AK, Collins SA. 2007. Entomopathogenic fungi and their role in pest control. 159-187, in:

CP Kubisek, IS Druzhinina (eds). The Mycota IV, Environmental and Microbial Relationships.

Second edition. New York: Springer.

CONAF. 2006. Normas de manejo del Sistema Nacional de Areas Silvestres Protegidas del Estado.

Chile. 42 p.

Delalibera I, Sosa-Gémez D, de Moraes G. 1992. Infection of Mononychellus tanajoa (Acari:

Tetranychidae) by the fungus Neozygites sp. (Entomophthorales) in northeastern Brazil. Fla.

Entomol. 75: 145-147. http://dx.doi.org/10.2307/3495493

Delalibera I, Humber R, Bento J, de Matos A. 1997. First record of the entomopathogenic fungus

Neozygites fumosa on the cassava mealybug Phenacoccus herreri. J. Inverteb. Pathol. 69:

276-278. http://dx.doi.org/10.1006/jipa.1996.4638

Delalibera I, de Moraes G, Lapointe S, Da Silva C, Tamai M. 2000. Temporal variability and

progression of Neozygites sp. (Zygomycetes: Entomophthorales) in populations of Mononychellus

tanajoa (Bondar) (Acari: Tetranychidae). Anais da Sociedade Entomoldgica do Brasil 29:

523-535.

Delalibera I, Hajek A, Humber R. 2004. Neozygites tanajoae sp. nov., a pathogen of the cassava

green mite. Mycologia 96: 1002-1009. http://dx.doi.org/10.2307/3762084

Neozygites species associated with aphids (Chile) ... 243

Elkinton JS, Hajek AE, Boettner GH, Simons EE. 1991. Distribution and apparent spread of

Entomophaga maimaiga (Zygomycetes: Entomophthorales) in gypsy moth (Lepidoptera:

Lymantriidae) populations in North America. Envir. Entomol. 20: 1601-1605.

Gomez de Picho H. 1991. Neozygites sp. (Zygomycotina: Neozygitaceae) hongo patégeno de Coccus

viridis Green (Homoptera: Diaspididae). Revista Peruana de Entomologia 34: 71-73.

Hajek AE. 1997. Ecology of terrestrial fungal entomopathogens. Adv. Microb. Ecol. 15: 193-249.

http://dx.doi.org/10.1007/978-1-4757-9074-0_5

Hatting JL, Humber RA, Poprawski TJ, Miller RM. 1999. A survey of fungal pathogens of

aphids from South Africa, with special reference to cereal aphids. Biol. Control. 16: 1-12.

http://dx.doi.org/10.1006/bcon.1999.0731

Hechenleitner P, Gardner M, Thonas P, Echeverria C, Escobar B, Brownless P, Martinez C. 2005.

Plantas amenazadas del centro-sur de Chile. Distribucién, conservacién y propagacidén.

Primera edicién. Universidad Austral de Chile y Real Jardin Botanico de Edimburgo.

Humber R. 2012. Entomophthoromycota: a new phylum and reclassification for entomophthoroid

fungi. Mycotaxon 120: 477-492. http://dx.doi.org/10.5248/120.477

Humber R, de Moraes G, Dos Santos J. 1981. Natural infection of Tetranychus evansi [Acarina:

Tetranychidae] by a Triplosporium sp. [Zygomycetes : Entomophthorales] in northeastern Brazil.

Entomophaga 26: 421-425. http://dx.doi.org/10.1007/BF02374716

Keller S. 1987. Arthropod-pathogenic Entomophthorales of Switzerland. I. Conidiobolus,

Entomophaga and Entomophthora. Sydowia 40: 122-167.

Keller S. 1991. Arthropod-pathogenic Entomophthorales of Switzerland. II. Erynia, Eryniopsis,

Neozygites, Zoophthora and Tarichium. Sydowia 43: 39-122.

Keller S. 1997. The genus Neozygites (Zygomycetes, Entomophthorales) with special reference to

species found in tropical regions. Sydowia 49: 118-146.

Keller S. 2006. Entomophthorales attacking aphids with a description of two new species. Sydowia

58: 38-74.

Keller S. 2007. Arthropod-pathogenic Entomophthorales: biology, ecology, identification.

Luxembourg: Office for Official Publications of the European Communities

Keller S, Petrini O. 2005. Keys to the identification of the arthropod pathogenic genera of the

families Entomophthoraceae and Neozygitaceae (Zygomycetes), with descriptions of three new

subfamilies and a new genus. Sydowia 57: 23-53.

Klingen I, Westrum K. 2007. ‘The effect of pesticides used in strawberries on the phytophagous

mite Tetranychus urticae (Acari: Tetranychidae) and its fungal natural enemy

Neozygites floridana (Zygomycetes: Entomopthorales). Biological Control 43: 222-230.

http://dx.doi.org/10.1016/j.biocontrol.2007.07.013

Klubertanz TH, Pedigo LP, Carlsson RE. 1991. Impact of fungal epizootics on the biology and

management of the two-spotted spider mite (Acari: Tetranychidae) in soybean. Environ.

Entomol. 20: 731-735.

Latgeé JP, Papierok B. 1988. Aphid pathogens. 323-335, in: AK Minks, P Harrewijn (eds). Aphids.

Their biology, natural enemies and control. Vol. 2B. Amsterdam: Eds. Elsevier.

Lohr B, Varela A, Santos B. 1990. Exploration for natural enemies of the cassava mealybug,

Phenacoccus manihoti (Homoptera: Pseudococcidae), in South America for the

biological control of this introduced pest in Africa. Bull. Entomol. Res. 80: 417-425.

http://dx.doi.org/10.1017/S0007485300050677

Milner RJ, Holdom DG. 1986. First record of Neozygites fresenii (Nowakowski) Batko, a fungal

pathogen of aphids, in Australia. J. Aust. Entomol. Soc. 25: 85-86.

244 ... Montalva & al.

Milner RJ, Soper RS, Lutton FF. 1982. Field release of an Israeli strain of the fungus Zoophthora

radicans for biological control of Therioaphis maculata f. maculata. J. Austral. Entomol. Soc. 21:

113-118. http://dx.doi-org/10.1111/j.1440-6055.1982.tb01776.x

Montalva C, Rojas E, Ruiz C, Lanfranco D. 2010. El pulgon del ciprés en Chile: Una revision

de la situacién actual y antecedents del control biolégico. Bosque. 2010; 31: 81-88.

http://dx.doi.org/10.4067/S07 17-92002010000200001

Montalva C, Barta M, Rojas E, Valenzuela E. 2013. Neozygites osornensis sp. nov., a new fungal

species causing mortality to the cypress aphid Cinara cupressi in Chile. Mycologia 105:

661-669. http://dx.doi.org/10.3852/12-125

Montserrat M, Castafié C, Santamaria S. 1998. Neozygites parvispora (Zygomycotina:

Entomophthorales) causing an epizootic in Frankliniella occidentalis (Thysanoptera: Thripidae)

on cucumber in Spain. J. Invertebr. Pathol. 71: 165-168.

http://dx.doi.org/10.1006/jipa.1997.4730

Nielsen C, Eilenberg J, Harding S, Oddsdottir E, Halldérsson G. 2001. Geographical distribution

and host range of Entomophthorales infecting the green spruce aphid Elatobium abietinum

Walker in Iceland. J. Inverteb. Pathol. 78: 72-80. http://dx.doi.org/10.1006/jipa.2001.5045

Nielsen C, Jensen A, Eilenberg J. 2007. Survival of entomophthoralean fungi infecting aphids

and higher flies during unfavorable conditions and implications for conservation biological

control. 13-38, in: S Ekesi, N Maniania (eds). Use of entomopathogenic fungi in biological pest

managment 2007. Kerala, India.

Nowakowski L. 1883. Entomophthoreae. Przyczynek do znajomosci pasorzytnych grzybkow

sprawiajacych pomor owadow. Pamietn. Wydz. Akad. Umiej. w. Krakow. 8: 153-183.

Remaudiere G. 1977. Sur quelques Aphidoidea de la Polynésie francaise. Bull. Soc. Entomol. Fr.

$2: 151=155:

Remaudiere G, Keller S. 1980. Révision systématique des genres d’ Entomophthoraceae a potentialité

entomopathogeéne. Mycotaxon 11: 323-338.

Roggia S, Guedes J, Kuss R, Nascimiento G, Navia D, Delalibera I. 2009. Acaros predadores e 0

fungo Neozygites floridana associados a tetraniquideos em soja no Rio Grande do Sul. Pesq

Agropec Bras. 44: 107-110.

Roy HE, Brodie EL, Chandler D, Goettel MS, Pell Jk, Wajnberg E, Vega FE. 2010. Deep space

and hidden depths: understanding the evolution and ecology of fungal entomopathogens.

Biocontrol 55:1-6. http://dx.doi.org/10.1007/s10526-009-9244-7

Sandhu SS, Sharma AK, Beniwal V, Goel G, Batra P, Kumar A, Jaglan S, Sharma AK, Malhotra S.

2012. Myco-biocontrol of insect pests: factors involved, mechanism, and regulation. J. Path.

1-10. http://dx.doi.org/10.1155/2012/126819

Scorsetti A, Lopez-Lastra C. 2007. Primer registro del hongo Neozygites sp. (Zygomycota:

Entomophthorales), patogeno de Tetranychus urticae (Acari: Tetranychidae) en la Republica

Argentina. Boletin de la Sociedad Argentina de Botanica 42: 189-193.

Scorsetti A, Humber R, Garcia J, Lopez-Lastra C. 2007. Natural occurrence of entomopathogenic

fungi (Zygomycetes: Entomophthorales) of aphid (Hemiptera: Aphididae) pest of horticultural

crops in Argentina. Biocontrol 52: 641-655. http://dx.doi.org/10.1007/s10526-006-9045-1

Shah P, Pell J. 2003. Entomopathogenic fungi as biological control agents. Appl. Microbiol.

Biotechnol. 61:413-423. http://dx.doi.org/10.1007/s00253-003-1240-8

Sosa-Gomez D, Lopez C, Humber RA. 2010. An overview of arthropod-associated fungi from

Argentina and Brazil. Mycopathologia 170: 61-76.

http://dx.doi.org/10.1007/s11046-010-9288-3

Neozygites species associated with aphids (Chile) ... 245

Steinkraus DC, Boys GO. 2005. Mass harvesting of the entomopathogenic fungus, Neozygites

fresenii, from natural field epizootics in the cotton aphid, Aphis gossypii. J. Invertebr. Pathol. 88:

212-217. http://dx.doi.org/10.1016/j.jip.2005.01.008

Steinkraus D, Hollingsworth R, Slaymaker P. 1995. Prevalence of Neozygites fresenii

(Entomophthorales: Neozygitaceae) on cotton aphids (Homoptera: Aphididae) in Arkansas

cotton. Environ Entomol 24: 465-474.

Steinkraus DC, Hollingsworth RG, Boys GO. 1996. Aerial spores of Neozygites fresenii

(Entomophthorales: Neozygitaceae): density, periodicity, and potential role in cotton aphid

(Hemiptera: Aphididae) epizootics. Environ. Entomol. 25: 48-57.

Thaxter R. 1888. The Entomophthoreae of the United States. Mem. Boston Soc. Nat. Hist. 4:

133-201.

Van der Geest L, de Moraes G, Navia D, Tanzini M. 2002. New records of pathogenic fungi in mites

(Arachnida: Acari) from Brazil. Neotrop Entomol. 31: 493-495.

http://dx.doi.org/10.1590/S1519-566X2002000300025

Wekesa V, de Moraes G, Ortega E, Delalibera I. 2010. Effect of temperature on sporulation

of Neozygites floridana isolates from different climates and their virulence against

the tomato red spider mite, Tetranychus evansi. J. Invertebr. Pathol. 103: 36-42.

http://dx.doi.org/10.1016/j.jip.2009.10.003

MY COTAXON

http://dx.doi.org/10.5248/129.247

Volume 129(2), pp. 247-253 October-December 2014

First report of Pseudoveronaea ellipsoidea

causing sooty blotch and flyspeck in China

CHEN CHEN’, WENHUAN LI’, Liu GAo', RONG ZHANG’,

GUANGYU SUN’, & MARK L. GLEASON?

'State Key Laboratory of Crop Stress Biology in Arid Areas and College of Plant Protection,

Northwest A&F University, Yangling, Shaanxi, 712100, P. R. China

*Department of Plant Pathology and Microbiology, Iowa State University,

Ames, Iowa 50011, USA

“CORRESPONDENCE TO: sgy@nwsuaf.edu.cn

ABSTRACT — Pseudoveronaea ellipsoidea is reported and described as a newly recorded

species from China. This fungus was isolated from peels of hawthorn and apple fruit collected

from Shaanxi and Shanxi Provinces, and caused sooty blotch and flyspeck signs on these

hosts. Based on morphological characteristics and phylogenetic analysis, we identified our

isolates as P. ellipsoidea. This is also the first report of the genus Pseudoveronaea in China.

Key worps — Dissoconiaceae, taxonomy, morphology, phylogeny

Introduction

Sooty blotch and flyspeck (SBFS) fungi colonize the waxy cuticle of a

wide range of plant hosts in humid regions worldwide. They cause cosmetic

damage to cultivated fruits and ornamental plants due to the smudges formed

on the surface, resulting in economic losses to growers (Williamson &

Sutton 2000, Batzer et al. 2005, Gleason et al. 2011). For nearly 75 years this

disease was considered to be two different diseases, each caused by a single

species (Colby 1920, Williamson & Sutton 2000). Based on analysis of rDNA

sequences combined with traditional morphology, researchers obtained a

better understanding of SBFS, the number of species in the complex increased

to more than 80 (Diaz Arias et al. 2010; Frank et al. 2010; Gleason et al. 2011;

Li et al. 2011, 2012), and many new genera and species were described in the

discovery process.

Among these fungi, Pseudoveronaea was recently established as a new

genus and named after its morphological similarity to Veronaea (Li et al.

248 ... Chen & al.

2012). A member of the family Dissoconiaceae, Pseudoveronaea is nevertheless

phylogenetically and morphologically distinct from other genera in this family,

in that it produces only one type of conidia and these are septate. Currently

there are only two species in this genus, P ellipsoidea and P. obclavata, both

associated with SBFS on apple (Gleason et al. 2011, Li et al. 2012).

In our recent study on SBFS, four isolates obtained from hawthorn and

apple were identified as P. ellipsoidea, based on morphological comparison and

phylogenetic analysis of the ITS region.

Materials & methods

Isolates and morphology

Hawthorn (Crataegus pinnatifida Bunge) and apple (Malus domestica Baumg.)

fruit with SBFS signs were collected from Shaanxi and Shanxi Provinces in 2011. Peels

with SBFS were cut off, examined and photographed under the dissecting microscope,

then pressed between paper towels until dry and preserved. Pieces of SBFS colonies on

the peels were transferred to potato-dextrose agar (PDA) and cultured at 25°C in the

dark (Sun et al. 2003). After one month, pure isolates were obtained and transferred

to synthetic nutrient-poor agar (SNA) for morphological examination and description

using the method of Zhang et al. (2009). Colony descriptions were based on cultures

on PDA and oatmeal agar (OA) after one month in the dark at 25°C. Specimens of

representative dried cultures were deposited in the Fungal Herbarium of Northwest

A&F University, Yangling, Shaanxi Province, China (HMUABO).

DNA extraction, PCR and sequencing

Genomic DNA was extracted from fungal mycelium according to the protocol of

Barnes et al. (2001). The primers ITS1-F (Gardes & Bruns 1993) and ITS4 (White et

al. 1990) were used to amplify the internal transcribed spacer (ITS) region of nuclear

ribosomal DNA. The PCR conditions followed the methods of Zhuang et al. (2010).

PCR products were sequenced by Sangon Biotech (Shanghai), China.

Sequence alignment and phylogenetic analysis

Sequences in this study and other sequences downloaded from GenBank were

put into CLUSTAL X to perform preliminary alignment (Thompson et al. 1997),

and then manually adjusted in BioEdit v. 5.0.9.1 (Hall 1999). Maximum parsimony

(MP) analysis of aligned sequences was conducted using PAUP v. 4.0b10 (Swofford

2003). Heuristic searches were performed with a 1000 random taxa addition and tree

bisection-reconnection (TBR) as the branch-swapping algorithm. All characters were

unordered and given equal weight and gaps were treated as missing data. The robustness

of clades and internal branches was evaluated by 1000 bootstrap replications. Tree

length (TL), consistency index (CI), retention index (RI) and rescaled consistency

index (RC) were also calculated. Sequences generated in this study were deposited

in GenBank. Alignment and the representative tree were deposited in TreeBase

(http://purl.org/phylo/treebase/phylows/study/TB2:S13736).

Pseudoveronaea ellipsoidea in China ... 249

ZZSZ16 KC329918

Pseudoveronaea ellipsoidea FJ425205

“1 ZZSZ21 KC329919

99 SXGL5 KC329920

PCPG10 KC329921

Pseudoveronaea obclavata AY598877

es Dissoconium eucalypti EF394855

100| "~- Dissoconium aciculare AF173308

75 Dissoconium proteae EU707897

ggf Ramichlondium apiculatum EU041 794

67} * Ramichloridium cucurbitae JQ622087

sei 101 + Ramichloridium luteum EU329730

Ramichloridium punctatum JQ622086

Uwebraunia dekkeri FJ425204

Uwebraunia musae EU514225

Uwebraunia australiensis EF394854

Uwebraunia communis JQ622085

gs) _Veronaea compacta EU041819

100 Veronaea japonica EU041818

100 Veronaea botryosa JF747142

400 Rhinocladiella aquaspersa GU017733

Rhinocladiella phaeophora EU041811

7 Ophiostoma stenoceras JX028583

Fic. 1. One of four equally most parsimonious trees obtained from a heuristic search with 1000

random taxon additions of the ITS sequence alignment. The bootstrap support values (>50%)

based on 1000 replicates are shown at the nodes. The tree was rooted to Ophiostoma stenoceras. The

scale bar shows 10 changes. Strains investigated in this paper are presented in bold.

Results

Phylogenetic analysis

The manually adjusted ITS alignment contained 23 sequences (including

the outgroup sequence) and 604 characters including alignment gaps. Of these

characters, 223 were parsimony-informative, 102 were variable and parsimony-

uninformative, and 279 were constant. Four equally most parsimonious trees

were obtained from the analysis, the first of which is shown in Fic. 1 (TL =

601 steps, CI = 0.7920, RI = 0.8808, RC = 0.6976). Two major clades were

resolved from the analysis. One clade included five species in Veronaea and

250 ... Chen & al.

Rhinocladiella. The other clade, with 100% bootstrap support, contained 17

strains in Pseudoveronaea, Dissoconium, Ramichloridium, and Uwebraunia.

Our strains clustered together with P ellipsoidea in the first subclade with a

bootstrap value of 93%, indicating that they might represent the same species.

Taxonomy

Pseudoveronaea ellipsoidea Batzer & Crous, Persoonia 28: 119, 2012 Fig. 2

HyPHaeE septate, branched, smooth, hyaline and turning to brown as

culture aged, 2-3.5 um diam. CoNIDIOPHORES erect, solitary, arising from

aerial hyphae, straight to flexuous, unbranched, 3-11-septate, subcylindrical,

smooth, thick-walled, brown, 35-115 x 3-4.5 um. CONIDIOGENOUS CELLS

terminal, integrated, subcylindrical, with apical taper to acutely rounded apex,

pale brown to brown, 6-21 x 3-5 um, sympodially proliferating, rachis straight

or flexuous, scars slightly darkened and unthickened, 0.5-1 um diam. ConrIpDIA

solitary, 0-1-septate, ellipsoid to obclavate, hyaline to pale brown, (6—-)8-11

(-16) x 3-4 um, base truncate, slightly darkened and thickened, 1-1.5 um diam.

CULTURAL CHARACTERISTICS — Colonies after 1 month on PDA at 25°C

flat, raised at the middle, slightly folded, with lobate, smooth margin and sparse

aerial mycelium, surface iron-gray, reverse black, reaching (18-)20(-23) mm

diam. On OA faster growing, flat, spreading, with lobate, feathery margin and

moderate aerial mycelium, surface olivaceous gray, reverse black, reaching 32

mm diam.

On peels of hawthorn fruit, showing the ridged honeycomb mycelial

type characterized by clumps and ridges of mycelia, olivaceous gray, without

distinct edges. On peels of apple fruit, forming the fuliginous mycelial type

characterized by uniform mats of mycelia with distinct, feathered edges.

SPECIMENS EXAMINED: CHINA, SHAANXI PROVINCE: Xian City, Zhouzhi County,

on fruit surface of hawthorn (Crataegus pinnatifida), 15 Oct. 2011, Chen C & Dang JL

ZZSZ16 (HMUABO; GenBank KC329918), ZZSZ21 (HMUABO; GenBank KC329919);

Weinan City, Pucheng County, on fruit surface of apple (Malus domestica), 25 Sept.

2011, Li WH & Gao L PCPG10 (HMUABO; GenBank KC329921); SHANXI PROVINCE:

on fruit surface of apple (Malus domestica), 16 Oct. 2011, Li WH & Gao L SXGL5

(HMUABO; GenBank KC329920).

Discussion

Based on phylogenetic analysis and morphological characteristics, we

identified our isolates as P. ellipsoidea, a newly recorded species for China. This

is also the first time that the genus Pseudoveronaea has been reported from

China.

Previous study showed P ellipsoidea causing SBFS on apple as displaying the

fuliginous mycelial type (Li et al. 2012). In our study, P. ellipsoidea formed the

Pseudoveronaea ellipsoidea in China ... 251

Fic. 2. Pseudoveronaea ellipsoidea (ZZSZ16). A. Signs on hawthorn. B. Colony on PDA. C. Colony

on OA. D-H. Conidiophores showing rachis, scars and conidia. I-J. Conidia. Bars: D-J = 10 um.

same mycelial type on apple, but caused a different type, ridged honeycomb, on

hawthorn. Hemnani et al. (2008) also found a similar phenomenon in that the

same SBFS species produced different mycelial types on different hosts. More

investigations are needed to clarify the relationships between hosts and SBFS

mycelial types.

So far, only two Pseudoveronaea species have been reported in USA (Li et

al. 2012) and China. More collections and reports on the genus are required

to help us understand the taxonomy, biogeography, and ecology of this genus.

252 ... Chen & al.

Acknowledgments

This work was supported by National Natural Science Foundation of China

(31170015, 31171797), the 111 Project from Education Ministry of China (B07049), and

the earmarked fund for China Agriculture Research System (CARS-28). The authors

wish to thank Dr Eric H.C. McKenzie (Landcare Research, Auckland, New Zealand)

and Prof Zhongyi Zhang (deceased, Yunnan Agricultural University, Kunming, China)

for reviewing the manuscript.

Literature cited

Barnes I, Roux J, Wingfield MJ, Coetzee MPA, Wingfield BD. 2001. Characterization of Seiridium

spp. associated with cypress canker based on $-tubulin and histone sequences. Plant Disease

85: 317-321. http://dx.doi.org/10.1094/PDIS.2001.85.3.317

Batzer JC, Gleason ML, Harrington TC, Tiffany LH. 2005. Expansion of the sooty blotch and

flyspeck complex on apples based on analysis of ribosomal DNA gene sequences and

morphology. Mycologia 97: 1268-1286. http://dx.doi.org/10.3852/mycologia.97.6.1268

Colby AS. 1920. Sooty blotch of pomaceous fruits. Transactions of the Illinois State Academy of

Science 13: 139-175.

Diaz Arias MM, Batzer JC, Harrington TC, Wong AW, Bost SC, Cooley DR, Ellis MA,

Hartman JR, Rosenberger DA, Sundin GW, Sutton TB, Travis JW, Wheeler MJ, Yoder

KS, Gleason ML. 2010. Diversity and biogeography of sooty blotch and flyspeck fungi

on apple in the eastern and midwestern United States. Phytopathology 100: 345-355.

http://dx.doi.org/10.1094/PHY TO-100-4-0345

Frank J, Crous PW, Groenewald JZ, Oertel B, Hyde KD, Phengsintham P, Schroers HJ. 2010.

Microcyclospora and Microcyclosporella: novel genera accommodating epiphytic fungi causing

sooty blotch on apple. Persoonia 24: 93-105. http://dx.doi.org/10.3767/003158510X510560

Gardes M, Bruns TD. 1993. ITS primers with enhanced specificity for basidiomycetes —

application to the identification of mycorrhizae and rusts. Molecular Ecology 2: 113-118.

http://dx.doi.org/10.1111/j.1365-294X.1993.tb00005.x

Gleason ML, Batzer JC, Sun GY, Zhang R, Diaz Arias MM, Sutton TB, Crous PW, Ivanovic

M, McManus PS, Cooley DR, Mayr U, Weber RWS, Yoder KS, Del Ponte EM, Biggs AR,

Oertel B. 2011. A new view of sooty blotch and flyspeck. Plant Disease 95: 368-383.

http://dx.doi.org/10.1094/PDIS-08- 10-0590

Hall TA. 1999. BioEdit: a user-friendly biological sequence alignment editor and analysis program

for Windows 95/98/NT. Nucleic Acids Symposium Series 41: 95-98.

Hemnani K, O’Malley PJ, Tanovi¢ B, Batzer JC, Gleason ML. 2008. First report of seven species

of sooty blotch and flyspeck fungi on Asimina triloba in Iowa. Plant Disease 92: 1366.

http://dx.doi.org/10.1094/PDIS-92-9-1366C

Li HY, Sun GY, Batzer JC, Crous PW, Groenewald JZ, Karakaya A, Gleason ML. 2011. Scleroramularia

gen. nov. associated with sooty blotch and flyspeck of apple and pawpaw from the Northern

Hemisphere. Fungal Diversity 46: 53-66. http://dx.doi.org/10.1007/s13225-010-0074-9

Li HY, Sun GY, Zhai XR, Batzer JC, Mayfield DA, Crous PW, Groenewald JZ, Gleason ML. 2012.

Dissoconiaceae associated with sooty blotch and flyspeck on fruits in China and the United

States. Persoonia 28: 113-125. http://dx.doi.org/10.3767/003158512X651157

Sun GY, Zhang R, Zhang Z, Zhang M. 2003. Isolation of sooty blotch and flyspeck fungi from apple

surface by picking up the thalli. Acta Phytopathologica Sinica 33: 479-480.

Swofford DL. 2003. PAUP’. Phylogenetic analysis using parsimony (‘and other methods). Version

4.0. Sinauer Associates, Sunderland, Massachusetts, USA.

Pseudoveronaea ellipsoidea in China ... 253

Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG. 1997. The CLUSTAL_X windows

interface: flexible strategies for multiple sequence alignment aided by quality analysis tools.

Nucleic Acids Research 25: 4876-4882. http://dx.doi.org/10.1093/nar/25.24.4876

White TJ, Bruns T, Lee S, Taylor J. 1990. Amplification and direct sequencing of fungal ribosomal

RNA genes for phylogenetics. 315-322, in: MA Innis et al. (eds). PCR Protocols: a guide to

methods and applications. San Diego, Academic Press.

Williamson SM, Sutton TB. 2000. Sooty blotch and flyspeck of apple: etiology, biology, and control.

Plant Disease 84: 714-724. http://dx.doi.org/10.1094/PDIS.2000.84.7.714

Zhang R, Yang HL, Sun GY, Li HY, Zhuang JL, Zhai XR, Gleason ML. 2009. Strelitziana

mali, a new species causing sooty blotch on apple fruit. Mycotaxon 110: 477-485.

http://dx.doi.org/10.5248/110.477

Zhuang JL, Zhu MQ, Zhang R, Yin H, Lei YP, Sun GY, Gleason ML. 2010. Phialophora sessilis, a

species causing flyspeck signs on bamboo in China. Mycotaxon 113: 405-413.

http://dx.doi.org/10.5248/113.405

MY COTAXON

http://dx.doi.org/10.5248/129.255

Volume 129(2), pp. 255-268 October-December 2014

Molecular annotation of type specimens of Russula species

described by W.A. Murrill from the southeast United States

BRIAN P. LOONEY

Department of Ecology & Evolutionary Biology, University of Tennessee

32 Hesler Biology Bldg., Knoxville TN 37996 USA

* CORRESPONDENCE TO: blooney@vols.utk.edu

ABSTRACT — Twenty-five historical type collections of North American Russula species

described by W.A. Murrill were sequenced and analyzed for placement in a phylogenetic

and taxonomic framework. Molecular data was successfully obtained from sixteen type

specimens (66%), of which eight species have never before been placed in an infrageneric

classification system. Classifications and synonymization of taxa proposed by Rolf Singer and

others are evaluated. Russula albimarginata and R. emeticiformis are suggested as synonyms

of R. vinacea, a widespread and common species in the eastern United States. Four taxa of

unknown classification (R. albimarginata, R. pinophila, R. subcremeiceps, R. subrubescens)

have been classified to subsection level, and an alternative classification is proposed for

R. westii. A morphological comparison of a collection from British Columbia (Canada) with

99% sequence similarity to R. levyana in the Internal Transcribed Spacer 1 (ITS1) region

indicates that this region alone may be inadequate for species barcoding in Russula.

Key worps —phylogenetics, TENN herbarium, high performance DNA extraction, Florida

Introduction

Between 1938 and 1948, W.A. Murrill described 110 species in the genus

Russula Pers. almost exclusively from the area surrounding Gainesville, Florida

(Mycobank 2014). Of the 334 Russula species described from the United States,

Murrill’s work comprises the largest number described by a single mycologist

(Buyck 2007). Unfortunately, most of the taxa are poorly known, due partly

to Murrill’s short diagnoses that focused on macromorphological characters

and partly to the lack of extensive studies of Russula in the southeast U.S. One

notable exception to the lack of attention given to Russula in the region is a

series of type studies by Hesler (1960, 1961). These studies treat 191 Russula

species, including 87 species described by Murrill, and they contribute data

from microscopic descriptions of the pileus cuticle, lamellar cystidia, and

256 ... Looney

spore morphology with some discussion of taxonomic affinities proposed by

Rolf Singer. Hesler received authentic and type material from major herbaria

(FH, MICH, NY, NYS, and FLAS) for these studies, and in the case of Murrill’s

collections, many were sent as isotypes to be accessioned at TENN (herbarium

abbreviations per Thiers 2013).

Central to our current understanding of Russula in North America are the

studies of Rolf Singer (Singer 1986), which include the most comprehensive

infrageneric classification system to include North American taxa. Singer

treated a number of Murrill’s taxa in various editions of “The Agaricales in

Modern Taxonomy” (Singer 1951, 1975, 1986), and it is my objective to test these

proposed relationships in a phylogenetic context. A recent initiative to revive

the taxonomy of North American Russula has led to a number of published

type studies using advanced modern microscopy and morphometrics and

unpublished phylogenies upon which a number of taxonomic affinities have

been proposed (Adam¢ik & Buyck 2010, Adam¢ik & Buyck 2011a, Adamcik &

Buyck 2011b, Adaméik et al. 2013, Buyck & Adam¢cik 201 1a, Buyck & Adam¢cik

2011b, Buyck & Adam¢ik 2013). With these renewed concepts, new reports of

historical Russula taxa are emerging (Adam¢ik et al. 2010, Buyck et al. 2008,

Buyck et al. 2011). The only other comprehensive treatment of a majority of

Murrill’s taxa can be found in the appendix of Kibby & Fatto (1990), who coded

features of southern species for use in their synoptic key system, which also

included abbreviated species descriptions. The objective of this study is to

contribute to the revival of Russula taxonomy in North America by determining

whether Murrill’s type material at TENN can be molecularly annotated and by

exploring what sort of taxonomic inferences can be drawn from these data.

Molecular methods

Following an inventory of Russula type material deposited at TENN, 25 type

collections of species designated by W.A. Murrill were selected for DNA extraction and

molecular annotation (TABLE 1). In most cases, it was determined that ample material

was available to allow using 50 mg of dried material for DNA extraction. The sporocarp

material and a pinch of sterile sand were placed in a mortar to which liquid nitrogen was

added. The frozen material was ground to a fine powder with a pestle and scraped into a

1.5 mL microtube with a metal spatula. DNA extraction protocols followed the E.Z.N.A.

HP Fungal DNA Kit (Omega Bio-Tek, Norcross, Georgia) with the following notable

exceptions to improve end-product DNA concentration: 1) 10 wL 2-mercaptoethanol

was added to the samples in buffer, and they were allowed to incubate at 65°C for 24

hours (vortexing at the beginning of incubation and intermittently during the final 30

minutes). 2) After the prescribed 300 ul of supernatant and associated reagents were

centrifuged through the HiBind DNA column, the remaining supernatant was run

through a second round of centrifugation. 3) Two rounds of DNA elution using 50 uL

of Elution buffer were performed after a 5 minute incubation at 65°C with the buffer

added. Dilutions of 1:10 were made from the genomic DNA product.

257

Sequence analyses of Murrill’s Russula types (U.S.A.) ...

TABLE 1. Sequence data from Russula taxa selected for phylogenetic comparison with Singer’s infrageneric classification

COLLECTIONS!

. westii

. brunneipes

. subrubescens

. variicolor

. subcremeiceps

. alachuana

. albimarginata

. levyana

. vinosirosea

. mutabilis

. fragiloides

. australirosea

. emeticiformis

. pinophila

. subsulphurea

. subrubescens

. pervirginea Murrill

. subflava Murrill

. venusta Murrill

. subglauca Murrill

. alutaceiformis Murrill

. testaceiceps Murrill

. subobscura Murrill

. patriotica Murrill

. subpusilla Murrill

. mutabilis

. cf. aquosa Leclair

APRA ARPARARARAA PAA RAARAARPAARAARAARDDAD DA

. vinacea

TYPE DESIG.

Isotype

Paratype

Isotype

Paratype

Isotype

none

COLL. #

F16404

F19537

F18349

F9513

F38920

F9510

F19447

F15859

F18677

F17943

F18001

F38859

F9535

F17982

F18743

F18339

F17266

F16256

F17836

F17781

F17703

F15916

F18301

F18066

F9512

DPL10654

BPL271

BPL257

TENN #

21262

21229

21254

21259

21230

21221

21225

21235

21261

21237

21232

21228

21231

21240

21255

21253

21238

21249

21260

21250

21226

21257

21251

21258

21252

ITS1 ITS2 GENBANK MATCH

Won Ses

SSeS eye SSN SE NN

V 93% R. aeruginea CAN

98% R. sp. MA

97% orchid root tip OH

99% Uncultured Eur

98% R. sp. VT

99% orchid root tip MEX

99% R. atropurpurea TN

99% R. xerampelina BC

94% R. sp. BC

98% orchid root tip THA

90% R. raoultii Eur

92% R. sp. CA

96% R. atropurpurea TN

96% R. xerampelina BC

100% environ NC

99% Uncultured MEX

FAILED

J N/A

INFRA. CLASS.

Lilaceinae

unknown

Amoeninae

unknown

Amoeninae

unknown

Xerampelinae

unknown

Subvelatae

subsect. Russula

Lilaceinae

subsect. Russula

unknown

Subvelatae

subsect. Russula

PHYLO. CLASS.

Subcompactinae

uncertain

Urentes

Amoeninae

Chamaeleontinae

Amoeninae

subsect. Russula

Xerampelinae

uncertain

Ingratae

subsect. Russula

uncertain

subsect. Russula

Xerampelinae

uncertain

Urentes

Ingratae

GENBANK #

KF810121

KF810122

KF810123

KF810124

KF810125

KF810126

KF810127

KF810128

KF810129

KF810130

KF810131

KF810132

KF810133

KF810134

KF810135

KF810136

KF810137

KF810138

KF810139

‘Type collections accessioned at TENN not sampled (out on loan) were: Russula albiduliformis Murrill, R. heterosporoides Murrill, R. lutescentifolia Murrill, R. pseudofoetens

Murrill, R. rooseveltiana Murrill, R. subalbidula Murrill, R. subbrunneipes Murrill, and R. subgraminicolor Murrill.

258 ... Looney

Initially, products were screened using the primer pair ITS1F-ITS4 (Gardes & Bruns

1993, White et al. 1990). PCR amplification protocols and controls used the requisite

reagents of sterile water, 5x GoTaq buffer (Promega, Madison, Wisconsin), GoTaq,

and 10 mM solution of dNTPs (Invitrogen Corp., Carlsbad, California). The DNA

concentration in master mix solution was increased from 1:24 to 4:21 for improved

PCR product concentration, and the samples were run using an ITS protocol on a

Bio-Rad C1000 thermal cycler (Bio-Rad, Hercules, California). PCR product quality

was visualized using gel electrophoresis on a 1% agarose gel prepared with ethidium

bromide and then transilluminated using UV light. Specimens for which no banding

was present were re-screened using the primer pairs ITS1F-ITS2 and 5.8SR-ITS4.

Specimens for which a band was produced were cleaned with a QIAquick PCR

purification kit (QIAGEN, Valencia, California). Sequence reactions were prepared and

purified following Birkebak et al. (2013). Purified samples were sent to the Molecular

Biology Resource Facility at U.T. to be sequenced using a 3730 DNA analyzer (Applied

Biosystems, Grand Island, NY).

Sequences were assembled using Sequencher 4.9 (Gene Codes Corp., Ann Arbor,

Michigan). For types where ITS] and ITS2 sequences were produced separately, a

bridge of repeated N’s was used to form a complete ITS sequence. Using the BLAST

program (Altschul et al. 1990) sequences were queried against the GenBank database

(NCBI, Bethesda, Maryland) and visualized using the distance tree of results. Sequences

that blasted with 98% similarity or higher were selected for phylogenetic analysis as

candidate conspecifics or closely related taxa as well as identified sequences with at

least 90% similarity for placement in an infrageneric group. Additional vouchered

sequences of nomenclatural type species or representatives of infrageneric groups based

on morphological synapomorphies were selected for testing infrageneric hypotheses.

Alignments were formed using MAFFT version 7 (Katoh & Standley 2013) using default

options and then adjusted manually in MacClade 4.08 (Maddison & Maddison 2005).

Phylip file formats were created in Seaview 4.4.2 (Gouy et al. 2010). Phylogenies were

reconstructed using maximum likelihood (ML) in raxmlGUI 1.2 (Silvestro & Michalak

2012, Stamatakis et al. 2008) with 1000 bootstrap replicates. Bootstrap values 70%

and above are considered good support for clades. State/provincial abbreviations for

the U.S. and Canada are according to national postal codes and country abbreviations

use the three-letter ISO code (International Organization for Standardization, Geneva,

Switzerland). All sequences are deposited in GenBank.

Molecular results

Sequence data were recovered from sixteen of twenty-five type specimens

(64% screened) with collection dates ranging from 1932 to 1944 (TABLE 1).

Only R. westii Murrill (KF810121; 4% screened) was recovered as a full ITS

sequence using the ITS1F-ITS4 primer pair. When the spacer regions were

amplified separately, the ITS1 region was successfully amplified in fourteen of

fifteen instances (93%) using the ITS1F-ITS2 primer pair. The ITS2 region was

successfully amplified for three out of fifteen types (20%) using the 5.8SR-ITS4

primer pair. Both ITS1 and ITS2 were produced for R. subrubescens Murrill

Sequence analyses of Murrill’s Russula types (U.S.A.) ... 259

FJ454938 orchid root THA

Tree 1 (Ingratae,

Lilaceinae, and FJ454960 orchid root THA

Subcompactinae) 79 R. mutabilis ISOTYPE F17943

80!LR. mutabilis DPL10654 TX

DQ422024 R. illota Eur

9 71HQ677769 R. illota IRL

EU598184 R. laurocerasi TN

100P4Y 961736 R. pulverulenta NAm

EU598186 R. pulverulenta TN

EU598187 R. cf. ventricosipes TN

AY061677 R. foetens Eur

100 GQ924690 R. ionochlora GER

94 AY061679 R. grisea Eur

98 R. westii ISOTYPE F164.04

JQ711921 R. aeruginea BC

8 4

100 JF908698 R. anatina Eur

DQ422007 R. parazurea Eur

EU598178 R. subtilis TN

JN944004 R. zvarae FRA

JN944005 R. lilacea SVK

EU598174 R. peckii sensu Sing.TN

DQ422013 R. lepida BEL ae

R. australirosea ISOTYPE F38859

EU880225 Uncultured MN

Tree 2 (Urentes 95 |F]196957 Uncultured MEX

and Decolorantes) R. subrubescens ISOTYPE F18339

09 )UDB011222 R. globispora EST

UDB011235 R. sp. EST

R. subrubescens PARATYPE F18349|

JF908710 R. dryadicola FRA

UDB016122 R. globispora EST

AF495464 R. sp. CO

AF418639 R. sp. EUR

100 JF908653 R. maculata ITA

AY061688 R. maculata EUR

DQ422015 R. cf. maculata BEL

FJ845432 R. decolorans BC

AY194601 R. decolorans SWE

260 ... Looney

AY061654 R. atropurpurea NAm

Tree 3 (Russula) R. emeticiformis ISOTYPE F9535

R. albimarginata ISOTYPE F19447

GU328630 Uncultured MI

R. vinacea BPL257 TN

100/EU598181 R. vinacea TN

HE820322 Uncultured OH

JQ272366 “R. atropurpurea” TN

9 9 JF908691 “R. atropurpurea” | Nas

JF908660 “R. atropurpurea” ITA

AF418618 “R. atropurpurea” GER

R. fragiloides ISOTYPE F18001

AF418621 R. raoultii Eur

FJ845435 R. bicolor BC

R. aquosa BPL271 TN

KC581327 R. fragilis BC

HQ604846 R. crenulata BC

DQ421997 R. emetica Eur

AM113956 R. brunneoviolacea DEN (Xerampelinae,

AY061691 R. melzeri Eur Chamaeleontinae, and

UDB011113 R. olivobrunnea FIN ;

UDB016260 R. olivina FIN Incertae sedis)

UDB002420 R. cuprea GBR

AF349710 R. sp. NC

DQ777981 R. sp. MA

DQ777980 Uncultured MA

R. subsulphurea PARATYPE F18743

AY456357 R. sp. NC

JQ272402 R. sp. NC

UDB011200 R. veternosa EST

JX178490 R. umerensis NZL

100} 9 g,GU220376 R. sp. MA

FM999623 Uncultured OH

9 6|_R prunneipes ISOTYPE F19537

JQ888199 R. paludosa GBR

R. vinosirosea ISOTYPE F18677

7 4FJ845433 R. xerampelina BC

FJ803947 Uncultured CHI

R. pinophila ISOTYPE F17982

R. levyana ISOTYPE F15859

HQ667806 Uncultured MEX

AY061705 R. pascua EUR iS

GU998782 Uncultured AK eo

oe &

70 EU569274 R. sp. MEX & WN

UDB000906 R. cremeoavellanea SWE +

100! __R subcremeiceps ISOTYPE WAM21230 o SS w

AF349709 R. sp. VT x & &

100 UDB000350 R. vinosa GER we

AJ534938 R. vinosa EST «Vv

HM189949 R. velenovskyi Eur a e

UDB015052 R. vinososordida FIN

Sequence analyses of Murrill’s Russula types (U.S.A.) ... 261

EU598199 R. mariae TN

JX030253 R. aff. mariae NY

EU819426 R. mariae WI

EU598164 R. nitida TN

AY061726 R. violeipes Eur

99 JF908655 R. violeipes Eur

AY061655 R. amoenicolor Eur

GU371290 R. cf. violeipes CHI

HQ667813 Uncultured MEX

R. alachuana ISOTYPE F9510

R. variicolor ISOTYPE F9513

JQ975980 Uncultured Eur

Tree 5 (Amoeninae)

AY061704 R. parazurea Eur

EU598170 R. flavida TN

FiGuREs 1-3. Unrooted phylogenies derived from maximum likelihood analysis of complete or

partial ITS rDNA sequences of types of Russula taxa described by W.A. Murrill with included

publicly accessioned sequences for phylogenetic classification. Type sequences are highlighted and

bolded. Publicly accessioned sequences are included if they share a 98% sequence similarity to a

type or are identified to species and within 90% sequence similarity.

(KF810136) and R. subsulphurea Murrill (KF810135). Five phylogenetic trees

were constructed by utilizing nomenclatural type species or representatives

of infrageneric groups based on morphological synapomorphies to test

proposed infrageneric classifications and synonymizations of Murrill’s taxa.

Monophyletic clades with high bootstrap support that include type species

or representatives of known infrageneric classification were used to define

infrageneric groups; however, relationships between these clades should not be

considered significant due to the lack of gene and taxon sampling.

To test whether the ITS1 region can consistently and accurately separate

species in Russula, I examined collection FJ845433 from British Columbia,

Canada, which shares a 99% sequence similarity with 309/310 sites of the

ITS1 region from the isotype of R. levyana Murrill. A disjunct or continuous

range of over 4000 km across continental North America for one Russula

species would be surprising, especially considering the significant host,

habitat, and climate differences. Although both specimens certainly belong

in R. subsect. Xerampelinae Singer, in the pileus of the Canadian collection, I

found large spores, predominantly olive green tones, and no long, attenuated

hyphal terminations in the suprapellis — features inconsistent with modern

examinations of the type and recent collections of R. levyana and which support

separation based on a morphological species concept (Adamcik & Buyck 2010).

262 ... Looney

Tree 1 (Ingratae, Lilaceinae, and Subcompactinae) FIGURE 1

The isotype of R. mutabilis Murrill shares a 100% sequence similarity with a

recent specimen collected in Texas determined as R. mutabilis (KF810137). The

species is in a well-supported clade that includes sequences identified as R. illota

Romagn. and environmental sequences of an orchid associate from Thailand.

This suggests that R. mutabilis is in R. sect. Ingratae Queél., but its placement

in R. subsect. Foetentinae Melzer & Zvara or R. subsect. Subvelatae (Singer)

Singer, represented by sequences labeled as R. pulverulenta Peck, cannot be

resolved. Russula pulverulenta sequences were chosen because no sequences of

R. subvelata Singer are available, and R. pulverulenta is a commonly collected

species with the floccose pilear patches characteristic of R. subsect. Subvelatae.

Russula mutabilis and R. illota share a benzaldehyde odor (characteristic of

R. fragrantissima Romagn.), acrid taste, and a staining context, although Singer

(1958), who considered its staining an artifact of the drying process, placed

R. mutabilis in R. subsect. Subvelatae based on its velar remnants that showed

a positive red reaction to potassium hydroxide. A similar species that exhibits

the same extreme red staining on both the pileus and stipe is R. ventricosipes

Peck, which may be a close relative separated by host preference (Adam¢ik et

al. 2013).

The isotype of R. westii was recovered in a well-supported clade with

members of R. subsect. Subcompactinae Singer, separate from a clade containing

members of R. subsect. Lilaceinae Melzer & Zvara, including the type species.

Morphological characters of R. westii are not inconsistent with placement in

R. subsect. Subcompactinae, which includes a cream spore print, mild taste,

and verrucose spores that lack an amyloid suprahilar spot (Murrill 1941).

Singer (1958) noted an absence of pileocystidia and the presence of “none

or few” primordial hyphae, but a preliminary morphological examination

of the isotype by Adamcik revealed conspicuous pileocystidia. To determine

whether the type collection was mixed and the TENN material represents a

different species, the isotype was compared with a more detailed study of the

FLAS holotype and confirmed as conspecific (Adam¢cik pers. comm.). Singer

(1958) noted that R. westii was morphologically and ecologically very similar to

R. cremea (Murrill) Singer and differed only in spore morphology.

Contrasting with Singer's (1986) classification, the phylogeny places the

isotype of R. australirosea Murrill outside the clade representing R. subsect.

Lilaceinae, united by the presence of primordial hyphae, a brightly colored

pileus, and a pale spore print; unfortunately it is not possible at this time to place

R. australirosea in another group. In his type study, Singer (1958) characterized

R. australirosea with a completely mild taste and lacking dermatocystidia,

characters consistent with R. subsect. Lilaceinae but described the spore print

color as C to D, darker than expected for R. subsect. Lilaceinae. Singer (1958)

Sequence analyses of Murrill’s Russula types (U.S.A.) ... 263

proposed that R. australirosea might be conspecific with R. vinosirosea Murrill,

but we can reject this based on its phylogenetic placement.

Tree 2 (Urentes and Decolorantes) FIGURE 1

The R. subrubescens isotype and paratype form a clade with closely related

sequences, but the sequences share only a 94% ITS similarity and are separated

in two distinct clades with strong support. The BLAST results support the

R. subrubescens type as closely related to R. globispora (J. Blum) Bon while

placing the paratype in a clade with a sequence labeled “R. dryadicola” and

another R. globispora sequence. Singer (1986) did not treat either of these taxa

specifically, although he might have included them as varieties of R. maculata

Quél., united by acrid taste, ochre spore print, absence of iodoform odor,

and numerous dermatocystidia. Therefore, both entities should be included

in R. subsect. Urentes Maire, not in R. sect. Decolorantes (Maire) Singer as

represented by the type species R. decolorans (Fr.) Fr. Metadata associated with

environmental sequences suggest that R. subrubescens is distributed throughout

temperate North America as an associate of Quercus.

Tree 3 (Russula) FIGURE 2

Russula albimarginata Murrill and R. emeticiformis Murrill are both

recovered in a clade with R. vinacea Burl., which is separate from the European

representatives of R. krombholzii Shaffer’ labeled “R. atropurpurea”. Shaffer

(1970) noted that R. vinacea reportedly has a less acrid taste and stronger

yellowing flesh than R. krombholzii. The name R. albimarginata likely refers

to the pallid margin described by Murrill (1945a) and observed in fresh

collections (particularly in young fruitbodies) of R. vinacea. Singer's (1958)

proposed placement of R. emeticiformis in R. subsect. Russula as a synonym of a

nominal subspecies of R. emetica (Schaeff.) Pers. is not supported, as R. emetica

is distantly related in Russula stirps Emetica. Singer’s later (1975) placement of

R. emetica subsp. lacustris Singer in Russula stirps Atropurpurea does, however,

correspond with the current phylogenetic placement of R. emeticiformis. Both

R. albimarginata and R. emeticiformis should be considered later taxonomic

synonyms or closely related species to R. vinacea.

The isotype of R. fragiloides Murrill falls in a well-supported clade with

members of R. subsect. Russula stirps Russula. Singer (1958) suggested that

R. fragiloides is synonymous with R. emetica subsp. emeticella (Singer) Singer

[= R. emeticella (Singer) Romagn.], but a recent revision by Hampe et al. (2013)

‘Singer (1986: 824) used a heterotypic synonym, R. bresadolae Schulzer 1886, for the illegitimate

later homonym R. atropurpurea (Krombh.) Britzelm. 1893, non Peck 1888. However the

correct name for this taxon is Russula krombholzii, which was published as a replacement

name (nom. nov.) based on the legitimate synonym Agaricus atropurpureus Krombh. 1845,

and which therefore has priority over R. bresadolae.

264 ... Looney

found this taxon’s position to be uncertain due to confusion over assigning a

lectotype.

Tree 4 (Xerampelinae and Incertae sedis) FIGURE 2

Murrill (1945a,b) described R. levyana and R. pinophila Murrill with

unchanging context, no odor, and — in the case of R. pinophila — a white

gill color, features that would generally exclude these taxa from R. subsect.

Xerampelinae, although he regarded R. levyana as related to R. xerampelina

(Schaeff.) Fr. Adamcik & Buyck (2010), who evaluated the type and recent

collections of R. levyana, noted these inconsistencies but explained that some

American representatives of this group are known for having weak odors,

especially in young fruitbodies. Younger fruitbodies might also explain gill

whiteness, as R. subsect. Xerampelinae is characterized by ochraceous spore

prints that make mature gills appear yellow. Phylogenetically, R. levyana and

R. pinophila are in a well-supported clade with representatives of R. subsect.

Xerampelinae. These two species share a 98% ITS sequence similarity and may

represent the same species, a conclusion supported by their association with

pine (Adam¢ik 2010). Ifso, R. pinophila should be regarded as a later taxonomic

synonym of R. levyana. However, Buyck (pers. comm.) has suggested that

species delimitation in this particular group should use a sequence similarity

cutoff greater than 98%.

The isotype of R. subcremeiceps Murrill is closely related to R. cremeoavellanea

Singer, which Singer (1986) placed in R. subsect. Chamaeleontinae Singer based

on its eventual chocolate brown reaction to phenol, deep ochraceous spore

print, and mild taste. Murrill (1946) did not test specimens for their phenol

reaction, but described R. subcremeiceps as having a mild taste and pale yellow

spore print. Russula subcremeiceps is somewhat allied with two other groups:

R. subsect. Vinosinae Singer represented by R. vinosa Lindblad and R. subsect.

Integrae Maire represented by R. velenovskyi Melzer & Zvara (Singer 1986).

Both R. cremeoavellanea and R. vinosa are species that discolor, but Murrill

(1946) described R. subcremeiceps as having unchanging flesh. He also noted

that R. subcremeiceps closely resembled R. albidicremea Murrill but differed

by having closer unforked gills and rounder spores. Russula subcremeiceps is

a species widely distributed across eastern North America ranging from the

northeast U.S. to Mexico.

Morphologically, R. vinosirosea is distinguished by its rose color with pale

vinose tint and subequal stipe that is shorter than pileus width (Murrill 1943),

two characters consistent with R. subsect. Integrae sensu Singer, of which

R. paludosa Britzelm. is the type species. Although the isotype of R. vinosirosea

shares 92% identity with R. paludosa, as well as R. olivobrunnea Ruots. &

Vauras, R. olivina Ruots. & Vauras, and R. veternosa Fr., it cannot be confidently

assigned to a taxonomic group due to low phylogenetic resolution.

Sequence analyses of Murrill’s Russula types (U.S.A.) ... 265

Russula subsulphurea is distinguished by its large (9-10 cm) solitary habit,

pallid to yellow-tinted to slightly rosy pileus, and white spore print (Murrill

1945b). The phylogenetic placement of the paratype specimen is unknown, but

multiple environmental studies have frequently sampled R. subsulphurea or a

closely related species in eastern North America.

The isotype of R. brunneipes Murrill is closely related to R. umerensis McNabb

from New Zealand. These two species share many characters such as presence

of dermatocystidia, vinaceous pileus color, a 7.5-10 um spore size, and spores

ornamented by mostly isolated echinulate 0.5-1 um tall spines (Hesler 1960,

McNabb 1973, Murrill 1945a). The R. brunneipes isotype shares a 92% identity

with R. cuprea (Krombh.) J.E. Lange, R. vinososordida Ruots. & Vauras, and

R. vinosa, but taxonomic placement is impossible at this time.

Tree 5 (Amoeninae) FIGURE 3

Russula alachuana Murrill and R. variicolor Murrill form a well-supported

clade with other members of R. subsect. Amoeninae Singer. No reliable sequence

of R. amoena Quél. was available, but all included taxa are united by having

cheilocystidia without contents, no dermatocystidia, and a pale yellow to

yellow spore print (Singer 1986). Both species form well-supported subclades

separated from a well-sampled clade of specimens identified as R. mariae

Peck, indicating that these are discrete phylogenetic species. Morphologically,

R. alachuana is distinct by its large size, velvety pileus, and pulverulent stipe with

rosy hue (Murrill 1938). An environmental sample of R. alachuana or closely

related species has been sampled from Mexico. Murrill (1942) distinguished

R. variicolor, potentially a trans-continental species sharing a close genetic

affinity (99% similarity) to a Pinus pinaster-associated root tip sample from

Europe, by its many-colored pileus that includes greenish hues and a completely

white stipe.

Discussion

A molecular sequence of type material is an invaluable tool for modern

systematics because it gives the taxonomist somewhat more objective criteria

for naming phylogenetic groups and delimiting new species. The ITS region

has been suggested as a barcoding region for ectomycorrhizal fungi due

to the ease of its amplification and its ability to separate intraspecific from

interspecific taxa (Schoch et al. 2012). My study illustrates a major hurdle in

the revival of North American Russula taxonomy, which is that Murrill and his

predecessors, G.S. Burlingham and C.H. Peck, described most Russula species

in the 1910’s—1940’s. This means that North American type material is at the

cusp of what can be successfully sequenced, and given the large number of

species described and even more yet to be described, sequences of these taxa

can greatly aid the revival of North American Russula systematics.

266 ... Looney

In this study I was able to produce ITS data from type collections designated

by W.A. Murrill between 1938 and 1948 with a 66% success rate for collections

accessioned at TENN. However, only the ITS1 region was successfully

sequenced for a majority of Murrill’s type material, and there are questions as

to the value of using only ITS1 or ITS2 as a species barcode to separate closely

related species. Either ITS region has been considered adequate for delimiting

species in some groups of fungi, but this might not work as well in speciose

groups, where species delimitation is performed at a fine scale with closely

related taxa (Bengtsson-Palme et al. 2013).

Comparison of the isotype collection of R. levyana with FJ845433 (the

specimen that shares a 99% sequence similarity) highlights the problem of

using morphological or phylogenetic approaches separately in taxonomy.

Either we must accept morphological variation in widespread species that may

have been over-split and over-described or regard the ITS1 region as inadequate

for separating closely related Russula species. To overcome these limitations,

I suggest combining morphological recognition with molecular data to

designate recently collected material as epitypes for historical North American

Russula species. A good example can be found in R. mutabilis, where a partial

sequence of the type shows a 100% sequence similarity with a recent collection

from Texas that also agrees with the morphological diagnosis of the type.

Although partial sequence data may not allow for complete confidence in

species delimitation, we are able to use such data to infer placement in clades,

thereby supporting or refuting past classifications based on morphology. From

the eight sampled taxa that have never before been placed in a group, I can now

confidently place four: R. albimarginata in R. subsect. Russula, R. pinophila in

R. subsect. Xerampelinae, R. subcremeiceps in R. subsect. Chamaeleontinae, and

R. subrubescens in R. subsect. Urentes (TABLE 1). The results indicate a different

or refined placement of R. westii in R. subsect. Subcompactinae, and we now

have a more refined placement of R. fragiloides, R. alachuana, R. variicolor, and

R. emeticiformis within their subsections (TABLE 1). Two taxa, R. albimarginata

and R. emeticiformis, are proposed as possible synonyms with R. vinacea, and

R. pinophila may be considered a possible synonym of R. levyana. Knowledge

of group placement will aid in future systematic revisions, and those species not

yet referable to an identifiable clade may be placed with additional molecular

sampling and reliably identified sequences accessioned in GenBank and curated

databases like UNITE.

Acknowledgments

I would like to acknowledge the input of Dr. Slavomir Adam¢ik for consultation

on morphological comparisons, reading the manuscript, and providing access to a

bibliographic database of North American Russula. I would also like to thank the

TENN Herbarium for permission to sequence historical types. Also, I thank Dr. Shaun

Sequence analyses of Murrill’s Russula types (U.S.A.) ... 267

Pennycook for reviewing and giving comments on nomenclature. Finally, I wish to

thank Dr. Bart Buyck and Dr. Jorinde Nuytinck for reviewing the manuscript and

providing important feedback.

Literature cited

Adamé¢ik S, Buyck B. 2010. Re-instatement of Russula levyana Murrill as a good and distinct

American species of Russula section Xerampelinae. Cryptogamie. Mycologie 31(2): 119-135.

Adam¢ik S, Buyck B. 2011a. The species of Russula subsection Xerampelinae described by C.H.

Peck and Miss G.S. Burlingham. Cryptogamie, Mycologie 32(1): 63-81.

http://dx.doi.org/10.7872/crym.v32.iss1.2012.063

Adam¢ik S, Buyck B. 2011b. Type-studies in American Russula (Russulales, Basidiomycota): species

of subsection Decolorantinae described by H.C. Beardslee, G.S. Burlingham and W.A. Murrill.

Cryptogamie, Mycologie 32(3): 323-339. http://dx.doi.org/10.7872/crym.v32.iss3.2011.323

Adam¢ik S, Mitchell D, Buyck B. 2010. Russula ochrifloridana sp. nov., a new yellowish fishy Russula

from Florida and its comparison with R. grundii. Cryptogamie. Mycologie 31(4): 363-372.

Adam¢ik S, Carteret X, Buyck B. 2013. Type studies on some Russula species described by C.H. Peck.

Cryptogamie, Mycologie 34(4): 367-391. http://dx.doi.org/10.7872/crym.v34.iss2.2013.367

Altschul SE, Gish W, Miller W, Myers EW, Lipman DJ. 1990. Basic local alignment search tool. Journal

of Molecular Biology 215(3): 403-410. http://dx.doi.org/10.1016/S0022-2836(05)80360-2

Bengtsson-Palme J, Ryberg M, Hartmann M, Branco S, Wang Z, Godhe A, De Wit P, Sanchez-

Garcia M, Ebersberger I, de Sousa F. Amend A, Jumpponen A, Unterseher M, Kristiansson

E, Abarenkov K, Bertrand YJK, Sanli K, Eriksson KM, Vik U, Veldre V, Nilsson RH. 2013.

Improved software detection and extraction of ITS1 and ITS2 from ribosomal ITS sequences of

fungi and other eukaryotes for analysis of environmental sequencing data. Methods in Ecology

and Evolution 4(10): 914-919.

Birkebak JM, Mayor JR, Ryberg M, Matheny PB. 2013. A systematic, morphological and

ecological overview of the Clavariaceae (Agaricales). Mycologia 105(4): 896-911.

http://dx.doi.org/10.3852/12-070

Buyck B. 2007. A new initiative towards the study of Russula in the eastern USA. Pagine Micol.

27: 81-86.

Buyck B, Adam¢ik S. 2011a. Type studies of Russula species described by WA Murrill, 1. R. rosei-

isabellina, R. sericella, and R. obscuriformis. Mycotaxon 115: 131-144.

http://dx.doi.org/10.5248/115.131

Buyck B, Adam¢ik S. 2011b. Type studies in Russula subgenus Heterophyllidia from the eastern

United States. Cryptogamie, Mycologie 32(2): 151-169.

http://dx.doi.org/10.7872/crym.v32.iss2.2011.151

Buyck B, Adam¢ik S. 2013. Type studies in Russula subsection Lactarioideae from North America

and a tentative key to North American species. Cryptogamie, Mycologie 34(3): 259-279.

http://dx.doi.org/10.7872/crym.v34.iss2.2013.259

Buyck B, Adamcik S, Lewis D. 2008. Russula section Xerampelinae in Texas. Cryptogamie.

Mycologie 29(2): 121-128.

Buyck B, Bessette A, Adamcik S. 2011. Russula hixsonii Murrill, a rare and intriguing southern

species of uncertain systematic position, rediscovered in Georgia, USA. Cryptogamie,

Mycologie 32(4): 403-412. http://dx.doi.org/10.7872/crym.v32.iss4.2011.403

Gardes M, Bruns TD. 1993. ITS primers with enhanced specificity for basidiomycetes-

application to the identification of mycorrhizae and rusts. Molecular Ecology 2(2): 113-118.

http://dx.doi.org/10.1111/j.1365-294X.1993.tb00005.x

268 ... Looney

Gouy M, Guindon S, Gascuel O. 2010. SeaView version 4: a multiplatform graphical user interface

for sequence alignment and phylogenetic tree building. Molecular Biology and Evolution 27(2):

221-224. http://dx.doi.org/10.1093/molbev/msp259

Hampe F, Eberhardt U, Kleine J, Verbeken A. 2013. Russula rhodomelanea und die Russula-

emeticella-Frage. Deutsche Gesellschaft fir Mykologie 79(2): 377-403.

Hesler LR. 1960. A study of Russula types. Memoirs of the Torrey Botanical Club 21(2): 1-59.

Hesler LR. 1961. A study of Russula types, II. Mycologia 53(6): 605-625.

http://dx.doi.org/10.2307/3756461

Katoh K, Standley DM. 2013. MAFFT multiple sequence alignment software version 7:

improvements in performance and usability. Molecular Biology and Evolution 30(4): 772-780.

http://dx.doi.org/10.1093/molbev/mst010

Kibby G, Fatto R. 1990. Keys to the species of Russula in northeastern North America. Kibby-Fatto

enterprises.

Maddison D, Maddison W. 2005. MacClade 4: Analysis of phylogeny and character evolution.

Version 4.08a., Sinauer Assoc.

McNabb R. 1973. Russulaceae of New Zealand 2. Russula Pers. ex S.F. Gray. New Zealand Journal of

Botany 11(4): 673-730. http://dx.doi.org/10.1080/0028825X.1973.10430308

Murrill W. 1938. New Florida agarics. Mycologia 30(4): 359-371. http://dx.doi.org/10.2307/3754461

Murrill WA. 1941. More Florida novelties. Mycologia 33(4): 434-448.

http://dx.doi.org/10.2307/3754898

Murrill WA. 1943. More new fungi from Florida. Lloydia 6(3): 207-221.

Murrill WA. 1945a. More fungi from Florida. Lloydia 7(4): 175-189.

Murrill WA. 1945b. New Florida fungi. Quarterly Journal of the Florida Academy of Sciences 8(2):

171-189.

Murrill WA. 1946. More Florida fungi. Lloydia 8(4): 263-290.

MycoBank. 2013. Fungal databases: nomenclature and species banks. MycoSearch. http://www.

mycobank.org/ [Accessed March 2014]

Schoch CL, Seifert KA, Huhndorf S, Robert V, Spouge JL, Levesque CA, Chen W, Bolchacova E,

Voigt K, Crous PW. 2012. Nuclear ribosomal internal transcribed spacer (ITS) region as a

universal DNA barcode marker for Fungi. Proceedings of the National Academy of Sciences

109(16): 6241-6246. http://dx.doi.org/10.1073/pnas.1117018109

Shaffer RL. 1970. Notes on the subsection Crassotunicatinae and other species of Russula. Lloydia

33: 49-96.

Silvestro D, Michalak I. 2012. raxmlGUI: a graphical front-end for RAxML. Organisms Diversity &

Evolution 12(4): 335-337. http://dx.doi.org/10.1007/s13127-011-0056-0

Singer R. 1951. The “Agaricales” (mushrooms) in modern taxonomy. Lilloa 22: 832.

Singer R. 1958. New and interesting species of Basidiomycetes. V. Sydowia 11: 141-272.

Singer R. 1975. The Agaricales in modern taxonomy. Vaduz, J. Kramer.

Singer R. 1986. The Agaricales in modern taxonomy. Koeltz Scientific Books Koenigstein.

Stamatakis A, Hoover P, Rougemont J. 2008. A rapid bootstrap algorithm for the RAxML Web

servers. Systematic biology, 57(5): 758-771. http://dx.doi.org/10.1080/10635 150802429642

Thiers B. 2013 [continuously updated]. Index herbariorum: a global directory of public herbaria

and associated staff. New York Botanical Garden’s virtual herbarium.

http://sweetgum.nybg.org/ih/ [Accessed Nov 2013]

White TJ, Bruns T, Lee S, Taylor J. 1990. Amplification and direct sequencing of fungal ribosomal

RNA genes for phylogenetics. 315-322, in: MA Innis et al. (eds). PCR protocols: a guide to

methods and applications. Academic Press.

MY COTAXON

http://dx.doi.org/10.5248/129.269

Volume 129(2), pp. 269-282 October-December 2014

Molecular phylogeny of placodioid lichen-forming fungi

reveal a new genus, Sede/nikovaea

S.Y. KONDRATYUK"™, M.-H. JEONG’, I.A. GALANINA3,

L.S. YAKOVCHENKO*, A.P. YATSYNA’®, & J.-S. HUR?

'M.H. Kholodny Institute of Botany of National Academy of Sciences of Ukraine, Kiev, Ukraine

? Korean Lichen Research Institute, Sunchon National University, Sunchon, Korea

> Institute of Biology and Soil Science FEB RAS,

Stoletiya Vladivostoka Avenue 159, 690022 Vladivostok, Russia

“Botanical Garden-Institute FEB RAS, Makovskogo st. 142, 690024 Vladivostok, Russia

°V.E. Kuprevich Institute of Experimental Botany of National Academy of Sciences of Belarus,

Akademicheskaya str. 27, 220072 Minsk, Belarus

CORRESPONDENCE TO *: ksya_net@ukr.net

ABsTRACT — A molecular phylogeny based on ITS1/ITS2 and mtSSU DNA sequences is

presented for lichen-forming fungi in Protoparmeliopsis and its distant relative Sedelnikovaea

gen. nov. The positions of the two genera in the phylogenetic tree of the families Lecanoraceae

and Lecideaceae are discussed. The new combination Sedelnikovaea baicalensis (= Lecanora

baicalensis) is proposed. The unique Sedelnikovaea-type ascus is described and illustrated.

Sedelnikovaea baicalensis and Zwackhiomyces zarei are for the first time recorded from China.

Key worps — Biatora, Rhizoplaca, Protoparmelia, Candelariella, Steinia, Pycnora

Introduction

The lichen-forming fungal genus Protoparmeliopsis comprises a total of

ca. 44 species (Santesson 2004; Kondratyuk 2010; Kondratyuk et al. 2012,

2013 b). After Arup & Grube (2000) provided the first molecular data for

Protoparmeliopsis, five new species and 42 new combinations have been

proposed in the genus (Kondratyuk 2010; Kondratyuk et al. 2012, 2013b;

Santesson 2004).

Protoparmeliopsis is not currently classified in the Lecanoraceae (Lumbsch

& Huhndorf 2007, 2010) due, in part, to lack of molecular data and unresolved

relationships within Lecanora sensu lato. Sequences from only three species —

P. achariana (A.L. Sm.) Moberg & R. Sant., P. macrocyclos (H. Magn.) Moberg

270 ... Kondratyuk & al.

& R. Sant., and P. muralis (Schreb.) M. Choisy — are available in GenBank.

Several species recently described in Protoparmeliopsis (or recently combined

in the genus based on morphology; Kondratyuk 2010, Kondratyuk et al. 2012,

2013 b) are here included in a molecular analysis.

Material & methods

Specimens were prepared using standard microscopical techniques: specimens were

hand-sectioned under Nikon SMZ645 dissecting microscope, examined under Nikon

E200 and Olympus BX51 microscope, and photographed using the Olympus DP-Soft

photo program.

TABLE 1. Specimens included in the phylogenetic analysis

TAXON NAME COUNTRY / VOUCHER : ITS1/ITS2 sae mt

_Adelolecia pilati = Norway?, Ekman 3373(BG) oo AY567713

Wide vere ns) Ae ah other I, oo Norway?, Ekman 3373 (BG) ooo dc cccccssecsusssessaseeeein tt 300874

___Aspicilia caesiocinerea FER O) USER se ROW LCA RRC MRD. Ae Mi Wee ART Cke Lc) o- ant

_Aspicilia cimereg cc ccsseussesien Oe Oe 5 ce coe Wh aa, Mia A tee 2 Qe SOR Ie,

Pere, Mansell eRe al Soh ne PEON Me OP ieee ke cytes ser EOS (B99 Me yl ets oe

.Boreoplaca ultrafrigida os Cnt cssccssssssassesnsessssnsssnareedossstssansessssssesaseee at! QO26247

: AFTOL-ID 1702

: Russia

Circinaria CaNCAT CA cc ccsake SOMO cc ccccccssassessssseressereeesestee 057898 ec ccceccssssee

We PS Pee A oe. Fee SL a ie Rm eee ek ee

aerate A ct ee to Austria?/ Wilfling (GZU) oo AF332108 cen

Schau eet ee ee PTO TSS ge craecacttvcecthedate nat snestosarebcothnendis ee Re BO8T OM

_.Circinaria desertorum od. Pou sehen eR, eRe oon eR BS Oe on. ill oi arn

RAN Pe el tek SRS, See. es. PRUSSIA Wi. Ta creer atch tcate tale teda Wey st Stn Ua eeu OCOD

_Circinaria hispida cesses TAT ee mgs, OR Bh ee EU a ate SSP LAT oer Me, Rye

i SK A12, Ukraine, Kharkiv : KP059052

oblast, Dvorychansky district,

‘Korobchyno’ zakaznik, 19.vi.2013 M.

Kryvokhyzhaya (KW-L)

i SK A15, Ukraine, Kharkiv : KP059053

: oblast, Dvorychansky district,

‘Korobchyno’ zakaznik, 19.vi.2013 M.

Kryvokhyzhaya 2 (KW-L)

Fh WO AN I aa Ae LY Trey sccscccsssecsesnseccesnsecsvsnsecsnsnsecsneneesansesscesnnsesecsannnseei ved VL060722.

_ Parnoldia jurang cc ccscccssecbe USHOANE oy A oh to ee Nhl Bee GU074511

ypocenomyce scalaris AFTOLID 1025 oc ccccsssessssseseeusseeeon FIQ650632 i ccccccsssesseesssssee

et Ae RP OS SWI cl dec tlle petecethea Becta cts teanlisiomncisdeeas ph OO eats,

4 eel Aa, Aa Lae aes Pe United Kimgdom ic ccccccssssseseseeeete RZ 99187 Eo ccccceussseseen

seta A var ete blend Saket 8 Pion, B® Sweden, 3 6) 0 Fe Eh Ge a NL eee OOo 20s

Bt Pe ee eh Ate UE cat ee OEIC BAN 1 cacticn pan ie ca Pe A Mevpcec Bates See eee

I GPEWIGHOTMOCNSIS. i ¥A81 58 44 pence CANAMay |W, Ucn Ae, ok beh ea EE USS. aig! Be 6 cael.)

nett cs UE ns td oe van nade, Re RIESE IO” yo aly oF Pe PR TOOSON SOS i Ode

TE ee RBs WO BO eecaliaddy es te a Po Ee, aE 0Gl ooo §

_Hbecanora achrog. oreo py Mima oie a cadatleatd dnttnentensct UN QASTE Sh aac Waleed

: Australia : JN943719

Lecanora albescens : Poland?/ Kubiak Jan. 2009 (KRAM) JQ993727

Lecanora argentata : Thailand : JQ782704 :

Sedelnikovaea baicalensis gen. & comb. nov. ... 271

TAXON NAME : COUNTRY / VOUCHER ITS1/ITS2 eo oka mt

: : DNA

Lecanora bicincta Austria?/ U. Trinkaus 102 (GZU) AY541264

Lecanora caesiorubella : Australia ? JQ782672

Lecanora swartzii subsp.

caulescens

: Sweden?/ Wedin 6860 (UPS) : AY756401

: AFTOL-ID 589 DQ912275

bobothalligalphoplaca oP) USA subi stvsssuistoncehctcuracarirnisscna te I RBOOTBT ii sccchotmsacninned

Os emetic te heehee UUM Lee E a, ted ls ef OO os ae MULL ce

Lobothallia melanaspis oS NOPWay oo ccccccsssseessssseesassseesseseeransie dE 820524 Essen

Pion Py L ik Welt Wed Sie. SWEAEI a ccssssessssssevsnsseevsnsstsvesnnseevnnssecsssnssseseasessessassnssese tt V060688

Lobothallia radiosa dan SY ECOCID A Jom. marek gs. Ln en FEA cee am A lS

Switzerland DQ780274

Lobothallia recedens : Sweden : : HM060724

272 ... Kondratyuk & al.

TAXON NAME CouUNTRY / VOUCHER ITS1/ITS2 ARNE: oy

.Megaspora verrucosa, Austria? /1996, Trinkaus (GZU) 002 AF332120

Austria?/ Hafellner 48544 & Ivanova:

ees er Snes e a sunt CA) peed ee eae ae ange tae

Se: AP ish Rhee apnea Ne SSO CME pe. Nir ML eS 8, SRI. te EIUOUCS et

AME ee ee eee tt Te: ARRE Se RE ata Mert he Riese 5-0) 1 ones, Pewee adver MI) Oded LE,

__Protoparmelia atriseda RISBA AP res We This On ENG, C953 Wied KE SG2T IU Aion Suet -Feea)

=Protoparmelia badia = |, suistriay ee ee RBBO2O DP a cacneacea

Ue Fan cee: Sc ata cee hosed eacita ROMEO CN NOD 8 ae Se cele aa ot EU IOO2Ss 2 6 et ue

Vee Tee aa a8 RR cara. xo Bosh, ek sia te eMEL SULA gs Iolo eeews. 12h We Ba. 8S Sets, Cet, EEE BAS AF351179

: Austria ? EU075540

: ?/ Lumbsch s.n. (F)

: EF105420

Protoparmeliopsis zarei i SK 480, Iran, Esfahan Province,

i Mooteh Wildlife refuge, 50°39°56”E

33°25'34’'N, 2510 m alt., 21. VII.2010

B. Zarei-Darki (1111) (KW-L)

SK 481, Iran, Esfahan Province,

i Mooteh Wildlife refuge, 50°39°56”E

33°25'34’'N, 2510 m alt., 21. VII.2010

B. Zarei-Darki (1108) (KW-L)

Sweden, Hermansson 7903a (UPS) FJ959357

? China, Wei, 3, 8/5/2002 ? AY509799

: USA ? HM577302

: USA : HM577297

: Switzerland : JX948232

? Sweden?/ U519

:_ China, Guo, 3557, 8/1/2003 i AY509802

Rhizoplaca polymorpha : USA : JX948194

Sedelnikovaea baicalensis gen. & comb. nov. ... 273

TAXON NAME CouUNTRY / VOUCHER ITS1/ITS2 ea ay

_ Rhizoplaca portert oo ccctecca ti OM csccccccscssslssienrssefrteceeatcetaesecssntedi PE MIST7 381 ooo ccccesteete

Rhizoplaca shusharit cd ISA cc cccccccsssssesssssssassssseessesseessaseeesnstee tH MAS 77294 ee

__Rhizoplaca subdiscrepans —_, China, Wang, 13-2, 8/4/2002, AYS09789

__Scoliciosporum chlorococcum i, United Kingdom de PRII9329: ts en Bs.

Oe, A Le, Week ee tn Norway?, Ekman 3390 (BG) oo Ecsssseccsssssssesessseeetee ft 967768

.Scoliciosporum umbrinum Austria?, A. Wilfling 2873 (GZU) ood BN OMQ7L Pet wns 8 Gs

Fi Eo eae Oe co, atk 2g aE Norway?, Ekman 3005 (BG) od ccccsssccssssssessseedttY 967719

Rs eal A ee et A Norway?, Ekman 3005 (BG) oi csssscssssssseesesseeeetae ft 300911

Sedelnikovaea baicalensis KP059050 KP059057

42°01’011’N, 116°17’738’E, 1434 m

i alt., 8.viii2009 J.-S. Hur & X.Y. Wang :

SRT 5 SANT PRCT om By, PREG HOI0322, OLR OLOOFOS Wa. (Mi SAO SOP ORR,.. 0 E FeR Wen

: SK 776, China, Inner Mongolia, : :

i 42°01’011"N, 116°17°738"E, 1434m | KPOS9051 | KPO59058

alt., 8.viii.2009 J.-S. Hur & X.Y. Wang

CH090322 KoLRI 010936

i SK A18, China, Inner Mongolia,

i 43°36’272”N, 116°43’242”E, 1260

m alt., 10.viii.2009 J.-S. Hur & X.Y.

Wang CH090352 (KoLRI 010966)

' KP059059

_ Steinia geophang a ccccccssesie TATE Ga yh, og es RN Dee ome Ae ee een, ale JN222812

__Xanthomendoza mendozae i. hile y's. ys ee eee EU68135 1 i ccsccesssseeee

2, Me on eile ee, NRL bk Wn 10S Cente cssssssssssessssstsssstseuartessssseusesensiee HV681349 | EU680937

Rd eee ee ee Ae eee he AT gem oc cccccccssesssssseseseserseesssseeeeeeis. A U681350 : EU680938

__Xanthomendoza kashiwadani 3 (SSI Ad oe ge eA SRE Petes No ee EU680936

Xylopsora friesii United Kingdom FR799184

: Sweden : AY853324

Results

We submitted data on P muralis and P. zarei S.Y. Kondr. into GenBank for

the first time, as ITS rDNA and 12SSU mtDNA sequence analyses confirm both

species within Protoparmeliopsis. Phylogenetic analysis also shows that another

Asian species, P. baicalensis (Zahlbr.) S.Y. Kondr., is outside the genus and

very distant from Protoparmeliopsis. Our anatomical examination of material

of ‘Protoparmeliopsis’ baicalensis revealed that this taxon is characterized by a

unique ascus-type (PL. 3), which we henceforth refer to as the Sedelnikovaea-

type. Consequently we propose a new genus, Sedelnikovaea, for this species,

S. baicalensis.

Sedelnikovaea baicalensis and Zwackhiomyces zarei S.Y. Kondr. (KoLRI

10971, on Rhizoplaca chrysoleuca, see below under specimens examined) are

for the first time recorded from China.

Taxonomy & phylogeny

Protoparmeliopsis M. Choisy, Bull. Soc. Bot. France 76: 523 (1929).

TYPE SPECIES — Lichen muralis Schreb. [= Protoparmeliopsis muralis]

Thallus commonly distinctly placodioid. Apothecia lecanorine. Asci of

Lecanora-type. Ascospores hyaline, simple.

274 ... Kondratyuk & al.

Protoparmeliopsis comprises approximately 44 species. For a detailed

description of the genus and its history, see Kondratyuk (2010).

MOLECULAR PHYLOGENY — As a monophyletic group Protoparmeliopsis

has high bootstrap support based on results from nuclear (ITS1/ITS2)

and mitochondrial (12S SSU) DNA sequence analyses that included four

Protoparmeliopsis species (Fics 1, 2).

The status of such species as P. macrocyclos and P. garovaglii (K6rb.) S.Y. Kondr.

still requires further study.

Protoparmeliopsis is not generally accepted (Lumbsch & Huhndorf 2007,

2010), probably because the Lecanora s. lat. clade itself has a rather high

bootstrap support (see PL. 2). However, relationships among lineages within

Lecanora s. lat. remain largely unresolved, and improved sampling is required

to circumscribe genera within this group adequately.

A parallel case is the Dufourea s.lat. clade in Xanthorioideae, Teloschistaceae

(Kondratyuk et al. 2014b, and Fedorenko et al. 2009, 2012; Kondratyuk et al.

2013a as Xanthodactylon s.lat. clade). However within this clade Dufourea Ach.

s.str., Jackelixia S.Y. Kondr. et al., Ovealmbornia S.Y. Kondr. et al., Langeottia

S.Y. Kondr. et al., and Xanthokarrooa S.Y. Kondr. et al. have strong individual

bootstrap support values, with each monophyletic branch also characterized by

a unique set of morphological, anatomical, and biochemical characters.

Similarly ITS1/ITS2 nrDNA (PL. 1) and 12S SSU mtDNA (PL. 2) analyses

strongly support Protoparmeliopsis within the Rhizoplaca/Protoparmeliopsis

clade. The genus is probably as polyphyletic as Rhizoplaca Zopf (Arup & Grube

2000, Arup et al. 2007, Leavitt et al. 2011, 2013), Protoparmelia M. Choisy

(Lendemer & Lumbsch 2008, Papong et al. 2011, Sing et al. 2013), and Lecanora

Ach. (Grube & Blaha 2003, Grube et al. 2004, Lumbsch et al. 2012, Sliwa et al.

2012).

Sedelnikovaea S.Y. Kondr., M.H. Jeong & Hur, gen. nov.

MycoBAnk 810705

Differs from Protoparmeliopsis in having a unique type of ascus in some respects similar

to Candelariella- and Biatora-type, and by its close molecular similarity to members of

the Lecideaceae.

TYPE SPECIES — Lecanora baicalensis Zahlbr. [= Sedelnikovaea baicalensis]

EtymMo.Locy — honouring lichenologist Nellia Vasiljevna Sedelnikova (Novo-sibirsk,

Russia) in recognition of her contribution to our knowledge of Asian lichen flora.

Thallus distinctly placodioid, rosette-like, apothecia lecanorine, asci of

Sedelnikovaea-type, 8-spored; ascospores simple, hyaline.

TAXONOMIC POSITION — This new genus is proposed for the single species

Sedelnikovaea baicalensis, morphologically similar but only distantly related

to species in the putative genus Protoparmeliopsis. Molecular data (see

99 100

4) 100

100

100 an

104 9g 84

100 61

100 56.90

64] 27

= 100

A 99

61| 66

T00

65 a

19| Mean

99 100

100

3 100

98 [Lo

80 100

100

T00

¥ 10

Sedelnikovaea baicalensis gen. & comb. nov. ...

EU681349 X. mendoza

EU681350 X. mendoza Xanthomendoza

EU681351 X. mendoza

SK776 S. baicalensis 5

SK743 S. baicalensis Sedelnikovaea

so Pen

7 P. sorophora

eee ES uk pitioagee Pycnora

. scalaris

HQ650632 H. scalaris Hypocenomyce

ER799 1 84 x friesii Xylopsora

75549 R. stuartii °

EU075542 R. brunneocarpa Ramboldia

EU057905 © deserto

. desertorum ICL 7

EU057898 C. calcarea Circinaria

AF332108 C. contorta ey.

AP332129 M4, vernuc Puletens

. verrucosa

AF332121 M. verrucosa Megasp ora

eens i pipnopinee

737 L. alphoplaca i,

3F825524 L. melanaspis Lobothallia

eae ie radiosa

. tornoensis °

HQ650656 J. tornoensis J apewla

Ae oere - ee ort Lecanora fuscescens

EU . sanguinolenta

AF517930 P. quernea Pyrrhospora

AY541261 L. farinacea Lecanora

JN943723 L. flavopallida

JN943725 L. farinacea

JX948232 R. melanophthalma

HM577255 'L.' novomexicana

HM577381 R. porterii

HM577302 R. haydenii

HM577297 R. idahoensis

JX948227 R. parilis

JX948194 R. polymorpha

HM577307 R. occulta

HM577294 R. shushanii

HM577253 R. chrysoleuca

AY509799 R. chrysoleuca

AY509789 R. subdiscrepans

SK480 P. zarei

SK765 P. muralis

HQ650653 P. muralis

HM209239 P. muralis

AF070019 P. achariana

AF159933 P. macrocyclos

ee Ab s screws

5 7 R. peltata :

AY309802 R. peltata Rhizoplaca peltata gr.

AF070016 L. dispersoareolata

AY398703 L. intricata

DQ534470 L. polytropa

AF070018 L. pruinosa

JQ993727 L. albescens

JQ993733 L. dispersa

JQ993735 L. hagenii

JQ993755 L. semipallida

JQ993741 L. reuterii

JQ993725 L. crenulata

HQ650604 L. contractula

AF070025 L. perpruinosa

AY541247 L. carpinea

DQ451657 L. subcarpinea

DQ451670 L. rupicola

AY541264 L. bicincta

AY541272 L. sw. spp. caulescens

DQ451655 L. swarizii

AF070031 L. allophana

AF159939 L. allophana

AF159930 L. campestris

KF562194 P. picea

AF101279 P. psarophana

AF101277 P. montagnei

KF562191 P. badia

AF070023 P. badia

EF495164 P. ochrococca

JN943720 L. tropica

JN943718 L. tropica

JQ900618 L. kenyana

JF821184 P. isidiata Protoparmelia isidiata gr.

FR799323 S. chlorococcum ow

AY541277 S. umbrinum Scoliciosporum

Lecanora achroa gr.

farinacea gr.

Rhizoplaca

chrysoleuca gr.

Protoparmeliopsis

Lecanora

polytropa gr.

Lecanora

dispersa gr.

Lecanora

carpinea gr.

Lecanora

Protoparmelia

Lecanora tropica gr.

JN943719 L. achroa

JN943715 L. achroa

JQ782704 L. argentata

KF562192 P. cupreobadia Protoparmelia

KF562190 P. atriseda airiséda gr

HQ650707 L. fuscoatra gr.

HQ605929 L. fuscoatra Lecidea

HQ605927 L. fuscoatra

2/9

Pate 1. The phylogenetic tree of Protoparmeliopsis and Sedelnikovaea and representatives of

Lecanoraceae and Lecideaceae, based on the ITS1/ITS2 gene of nuclear DNA.

276 ... Kondratyuk & al.

100

A 100

100 be

100

100 100

89 86

100

100 83

91 98

100

100 ~

100

94 96

100

99 96

EU680937 X. mendoza

EU680938 X. mendoza Xanthomendoza

EU680936 X. kashiwadanii

SK 743 S. baicalensis Sedelnikovaea

SK 776 S. baicalensis

JN222812 S. geophana Steinia

AY853338 P. sorophora

AY853339 P. xanthococca P¥Chora

AY853325 H. scalaris

AY853326 H. scalaris

DQ912274 H. scalaris Hy pocenomyce

AY853324 X. friesii Xylopsora

HQ026247 B. ultrafrigida

AY853312 B. ultrafrigida Boreoplaca

DQ986813 B. ultrafrigida

DQ986892 A. caesiocinerea

DQ986890 A. cinerea A spicilia

DQ986876 C. contorta

HM060688 L. melanaspis

HM060724 L. recedens Lobothallia

DQ780274 L. radiosa

HM060722 C. hispida

SK Al2 C. hispida

SK A15 C. hispida —_ /

HM060689 C. desertorum Circinaria

AY853310 C. calcarea

HM060687 M. verrucosa

JQ797483 M. verrucosa Megaspora

JQ797482 M. verrucosa

HQ660567 L. fuscogrisea

AY756401 L. fuscoatra :

DQ912275 L. fuscoatra Lecidea

GU074511 F. jurana Farnoldia

AY464070 R. bullata

DQ787354 R. chrysoleuca Rhizoplaca

DQ787352 R. melanophthalma

SK 765 P. muralis

SK 480 P. zarei

SK 481 P. zarei

DQ787340 P. muralis

DG787342 P. achariana

DQ787350 L. valesiaca

DQ787356 L. sulphurea

DQ986898 L. contractula

JQ782672 L. farinacea TLecanora

JQ782667 L. caesiorubella farinacea gr.

JQ782670 L. farinacea

DQ787364 L. carpinea

Protoparmeliopsis

Lecanora

sulphurea gr.

AY567715 L. intumescens eeanaaek

AY300892 L. intumescens :

AF351179 P. badia a

BF105420 P. badia -Protoparmelia

AY567710 L. allophana

JQ782699 L. tropica

DQ787362 L. campestris Lecanora

DQ787360 L. glabrata

EF105417 L. hybocarpa Lecanora

EF105418 L. paramerae gangalaeoides gr.

JQ782676 L. gangalaeoides

AY567713 A. pilati

AY300874 A. pilati Adelolecia

HQ660559 J. tornoensis Japewia

AY567712 P. quernea

AY300908 P. quernea Pyrrhospora

JQ782674 L. flavopallida Lecanora

JQ782673 L. flavopallida flavopallida gr.

AY567719 S. umbrinum

AY300911 S. umbrinum Scoliciosporum

AY567768 S. chlorococcum

Pate. 2. The phylogenetic tree of Protoparmeliopsis and Sedelnikovaea and representatives of

Lecanoraceae and Lecideaceae, based on the 12S SSU gene of mitochondrial DNA.

Sedelnikovaea baicalensis gen. & comb. nov. ... 277

below) indicate a close relationship with the Lecideaceae or Megasporaceae.

Morphological and anatomical comparisons show important differences

from Protoparmeliopsis species, primarily a somewhat grey-brownish (instead

of greenish) thallus and possession of a unique Sedelnikovaea-type ascus,

described below.

Sedelnikovaea baicalensis (Zahlbr.) S.Y. Kondr., M.H. Jeong & Hur, comb. nov.

MycoBAnk 810706 PL. 3-4

= Lecanora baicalensis Zahlbr., Trav. Sous-Sect. Troitzkossawsk.-Khiakta,

Sect. Pays d'Amour, Soc. Imp. Russe de Géogr. 12: 85. 1911 [“1909”].

= Placolecanora baicalensis (Zahlbr.) Kopach., Nov. Sist. Niz. Rast. 9: 293. 1972.

= Protoparmeliopsis baicalensis (Zahlbr.) S.Y. Kondr.,

Ukr. Botan. Zhurn. 96 (6): 876. 2012.

Thallus crustose, rosette-like, up to 1-2.5 cm across, often forming larger

aggregations, rather thick, in section to 1 mm thick, densely attached to the

substrate, distinctly lobate in peripheral zone and sometimes areolate in the

centre, upper surface rough to slightly verruculose, matt to slightly shiny,

grayish brown to brownish, yellow-brownish or ochraceous-brown. Lower

surface dark brown to blackish-brown. Apothecia up to 1-1.5 mm diam.,

lecanorine (in section zeorine), mainly rounded, attenuated at the basis, disc

shiny at first, subconvex to convex when mature, light-brown to light reddish

brown; thalline margin narrow, entire to crenulate, sometimes disappearing in

senescence. True exciple up to 30 um thick in lateral portion and somewhat

indistinct in basal portion. Hymenium up to 50-70 um high, epihymenium

15-20 um thick, brownish. Subhymenium up to 100 um thick, underlying an

algal layer up to 50-80 um thick. Paraphyses rather lax. Asci 50-57 x (13-)

15-18(-20) um. Ascospores ellipsoid, 11-18 x 5-7 um. Thallus K-, C-, Pd+

slightly reddish.

SPECIMENS EXAMINED — RUSSIA. CHITA REGION (OBLAST), KYRINSKY DISTRICT, 2 km

SW of Chita-Khapcheranga junction, 49°45’21.40”N 112°20’08.60”E, 1184 m alt., shale

rocks, 28 July 2008 Yakovchenko (VBGI); Sokhondinskiy biosphere reserve, vicinity of

‘Buninda’ Forest Station, 49°42’20.46”N 111°21’55.36”E, 1294 m alt., experimental plot

31, Steppe S-facing rocky slope with granitoid stones, 24 July 2008 Yakovchenko (VBGI);

Sokhondinskiy biosphere reserve, vicinity of ‘Agutsa Forest Station, 49°39’57.16”N

111°25’39.11”E, 1130 m alt., Steppe S-facing rocky slope with Pinus sylvestris in the

upper part of the slope near the Station, 02 September 2009 Yakovchenko (VBGI);

Sokhondinskiy biosphere reserve, vicinity of ‘Agutsa Forest Station, 49°39’55.03”N

111°25’39.42”E, 1115 m alt., S-facing rocky slope with granitic gneissic rocks near the

Station, 20 July 2008 L. Yakovchenko (VBGI); “Gornaya Step” Protected Area, division

of Sokhondinskiy biosphere reserve, 14 km S of Kira Village, 49°24’N 112°03’E, 970

m. alt., vicinity of ‘Gazultiy’ Forest Station, ‘Sukhaya Padj’ locality, Multiherb prairie

with fragments of kharganat-type of vegetation on the S-facing rocky slope, 03 August

2005 Yakovchenko (VBGI); ZABAIKALSKY TERRITORY (KRAY), ONONSKIY DISTRICT,

road from Verkhniy Sharanay Village to Undino Posele Village, 3.5 km W of Zarya

278 ... Kondratyuk & al.

Village, 51°08’51.10”N 116°01’35.70’E, 606 m alt., steppe S-facing slope with shrubs

and rocks, 01 August 2008 Yakovchenko (VBGI); TsasucHEyskly DistTrRict, road

from Nizniy Tcasuchey to Aginskoe, 500 m N of the Bridge on Onon River, vicinity of

“Krasny Bator” nature monument, 50°31’45.00”N 115°01’19.10”E, 631 m alt., steppe

slope with granitic rocks, 30 July 2008 Yakovchenko (VBGI); AKsHINSKIy DISTRICT,

11 km NE of Mogoytuy Village and 10 km N of the junction to Nizniy Zasuchey,

50°24’35,10”N 113°58’04,40”E, 768 m alt., vegetation of kharganat-type (cereals -

Filifolium - herb steppe with Armeniaca sibirica) and shale rocks on S-facing slope,

28 July 2008 Yakovchenko (VBGI); KRASNOKAMENSKIY DisTRICT, 6.9 km NNE of

Tselinniy Village, 50°19’00.60’N 118°08’41.30”E, 710 m alt., Spurs of Klichkinskiy

Range, steppe top of spur with granitic gneissic rocks, 05 August 2008 Yakovchenko

(VBGI). CHINA. NEIMENGGU PROVINCE, DUOLUN COUNTY, Shisanlitan, 42°02.560’N

116°17.014’E, 1324 m alt., on rock, growing together with Caloplaca subsoluta, 2009

Hur CH090318 (KoLRI 10932); CH090321 (KoLRI 10935); CH090322 (KoLRI 10936);

42°01.011’N 116°17.738’E, 1434 m alt., on rock growing together with Protoparmeliopsis

muralis, 2009 Hur CH090332 (KoLRI 10946); X1LIN CouNTyY, Mountain, 43°36.272’N

116°43.242’E, 1260 m alt., on rock, 2009 Hur CH090347 (KoLRI 10961), CH090352

(KoLRI 10966); growing together with Candelariella sp. and Rhizoplaca chrysoleuca

damaged by Zwackhiomyces zarei, CH090357 (KoLRI 10971). UZBEKISTAN. Along

Dugave River, 23 August 1950 N. Shafeev 97 (MSK-L).

MOLECULAR PHYLOGENY — Our ITS rnDNA analyses (PL. 1) show

Sedelnikovaea baicalensis as closely related to Steinia Korb. and Pycnora

Hafellner (Lecideaceae or Lecanoromycetidae incertae sedis), while our mt

DNA analyses (PL. 2) place the species with Boreoplaca Timdal (Lecanoraceae)

Pate. 3. Asci of Sedelnikovaea type.

Sedelnikovaea baicalensis gen. & comb. nov. ... 279

Pate. 4. Sedelnikovaea baicalensis. a-d. General habit; e. Enlarged portion of thallus; f. Apothecia.

Scale bars: a-d = 3 mm; e, f = 1 mm.

and Lobothallia (Clauzade & Cl. Roux) Hafellner (Megasporaceae). Our

combined ITS + 12S mt SSU phylogenetic analysis suggest that Sedelnikovaea

is most closely related to genera representing several different families:

Boreoplaca (Lecanoraceae), Steinia (Lecideaceae or Aphanopsidaceae), Pycnora

(Lecideaceae), and Lobothallia (Megasporaceae). Unfortunately GenBank does

not contain both nuclear and mitochondrial sequences for the same species in

each of the four genera so that we can only infer that the new genus is unique.

Nonetheless, our data confirm that Sedelnikovaea is only distantly related to

Protoparmeliopsis.

280 ... Kondratyuk & al.

TYPE OF ascus — The Sedelnikovaea-type ascus shares some characters with

the Lecanora-, Candelariella-, and Biatora-type asci. The Sedelnikovaea-type

shares the strongly thickened apex with a K/I+ pale blue apical dome and a

broad K/I- apical cushion found in the Lecanora and similar Candelariella

types (Edwards et al. 2009). Furthermore, the apical cushion is surrounded

by a narrow, deeply K/I+ blue zone as seen in the Biatora-type (Printzen &

Coppins 2009) (Px. 3). Unlike the Candelariella-type, Sedelnikovaea asci

have only a narrow deeply K/I+ blue zone around the basal part of the apical

cushion, unlike the uniformly deep blue apical cushion of Candelariella. The

Biatora-type ascus is distinguished by a narrowly conical K/I- apical cushion

that contrasts with the broad cushion found in Sedelnikovaea.

As shown above, Sedelnikovaea baicalensis shows the highest affinity with

Steinia and Pycnora according to nuclear DNA and with Boreoplaca and

Lobothallia according to mitochondrial DNA. Pycnora and Boreoplaca are

characterized by a Lecanora-type ascus (Coppins 2009), while Megaspora

and Lobothallia are characterized by the Biatora-ascus type (James & Fletcher

2009). Among the genera mentioned, Steinia alone possesses a thin-walled

Aphanopsis-type ascus (Fletcher et al. 2009). According to Wolseley & Purvis

(2009) the Aphanopsis ascus is similar to the Trapelia-type, but this possibly

requires confirmation with fresh material. Aphanopsis and Steinia represent

a separate family, Aphanopsidaceae. Finally, we should note that correlation

between ascus type and DNA-phylogeny is generally poor in the Lecanorales

(Ekman et al. 2008).

Our data on Russian, Uzbek, and Chinese material of Sedelnikovaea

baicalensis completely agrees with the description published in the ‘Handbook

of the Lichens of the USSR’ (Kopachevskaya 1971, as Placolecanora baicalensis),

with the exception that the asci we measured were much wider: (13-)15-18

(-20) um vs. 8.5-10.2 um according to Kopachevskaya (1971).

Sedelnikovaea baicalensis usually grows on open, well-illuminated rock

surfaces where it is often associated with Dimelaena oreina, Lecanora frustulosa,

Pleopsidium chlorophanum, Aspicilia transbaicalica, Aspicilia cinerea,

Candelariella vitellina, and Rhizoplaca chrysoleuca.

Additional molecular analyses of Protoparmeliopsis and Sedelnikovaea will

most likely reveal further species that belong in Sedelnikovaea.

Acknowledgements

We would like to express our deep thanks to Dr. A. Thell (Lund, Sweden) for kind

help with some literature and herbarium specimens as well as for correction of English;

to Dr. A.B. Bochka and Mrs M. Kryvokhyzhaya (Kharkiv, Ukraine), Dr. B. Zarei-

Darki (Tehran, Iran) for providing herbarium specimens. The authors are grateful to

Dr. Steven Leavitt, Dr. Arne Thell, Dr. Shaun Pennycook, and Dr. Lorelei L. Norvell

Sedelnikovaea baicalensis gen. & comb. nov. ... 281

for their valuable comments on manuscript. This work was supported in part by The

State Agency on Science, Innovations and Information of Ukraine (M317-2011-409 and

M111-2012-409) for SK, The Russian Foundation for Basic Research (No. 13-04-01453)

for LY, and by The Forest Science & Technology Projects (No. $111212L030100) provided

by the Korea Forest Service for JSH.

Literature cited

Arup U, Grube M. 2000. Is Rhizoplaca (Lecanorales, lichenized Ascomycota) a monophyletic genus?

Canadian Journal of Botany 78: 318-327. http://dx.doi.org/10.1139/b00-006

Arup U, Ekman S, Grube M, Mattsson JE, Wedin M. 2007. The sister group relation of Parmeliaceae

(Lecanorales, Ascomycota). Mycologia 99: 42-49. http://dx.doi.org/10.3852/mycologia.99.1.42

Coppins BJ. 2009. Pycnora Hafellner. 771-772, in: OW Purvis et al. (eds). The lichen flora of Great

Britain and Ireland. London. Nat. Hist. Mus. Publ.

Edwards B, Aptroot A, Hawksworth DL, James PW. 2009. Lecanora Ach. in Luyken. 465-502, in:

OW Purvis et al. (eds). The lichen flora of Great Britain and Ireland. London. Nat. Hist. Mus.

Publ.

Ekman S, Andersen HL, Wedin M. 2008. The limitations of ancestral state reconstruction and the

evolution of the ascus in the Lecanorales (lichenized Ascomycota). Syst. Biol. 57(1): 141-156.

http://dx.doi.org/10.1080/10635150801910451

Fedorenko NM, Stenroos S, Thell A, Karnefelt I, Kondratyuk S. 2009. A phylogenetic analysis

of xanthorioid lichens (Teloschistaceae, Ascomycota) based on ITS and mtSSU sequences.

49-84, in: A Thell et al. (eds). Diversity of Lichenology - anniversary volume. Bibliotheca

Lichenologica 100. J. Cramer, Berlin, Stuttgart.

Fedorenko NM, Stenroos S, Thell A, Karnefelt I, Elix JA, Hur J-S, Kondratyuk SY. 2012. Molecular

phylogeny of xanthorioid lichens (Teloschistaceae, Ascomycota), with notes on their morphology.

58-76, in: I Karnefelt et al. (eds). Systematics, biodiversity and ecology of lichens. Bibliotheca

Lichenologica 108. J. Cramer, Stuttgart.

Fletcher A, Aptroot A, Purvis OW. 2009. Steinia Korb. 856, in: OW Purvis et al. (eds). The lichen

flora of Great Britain and Ireland. London. Nat. Hist. Mus. Publ.

Grube M, Blaha J. 2003. On the phylogeny of some polyketide synthase genes in the lichenized

genus Lecanora. Mycol. Res. 107: 1419-1426. http://dx.doi.org/10.1017/S0953756203008724

Grube M, BalochE, Arup U. 2004. A phylogenetic study of the Lecanora rupicola group (Lecanoraceae,

Ascomycota). Mycol. Res. 108: 506-514. http://dx.doi.org/10.1017/S0953756204009888

James PW, Fletcher A. 2009. Megaspora (Clauzade & Cl. Roux) Hafellner & V. Wirth. 568-569, in:

OW Purvis et al. (eds). The lichen flora of Great Britain and Ireland. London. Nat. Hist. Mus.

Publ.

Kondratyuk SY. 2010. Protoparmeliopsis M. Choisy. 130-149, in: SY Kondratyuk, OG Roms (eds).

Flora of the lichens of Ukraine 2(3). Kiev: Nauk. dumka.

Kondratyuk SY, Zarei-Darki B, Khajeddin SJ. 2012. New species and combinations in the genus

Protoparmeliopsis (Lecanoraceae, Lichenized Ascomycota). Ukr. Botan. Zhurn. 96(6): 869-879.

Kondratyuk SY, Fedorenko NM, Jeong MH, Yu N-N, Stenroos S, Karnefelt I, Thell A, Elix JA, Kim

J, Kondratyuk AS, Hur J-S. 2013a. Molecular phylogeny and exchanges of current taxonomy of

lichen family Teloschistaceae (Teloschistales, Ascomycota). 5-74, in: Problems of experimental

botany. Nine Kuprevych reading. Minsk: Division of biological sciences of NAS of Belarus.

Kondratyuk S, Lé6k6s L, Tschabanenko S, Haji Moniri M, Farkas E, Wang XY, Oh S-O, Hur J-S.

2013b. New and noteworthy lichen-forming and lichenicolous fungi. Acta Botanica Hungarica

55(3-4): 275-347. http://dx.doi.org/10.1556/ABot.55.2013.3-4.9

282 ... Kondratyuk & al.

Kondratyuk S, Jeong M-H, Yu N-H, Karnefelt I, Thell A, Elix JA, Kim J, Kondratyuk AS,

Hur J-S. 2014a. A revised taxonomy for the subfamily Caloplacoideae (Teloschistaceae,

Ascomycota) based on molecular phylogeny. Acta Botanica Hungarica 56 (1-2): 93-124.

http://dx.doi.org/10.1556/ABot.56.2014.1-2.10

Kondratyuk S, Karnefelt I, Thell A, Elix JA, Kim J, Jeong M-H, Yu N-H, Kondratyuk AS, Hur

J-S. 2014b. A revised taxonomy for the subfamily Xanthorioideae (Teloschistaceae, Ascomycota)

based on molecular phylogeny. Acta Botanica Hungarica 56(1-2): 141-178. http://dx.doi.

org/10.1556/ABot.56.2014.1-2.12

Kopachevskaya YG. 1971. Genus Placolecanora. 89-136, in: Handbook of the lichens of the USSR.

Issue 1. Leningrad: Nauka, Leningrad devision.

Leavitt SD, Fankhauser JD, Leavitt DH, Porter LD, Johnson LA, St Clair LL. 2011. Complex patterns

of speciation in cosmopolitan ‘rock posy’ lichens - Discovering and delimiting cryptic fungal

species in the lichen-forming Rhizoplaca melanophthalma species-complex (Lecanoraceae,

Ascomycota). Mol. Phylogenet. Evol. 59(3): 587-602.

http://dx.doi.org/10.1016/j.ympev.2011.03.020.

Leavitt SD, Fernandez-Mendoza F, Perez-Ortega S, Sohrabi M, Divakar PK, Vondrak J, Lumbsch

HT, St. Clair LL. 2013. Local representation of global diversity in a cosmopolitan lichen-

forming fungal species complex (Rhizoplaca, Ascomycota). J. Biogeogr. 40(9): 1792-1806.

http://dx.doi.org/10.1111/jbi.12118

Lendemer J, Lumbsch T. 2008. Protoparmelia capitata sp. nov. and Protoparmelia

isidiata Diederich, Aptroot & Serusiaux, two species of Protoparmelia (Lecanorales,

Ascomycota) from southeastern North America. Lichenologist 40(4): 329-336.

http://dx.doi.org/10.1017/S0024282908007810

Lumbsch HT, Huhndorf S. 2007. Outline of Ascomycota - 2007. Myconet 13: 1-58.

Lumbsch HT, Huhndorf SM. 2010. Myconet Volume 14. Part one. Outline of Ascomycota - 2009.

Part two. Notes on ascomycete systematics. Nos. 4751-5113. Fieldiana Life and Earth Sciences

1: 1-64. http://dx.doi.org/10.3158/1557.1

Lumbsch T, Parnmen S, Papong K. 2012. Two new species of Lecanora sensu stricto (Lecanoraceae,

Ascomycota) from east Africa. MycoKeys 3: 37-47. http://dx.doi.org/10.3897/mycokeys.3.3201

Papong K, Kantvilas G, Lumbsch TH. 2011. Morphological and molecular evidence places

Maronina into synonymy with Protoparmelia (Ascomycota: Lecanorales). Lichenologist 43(6):

561-567. http://dx.doi.org/10.1017/S0024282911000284

Printzen C, Coppins BJ. 2009. Biatora Fr. 213-216, in: OW Purvis et al. (eds). The lichen flora of

Great Britain and Ireland. London. Nat. Hist. Mus. Publ.

Santesson R. 2004. The lichens and lichenicolous fungi of Sweden and Norway. Lund: SBT-forlaget.

309 p.

Singh G, Divakar PK, Dal Grande F, Otte J, Parnmen S, Wedin M, Crespo A, Lumbsch HT, Schmitt

I. 2013. The sister-group relationships of the largest family of lichenized fungi, Parmeliaceae

(Lecanorales, Ascomycota). Fungal Biol. 117(10): 722-730.

http://dx.doi.org/10.1016/j.funbio.2013.08.001

Sliwa L, Miadlikowska J, Redelings BD, Molnar K, Lutzoni F. 2012. Are widespread morphospecies

from the Lecanora dispersa group (lichen-forming Ascomycota) monophyletic? Bryologist

115(2): 265--277.

Wolseley PA, Purvis OW. 2009. Aphanopsis Nyl. Ex. P. Syd. 151, in: OW Purvis et al. (eds). The

lichen flora of Great Britain and Ireland. London. Nat. Hist. Mus. Publ.

MY COTAXON

http://dx.doi.org/10.5248/129.283

Volume 129(2), pp. 283-292 October-December 2014

A new species of Leptostroma on Pinus henryi from China

LAN ZHANG}, D.W. MINTER’, QING LI’, & YING-REN LIN?

' School of Life Science &°* School of Forestry & Landscape Architecture,

Anhui Agricultural University, West Changjiang Road 130, Hefei, Anhui 230036, China

? CAB International, Bakeham Lane, Egham, Surrey, TW20 9TY, UK

*CORRESPONDENCE TO: yingrenlin@yahoo.com

ABSTRACT — A coelomycetous fungus found on dead fallen secondary needles of Pinus

henryi from Shennongjia forestry region of Hubei Province, China, is described, illustrated,

and discussed. Habitat, morphology, and conidial development place it in the Rhytismataceae.

It differs from other members of this family on the same substratum by having conidiomata

with upper walls more than one cell thick, and is formally named as Leptostroma magnum sp.

nov. Its conservation status is evaluated as Data Deficient. The type specimen is deposited in

the Reference Collection of Forest Fungi of Anhui Agricultural University, China (AAUF).

KEY worDs — anamorph, anatomy

Introduction

During a study of microscopic fungi associated with trees in the Shennongjia

forestry region (Hubei, China) in 2010, an interesting coelomycetous species

was found on dead fallen needles of Pinus henryi. The fungus was collected

again in repeat visits to the same site made in 2012 and 2013. It is described,

illustrated, and discussed below.

Materials & methods

Specimens were collected from the leaf litter of Pinus henryi trees, which were

more than 20 years old in 2010. All collections were made at the same location, at an

altitude of 1320 m, in Shennongijia forestry region, Hubei, China, in July 2010, October

2012, and April 2013. Specimens were deposited in in the Reference Collection of

Forest Fungi of Anhui Agricultural University, Hefei, China (AAUF). For microscopic

examination, only needles with sporulating conidiomata were used. External features

and arrangement of the conidiomata and associated zone lines were observed with a

dissecting microscope at 10-50x magnification. Specimens were rehydrated in water

for 10 minutes, and 10-15 um thick vertical transverse sections of conidiomata were

284 ... Zhang & al.

made using a freezing microtome (YD-202, China) and mounted in 0.1% (w/v) cotton

blue with lactic acid. Measurements and drawings of conidiogenous cells and conidia

were made from squash mounts in 5% KOH, with more than 30 conidiogenous cells and

conidia measured for each specimen. Colors of the various structures were observed

in water. Illustrations of external shapes and internal structures of the conidiomata

were prepared using the Panasonic XSJ-2 microscope drawing device. Comparisons

with other similar species were based on protologue descriptions and illustrations, and,

where possible, examination of type material.

Taxonomy

Leptostroma magnum Y.R. Lin & Lan Zhang, sp. nov. FIGS 1,.2

MycoBank MB805599

Differs from other species of the Rhytismataceae on dead fallen secondary needles of

diploxylon pines by having subcuticular conidiomata with upper walls more than one

cell thick.

Type: China, Hubei, Shennongjia forestry region, Guanmenshan, alt. 1320m, on dead

needles of Pinus henryi Mast. (Pinaceae), 10 July 2010, Y.R. Lin, J.L. Chen & Q. Zheng

2455 (Holotype, AAUF 68563).

ETYMOLOGY: magnum (Latin = large), referring to the size of the conidiomata.

ZONE LINES black, thin, frequent, with the black fungal tissue penetrating

the stele. Conrpromata on both surfaces of the needle, though usually much

more frequent on the abaxial side, scattered, sometimes confluent in groups

of two or three (especially longitudinally), 350-740 x 110-240 um, oblong or

elliptical, rugged, with a clearly marked outline, shiny black, slightly raising

the substratum surface, opening by at least four more or less circular ostioles

and one to several irregular tears in the upper wall. IN VERTICAL TRANSVERSE

SECTION located underneath the needle cuticle and above the epidermal cells

which are largely intact and yellow-brown to brown, with some hypodermal

cells invaded by dark grey-brown angular fungal hyphae 2.5—4.5 um diam. and

others strongly brown perhaps as a result of accumulated tannins; somewhat

domed, 70-80 um deep, with an upper wall 8.5-13 um thick, composed of

blackish-brown angular-aliform thick-walled cells below the needle cuticle,

becoming slightly pale (yellow-brown) and thin-walled towards the edge

and not connecting to the basal wall which is extremely poorly developed.

SUBCONIDIOGENOUS LAYER colourless, 6—9.5 um thick, consisting of textura

angularis composed of cells 2—3.5 um diam. with rather thick walls; a similarly

structured but grey-brown semicircular raised area 40—70 um wide sometimes

also present in the centre. CONIDIOPHORES not observed. CONIDIOGENOUS

CELLS colourless, thin-walled, smooth, either ampulliform or cylindrical

and tapering towards the apex, 12.5-20 x 2.5—4 um, usually proliferating

sympodially, rarely percurrently. Conrp1a 7.5-11.5 x 0.9-1.3 um, colourless,

Leptostroma magnum sp. nov. (China) ... 285

4 ‘ e ‘

' |

, P

FIGURE 1. Leptostroma magnum (ex holotype AAUF 68563): a. Conidiomata on a needle.

b. Detail of conidiomata. c. Conidioma in vertical section. d. Opening conidioma in vertical section.

e. Conidiogenous cells, conidia, subconidiogenous layer and basal wall. f. Enlargement of conidia.

Bars: a, b = 500 um; c, d = 20 um; e, f= 10 um.

286 ... Zhang & al.

FIGURE 2. Leptostroma magnum (ex holotype AAUF 68563): a. Habit on needles. b. Detail

of conidiomata and a zone line. c. Conidiomata in vertical section. d. Conidiogenous cells

and conidia. Bars: a = 10 mm;, b = 250 um; c = 50 um; d = 10 um.

aseptate, smooth, cylindrical, sometimes slightly curved, obtuse or round at the

ends. TRICHOGYNES not observed. TELEOMORPH not observed.

Eco.ocy: The fungus was observed on 2-3-year old secondary needles,

which had turned yellow-brown to straw yellow, died, and fallen into the litter.

ADDITIONAL SPECIMENS EXAMINED: On Pinus henryi: CHINA, HuBe1, Shennongjia

forestry region, alt. 1320 m, October 2012, L. Zhang & S.J. Wang 2864 (AAUF 68972);

April 2013, Y.R. Lin & Q. Li 2971 (AAUF 69079).

Leptostroma magnum sp. nov. (China) ... 287

Discussion

From their habitat and general form, there can be little doubt that the zone

lines and conidiomata in these collections are of a member of the ascomycete

family Rhytismataceae Chevall. These fungi are seen on dead leaves, twigs, and

sometimes other woody material of a wide range of plants. Some are plurivorous.

Others are associated with only one or a few closely related species or genera of

plants. Many exist as symptomless endobionts in living plant tissues. Some may

be mutualistic. Others can be parasitic. A few are pathogens. Fruitbodies are

only seen after death of colonized plant tissue. Zone lines defining the extent

of individual colonies occur in many species. In some only a teleomorph is

observed. In many, perhaps most, an anamorph precedes and accompanies the

teleomorph. In a few an anamorph is produced but no teleomorph. Species of

this family occurring on pines are generally not found on other plants.

The biology of this fungus poses interesting questions. Why is it only

known as an anamorph? Is it possible that the teleomorph has simply not yet

been found? In many of the Rhytismataceae on pine needles, the anamorph is

produced before the teleomorph and, at the correct time of year, the anamorph

can be seen but not the teleomorph. The time in this condition is generally

rather short. Ascomatal initials usually appear within a month of the first

conidiomata appearing (Minter 1977). The three collections of the current

species were made at very different times of year, and none had evidence of

a teleomorph, so this may be a species with no teleomorph. That would have

significant implications for the fungus.

In general, for pine needle inhabiting members of the Rhytismataceae,

ascomata on dead needles release ascospores, which disperse and colonize

young living needles. The fungus then lives there as a parasite or symptomless

endobiont until death of the colonized tissues. Then new conidiomata

and/or ascomata form, completing the life cycle. Most anamorphs of the

Rhytismataceae are thought to have a spermatial function (Jones 1935): they are

there for sexual reproduction, not dispersal. The conidiomata, conidiogenous

cells, and conidia of the present fungus look very similar to many of those

anamorphs. If this fungus does not produce ascomata (the usual dispersal state

in the Rhytismataceae), then what function do these conidiomata have, and

how does the fungus colonize new substrata? The search for answers only raises

further questions. Does this fungus only occur on P. henryi, or is it also on

other pines nearby? If so, perhaps there is some reason why the fungus only

produces conidiomata on P. henryi (for example some interaction within the

needle may suppress teleomorph formation), and perhaps the fungus produces

the teleomorph on those other pines. Alternatively, the fungus may occur only

288 ... Zhang & al.

on P. henryi, and may as a result have evolved different strategies to spread.

One possible way might be if the fungus were systemic within the tree, ie. not

just in the needles but also in living twigs. At least one pine needle inhabiting

member of the Rhytismataceae, Elytroderma deformans (Weir) Darker, has

adopted that strategy (Lightle 1954). Such a fungus could at least theoretically

spread as a symptomless endobiont of seeds. It is clear that the current fungus

offers several interesting lines of investigation.

To establish its identity, the current fungus must be compared with other

known species. Many members of the Rhytismataceae are known from pines,

and more than 40 species have been observed on needles. Some fruit on dead

portions of still attached living needles (species of the genera Bifusella Hohn.,

Canavirgella W. Merr. et al., Davisomycella Darker, Elytroderma Darker,

Lophodermella Hohn., Ploioderma Darker, Soleella Darker). The present fungus,

collected on dead fallen needles is very unlikely to be one of them.

Some of the litter-inhabiting species have no known anamorph

(Lophodermium canberrianum W. Stahl ex Minter & Millar, L. durilabrum

Darker, L. erlangshanense Y.G. Liu, L. indianum Suj. Singh & Minter, L. macci

Sokolski & Bérubé, L. maximum B.Z. He & D.Q. Yang, L. puerense C.L. Hou

& M. Piepenbr., L. sichuanense D.X. Qiu & Y.G. Liu, Meloderma sharmarum

P.E Cannon & Minter). The present fungus, being known only as an anamorph,

is also unlikely to be one of these species.

Although exceptions are known, there is a tendency for species of the

Rhytismataceae on pine needles to occur on either haploxylon pines (i.e.

species with one vascular bundle per leaf) or diploxylon pines (species with two

vascular bundles per leaf), but not both (Minter 1977). As the current fungus

was found on a diploxylon pine, it is likely to differ from species known on

haploxylon pines (Lophodermium anhuiense Y.R. Lin, L. confluens Y.R. Lin et

al., L. ellipticum Y.R. Lin, L. himalayense P.F. Cannon & Minter, L. kumaunicum

Minter & M.P. Sharma, L. mirabile Y.R. Lin, L. nitens Darker, L. orientale

Minter, L. pini-bungeanae Y.R. Lin, L. pini-excelsae S. Ahmad, L. pini-sibiricae

C.L. Hou & S.Q. Liu).

Of the species known to have an anamorph and to occur on secondary

needles of diploxylon pines, the following have conidiomata immersed beneath

the epidermis: Leptostroma pineae (Bubak) Arx, Lophodermium australe

Dearn., L. baculiferum Mayr, L. conigenum (Brunaud) Hilitzer, L. guangxiense

Y.R. Lin, L. iwatense Sakuyama, L. pinastri (Schrad.) Chevall., L. ravenelii Minter,

L. seditiosum Minter et al., L. staleyi Minter, L. yuexiense C.L. Hou et al. As the

depth of embedding of conidiomata is a rather constant character at species

level in the Rhytismataceae (Minter 1980), all of those species are also likely

to be different from the current fungus, which has subcuticular conidiomata.

Leptostroma magnum sp. nov. (China) ... 289

There remain four members of the Rhytismataceae known to produce

subcuticular conidiomata on fallen secondary needles of diploxylon pines:

Lophodermium molitoris Minter, L. pini-mugonis C.L. Hou & M. Piepenbr.,

L. yanglingense Z.M. Cao & C.M. Tian, Meloderma desmazieresii (Duby)

Darker. All of them have conidiomata with either no upper wall, or with an

upper wall only one cell thick. This is another rather constant species-level

character in conidial states of the Rhytismataceae, and distinguishes those four

species from the present fungus that produces conidiomata with upper walls

several cells thick.

A few other names in the Rhytismataceae have been used for species

inhabiting pine needles: Cryocaligula hedgcockii (Dearn.) Minter, Leptostroma

ahmadii Petr., L. austriacum Oudem., L. durissimum Cooke, L. decipiens Petr.,

L. pinorum Sacc., L. rostrupii Minter, L. strobicola Hilitzer. These are all in

genera traditionally considered as anamorphic, and are all known synonyms

of species already discussed above. The present fungus thus seems different

from all previously described members of the Rhytismataceae known from

pines, because of its unique combination of characters: it inhabits dead fallen

secondary needles of a diploxylon pine, producing subcuticular conidiomata

where the upper wall is more than one cell thick.

Fallen needles of pine are known to be a hotspot substratum for sympatric

evolution of the Rhytismataceae (Burnett 1983): they provide a remarkable

range of habitats (the location of a needle on the tree can be significant, needles

may be retained by a tree for different numbers of years, and they may die

for different reasons). Subtle morphological variations can provide clues to

the presence of different species, and molecular evidence is starting to show

cryptic speciation where there is little or no morphological variation. There is

also an increasing body of evidence that some members of the Rhytismataceae

are highly specific in their plant associations. It is not surprising therefore to

encounter new species in this environment.

Unfortunately, it is generally impossible to predict the teleomorphic genus

to which conidiomata belong purely from appearance. Molecular techniques

may help to resolve this dilemma. At least one species on pine needles recently

found and known only as an anamorph (with subepidermal conidiomata) is

being described in Lophodermium on the basis of molecular evidence (Koukol

& Pusz, pers. comm.). Without molecular information, the only realistic option

still remains to place the species in an anamorphic genus.

There are several anamorphic genera in the Rhytismataceae. Those accepted

in the tenth edition of Ainsworth & Bisby’s Dictionary of the Fungi (Kirk et

al. 2008) are Conostroma Moesz, Crandallia Ellis & Sacc., Cryocaligula Minter,

Hysterodiscula Petr., Leptostroma Fr., Melasmia Lév., and Tryblidiopycnis Hohn.

290 ... Zhang & al.

Conidiogenous cells in all these genera are more or less compatible with the

present species in shape, colour, size, and developmental characteristics. Ways

in which the genera differ are reviewed in the following paragraph.

The type of Conostroma is the anamorph of Colpoma quercinum (Pers.)

Wallr. It characteristically produces rather large multiloculate stromatic

conidiomata in bark on twigs of Quercus (Sutton 1980). The type of Crandallia

is the anamorph of Bifusella acuminata (Ellis & Everh.) Bonar & W.B. Cooke.

It produces multiloculate conidiomata with an upper wall only one cell thick

on dead leaves and dead portions of living stems of Juncus (Sutton 1980).

The type of Cryocaligula is the anamorph of Ploioderma hedgcockii (Dearn.)

Darker. It produces conspicuous unilocular conidiomata with upper walls

several cells thick on dead portions of attached living needles of Pinus. It is also

unusual in having large 1-septate characteristically snowshoe-shaped conidia

with possibly a dispersal function, rather than the small aseptate conidia with

a spermatial function found in most other anamorphs of the Rhytismataceae

(Minter 1985). Hysterodiscula was originally described with two species, H.

empetri Petr. and H. kalmiae Petr. Index of Fungi (1947) selected H. empetri as

the generic type, and this choice was endorsed by Sutton (1977, 1980). Based

on the information in Sutton (1980), conidiomata in this genus have an upper

wall several cells thick, and large 0-1-septate cylindrical to clavate conidia

each with a truncate base. The lectotype of Leptostroma is L. scirpinum Fr., the

anamorph of Hypohelion scirpinum (DC.) P.R. Johnst. It produces uniloculate

often coalescing conidiomata with an upper wall only one cell thick on dead

stems usually of Schoenoplectus lacustris (L.) Palla (Sutton 1980). The type

of Melasmia is the anamorph of Rhytisma acerinum (Pers.) Fr. It produces

multiple conidiomata inside large black stromata on dead portions of living

leaves and on dead attached or recently fallen leaves of Acer. The conidioma

upper wall is only one cell thick. The type of Tryblidiopycnis is the anamorph

of Tryblidiopsis pinastri (Pers.) P. Karst. It produces large complex often

multilocular conidiomata with upper walls only one cell thick on twigs of Picea.

Its conidia are long, thin and curved.

From this review, it is evident that the present species has a combination of

characters not perfectly matched by any one of these genera: those producing

conidiomata with thick upper walls are on woody material or have large septate

conidia with a dispersal function; those on leaves produce conidiomata with

thin upper walls and have conidia with a spermatial function. In external

appearance, and in shape, size and development of conidiogenous cells and

conidia, the present fungus looks like a species of Leptostroma. This is by far the

largest anamorphic genus of the Rhytismataceae, and it has already been used

for anamorphs of many members of the family inhabiting pine needles (Minter

Leptostroma magnum sp. nov. (China) ... 291

1980; Sutton 1980; Cannon & Minter 1986; Lin et al. 2012). The only significant

difference between the present fungus and L. scirpinum, the type of the genus,

is that its conidiomatal upper walls are several cells thick.

Closer inspection of other pine needle inhabiting Leptostroma anamorphs

shows that there is, in fact, considerable variation in conidiomatal upper walls.

Where conidiomata are subepidermal, the upper wall is often paler, may be

limited to an area round each ostiole (where present), or may even apparently

be absent. Where conidiomata are subcuticular, the upper wall is often much

blacker and much more extensive. The extent and pigmentation of conidiomatal

upper walls may therefore be related, at least partly, to depth of embedding in

the plant tissue, and that may reflect water availability and the need to protect

against UV radiation (Sherwood 1981). In the present species, where the

conidiomata are subcuticular, the upper wall is even more strongly developed.

Until the biological implications of a thicker upper wall are better understood,

upper wall thickness seems insufficient reason to establish a new genus, the

fungus is therefore formally named below as a new species of Leptostroma.

Conservation status

Data Deficient. The species is poorly known and has been recorded from

only one location (in a protected area which is part of the UNESCO Man

& Biosphere network). If an obligate association with P henryi were to be

demonstrated, the conservation status of this fungus would automatically be at

least Near Threatened, the current IUCN red list conservation status of its only

known associated plant.

Acknowledgments

We are grateful to Dr Z. Wang (Yale University, USA) and Dr M. Ye (Hefei University

of Technology, China) for serving as pre-submission reviewers, to Dr S.J. Wang,

Ms J.L. Chen and Q. Zheng for the field investigations. This study was supported by the

National Natural Science Foundation of China (No. 31270065, 31170019).

Literature cited

Burnett JH. 1983. Speciation in fungi. Transactions of the British Mycological Society 81(1): 1-14.

Cannon PF, Minter DW. 1986. The Rhytismataceae of Indian subcontinent. Mycol. Pap. 155: 1-123.

Index of Fungi. 1947. Review of applied mycology, supplement 14. Index of Fungi 1(14): 161-176.

Jones SG. 1935. The structure of Lophodermium pinastri (Schrad.) Chev. Annals of Botany 49:

699-728.

Kirk PM, Cannon PF, Minter DW, Stalpers JA. 2008. Dictionary of the fungi. 10th ed. CAB

International. Wallingford. 771 p.

Lightle PC. 1954. The pathology of Elytroderma deformans on ponderosa pine. Phytopathology

44: 557-569.

Lin YR, Liu HY, Hou CL, Wang SJ, Ye M, Huang CL, Xiang Y, Yu SM. 2012. Flora fungorum

sinicorum, vol. 40, Rhytismatales [in Chinese]. Science Press. Beijing. 261 p.

292 ... Zhang & al.

Minter DW. 1977. Lophodermium on pines with special reference to species occurring on Pinus

sylvestris in North-east Scotland. [Ph.D. thesis; in English]. University of Aberdeen. Aberdeen.

401 p.

Minter DW. 1980. Leptostroma on pine needles. Can. J. Bot. 58: 906-917.

http://dx.doi.org/10.1139/b80-115

Minter DW. 1981. Lophodermium on pines. Mycol. Pap. 147: 1-54.

Minter DW. 1985. Some members of the Rhytismataceae (Ascomycetes) on conifer needles from

central and North America. USDA Forest Service General Technical Report WO-50: 71-106.

Sherwood MA. 1981. Convergent evolution in discomycetes from bark and wood. Journal of the

Linnean Society Botany 82: 15-34.

Sutton BC. 1977. Coelomycetes VI. Nomenclature of generic names proposed for coelomycetes.

Mycol. Pap. 141: 1-253.

Sutton BC. 1980. The Coelomycetes: Fungi imperfecti with pycnidia, acervuli and stromata.

Commonwealth Mycological Institute. Kew. 696 p.

MY COTAXON

http://dx.doi.org/10.5248/129.293

Volume 129(2), pp. 293-296 October-December 2014

A new Stemonitis species from Turkey

HasAN HUSEYIN DOGAN' & GONUL EROGLU!

University of Selcuk, Faculty of Science, Department of Biology, 42075 Konya, Turkey

Bi ¢ 3 P Sy: 4

* CORRESPONDENCE TO: hhuseyindogan@yahoo.com

ABSTRACT — Stemonitis mediterraneensis is described herein as a new species of myxomycete

from material collected on a rotten wood remnant of Abies cilicica subsp. cilicica from

Adana-Aladag in Turkey. The new species is characterized by rough warty pale brownish to

yellowish-brown spores and smooth capillitia lacking spines.

Key worps — Mycetozoa, Protista, slime moulds, Stemonitidaceae

Introduction

Myxomycetes are a small, relatively homogeneous group of eukaryotic

organisms (Stephenson & Stempen 1994). They occur on a variety of substrates

such as living plants as well as decaying logs, dead leaves, and other organic

matter (Alexopoulos et al. 1996). Since Gleditsch (1753) first described

Stemonitis Gled., 17 species have been identified (Lado 2005-2014), including

nine recorded in Turkey (Sesli & Denchev 2014). Recent morphological

examination has revealed a new species, S. mediterraneensis, bringing world’s

total Stemonitis species to 18.

Materials & methods

Morphological observations and measurements were conducted under a Leica Z 6

APO-A stereoscopic microscope (Leica Microsystems CMS GmbH, Mannheim, DE).

The total height of the sporocarps as well as height and width of the sporotheca were

measured. Whenever possible, 15 sporocarps of each collection were checked (110

sporocarps in total). Microscopic characteristics were observed and measured from

material mounted in Hoyer’s medium under a Leica DM-750 light microscope. A

total of 110 spores (including ornamentation) were measured using an oil immersion

objective; spore color and ornamentation also were determined. SEM images were

prepared with a Zeiss DSM-950 scanning electron microscope (Micronova, Espoo,

Finland). Specimens are conserved in the Fungarium, Mushroom Application and

Research Center, University of Selcuk, Konya, Turkey (KONF).

294 ... Dogan & Eroglu

Taxonomy

Stemonitis mediterraneensis H.H. Dogan & Eroélu, sp. nov. FIGURE 1

MycoBank MB 804345

Differs from Stemonitis mussooriensis by its smaller spores with shorter rougher warts.

Type: Turkey. Adana: Aladag, Meydan Yaylasi, Ceritler, 37°29’01”N, 035°19’45”E, 1265

m, on rotten wood of Abies cilicica (Antoine & Kotschy) Carrriére subsp. cilicica, 27

Oct. 2011, leg. HLH. Dogan 9315 (Holotype, KONF HD 9315; Isotype, KONF GE 560).

Erymo oey: referring to the Mediterranean region where the specimens were collected.

SPOROCARPS in groups, cylindrical, total height 2.2-3.5 mm, dark brown.

Sporotheca 1.5-2.8 mm in length and 0.2-0.4 mm in diameter. STALK short,

about 1/2-1/3 of the total height, 0.7-0.8 mm in length, shining, dark brown.

PERIDIUM fugacious. COLUMELLA 7-8 um wide, cylindrical, dark brown,

opaque, similar in color to the stalk, tapering gradually upwards and merging

into the capillitium just below the apex. CAPILLITIUM 1-1.5 um in diameter,

pale brown, with a surface net consisting of variable-sized meshes (4-110

um), smooth, not bearing spines and the internal reticulum 3-4 meshes over

the radius; the surface net delicate with angular meshes and attached to the

columella. Spores (8.5-)9-10.6 um in diameter, globose, covered with rough

warts, pale brownish, yellowish brown under the light microscope. By SEM,

the ornamentation consisting of regularly distributed and isolated spiny warted

0.3 um long projections. HyPOTHALLUs discoid under the separate sporangia,

yellowish brown.

EcoLoGy & DISTRIBUTION: Found on rotten wood of Abies cilicica subsp.

cilicica in a mixed forests of A. cilicica subsp. cilicica and Cedrus libani A. Rich.

Discussion

Stemonitis is characterized by stalked cylindrical sporocarps, an evanescent

peridium, a columella that almost reaches the top of the sporocarp, a pale

brown, red brown or dark brown capillitium that arises from the entire length

of the columella, the absence of capillitial spines, spores that are dark brown,

rust brown or lilac grey in mass and usually pale brown in transmitted light.

Morphologically and anatomically, six species are very close to Stemonitis

mediterraneensis; the seven similar species can be distinguished as follows:

Key to Stemonitis mediterraneensis and related species

IP SPObeS Len cu late ty... le ta wate te te Ute tain rae My, Urea, Wr, re Be ee 2

Le Spores: with isolatedswarts-or, Bluintzspines...\.. vd sence Beets omy Monde mtente eg 3

2 OPOESUSUAll Y= SUNY tema, worse, terete ain ede ede eee eats S. virginiensis

2 Sporeswuisuallyees iia: wes 2) SNe SNe OP Oe aie o aGls eh icine cele sees Se S. fusca

SHSPOLes WStial ly iss6: PATS 4 bee eg dont e g discwreck dated 4 berg Wade BA iaber BW bee were S. axifera

DeeS POLES US Ural N72 Ge (TN ee Fay Sete Pans rtaays tap laid as Oar la Bar alata Mer alates wth abt heath AG: ah +:

Stemonitis mediterraneensis sp. nov. (Turkey) ... 295

FIGURE 1 — Stemonitis mediterraneensis (holotype). A: Sporocarps (SM). B, C: An entire sporocarp

(SM). D: Capillitial meshes and spores (SEM). E: Detail of spore ornamentation (SEM). F: Spores

(SEM). G, H: Capillitial meshes (LM). I: Expanded columella apices (LM). J: Spores (LM).

A pour facesite tsiniall tives ede Lie oT Pea EI Nag LU Sag INL Sy Be Nae FN 5

Ar Subiacetietlarce-mesiede 8 .e:tt Rents Mert neley hale hheole rhe mele hee EN ae ER es Old 6

>. stalk 41/3 ‘onthe: total ere tite, 0.) i 2 9 tes oo emo seg 3 nee ale S. pallida

5. Stalk 1/5-1/4 of the total height, sometimes almost lacking............. S. herbatica

GnSpotes bearing fineipalewarte yet, sete pets epee agen nett. S. mussooriensis

Ox spores bearitigirough spiny wa4ts.. ete lie a Lion cet one S. mediterraneensis

Its rough warty spores distinguish Stemonitis mediterraneensis from S. fusca

Roth and S. virginiensis Rex, both characterized by reticulate spores with rows

296 ... Dogan & Eroglu

of spines, some of which are connected by ridges (Nannenga-Bremekamp 1991,

Ing 1999). Stemonitis mediterraneensis is close to S. axifera (Bull.) T. Macbr.,

S. pallida Wingate, S. herbatica Peck, and S. mussooriensis G.W. Martin et al.

in its spore ornamentation, but its spore morphology easily distinguishes the

new species. The colorless spores of S. axifera are smaller (5-6 um) and almost

smooth to covered with very small, pale warts. The pale rust spores in S. pallida

are also smaller (6.5-7.5(-8) um) and ornamented with pale warts, sometimes

appearing almost smooth (Nannenga-Bremekamp 1991; Stephenson 2003). In

spore size, S. herbatica and S. mussooriensis are the most closely related species

to S. mediterraneensis but can be distinguished by the lilac brown color of the

spores.

The absence of a spinose surface net, shared by Stemonitis mediterraneensis

and S. herbatica, distinguishes those two species from S. fusca, S. virginiensis,

S. pallida, and S. mussooriensis, all of which have a spinose surface net. Its

unique combination of macroscopic, capillitial, and spore features easily

support S. mediterraneensis as an independent Stemonitis species.

Acknowledgments

The authors are indebted to the Scientific Research Projects (BAP) Coordinating

Office (BAP/11401021) at Selcuk University and The Scientific & Technical Research

Council of Turkey (TUBITAK TOVAG 1060496) for their financial support of the

current work. We would like to thank Dr. Gabriel Moreno, Spain for the SEM pictures.

We would like to thank Dr. Steven L. Stephenson and Dr. Hakan Alli for reviewing this

article.

Literature cited

Alexopoulos CJ, Mims CW, Blackwell M. 1996. Introductory mycology, 4th edition. John Wiley

and Sons Inc., New York.

Gleditsch JG. 1753. Methodus fungorum exhibens genera, species et varietates cum charactere,

differentia specifica, synonomis, solo, loco et observationibus. Sumtibus Scholae Realis,

Berolini.

Ing B. 1999. The Myxomycetes of Britain and Ireland. The Richmond Publishing Co., Slough,

England.

Lado, C. (2005-2014). An online nomenclatural information system of Eumycetozoa.

http://www.nomen.eumycetozoa.com (accessed October 2014).

Nannenga-Bremekamp NE. 1991. A guide to temperate Myxomycetes. Biopress Limited, Bristol,

England.

Sesli E, Denchev CM. 2014. Checklists of the myxomycetes, larger ascomycetes, and larger

basidiomycetes in Turkey (accessed October 2014).

http://www.mycotaxon.com/resources/checklist/sesli-v106-checklist.pdf

Stephenson SL. 2003. Myxomycetes of New Zealand. Fungal Diversity Press, Hong Kong.

Stephenson SL, Stempen H. 1994. Myxomycetes: a handbook of slime molds. Timber Press,

Portland, Oregon, USA.

MY COTAXON

http://dx.doi.org/10.5248/129.297

Volume 129(2), pp. 297-302 October-December 2014

A new species of Coccomyces with dimorphic paraphyses

XIAO-YAN WANG’, HAI-LIN Gu’, QING LI’,

LEI-HONG WANG?, & YING-REN LIN?

' School of Life Science &? School of Forestry & Landscape Architecture,

Anhui Agricultural University, West Changjiang Road 130, Hefei, Anhui 230036, China

*CORRESPONDENCE TO: yingrenlin@yahoo.com

ABSTRACT — A new Coccomyces species found on fallen leaves of Millettia dielsiana at Mt

Sanqingshan, Jiangxi province, China, is described as Coccomyces alienus. This taxon is

distinguished from all other Coccomyces taxa by having two types of paraphyses. The type

specimen is deposited in the Reference Collection of Forest Fungi of Anhui Agricultural

University, China (AAUF).

Key worps — foliicolous fungus, taxonomy, Fabaceae, fungal diversity

Introduction

Coccomyces De Not. is the second-largest genus in the Rhytismataceae

(Rhytismatales, Leotiomycetes, Ascomycota), of which at least 116 species have

been recorded (Kirk et al. 2008, Lin et al. 2012). They are widely distributed

and occur on leaves, herbaceous stems, bark, and wood (Sherwood 1980). The

study of Coccomyces in China began in 1934 with the records of C. dentatus on

Quercus and Castanea, and C. delta (Kunze) Sacc. on Lauraceae (Teng 1934).

Subsequently, twenty-eight additional species have been reported (Korf &

Zhuang 1985; Lin et al. 1994; Lin 1998; Hou et al. 2006, 2007; Jia et al. 2012;

Zheng et al. 2012; Wang et al. 2013; Yang et al. 2013). Of the 159 fungal species

known to inhabit Millettia, no Coccomyces species has yet been reported on this

plant genus (Farr & Rossman 2014).

The present study, which is based on a specimen collected from Mount

Sanqingshan National Park (Jiangxi Province, China) describes a new species

of Coccomyces on a Millettia dielsiana vine.

Materials & methods

The external shape, size, colour, opening mechanisms of ascomata and conidiomata,

and zone lines were observed on dried reference materials using a stereoscope with

298 ... Wang & al.

10-50x magnification. After rehydration in water for ca 10 min, the fruitbodies were

cut into 10-20 um thick sections with a cryo-microtome. The sections were examined

microscopically in water, 5% KOH, Melzer’s reagent, or lactophenol-cotton blue.

Sections pretreated in water were mounted in lactic acid or cotton blue to observe

ascomatal and conidiomatal outlines in vertical section. Colours of various structures

and ascospore contents were determined in water. Asci, ascospores and paraphyses were

measured and drawn from tissues mounted in 5% KOH. At least thirty asci, ascospores,

and paraphyses were measured for each specimen. Figures of the external features

and internal structures were drawn using a microscopic painting device. The type

specimen is deposited in the Reference Collection of Forest Fungi of Anhui Agricultural

University, Hefei, China (AAUF).

Taxonomy

Coccomyces alienus Y.R. Lin & Xiao Y. Wang, sp. nov. Fics 1-6

MycoBank MB 807573

Differs from Coccomyces dentatus by triangular to pentagonal ascomata with lip cells

and periphysoids, loose-textured and fawn-coloured internal stromatic matrix (textura

angularis-globulosa), two types of paraphyses with very different shapes, cylindrical and

thinner asci, and narrower ascospores.

Type: China, Jiangxi, Mount Sanqingshan National Park, Bingyudong, alt. 1550 m,

on fallen leaves of Millettia dielsiana Harms (Fabaceae), 21 August 2012, Y.R. Lin,

X.Y. Wang & L. Zhang 2661 (Holotype, AAUF 68769).

EryMo_oey: alienus (Latin = alien), referring to two types of paraphyses with very

different shapes.

ZONE LINES generally frequent, dark brown or black, thin, entirely or partly

surrounding bleached spots of 8-20 mm dam.

CONIDIOMATA in similar positions to ascomata on the host, scattered

to clustered. In surface view, conidiomata 110-260 um diam., rounded or

subrounded, black-brown in the centre and the perimeter line of the conidioma,

hazel to brown elsewhere, flattened, discharging spores through an apical

ostiole. In median vertical section, conidiomata intraepidermal, more or less

lenticular. UPPER WALL extremely poorly developed, yellow-brown but black-

brown near the ostiole, composed of tiny angular cells beneath the cuticle.

Sometimes individual epidermal cells conjoining the cuticle. BASAL WALL

12—25 um wide, of dark brown textura angularis with thick-walled cells 3—7

um diam. SUBCONIDIOGENOUS LAYER 10-15 um wide, comprised of colourless,

thin-walled angular cells 2-3 um diam. CONIDIOGENOUS CELLS and CONIDIA

not present in material available.

AscoMaTA developing on both sides of leaves, mostly hypophyllous,

scattered or somewhat clustered, sometimes two or three coalescent; occurring

in subcircular or irregular pale areas, sandy-brown in colour with obvious

edges. In surface view, ascomata 520-900 um diam., triangular to pentagonal,

Coccomyces alienus sp. nov. (China) ... 299

Ps ‘)

CED ae

EXO

: OG as

CURES

50 um

Figs 1-6. Coccomyces alienus (holotype) on Millettia dielsiana. 1. Habit on a leaf. 2. Detail of

fruitbodies and a zone line. 3. Ascoma in median vertical section. 4. Portion of ascoma in median

vertical section. 5. Paraphyses, asci and ascospores. 6. Conidioma in vertical section.

300 ... Wang & al.

black-brown to black, slightly shiny, moderately raising above the surface

of leaf but flat or slightly hollow in the centre portion, edge defined, with

an obvious preformed dehiscence mechanism, opening by 3-5 radial splits

which extend nearly to the edge of ascoma to expose the cream yellow top

of hymenium. Lips poorly developed. In median vertical section, ascomata

intraepidermal. COVERING STROMA 15-25 um thick, composed of blackish

brown textura angularis-epidermoidea with thick-walled cells 3—5.5 um diam.,

connecting to the basal stroma. There is a small thin mulch that comprises

strongly carbonized tissue with no obvious cellular structure on top of the

covering stroma, sometimes a few epidermal cells connecting with the cuticle.

Lip CELLS hyaline, 5-8 x 2—2.5 um, cylindrical, slightly swelling at the apex,

thin-walled, 0—1-septate. PERIPHysorps lining the inner face of the covering

stroma, 8-15 x 2-3 um, cylindrical, often indistinctly swollen at the top,

hyaline, straight or curved, 1—3-septate. BASAL STROMA consisting of 1—2(—3)

layers of black-brown, angular, thick-walled cells 4—6 um diam. 25—40 um thick

textura angularis with colorless, thin-walled cells 5-8(—10) um diam. existing

between the covering stroma and basal stroma. INTERNAL MATRIX OF STROMA

moderately developed, 20-35 um thick, comprised of loose, gelatinized,

nearly colourless or fawn-coloured textura angularis-globulosa with thin-

walled cells 4-8 um diam. SUBHYMENIUM 10-15 um thick, slightly concave,

consisting of colourless textura porrecta. PARAPHYSES exceeding height of

asci by 10-20 um, of two types: A-paraphyses frequent, 3—4.5 um wide, thick

and strong, cylindrical, straight or occasionally curved, mainly gradually or

abruptly swollen to 4.5—-6 um at the apex, sometimes branching near the top

or at lower positions, thinly septate, smooth-walled, with no gelatinous matrix;

B-paraphyses sparse, 0.8-1.4 um wide, narrow, filiform, slightly curved,

occasionally gradually swollen to 1.5—2.5 um at the apex, unbranched, septate,

covered with a ca. 0.5 um thick mucous coating. Ascr ripening sequentially,

somewhat sparse, 85-120 x 4.5—6 um, cylindrical, apex rounded, thin-walled,

without a circumapical thickening, short-stalked, J-, 8-spored. ASCOSPORES

arranged in a fascicle, 45-75 x 1-1.2 um, filiform, slightly tapered towards

the base, straight or slightly curved, hyaline, aseptate, containing oil drops,

surrounded by a 0.5—1 um thick gelatinous sheath, sometimes with subglobose

gelatinous caps at the apex or both ends.

HOST SPECIES, HABITAT, & DISTRIBUTION: Producing conidiomata and

ascomata on fallen leaves of Millettia dielsiana. Known only from the type

locality, Jiangxi Province, China.

COMMENTS—Coccomyces alienus is very similar to C. dentatus (J.C. Schmidt)

Sacc. in the way ascomata and conidiomata are embedded, the opening form of

fruitbodies, and the lack of excipulum. However, C. dentatus has quadrangular

to hexagonal ascomata with nonexistent lip cells, periphysoids, and a black

Coccomyces alienus sp. nov. (China) ... 301

mulch that occurs on top of the stromal covering, a virtually absent internal

stromal matrix, monomorphic, shorter (80-105 um long) paraphyses, shorter

and thicker (70-105 x 8-10 um) cylindric-clavate asci, and thicker (2 um wide)

ascospores (Sherwood 1980).

Coccomyces limitatus (Berk. & M.A. Curtis) Sacc. which produces

macroscopically similar zone lines, ascomata, and conidiomata and similarly

shaped periphysoids, asci, and ascospores, differs from C. alienus in its absence

of lips, a well developed excipulum, a covering stroma not extending as far as

the basal stroma, a subhymenium of globose cells, and a much thicker (up to

50 um thick) internal stromal matrix composed of loose hyphae and crystals

(Johnston 1986). Sherwood’ earlier (1980) observation that the excipulum and

periphysoids of C. limitatus are absent was contradicted by Johnston (1986),

who suggested that the excipulum of C. limitatus form after the ascomata open

and may not be obvious when immature. Young ascomata have the appearance

of a well-developed layer of short, hyaline, cylindric periphyses that lengthen

and develop septa to form the excipulum.

Coccomyces cyclobalanopsidis Y.R. Lin & Z.Z. Li, which resembles C. alienus

in the zone lines, the way the ascomata open, and the shapes and sizes of the

asci and ascospores, is distinguished from the new species by its triangular to

hexagonal ascomata, covering stroma and basal stroma consisting of textura

globulosa, the longer and thicker (12-20 x 3—4 um) periphysoids, absent lip

cells, excipulum arising from the inner layer of the covering stroma, and the

extremely poorly developed internal stromal matrix of textura intricata (Lin

et al. 2000).

We observed that C. alienus, which produced mature or overripe conidiomata

and ascomata on recently dead leaves, was often confused with Terriera minor

(Tehon) P.R. Johnst., whose immature or barely mature ascomata could be

easily distinguished on a same leaf, suggesting that C. alienus fruits earlier than

T. minor. This new fungus may bea weak pathogen causing leaf spot. It is possible

that it first develops latently within the living leaves but exhibits no obvious

symptoms, after which the asexual and sexual stages appear sequentially on

the brown lesions of the infected leaves as the bioenergy of the host declines.

Thereafter diseased leaves become reddish brown or sandy-brown and fall

during July and August. Sherwood’s (1980) observation that most Coccomyces

species fruit on recently dead plant materials is largely consistent with our

observations.

Acknowledgements

The authors are grateful to Dr J.E. Taylor for giving valuable suggestions and

editing English for the manuscript, and to Dr C.L. Hou and Dr M. Ye for serving as

pre-submission reviewers. This study was supported by the National Natural Science

Foundation of China (No. 31270065, 31170019).

302 ... Wang & al.

Literature cited

Farr DF, Rossman AY. 2014. Fungal databases, Systematic Mycology and Microbiology Laboratory.

ARS, USDA. Retrieved 25 January 2014.

Hou CL, Piepenbring M. 2007. Two new species of Rhytismataceae on twigs of conifers from

Yunnan Province, China. Mycotaxon 102: 165-170.

Hou CL, Kirschner R, Chen CJ. 2006. A new species and new records of Rhytismatales from

Taiwan. Mycotaxon 95: 71-79.

Jia GJ, Lin YR, Hou CL. 2012 [“2011”]. A new species of Coccomyces (Rhytismatales, Ascomycota)

from Mt. Huangshan, China. Mycotaxon 118: 231-235. http://dx.doi.org/10.5248/118.231

Johnston PR. 1986. Rhytismataceae in New Zealand 1. Some foliicolous species of Coccomyces de

Notaris and Propolis (Fries) Corda. New Zealand J. Bot. 24: 89-124.

Kirk PM, Cannon PF, Minter DW, Stalpers JA. 2008. Dictionary of the fungi, 10th ed. CAB

International. Wallingford. 771 p.

Korf RP, Zhuang WY. 1985. Some new species and new records of Discomycetes in China.

Mycotaxon 22: 483-514.

Lin YR. 1998. Studies on Coccomyces de Notaris and Neococcomyces gen. nov. in China. [Ph.D.

thesis; in Chinese]. Northeast Forestry University. Harbin. 98 p.

Lin YR, Liu HY, Tang YP, Hu BE 1994. Two new species of the genus Coccomyces from China

[in Chinese]. Acta Mycol. Sin. 13: 8-12.

Lin YR, Li ZZ, Huang CL, Xiang CT. 2000. Studies on the genus Coccomyces from China II

[in Chinese]. Mycosystema 19: 297-301.

Lin YR, Liu HY, Hou CL, Wang SJ, Ye M, Huang CL, Xiang Y, Yu SM. 2012. Flora fungorum

sinicorum, vol. 40, Rhytismatales [in Chinese]. Science Press. Beijing. 261 p.

Sherwood MA. 1980. Taxonomic studies in the Phacidiales: The genus Coccomyces (Rhytismataceae).

Occ. Pap. Farlow Herb. Crypt. Bot. 15: 1-120.

Teng SC. 1934. Notes on Discomycetes from China. Sinensia 5: 431—465.

Wang SJ, Lin YR, Tang YP, Li K. 2013 [“2012”]. A new species of Coccomyces (Rhytismatales,

Ascomycota) on Ilex elmerrilliana. Mycotaxon 122: 287-291. http://dx.doi.org/10.5248/122.287

Yang MS, Lin YR, Zhang L, Wang XY. 2013 [“2012”]. Coccomyces hubeiensis, a new fungus of

Rhytismatales from China. Mycotaxon 122: 249-253. http://dx.doi.org/10.5248/122.249

Zheng Q, Lin YR, Yu SM, Chen L. 2012 [“2011”]. Species of Rhytismataceae on Lithocarpus spp.

from Mt Huangshan, China. Mycotaxon 118: 311-323. http://dx.doi.org/10.5248/118.311

MY COTAXON

http://dx.doi.org/10.5248/129.303

Volume 129(2), pp. 303-315 October-December 2014

Cytospora from Salix in northern China

XIN-LEI FAN’, CHENG-MING TIAN’, QIN YANG’, YING-MEI LIANG’,

CHONG-JUAN YoU’, & YU-BO ZHANG"

'The Key Laboratory for Silviculture and Conservation of Ministry of Education,

Beijing Forestry University & *Museum of Beijing Forestry University

Beijing 100083, China

* CORRESPONDENCE TO: chengmt@bjfu.edu.cn

ABSTRACT — Fungal specimens from Salix causing willow Cytospora canker in northern

China were evaluated through morphological observation and phylogenetic analysis. They

were identified as four species — C. rostrata sp. nov., C. nivea, C. chrysosperma, and Valsa

populina. Cytospora rostrata is a new beaked species parasitizing Salix cupularis in Gansu

Province, China. It is morphologically distinguished from other Cytospora species by its

fruiting bodies with a single locule and a thorn-like beak. Molecular evidence obtained from

the internal transcribed spacer (ITS) region also suggests that C. rostrata represents a distinct

lineage within Cytospora. ‘This is the first Chinese record of C. nivea on Salix.

Key worps — Diaporthales, morphology, phylogeny, taxonomy

Introduction

Willow (Salix) is a crucial commercial and ecologically important forest

tree that is grown as a major biomass energy source for its yield potential and

coppicing ability in Netherland (Londo 2002). In China, willows are one of

the most important trees for their economic and ecological value, and are

also widely planted in courtyards, residential districts, and parks as a highly

adversity-resistant perennial tree. However, most willow trees are under serious

threat in northern China from dieback and stem canker caused by Cytospora

species. A systematic study of pathogenic fungi on Salix is badly needed.

The genus Cytospora was established by Ehrenberg in 1818 and has been

shown to be the anamorph of the ascomycetous genus Valsa Fr. (Diaporthales)

(Adams et al. 2005), causing serious dieback and stem canker diseases

commonly found on woody plants (Schreiner 1931, Spielman 1984, Wang et

al. 2014). About 110 species of Cytospora have been described on more than

85 host species (Adams et al. 2005, Kirk et al. 2008). Traditional taxonomy

304 ... Fan & al.

of Cytospora has been based heavily on host species, but Adams et al. (2006)

and Mehrabi et al. (2011) have recently stressed the taxonomic significance

of morphology (including presence or absence of a conceptacle, number and

arrangement of locules, size and shape of conidiophores, and conidial size) in

this group.

Twenty-six species of Cytospora and Valsa have been recorded as pathogenic

fungi of willow canker worldwide, of which nine have been reported in China:

C. capreae Fuckel, C. salicis (Corda) Rabenh., C. fugax (Bull.) Fr., C. fertilis

Sacc., C. translucens Sacc., C. germanica Sacc., C. chrysosperma, C. ambiens

(Pers.) Sacc., and V. populina (Deng 1963, Tai 1979, Wei 1979, Zhuang 2005).

However, morphological data for some of these records were inadequate for

identification, and taxonomic studies of Cytospora species from Salix in China

are lacking. We therefore collected 116 isolates from ca. 200 specimens on willow

trees from nine provinces of northern China and used 14 representative isolates

in a systematic study based on morphology and DNA sequence variation. Our

specimens included a new taxon, which we describe as Cytospora rostrata.

Materials & methods

Sample collection and isolation

A total of 116 Cytospora isolates were obtained from lesions on infected twigs of

willow trees in nine Chinese provinces: Xinjiang, Tibet, Shaanxi, Gansu, Ningxia,

Qinghai, Heilongjiang, Beijing, and Liaoning. Plant tissues were cut into ca. 5 mm

diameter discs, placed in 70% ethanol for 1 min, and then rinsed with sterile water three

times. The twig segments were then placed onto the surface of potato dextrose agar

(PDA) and incubated at 28°C. The growing edges of hyphae were transferred onto a new

PDA plate after 1-2 days. Each isolate was purified through single spore isolation and

maintained according to Fan et al. (2014).

Morphological observation

Cytospora specimens used for morphological observation have been conserved in

the Museum of Beijing Forestry University (BJFC). Morphological characteristics of the

fruiting body, including the size of stromata, color, size and shape of disc, presence or

absence of a conceptacle, number and diameter of ostioles per disc, arrangement of

locules, and conidiophore size and shape, were recorded. Over 20 conidiomata were

sectioned and diameters of 50 conidia were randomly chosen to measure the length and

width by a Leica light microscope (DM 750). Cultural characteristics including colony

color and pycnidial structure were recorded after 3, 7, and 30 days.

DNA extraction and PCR amplification

Genomic DNA of all cultures was extracted using a modified CTAB method (Doyle

& Doyle 1990). Extracted DNA was electrophoresed on 1% agarose gel to check genomic

DNA quality. The ITS1-5.8S-ITS2 (ITS) region of the rDNA was amplified by PCR using

the primer pair ITS1 and ITS4 (White et al. 1990) in 20 uL volumes with 1 uL (20 ng)

DNA template, 1.6 uwL dNTP (2.5 mmol/L), 2 uL 10xPCR buffer (15 mM/mL MgCl),

Cytospora rostrata sp. nov. (China) ... 305

TABLE 1. New ITS sequences of Cytospora and Valsa spp. included in phylogenetic

analyses.

SPECIES Host LOCATION ISOLATE SEQUENCE

C. rostrata Salix cupularis China, Gansu BJFC CGLs251 HT KC313890

S. cupularis China, Gansu BJFC CGLs252 KC313891

S. cupularis China, Gansu BJFC CGLs280 KC313892

C. chrysosperma S. babylonica China, Ningxia §BJFC CNLs2 KC787357

S. babylonica China, Shaanxi = BJFC CSLs3 KC787358

S. carmanica China, Gansu BJFC CGLs4 KC787359

S. cupularis China, Gansu BJFC CGLs8 KC787360

S. matsudana China, Gansu BJFC CGLs2 KC787361

S. matsudana f. tortuosa China, Gansu BJFC CGLs7 KC787362

S. babylonica China, Ningxia BJFC VNLs2 KC787365

C. nivea S. cupularis China, Gansu BJFC CGLs32 KC787364

S. cupularis China, Gansu BJFC VGLs33 KC787363

V. populina S. matsudana f. tortuosa China, Gansu BJFC VGLs7 KC787366

S. cupularis China, Gansu BJFC VGLs24 KC787367

HT = holotype.

1 uL of each primer (10 uLmol/L), 0.1 uL Taq DNA polymerase (5 U/uL), and 13.3 uL

ddH,O. The PCR protocol comprised denaturation at 94°C for 10 min, followed by 35

cycles of 94°C for 30 s, 51°C for 30 s, 72°C for 40 s, and a final cycle at 72°C for 10 min.

PCR products were electrophoresed on 1% agarose gel and the DNA was sequenced by

the Shanghai Invitrogen Biological Technology Corporation.

Phylogenetic analyses

Our 14 sequences representing the four species isolated from Salix were used in the

phylogenetic analyses (TABLE 1) with 49 GenBank ITS sequences representative of a

cross-section of Cytospora/Leucostoma/Valsa spp., and Rostraureum tropicale Gryzenh.

& Wingf. as outgroup. The sequences were assembled using SeqMan (7.1.0) and

aligned using Clustal X (1.81). Alignments were manually adjusted to allow maximum

alignment and maximum sequence similarity, and gaps were treated as missing data.

The phylogenetic tree was constructed with the neighbor-joining (NJ) method using

PAUP (4.10), and the maximum likelihood (ML) method using Phylip (3.6.9) (Swofford

2000, Tamura et al. 2011). The stability of relationships was assessed by bootstrap

analyses comprising 1000 data settings. Constructed trees were viewed by Treeview

(1.6.6) software.

Results

Phylogenetic analyses

The alignment of ITS regions consisted of 510 base pairs. The ML analysis

produced a similar topology to the NJ analysis presented in PLATE 1.

306 ... Fan & al.

-/52

87/-

100/100

aro

0.02

PL. 1 Phylogenetic tree based on aligned sequences of ITS1-5.8S-ITS2 region of 63 isolates

of Cytospora, Leucostoma, and Valsa generated from neighbor joining (NJ) and maximum

likelihood (ML) analyses. NJ and ML bootstrap values greater than 50% are indicated

(separated bya slash from 1000 heuristic replicates). Tree is rooted with outgroup Rostraureum

Cytospora ribis EF447386

C. gutnerae EF447365

C. terebinthi EF447402

100}g1 | C. rhodophila DQ243809

C. carbonacea DQ243805

00/109 V. populina KC787366

V. populina KC787367

V. ceratosperma EF447409

C. sacculus EF447389

C. mougeotii AY347329

V, friesii AY347319

1001001} tiesii AY347317

99/98| V. salicina EF447364

V. salicina EF447363

C. schulzeri EF447399

C. schulzeri JQ086558

V. malicola EF447415

69- C. schulzeri JQ086562

V. malicola EF447414

V. malicola DQ243802

C. schulzeri EF447401

V. germanica DQ243793

C. kantschavelii EF447367

C. hariotii DQ243807

C. rostrata KC313891

C. rostrata KC313892

C. rostrata KC313890 (Holotype)

C. chrysosperma EF447310

C. chrysosperma KC787357

C. chrysosperma KC787358

C. chrysosperma KC787359

C. chrysosperma KC787360

C. chrysosperma KC787361

C. chrysosperma KC787362

C. chrysosperma KC787365

V. sordida EF447419

C. chrysosperma EF447317

C. chrysosperma EF447309

C. eriobotryae AY347327

V. ambiens JQ618018

V. ambiens EU310398

V. ceratosperma DQ241769

C. nivea KC787363

Leucostoma niveum DQ243795

C. nivea KC787364

Leucostoma niveum AF 191174

og/64r L. persoonii EF447375

L. persoonii EF447373

C. pruinosa EF447380

V. cypri EF447411

V. pini AY347316

L. cinctum EF447406

L. cinctum EF447359

98/100| | L. cinctum EF447408

90/76" L, cinctum EF447361

C. cedri AF192311

V. ceratosperma AY347335

C. mali AB470827

C. austromontana AY 347362

V. fabianae AY347360

V. fabianae AF260265

100/98

82/100

100/99

56/53' Y fabianae AF260266

Valsa subclypeata AY347331

V. populina

C. rostrata

C. chrysosperma

C. nivea

Rostraureum tropicale AY 167436

tropicale. The new sequences are represented in bold font.

Cytospora rostrata sp. nov. (China) ... 307

The sequences grouped into 29 clades. Cytospora rostrata had a high

similarity of several nucleotides variation with C. chrysosperma. However, the

phylogenetic tree and the morphological characteristics indicated C. rostrata

represents an independent taxon.

Taxonomy

PL. 2 Cytospora rostrata (Holotype, BJFC-CGLs251). a Conidiomata on twig. b, c Transverse

sections through conidiomata. d Detail of beak on twig. e, f Longitudinal sections through

conidiomata. g Conidiophores and conidiogenous cells. h Conidia. i Colonies on PDA at 3 d (left)

and 30 d (right). Scale bars: a = 1 mm; b-f = 0.5 mm; g = 20 um; h = 5 um.

Cytospora rostrata C.M. Tian & X.L. Fan, sp. nov. PLATE 2

MycoBank MB 810728

Differs from Cytospora chrysosperma by its conidial stromata with a single simple locule

and thorn-like ostiolar beak.

308 ... Fan & al.

Type: CHINA, GANSU PROVINCE, Gannan, Diebu, 34°04’48.85”N 103°23’34.20’E, elev.

2989 m asl, on stems of Salix cupularis Rehder, 9 August 2012, Y.M. Liang & X.L. Fan

(Holotype, BJFC-S726; culture BJFC-CGLs251; GenBank KC313890).

EryMo_oey: rostrata, referring to the beaked fruiting body.

Conidiomatal stromata immersed in bark, erumpent, beaked, discoid to

conical, with a single locule. Disc absent. Beak slim, spiny, 0.87-1.20 mm (x = 0.98,

n = 20), with one ostiole per beak. Ostiole in the center of the beak, 22-24

um (x = 23 um, n = 20) in diameter. Locules discoid, simple-loculed, 455-851

um (x = 668 um, n = 20) in diameter. Conidiophores hyaline, unbranched or

occasionally branched at the bases, 15.3-25.1 um (x = 18.9 um, n = 20). Conidia

hyaline, eguttulate, elongate-allantoid, aseptate, 3.6-4.8 x 1.0-1.6 um (x = 4.1

x 1.38 um, n = 50). Cultural characteristics were white originally, became pale

yellow after 6-7 d. The colony was flat, thin, and with a uniform texture. The

arrangement of conidiomata on PDA was circular in the center and radial to

colony edge.

Teleomorph not found.

ADDITIONAL SPECIMENS EXAMINED: CHINA, GANSU PROVINCE, Diebu, 34°04’48.35”N

103°23’36.60’E, elev. 2973 m asl, on stems of Salix cupularis, 9 August 2012, Y.M.

Liang & X.L. Fan (BJFC-S727; culture BJFC-CGLs252; GenBank KC313891); Diebu,

34°04’44.65”"N 103°23’39.42”E, elev. 2977 m asl, on stems of Salix cupularis, 9 August

2012, Y.M. Liang & X.L. Fan (BJFU-S733; culture BJFC-CGLs280, GenBank KC313892).

Cytospora chrysosperma (Pers.) Fr. Syst. Mycol. 2: 542, 1823. PLATE 3

Conidiomata stromata immersed in bark, erumpent, discoid, flask-

shaped to conical, with a large multiple locules. Disc grey to black, nearly flat,

circular to ovoid, 0.21-0.33 mm (x = 0.22 mm, n = 20) in diameter, with one

ostiole per disc. Ostiole medium grey, prominent, 62-85 um (x = 76 um, n

= 20) in diameter. Locules complex multi-loculed, subdivided frequently by

invaginations, sharing common walls, 0.64-1.26 mm (x = 1.01 mm, n = 20)

in diameter. Conidiophores hyaline, unbranched or occasionally branched at

the bases, 17.3-28.2 um (x = 22.6 um, n = 20). Conidia hyaline, eguttulate,

elongate-allantoid, aseptate, 4.0-5.6 x 0.8-1.3 um (x = 4.7x 1.2 um, n = 50).

Cultural characteristics were white originally, becoming partially pale yellow

after 6-7 d although primarily white. The colony was flat, felt-like, and the

texture was uniform. Conidiomata formed on PDA randomly.

PL. 3 Cytospora chrysosperma (BJFC-CNLS2). a Conidiomata on twig. b, c Longitudinal sections

through conidiomata. d Detail of beak on twig. e, f Transverse sections through conidiomata.

g Conidiophores and conidiogenous cells. h Conidia. i Colonies on PDA at 3 d. j, 1 Habit of

ascomata on twig. k Longitudinal sections through ascomata. m Transverse sections through

ascomata. n Asci and immature ascospores. o Ascospores. Scale bars: a = 1 mm; b-f, j-m = 0.5 mm;

g=20um;h=5 um;n,o= 10 um.

Cytospora rostrata sp. nov. (China) ... 309

310... Fan & al.

A huge ascostromatal region immersed in the bark, erumpent, extending

basally up to 1.62-2.46 mm (x = 2.13 mm, n = 20) in diameter. Ascostromata

usually scattered, erumpent, circular to ovoid, 0.86-1.19 mm (x = 0.97 mm,

n=20) diameter. Disc was usually obscured by tightly ostiolar necks, when

apparent pale brown to black, nearly hemispherical, circular to ovoid,

0.20-0.34 mm (x = 0.30 mm, n = 20) in diameter, with 4-8 ostioles arranged

circinately in a disc, brown to black. 4-8 perithecia arranged circinately in

black entostromata, flask-shaped to spherical, 0.26-0.34 mm (x = 0.30 mm,

n=20) in diameter. Asci free, clavate to elongate obovoid, 38.6-43.8 x 5.1-6.2 um

(x = 40.3 x 5.4 um, n = 20), 8-spored. Ascospores, elongate-allantoid, thin-

walled, hyaline, aseptate, 8.3-13.1 x 2.0-2.8 um (x = 11.1 x 2.4 um, n = 50).

Cultural characteristics were white. The colony was flat, felt-like, and the

texture was uniform. Conidiomata formed on PDA uniformly.

MATERIALS EXAMINED: CHINA, GANsu PROVINCE, Gannan, Lintan, Yeliguan,

34°57’36.48”N 103°39’55.12’E, elev. 2187 masl, on stems of Salix carmanica Bornmiieller,

5 August 2012, X.L. Fan (BJFC-S702; culture BJFC-CGLs4; GenBank KC787359);

Gannan, Zhuoni, Dayu National Forestry Park, 34°22’19.82”N 103°35’55.45”E, elev.

2181 m asl, on stems of Salix cupularis, 7 August 2012, X.L. Fan (BJFC-S706; culture

BJFC-CGLs8; GenBank KC787360); Gannan, Lintan, Yeliguan, 34°57’43.41”N

103°39’44.82’E, elev. 2189 m asl, on stems of Salix matsudana Koidzumi, 5 August 2012,

X.L. Fan (BJFC-S700; culture BJFC-CGLs2; GenBank KC787361); Gannan, Lintan,

Yeliguan, 34°57'36.48"N 103°39’59.70’E, elev. 2187 m asl, on stems of Salix matsudana

f. tortuosa (Vilm.) Rehder, 5 August 2012, X.L. Fan (BJFC-S705; culture BJFC-CGLs7;

GenBank KC787362). NINGXIA PROVINCE, Yinchuan, 38°31’51.80”N 106°17’48.05”E,

elev. 1231 m asl, on stems of Salix babylonica L., 13 February 2012, X.L. Fan (BJFC-S508;

culture BJFC-CNLs2; GenBank KC787357; culture BJFC-VNLs2; GenBank KC787365).

SHAANXI PROVINCE, Weinan, Hua, Xiamiao, 34°34’47.66”N 109°45’41.61”E, elev. 335

m asl, on stems of Salix babylonica L., 26 May 2012, X.L. Fan (BJFC-S571; culture BJFC-

CSLs3; GenBank KC787358).

Cytospora nivea (Hoftm.) Sacc., Michelia 2: 264, 1881, nom. illegit., non Fuckel 1860.

PLATE 4

Conidiomata immersed in bark, erumpent, discoid, flask-shaped to conical,

conceptacles dark, with a large multiple locules. Disc grey, nearly flat, became

white when old, raised, circular to ovoid, 0.20-0.34 mm (x = 0.29 mm, n = 20)

in diameter, with one ostiole per disc. Ostioles medium grey, prominent, 60-92

um (x = 77 um, n = 20) in diameter. Locules complex multi-loculed, subdivided

frequently by invaginations, sharing common walls, 0.69-1.04 mm (x = 0.88

Pi. 4 Cytospora nivea (BJFC-CGLs32). a Conidiomata on twig. b, c Longitudinal sections

through conidiomata. d Detail of beak on twig. e, f Transverse sections through conidiomata

g Conidiophores and conidiogenous cells. h Conidia. i Colonies on PDA at 3 d. j, 1 Habit of

ascomata on twig. k Longitudinal sections through ascomata. m Transverse sections through

ascomata. n Asci and immature ascospores. 0 Ascospores. Scale bars: a, j = 1 mm; b-f, k-m = 0.5

mm; g = 20 um;h = 5 um; n, o = 10 um.

Cytospora rostrata sp. nov. (China) ... 311

312 ... Fan & al.

mm, n = 20) in diameter. Conidiophores hyaline, unbranched or occasionally

branched at the bases, 19.3-36.6 um (x = 27.2 um, n = 20). Conidia hyaline,

eguttulate, elongate-allantoid, aseptate, 6.3-8.0 x 1.1-1.4 um (x =7.1 x 1.2 um,

n = 50). Cultural characteristics were white originally, became dark brown and

black after 14 d. The colony was flat, felt-like, and the texture was uniform.

Conidiomata formed on PDA uniformly.

Ascostromata immersed in the bark, erumpent, circular to ovoid, 0.72-

1.15 mm (x = 1.03 mm, n = 20) diameter, with a well defines black zone of

demarcation, which was known as the conceptacle. Disc was usually obscured

by tightly ostiolar necks, when apparent pale brown to beige, nearly flat,

circular to ovoid, 0.19-0.27 mm (x = 0.24 mm, n = 20) in diameter, with 3-8

ostioles arranged circinately in a disc, brown to black. 3-8 perithecia arranged

circinately in beige entostromata, flask-shaped to spherical, 0.23-0.31 mm

(x=0.28 mm, n= 20) in diameter. Asci free, clavate to elongate obovoid, 33.6-42.1

x 6.0-7.1 um (x = 40.4 x 6.9, n = 20), 8-spored. Ascospores, elongate-allantoid,

thin-walled, hyaline, aseptate, 10.7-14.8 x 2.4-2.9 um (x = 12.9 x 2.6 um,

n = 50). Cultural characteristics were white originally, became dark brown and

black after 14 d. The colony was flat, felt-like, and the texture was uniform.

MATERIALS EXAMINED: CHINA, GANSU PROVINCE, Gannan, Zhuoni, 34°35’16.64”N

103°09’39.53”E, elev. 2758 m asl, on stems of Salix cupularis, 8 August 2012, Y.M. Liang

& X.L. Fan (BJFC-S730; culture BJFC-CGLs32; GenBank KC787364); Gannan, Zhuoni,

34°29'23.60”N 103°06’40.18”E, elev. 2540 m asl, on stems of Salix cupularis, 8 August

2012, Y.M. Liang & X.L. Fan (BJFC-S736; culture BJFC-VGLs33; GenBank KC787363).

Valsa populina (Pers.) Fuckel, Jahrb. Nassauischen Vereins Naturk. 25-26: 314, 1871.

PLATE 5

Ascostromata immersed in the bark, erumpent, circular to ovoid, 0.90-1.27

mm (x = 1.07 mm, n = 20) diameter. Disc was usually obscured by tightly

ostiolar necks, when apparent pale brown to beige, nearly flat, circular to ovoid,

0.19-0.24 mm (x = 0.22 mm, n = 20) in diameter, with 6-8 ostioles arranged

circinately in a disc, brown to black. 6-8 perithecia arranged circinately in

beige entostromata, flask-shaped to spherical, 0.25-0.26 mm in diameter. Asci

free, clavate to elongate obovoid, 55.0-68.3 x 8.2-12.7 um (x = 62.5 x 10.5 um,

n= 20), 4-spored. Ascospores, elongate-allantoid, thin-walled, hyaline, aseptate,

18.7-24.1 x 4.3-5.8 um (x = 20.4 x 5.1 um, n = 50). Cultural characteristics were

white. The colony was flat, felt-like, and the texture was uniform. Conidiomata

formed on PDA uniformly.

MATERIAL EXAMINED: CHINA, GANSU PROVINCE, Gannan, Lintan, Yeliguan,

34°57'36.48”N 103°39’59.70’E, elev. 2181 m asl, on stems of Salix matsudana f. tortuosa,

5 August 2012, Y.M. Liang & X.L. Fan (BJFC-S716; culture BJFC-VGLs7; GenBank

KC787366); Gannan, Pucigou, 34°04’51.38”N 103°23’34.33’E, elev. 3041 m asl, on stems

of Salix cupularis, 9 August 2012, Y.M. Liang & X.L. Fan (BJFC-S725; culture BJFC-

VGLs24; GenBank KC787367).

Cytospora rostrata sp. nov. (China) ... 313

Sw és

Pi. 5 Valsa populina (BJFC-VGLs7). a Ascomata on twig. b, c Longitudinal sections through

ascomata. d Disc of ascomata on twig. e, f Transverse sections through ascomata g Asci and

immature ascospores. h Ascospores. i Colonies on PDA at 3 d. Scale bars: a = 1 mm; b-f = 0.5 mm;

g-h = 20 um.

Discussion

Previously nine species of Cytospora and Valsa had been recorded on Salix

in China. However, the identification of some of these species is questionable:

the records of C. capreae, C. salicis, C. fertilis, C. translucens, C. fugax, and

C. germanica share similar morphological characteristics but lack detailed

descriptions and molecular analyses, and we agree that they should be

unified under C. fugax (the name with priority) as suggested by Tai (1979).

Consequently, based on earlier records only four species of Cytospora and

Valsa from Salix can be accepted in northern China: C. chrysosperma, C. fugax,

C. ambiens, and V. populina. In this study, we confirm the earlier reports of

314 ... Fan & al.

C. chrysosperma and V. populina from Salix in China and add C. nivea as a new

host record and C. rostrata as a new species.

Cytospora rostrata has a single locule without invaginations. Cytospora

pruinosa (Fr.) Sacc., C. eucalypticola Van der Westh, C. variostromatica G.C.

Adams & M.J. Wingf. also have single locules but differ from C. rostrata by

having an ostiolar disc, rather than a thorn-like beak. All multi-locular species

of Cytospora are further differentiated from C. rostrata by their lack of a thorn-

like beak.

Phylogenetic analyses also support the proposal of C. rostrata as a

new species. Cytospora rostrata has its closest genetic relationship with

C. chrysosperma, which is the most widespread species of Cytospora.

Acknowledgments

This study is financed by the National Natural Science Foundation of China (Project

No.: 31170603) and Fundamental Research Funds for the Central Universities (Project

No.: BLYJ201404). We are grateful to Dr. K. Tanaka (Hirosaki University) and Dr. L.L.

Huang (Northwest A & F University) for the manuscript amendments. Thanks are also

due to L. Zhao (Bailong River Forestry Administration Bureau, Gansu) for collection of

specimens in this study.

Literature cited

Adams GC, Wingfield MJ, Common R, Roux J. 2005. Phylogenetic relationships and morphology

of Cytospora species and related teleomorphs (Ascomycota, Diaporthales, Valsaceae) from

Eucalyptus. Studies in Mycology 52: 1-144.

Adams GC, Roux J, Wingfield MJ. 2006. Cytospora species (Ascomycota, Diaporthales, Valsaceae):

introduced and native pathogens of trees in South Africa. Australasian Plant Pathology 35:

521-548. http://dx.doi.org/10.1071/AP06058

Deng SQ. 1963. Fungi of China. Beijing, China.

Doyle JJ, Doyle JL. 1990. Isolation of plant DNA from fresh tissue. Focus 12: 13-15.

Fan XL, Liang YM, Ma R, Tian CM. 2014. Morphological and phylogenetic studies of Cytospora

(Valsaceae, Diaporthales) isolates from Chinese scholar tree, with description of a new species.

Mycoscience 55: 252-259. http://dx.doi.org/ 10.1016/j.myc.2013.10.001

Kirk PM, Cannon PF, Minter DW, Stalpers JA. 2008. Dictionary of the Fungi. 10th ed. Wallingford,

UK.

Londo HM. 2002. Energy farming in multiple land use: An opportunity for energy crop

introduction in the Netherlands. Utrecht, Netherlands.

Mehrabi ME, Mohammadi GE, Fotouhifar KB. 2011. Studies on Cytospora canker disease of apple

trees in Semirom region of Iran. Journal of Agricultural Technology 7: 967-982.

Schreiner EJ. 1931. Two species of Valsa causing disease in Populus. American Journal of Botany

18: 1-26. http://dx.doi.org/10.2307/2435722

Spielman LJ. 1984. A monograph of Valsa on hardwoods in North America. Canadian Journal of

Botany 63:1355-1378. http://dx.doi.org/10.1139/b85-190

Swofford DL, 2000. PAUP*: phylogenetic analysis using parsimony, *and other methods. Version

4.0b10. Sinauer Associates, Sunderland.

Tai FL. 1979. Sylloge Fungorum Sinicorum. Beijing, China.

Cytospora rostrata sp. nov. (China) ... 315

Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S. 2011. MEGAS: molecular

evolutionary genetics analysis using maximum likelihood, evolutionary distance, and

maximum parsimony methods. Molecular Biology and Evolution 28: 2731-2739.

Wang X, Zang R, Yin Z, Kang Z, Huang L. 2014. Delimiting cryptic pathogen species causing apple

Valsa canker with multilocus data. Ecology and evolution 4: 1369-1380.

http://dx.doi.org/10.1002/ece3.1030

Wei JC. 1979. Identification of fungus handbook. Shanghai, China.

White TJ, Bruns T, Lee S, Taylor JW. 1990. Amplification and direct sequencing of fungal ribosomal

RNA genes for phylogenetics. 315-322, in MA Innis et al. (eds). PCR protocols: a guide to

methods and applications. Academic Press, New York.

Zhuang WY. 2005. Fungi of northwestern China. Ithaca, New York.

MY COTAXON

http://dx.doi.org/10.5248/129.317

Volume 129(2), pp. 317-327 October-December 2014

New record of Melanoleuca cinereifolia in

Himalayan moist temperate forests of Pakistan

M. SABA'* & A.N. KHALID’

'Department of Botany, University of the Punjab, Quaid-e-Azam Campus, Lahore, 54590, Pakistan

P Vs y: 4] Pp

* CORRESPONDENCE TO: rustflora@gmail.com

ABSTRACT — Basidiomata of Melanoleuca cinereifolia have been collected underneath Pinus

wallichiana trees in Himalayan moist temperate forests of Pakistan. Its identification was

confirmed by nrDNA ITS sequence analysis. A detailed morphological description and

illustrations are provided. This is the first report of M. cinereifolia from Asia.

Key worps —Khyber Pakhtoonkhaw, pine forest, saprobic fungi, Tricholomataceae

Introduction

The Himalayan moist temperate forests of Pakistan, located at elevations of

1373-3050 m, are characterized by vigorous thick vegetation of conifers and

some deciduous trees. These mixed forests have an average rainfall of 59.3 cm

with humidity up to 57% (Champion et al. 1968). Although the Himalayas are

considered one of twenty-five biodiversity hotspots, most mushroom species

are yet to be reported (Myers et al. 2000; Razaq et al. 2013). During research on

saprobic fungi from pine-dominated forests of Western Himalayas, Melanoleuca

cinereifolia was identified as a new record for Pakistan. Melanoleuca Pat. is a

cosmopolitan genus characterized by collybioid to tricholomatoid basidiomata,

convex to slightly depressed pilei (often with a shallow umbo), emarginate to

adnate to shortly decurrent lamellae, absence of a veil, white to pale-yellowish

spore print, cutis to trichoderm pileipellis, hyaline spores with amyloid

ornamentations, and absence of clamp connections (Singer 1986, Boekhout

1988, Vizzini et al. 2012).

Singer (1935, 1943, 1986), Kuhner (1978), Bon (1978), and Boekhout (1988)

gave much importance to morphology (e.g., pileus, stipe and lamellar colour,

and stipe ornamentation) and anatomy (cystidial shape and the basidiospore

Q value) for infrageneric classification in Melanoleuca. Vizziniet al. (2012) used

ITS sequence analyses to check the traditional tripartite Melanoleuca subgeneric

classification (M. subg. Acystis without cystidia; M. subg. Urticocystis with

318 ... Saba & Khalid

urticiform cystidia; M. subg. Melanoleuca with macrocystidia). He concluded

that Melanoleuca is a monophyletic genus in which only two emended

subgenera, Urticocystis and Melanoleuca, are supported; species previously

placed in M. subg. Acystis, shown to be polyphyletic and no longer accepted,

are now supported in M. subg. Urticocystis.

Two other Melanoleuca species — M. exscissa (Fr.) Singer and M. angelesiana

A.H. Sm. — have previously been reported from Pakistan (Ahmad 1980, Razaq 2013).

Materials & methods

Collection and morphological examination

The basidiome (MSM#005) was collected, photographed, vouchered, dried under

fan heater, and characterized morphologically. Specimen sections were mounted in 5%

KOH for examination under a MX4300H biological microscope (Meiji Techno Co.,

Ltd., Japan); phloxine was used to increase contrast, and Melzer’s reagent was used to

test for amyloidity of the basidiospores.

Measurements of anatomical features (basidiospores, basidia, cystidia, pileus hyphae

and stipe hyphae) were calculated from at least 20 measurements made with an ocular

micrometer and 100x oil-immersion objective; abbreviations include x = mean spore

length and width for all spores measured, Q = spore length / width ratio. Line drawings

were made with a Lucida camera. Color designations are from Munsell (1975).

DNA extraction, PCR amplification, DNA sequencing

Genomic DNA was extracted from a small piece of pileus by a modified CTAB

method (Bruns 1995). The internal transcribed spacers (ITS1, ITS2) + 5.8S region of

the nuclear ribosomal RNA gene were targeted by the ITS1F/ITS4 primer pair (White

et al. 1990; Gardes & Bruns 1993) using an Extract-N-Amp plant DNA extraction Kit

(Sigma-Aldrich, St. Louis, MO, USA). The PCR amplification parameters were: initial

denaturation (94 °C for 1 min), 35 cycles (94 °C for 1 min, 53 °C for 1 min, and 72 °C

for 1 min), and final extension 72 °C (8 min). Amplified PCR products were sent for

purification and bidirectional sequencing to Macrogen (Korea).

Sequence alignment and phylogenetic analysis

The nrITS sequence from MSM#005 comprised 740 base pairs. Initial blast analysis

of nucleotide sequences revealed that the specimen showed 99% maximum identity

with Melanoleuca aff. cinereifolia, M. cinereifolia, M. communis (GenBank JX429210,

JX429108, JX429197, JX429207). GenBank sequences were selected based on the

phylogenetic studies on Melanoleuca by Vizzini et al. (2012) and Sanchez-Garcia et al.

(2013).

Sequences were manually edited and assembled using BioEdit (www.mbio.ncsu.edu/

bioedit/bioedit.html). Following Dentinger et al. (2011) for complete ITS sequences,

all sequences were trimmed with the conserved motifs 5’-(...GAT)cATTA-— and

—GACCT(CAAA...)-3" and the alignment portion between them were included in the

analysis. Gaps were treated as data for construction of phylogeny.

The sequence from MSM#005 was aligned with GenBank sequences of M. cinereifolia

and other related taxa (TABLE 1) by Muscle using the default setting in Molecular

Melanoleuca cinereifolia new to Pakistan ... 319

Evolutionary Genetics Analysis (MEGA) software (Tamura et al. 2011). A phylogenetic

tree was constructed with the Maximum Likelihood (ML) algorithm and Neighbor-

Joining method, Jukes & Cantor (1969) model of nrITS sequences, and nearest-

neighbor-interchange (NNI) as ML heuristic search method using MEGAS software

(Tamura et al. 2011). Phylogeny was tested by a bootstrap value of 1000 replicates, and

bootstrap values >50% are cited in the tree.

Sequence of M. cinereifolia generated for this study was submitted to the Genbank

and the accession numbers for this, as well as for other closely related taxa used in the

phylogenetic analysis are cited in phylogenetic tree. Percent Identities (PID) and DNA

divergence were calculated by DNAStar.

TABLE 1. Limnoperdon and Melanoleuca ITS-rDNA sequences used in the phylogenetic

analysis. New sequence indicated in bold.

TAXON COLLECTION/ VOUCHER COUNTRY GENBANK #

L. incarnatum IF030398 Japan DQ097363

M. angelesiana ANC M0203 Italy JN616420

ANC M0203 Italy JN616421

M. brevipes 4574 Italy JF908352

M. aff. cinereifolia FCME11225 Mexico JX429108

IBUGJS89 Mexico JX429210

M. cinereifolia 319 Italy JN052137

1471 Italy JN052138

MSM#005 Pakistan KJ 182965

M. cognata 13939 Italy JF908360

ANC M0170 Italy JN616425

M. communis XAL Murrietal025 Mexico JX429204

FCME 17116 Mexico JX429207

ENCB Guzman 6329 Mexico JX429226

M. decembris ANC M0197 Italy JN616426

ANC M0200 Italy JN616428

M. exscissa ANC M0210 Italy JN616433

ANC M0213 Italy JN616436

M. grammopodia ANC M0217 Italy JN616439

ANC M0218 Italy JN616440

M. heterocystidiosa ANC M0174 Italy JN616444

ANC M0175 Italy JN616445

M. nivea MCVE 9578 Italy JN392452

ANC M0177 Italy JN616450

M. paedida ANC M0189 Italy JN616452

ANC M0190 Italy JN616453

M. pseudoluscina ANC M0194 Italy JN616458

ANC M0195 Italy JN616459

M. strictipes ANC M0172 Italy JN616465

ANC M0173 Italy JN616466

M. subpulverulenta ANC M0004 Italy JN616472

ANC M0178 Italy JN616473

M. substrictipes ANC M0214 Italy JN616474

320 ... Saba & Khalid

Fic. 1. Melanoleuca cinereifolia (MSM#005). Basidiomes. Scale bars: 10 mm.

Taxonomy

Melanoleuca cinereifolia (Bon) Bon, Docum. Mycol. 9(33): 71 (1978). Figs 1-3

DESCRIPTION BASED ON A SINGLE BASIDIOME:

PILEUs 21 mm diam., plano-convex, flat, thin (collybioid); margin straight or

flaring, smooth; surface dull, off-white to grayish near margin (5R9/2), central

disc chocolate brown (5YR1/2). LAMELLAE free or approximate, close, off-white

to gray (SYR9/2). STIPE 26 mm, central, subequal, often with subbulbous base,

hollow, fibrillose, light brown (5YR7/4). RH1zOMORPHS white, few. ODoR and

TASTE not recorded. BASIDIOSPORES 4.3-6.1 x 7.2-9.5 um [x = 5.4 x 8.5 um,

Q = 1.57], oblong or ellipsoid, apiculus absent, warty, thin-walled, hyaline in

KOH, amyloid. Basip1a 8.5-11.8(-13) x 28.7-38.2(-44.8) um, clavate, mono-

, bi-, or tetra-spored, thin-walled, hyaline in KOH; sterigmata 1.8-4.2(-5.1)

um. Trama hyphal, 6.1-8.9 um, thin-walled, hyaline in KOH. CHEILOCYSTIDIA

11.8-14(-16.9) x 47-71(-77.4) um, lageniform with crystals at the apex,

sometimes septate, hyaline, slightly thick walled, up to 1 um. PLEUROCYSTIDIA

similar to cheilocystidia. PILEIPELLIS a cutis, hyphae cylindrical, 9-16 um,

thin-walled, hyaline in KOH. Stipe Hypwae cylindrical, 5-11.8(-14.8) um,

hyaline to pale yellow in KOH. CLAMP CONNECTIONS absent.

Melanoleuca cinereifolia new to Pakistan ... 321

MATERIAL EXAMINED: PAKISTAN, KHYBER PAKHTOONKHAW, Shangla, Yakh Tangay,

under Pinus wallichiana A.B. Jacks., 2 September 2013, coll. M. Saba & A.N. Khalid,

MSM#005 (LAH; GenBank KJ182965).

B aa

Fic. 2. Melanoleuca cinereifolia (MSM#005). A, pileipellis; B, stipitipellis.

Scale bars: A = 20 um, B = 16 um.

322 ... Saba & Khalid

Fic. 3. Melanoleuca cinereifolia (MSM#005). A, basidia; B, cheilocystidia;

C, basidiospores. Scale bars: A, B = 10 um; C = 3.6 um.

Melanoleuca cinereifolia new to Pakistan ... 323

Results

The phylogenetic trees were constructed with the Maximum Likelihood

algorithm (Fic. 4) and Neighbour-Joining method (Fic. 5), using Jukes &

Cantor (1969) model of nrITS sequences and nearest-neighbor-interchange

(NNI) as ML heuristic search method using MEGAS software (Tamura et

al. 2011). Codon positions included were Ist+2nd+3rd+Noncoding. The

Melanoleuca_communis_JX429207

$8 Melanoleuca_communis_JX429204

Melanoleuca_communis_JX429226

M@ Melanoleuca_cinereifolia_KJ182965

Melanoleuca_cinereifolia_JN052137

Melanoleuca_cinereifolia_JN052138

Melanoleuca_aff_cinereifolia_JX429210

7)! Melanoleuca_aff_cinereifolia_JX429108

73 Melanoleuca_subpulverulenta_JN616472

Clade A

Melanoleuca_subpulverulenta_JN616473

100 Melanoleuca_nivea_JN616450

gg | Melanoleuca_heterocystidiosa_JN616444

Melanoleuca_heterocystidiosa_JN616445

Melanoleuca_nivea_JN392452

Melanoleuca_strictipes_JN616465

95 Melanoleuca_strictipes_JN616466

g3 Melanoleuca_exscissa_JN616436

100} | Melanoleuca_exscissa_JN616433

Melanoleuca_substrictipes_JN616474

Melanoleuca_grammopodia_JN616439

83! Melanoleuca_grammopodia_JN616440

Melanoleuca_brevipes_JF908352

109 ; Melanoleuca_pseudoluscina_JN616459

Melanoleuca_pseudoluscina_JN616458

gg, Melanoleuca_paedida_JN616453

75 Melanoleuca_paedida_JN616452

409 ; Melanoleuca_cognata_JF908360

Melanoleuca_cognata_JN616425

83 Melanoleuca_decembris_JN616428

Melanoleuca_decembris_JN616426

56 Melanoleuca_angelesiana_JN616421

Clade B

100 '! Melanoleuca_angelesiana_JN616420

Limnoperdon_incarnatum_DQ097363 “]Outgroup

0.02

Fic. 4. Phylogenetic relationship of Melanoleuca cinereifolia with other Melanoleuca spp. based

on Maximum Likelihood method inferred from nrITS sequences. The tree with the highest log

likelihood (-3381.2856) is shown. Bootstrap values >50 (1000 replicates) are shown below the

branches, and the percentage of trees in which the associated taxa cluster together is shown

next to the branches. The tree is drawn to scale, with branch lengths measured in the number of

substitutions per site.

324 ... Saba & Khalid

Melanoleuca_communis_JX429226

$3 Melanoleuca_communis_JX429207

92 Melanoleuca_communis_JX429204

Melanoleuca_cinereifolia_JN052137

92 57 @ Melanoleuca_cinereifolia_KJ182965

Melanoleuca_cinereifolia_JN052138

Melanoleuca_aff_cinereifolia_JX429210

90 30 Melanoleuca_aff_cinereifolia_JX429108

738 Melanoleuca_subpulverulenta_JN616472

51 Melanoleuca_subpulverulenta_JN616473

Melanoleuca_nivea_JN616450

94 Melanoleuca_nivea_JN392452

Melanoleuca_heterocystidiosa_JN616445

Melanoleuca_heterocystidiosa_JN616444

Melanoleuca_strictipes_JN616465

99 Melanoleuca_strictipes_JN616466

52 Melanoleuca_grammopodia_JN616439

Clade A

100

36 Melanoleuca_grammopodia_JN616440

96 Melanoleuca_brevipes_JF908352

Melanoleuca_substrictipes_JN616474

100 Melanoleuca_exscissa_JN616436

98 Melanoleuca_exscissa_JN616433

100 Melanoleuca_pseudoluscina_JN616459

Melanoleuca_pseudoluscina_JN616458 Clade B

100 Melanoleuca_cognata_JF908360

Melanoleuca_cognata_JN616425

24 Melanoleuca_decembris_JN616428

Melanoleuca_decembris_JN616426

100 Melanoleuca_angelesiana_JN616421

Melanoleuca_angelesiana_JN616420

Melanoleuca_paedida_JN616453

98 Melanoleuca_paedida_JN616452

Limnoperdon_incarnatum_DQ097363 “]Outgroup

Fic. 5. Phylogenetic relationship of Melanoleuca cinereifolia with other Melanoleuca spp. based

on Neighbor-Joining method. The bootstrap consensus tree inferred from 1000 replicates is taken

to represent the evolutionary history of the taxa analyzed. Branches corresponding to partitions

reproduced in less than 50% bootstrap replicates are collapsed. The percentage of replicate trees in

which the associated taxa clustered together in the bootstrap test (1000 replicates) is shown next to

the branches. Evolutionary distances were computed using the Jukes-Cantor method and are in the

units of the number of base substitutions per site.

final dataset contains 33 nucleotide sequences. Limnoperdon incarnatum

(DQ097363) was used as outgroup based on results from Vizzini et al. (2012).

After removing and editing the ambiguous letters from aligned datasheet, a

total of 711 characters were subjected for phylogenetic analysis, of which 435

characters were conserved, 274 were variable, 186 were parsimony informative,

and 87 were singletons.

Melanoleuca cinereifolia new to Pakistan ... 325

The percentage similarity was calculated by MegAlign (DNA Star Inc.).

Melanoleuca cinereifolia showed 99.4% identity and 0% genetic divergence with

sequences retrieved from GenBank (JN052137, JN052138), 99.4% identity and

0.5% divergence with M. aff. cinereifolia sequences (JX429108, JX429210), and

99.3% identity and 0.5% divergence with M. communis (JX429226, JX429204,

JX429207).

Discussion

Melanoleuca cinereifolia and M. communis M. Sanchez-Garcia & J. Cifuentes

closely resemble each other. Both species show 99% maximum similarity to the

present species in the Blast search and probably represent the same taxon. In

the phylogeny MSM#005 clusters with M. cinereifolia and morphologically, it

differs from M. communis in basidiome and pileus size and pileus and lamellar

colour. Melanoleuca cinereifolia is usually characterized as a basidiome with

a short stipe, grey lamellae, lageniform cystidia that grows in sand dunes

(Vizzini et al. 2012). Lantieri et al. (2009), describe it from Italy as frequent on

sand dunes characterized by Cypero capitati-Agropyretum juncei and as well

as at higher elevations characterized by Medicagini marinae-Ammophiletum

australis. In Pakistan, our M. cinereifolia specimen was collected growing on

forest floor rich in organic matter in a pure Pinus wallichiana forest. Sanchez-

Garcia et al. (2013) collected M. cf. cinereifolia in mountainous mesophilic

forests in Mexico.

Other Melanoleuca species known from Pakistan are M. exscissa and

M. angelesiana (Ahmad 1980; Razaq 2013). Unlike M. cinereifolia, M. exscissa

has a urticiform exscissa cystidial type while M. angelesiana lacks cystidia.

Identification of Melanoleuca species is difficult because most morphological

characters are homoplastic and influenced by environmental factors (Vizzini et

al. 2012), making molecular analysis necessary for species identification.

Our phylogenetic trees (Fics 4, 5) divide Melanoleuca into two major clades,

supporting recognition of two subgenera (Melanoleuca and Urticocystis). Clade

A in which the Pakistani specimen is nested is supported by a robust (100)

bootstrap and includes taxa mainly with lageniform or fusiform macrocystidia.

Clade B, which is supported by only 56 bootstrap, includes all taxa with urticoid

cystidia, some species lacking cystidia, and the macrocystidiate M. cognata

complex. Melanoleuca exscissa and M. angelesiana reported from Pakistan

cluster in clade B. Our phylogeny agrees closely with the results reported by

Vizzini et al. (2012). Moreover, percent identity and genetic divergence support

the identity of MSM#005 as M. cinereifolia, as inferred from the phylogenetic

tree.

326 ... Saba & Khalid

Acknowledgments

We are highly indebted to Higher Education Commission (HEC), Pakistan, for

funding this project under Phase II, Batch I, Indigenous PhD fellowships Program for

5000 scholars. We are cordially grateful to Dr. Alfredo Vizzini (Dipartimento di Scienze

della Vita e Biologia dei Sistemi-Universita degli Studi di Torino, Viale Mattioli, Torino,

Italy), Dr. Omar Perdomo (Dominican Society of Mycology, Dominican Republic), and

Dr. Vladimir Antonin (Moravian Museum, Czech Republic) for critically reviewing

the manuscript and their valuable comments. We are thankful to all lab fellows for

accompanying us on the field trips.

Literature cited

Ahmad S. 1980. A contribution to the Agaricales of Pakistan. Bulletin of Mycology 1: 35-89.

Boekhout T. 1988. Notulae ad floram agaricinam neerlandicam, XVI - new taxa, new combinations

in Melanoleuca Pat. and notes on rare species in the Netherlands. Persoonia 13(4): 397-431.

Bon M. 1978. Tricholomataceae de France et d’Europe occidentale (Leucopaxilloideae). Documents

Mycologiques 9(33): 1-79.

Bruns TD. 1995. Thoughts on the processes that maintain local species diversity of ectomycorrhizal

fungi. Plant and Soil 170: 63-73. http://dx.doi.org/10.1007/BF02183055

Champion HG, Sethi SK, Khattak GM. 1968. Forests types of Pakistan. Pakistan forest institute,

Peshawar.

Dentinger BTM, Didukh MY, Moncalvo JM. 2011. Comparing COI and ITS barcode

markers for mushrooms and allies (Agaricomycotina). PLoS One 6(9): e25081.

http://dx.doi.org/10.1371/journal.pone.0025081

Gardes M, Bruns TD. 1993. ITS primers with enhanced specificity for Basidiomycetes:

application to the identification of mycorrhizae and rusts. Molecular Ecology 2: 113-118.

http://dx.doi.org/10.1111/j.1365-294X.1993.tb00005.x

Jukes TH, Cantor CR. 1969. Evolution of protein molecules. 21-132, in: HN Munro (ed.).

Mammalian Protein Metabolism, vol. 3. Academic Press, New York.

http://dx.doi.org/10.1016/B978-1-4832-3211-9.50009-7

Kithner R. 1978. Agaricales de la zone alpine. Genre Melanoleuca. Bulletin de la Société Linnéenne

de Lyon 47: 12-52.

Lantieri A, Gargano ML, Venturella G. 2009. The sabulicolous fungi from Sicily (southern Italy):

additions and critical review. Mycotaxon 110: 151-154. http://dx.doi.org/10.5248/110.151

Munsell ™. 1975. Soil color charts. Baltimore.

Myers N, Mittermeier RA, Mittermeier GAB, Fonseca D, Kent J. 2000. Biodiversity hotspots for

conservation priorities. Nature 403: 853-858. http://dx.doi.org/10.1038/35002501

Razaq A. 2013. Molecular characterization and identification of gilled fungi from Himalayan moist

temperate forests of Pakistan using internal transcribed spacers (ITS) of rDNA. PhD thesis,

Department of Botany, University of the Punjab, Lahore.

Razaq A, Khalid AN, Ilyas S. 2013. Molecular identification of Lepiota acutesquamosa and

L. cristata (Basidiomycota, Agaricales) based on ITS-rDNA barcoding from Himalayan Moist

Temperate forests of Pakistan. International Journal of Agriculture and Biology 15: 313-318.

Sanchez-Garcia M, Cifuentes-Blanco J, Matheny PB. 2013. Revision taxondmica del género

Melanoleuca en México y descripcion de especies nuevas. Revista Mexicana de Biodiversidad

84: 111-117. http://dx.doi.org/10.7550/rmb.31569

Singer R. 1935. Etude systématique sur les Melanoleuca d’Europe et clé des espéces observées en

Catalogne. Cavanillesia 7: 122-132.

Melanoleuca cinereifolia new to Pakistan ... 327

Singer R. 1943. Das system der Agaricales III. Annales Mycologici 41: 1-189.

Singer R. 1986. The Agaricales in modern taxonomy, 4th edn. Koeltz Scientific Books, Koenigstein.

Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S. 2011. Molecular

evolutionary genetics analysis using maximum likelihood, evolutionary distance, and

maximum parsimony methods. Molecular Biology and Evolution 28(10): 2731-2739.

http://dx.doi.org/10.1093/molbev/msr121

Vizzini A, Para R, Fontenla R, Ghignone S. Ercole E. 2012 [“2011”]. A preliminary ITS phylogeny

of Melanoleuca (Agaricales), with special reference to European taxa. Mycotaxon 118: 361-381.

http://dx.doi.org/10.5248/118.361

White TJ, Bruns T, Lee S, Taylor JW. 1990. Amplification and direct sequencing of fungal ribosomal

RNA genes for phylogenetics. 315-322, in: MA Innis et al. (eds). PCR Protocols: a guide to

methods and applications. Academic, New York.

MY COTAXON

http://dx.doi.org/10.5248/129.329

Volume 129(2), pp. 329-359 October-December 2014

Entoloma species from New South Wales and

northeastern Queensland, Australia

Davip L. LARGENT**, SARAH E. BERGEMANN’, & SANDRA E. ABELL-DAVIS?

‘Biological Sciences, Humboldt State University, 1 Harpst St, Arcata CA 95521, USA

Evolution and Ecology Group, Biology Department, Middle Tennessee State University,

PO Box 60, Murfreesboro TN 37132, USA

School of Marine and Tropical Biology, Australian Tropical Herbarium and Centre for Tropical

Environmental & Sustainability Science, James Cook University,

PO Box 6811, Cairns QLD 4870 AU

* CORRESPONDENCE TO: mrp@humboldtl.com

ABSTRACT — Seven new species in the Prunuloides clade of the Entolomataceae are described

here: Entoloma hymenidermum is diagnosed by blackish blue basidiomata, isodiametric

basidiospores and moderately broad pileocystidia; E. violaceotinctum has a violet-tinged

pileus, violaceous-tinged stipe, and broad inflated pileocystidia; E. discoloratum possesses

a subviscid yellow-tinged white pileus; E. kewarra is distinguished by its yellow pileus and

stipe, both with a white and then eventually greenish yellow context; E. pamelae has a smooth,

bright yellow, dry pileus; E. rugosiviscosum has a yellow-brown, rugose viscid pileus; and

E. guttulatum is distinguished by lamellae with droplets that become reddish brown on

drying.

Key worps — Basidiomycota, phylogeny, taxonomy

Introduction

A morphologically based classification has given rise to two interpretations

of the genus Entoloma (Agaricales, Entolomataceae). In one approach, taxa are

placed into a single genus and the subgenera are defined mostly by the pileus

surface, the pileipellis structure, and the basidiospore size, shape, and angularity

(Noordeloos 1992, 2005; Co-David et al. 2009; Noordeloos & Gates 2012). In

a second approach, Entoloma is accepted in a restricted sense with additional

genera recognized based on biochemical features, pigmentation, location of

clamp connections, and stature types in addition to the pileus surface, the

pileipellis structure, and the basidiospore size, shape, and angularity (Orton

1991, Largent 1994, Baroni & Matheny 2011, Baroni et al. 2011). Recent

comprehensive phylogenetic analyses of the Entolomataceae have highlighted

330 ... Largent, Bergemann, & Abell-Davis

many issues inherent with the morphologically based classifications by showing

that many of the segregate genera or infrageneric groups are not monophyletic

(Co-David et al. 2009; Baroni & Matheny 2011), suggesting the need to

re-evaluate key characters used for delineation of segregate genera.

The majority of species of Entoloma s.s. are supported in one of two clades,

either the Rhodopolioid or Prunuloides clade (Co-David et al. 2009, Baroni

& Matheny 2011). The species described in this paper share similar features

with many members of the Prunuloides clade: a tricholomatoid or naucorioid

stature (Largent 1986), isodiametric basidiospores, rather short and broad

hyphae in the pileus trama, and abundant clamp connections in the pileipellis.

The Prunuloides clade also accommodates segregate genera. Entocybe

TJ. Baroni et al. was erected for species with finely pustulate basidiospores

with bumps and 6-10 facets when viewed in polar view (Baroni et al. 2011).

Calliderma (Romagn.) Largent was erected for species with a tricholomatoid

stature, a pruinose, tomentose, velutinous, or rivulose pileus corresponding to a

hymeniform pileipellis, and septate hyphae with or without clamp connections.

Australian field investigations from 2009-2012 within northeastern

Queensland’s Wet Tropics Bioregion and from 2010-2012 in the temperate

rainforests of central New South Wales have uncovered several novel

entolomataceous species (Largent & Abell-Davis 2011; Largent et al. 2011a,b,

2013a,b; Bergemann et al. 2013). Here, we describe seven new species

representing Entoloma s.s.: E. discoloratum, E. guttulatum, E. hymenidermum,

E. kewarra, E. pamelae, E. rugosiviscosum, and E. violaceotinctum.

Materials & methods

Macromorphological and micromorphological features

Techniques and equipment for collecting and describing basidiomata in the field,

GPS coordinates, microscopy of dried collections including how basidiospores were

measured, and digital microphotographs have been described in Largent et al. (201 1a, b),

while techniques for color descriptions using Kornerup & Wanscher (1978) and factors

determined from mathematical analyses in the descriptions are covered in Largent et

al. (2013a,b). All collections for New South Wales cited in the ‘Additional collections

examined’ were deposited in The Plant Pathology Herbarium, Orange Agricultural

Institute (DAR). Collections made in Queensland were deposited in the Australian

Tropical Herbarium (CNS). All holotype and isotype collections are deposited in the

herbaria designated using acronyms from Thiers (2012).

DNA sequences and phylogenetic analyses

Thirty-four sequences from three partial genes including the mitochondrial small

subunit ribosomal RNA (mtSSU), the nuclear large subunit ribosomal RNA (LSU), and

second largest subunit of RNA polymerase II (RPB2) were generated for this phylogenetic

analysis along with sequences obtained from GenBank (TaBLE 1). The extraction and

Polymerase Chain Reaction (PCR) protocols for the mtSSU, LSU and the RPB2 follow

Entoloma spp. nov. (Australia) ... 331

TABLE 1. Collections used in the phylogenetic analyses. New sequences generated for

this study are shown in bold. Square bracketed annotations indicate species

names applied in GenBank that differ from those in the phylogram (Fie. 1).

COLLECTION GENBANK ACCESSION NUMBERS

SPECIES FROM GENBANK

IDENTIFIER mtSSU LSU RPB2

Entocybe haastii DLL9868 JQ793644 JQ793651 JQ793658

DLL10087 JQ793645 JQ793652 JQ793659

Ec. nitida [E. nitidum| 7526 TJB GU384602 GU384626 GU384655

Ec. nitida [E. alcedicolor] 210 GQ289292 GQ289152 GQ289224

Ec. trachyospora [R. trachyospora] 5856 TJB GU384605 GU384629 GU384658

Ec. turbida [E. turbidum] 27 GQ289341 GQ289201 GQ289269

6949 TJB GQ289341 GQ289201 GQ289269

Ec. vinacea |E. vinaceum| 8870 TJB GU384598 GU384631 GU384651

Clitopilus aff. hobsonii DLL9586 KJ021688 KJ021698 KC816912

Entoloma albomagnum 427 KC710165 KC710137 —

E. bloxamii 219 GQ289294 GQ289154 GQ289226

E. aff. bloxamii 628 KC710189 KC710159 —

Thiers 53901 KC710168 KC710139 —

E. aff. whiteae [E. prunuloides] 4765 TJB — AY700180 DQ385883

Entoloma sp. [E. haastii] BY21 — AF261309 —

Entoloma sp. 292, 9895 TJB GQ289296 GQ289156 GQ289228

E. caccabus 17 GQ289295 GQ289155 GQ289227

E. caesiolamellatum [E. bloxamii] 6117 TJB = AF261289 =

E. callidermum 609 KC710183 KC710153 —

E. calongei 322 GQ289298 GQ289158 —

E. clypeatum 41 KC710164 KC710136 —

E. coeruleogracilis [E. haastii] 216 GQ289308 GQ289168 GQ289239

217 GQ289309 GQ289169 GQ289240

E. coeruleoviride 609 KC710162 KC710134 KC710057

E. cretaceum 213 GQ289302 GQ289162 GQ289233

E. discoloratum DLL10217 JQ793646 JQ793653 JQ793660

E. ferruginans 11CA032 KJ021689 KJ021699 KJ021693

E. fibulatum [Calliderma fibulatum| SP393751 — FJ973677 —

E. flavifolium 6215 TJB GU384597 AF261301 GU384644

621 KC710179 KC710150 —

E. fragilum MCA2415 KJ021690 KJ021700 KJ021694

E. fumosobrunneum [Entoloma sp.] 2005113 KC710185 KC710155 _

2005120 KC710186 KC710156 —

E. gelatinosum 212 GQ289305 GQ289165 GQ289236

E. griseolazulinum ill GQ289306 GQ289166 GQ289237

E. guttulatum DLL9791 — _ JQ793656

E. hymenidermum DLL10025 JQ793648 JQ793642 —

DLL10054 — JQ793649 —

E. illinitum MCA2488 — KJ021701 KJ021695

E. indigoticoumbrinum 83 GQ289318 GQ289178 GQ289249

E. kermandii 222 GQ289313 GQ289173 GQ289244

E. kewarra DLL10055 — JQ793655 JQ793662

E. lividoalbum 233 KC710182 KC710152 —

332 ... Largent, Bergemann, & Abell-Davis

TABLE 1, concluded

COLLECTION GENBANK ACCESSION NUMBERS

SPECIES FROM GENBANK

IDENTIFIER mtSSU LSU RPB2

E. luridum 2005108 KC710175 KC710146 KC710192

634 KC710170 KC710141 —

YL3952 KC710177 KC710148 —

E. manganaense 215 KC710172 KC710143 —

E. myrmecophilum 231 GQ289314 GQ289174 GQ289245

E. nidorosum 9971 TJB GU384596 GU384617 GU384643

E. ochreoprunuloides [Entoloma sp.] 632 KC710176 KC710147 —

E. pallidocarpum GDGM 28828 JQ993074 JQ410331 JQ993080

E. pamelae DLL10352 = KJ021702 KJ021696

E. perbloxamii 71 GQ289318 GQ289178 GQ289249

E. pluteisimilis 200 GQ289320 GQ289180 GQ289251

E. politum 15 GQ289321 GQ289181 GQ289252

E. prismaticum K381 = AB692006 AB692016

E. prunuloides 40 GQ289324 GQ289184 GQ289255

E. pseudoprunuloides [Entoloma sp.] 627 KC710169 KC710140 —

E. rhodopolium 8 GQ289327 GQ289187 GQ289258

E. rugosiviscosum DLL9676 JQ793647 JQ793654 JQ793661

E. sarcitum 235 GQ289328 GQ289188 GQ289259

E. sericatum 28 GQ289329 GQ289189 —

E. sinuatum 50 GQ289333 GQ289193 GQ289264

182 KC710184 KC710154 —

E. sordidulum 1 GQ289334 GQ289194 GQ289265

E. subsaundersii 11CA047 — KJ021703 KJ021697

E. violaceotinctum DLL10088 JQ793643 JQ793650 JQ793657

Lepista nuda 11CA041 KJ021692 KJ021705 KJ136110

Rhodocybe pruinosistipitata MCA1492 GU384608 GU384627 GU384653

R. spongiosa MCA2129 GU384604 GU384628 GU384657

Tricholoma flavovirens 11CA038 KJ021691 KJ021704 KC816997

Largent et al. (2011b). The primer combinations rpb2-i6f and rpb2-i7r (Co-David et al.

2009), rpb2-EntF2 and rpb2-EntR4 (Largent et al. 2013b) and fRPB2-5F and bRPB2-7R

(Liu et al. 1999; Matheny 2005) were also used. Sequences were generated on an Applied

Biosystems 3130xI Genetic Analyzer at Middle Tennessee State University using the

sequencing protocols outlined in Largent et al. 2011b.

Sequences were edited using Sequencher 4.2.2 (Gene Codes Corporation, Ann

Arbor, MI) and an automated alignment performed using MAFFT v. 7 (Katoh &

Standley 2013). Introns in the RPB2 and hypervariable regions and large introns in the

mtSSU were excluded prior to analysis. The alignment lengths were 570 bp (mtSSU),

1506 bp (LSU), and 1165 bp (RPB2). A Maximum Likelihood analysis based on 100

FIGuRE 1. Topology of the maximum-likelihood phylogram (mtSSU+LSU+RPB2) highlighting

the subclades within the Prunuloides clade that included the new species proposed here (P1 and

P2). Each sequence is labeled with the GenBank-listed isolate identifier and species placement for

clusters (A-D) are described in the text. Branches with >70% support from 1000 rapid bootstraps

are shown. Lepista nuda, Clitopilus aff. hobsonii, Rhodocybe pruinosistipitata, R. spongiosa, and

Tricholoma flavovirens, served as outgroups, and the tree was rooted with L. nuda.

Entoloma spp. nov. (Australia) ... 333

400) Entocybe haastii DLL10087

rl i P1

o1| 'Entocybe haastii DLL9868

er Entoloma coeruleogracilis 217

Entoloma coeruleogracilis 216

Entocybe nitida 7526 TJB pe ear

Entocybe nitida 210 0.03 substitutions / site

os Entocybe turbida 27

oo] | Entocybe vinacea 8870 TJB

|_Entocybe trachyspora 5856 TJB

\_ Entocybe turbida 6949 TJB

Entoloma luridum 634

: 100 Entoloma luridum 2005108

99 Entoloma luridum YL3952

Entoloma manganaense 215

Entoloma hymenidermum DLL10025

Entoloma griseolazulinum i11 A

5) i eee ce er 8 at

Entoloma sp. 292 8985 TJB

7 98 |__________ Entoloma fibulatum SP393751

Entoloma callidermum 512

Entoloma coeruleoviride 609

Entoloma sp. BY21

Entoloma kermandii 222

i -_—Entoloma pseudoprunuloides 627

too | 'Entoloma prunuloides 40 i

jo -Entoloma aff. prunuloides 628 H

\_Entoloma aff. prunuloides Thiers 53901;

100

sopiojnunid

Entoloma ochreoprunuloides 632

L_Entoloma bloxamii 219

rf 7) W-Entoloma rugosiviscosum DLL9676

H Entoloma gelatinosum 212 B:

100 to0 + Entoloma fumosobrunneum 2005120

100 Entoloma fumosobrunneum 2005113

Entoloma caesiolamellatum 6117 TJB

# ee Entoloma albomagnum 427

Entoloma cretaceum 213

100

Entoloma discoloratum DLL10217

i _____ Entoioma guttulatum DLL9791 i

a i L__Entoloma illinitum MCA2488 :

-—Entoloma kewarra DLL10055

i _Entoloma pamelae DILI10352 :

Entoloma indigoticoumbrinum 83

L__Entoloma perbloxamii 71

Entoloma fragilum MCA2415

too jEntoloma sinuatum 182

Entoloma sinuatum 50

79|Entoloma subsaundersii 11CA047

-——Entoloma aff. whiteae 4765 TJB

|____Entoloma pallidocarpum GDGM 28828

Entoloma lividoalbum 238

Entoloma sordidulum 1

Entoloma caccabus 17

—Entoloma sericatum 28

L_Entoloma myrmecophilum 231

Entoloma ferruginans 11CA032

a7|[ss3-—ENntoloma politum 15

[L_Entoloma nidorosum 9971 TJB

Entoloma rhodopolium 8

—Entoloma flavifolium 621

Entoloma flavifolium 6215 TJB

Entoloma clypeatum 41

100

Entoloma prismaticum K381

Entoloma sarcitum 235

Entoloma calognei 322

Entoloma pluteisimilis 200

Rhodocybe spongiosa MCA2129

Rhodocybe pruinosistipitata MCA1492

ploljodopoyuy

Clitopilus aff. hobsonii DLL9586

Tricholoma flavovirens 11CA038

Lepista nuda 11CA041

334 ... Largent, Bergemann, & Abell-Davis

ML replicates and specifying a GTRGAMMA model in RAxML-HPC v. 7.2.8 ALPHA

(Stamatakis 2006) was carried out with a partitioned dataset: RPB2 (across each codon

position for three partitions), and separate partitions for the mtSSU and LSU. Support

values were based on bootstrap proportions on 1000 bootstrap replicates (Stamatakis

et al. 2008).

Results

The phylogenetic analysis based on combined mtSSU+LSU+RPB2 genes

place all seven Entoloma species into the Prunuloides clade (Fic. 1). Clade

P1 contains a cluster of species recognized as Entocybe (BS = 98) and a sister

clade including E. Iuridum Hesler and E. manganaense G.M. Gates & Noordel.

(BS = 99), a relationship that remains unsupported here. Clade P2 contains all

the Australian species described in this report: 1) Entoloma hymenidermum

and E. violaceotinctum cluster with several Calliderma species and share a

close affinity with E. griseolazulinum Manim. & Noordel. (BS = 100, Fie. 1, A);

2) Entoloma rugosiviscosum clusters with E. bloxamii (Berk. & Broome) Sacc.,

E. gelatinosum E. Horak, and species within the E. prunuloides species complex

(BS = 98, Fig. 1, B), whereas E. discoloratum cluster with E. cretaceum G.M.

Gates & Noordel. and E. albomagnum G.M. Gates & Noordel. (BS = 100, Fic.

1, C); and 3) E. guttulatum, E. pamelae and E. kewarra cluster together with

E. illinitum Largent & Aime (BS = 100, Fie. 1, D).

Taxonomy

Entoloma hymenidermum Largent, sp. nov. PLATES 1-2

MycoBank MB804379

Differs from Entoloma griseolazulinum by the broader stipe, smaller basidia, and shorter

basidiospores.

Type — Australia, Queensland, Cook Region, Yorkeys Knob coastal vine forest, within

20m of 16°49’00.3”S 145°43’49.3”E, 20 March 2011, DL Largent 10054 (holotype BRI;

isotype CNS).

EryMoLocy — named for the hymeniderm pileipellis.

PiLEus 29-70 mm broad, 6.5-15.0 mm high; opaque, not hygrophanous, not

translucent-striate; at first convex to broadly convex, sometimes campanulate-

convex, upon expansion and maturity becoming plane and finally uplifted and

undulate, sometimes broadly umbonate; entirely minutely velvety to suede-like

but forming irregularly radial cracks near the margin when mature, always

dull; when young blackish blue with the bluish color dominant (20F8) in the

center, slightly lighter (20F6) elsewhere, on drying dark blue (20E-F6), when

old blackish blue with the black color dominating (19F4); margin incurved at

first, then decurved, finally plane then uplifted, entire then eroded; context

1-3 mm thick above the stipe, bluish. ODoR mild, at times somewhat fragrant.

Entoloma spp. nov. (Australia) ... 335

PLaTE 1. Entoloma hymenidermum (DLL10054 holotype). A. Basidiomata. B. Stipe surface. C.

Pileus surface. Bar scale: A = 15 mm; B= 5 mm; C = 8 mm.

TasTE mild to at times somewhat bitter. LAMELLAE 11-29 mm long, 2.7-10.5

mm deep, narrow to moderately broad and eventually nearly broad; uncinate

to sinuate with a decurrent tooth; close then subdistant; white to between white

and orange white (5A1-2) when young becoming greyish orange (5A-B4-5)

or (5A3-4) with basidiospore maturation; margin smooth and concolorous.

STIPE 27-79 mm long, 4-10.5 mm broad at the apex, 4-14.5 mm broad at

base, equal to clavate; longitudinally appressed fibrillose from the apex to

the basal tomentum, markedly striate, and sometimes twisted; fibrils various

336 ... Largent, Bergemann, & Abell-Davis

: o Pia! o] ™ yay A

Ral Mela zie)

ie ey twee a NER fei

e* J va tee

tsoe me ®

Pw '

P “) J ’ . , *

me ee —2F- oa |

A vs ; : =" — ei

PLaTE 2. Entoloma hymenidermum (DLL10054 holotype). A. Basidiospores. B. Basidium,

basidioles. C. Pileipellis radial section at pileus disc; pileal trama cylindric hyphae (top arrows),

broad hyphae (bottom arrows). D. Pileipellis (calliderm and outer pileal trama). E. Pileipellis, a

calliderm (section slightly squashed). Bars: A = 8 um; B = 12 um; C1 = 190 um; C2 = 230 um;

D=5 um; E= 10 um.

shades of blackish blue, more bluish (20F6) when young, fading slightly (20E4)

when drying, and more blackish or dark bluish grey (19E-F3 or 4) when old,

background color white, base of the stipe white; context stuffed at first, quickly

becoming hollow with maturity; basal tomentum scarce, white. BRUISING

REACTIONS none.

Entoloma spp. nov. (Australia) ... 337

BasIpiIospores distinctly 5-6-angular, others are typically collapsed

when mounted in 3% KOH, in profile and dorsiventral views isodiametric to

subisodiametric, rarely heterodiametric, isodiametric in polar view, 6.8-10.0

(-10.4) x (5.9)6.2-8.8(-9.0) um (x = 8.3 + 0.6 x 7.4 + 0.6 um; E = 1.00-1.31;

Q = 1.12 + 0.07 (isodiametric); n = 125/4). Basip1a narrowly clavate and

tapered, relatively long and narrow, 38.9-48.9 x 9.7-14.5 um (x = 42.8 + 3.1

x 11.4 + 1.32 um; E = 3.25-4.00; Q = 3.78 + 0.33; n = 16/2); 4-sterigmate.

SCLEROBASIDIA absent. HYMENIAL CYSTIDA absent. LAMELLAR EDGE fertile.

CHEILOCYSTIDIA and PLEUROCYSTIDIA absent. LAMELLAR TRAMAL HYPHAE

subparallel, in the center to near the subhymenium, mostly relatively long and

broad and inflated with rounded end walls, 41.0-170 x 8.5-42.1 um, rarely long

and narrow, not inflated and then 170 x 4 um. PILEIPELLIS 28.6-89.7 um in

depth, composed of a chain of 3-5 entangled cells, the basal portion developing

from the outer layer of the pileal trama and the terminal cells erect, distinct,

forming a palisade of pileocystidia that are not tightly packed. PILEOCYSTIDIA

versiform (cylindric, narrowly cylindro-clavate, clavate, obclavate, to

ventricose-rostrate with the rostrum up to 2 um long), 22.4-58.6 x 4.4-18.6

(-21.3) um (x = 41.6 + 10.5 x 10.6 + 4.5 um; E = 1.20-6.3 (rarely 8.8-9.8);

Q =4.60 + 2.17;n = 20/2). PILEAL TRAMA composed of 2 layers, a layer adjacent

to the subhymenium, 228-312 um in depth composed of inflated, relatively

short and broad hyphae, 34.4-137.6 x 9.4-24.2 um (n = 11/1), and a narrow

layer, 119-163 um deep, composed of cylindric entangled hyphae, 30-110 x

3-6 um, the outer most portion forming the pileipellis. StrprTIPELLis a cutis.

CAULOCYSTIDIA and HYMENIAL CLUSTERS absent. OLEIFEROUS HYPHAE present

in the subhymenium. BRILLIANT GRANULES and LIPOID GLOBULES absent.

PIGMENTATION cytoplasmic in the pileipellis, in 3% KOH at first bluish brown

and then colorless due to its solubility in the medium. CLamMP CONNECTIONS

present in all tissues, small and inconspicuous, easily observed in the branched

portion of the pileal tramal hyphae just beneath the pileipellis.

ECOLOGY & DISTRIBUTION — At times abundant, scattered, or solitary in

sandy humus with leaf litter, in coastal vine forest near Maytenus fasciculiflora

Jessup, Canarium australianum F. Muell. var. australianum, Polyalthia

nitidissima Benth., and Guioa acutifolia Radlk., often adjacent to Drynaria

sparsisora (Desv.) T. Moore, and at times sheltered by decaying logs and at

other times near the mangrove zone, Clifton Beach to Yorkeys Knob Beach,

northern Queensland, early to late March.

ADDITIONAL COLLECTIONS EXAMINED — AUSTRALIA. QUEENSLAND, Cook Region,

Clifton Beach, 16°48’50.6”S 145°43’46.6”E, 9 March 2010, PN 016, PN 017; Yorkeys

Knob, 16°45’45.7”S 145°40’29.6”E, 29 January 2011, PN 51 3 February 2011, PN 56;

16°48’33”S 145°40’32”E, 8 March 2010, PN 012; 10 March 2010, PN 012A; 16°45’43.8”S

145°40’27.5”E, 14 March 2011, DL Largent 10025.

338 ... Largent, Bergemann, & Abell-Davis

DISTINCTIVE CHARACTERS — Tricholomatoid basidiomata, minutely velvety

bluish black pileus, longitudinally appressed fibrillose dark bluish grey stipe,

2-layered pileal trama, and basidiospores averaging 8 x 7 um.

ComMEnNts — In the phylogeny, E. hymenidermum and E. violaceotinctum

cluster with species that have bluish basidiomata with a finely to distinctly

velutinous or tomentose pileus surface, a hymeniform-type (calliderm or

erect trichoderm) pileipellis, strongly and distinctly angular basidiospores,

and an outer pileus trama of cylindric cells, at least on the disc. Species with

a hymeniform type pileipellis are often recognized in Calliderma (Aime et

al. 2010; Karstedt & Capelari 2010). However, E. kermandii G.M. Gates &

Noordel., which also falls in this clade, possesses a cutis, a feature inconsistent

with the generic circumscription of Calliderma. Furthermore, C. pruinatocutis

(E. Horak) Karstedt & Capelari and C. rimosum Karstedt & Capelari (Karstedt

et al. 2010) are placed in a distant and unrelated clade (Baroni & Matheny

2011). Without support for the monophyly of Calliderma, we classify the

species described here as Entoloma.

Entoloma hymenidermum is morphologically similar to several species that

have a tricholomatoid habit, dark bluish or bluish-black tinged pileus, fibrillose

stipe, 5-6 angled basidiospores, clavate to versiform pileocystidia, clamp

connections on the pileipellis hyphae, and lack of cheilocystidia.

Entoloma violaceotinctum from New South Wales differs from E. hymeni-

dermum in its larger basidiomata with violaceous tones, a stipe that discolors

purplish and then brownish when handled, basidiospores averaging 9 x 8 um,

longer (47-73 um) basidia, and occurrence in deep leaf humus and organically

rich soils in a northern warm temperate rain forest. Entoloma griseolazulinum,

described from Kerala State (India), has a more slender (3-5 mm wide) stipe,

longer (10-13 um) basidiospores, and larger (47-71 x 11-16 um) basidia

(Manimohan et al. 2006). Calliderma indigoferum (Ellis) Largent, collected in

a swamp among mosses in New Jersey, has an indigo blue (18F3) pileus (75-

100 mm broad), white + blue-tinged blue stipe, and large clamp connections

(Largent 1994). Calliderma caeruleosplendens Largent et al. (Pakaraima

Mountains, Guyana) is distinguished by its densely matted tomentose dark

blue pileus, very dark blue stipe, and its broadly clavate, napiform, or broadly

obclavate pileocystidia (Aime et al. 2010). Entoloma rugosopruinatum Corner

& E. Horak, reported from Sabah (Malaysia) and Kerala State (India), differs

from all of these species primarily by the clampless hyphae (Horak 1980;

Manimohan et al. 1995, 2006).

Entoloma callidermum (Romagn.) Noordel. & Co-David from Africa and

Malaysia can be differentiated from E. hymenidermum by the violet and/or

Entoloma spp. nov. (Australia) ... 339

bister tinges in the pileus and the stipe, 5-7-angled basidiospores, smaller

basidia (27-35 x 6-11 um), and the absence of a 2-layered lamellar tramal

hyphae (Romagnesi 1956, as Rhodophyllus callidermus; Morgado et al. 2013).

However, the original description provided by Romagnesi (1956) may include

segregate species (Eyssartier et al. 2012).

Calliderma fibulatum Karstedt & Capelari, Entoloma simillimum Corner

& E. Horak, E. marinum Corner & E. Horak, E. ducale E. Horak, E. divum

Corner & E. Horak, and E. burkilliae Massee resemble E. hymenidermum and

E. violaceotinctum in basidiome color and the pileipellis but differ in their 5-11

um broad fusoid to cylindric pileocystidia and the presence of cheilocystidia

(Horak 1980, Karstedt & Capelari 2010). Entoloma coeruleomagnum G.M.

Gates & Noordel. differs by its palisadoderm pileipellis and E. kermandii by its

cutis (Noordeloos & Gates 2012).

Entoloma violaceotinctum Largent, sp. nov. PLATES 3-4

MycoBank MB804380

Differs from Entoloma hymenidermum by its pileus and stipe with violet or violaceous

tones, stipe that bruises when handled, larger basidiospores, and longer basidia.

Type — Australia, New South Wales, Strickland State Forest, lower track, central Hunter

District, within 20m of 33°22’49”S 151°19’32”E, 14 April 2011, DL Largent 10088

(holotype DAR).

EryMoLocy — Derived from the Latin violaceus (= violet) + tinctus (= tint) for the

violet tinted pileus and stipe.

PrLEus 40-100 mm broad, 7-40 mm high; opaque and even, hygrophanous, not

translucent-striate; broadly umbonate at all times, convex to broadly convex at

first, then plane to uplifted with age; velvety, at times appearing stippled with

minute, pointy hairs; dark violet to blackish blue (18F4-5 or 19F5-6) fading

to dull violet to dark blue (18-19E4-5) with a darker center (dark violet grey

17F2), dull; margin incurved then decurved, entire; context cream-colored

except for the pellicle that is concolorous with the surface, 6-10 mm broad

above the stipe, 1-3 mm broad at the margin. Opor mild and fragrant in

young specimens, strong but undefined in older specimens. Taste indistinct

in younger specimens, unpleasant in older specimens. LAMELLAE 15-45 mm

long, 6-15 mm deep, moderately broad; adnexed and close then sinuate and

subdistant, seceding with age; at first white or off-white to pale orange (5A3

or 15A2-3) then light orange (5A4) with basidiospore maturation; margin

smooth and concolorous. STIPE 40-92 mm long, 6-16 mm broad at apex, 5-13

mm broad at base, at times flattened and then 17 x 7 mm at the apex and 13

x 10 mm at the very base, versiform in shape (equal, subclavate, clavate or

broad at apex and more narrow at base); appressed fibrillose; fibrils medium

340 ... Largent, Bergemann, & Abell-Davis

PiaTE 3. Entoloma violaceotinctum (DLL10088 holotype). A. Basidiomata. B. Stipe surface.

C. Pileus surface. Bar: A = 20 mm; Bar scale: B. = 16 mm; C = 35 mm.

violet grey (18D2-3), base color yellowish white to light orange (4A2 or 5B3);

context white, at first stuffed with maturity becoming hollow and then snaps

easily; basal tomentum absent. BRUISING REACTIONS on the stipe occurs after

handling, becoming purplish (18E4) and then brownish (5E4).

BASIDIOSPORES 5-6-angled, angles typically distinct, often with irregularly

wavy walls in dried specimens, isodiametric to subisodiametric in profile and

dorsiventral views, isodiametric in polar view, 7.8-10.2 x 7.1-9.2 um (x = 8.9

Entoloma spp. nov. (Australia) ... 341

Prate 4. Entoloma violaceotinctum (DLL10088 holotype). A. Basidiospores. B. Basidia, basidioles.

C. Pileipellis branching pattern. D. Pileipellis (calliderm and outer pileal trama). Bars: A = 8 um;

B = 13 um; C, D = 30 um.

342 ... Largent, Bergemann, & Abell-Davis

+ 0.7 x 8.0 + 0.7 um; E = 1.00-1.24; Q = 1.11 + 0.06 (isodiametric); n = 56/2).

BasIpIA clavate, very long and relatively narrow, often tapered to 2-3 um, usually

with rather large droplets or globules in mature forms, 4-sterigmate, 47.1-73.1

x 10.8-14.2 um (x = 57.4 + 7.8 x 12.6 + 0.9 um; E = 3.63-5.77; Q = 4.57 + 0.58;

n = 25/2). SCLEROBASIDIA absent. HYMENIAL CYSTIDIA absent. LAMELLAE in

section composed of two layers, a hymenium 31-45 um thick, subhymenium

13-22 um thick with two lateral strata, each 17-30 um thick, and a central

layer (86-95 um wide). LAMELLAR TRAMA HYPHAE Subparallel, relatively short,

consisting of two types, slender, narrow hyphae, 61.2-113.1 x 3.3-8.0 um

(n = 9/2) forming between the subhymenium and the center of the lamellae

and inflated hyphae, 39.5-105.3 x 9.5-22.2 um (n = 22/2) found in the center.

PILEIPELLIS 36-54 um in depth, a hymeniderm of pileocystidia, composed of

entangled layer of hyphae, typically 3 cells long, with the pileocystidia as 1-3

terminal cells branching from a subterminal cell. PrLeocystip1A the terminal

cells of branched hyphae that originate from the outer pileal trama, clavate to

broadly obclavate, 29.8-82.7 x 8.2-39.3 um (x = 48.9 + 12.3 x 14.9 + 4.5 um;

E = 1.01-6.36; Q = 3.55 + 1.14; n = 38/2). PILEAL TRAMA composed of two

layers, an entangled layer (61-210 um thick) with slender hyphae, 68.6-112.6

x 3.7-9.1 um; (n = 7/1) the outer portion of which produce the pileipellis and

an adjacent layer (475-539 um thick) composed mostly of inflated hyphae,

50-237 x 12.5-26.0 um (n = 15/2). STIPITIPELLIS a cutis. CAULOCYSTIDIA and

HYMENIAL CLUSTERS absent. OLEIFEROUS HYPHAE present in the trama of

the lamellae and the pileus. BRILLIANT HYPHAE and LIPOID GLOBULES absent.

PIGMENTATION cytoplasmic in the pileipellis, in 3% KOH at first bluish brown

and then colorless due to its solubility in the medium. CLamMP CONNECTIONS

rather large and present in all tissues.

ECOLOGY & DISTRIBUTION — Typically gregarious in mid-April, then

scattered in early May, in deep leaf humus and organic rich soil, near Syncarpia

glomulifera (Sm.) Nied., Ceratopetalum apetalum D. Don, and Doryphora

sassafras Endl.), and Schizomeria ovata D. Don hosts, New South Wales, central

Hunter District, Strickland State Forest, mid April through early May.

ADDITIONAL COLLECTIONS EXAMINED — AUSTRALIA. New SouTH WALEs, central

Hunter District, Strickland State Forest, lower car park area, 19 April 2011, DL Largent

10128; 33°22’47”S 151°19’28”E, DL Largent 10129; 33°22’47”S 151°19’27”E, 28 April

2011, DL Largent 10168; 33°22’47”S 151°19’28”E; 8 May 2011, DL Largent 10215;

33°22’48’S 151°19'30’E, Banksia loop path, 19 April 2011; 33°22’19”S, 151°19’11”E, 3

May 2011, DL Largent 10183.

DISTINCTIVE CHARACTERS — Tricholomatoid basidiomata with violaceous

tones, a stipe that discolors purplish then brownish when handled, basidiospores

averaging 9 x 8 um, long (47-73 um) basidia, and its occurrence in deep leaf

humus.

Entoloma spp. nov. (Australia) ... 343

ComMENTS — Refer to the Comments section under E. hymenidermum for a

discussion of the differences among morphologically similar species.

Entoloma discoloratum Largent, sp. nov. PLATE 5

MycoBank MB804381

Differs from Entoloma albomagnum by its orangish pileus disc, appressed fibrillose

stipe discoloring yellowish then brownish, broader lamellae, farinaceous odor without a

soapy component, and sclerobasidia.

Type — Australia, New South Wales, Central Hunter District, Barrington Tops National

Park, within 20 m of 32°14’25.2”S 151°43’29.8”E, 25 April 2010, DL Largent 9936

(holotype DAR).

EtryMo.Locy — Derived from the Latin discoloratus (= discolored) for the discoloration

of the pileus and stipe.

PiLEus 64-80 mm broad, 11-18 mm high; not hygrophanous, not translucent-

striate; broadly convex and broadly umbonate becoming irregularly uplifted

with age; mostly glabrous but obscurely fibrillose in places particularly near the

margin, dull, sticky to the touch; orange white to pale orange to light orange

(5A3-4or 5A2-3) in the center, off-white elsewhere; margin decurved at first

becoming plane and then uplifted, entire and rimose then quickly eroded;

context white, 3-8 mm deep above the stipe, nearly non-existent at the margin.

Opor of mature basidiomata distinctly farinaceous, particularly after being

crushed. TasTE strongly farinaceous. LAMELLAE 20-35 mm long, 7-15 mm

deep; moderately broad to broad at first and when expanded and mature,

mostly sinuate, in a few places broadly adnexed or adnate; at first crowded

or close and then nearly subdistant; off-white (5A2-3); margin smooth and

concolorous. STIPE 67-148 mm long, 10-21 mm broad at apex, 10-14 mm

broad in the middle, 13-15 mm broad at base, tapered at the base or ventricose

in the middle and then tapered; decidedly and deeply appressed fibrillose along

the entire surface; off white, solid and stuffed; basal tomentum absent. BRUISING

REACTIONS pileus stains or bruises greyish orange (5B4); stipe discoloring

yellowish then brownish (5A2 to 5B3 or 5E-F4) when handled.

BasipiosPorss distinctly angular, isodiametric to subisodiametric in profile

view, 6.7-8.3 x 5.8-8.5 um (x = 7.5 + 0.48 x 7.0 + 0.59 um; E = 0.92-1.20;

Q = 1.07 + 0.08 (isodiametric); n = 31/1). Basrp1a cylindro-clavate, slender,

42.1-50.3 x 7.6-10.3 um (x = 46.0 + 3.04 x 8.9 + 0.89 um; E = 4.43-6.09; Q =

5.2 + 0.61; n = 7/1), full of large irregularly shaped globules; 2- or 4-sterigmate.

SCLEROBASIDIA rare, aborted 51-53.9 x 8.4-10.7 um, wall 1.7-2.3 um thick

(n = 2/1), filled with large globules. HyMENIAL cysTIpIA absent. LAMELLAR

TRAMAL HYPHAE Subparallel, relatively long and broad, 63.2-199.7 x 13.0-29.9

uum (n = 7/1). PILEIPELLIS 140 um thick, an entangled ixocutis, particularly

evident near the center of the pileus, hyphae just below the surface, 3.3-5.8

um wide (n = 4/1). PILEOcysTIDIA mostly prostrate, a few semi-erect, slender,

344 ... Largent, Bergemann, & Abell-Davis

PiaTE 5. Entoloma discoloratum (DLL 9936 holotype). A. Basidiomata. B. Basidiospores.

C. Sclerobasidium on left; elsewhere thin-walled basidia and basidioles (1% phloxine stain).

D. Hyphae of lamellar trama, large clamp on septum of top hyphae. E. Pileipellis, an ixocutis.

Bars: A = 20 mm; B = 7 um; C = 10 um; D = 15 um; E = 35um.

Entoloma spp. nov. (Australia) ... 345

cylindric, 40.9-45.0 x 2.0-3.0 um (n = 4/1). PILEAL TRAMAL HYPHAE Similar

to the lamellar trama, 76.2-169.3 x 16.9-35.3 um (n = 7/1). STIPITIPELLIS a

cutis. CAULOCYSTIDIA and HYMENIAL CLUSTERS absent. OLEIFEROUS HYPHAE

abundant in the lamellar trama, end cells may extend into the subhymenium.

BRILLIANT GRANULES and LIPOID BODIES absent. PIGMENTATION non-existent.

CLAMP CONNECTIONS abundant at the base of the basidia, on the hyphae of the

lamellar trama and pileipellis.

ECOLOGY AND DISTRIBUTION - Scattered to gregarious and buried in the

soil and leaf litter, forming a fairy ring around Syncarpia glomulifera and near

Allocasuarina torulosa (Aiton) L.A.S. Johnson, New South Wales, Barrington

Tops National Park, Jerusalem Creek Track, late April to early-mid May.

ADDITIONAL COLLECTIONS EXAMINED — AUSTRALIA. New SouTH WALEs, central

Hunter District, Barrington Tops National Park, Jerusalem Creek Track. 32°14’25.2”S

151°43’29.8”E, 10 May 2010, DL Largent 9970 (topotype DAR); 32°14’23”S 151°43’29’E,

10 May 2011, DL Largent 10217.

DISTINCTIVE CHARACTERS — Large off-white, subviscid pileus with pale yellow

orange colors in the center; strongly fibrillose stipe without fibrillose flocks

that eventually bruises yellowish then brown, sclerobasidia, distinctly angular

basidiospores, and an ixocutis-type pileipellis.

ComMEnts — Entoloma discoloratum, which clusters with E. albomagnum and

E. cretaceum, can be classified in Entoloma sect. Albida Noordel. based on its

white basidiomata (Noordeloos & Gates 2012).

Entoloma albomagnum is distinguished by its white pileus with faint yellow

tinge at the center, silky fibrillose-scurfy white stipe that does not discolor,

soapy odor when first collected, weakly angular basidiospores, lamellar and

pileal trama composed of short barrel-shaped elements (20-90 x 4-15 um), and

the absence of sclerobasidia (Noordeloos & Gates 2012). The similarly white

E. cretaceum from Tasmania differs in its smaller chalky white basidiomata, the

white watery fibrous stipe, and rancid-farinaceous odor (Gates & Noordeloos

2007, Noordeloos & Gates 2012).

Entoloma kewarra Largent, sp. nov. PLATE 6

MycoBank MB804382

Differs from Entoloma praeluteum by its subviscid pileus with a pruinose bloom at

first, fibrillose stipe base, white context becoming greenish yellow, ixocutis, and the

intracellular pigment in the pileipellis.

Type — Australia, Queensland, Cook region, Kewarra Beach coastal vine forest. Within

20 m of 16°46’50.7”S 145°41’24.8”E, 12 March 2012, DL Largent 10275 (holotype BRI,

isotype CNS).

Erymo.ocy — referring to the type locality, Kewarra Beach coastal vine forest.

346 ... Largent, Bergemann, & Abell-Davis

PILEUS 6-52 mm broad, 4-12 mm high; opaque, not hygrophanous, not

translucent-striate; when immature conic-campanulate then convex, finally

broadly convex to plane, at times broadly umbonate; at first entirely covered

with a thin layer of yellowish white fibrils (2-3A-C7-8), with maturity the

bloom showing spots when damaged or dry, eventually the surface becomes

glabrous but sticky to the touch when bloom disappears; surface without bloom

yellow (4A-B7) except light yellow (3 or 4A2-4) on the margin, when old and

overly mature becoming entirely olive yellow (3C8); margin entire, at first

incurved, when mature remaining incurved or becoming decurved; context

yellowish white (3A3) becoming vivid yellow (2-3A7-8) above the stipe and

slightly lighter (3A7) elsewhere, solid, and 2-6 mm thick above the stipe, 1 mm

thick at the margin. Opor farinaceous and somewhat pungent. Taste bitter

and farinaceous. LAMELLAE 2-17 mm long, 1-5 mm deep; in some narrowly

adnate at first then adnexed, in most adnexed at all times, crowded then close,

narrow at first then slightly more broad in some; when immature yellowish

white to light yellow (4A2-3) becoming orange yellow (5A2) then light orange

(4-5A4-5) with basidiospore production; margin smooth and concolorous.

STIPE 25-100 mm long, 4-10 mm broad at the apex, in some flattened and

5 x 8, 10 x 13, or 13 x 15 mm broad at the apex, 5-15 mm broad in the middle,

and 4-13 mm broad at the base, tapered; distinctly appressed-fibrillose; pale

yellow to light yellow (2A2-3, 3A3, 4A4) becoming olive yellow (3C8) with

age; context solid and stuffed then becoming hollow from the apex downward,

white at first becoming greenish yellow on exposure and when cut becoming

yellow (3A6-7) where hollow; basal tomentum absent. BRUISING REACTIONS

stipe surface becoming pale yellow to light yellow (4A3-4) then olive or olive

brown (3E-F2) where handled.

Basiprospores distinctly angular in larger basidiospores, indistinctly

angular in smaller basidiospores, isodiametric to barely subisodiametric in

all views, short hilar appendage, 5.3-6.8 x 5.0-6.4 um (x = 6.4 + 0.3 x 6.0

+ 0.3 um; E = 1.00-1.19; Q = 1.07 + 0.05 (isodiametric); n = 32/1). BAsiDIA

with abundant granules, narrowly clavate and hardly tapered, 25.7-37.5 x

7.2-11.8 um (x = 31.5 + 4.9 x 8.9 + 1.2 um; E = 2.74-4.94; Q = 3.61 + 0.71;

n= 14/2); 4-sterigmate, the sterigma short and up to 2 um long. SCLEROBASIDIA

absent. HYMENIAL CYSTIDIA absent. LAMELLAR TRAMAL HYPHAE Subparallel,

slightly narrow, 63.3-101.2 x 5.4-9.7 um (x = 76.0 + 11.6 x 7.5 + 1.62 um;

E = 7.28-15.86; Q = 10.6 + 2.78; n = 10/1). PILEIPELLIs an entangled ixocutis,

29.0-120.0 um in depth. PiLeocystTip1a narrowly cylindro-clavate, at times

with a slightly inflated and minutely rostrate apex, 13.9-29.5 x 1.6-2.9 um

(n/1 = 6). PILEAL TRAMAL HYPHAE relatively short and narrow, 33.9-106.9 x

3.7-14.9 um (x = 55.8 + 24.0 x 9.4 + 3.2 um; E = 3.16-12.65; Q = 6.67 + 3.37;

n = 12/1). OLEIFEROUS HYPHAE and LIPOID GLOBULES absent. PIGMENTATION

Entoloma spp. nov. (Australia) ... 347

PLATE 6. Entoloma kewarra (A-B DLL 10275 holotype; C-E DLL 10055). A. Basidiomata. B. Pileus

surface with bloom of very finely appressed fibrils on pileal margin. C. Basidiospores. D. Basidium

and basidioles (1% phloxine stain). E. Pileipellis, an ixocutis. Bar scale: A, B = 15; mm; Bars:

C= 6 um; D = 9 um; E = 30 um.

348 ... Largent, Bergemann, & Abell-Davis

intracellular and yellowish in the pileipellis, exudes a yellowish green exudate

in 70% ethanol, water and 3% KOH. CLAMP CONNECTIONS present on all the

tissues, narrow and small.

ECOLOGY AND DISTRIBUTION —Solitary to scattered in the open and in

sand between a poorly defined heath community and coastal vine forest near

a mangrove community in northern Queensland, Kewarra Beach, late January

through late March.

ADDITIONAL COLLECTIONS EXAMINED — AUSTRALIA. QUEENSLAND, Cook Region,

Kewarra Beach, 16°46’49.7”S 145°41’23.1”E, 28 January 2011, PN 50; 4 February 2011,

PN 57; 16°49’00.3”S 145°43’49.3”E, 21 March 2011, DL Largent 10055; 16°46’50.8”S

145°41’24.6”E 18 March 2012, DL Largent 10289, PN 70; DL Largent 10290, PN 72.

DISTINCTIVE CHARACTERS — Basidiomata tricholomatoid and buried in

the sandy substrate up to two-thirds the stipe length, pileus initially with a

yellowish white pruinose bloom, subviscid pileus surface, stipe surface light or

pale yellow becoming darker where handled, stipe context becoming greenish

yellow with exposure, an ixocutis-type pileipellis, and narrowly clavate basidia.

ComMENTs — Entoloma pamelae and E. kewarra are strongly supported as

sister taxa in our analyses. With its bright yellow colors and tricholomatoid

basidiomata, E. pamelae (also from northeastern Queensland) differs in its

shiny dry egg-shell smooth glabrous pileus, diffracted-scaly stipe base, white

context turning yellow, narrower basidia, minutely externally encrusted

pigmentation in the pileipellis, and cutis-type pileipellis. Additional Entoloma

species with similar morphological features are discussed in the Comments

section under E. pamelae.

Entoloma pamelae Largent, sp. nov. PLATE 7

MycoBank MB804383

Differs from Entoloma praeluteum by its smaller pileus and stipe, fibrillose-squamulose

stipe base, longer basidia, mild and indistinct odor and taste, and narrow cheilocystidia.

Type — Australia, Queensland, Cook region, Danbulla National Park, Kauri Creek

Track, within 20 m of 17°07’50.6”S 145°35’57.7”E, 28 February 2010, DL Largent 9753

(holotype BRI, isotype CNS).

ErymMoLocy — in honor of Pamela Largent, the collector of the holotype.

Piteus 11-40 mm broad, 5-15 mm high; opaque, not translucent-striate,

hygrophanous; convex when young and not umbonate, upon expansion

and maturity becoming broadly umbonate to faintly gibbous and either

remaining convex or becoming broadly convex; shiny, moist, at all times

decidedly glabrous and smooth like an egg-shell; at first yellow (3A6 or 7)

when hygrophanous darker yellow (3B6 or 6-7, “mustard yellow” or “Nankeen

yellow”); margin entire becoming broadly lobed, decurved; context (beneath

a thick yellow rind) 5-7 mm deep above the stipe, <1 mm at the very margin,

Entoloma spp. nov. (Australia) ... 349

——

PiaTE 7. Entoloma pamelae (DLL9753 holotype) A. Basidiomata. B. Basidiospores. C. Basidia

(1% phloxine stain). D. Cheilocystidia (1% phloxine stain). E. Pileipellis, a cutis. F Pileocystidium

with faint external incrustations (arrow). Bars: A = 20 mm; B, C = 5 um; D = 20 um; E = 15 um;

F=3 um.

350 ... Largent, Bergemann, & Abell-Davis

yellow white (3A2) becoming brighter yellow (3A2-3) then pale yellow (3A3)

upon exposure to air. ODOR and TASTE indistinct and mild. LAMELLAE 2-20

mm long, 0.5-4.0 mm deep, when young consistently narrow then moderately

broad to sigmoid; at first adnexed with maturity remaining adnexed but at times

becoming sinuate, crowded then close; yellowish at first becoming pinkish with

basidiospore maturity and then distinctly contrasted when compared with the

pileus and stipe colors; margin concolorous with the face, smooth, at times

faintly denticulate. Stipe 32-60 mm long, 5-15 mm broad at apex, 4-8 mm

broad at base; consistently narrowing to the base; at first faintly appressed-

fibrillose becoming distinctly-appressed fibrillose with the fibrils matted and

becoming + fibrillose-squamulose towards base; pale yellow to pastel yellow

then dull yellow (3 or 4A3-4), the darker colors more evident on handling;

context at first hollow in the apical % to % and stuffed elsewhere, upon maturity

becoming mostly hollow, white when first cut, quickly becoming yellow upon

exposure to air; basal tomentum absent. BRUISING REACTIONS stipe bruises

brownish on handling.

BASIDIOSPORES 5-angled in profile view, 4-5-angled in dorsiventral view,

distinctly angular, small, isodiametric to heterodiametric, 5.0-7.6 x 5.1-7.3 um

(x = 6.4 + 0.53 x 6.0 + 0.49 um E = 0.83-1.35; Q = 1.07 + 0.11 [isodiametric];

n = 30/1). Basrp1a cylindric to cylindro-clavate and very narrow 33.7-38.7

x 5.1-7.7 um (x = 35.6 + 1.8 x 6.6 + 0.8 um; E = 4.60-6.56; Q = 5.4 + 0.55;

n = 9/1); 4-sterigmate. SCLEROBASIDIA absent. HYMENIAL CYSTIDIA absent.

CHEILOCYSTIDIA cylindric to narrowly obclavate, 24-37.7 x 0.8-4.1 um

(n = 6/1), the ends of lamellar tramal hyphae protrude through the hymenium.

LAMELLAR TRAMAL HYPHAE subparallel, very narrow, 55.1-113.1 x 2.7-12.9

um (n = 6/1). PILEIPELLIS a periclinally arranged cutis composed of narrow,

cylindric hyphae, 10-30 um deep and with an unknown material on the outer

surface that stains golden yellow in 3% KOH; prteocystip1a cylindric to

narrowly cylindro-clavate, 24-40 x 2.2-3.6 um. PILEAL TRAMAL HYPHAE not

studied. STIPITIPELLIS a cutis. CAULOCYSTIDIA and HYMENIAL CLUSTERS absent.

OLEIFEROUS HYPHAE, BRILLIANT GRANULES, and LIPOID GLOBULES absent.

PIGMENTATION minutely incrusting the outer walls of the pileipellis hyphae.

CLAMP CONNECTIONS present at the base of the basidia, the cheilocystidia, and

on the hyphae of the lamellar trama and pileipellis.

ECOLOGY AND DISTRIBUTION — Scattered to gregarious in leaf humus

beneath an unidentified species of Casuarina or Acacia in a dry sclerophyll

forest in northern Queensland, Danbulla National Park, Kauri Creek Track

from mid-February to late March.

ADDITIONAL COLLECTIONS EXAMINED—AUSTRALIA. QUEENSLAND, Cook Region,

Danbulla National Park, Kauri Creek Track, 17°07’59.1”S 145°35’55.6”E, 2 March

Entoloma spp. nov. (Australia) ... 351

2010, DL Largent 9763; 17°07’56.3”S 145°35’54.3”E 17 March 2010, DL Largent 9787;

17°08’02.4’S 145°35’51.7”E, 27 March 2012, DL Largent 10352.

DISTINCTIVE CHARACTERS — Basidiomata yellow in contrast to the pinkish

mature lamella, narrow yellowish immature lamellae, pileus glabrous and

smooth like the surface of an egg, pileal context pale yellow beneath a thick

bright yellow rind, small (x = 6.4 x 6.0 um) distinctly angular basidiospores,

cylindric to obclavate cheilocystidia, minutely encrusted pigment on the

external pileipellis hyphal walls.

ComMENTs — Entoloma praeluteum Corner & E. Horak (from Malaysia) differs

from E. pamelae by the larger (40-80 mm broad) pileus, longer (50-80 mm)

stipe, farinaceous odor, shorter (25-30um) basidia, and lack of cheilocystidia

(Horak 1980).

Entoloma flavidum (Massee) Corner & E. Horak (from Singapore, Malaysia,

and Kerala, India) and E. xanthomyces Corner & E. Horak (from Singapore)

have somewhat similar basidiome colors and stature as E. pamelae but differ

in their broader (2140 mm) pileus and white to pallid immature lamellae that

become subdecurrent or decurrent with age. Additionally, in E. flavidum the

basidiospores are larger (9-11 x 6.5-7.5 um) and the basidia are longer (25-65

mm long) than those of E. pamelae (Horak 1980; Manimohan et al. 1995, 2006).

Entoloma rugosiviscosum Largent sp. nov PLATES 8-9

MycoBank MB804384

Differs from Entoloma conspicuum by its smaller basidiomata, the radial rugose viscid

pileus, brown stipe, and ixotrichodermium.

Type — Australia, Queensland, Cook region, Daintree National Park, Tribulation

section, Marrjda Track, within 20 m of 16°08’18.5”S 145°26’26.0’E, 20 March 2009, DL

Largent 9676 (holotype BRI, isotype CNS).

EryMoLocy — from the Latin adjectives rugosus + viscosus referring to rugose viscid

pileus surface.

PitEus 55-75 mm broad, 15 mm high; not hygrophanous, not striate, and not

translucent; broadly convex and umbonate; shiny, viscid and sticky to the touch

and remaining so 14 hours after collecting; scummy on the disc, radially-rugose

elsewhere; medium dark yellow brown (5E-F5) on the disc and brownish orange

to light brown (5C-D4-5) elsewhere, eventually entirely greyish orange with

age and fading; margin greyish orange (5B2-3), decurved and even, elastic, and

extends past lamellae up to 2 mm; context solid, brownish beneath the cuticle,

white elsewhere, 10 mm broad above the stipe apex, 1-1.5 mm at the margin.

opor mild to mildly pungent. Taste mild. LAMELLAE 25-26 mm long, 11-13

mm deep; uncinate to adnexed, at times nearly free; white at first; subdistant,

more or less ventricose and broad; margin smooth then eroded with age,

352 ... Largent, Bergemann, & Abell-Davis

PiaTE 8. Entoloma rugosiviscosum (DLL9676 holotype). A. Basidiome with rugose pileus.

B. Lamellae with eroded margins and stipe apex. Bar = 10 mm.

concolorous. STIPE 50-51 mm long, 11-14 mm broad at the apex, 9.5-14 mm

broad in the middle, 7-15 mm broad at the base, more or less clavate-tapered,

white at first discoloring yellowish (3A2-3) with age and handling, minutely

Entoloma spp. nov. (Australia) ... 353

7 "as -

Fs 2 se : a?” 4 Pul at et 8

PiaTE 9. Entoloma rugosiviscosum (A. DLL9676 holotype; B-D. DLL 9644, 1% phloxine stain).

A. Basidiospores. B. Basidia, basidioles, clamp connection. C. Lamellar trama with gelatinized

subhymenium, section slightly squashed. D. Pileipellis, an ixotrichodermium. Bars: A = 5 um;

B= 8 um; C, D = 40 um.

—~ 4

appressed-fibrillose; flesh white, fibrous-stuffed then hollow in the center with

age. BRUISING REACTIONS basidiomata discoloring yellowish with handling.

BASIDIOSPORES typically cuboidal and distinctly angular in all views, rarely

5-sided, small, 4.8-7.0 x 4.1-6.6 um (x = 5.8 + 0.4 x 5.4 + 0.5 um; Q 0.87-1.25;

E 1.07 + 0.08; n = 52/2). Basip1a 4-sterigmate, long and narrow, 33.1-44.7 x

6.4-9.7 um (x = 38.1 + 3.4 x 7.99 + 1.1 um; Q 3.76-6.23; E 4.85 + 0.8; n=10/1.

SCLEROBASIDIA absent. HYMENIAL CYSTIDIA absent. LAMELLAR TRAMA 318-

400 um wide; subhymenium gelatinized and broad, 40-56 um wide. LAMELLAR

TRAMAL HYPHAE subparallel to nearly parallel, short and broad, 38.5-102.2 x

12.7-17.6 um. PILEIPELLIS an ixotrichodermium with cylindric hyphae 2-3

354 ... Largent, Bergemann, & Abell-Davis

um diam., walls irregular and gelatinized. PILEAL TRAMAL HYPHAE Subparallel,

relatively short and broad, 61.2-133.5 x 15.2-31.0 um. STIPITIPELLIS a cutis but

with solitary but scattered basidia-like cells at the apex. STIPE TRAMAL HYPHAE

nearly parallel, narrow in the outer portions, short and broad in central area and

then 81.7-169.8 x 15.2-36.9 um. OLEIFEROUS HYPHAE abundant in the trama

of the pileus, lamellae, and stipe. BRILLIANT GRANULES and LIPOID GLOBULES

absent. PIGMENTATION cytoplasmic, brownish and uniform in the hyphae of

the pileipellis. CLAamMp CONNECTIONS abundant and present in all tissues.

ECOLOGY & DISTRIBUTION — Solitary, humicolous, adjacent to the

pavement on the Marrdja Track, 15-50 m from parking lot, Daintree National

Park, Tribulation Section.

ADDITIONAL COLLECTIONS EXAMINED — AUSTRALIA. QUEENSLAND, Cook Region,

Daintree National Park, Tribulation section, Marrjda Track, 16°08’18.5”S 145°26’26.0’E,

12 March 2009, DL Largent 9644.

DISTINCTIVE CHARACTERS — Radially rugose viscid yellow brown pileus

that fades to brownish orange or light brown, white stipe discoloring

yellowish, basidiospores mostly cuboidal and small (4.8-7.0 x 4.1-6.6 um), an

ixotrichodermium-type pileipellis, long narrow basidia.

ComMENTsS — Phylogenetically E. rugosiviscosum clusters with several species

including E. prunuloides (Fr.) Quél., E. albomagnum, and E. cretaceum.

Excluding E. ochreoprunuloides Morgado & Noordel.), most of these species

possess a tacky, sticky, subviscid, viscid, or glutinous pileus surface with an

ixocutis- or ixotrichodermium-type pileipellis. Entoloma conspicuum E. Horak

from Papua New Guinea resembles E. rugosiviscosum in its tricholomatoid

basidiomata, brown rugose to subvenous pileus, and cuboidal basidiospores

but differs by the cutis-type pileipellis and larger basidiomata (Horak 1976,

1980). Entoloma rugosostriatum Largent & T. W. Henkel from Guyana also has

a brown rugose pileus and tricholomatoid stature but differs in its isodiametric

nodulose basidiospores (Largent et al. 2008).

Entoloma guttulatum Largent, sp. nov. Piates 10-11

MycoBank MB 807410

Differs from Entoloma pingue by the brown pileus, lamellae with discolored spots, larger

basidia, and unusual pileipellis.

Type — Australia, Queensland, Cook region, Mossman National Park, 20 m from

16°28'17.1”S 145°19’51.2”E, 18 March 2010, DL Largent 9791 (holotype BRI, isotype

CNS).

EryMoLocy — from the Latin adjective guttulatus referring to droplets on the lamellar

edge that dry as reddish brown spots

PitEus 80-82 mm broad, 20 mm high; opaque, not hygrophanous, not

translucent-striate; not umbonate, convex then uplifted; moist, glabrous to

Entoloma spp. nov. (Australia) ... 355

PLATE 10. Entoloma guttulatum (DLL9791 holotype). A. Basidiome. B. Pileus surface. Bar = 13 mm.

356 ... Largent, Bergemann, & Abell-Davis

the eye but under 4x to 20x subfelty or suede-like and with minute colorless

pruinae; between greyish brown and yellowish brown (5D-E3-4) fading to

between light orange and grayish orange (5A-B5-6) when old; margin plane

then uplifted, wavy and crenulate. Opor and TasTeE unpleasant and slightly

farinaceous. LAMELLAE 30-35 mm long, 9-10 mm deep, broad; adnate but a

decurrent or subdecurrent tooth or subdecurrent; greyish orange (6B4) when

mature; margin smooth and concolorous, with droplets as in some Hebeloma

sp. that are abundant and dry as reddish brown spots. St1PE 60-100 mm long,

20 mm broad at the apex, 14 mm broad at base; longitudinally appressed

fibrillose; grayish orange (6B4); context stuffed becoming hollow with age; basal

tomentum absent. BRUISING REACTIONS stipe surface developing brownish

orange to light brown (6C-D4) areas with age and handling.

Basipiospores' distinctly angular, 5-6-angled, isodiametric to

subisodiametric in profile view, isodiametric in polar view, 7.0-9.2 x 6.7-8.8

um (x = 8.1 + 0.6 x 7.8 + 0.5 um; E = 0.9-1.3, Q = 1.04 + 0.08 (isodiametric);

n = 60/2). Bastp1a long and cylindro-clavate, 45.7-60.9 x 7.4-11.2 um

(x = 53.5 + 5.7 x 9.6 + 1.3 um; E = 4.79-6.16, Q = 5.64 + 0.56; n = 6/1); 2- or

4-sterigmate. SCLEROBASIDIA absent. HYMENIAL CYSTIDIA absent. LAMELLAR

TRAMAL HYPHAE 57.6-112.2 x 8.8-13.6 um (n = 7/1). PILEIPELLIS 48-70 um

deep, a colorless anticlinal layer of slender, entangled hyphae, 3-4 um in diam,

beneath which is a pigmented, densely entangled layer of hyphae; terminal

cells cylindric to narrowly obclavate to rostrate-ventricose, 28.2-43.0 x 3.2-5.3

um (n = 6/1). PILEAL TRAMAL HYPHAE composed of broad cells. 34.9-59.9 x

7.4-16.8 um (E = 2.54-6.13; n = 6/1). STIPITIPELLIS a cutis. CAULOCYSTIDIA

and HYMENIAL CLUSTERS absent. OLEIFEROUS HYPHAE, LIPOID GLOBULES and

BRILLIANT GRANULES absent in the trama; basidia with abundant granules.

PIGMENTATION colorless in the pileipellis, parietal in the entangled layer.

CLAMP CONNECTIONS present in all tissues.

ECOLOGY & DISTRIBUTION — Solitary in leaf humus, Mossman and Daintree

National Parks, far Northern Queensland, mid-March.

ADDITIONAL COLLECTIONS EXAMINED — AUSTRALIA. QUEENSLAND, Cook Region,

Daintree National Park, Tribulation section, Emmagen Creek Track, 16°02’19.7”S

145°27’41.7”E, 15 March 2010, DL Largent 9783.

DISTINCTIVE CHARACTERS — Tricholomatoid basidiomata, subfelty brown

pileus, lamellae with droplets drying as reddish brown spots, unpleasant taste

and odor, and a pileipellis as an anticlinal layer of entangled slender colorless

hyphae with narrow 3-5 um wide terminal cells.

CoMMENTS — Phylogenetically Entoloma guttulatum clusters with E. kewarra,

E. pamelae, and E. illinitum (from Guyana) but few morphological features

Entoloma spp. nov. (Australia) ... 357

NGS

Wasa t*

Cine

eee ioe

eek

PiaTE 11. Entoloma guttulatum (DLL9791 holotype). A. Basidiospores. B. Basidia, basidioles.

C. Pileipellis (radial section at pileus disc). D. Pileipellis (squash mount) E. Pileipellis (squash

mount) EF. Pileocystidium. Bars: A = 8 um; B = 20 um; C = 25 um; D = 35 um; E = 10 um; F=5 um.

are shared between species. For example, the basidiospores are very small and

distinctly angular in E. kewarra and E. pamelae, small and obscurely angular in

E. illinitum, and large and distinctly angular in E. guttulatum.

Entoloma pingue Corner & E. Horak (from a Casuarina forest in the Solomon

Islands), which shares a large fleshy stature, decurrent lamellae, and isodiametric

basidiospores (7-9 um diam.) with E. guttulatum, can be distinguished by

its pale fuscous pileus, stipe changing to pale brown, white lamellae without

reddish spots, 30-40 um long basidia, and pigmented pileipellis (Horak 1980).

358 ... Largent, Bergemann, & Abell-Davis

Acknowledgments

Fieldwork in Australia was supported by the Largent family trust and we are

particularly grateful for the support of Pamela Largent. The Australian Tropical

Herbarium and the School of Marine and Tropical Biology, James Cook University

provided fieldwork and logistical support. The DNA sequences generated in this study

are based upon work supported by the National Science Foundation under Grant No.

DRI 0922922 awarded to Dr. Sarah Bergemann. Comments by Dr. Fernanda Karstedt on

an earlier draft, the two reviewers, Dr. Joseph EF Ammirati and Dr. Andrew S. Methven,

and by the Nomenclature Editor, Dr. Shaun Pennycook, were also helpful. We wish to

thank Peter Newling for sharing his collections from northern Queensland and for

providing the identification of vascular plants and habitat descriptions for taxa collected

in Mt. Hypipamee National Park, Ms. Pam O’Sullivan and Ms. Skye Moore for sharing

their collections and descriptions from New South Wales, and Ms. Kerri Kluting and

Dr. Fernanda Karstedt for assisting with sequencing.

Literature cited

Aime MC, Largent DL, Henkel TW, Baroni TJ. 2010. The Entolomataceae of the Pakaraima

Mountains of Guyana IV: new species of Calliderma, Paraeccilia and Trichopilus. Mycologia

102: 633-649. http://dx.doi.org/10.3852/09-162

Baroni TJ, Matheny PD. 2011. A re-evaluation of gasteroid and cyphelloid species of

Entolomataceae from Eastern North America. Harvard Papers in Botany 16: 293-310.

http://dx.doi.org/10.3100/0.25.016.0205

Baroni TJ, Hofstetter V, Largent DL, Vilgalys R. 2011. Entocybe is proposed as a new genus in

the Entolomataceae (Agaricomycetes, Basidiomycota) based on morphological and molecular

evidence. North American Fungi 6: 1-19. http://dx.doi:10.2509/naf2011.006.012

Bergemann SE, Largent DL, Abell-Davis SE. 2013. Entocybe haastii from Watagans National Park,

New South Wales, Australia. Mycotaxon 126: 61-70. http://dx.doi.org/10.5248/126.61

Co-David D, Langeveld D, Noordeloos ME. 2009. Molecular phylogeny and spore evolution of

Entolomataceae. Persoonia 23: 147-176. http://dx.doi.org/10.3767/003158509X480944

Eyssartier G, Randrianjohany E, Buyck B. 2012. Trois entolomes (Entolomatales, Basidiomycota)

a épicutis hymeniforme de la réserve spéciale dAmbohjitantely, Madagascar. Cryptogamie

Mycologie 33(2): 157-166. http://dx.doi.org/10.7872/crym.v33.iss2.2012.157

Gates GM, Noordeloos ME. 2007. Preliminary studies in the genus Entoloma in Tasmania I.

Persoonia 19: 157-226.

Horak E. 1976. On cuboid-spored species of Entoloma (Agaricales). Sydowia 28: 171-236.

Horak E. 1980. Entoloma (Agaricales) in Indomalaya and Australasia. Beihefte zur Nova Hedwigia

65: 1-352.

Karstedt F, Capelari M. 2010. New species and new combinations of Calliderma (Entolomataceae,

Agaricales). Mycologia, 102: 163-173. http://dx.doi.org/10.3852/09-019

Katoh K, Standley DM. 2013. MAFFT Multiple Sequence Alignment Software Version 7:

Improvements in Performance and Usability. Molecular Biology and Evolution: 30: 772-780.

http://dx.doi.org/10.1093/molbev/mst010

Kornerup A, Wanscher JH. 1978. Methuen handbook of colour 3rd ed. Richard Clay Ltd:

Chichester, Sussex.

Largent DL. 1986. How to Identify Mushrooms to Genus 1. Mad River Press Inc: Eureka, California.

Largent DL. 1994. Entolomatoid fungi of the western United States and Alaska. Mad River Press

Inc: Eureka, California.

Entoloma spp. nov. (Australia) ... 359

Largent DL, Abell-Davis SE. 2011. Observations on Inocephalus virescens comb. nov. and

Alboleptonia stylophora from northeastern Queensland. Mycotaxon 116: 231-245.

http://dx.doi.org/10.5248/116.231

Largent DL, Henkel TW, Aime MC, Baroni TJ. 2008. The Entolomataceae of the Pakaraima

Mountains of Guyana I: four new species of Entoloma s. str. Mycologia 100: 132-140.

http://dx.doi.org/10.3852/mycologia.100.1.132

Largent DL, Abell-Davis SE, Cummings GA, Ryan KL, Bergemann SE. 201la. Saxicolous

species of Claudopus (Agaricales, Entolomataceae) from Australia. Mycotaxon 116: 253-264.

http://dx.doi.org/10.5248/116.253

Largent DL, Bergemann SE, Cummings GA, Ryan KL, Abell-Davis SE, Moore S. 2011b. Pouzarella

(Agaricales, Entolomataceae) from New South Wales (Barrington Tops National Park) and

northeastern Queensland. Mycotaxon 117: 435-483. http://dx.doi.org/10.5248/117.435

Largent DL, Bergemann SE, Abell-Davis SE, Kluting KL, Cummings GA. 2013a. Three new

Inocephalus species with cuboid basidiospores from New South Wales and Queensland,

Australia. Mycotaxon 123: 301-309. http://dx.doi.org/10.5248/123.301

Largent DL, Bergemann SE, Abell-Davis SE, Kluting KL, Cummings GA. 2013b. Five Leptonia

species from central New South Wales and Queensland, Australia. Mycotaxon 125: 11- 35.

http://dx.doi.org/10.5248/125.11

Liu YJ, Whelen S, Hall BD. 1999. Phylogenetic relationships among Ascomycetes: evidence from an

RNA polymerase II subunit. Molecular Biology and Evolution 16: 1799-1808.

Manimohan P, Joseph AV, Leelavathy KM. 1995. The genus Entoloma in Kerala State, India.

Mycological Research 99: 1083-1097. http://dx.doi.org/10.1016/S0953-7562(09)80777-6

Manimohan P, Noordeloos, ME, Dhanya AM. 2006. Studies in the genus Entoloma (Basidiomycetes,

Agaricales) in Kerala State, India. Persoonia 19(1): 45-93.

Matheny PB. 2005. Improving phylogenetic inference of mushrooms with RPB1 and RPB2

nucleotide sequences (Inocybe; Agaricales). Molecular Phylogenetics and Evolution 35: 1-20.

http://dx.doi.org/10.1016/j.ympev.2004.11.014

Morgado LN, Noordeloos ME, Lamoureux Y, Geml J. 2013. Multi-gene phylogenetic

analyses reveal species limits, phylogeographic patterns, and evolutionary histories of key

morphological traits in Entoloma (Agaricales, Basidiomycota). Persoonia 31: 159-178.

http://dx.doi.org/10.3767/003158513X673521

Noordeloos ME. 1992. Entoloma s.l. in Fungi Europaei vol. 5. Ed. Candusso: Alassio, Italy.

Noordeloos ME. 2005. Entoloma s.l. in Fungi Europaei vol. 5a. Ed. Candusso: Alassio, Italy.

Noordeloos ME, Gates GM. 2012. The Entolomataceae of Tasmania. Fungal Diversity Research

Series 22: 1-399. http://dx.doi.org/10.1007/978-94-007-4679-4

Orton PD. 1991. A revised list of the British Species of Entoloma sensu lato. The Mycologist

5; 123-138.

Romagnesi H. 1956. Les Rhodophylles du Congo Belge d’aprés les recoltes de Mme Goossens-

Fontana. Bulletin du Jardin Botanique de Etat a Bruxelles 26: 137-182.

Stamatakis A. 2006. RAxML-VI-HPC: maximum likelihood-based phylogenetic analyses with

thousands of taxa and mixed models. Bioinformatics 22: 2688-2690.

http://dx.doi.org/10.1093/bioinformatics/btl446

Stamatakis A, Hoover P, Rougemont J. 2008. A rapid bootstrap algorithm for RAxML web servers.

Systematic Biology 57: 758-771. http://dx.doi.org/10.1080/10635 150802429642

Thiers B. 2012. Index Herbariorum: a global directory of public herbaria and associated staff.

New York Botanical Garden's Virtual Herbarium. http://sweetgum.nybg.org/ih/[accessed May

2012].

MY COTAXON

http://dx.doi.org/10.5248/129.361

Volume 129(2), pp. 361-364 October-December 2014

Leptocorticium indicum sp. nov. from India

SAMITA, S.K. SANYAL , & G.S. DHINGRA

Department of Botany, Punjabi University, Patiala 147 002, India

*CORRESPONDENCE TO: skskumar731@gmail.com

ABSTRACT — A new corticioid species, Leptocorticium indicum, is described on a decaying

log of Rhododendron arboreum from Uttarakhand state in India.

KEY worps — Agaricomycetes, Tehri Garhwal, Kaddukhal

While conducting fungal forays in Kaddukhal area of district Tehri Garhwal,

Uttarakhand, India, Samita and Sanyal collected an unknown corticioid fungus

on a decaying log of Rhododendron arboreum. On the basis of macroscopic

and microscopic features and comparison with available literature (Hjortstam

& Ryvarden 2002, Nakasone 2005, Bernicchia & Gorjon 2010, Gorjon & Saitta

2014), we found the species different but closely related to the known species

of Leptocorticium, hence the description of a new species in this genus. The

material was also analyzed by Prof. Nils Hallenberg, who supported the concept

of a new species within Leptocorticium.

Leptocorticium indicum Samita, Sanyal & Dhingra, sp. nov. PratEs 1, 2

MycoBANnkK 808626

Differs from Leptocorticium tenellum in having larger basidia, smaller broadly

ellipsoid to ovate to subglobose basidiospores, and presence of gloeocystidia and from

L. gloeocystidiatum in having broadly ellipsoid to ovate to subglobose basidiospores.

Type: India, Uttarakhand, Tehri Garhwal, Kaddukhal, on decaying log of Rhododendron

arboreum Sm. (Ericaceae), 20 August 2010, Samita 6092 (PUN, holotype).

ErymMo oey: The epithet refers to the country of the type collection.

Basidiocarp resupinate, effused, adnate, <80 um thick in section; hymenial

surface smooth, grayish orange; margins thinning, fibrillose, paler concolorous,

or indeterminate. Hyphal system monomitic. Generative hyphae <4.5 um

wide, septate, clamped, thin- to thick-walled, with or without oily contents;

362 ... Samita, Sanyal, & Dhingra

PLATE 1. Leptocorticium indicum (holotype). Basidiocarp.

basal hyphae parallel to substrate, less branched; subhymenial hyphae vertical,

more densely branched. Dendrohyphidia abundant, irregularly branched,

non-dextrinoid, with thin to slightly thickened walls. Gloeocystidia 61-72 x

7.5-9 um, flexuous, with round to moniliform tips, embedded to projecting

<40 um out of hymenium, with basal clamp and oily contents negative to

sulphovanillin. Basidia 41-51 x 6.6-7.7 um, clavate, 4-sterigmate, with basal

clamp, with or without oily contents; sterigmata <7.5 um long. Basidiospores

5.5-6.6 x 3.3-4.4 um, broadly ellipsoid to ovate to subglobose, thin-walled,

inamyloid, acyanophilous.

REMARKS— ‘This species is placed in Leptocorticium based on the presence

of a thin and poorly developed subiculum, catahymenium with abundant

dendrohyphidia, flexuous gloeocystidia, and clavate basidia. Leptocorticium

tenellum Nakasone differs from L. indicum by its larger, cylindrical to ellipsoid

to sub-fusiform basidiospores (7-9.5 x 3-5 um), absence of gloeocystidia,

and smaller, usually utriform basidia (15-26 x 4.5-8 um; Nakasone 2005).

Leptocorticium gloeocystidiatum Gorjon & Saitta differs from L. indicum by its

smaller, ellipsoid basidiospores (4-5 x 2.5-3 um) and smaller basidia (15-20 x

3-4 um; Gorjon & Saitta 2014).

Acknowledgements

The authors thank Head, Department of Botany, Punjabi University, Patiala, for

providing research facilities, Prof. Nils Hallenberg (Gothenburg, Sweden) for expert

comments and peer review, and Prof. B.M. Sharma (Department of Plant Pathology,

COA, CSKHPAU, Palampur, H.P., India) for peer review.

Leptocorticium indicum sp. nov.(India) ... 363

PLATE 2. Leptocorticium indicum (holotype).

1. Basidiospores. 2. Basidia. 3. Cystidium. 4. Dendrohyphidium.

5. Hyphae. 6. Vertical section through basidiocarp.

364 ... Samita, Sanyal, & Dhingra

Literature cited

Bernicchia A, Gorjon SP. 2010. Corticiaceae s.1. Fungi Europaei 12. Edizioni Candusso. Alassio,

Italia. 1008 p.

Gorjon SP, Saitta A. 2014. Leptocorticium gloeocystidiatum sp. nov. (Basidiomycota), a new

corticioid fungus from Sicily, Italy. Mycosphere 5(3): 406-409.

Hjortstam K, Ryvarden L. 2002. Leptocorticium, a new genus among the corticoid fungi

(Basidiomycotina, Aphyllophorales). Synopsis Fungorum 15: 22-25.

Nakasone KK. 2005. Leptocorticium (Corticiaceae s.l., Basidiomycota): new species and

combinations. Mycological Progress 4(3): 251-256.

http://dx.doi.org/10.1007/s11557-006-0128-2

MYCOTAXON

http://dx.doi.org/10.5248/129.365

Volume 129(2), pp. 365-372 October-December 2014

A new species of Podosphaera sect. Sphaerotheca

from China

Lu-CHAO BAI & ZHI-MIN CAo

College of Forestry, Northwest A&F University

Taicheng Road, Yangling, Shaanxi 712100 China

* CORRESPONDENCE TO: zmcao@nwsuaf.edu.cn

ABSTRACT — Podosphaera girardiniae (Erysiphaceae, Podosphaera sect. Sphaerotheca),

identified in China on Girardinia suborbiculata, is described, illustrated, and compared with

allied species. The phylogeny of this new species has been inferred from internal transcribed

spacer (ITS) and 28S rDNA sequence analyses.

Key worps — molecular phylogeny, morphology, taxonomy

Introduction

In the autumn of 2013, Girardinia suborbiculata (Urticaceae) was identified

in Shaanxi, China, that was heavily infected by a powdery mildew. Based on

the asexual morph characterized by catenescent conidia with distinct fibrosin

bodies and chasmothecia with a single ascus and mycelioid appendages, this

collection was easily assignable to Podosphaera sect. Sphaerotheca (Lév.) de Bary,

as currently circumscribed and recognized (Braun & Cook 2012). This section

consists of 75 species, with 57% of the host plants belonging to the Rosaceae

(Takamatsu et al. 2010). Given the relatively large peridial cells, the powdery

mildew on Girardinia can be placed in Podosphaera subsect. Magnicellulatae

(U. Braun) U. Braun & Shishkoff (Braun & Cook 2012).

Podosphaera parietariae (Schwarzman) U. Braun & S. Takam. on Parietaria

spp. in Asia and Europe is the only species on an urticaceous host assigned to

Podosphaera sect. Sphaerotheca, where it has been placed based on its small

(8-25 um diam.) peridial cells (Braun 1987, Braun & Takamatsu 2000, Braun &

Cook 2012). Its morphology easily eliminates P. parietariae as the causal agent

of the Girardinia infection in China. However, in subsect. Magnicellulatae,

plurivorous races must be taken into consideration, particularly the P xanthii

(Castagne) U. Braun & Shishkoff complex to which the collection on Girardinia

366 ... Bai & Cao

appears to belong. Therefore, it was necessary to perform molecular sequence

analyses to elucidate the taxonomic status of the powdery mildew involved.

Materials & methods

Living leaves of Girardinia suborbiculata bearing the holomorph of a powdery mildew

were collected in October 2013 in the Qinling Mountains within Lueyang County in

China. Herbarium specimens were deposited in the Mycological Herbarium of Forestry

College, Northwest A & F University, Yangling, Shaanxi Province, China (HMNWAFU-

CF) and the herbarium of Martin Luther University, Halle (Saale), Germany (HAL).

The specimen was mounted in distilled water and examined using light microscopy

(Olympus, CX31RTSF, Japan). The teleomorphic features of the fungus, including

chasmothecia, appendages, asci, and ascospores, were described, measured, and

photographed. A scanning electron microscope (JEOL, JSM-6360LV) was used to

observe the anamorph ultrastructure, particularly the surface features of conidia (Cook

et al. 1997) and appressoria of this fungus; SEM images were taken.

Genomic DNA was extracted from chasmothecia using Chelex-100 (Walsh et al.

1991; Hirata & Takamatsu 1996). The ITS region of the nuclear rDNA (including 5.8S

and 28S rDNA sequences with domains D1 and D2) were amplified via polymerase

chain reaction (PCR) using primers designed for each region: ITS1 and ITS4 (White et

al. 1990) were used to amplify the ITS region, while LSU1 and LSU2 (Scholin et al. 1999)

were used to amplify the 28S rDNA sequence.

The PCR assays were conducted in a 50 uL final volume (Hirata & Takamatsu 1996)

containing 27 uL of 2x BoisTaq PCR MasterMix, 1 uL of each of primer, 1 uL of the

extracted DNA and 20 uL of ddH20 (Hirata & Takamatsu 1996). Thermal cycling in

a PTC-200 thermal cycler (BioRad) comprised an initial denaturation step at 95°C for

5 min, 35 cycles of 94°C for 1 min + 60°C for 1 min + 72°C for 1 min, and a final

elongation step at 72°C for 8 min. A negative control for each set of reactions replaced

template DNA with ddH,O. The PCR products were separated by electrophoresis on a

2% agarose gel in TAE buffer and purified using the Zymoclean™ Gel DNA Recovery

Kit, according to the manufacturer's instructions. The purified DNA products were

ligated into the pMD18-T vector (Takara) and transformed into E. coli DH5a cells. The

cloned fragments were sequenced by Sangon Biotech (Shanghai) Co., Ltd.

All DNA sequences were aligned using Clustal X 1.81 (Thompson et al. 1997), and

the alignments were adjusted following Nei & Kumar (2000). All positions containing

gaps or missing data were eliminated from the dataset. Cladistic trees were constructed

using the neighbor-joining method with the Kimura 2-parameter substitution model in

MEGA 4.0 (Tamura et al. 2007). Branch robustness was assessed by bootstrap analysis

with 1,000 replicates.

Taxonomy

Podosphaera girardiniae Z.M. Cao & L.C. Bai, sp. nov. PLATES 1-3

MycoBank MB 810810

Morphologically and phylogenetically similar to Podosphaera xanthii but sufficiently

distinct genetically to be considered a separate species.

Podosphaera girardiniae sp. nov. (China) ... 367

PL. 1. Podosphaera girardiniae (Holotype). A. asci; B. chasmothecia and appendages.

368 ... Bai & Cao

Px. 2. Podosphaera girardiniae (Holotype). A. conidia and conidiophore; B. conidium;

C. conidiophore, conidia, and hyphae. Scale bars: A, B = 10 um; C = 50 um.

Type: China, Shaanxi, Qinling Mountains, Lueyang County, 33°19’29”N 106°08’29’E;

alt. 840 m, on living leaves of Girardinia suborbiculata C.J. Chen (Urticaceae), Oct.

2013, L.C. Bai (Holotype, HMNWAFU-CE 2013125; isotype, HAL 2650 F; GenBank

KJ540945, KJ540944).

EryMo_oey: referring to the host genus.

Mycelium mainly foliicolous, amphigenous, effuse or in irregular patches,

later confluent, often covering the entire leaf surface, white, persistent. Hyphal

appressoria indistinct to slightly nipple-shaped. Hyphae smooth or almost

smooth, 3-8 um wide, colorless. Conidiophores arising from the upper

surface of hyphal mother cells, erect, straight to somewhat curved, foot-

cells cylindrical, about 34-76(-92) x 9-15 um, followed by 1-3 shorter cells,

forming catenescent conidia. Conidia ellipsoid-ovoid, doliiform, 25-36 x 15-

21 um, with a length/width ratio of 1.4-2.1. Chasmothecia mainly epiphyllous,

gregarious, 82-133 um diam. , subglobose; peridium cells conspicuous, large,

irregularly shaped, 14-44 um diam, appendages usually few, about 2-14, in

the lower half, mycelioid, simple, unbranched, sometimes interlaced with each

other, about 0.4-3.9 times as long as the chasmothecial diam. (up to about

410um), 5-9 um wide, brown below and paler towards the tip or most of

the short appendages brown throughout, 0-4-septate, walls thin, smooth or

almost so; ascus broadly ovoid or broadly ellipsoid-subglobose, 71-96 x 61-

77 um, almost sessile, terminal oculus 15-22 um diam., wall 1.5-3 um thick,

Podosphaera girardiniae sp. nov. (China) ... 369

PL. 3. Podosphaera girardiniae (Holotype). A. chasmothecia and appendages; B. asci;

C. conidia; D. conidia and conidiophore. Scale bars = 50 um.

6-8-spored; ascospores broadly ellipsoid-ovoid, 16-23 x 13-17 um, colorless,

development relatively late.

Phylogeny

28S ANALYSIS PL. 4

The 28S rDNA sequence comprised 632 total characters and was deposited

in GenBank under accession number KJ540945. The sequence was aligned

370 ... Bai & Cao

Erysiphe mori AB022418

Erysiphe carpinicola AB252472 | Erysipheae

Erysiphe adunca var. adunca AB022374

Golovinomyces cichoracearum vat. cichoracearum AB022360

Arthrocladiella mougeotii AB022379 | Gol lovinomyc eteae

Neoerysiphe galeopsidis AB022369

Pleochaeta turbinata AB218773

Phyllactinia moricola AB022401 | Phyllactinieae

Leveillula taurica AB022387

a as: Cystotheca wrightii AB022355

100| > Sawadaea tulasnei AB022366

L| 100) . Podosphaera tridactyla var. tridactyla AB022393

99 Podosphaera longiseta AB022423 Cystotheceae

3g Podosphaera fusca AB525914

3—_ Podosphaera girardiniae KJ540945

Podosphaera xanthii AB462786

Blumeria graminis AB022399 Blumerieae

Byssoascus striatosporus U17912

Px. 4. Neighbor-joining tree based on distances derived from sequences of the 28S rRNA genes

from 17 taxa of Erysiphaceae, with Byssoascus striatosporus as outgroup. The bar indicates a

distance of 0.01.

with 15 sequences representing the five tribes of Erysiphaceae. The 28S rDNA

phylogenetic tree places the Podosphaera girardiniae sequence in a strongly

supported clade (bootstrap value = 98%) where it clusters with P xanthii,

clearly supporting its placement in the genus Podosphaera, probably within

subsect. Magnicellulatae. Byssoascus striatosporus (G.L. Barron & C. Booth)

Arx was used as outgroup.

ITS ANALYSIS PL. 5

The ITS rDNA sequence analysis comprised 499 total characters and was

deposited in GenBank under accession number KJ540944. The sequences were

aligned with 14 sequences representing Podosphaera Kunze emend. U. Braun

& S. Takam. Cystotheca wrightii Berk. & M.A. Curtis was used as outgroup.

The ITS phylogenetic tree placed Podosphaera xanthii and P. girardiniae in one

clade with 79% bootstrap support. The P. girardiniae sequences differ from all

available sequences representing other P. sect. Sphaerotheca species, including

P. xanthii. The phylogenetic analyses of the rDNA sequences indicated that

Podosphaera on Girardinia suborbiculata represents a new species.

Discussion

The only other Podosphaera known on an urticaceous host, P parietariae,

clearly differs morphologically from P girardiniae, and its chasmothecia

Podosphaera girardiniae sp. nov. (China) ... 371

Podosphaera leucotricha AB027231

Podosphaera clandestina AB026137

Podosphaera spiraeae AB026143

Podosphaera ferruginea var. ferruginea AB026152

Podosphaera aphanis var. aphanis AB026141

Podosphaera pannosa AF011323

Podosphaera fugax AB026134

Podosphaera cercidiphylli AB026140

Podosphaera fuliginea AB026144

Podosphaera girardiniae KJ540944

Podosphaera xanthii D84387

Podosphaera elsholtziae AB026142

Podosphaera fusca AFO11320

Podosphaera longiseta AB000945

Podosphaera tridactyla var. tridactyla AB000943

Cystotheca wrightii AB000932

0.01

Px. 5. Neighbor-joining tree based on distances derived from the ITS1, ITS2, and 5.88 rRNA gene

sequences from 15 Podosphaera taxa, with Cystotheca wrightii as outgroup. The bar indicates a

distance of 0.01.

with small peridial cells place it in subsect. Sphaerotheca. Our phylogenetic

analyses support a close affinity of P girardiniae with P. xanthii, from which it

is morphologically barely distinguishable. Nonetheless the genetic differences

are too significant to assign the Girardinia powdery mildew to P. xanthii.

The sequence selected for P. xanthii was previously used by other authors

(e.g., Kousik et al. (2011), who demonstrated that this sequence and other

sequences of P xanthii from several cucurbits, beans and exotic impatiens

species share 100% ITS similarity). These genetic differences support the

Girardinia powdery mildew as a new species.

Acknowledgments

This paper was supported by the Natural Science Foundation of Shaanxi Province

(No. 2010JZ003). We wish to thank U. Braun (Herbarium, Martin Luther University,

Germany) for his valuable suggestions based on the examination of type material of

Podosphaera girardiniae. We also thank Prof. Guangyu Sun (Northwest A&F University,

China) and Dr. Braun for their expert reviews.

Literature cited

Braun U. 1987. A monograph of the Erysiphales (powdery mildews). Beihefte zur Nova Hedwigia

89: 1-700.

Braun U, Cook RTA. 2012. Taxonomic manual of the Erysiphales (powdery mildews). CBS

Biodiversity Series 11: 1-707.

372 ... Bai & Cao

Braun U, Takamatsu S. 2000. Phylogeny of Erysiphe, Microsphaera, Uncinula (Erysiphaceae) and

Cystotheca, Podosphaera, Sphaerotheca (Cystotheceae) inferred from rDNA ITS sequences —

some taxonomic consequences. Schlechtendalia 4: 1-33.

Cook RTA, Inman AJ, Billings C. 1997. Identification and classification of powdery mildew

anamorphs using light and scanning electron microscopy and host range data. Mycol .Res. 101:

975-1002. http://dx.doi.org/10.1017/S095375629700364X

Hirata T, Takamatsu S. 1996. Nucleotide sequence diversity of rDNA internal transcribed spacers

extracted from conidia and cleistothecia of several powdery mildew fungi. Mycoscience 37:

283-288. http://dx.doi.org/10.1007/BF02461299

Kousik CS, Danahoo RS, Webster CG, Turechek WW, Atkins ST, Roberts PD. 2011. Outbreak of

cucurbit powdery mildew on watermelon fruit caused by Podosphaera xanthii. Disease Notes

95: 1586.

Nei M, Kumar S. 2000. Molecular evolution and phylogenetics. Oxford University Press. New York.

Scholin CA, Marin III R, Miller PE, Doucette GJ, Powell CL, Haydock P, Howard J, Ray J. 1999.

DNA probes and a receptor-binding assay for detection of Pseudonitzschia (Bacillariophyceae)

species and domoic acid activity in cultured and natural samples. J. Phycol. 35: 1356-1367.

http://dx.doi.org/ 10.1046/j.1529-8817.1999.3561356.x

Takamatsu S, Niinomi S, Harada M, Havrylenko M. 2010. Molecular phylogenetic analyses reveal

a close evolutionary relationship between Podosphaera (Erysiphales: Erysiphaceae) and its

rosaceous hosts. Persoonia 24: 38-48. http://dx.doi.org/ 10.3767/003158510X494596

Tamura K, Dudley J, Nei M, Kumar S. 2007. MEGA4: Molecular Evolutionary

Genetics Analysis (MEGA) software version 4.0. Mol. Biol. Evol. 24: 1596-1599.

http://dx.doi.org/10.1093/molbev/msm092

Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG. 1997. The CLUSTAL X windows

interface: flexible strategies for multiple sequence alignment aided by quality analysis tools.

Nucleic Acids Res. 24: 4876-4882. http://dx.doi.org/10.1093/nar /25.24.4876

Walsh PS, Metzger DA, Higuchi R. 1991. Chelex 100 as a medium for simple extraction of DNA for

PCR-based typing from forensic material. Biotechniques 10: 506-513.

White TJ, Bruns TD, Lee S, Taylor J. 1990. Amplification and direct sequencing of fungal ribosomal

genes for phylogenetics. 315-322, in: MA Innis et al. (eds). PCR Protocols; a guide to methods

and applications. San Diego, Academic Press.

MY COTAXON

http://dx.doi.org/10.5248/129.373

Volume 129(2), pp. 373-386 October-December 2014

Rhizoglomus, a new genus of the Glomeraceae

EWALD SIEVERDING’, GLADSTONE ALVES DA SILVA’,

REINHARD BERNDT, & FRITZ OEHL?*

‘Institute for Plant Production and Agroecology in the Tropics and Subtropics,

University of Hohenheim, Garbenstrasse 13, D-70599 Stuttgart, Germany

*Departamento de Micologia, CCB, Universidade Federal de Pernambuco,

Av.da Engenharia s/n, Cidade Universitaria, 50740-600, Recife, PE, Brazil

3ETH Zurich, Plant Ecological Genetics, Universitatstr. 16, CH-8092 Ziirich, Switzerland

‘Agroscope, Federal Research Institute for Sustainability Sciences, Plant-Soil-Interactions,

Reckenholzstrasse 191, CH-8046 Ziirich, Switzerland

“CORRESPONDENCE TO: fritz.oehl@agroscope.admin.ch, fritz.oehl@gmail.com

Asstract — Rhizoglomus gen. nov. (Glomeraceae, Glomeromycetes) is proposed, typified

by Glomus intraradices [= Rhizoglomus intraradices]. The genus encompasses species of

arbuscular mycorrhizal fungi that frequently form abundant spores in soil and roots and

is morphologically characterized by spores with cylindrical subtending hyphae (usually

with an open pore at the base) and at least two or three (rarely up to five) distinct wall

layers. Phylogenetically, the genus forms a separate clade in the Glomeraceae. In addition to

R. intraradices, the genus includes R. aggregatum, R. antarcticum, R. arabicum, R. clarum,

R. custos, R. fasciculatum, R. invermaium, R. irregulare, R. manihotis, R. microaggregatum,

R. natalense, and R. proliferum. Some of these species were previously assigned to Rhizophagus

(type: R. populinus), a pathogenic genus that does not belong in the Glomeromycota.

Key worps — Glomerales, Glomus aggregatum, Glomus clarum, Glomus fasciculatum,

Glomus irregulare

Introduction

Glomus Tul. & C. Tul. s.l. (Glomeraceae, Glomeromycetes) comprises a

heterogeneous assembly of arbuscular mycorrhizal (AM) fungi (e.g., Morton

& Benny 1990, Schwarzott et al. 2001). In an attempt to redefine the genus

more naturally, Schifler & Walker (2010) reassigned several Glomus species

characterized by the frequent formation of intraradical spores to Rhizophagus

PA. Dang. Dangeard (1896), who identified Rhizophagus populinus as the

causal agent of a severe root disease of poplar (Populus spp.) in northern France,

374 ... Sieverding & al.

later described the root infection structures of the species (Dangeard 1900) but

neither designated nor deposited type specimens, as this was not the procedure

at that time. Consequently, the vague type description (Dangeard 1896) and

detailed drawings (Dangeard 1900) remain the only reference for Rhizophagus

and its type species, R. populinus.

Several decades later, Gerdemann & Trappe (1974) synonymized Rhizophagus

with Glomus. Later, reviewing the type description (Dangeard 1896), Schiifler

& Walker (2010) noted, “examination of the protologue of R. populinus reveals

that the species is an arbuscular mycorrhizal fungus that produces abundant

spores in roots.” For this reason they resurrected Rhizophagus, placing the

genus in the Glomeraceae. Here, we argue that the name Rhizophagus should

not be assigned to any species in the Glomeromycota.

First: Dangeard (1896) described (without providing any illustrations) a

disease of poplar roots occurring under wet soil conditions, which produced

“sporangia and finely ramified hyphae inside the root epidermal cells. The

morphology he observed led him to propose that the organism represented

a new genus in the Chytridineae. He related the death of the poplar trees to

this pathogen and not to a foliar disease. He did not observe germination of

the “sporangia” but explained that there were durable spores or cysts in the

root cells with a thick membrane and with protoplasm containing abundant

oil. Dangeard (1896) did not describe that numerous spores (or sporangia or

cysts) were formed in the infected poplar roots. The diagnosis of Rhizophagus

therefore, published by Schiifler & Walker (2010), does not correspond with

the observations described by Dangeard (1896).

SECOND: Dangeard (1900), through illustrations, showed that the infection

structures of R. populinus in poplar roots comprised two structural types:

i) those resembling vesicles (which he referred to as sporangia with dense

protoplast) and ii) arbuscules (termed ramified hyphae in cells) of mycorrhizal

root colonization. He did not recognize these structures as endomycorrhizae,

however. He also illustrated spores, which were not connected to the vesicles or

arbuscules. These spores were spherical, small (in relation to the root cells), and

with an apparently single wall layer and one or two hyphal connections formed

terminally or intercalary (see plate IX, D, E). Dangeard (1900) called these cells

‘cysts’ (‘kystes’ in French) because they contained a lipid globule. Recognizing

these structures as characteristic of chytrids, Dangeard (1900) assigned

R. populinus to the Chytridineae, a suborder containing root pathogenic fungi.

THIRD: Saccardo & Trotter (1912) transferred Rhizophagus to the

Peronosporaceae' and stated that “sporae perdurantes intercalares v. terminales,

‘In 1912 mycologists placed root-borne and root disease-causing fungi such as Pythium into the

Peronosporaceae (see Middleton 1952, for historic overview). Today Peronosporaceae contain

only foliar disease-causing fungi.

Rhizoglomus gen. & combs. nov. ... 375

globosae, membrana levi et guttulis pluribus praeditae.” Here, reference

is clearly made to the lipid globules in the globose spores (also drawn by

Dangeard (1900) in plate IX, Figs. D&E), a characteristic feature for spores

of some representatives of the Chytridiales and Oomycetes (Hawksworth et al.

1983). Saccardo & Trotter (1912) referred to R. populinus: “Characteres generis.

Hab. in radicibus junioribus Populi in Gallia occid. - Profunde penetrat et

plantas necat,’ which clearly states that R. populinus causes death of the host

plants. It must be noted, however, that early in the 20" century, fungal root

pathogens were separated from Peronosporaceae and placed in families such

as Pythiaceae (Middleton 1952). To our knowledge, no one ever officially

transferred Rhizophagus to Pythiaceae or any other family in the Oomycetes.

FourtTH: Petri (1919) argued that the structures depicted by Dangeard (1900)

in poplar tree roots belonged to the endomycorrhizae and were vesicles and

arbuscules, not the cause of death of the poplar trees. However, Petri ignored

the cysts with lipid globules illustrated by Dangeard (1900). He believed that

the poplar tree deaths in Northern France were resulted from a foliar disease

and that R. populinus was not causing root disease.

FirtH: Butler (1939), who reviewed vesicular-arbuscular mycorrhizal

fungi, showed that extramatrical fruiting bodies of Endogonaceae (including

some species currently assigned to Glomeraceae) were connected with internal

structures resembling those depicted by Dangeard (1900) that were later

identified as vesicles and arbuscules. Butler concluded that the vesicles and

arbuscules formed in roots represented the “imperfect form” of Endogonaceae,

a form named by that Dangeard (1900) as R. populinus. His statement had far-

reaching consequences: for a decade Rhizophagus was considered the most

widely distributed fungus in the world, until Mosse (1953) successfully isolated

the mycorrhizal spores and vesicular-arbuscular mycorrhizal structures

from roots during inoculation experiments. Mosse (1953, 1970) showed that

different spore “types” (i.e., species), such as “yellow vacuolated” and “honey-

colored” spores, formed arbuscules and vesicles in roots.

SIXTH: Gerdemann (1968) concluded, “..there is little reason to continue

to use the name Rhizophagus [for a fungus forming vesicles and arbuscules in

roots] ... as the genus Endogone was described much earlier.’ It should be noted

that by 1968 vesicular-arbuscular mycorrhiza-forming fungi were organized

in Endogone (Endogonaceae). A few years later Gerdemann & Trappe (1974)

separated many species from the Endogonaceae, placing them into the genus

Glomus, earlier described by Tulasne & Tulasne (1844). Following Gerdemann’s

(1968) reflections, the vesicles and arbuscules depicted by Dangeard (1900)

could not be a morphological character restricted to Rhizophagus, as Glomus,

Sclerocystis (described by Berkeley & Broome in 1873), and most other

glomeromycete genera form such mycorrhizal structures.

376 ... Sieverding & al.

SEVENTH: Gerdemann & Trappe (1974) synonymized Rhizophagus with

Glomus without providing a specific justification, which is intriguing, given

that Gerdemann (1968) had earlier doubted the validity of applying the name

R. populinus to a vesicular-arbuscular fungal species. Indeed, at this point

Gerdemann & Trappe (1974) should have eliminated the name Rhizophagus

from the Endogonaceae.

Although Dangeard (1896, 1900) interpreted vesicles of a vesicular-

arbuscular mycorrhizal fungus in poplar roots as sporangia, it is now clear that

this was incorrect and that the spores (cysts) he described were not, in fact,

released from these sporangia. ‘This observation was also made after Schlicht

(1889) had already described AM type endomycorrhizae and determined that

numerous fungal species could be involved. It has since been established that

AM vesicles and arbuscules are so morphologically uniform that they cannot be

used to differentiate species, genera, families, or orders of in the Glomeromycota

(Oehl et al. 2011a,b,c). Further, molecular techniques have demonstrated that

single roots can be colonized simultaneously by numerous glomeromycete

species (e.g., Lekberg et al. 2007, Tchabi et al. 2009, Borriello et al. 2012, Wetzel

et al. 2014). Consequently, we assume that the vesicle and arbuscule structures

delineated by Dangeard (1900) most likely resulted from combined infections

of a number of species representing different orders of the Glomeromycota.

It is well established that poplar trees host non-pathogenic vesicular-

arbuscular mycorrhizal fungi (Clark 1963), and it is likely that Dangeard (1900)

observed such vesicles and arbuscules in poplar root cells in addition to spores

of a disease-causing fungus such as R. populinus. Dehne (1982) and Linderman

(1994) state that fungal root pathogens can occupy root cortical cells adjacent

to those colonized by AM fungi, and that mycorrhizal colonization of a root

can never confer complete immunity against any root disease (see Bagyaraj

1984). Such colonization by AM fungi in close vicinity of pathogens such

as Pythium or Rhizoctonia has frequently been reported (e.g., Dehne 1982,

Linderman 1994).

Additionally confusing, with respect to Rhizophagus and its type species,

is that to date, no vesicular-arbuscular mycorrhiza-forming species of the

Glomeromycota has been recorded as a root pathogen that kills a plant. This

would represent a fundamental change to the current standing of vesicular-

arbuscular mycorrhiza as the most widespread mutualistic symbiosis between

flowering plants and soil-borne microorganisms.

It is actually quite remarkable that researchers and taxonomists have paid

attention to only the vesicle and arbuscule structures described by Dangeard

(1900) while after 1912 ignoring or overlooking the cysts he described (Dangeard

1896, 1900). According to our historical analysis, Dangeard (1900) described

the real species in the last paragraph: “The cysts of Rhizophagus populinus are

Rhizoglomus gen. & combs. nov. ... 377

spherical; the content is oily (P. IX, E); some even contain a large drop of oil

(P. IX, D).” Saccardo & Trotter (1912) identified these cysts (spores) as the

causal agent of a mortal poplar disease. ‘Thus the actual Rhizophagus populinus

cannot belong to the Glomeromycota, but rather represents Oomycetes.

We therefore conclude that the genus name Rhizophagus cannot be

applied to a group of glomeromycotan fungi that form spores in plant roots,

as suggested by SchiifSler & Walker (2010). Hence, we propose a new genus,

Rhizoglomus, to accommodate vesicular-arbuscular mycorrhizal fungal species

of the Glomeraceae that often form abundant spores in the soil and roots of

vascular plants.

Materials & methods

Specimens and original species descriptions of ten Glomus spp., one Endogone sp.,

and two recently described Rhizophagus spp. (Symanczik et al. 2014, Blaszkowski et al.

2014) were re-examined. Type specimens, ex-type material (specimens or single species

cultures either separated or propagated from original type material, and deposited at

a herbarium or collection that was not mentioned in the protologue), and non-type

specimens were provided by the herbaria OSC (Corvallis, Oregon), Z+ZT (Zurich),

DEPE (Szczecin, Poland), URM (Recife, Brazil), Culture Collection of Vesicular-

Arbuscular Mycorrhizal Fungi (INVAM), Swiss collection of Arbuscular mycorrhizal

Fungi (SAF, Agroscope), and the private AM fungal collections of Sieverding, Oehl, and

Blaszkowski. The Hall & Abbott (1979) photographic slide collection was also reviewed.

For this paper, all original species descriptions and published species emendations were

also considered.

Specimens were either mounted in lactophenol, in polyvinyl alcohol-lactic acid-

glycerol (PVLG), in Melzer’s reagent, in a mixture of PVLG + Melzer’s reagent (1:1;

Brundrett et al. 1994), in a mixture of lactic acid to water (1:1), or in water. When

available, spores freshly isolated from soils or bait cultures were also mounted and

analyzed. Spore wall terminology follows the nomenclature of Oehl et al. (2005,

2011b). Spore walls, germination structures, and all other mycorrhizal structures were

investigated using a compound microscope at 100-1000x magnifications. All spore

observations and all information on spore characteristics are based on spores extracted

from soil, bait cultures, or single species cultures. No information is provided from in

vitro-cultured material.

Results

Taxonomic analyses

Rhizoglomus Sieverd., G.A. Silva & Oehl, gen. nov. FIGS 1-15

MycoBank MB 803191

Differs from Glomus by regularly open pores at the spore bases, and usually wide open

pore channels within the frequently cylindrical subtending hyphae.

TYPE SPECIES: Glomus intraradices N.C. Schenck & G.S. Sm. [= Rhizoglomus intraradices].

378 ... Sieverding & al.

ErymoLoey: from Latin: rhiza = root, referring to the often abundant hyphae,

arbuscules, vesicles and spores formed in roots; and Glomus, referring to the genus in

which the type species of Rhizoglomus was first described.

Spores formed in loose sporocarps, in clusters, or singly in soil, and frequently

in roots. Wall of the subtending hyphae continuous with the spore wall, and for

a certain distance concolorous with the spore wall, or slightly lighter in color.

Subtending hypha cylindrical or seldom slightly funnel shaped at spore base.

Pore at spore base regularly open, rarely closed by a septum. Spore walls show

more than one, and generally two to three (and up to five) distinct layers, of

which one or several of the outermost may separate when pressure is applied

to spores. Forming vesicular-arbuscular mycorrhizae, whose fungal structures

stain blue to dark blue in trypan blue.

New combinations

Rhizoglomus aggregatum (N.C. Schenck & G.S. Sm.) Sieverd., G.A. Silva & Oehl

comb. nov.

MycoBank MB 803193

= Glomus aggregatum N.C. Schenck & G.S. Sm., Mycologia 74: 80. 1982.

SPECIMEN EXAMINED: Type described from a pot culture by Schenck & Smith (1982).

Isolated from Florida State near Haines City. Holotype OSC #40255, examined by Oehl

at OSC in 2002.

Rhizoglomus antarcticum (Cabello) Sieverd., G.A. Silva & Oehl comb. nov.

MycoBank MB 803194

= Glomus antarcticum Cabello, Mycotaxon 51: 124. 1994.

SPECIMEN EXAMINED: Type described from rhizospheric soil of Dechampsia antarctica

at Danco Coast, Base Primavera, Antarctic Peninsula (Argentina), by Cabello et al.

(1984). Specimen was loaned by M. Cabello to B.T. Goto (UFPE, Recife, Brazil) in 2009

and examined by Oehl at URM.

Fics 1-12. Spores of Rhizoglomus spp. from soils or within roots. Rhizoglomus intraradices

(from Germany, ZT Myc 30479): 1. Spores formed within the host root, isolated from pot culture.

2. Spore with three wall layers (SWL1-3). Rhizoglomus irregulare (from Switzerland, ZT Myc

30483): 3. Spore with three wall layers (SWL1-3). Rhizoglomus aggregatum (type, OSC): 4. Spores

formed within the host root. 5. Spore with two wall layers (SWL1-2). Rhizoglomus microaggregatum

(type, OSC): 6. Spores formed within the host root; mature spores have two layers. Rhizoglomus

proliferum (ex type, ZT Myc 30484): 7. Spore with four wall layers (SWL1-4). Rhizoglomus clarum

(from an INVAM culture): 8. Spore with three wall layers (SWL1-3). Rhizoglomus natalense (from

Natal, Brazil, isotype, DEPE 3384): 9. Spore with four wall layers (SWL1-4). Rhizoglomus custos

(ex type, ZT Myc 49021): 10. Spore with three (SWL1-3) of four wall layers described visible.

Rhizoglomus manihotis (from type, OSC): 11. Spore with evanescent outer wall layer (SWL1),

a rather thick hyaline layer (SWL2) and up to two separable yellow wall layers (SWL3). Rhizoglomus

fasciculatum (from Switzerland, ZT Myc 30478): 12. Spore with three wall layers (SWL1-3) and

open pore. Rhizoglomus invermaium (isotype, OSC): 13. Spores with two wall layers (SWL1-2).

Rhizoglomus gen. & combs. nov. ... 379

Rhizoglomus antarcticum (from type, M. Cabello collection): 14. Spore with two (SWL1-2) layers

visible (of three described). Rhizoglomus arabicum (isotype, DEPE): 15. Spore with 2-5 wall layers,

depending on layer and laminae definition, not easily discerned.

380 ... Sieverding & al.

Rhizoglomus arabicum (Blaszk., Symanczik & Al-Yahyaei) Sieverd., G.A. Silva &

Oehl comb. nov.

MycoBank MB 809436

= Rhizophagus arabicus Blaszk., Symanczik & Al-Yahyaei, Mycologia 106: 253. 2014.

SPECIMEN EXAMINED: Type described from two pot cultures by Symanczik et al. (2014).

Isolated from an undisturbed natural field at Al-Kamel in Al-Sharqyia region of Oman;

examined by Oehl in 2014. One slide deposited at Z+ZT (ZT Myc 57169).

Rhizoglomus clarum (T.H. Nicolson & N.C. Schenck) Sieverd., G.A. Silva & Oehl

comb. nov.

MycoBank MB 803195

= Glomus clarum T.H. Nicolson & N.C. Schenck, Mycologia 71: 182. 1979.

= Rhizophagus clarus (T.H. Nicolson & N.C. Schenck) C. Walker

& A. Schiissler, The Glomeromycota: 19. 2010.

SPECIMEN EXAMINED: Type described from a pot culture by Nicolson & Schenck (1979)

from soil on the Agronomy Farm of the University of Florida. Type at OSC, examined by

Oehl at OSC in 2002, and by Sieverding at Centro Internacional de Agricultura Tropical

(CIAT; Cali, Colombia) culture collection.

Rhizoglomus custos (C. Cano & Dalpé) Sieverd., G.A. Silva & Oehl comb. nov.

MycoBank MB 803196

= Glomus custos C. Cano & Dalpé, Mycotaxon 109: 502. 2009.

= Rhizophagus custos (C. Cano & Dalpé) C. Walker & A.

Schiissler, The Glomeromycota: 19. 2010.

SPECIMEN EXAMINED: Ex type specimens (Cano et al. 2009) isolated from cultures

maintained at EEZ, CSIC in Granada, Spain (collection date 15.9.2013 by J. Palenzuela;

ZT Myc 49021; examined by Oehl).

Rhizoglomus fasciculatum (Thaxt.) Sieverd., G.A. Silva & Oehl comb. nov.

MycoBank MB 803197

= Endogone fasciculata Thaxt., Proc. Am. Acad. Arts & Sci. 57: 308. 1922.

= Glomus fasciculatum (Thaxt.) Gerd. & Trappe, Mycol. Mem. 5: 51. 1974.

= Rhizophagus fasciculatus (Thaxt.) C. Walker & A.

Schiissler, The Glomeromycota: 19. 2010.

= Endogone arenacea Thaxt., Proc. Am. Acad. Arts & Sci. 57: 317. 1922

= Rhizophagites butleri Rosend., Bull. Torrey Bot. Club 70: 131. 1943.

SPECIMEN EXAMINED: Non-type specimen (morphology following Thaxter 1922; sensu

Walker & Koske 1987) collected in bait cultures from Switzerland, near Basel (collection

date 1.10.2005); examined by Oehl (ZT Myc 30478).

Rhizoglomus intraradices (N.C. Schenck & G.S. Sm.) Sieverd., G.A. Silva & Oehl

comb. nov.

MycoBank MB 803192

= Glomus intraradices N.C. Schenck & G.S. Sm., Mycologia 74: 78. 1982.

= Rhizophagus intraradices (N.C. Schenck & G.S. Sm.) C. Walker

& A. Schiissler, The Glomeromycota: 19. 2010.

Rhizoglomus gen. & combs. nov. ... 381

SPECIMEN EXAMINED: Type described from a pot culture by Schenck & Smith (1982).

Isolated from Florida State at the USDA Horticultural Research Station in Orlando.

Type OSC #40255, examined by Oehl at OSC in 2002. Non-type from a pot culture

(SAF51) established by Oehl, isolated from a temporary Lolium perenne grassland in

Biengen (Germany), near Freiburg im Breisgau (ZT Myc 30479); examined by Oehl.

Rhizoglomus invermaium (LR. Hall) Sieverd., G.A. Silva & Oehl comb. nov.

MycoBank MB 808609

= Glomus invermaium LR. Hall, Trans. Br. Mycol. Soc. 68: 345. 1977.

SPECIMEN EXAMINED: Isotypes after Hall (1977) deposited at OSC #43941. Hall

& Abbott (1979) photographic slide collection representing the type specimens

deposited at PDD herbarium; examined by Oehl. Several greenhouse pot cultures

of Gl. invermaium, established from single spores on host plant Hieracium pilosella

(so-called monosporic cultures) at SAF (see _ http://www.agroscope.admin.ch/

bodenoekologie/08050/08067/08068/index.html?lang=en; accessions SAF205-208)

were examined. Specimens from three cultures were deposited at Z+ZT (ZT Myc 57170-

57172), and first sequences from these cultures at EMBL-EBI (LN624111-LN6241112;

Sale et al., unpublished).

Rhizoglomus irregulare (Btaszk., Wubet, Renker & Buscot) Sieverd., G.A. Silva &

Oehl comb. nov.

MycoBank MB 803198

= Glomus irregulare Blaszk., Wubet, Renker & Buscot, Mycotaxon 106: 252. 2009.

= Rhizophagus irregularis (Btaszk., Wubet, Renker & Buscot) C.

Walker & A. Schiissler, The Glomeromycota: 19. 2010.

SPECIMEN EXAMINED: Type described from a pot culture by Blaszkowski et al. (2009).

Isolated from Poland near Szczecin. Type (DPP, nowadays DEPE). Type specimen

examined by Oehl at SAF from a type culture maintained in Basel in 2008 (ZT Myc

30480). Non-types from pot cultures established by Oehl. Isolated from three Swiss

long-term experiments in Therwil near Basel, in Frick and in Zollikofen near Bern,

respectively (ZT Myc 30481; ZT Myc 30482 and ZT Myc 30483); examined by Oehl in

2008-2014.

Rhizoglomus manihotis (R.H. Howeler, Sieverd. & N.C. Schenck) Sieverd., G.A.

Silva & Oehl comb. nov.

MycoBank MB 803199

= Glomus manihotis R.H. Howeler, Sieverd. & N.C. Schenck, Mycologia 76: 695. 1984.

= Rhizophagus manihotis (R.H. Howeler, Sieverd. & N.C. Schenck)

C. Walker & A. Schiissler, The Glomeromycota: 19.2010.

SPECIMEN EXAMINED: Type described from a pot culture on Pueraria phaseoloides

(Roxb.) Benth. by Schenck et al. (1984). Isolated from Colombia near Quilichao

(Cauca). Type OSC #41495, #41498, specimen examined by Oehl at OSC in 2002, and by

Sieverding who cultivated the type under the accession C-1-1 at CIAT (ZT Myc 30484).

Rhizoglomus microaggregatum (Koske, Gemma & P.D. Olexia) Sieverd., G.A. Silva

& Oehl comb. nov.

MycoBank MB 803200

= Glomus microaggregatum Koske, Gemma & P.D. Olexia, Mycotaxon 26: 125. 1986.

382 ... Sieverding & al.

SPECIMEN EXAMINED: Holotype described from soil by Koske et al. (1986) from USA,

Michigan, Benzie Co. Holotype OSC #46719; examined by Oehl at OSC in 2002.

Rhizoglomus natalense (Btaszk., Chwat & B.T. Goto) Sieverd., G.A. Silva & Oehl

comb. nov.

MycoBank MB 810477

= Rhizophagus natalensis Blaszk., Chwat & B.T. Goto, Mycotaxon 129: 100. 2014.

SPECIMEN EXAMINED: Type described from pot-cultured Plantago lanceolata by

Blaszkowski et al. (2014). Isolated from a sand dune in Parque Estadual das Dunas de

Natal “Journalista Luiz Maria Alves” (Rio Grande do Norte, Brazil); examined by Oehl

in 2014. One slide deposited at URM (URM 87580).

Rhizoglomus proliferum (Dalpé & Declerck) Sieverd., G.A. Silva & Oehl comb. nov.

MycoBank MB 803201

= Glomus proliferum Dalpé & Declerck, Mycologia 92: 1180. 2000.

= Rhizophagus proliferus (Dalpé & Declerck) C. Walker &

A. Schiissler, The Glomeromycota: 19. 2010.

SPECIMEN EXAMINED: Type described from monoxenic culture by Declerck et al. (2000)

from Neufchateau, Guadeloupe, France. Isotype MUCL 41827, examined by Oehl from

a pot culture maintained in Basel in 2008 (ZT Myc 30485).

Some species that superficially resemble Rhizoglomus species (e.g., G. diaphanum,

G. pallidum, and G. vesiculiferum) were not considered, as their spore formation

might differ from that of Rhizoglomus and their molecular phylogeny has not

yet been fully determined or remains unclear.

Discussion

The name of the new genus, Rhizoglomus, is based on the ability of its

species to form abundant spores, spore clusters, or sporocarps in the bulk

soil and soil rhizosphere and additionally within roots. Formation of spores

within roots of host plants is, however, not unique to Rhizoglomus. Certain

species of other, phylogenetically distant, genera also form spores inside roots,

e.g., Claroideoglomus lamellosum (Dalpé et al. 1992, as Glomus lamellosum),

Septoglomus xanthium (Blaszkowski et al. 2004, as G. xanthium), Archaeospora

myriocarpa (Schenck et al. 1986, as Acaulospora myriocarpa), and Entrophospora

infrequens (Sieverding & Toro 1985, Sieverding & Oehl 2006).

Rhizoglomus species, however, differ from other Glomeraceae (sensu Oehl

et al. 2011a,b) in that they possess a pore or channel that is usually open at

the connection between spore and the cylindric subtending hypha. One or

a few septa may form in the hyphae at some distance from the spore base.

Mature spores of most Rhizoglomus spp. have two or up to five distinct layers

of which one to three hyaline to subhyaline outer layers may separate from the

inner, persistent (= structural), and usually pigmented layer(s) when spores

are squeezed. In some species, such as R. intraradices and R. irregulare, the

Rhizoglomus gen. & combs. nov. ... 383

outer hyaline to subhyaline layers may degrade or slough away from the inner

persistent layer(s).

In spores of Glomus spp., the basal channel or pore is generally obstructed by

wall material or proper septa (Oehl et al. 2011b). In Funneliformis, the hyphal

attachment is characteristically funnel-shaped (Oehl et al. 2011b). Simiglomus

resembles Rhizoglomus in its similarly open-pored hyphal attachment, but its

hyphal walls are substantially thicker and thicken over longer distances than in

Rhizoglomus. Septoglomus spp. have regular septa or strong plugs at the spore

base (Oehl et al. 2011b). The taxonomic separation of Rhizoglomus based on

morphological characteristics was corroborated by molecular studies that

placed Rhizoglomus spp. in a separate clade (SchiiBler & Walker 2010, Kriiger

et al. 2012); this clade was formerly called the ,,Glomus group Ab2“ (Oehl et al.

2011b).

To date, no Rhizoglomus species has been reported to cause a root disease

resulting in the dieback or death of a plant. Rhizoglomus comprises ‘key species’

of AM fungi with regard to scientific and economic importance, such as

R. intraradices and R. irregulare (e.g., Stockinger et al. 2010). The latter, formerly

often confused with R. intraradices (Stockinger et al. 2010), develops well on

axenic root cultures (Chabot et al. 1992) and has therefore been used as a model

organism for AM fungal genetics (e.g., Jansa et al. 2002, Borstler et al. 2008,

Croll et al. 2008, 2009). These AM fungi are also key components of many of

the commercially available AM fungal products. Further from our extensive re-

examination, both morphologically and molecularly, of the various specimens

and cultures and our re-evaluation of the literature, we are convinced that such

important AM fungal species should be placed correctly in Glomeromycota and

not erroneously assigned to a pathogenic genus, such as Rhizophagus.

Acknowledgments

Fritz Oehl thanks Joey Spatafora and Richard Halse for the possibility to visit the

Oregon State University herbarium in Corvallis in 2002 and 2005. We are also thankful

to the collections managers of DEPE, INVAM, OSC, SAF, URM, and Z+ZT as well as

Janusz Blaszkowski (Szczecin, Poland), Marta Cabello (La Plata, Argentina), Bruno

Tomio Goto (Natal, Brazil), Sarah Symanczik (FiBL, Frick, Switzerland), Mohamed Al-

Yahyaei (Sultanate of Oman) for providing type specimens. This study was supported

by the Swiss National Science Foundation (SNE, 315230_130764/1), and by CNPq

and the Fundacao de Amparo a Ciéncia e Tecnologia do Estado de Pernambuco

(FACEPE) which provided a grant to F Oehl as ‘visiting researcher’ We acknowledge the

valuable comments and revisions of Danny Coyne (International Institute of Tropical

Agriculture—East Africa, Nairobi, Kenya), Danielle Karla Alves da Silva (Universidade

Federal do Vale de Sao Francisco, Petrolina, Brazil), and Ivan Sanchez-Castro (University

of Granada, Spain) and appreciate the corrections by Shaun Pennycook, Nomenclatural

Editor, and suggestions by Lorelei L. Norvell, Editor-in-Chief.

384 ... Sieverding & al.

Literature cited

Bagyaraj DJ. 1984. Biological interactions with mycorrhizal fungi. 131-153,in: CL Powell,

DJ Bagyaraj (eds). VA Mycorrhiza. CRC Press, New York.

Berkeley MJ, Broome CE. 1873. Enumeration of the Fungi of Ceylon. Part II., containing the

remainder of the Hymenomycetes, with the remaining established tribes of Fungi. J. Linn. Soc.

London Bot. 14: 65-140. http://dx.doi-org/10.1111/j.1095-8339.1873.tb00302.x

Btaszkowski J, Blanke V, Renker C, Buscot F. 2004. Glomus aurantium and G. xanthium,

new species in Glomeromycota. Mycotaxon 90: 447-467.

Blaszkowski J, Czerniawska B, Wubet T, Schafer T, Buscot F, Renker C. 2009. Glomus irregulare,

a new arbuscular mycorrhizal fungus in the Glomeromycota. Mycotaxon 106: 247-267.

Blaszkowski J, Chwat G, Goralska A, Goto BT. 2014. Rhizophagus natalensis, a new species in the

Glomeromycota. Mycotaxon 129(1): 97-108, http://dx.doi.org/10.5248/129.97

Borriello R, Lumini E, Girlanda M, Bonfante P, Bianciotto V. 2012. Effects of different management

practices on arbuscular mycorrhizal fungal diversity in maize fields as determined by molecular

techniques. Biol. Fert. Soils 48: 911-922. http://dx.doi.org/10.1007/s00374-012-0683-4

Borstler B, Raab PA, Thiéry O, Morton JB, Redecker D. 2008. Genetic diversity of the arbuscular

mycorrhizal fungus Glomus intraradices as determined by mitochondrial large subunit rRNA

gene sequences is considerably higher than previously expected. New Phytol. 180: 452-465.

http://dx.doi.org/10.1111/j.1469-8137.2008.02574.x

Brundrett M, Melville L, Peterson L. 1994. Practical methods in mycorrhizal research. Mycologue

Publications, University of Guelph, Guelph.

Butler EJ. 1939. The occurrences and systematic position of the vesicular-arbuscular type of

mycorrhizal fungi. Trans. Br. Mycol. Soc. 22: 274-301.

http://dx.doi.org/10.1016/S0007-1536(39)80052-3

Cabello MN, Gaspar ML, Pollero RJ. 1994. Glomus antarcticum sp. nov., a vesicular-arbuscular

mycorrhizal fungus from Antarctica. Mycotaxon 51: 123-128.

Cano C, Bago B, Dalpé Y. 2009. Glomus custos sp. nov., isolated from a naturally heavy metal polluted

environment in southern Spain. Mycotaxon 109: 499-512. http://dx.doi.org/10.5248/109.499

Chabot S, Becard G, Piche Y. 1992. Life cycle of Glomus intraradix in root organ culture. Mycologia

84: 315-312. http://dx.doi.org/10.2307/3760183

Clark FB. 1963. Endotrophic mycorrhizae influence yellow poplar seedling growth. Science 140:

1220-1221. http://dx.doi.org/10.1126/science.140.3572.1220

Croll D, Wille L, Gamper HA, Mathimaran N, Lammers PJ, Corradi N, Sanders IR. 2008 Genetic

diversity and host plant preferences revealed by simple sequence repeat and mitochondrial

markers in a population of the arbuscular mycorrhizal fungus Glomus intraradices. New Phytol.

178: 672-687. http://dx.doi.org/10.1111/j.1469-8137.2008.02381.x

Croll D, Giovannetti M, Koch AM, Sbrana C, Ehinger M, Lammers PJ, Sanders IR. 2009 Nonself

vegetative fusion and genetic exchange in the arbuscular mycorrhizal fungus Glomus

intraradices. New Phytol. 181: 924-937. http://dx.doi.org/10.1111/j.1469-8137.2008.02726.x

Dalpé Y, Koske RE, Tews LL. 1992. Glomus lamellosum sp. nov.: a new Glomaceae associated with

beach grass. Mycotaxon 43: 289-293.

Dangeard PA. 1896. Une maladie du peuplier dans louest de la France. Le Botaniste 5: 38-43.

Dangeard PA. 1900. Le Rhizophagus populinus Dangeard. Le Botaniste 7: 285-289.

Declerck S$, Cranenbrouck S, Dalpé Y, Séguin $, Grandmougin-Ferjani A, Fontaine J, Sancholle M.

2000. Glomus proliferum sp. nov.: a description based on morphological, biochemical, molecular

and monoxenic cultivation data. Mycologia 92: 1178-1187. http://dx.doi.org/10.2307/3761485

Dehne H-W. 1982. Interaction between vesicular-arbuscular mycorrhizal fungi and plant

pathogens. Phytopath. 72: 1115-1119.

Rhizoglomus gen. & combs. nov. ... 385

Gerdemann JW. 1968. Vesicular-arbuscular mycorrhiza and plant growth. Ann. Rev. Phytopathol.

6: 397-418. http://dx.doi.org/10.1146/annurev.py.06.090168.002145

Gerdemann JW, Trappe JM. 1974. The Endogonaceae in the Pacific Northwest. Mycol. Memoirs

5. 76 p.

Hall IR. 1977. Species and mycorrhizal infections of New Zealand Endogonaceae. Trans. Br. Mycol.

Soc. 68: 341-356. http://dx.doi.org/10.1016/S0007-1536(77)80186-1

Hall IR, Abbott LK. 1979. Photographic slide collection illustrating features of the Endogonaceae.

3rd edition. Invermay Agricultural Research Centre and Soil Science Department, University

of Western Australia, 27 p., plus 400 color transparencies.

Hawksworth DL, Sutton BC, Ainsworth GC. 1983. Ainsworth and Bisby’s dictionary of the fungi.

7* edition, CMI, Kew

Jansa J, Mozafar A, Banke S$, McDonald BA, Frossard E. 2002. Intra- and intersporal diversity of

ITS rDNA sequences in Glomus intraradices assessed by cloning and sequencing, and by SSCP

analysis. Mycol. Res. 106: 670-681. http://dx.doi.org/10.1017/S0953756202006032

Koske RE, Gemma JN, Olexia PD. 1986. Glomus microaggregatum, a new species in the

Endogonaceae. Mycotaxon 26: 125-132.

Kriiger M, Kriiger C, Walker C, Stockinger H, Schii®ler A. 2012. Phylogenetic reference data for

systematics and phylotaxonomy of arbuscular mycorrhizal fungi from phylum to species level.

New Phytol. 193: 970-984. http://dx.doi.org/10.1111/j.1469-8137.2011.03962.x

Lekberg Y, Koide RT, Rohr JR, Aldrich-Wolfe L, Morton JB, 2007. Role of niche restrictions and

dispersal in the composition of arbuscular mycorrhizal fungal communities. Journal of Ecology

95: 95-100. http://dx.doi.org/10.1111/j.1365-2745.2006.01193.x

Linderman RG. 1994. Role of VAM fungi in biocontrol. 1-25, in: FL Pfleger, RG Linderman (eds).

Mycorrhiza in Plant Health. Am. Phytopath. Soc. Press, St Paul, Minnesota.

Middleton JT. 1952. Generic concepts in the Pythiaceae. Tijdschrift over Plantenziekten 58(6):

226-235. http://dx.doi.org/10.1007/BF01988210

Morton JB, Benny GL. 1990. Revised classification of arbuscular mycorrhizal fungi (Zygomycetes):

a new order, Glomales, two new suborders, Glomineae and Gigasporinae, and two families,

Acaulosporaceae and Gigasporaceae, with an emendation of Glomaceae. Mycotaxon 37:

471-491.

Mosse B. 1953. Fructifications associated with mycorrhizal strawberry roots. Nature 171: 974.

http://dx.doi.org/10.1038/171974a0

Mosse B. 1970. Honey-coloured, sessile Endogone spores. I. Life history. Arch. Microbiol. 70:

167-175. http://dx.doi.org/10.1007/BF00412208

Nicolson TH, Schenck NC. 1979. Endogonaceous mycorrhizal endophytes in Florida. Mycologia

71: 178-198. http://dx.doi.org/10.2307/3759231

Oehl F, Redecker D, Sieverding E. 2005. Glomus badium, a new sporocarpic arbuscular mycorrhizal

fungal species from European grasslands of higher soil pH. J. Appl. Bot. Food Qual., Angew.

Bot. 79: 38-43.

Oehl F, Sieverding E, Palenzuela J, Ineichen K, Silva GA. 2011a. Advances in Glomeromycota

taxonomy and classification. IMA Fungus 2: 191-199.

http://dx.doi.org/10.5598/imafungus.2011.02.02.10

Oehl F, Silva GA, Goto BA, Sieverding E. 2011b. Glomeromycota: three new genera, and glomoid

species reorganized. Mycotaxon 116: 75-120. http://dx.doi.org/10.5248/116.75

Oehl F, Silva GA, Goto BT, Maia LC, Sieverding E. 2011c. Glomeromycota: two new classes and a

new order. Mycotaxon 116: 365-379. http://dx.doi.org/10.5248/116.365

Petri L. 1919. Sopra una presunta malattia del pioppo. Firenze: M. Ricci. 10 p. [extract from Annali

del R. Istituto Superiore Forestale Nazionale, vol. 4].

386 ... Sieverding & al.

Saccardo PA, Trotter A. 1912. Supplementum uiversale, pars VIII. Hymenomycetae-Phycomycetae.

Sylloge Fungorum 21. 928 p.

Schenck NC, Smith GS. 1982. Additional new and unreported species of mycorrhizal fungi

(Endogonaceae) from Florida. Mycologia 74: 77-92. http://dx.doi.org/10.2307/3792631

Schenck, NC, Spain JL, Sieverding E, Howeler RH. 1984. Several new and unreported vesicular-

arbuscular mycorrhizal fungi (Endogonaceae) from Colombia. Mycologia 76: 685-699.

http://dx.doi.org/10.2307/3793226

Schenck NC, Spain JL, Sieverding E. 1986. A new sporocarpic species of Acaulospora

(Endogonaceae). Mycotaxon 25: 111-117.

Schlicht A. 1889. Beitrag zur Kenntniss der Verbreitung und der Bedeutung der Mykorhizen.

Landwirtschaftl. Jahrbiicher 18: 477-505. http://dx.doi.org/10.5962/bhI.title.63757

Schiifler A, Walker C. 2010. The Glomeromycota. A species list with new families and new genera.

Gloucester, UK. 56 p.

Schwarzott D, Schifler A, Walker C. 2001. Glomus, the largest genus of the arbuscular

mycorrhizal fungi (Glomales), is nonmonophyletic. Mol. Phylogenetics Evol. 21: 190-197.

http://dx.doi.org/10.1006/mpev.2001.1007

Sieverding E, Oehl F. 2006. Revision of Entrophospora, and description of Kuklospora and

Intraspora, two new genera in the arbuscular mycorrhizal Glomeromycetes. J. Appl. Bot. Food

Qual., Angew. Bot. 80: 69-81.

Sieverding E, Toro S. 1985. The genus Entrophospora in Colombia. 621-626, in: V Gianinazzi-

Pearson, S Gianinazzi (eds). Physiological and genetical aspects of mycorrhizae. Proceedings

of the Ist European Symposium on Mycorrhizae, Dijon, 1-5 July 1985, INRA, Service des

Publications, Versailles, France.

Stockinger H, Walker C, Schiifdler A. 2010. “Glomus intraradices DAOM197198; a model fungus

in arbuscular mycorrhiza research, is not Glomus intraradices. New Phytol. 183: 1176-1187.

http://dx.doi.org/10.1111/j.1469-8137.2009.02874.x

Symanczik S, Blaszkowski J, Chwat G, Boller T, Wiemken A, Al-Yahyaei M. 2014. Three new species

of arbuscular mycorrhizal fungi discovered at one location in a desert of Oman: Diversispora

omaniana, Septoglomus nakheelum and Rhizophagus arabicus. Mycologia 106: 243-259.

http://dx.doi.org/10.3852/106.2.243

Tchabi A, Burger S, Coyne D, Hountondji FE, Lawouin L, Wiemken A, Oehl F. 2009. Promiscuous

arbuscular mycorrhizal symbiosis of yam (Dioscorea spp.), a key staple crop in West Africa.

Mycorrhiza 19: 375-392. http://dx.doi.org/10.1007/s00572-009-0241-6

Thaxter B. 1922. A revision of the Endogonaceae. Proc. Am. Acad. Arts Sci. 57: 291-341.

Tulasne LR, Tulasne C. 1844. Fungi nonnulli hypogaei, novi v. minus cogniti. Giorn. Bot. Ital.

1(2(7-8)): 55-63.

Walker C, Koske RE. 1987. Taxonomic concepts in the Endogonaceae: IV. Glomus fasciculatum

redescribed. Mycotaxon 30: 253-262.

Wetzel K, Silva G, Matczinski U, Oehl F, Fester T. 2014. Superior differentiation of arbuscular

mycorrhizal fungal communities from till and no-till plots by morphological spore

identification when compared to T-RFLP. Soil Biology & Biochemistry 72: 88-96.

http://dx.doi.org/10.1016/j.soilbio.2014.01.033

MY COTAXON

http://dx.doi.org/10.5248/129.387

Volume 129(2), pp. 387-395 October-December 2014

First report of the pantropical species

Diploschistes rampoddensis from Europe

SAMANTHA FERNANDEZ-BRIME”?, XAVIER LLIMONA’,

NEsTOR HLADUN’, & ESTER GAYA?

‘Department of Plant Biology, Facultat de Biologia, Universitat de Barcelona,

Barcelona, 08028, Spain

*Department of Botany, Swedish Museum of Natural History,

Stockholm, SE-104 05, Sweden

3Mycology section, Jodrell Laboratory, Royal Botanic Gardens,

Kew, Richmond, Surrey, TW9 3DS, U.K.

* CORRESPONDENCE TO: sfernandezbrime@ub.edu

ABSTRACT — The lichen species Diploschistes rampoddensis, previously known only from

a few localities in tropical and subtropical Asia and Oceania, is reported here for the first

time in Europe. A detailed description, including macro- and microscopic characters, and

comparisons with closely related taxa are also provided. Molecular analyses based on the

nrITS were used to confirm this new record for the European lichen biota.

Key worps — Ascomycota, BAli-phy, disjunct distribution, lichenized-fungi, Ostropales

Introduction

The lichen genus Diploschistes Norman (Graphidaceae, Ostropales,

Ascomycota) currently includes 45 species (Kirk et al. 2008, Pérez-Vargas et

al. 2012, Fernandez-Brime et al. 2013), distributed mainly in temperate and

arid regions of both hemispheres, with only a few species restricted to the

tropics (Lumbsch 1989). One of the few species occurring in tropical areas is

D. rampoddensis, ataxon that was first described by Nylander (1900, as Urceolaria

rampoddensis) from the locality of Rampodde (Sri Lanka). According to the

literature (Nylander 1900, Lumbsch 1993, Pant & Upreti 1993, Martin et al.

2003) and unpublished herbarium specimens, this species appears to be quite

rare and has previously been recorded only from a few localities in tropical and

subtropical Asia and Oceania.

As part of an ongoing project to assess the lichen diversity of several protected

areas of Catalonia (NE Spain), we intensively surveyed the cliffs of Sant Lloreng

388 ... Fernandez-Brime & al.

de la Muga, a protected area included in the Plan of Sites of Natural Interest

(PEIN). Among several Diploschistes collections, three samples were initially

identified as D. rampoddensis, based on their morphological, anatomical,

and chemical features. As these specimens were, however, rather scant or

inconspicuous, we checked the identification using the nuclear ribosomal

internal transcribed spacer (nrITS), recently selected as the universal DNA

barcode marker for fungi (Schoch et al. 2012). Our molecular analyses further

confirmed that these specimens represent D. rampoddensis, which extends its

distributional range to Europe.

Materials & methods

Morphological analysis

The morphological study was carried out on fresh specimens collected from

the cliffs of Sant Lloreng de la Muga and on herbarium material of Diploschistes

rampoddensis lodged and loaned from UPS, as well as on other taxa of Diploschistes

used for comparison. Morphological characters were examined with an Olympus

VMZ dissecting microscope (40x). Anatomical characters were studied on hand-cut

sections mounted in water (40x) or oil immersion (100x) using an Olympus CH-2 light

microscope. Vouchers of new specimens were conserved in CeDocBiV , Universitat

de Barcelona, Spain (BCN), Institute of Botany, Academy of Sciences, Czech Republic

(PRA), and the first author’s private herbarium (Hb. Fdez.-Brime, SFB).

Selected specimens were photographed using a Pixera Prol150 ES camera mounted

on an Olympus SZ60 stereomicroscope. For each selected specimen, multiple images in

different focal planes were taken and subsequently assembled into a final unique picture

using the software Helicon Focus 5.2.

Spot tests were performed using standard lichenological chemical reagents (K, C)

along with thin-layer chromatography (TLC) following the protocols of Orange et al.

(2001).

Molecular analysis

Genomic DNA was obtained from small pieces of thallus and apothecia sampled

from fresh specimens using a phenol-chloroform-isoamyl alcohol extraction protocol

based on Lee et al. (1988). Approximately 560 bp including the complete nrITS region

was amplified using the primers ITSIF (Gardes & Bruns 1993) and ITS4 (White et

al. 1990). PCR mixtures were prepared as in Gueidan et al. (2007) and conditions for

thermocycling followed Fernandez-Brime et al. (2013). The nrITS region was sequenced

as detailed in Martin & Winka (2000), using the same amplification primers.

We aligned the nrITS of the three specimens tentatively identified as Diploschistes

rampoddensis with the only nrITS sequence of D. rampoddensis available from GenBank,

together with other sequences of closely related representatives of Diploschistes subg.

Diploschistes (Fernandez-Brime et al. 2013). Diploschistes actinostomus (Ach.) Zahlbr.

and D. candidissimus (Kremp.) Zahlbr., from Diploschistes subg. Limborina Fdez.-Brime

et al. (Fernandez-Brime et al. 2013) were selected as outgroups to root the phylogeny.

A total of 17 sequences were used for the phylogenetic analyses (TABLE 1). All newly

Diploschistes rampoddensis in Europe ... 389

TABLE 1. Specimens and sequences of Diploschistes used in the present phylogenetic

analysis.

SPECIES VOUCHER ORIGIN GENBANK*

D. actinostomus BCC-Lich 13394 Spain, Catalonia AF229194

D. candidissimus ESS 20699 Australia, South Australia AJ458281

D. cinereocaesius ESS 9364 Venezuela, Mérida AJ458282

DUKE 0047509 Costa Rica, San José HQ650715

Hb. Palice, Palice 4471 Ecuador KJ542542

D. rampoddensis Hb. Lumbsch, Aptroot 39679 Papua New Guinea AJ458286

BCC-Lich 18009 Spain, Catalonia KC166992

BCC-Lich 18011 Spain, Catalonia KC166993

BCC-Lich 18008 Spain, Catalonia KJ542543

D. scruposus BCN-Lich 19326 Spain, Catalonia KC167067

Hb. Fdez.-Brime, SFB 59 Spain, Catalonia KJ542545

Hb. Fdez.-Brime, SFB 103 Spain, Catalonia KJ542546

ESS 21508 Germany, Rheinland-Pfalz AJ458287

D. scruposus “interpediens” = BCN-Lich 19319 Spain, Catalonia KC166995

BCN-Lich 19355 Spain, Catalonia KC166999

BCN-Lich 19322 France, Languedoc- KC167003

Roussillon

Hb. Fdez.-Brime, SFB 104 Spain, Catalonia KJ542544

* Newly obtained sequences shown in bold.

generated DNA sequences are now deposited in GenBank (KJ542542-KJ542546) and

the alignment is available in TreeBASE (http://www.treebase.org; ID number 15496).

Sequences were manually aligned with Mesquite 2.75 (Maddison & Maddison 2011).

Terminal primers and partial nuSSU and nuLSU were excluded from the alignment.

Phylogenetic relationships were inferred using weighted maximum parsimony (wMP)

and Bayesian methods (B). The wMP analyses were carried out in PAUP* v.4.0b10

(Swofford 2002), with gaps treated as a fifth character state in unambiguously aligned

regions. These regions were subjected to symmetric step matrices (i-e., ITS1, 5.88, and

ITS2 being treated separately), using STMatrix v.3.0 (F Lutzoni & S. Zoller, Dept. of

Biology, Duke University), as outlined in Gaya et al. (2003). Ambiguously aligned

regions were removed from the wMP searches and recoded and incorporated into

the analyses using INAASE v.2.3b (Lutzoni et al. 2000), as in Gaya et al. (2008). All

wMP analyses were performed using heuristic searches with 1000 random addition

sequences (RAS), the tree bisection-reconstruction (TBR) algorithm, MULTREES in

effect, and collapsing branches with maximum branch length equal to zero. Branch

support (BS) was assessed with 1000 bootstrap replicates (Felsenstein 1985) with full

heuristic searches, and 2 RAS per bootstrap replicate. In all BS analyses, the same

parameters as in the original MP search were used, and constant sites were excluded

from all analyses. The Bayesian analyses (B) were done using BAli-Phy 2.1.1 (Suchard

& Redelings 2006), a software that simultaneously accounts for alignment uncertainty

and phylogeny, allowing the inclusion of the whole alignment. In this case, the nrITS

dataset was divided into three partitions (ITS1, 5.88, and ITS2) as suggested in Gaya et

390 ... Fernandez-Brime & al.

al. (2011). The analyses were run under the following settings: ITS1 and ITS2 sharing

the same model of evolution (HKY+G), indel model (RSO7; Redelings & Suchard

2007) and branch length estimation, while the EQU model and separate branch length

estimation were applied to the partition 5.88, which was fixed because the alignment was

unambiguous. We ran 6 independent Markov chains of 50,000 iterations each, sampling

the Markov chains every 10 iterations. After discarding the first 1250 samples of each

run (25%), we computed the majority rule consensus with the remaining trees (22,500).

Results & discussion

Molecular phylogenetics

The final nrITS alignment included 17 taxa and comprised 594 characters.

We excluded from the wMP analyses 557 characters, of which 407 were

constant sites and 150 corresponded to 19 ambiguously aligned regions. The

characters from the ambiguously aligned regions were included as 19 INAASE

recoded characters. From the 56 variable characters analyzed by wMP, 40 were

parsimony-informative. We included the whole alignment for the B analyses.

The wMP search yielded six most parsimonious trees of 159.27 steps, which

were found in one island hit 991 times out of 1000 RAS. The strict consensus

tree (Fic. 1) shows 13 resolved internodes, of which nine were significantly

supported (BS =70%). The 50% majority rule consensus tree from the B

analyses showed slightly less resolution, with nine resolved internodes, but, as

with wMP, nine of these were significantly supported (PP 20.95).

The three European specimens putatively representing Diploschistes

rampoddensis clustered with the only sequence of D. rampoddensis from

GenBank and formed a well-supported clade (BS = 100%; PP = 1). This

clade was sister to D. cinereocaesius (Sw.) Vain., which provided support for

this sister relationship for the first time. In contrast, specimens of other taxa

[e.g., Diploschistes scruposus (Schreb.) Norman, D. scruposus “morphotypus

interpediens’)], even if morphologically similar and collected from the same

locality of Sant Lloreng¢ de la Muga (specimens marked with an asterisk in Fic.

1), clustered separately from D. rampoddensis.

Taxonomy

Diploschistes rampoddensis (Nyl.) Zahlbr., Cat. Lich. Univers. 2: 665, 1924. Fic. 2A

THALLUS saxicolous, crustose, yellowish grey to greenish yellow, without

pruina but with a crystalliferous surface, 0.2-0.9 um thick, friable (easily

broken), from verrucose-areolate to mostly reduced into conglutinated

granules; epinecral layer 24-31 um thick, without a well-developed cortex;

photobiont trebouxioid.

APOTHECIA urceolate, 0.2-1.3(-1.7) mm in diam., never subdivided, with

a deeply concave black disc with white pruina. Excrputum black, 60-100

81/ 0.96

78/ 0.96

62/-

80 / -

100 / 0.86

96/1

100 /1

55/1

-/0,99

100 /1

96/1

Diploschistes rampoddensis in Europe ... 391

D. scruposus KJ542545*

D. scruposus KC167067*

D. scruposus KJ542546*

D. scruposus AJ458287

D. scruposus “interpediens”

KC167003

D. scruposus “interpediens”

KC166995*

D. scruposus “interpediens”

KC166999

D. scruposus “interpediens”

KJ542544*

D. rampoddensis KC166993*

. rampoddensis KJ542543*

. rampoddensis KC166992*

rampoddensis AJ458286

. cinereocaesius AJ458282

. cinereocaesius KJ542542

cinereocaesius HQ650715

candidissimus AJ458281

5 0 FS BD FF 8 BP

actinostomus AF229194

1.0

Fic. 1. Phylogenetic relationships of sequences of Diploschistes species summarized by a strict

consensus tree of six equally most parsimonious trees, as revealed by the wMP analyses of the

nrITS dataset, including 19 INAASE recoded characters. Bootstrap support =50% (BP; left) and

posterior probabilities 20.5 (PP; right) are shown above branches. Internodes with BP values =70%

or PP 20.95 are highlighted by thicker lines. Asterisk-marked specimens collected in the same

locality of Sant Lloreng de la Muga (Catalonia, Spain). Newly generated sequences are presented

in bold font.

(-130) um thick, lateral paraphyses present. HYMENIUM hyaline to pale brown,

(78—)98-128(-134) um high. Asci clavate, unitunicate, non-amyloid walls, (6-)8

spores per ascus, 100-170 x 15-30 um. Ascosporss ellipsoid, pale to dark

brown, non-amyloid, (19.5—)21- 26.5(-29) x (8.5-)9-12(-13.5) um (n = 59),

with 4—7(-8) transverse septa, and 1-2 longitudinal septa.

CHEMISTRY — Spot test reactions of the thallus K— or K+ pale yellow,

C+ and KC+ pink (brief reaction). TLC: lecanoric acid as major compound,

compounds in traces were not found.

DISTRIBUTION & HABITAT — Since Diploschistes rampoddensis was first

described by Nylander (1900) from material collected in Rampodde, Sri

392 ... Fernandez-Brime & al.

Fic. 2. A. Diploschistes rampoddensis (BCN-lich. 18011): habit. B. D. scruposus “interpediens”

(BCN-lich. 14404): habit. C. D. cinereocaesius (BCN-lich. 18010): habit. Scale bars = 1 mm.

Lanka, it has been recorded from a few Asian and Australasian localities:

China (see examined specimens), India and Nepal (Pant & Upreti 1993),

Indonesia (Lumbsch 1993), and Papua New Guinea (Martin et al. 2003). The

lichen usually occurs in lowlands and only rarely appears at high elevations of

the intertropical region (Lumbsch 1993). Our study significantly extends its

distribution to Europe, where it is reported only from a single locality, Sant

Lloreng de la Muga (Catalonia, Spain). This areas warm humid Mediterranean

climate (with relatively high temperatures and humidity) has allowed the

establishment of a rich foliicolous lichen flora (Llop & Gémez-Bolea 2006),

predominantly Atlantic bryophyte species such as Leucobryum juniperoideum

(Brid.) C. Mull. (Brugués et al. 1974), and the pteridophyte Pellaea calomelanos

(Sw.) Link earlier known from South Africa, northern India, and Macaronesia

(Terradas & Brugués 1973). The existence of these Atlantic and paleotropical

elements in the study area would agree with the presence of Diploschistes

rampoddensis outside its known pantropical distribution.

The European D. rampoddensis samples are saxicolous and grow on

Garumnian red beds (HCI-). Pant & Upreti (1993), however, reported the

species colonizing sandstones from India.

SPECIMENS EXAMINED — CHINA. YUNNAN. Simao Distr. Mojiang Co., Mojiang (km-

post 350 from Kunming), along the main road, 23°26’N 101°41’E, 1200 m, on hard

soil, 12 Sep. 1987, Moberg & Santesson 7702 (UPS L-11539); along the road E of the

river Bavian Jiang (km-post 433 from Kunming), 23°16’N 101°16’E, 1100 m, on wayside

rocks, 13 Sep. 1987, Moberg & Santesson 31970 (UPS L-11540). SPAIN. CATALONIA.

Sant Lloreng de la Muga, path to Can Gener, UTM 31TDG8388, 150 m, Garumnian red

beds, 9 Feb. 2007, Llimona & Hladun (BCN-lich. 18011); cliffs by the river la Muga, near

an old mine, UTM 31TDG8188, 280 m, Garumnian red beds 45° S-SW, 2 Mar. 2007,

Llimona, Hladun & Mufiz (BCN-lich. 18008, GenBank KJ542543; BCN-lich. 18009).

Diploschistes rampoddensis in Europe ... 393

REMARKS —Diploschistes rampoddensis resembles octosporate specimens

of D. scruposus (Fic. 2B; D. scruposus “morphotypus interpediens’, hereafter

abbreviated to D. scruposus “interpediens”), which also have (6—)8 ascospores

per ascus and a similar ascospore size [(21-)23.5-27 x (9.5—)10-13(-13.5) um].

However, a detailed morphological comparison reveals that D. rampoddensis

has a thinner and more delicate thallus than D. scruposus “interpediens.” Their

chemistry also differs: D. rampoddensis contains only lecanoric acid (major),

while D. scruposus “interpediens” contains diploschistesic acid (minor) and

orsellinic acid (minor), in addition to lecanoric acid (major) (Barbero 1998).

Morphological and chemical differences between these two taxa are nonetheless

really subtle, and the utilization of the nrITS was crucial in confirming the

identity of the European specimens of D. rampoddensis.

Within Diploschistes, there are two other pantropically distributed species

(D. cinereocaesius and D. hypoleucus Zahlbr.), which resemble D. rampoddensis

in having a yellowish thallus, octosporous asci, and a rather similar ascospore

size. These two species, however, are terricolous, have bigger apothecia that

are usually secondarily subdivided (see D. cinereocaesius, Fic. 2C), and have

different chemistry: D. cinereocaesius contains lecanoric acid (major) and

diploschistesic acid (minor), while D. hypoleucus contains gyrophoric acid

as a major compound. That Diploschistes rampoddensis and D. cinereocaesius

appear as sister taxa in our phylogeny suggests a biogeographic pattern.

ADDITIONAL SPECIMENS EXAMINED — Diploschistes cinereocaesius. COLOMBIA.

UrRRAO. Paramo de Frontino, 6°25’N 76°05’W, 3500 m, 30 June 1985, Churchill, Sastre-

De Jesus & Escobar (DUKE 0144448). ECUADOR. IMBABURA. Laguna Cuicocha - NE

edge, ca 9km W of town Cotacachi, 00°18’43” N, 78°21’15” W, on soil at trail-cutting,

3300-3350 m, 8 Oct. 2000, Palice 4471 & Soldan (PRA, GenBank KJ542542) PERU.

HvuaRAz. Route to Laguna Churup, 9°29’S 77°26’W, 4070 m, 25 Aug. 2010, Rothfels

4000 & Zylinski (BCN-lich. 18010).

Diploschistes scruposus. SPAIN. CATALONIA. Girona, Sant Lloreng de la Muga, cliffs by

the river la Muga, near an old mine, UTM 31TDG8188, 280 m, Garumnian red beds

45° S-SW, 2 Mar. 2007, Llimona, Hladun & Muniz (Hb. Fdez.-Brime, SFB 59, GenBank

KJ542545; BCN-Lich 19326); cliffs by the river la Muga, UTM 31TDG8188, 240 m,

granitic outcrops 40° S-SW, 3 June 2009, Fernandez-Brime (Hb. Fdez.-Brime, SFB 103,

GenBank KJ542546).

D. scruposus “interpediens. FRANCE. PYRENEES ORIENTALES: Albera, road from

Maurellans to Rinoguers, 31TDH85351, 300 m, granitic outcrops near the road, 13

June 2009, Llimona (BCN-Lich 19322). SPAIN. CaTatonta. Barcelona, Orrius, turd

de Céllecs, 31T DG4400, 530 m, granitic outcrops, 23 Jan. 2009, Llimona & Fernandez-

Brime (BCN-Lich 19355). Girona, Sant Lloreng de la Muga, path to Can Gener, 31T

DG8387, 140-150 m, Garumnian red beds, 9 Feb. 2007, Llimona & Hladun (BCN-lich.

19319); cliffs by the river la Muga, UTM 31TDG8188, 260 m, granitic outcrops 60°

E, 3 June 2009, Fernandez-Brime (Hb. Fdez.-Brime, SFB 104, GenBank KJ542544). La

Rioja. Anguiano, electric power station, 30T WM1878, 670 m, Quercus ilex forest, 8

Sep. 2004, Hladun & Muniz (BCN-Lich 14404).

394 ... Fernandez-Brime & al.

Acknowledgments

The authors thank the herbaria of BCN, DUKE, PRA, UPS and Dr. Diana Muniz

and Dr. Carl Rothfels for providing material used in the present study, Dr. Thorsten

Lumbsch for generating sequences used in this study, Dr. Ana Rosa Burgaz and Dr

Yogesh Joshi for presubmission review, Molly McMullen and Dr. Robert Lachlan for the

English revision, and Dr. Shaun Pennycook for the nomenclature review. This work was

supported by the projects ‘Filogenia molecular de les Teloschistales i Ostropales. Part IT

and ‘Biodiversitat dels Fongs i Liquens dels Paisos Catalans’ (PIN2012-S02) both funded

by the Institut d’Estudis Catalans, with the contribution of the “Fons Memorial Salvador

Llimona:

Literature cited

Barbero MM. 1998. Estudio floristico y quimiotaxondmico de los liquenes silicicolas del Maresme.

Ph.D. thesis, University of Barcelona.

Brugués M, Casas de Puig C, Cros RM. 1974. Aportacion a la brioflora catalana. Leucobryum

juniperoideum (Brid.) C Mull en los alcornocales del Alto Ampurdan. Anales Inst. Bot.

Cavanilles 31: 109-117.

Felsenstein J. 1985. Confidence limits on phylogenies: an approach using the bootstrap. Evolution

39: 783-791. http://dx.doi.org/10.2307/2408678

Fernandez-Brime S, Llimona X, Lutzoni F, Gaya E. 2013. Phylogenetic study of Diploschistes

(Graphidaceae, Ostropales, lichen-forming Ascomycota), based on morphological, chemical,

and molecular data. Taxon 62: 267-280. http://dx.doi.org/10.12705/622.10

Gardes M, Bruns TD. 1993. ITS primers with enhanced specificity for basidiomycetes: application

to the identification of mycorrhizae and rusts. Mol. Ecol. 2: 113-118.

http://dx.doi.org/10.1111/j.1365-294X.1993.tb00005.x

Gaya E, Lutzoni F, Zoller S, Navarro-Rosinés P. 2003. Phylogenetic study of Fulgensia and allied

Caloplaca and Xanthoria species (Teloschistaceae, lichen-forming Ascomycota). Amer. J. Bot.

90: 1095-1103. http://dx.doi.org/10.3732/ajb.90.7.1095

Gaya E, Navarro-Rosinés P, Llimona X, Hladun N, Lutzoni F. 2008. Phylogenetic reassessment of

the Teloschistaceae (lichen forming Ascomycota, Lecanoromycetes). Mycol. Res. 112: 528-546.

http://dx.doi.org/10.1016/j.mycres.2007.11.005

Gaya E, Redelings BD, Navarro-Rosinés P, Llimona X, de Caceres M, Lutzoni F. 2011. Align or

not to align? Resolving species complexes within the Caloplaca saxicola group as a case study.

Mycologia 103: 361-378. http://dx.doi.org/10.3852/10-120

Gueidan C, Roux C, Lutzoni F. 2007. Using a multigene phylogenetic analysis to assess generic

delineation and character evolution in Verrucariaceae (Eurotiomycetes, Ascomycota). Mycol.

Res. 111: 1145-1168. http://dx.doi.org/10.1016/j.mycres.2007.08.010

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

10th edition. CAB International, Wallingford.

Lee SB, Milgroom MG, Taylor JW. 1988. A rapid, high yield mini-prep method for isolation of total

genomic DNA from fungi. Fungal Genet. Newslett. 35: 23-24.

Llop E, Gémez-Bolea A. 2006 Foliicolous lichens and associated lichenicolous fungi in the

northeastern Iberian Peninsula: the effect of environmental factors on distribution.

Lichenologist 38: 55-65. http://dx.doi.org/10.1017/S0024282905014362

Lumbsch HT. 1989. Die holarktischen Vertreter der Flechtengattung Diploschistes

(Thelotremataceae). J. Hattori Bot. Lab. 66: 133-196.

Lumbsch HT. 1993. Studien tiber die Flechtengattung Diploschistes 1. Nova Hedwigia 56: 227-236.

Diploschistes rampoddensis in Europe ... 395

Lutzoni F, Wagner P, Reeb V, Zoller S. 2000. Integrating ambiguously aligned regions of DNA

sequences in phylogenetic analyses without violating positional homology. Syst. Biol. 49:

628-651. http://dx.doi.org/10.1080/106351500750049743

Maddison WP, Maddison DR. 2011. Mesquite: a modular system for evolutionary analysis. Version

2.75. http://mesquiteproject.org.

Martin MP, Winka K. 2000: Alternative methods of extracting and amplifying DNA from lichens.

Lichenologist 32: 189-196.

Martin MP, LaGreca S, Lumbsch HT. 2003. Molecular phylogeny of Diploschistes inferred from ITS

sequence data. Lichenologist 35: 27-32.

Nylander W. 1900. Lichenes ceylonenses et additamentum ad lichenes japoniae. Acta Soc. Sci.

Fenn. 26: 1-33.

Orange A, James PW, White, FJ. 2001. Microchemical methods for the identification of lichens.

British Lichen Society, London.

Pant G, Upreti DK. 1993. The lichen genus Diploschistes in India and Nepal. Lichenologist 25:

33-50. http://dx.doi.org/10.1017/S0024282993000040

Pérez-Vargas I, Hernandez-Padrén C, Pérez De Paz PL, Elix JA. 2012. A new saxicolous species

of Diploschistes (Thelotremataceae) from the Canary Islands. Lichenologist 44: 67-71.

http://dx.doi.org/10.1017/S0024282911000612

Redelings BD, Suchard MA. 2007. Incorporating indel information into phylogeny estimation for

rapidly emerging pathogens. BMC Evol. Biol. 7: 40. http://dx.doi.org/10.1186/1471-2148-7-40

Schoch CLet al. 2012. Nuclear ribosomal internal transcribed spacer (ITS) region asa universal DNA

barcode marker for fungi. PNAS 109: 6241-6246. http://dx.doi.org/10.1073/pnas.1117018109

Suchard M, Redelings BD. 2006. BAli-Phy: simultaneous Bayesian inference of alignment and

phylogeny. Bioinformatics 22: 2047-2048. http://dx.doi.org/10.1093/bioinformatics/btl175

Swofford DL. 2002. PAUP*: Phylogenetic analysis using parsimony (* and other methods), version

4.0b10. Sunderland, Massachusetts: Sinauer.

Terradas JA, Brugués M. 1973. Una nueva localidad de Pellaea calomelanos en Catalufia. Acta

Geobot. Barcinon. 8: 1-15.

White TJ, Bruns T, Lee S, Taylor JW. 1990. Amplification and direct sequencing of fungal ribosomal

RNA genes for phylogenetics. 315-322 in: MA Innis et al. (eds). PCR protocols: a guide to

methods and applications. Academic Press, San Diego.

MY COTAXON

http://dx.doi.org/10.5248/129.397

Volume 129(2), pp. 397-401 October-December 2014

New species of Graphium and Periconia from China

YUE-MING Wu?”, JUN-JIE XU3, JIN-HuAa KonG‘, & TIAN-YU ZHANG?”

' Department of Plant Pathology, Shandong Agricultural University, Taian, 271018, China

2 Key Laboratory of Agricultural Microbiology, Shandong Province, Taian, 271018, China

* College of Life Sciences, Linyi University, Shandong Province, Linyi, 276005, China

* Linyi Productivity Promotion Center, Shandong Province, Linyi, 276001, China

*CORRESPONDENCE TO: tyzhang1937@yahoo.com.cn

ABSTRACT—Three new species from soil in China, Graphium variabile, G. wuweiense, and

Periconia guangdongensis, are described and illustrated. Specimens (dried cultures) and

living cultures are deposited in the Herbarium of Shandong Agricultural University, Plant

Pathology (HSAUP), and the Herbarium of Institute of Microbiology, Academia Sinica

(HMAS).

KEY worps — dematiaceous hyphomycetes, anamorphic fungi, taxonomy

Introduction

Graphium was established by Corda (1837) and is characterized by

possession of synnematous conidiophores with each synnema capped by a slimy

conidial head. The conidiogenous cells are monoblastic, percurrent, subulate or

cylindrical. Conidia are produced from annellides, often aggregated in slimy

heads, cylindrical rounded at the apex, cuneiform or ellipsoidal, usually with a

flat base, and 0-septate. Although Index Fungorum (2014) lists 173 taxa (many

infraspecific), Seifert et al. (2011) estimates that the genus may contain only 19

authentic species.

Periconia was established by Tode (1791) and is characterized by

macronematous, mononematous conidiophores. The conidiogenous cells

are monoblastic, polyblastic, branched, and ellipsoidal. Conidia are catenate,

spherical or subspherical, verruculose or echinulate, 0-septate and aggregated

in heads. Seifert et al. (2011) estimates that the genus may contain 40 authentic

species.

During a survey of soil dematiaceous hyphomycetes in China, several

unusual species of Graphium and Periconia were collected. Two Graphium and

one Periconia species are described and illustrated as new.

AY.M. Wu and J.J. Xu contributed equally to this work.

398 ... Wu, Xu, &al.

Fia. 1. Graphium variabile (ex holotype HSAUP II ,,2789).

Conidia, conidiophores, conidiogenous cells, and synnemata. Scale bars = 25 um.

Graphium variabile J.J. Xu & T.Y. Zhang, sp. nov. FIG. 1

MycoBank MB 807387

Differs from Graphium eumorphum by its conidia that are smaller and contain no oil

droplets and from G. terricola by its larger conidia.

Type: China, Fujian Province, Xiamen National Forest Park, from a forest soil, 6 Oct.

2004, J.J. Xu (Holotype HSAUP II ,,2789; isotype HMAS 196278).

Erymo.oey: The epithet refers to the variably shaped conidia.

CoLonigs on CMA (Matsushima 1995) slow growing, effuse, felted, white at

initial stage, then darkish brown to black. Myce.ium partly superficial, partly

immersed in the substratum, hyphae branched, septate, smooth, subhyaline,

2-4 um wide. SYNNEMATA erect, scattered, rough, brown at base paler towards

the apex, 150-320 um long, 15-30 um wide at base, slightly splayed out at the

apex. CONIDIOGENOUS CELLS annellides, subhyaline, subulate or cylindrical,

integrated, terminal, smooth, 15-25 x 1.5-2.5 um. CONIDIA acrogenous,

subhyaline to hyaline, forming conidial chains and enveloped in mucilaginous

secretion, ellipsoidal, pyriform, obovoid to globose, smooth, 0-septate, globose

conidia 3-6.5 um in diameter, others 5.5-7 x 3.75-5 um, wide in the broadest

part, slightly truncate at the ends.

Comments: Morphologically, G. variabile resembles G. eumorphum (Sacc.)

Sacc. (Saccardo 1886) and G. terricola Manohar. et al. (Manoharachary et al.

1975). Graphium eumorphum is distinguished by longer conidia (6-8 x 3.5-4.5

Graphium & Periconia spp. nov. (China) ... 399

um, 2-guttulate) that contain oil droplets, while G. terricola produces smaller

conidia (4-6.5 x 2-3.2 um).

N {

anys P96 |

CY A

TSN (

Fic. 2. Graphium wuweiense (ex holotype HSAUP II ,.2647).

Conidia, conidiophores, conidiogenous cells, and synnemata. Scale bars = 25 um.

Graphium wuweiense J.H. Kong & T.Y. Zhang, sp. nov. FIG. 2

MycoBank MB 807385

Differs from Graphium adansoniae and G. penicillioides by its shorter conidiophores.

Type: China, Gansu Province, Wuwei, from a desert oasis soil, 18 Apr. 2005, J.H. Kong

(Holotype HSAUP II ,.2647; isotype HMAS 196279).

EryMo_oey: in reference to the type locality.

CoLoniges on CMA (Matsushima 1995) slow growing, effuse, white at

initial stage, then darkish brown, felted. MyceLtrum mostly immersed in the

substratum, hyphae branched, septate, smooth, subhyaline, 1.5-3 um wide.

SYNNEMATA erect, scattered, smooth, brown at base, paler towards the apex,

400 ... Wu, Xu, & al.

20-40 um long, 10-20 um wide at base, slightly splayed out at the apex.

CONIDIOGENOUS CELLS annellides, hyaline, subulate or cylindrical, integrated,

terminal, smooth, 7-18 x 1.5-2.5 um. Conip1a acrogenous, hyaline, forming

conidial chains enveloped in mucilaginous secretion, ellipsoidal, pyriform to

obovoid, smooth, 0-septate, truncate at the base, 4-6 um long, 3-5 um wide.

ComMENTs: Morphologically, Graphium wuweiense is most similar to

G. adansoniae Cruywagen et al. (Cruywagen et al. 2010) and G. penicillioides

Corda (Corda 1837). However, both of these species have much longer

conidiophores (G. adansoniae, 88-150 um; G. penicillioides, up to 200 um).

(

)

19-@)

C) CA Ys bs

Fia. 3. Periconia guangdongensis (ex holotype HSAUP II ,,3133).

Conidia, conidiophores, and conidiogenous cells. Scale bars = 25 um.

Periconia guangdongensis J.J. Xu & T.Y. Zhang, sp. nov. FIG. 3

MycoBank MB 807386

Differs from Periconia cookei and P. pseudobyssoides by its shorter globose conidiophores

and smaller conidia.

Type: China, Guangdong Province, Meizhou, from a grassland soil, 10 Aug. 2004, J.J. Xu

(Holotype HSAUP II ,,3133; isotype HMAS 196280).

EryMo_oey: in reference to the type locality.

Cotoniges on MEA (Lefebvre et al. 1949) effuse, initially white, then grey

to greyish brown, cottony or fine hairy. MycELtum superficial or immersed,

Graphium & Periconia spp. nov. (China) ... 401

hyphae branched, septate, sometimes rough, subhyaline, 1.5-4 um wide.

CONIDIOPHORES macronematous, mononematous, straight or slightly

flexuous, smooth although often verrucose at base, pale brown to mid-brown,

unbranched, 3-4-septate, 230-480 um long, 7-9 um wide at base, often swollen

at apex. CONIDIOGENOUS CELLS globose, directly arising from the swollen apex,

pale brown to dark brown, 5-9 um diam. Conip1A globose to subglobose,

0-septate, smooth initially but echinulate at maturity, brown to dark brown,

7-11 um diam.

ComMEnts: Morphologically, Periconia guangdongensis slightly resembles

P. cookei E.W. Mason & M.B. Ellis (Mason & Ellis 1953) and P. pseudobyssoides

Markovsk. & Kacergius (Markovskaja & Kacergius 2014). However, P. cookei

has larger non-globose conidia (13 x 16 um) and shorter conidiophores

(88-150 um), and P. pseudobyssoides has larger non-globose conidia (15 x 17

um) with conidiophores up to 600 um.

Acknowledgments

The authors are grateful for pre-submission comments and suggestions provided

by Dr. Eric McKenzie, Dr. Yong Wang, and Dr. Shaun Pennycook. This project was

supported by the National Science Foundations of China (no. 30970011 & 30499340).

Literature cited

Corda ACJ. 1837. Icones fungorum hucusque cognitorum 1. 32 p.

Cruywagen EM, de Beer ZW, Roux J, Wingfield MJ. 2010. Three new Graphium

species from baobab trees in South Africa and Madagascar. Persoonia 25: 61-71.

http://dx.doi.org/10.3767/003158510X550368

Index Fungorum. 2014. http://www.indexfungorum.org/Names/Names.asp (accessed 22 May.

2014).

Lefebvre CL, Johnson AG, Sherwin HS. 1949. An undescribed species of Periconia. Mycologia 41:

416-419. http://dx.doi.org/10.2307/3755235

Manoharachary C, Rao PR, Rehana AR, Ramarao P. 1975. Notes on microfungi from Andhra

Pradesh - I: a new species of Graphium from soil. Nova Hedwigia 26(2-3): 473-476.

Markovskaja S, Kacergius A. 2014. Morphological and molecular characterisation of Periconia

pseudobyssoides sp. nov. and closely related P byssoides. Mycological Progress 13: 291-302.

http://dx.doi.org/10.1007/s11557-013-0914-6

Mason EW, Ellis MB. 1953. British species of Periconia. Mycological Papers 56. 127 p.

Matsushima T. 1995. Matsushima mycological memoirs, no. 8.

Saccardo PA. 1886. Pheostibeae, Amerosporae, Graphium. Sylloge Fungorum 4: 609-619.

Seifert K, Morgan-Jones G, Gams W, Kendrick B. 2011. The genera of hyphomycetes. CBS

Biodiversity Series 9. 997 p. http://dx.doi.org/10.3767/003158511X617435

Tode HJ. 1791. Fungi mecklenburgenses selecti 2. 66 p.

MY COTAXON

http://dx.doi.org/10.5248/129.403

Volume 129(2), pp. 403-406 October-December 2014

Two new species of Myrothecium

from the Qinghai-Tibet Plateau Area, China

YUE-MING Wu?”’, Yu-LAN JIANG3’, YA-NAN MaA?? & TIAN- YU ZHANG?”

' Department of Plant Pathology, Shandong Agricultural University, Taian, 271018, China

? Key Laboratory of Agricultural Microbiology, Shandong Province, Taian, 271018, China

* Agriculture College, Guizhou University, Guiyang, Guizhou, 550025, China

*CORRESPONDENCE TO: tyzhang1937@yahoo.com.cn

ABSTRACT — Two new species from the Qinghai-Tibet Plateau Area (China), Myrothecium

variabile and M. xigazense, are described and illustrated. Specimens (dried cultures) and

living cultures are deposited in the Herbarium of Shandong Agricultural University, Plant

Pathology (HSAUP), and the Herbarium of Institute of Microbiology, Academia Sinica

(HMAS).

KEY worps — Ascomycota, dematiaceous hyphomycetes, soil fungi, Stachybotriaceae,

taxonomy

Introduction

During a survey of dematiaceous soil hyphomycetes in China, several

unusual Myrothecium specimens were collected, of which two represent new

species. They are described and illustrated from cultures grown on potato

dextrose agar (PDA) (Domsch et al. 2007).

Myrothecium Tode [type species: M. inundatum Tode (Tode 1790)] is

characterized by conspicuous sporodochia that are cupulate, pustulate or

stalked, synnema-like, formed of densely compacted conidiophores arising

from a developed stroma and bearing a mass of slimy, green to black conidia.

Sporodochia are surrounded by differentiated, often curling, marginal hairs.

Conidiophores are hyaline or slightly olivaceous, and branch repeatedly to the

ultimate branches, the phialides. Phialides are compacted in a dense parallel

layer, cylindrical with conically tapering tips and usually undifferentiated

collarettes. Conidia are 1-celled, subhyaline to pale greenish brown but dark

olivaceous in mass. Seifert et al. (2011) listed 30 taxa for the genus.

AYue-Ming Wu and Yu-Lan Jiang contributed equally to this work.

404 ... Wu, Jiang, & al.

Fra. 1. Myrothecium variabile (ex holotype HSAUP II 1083).

Conidia, conidiophores, and conidiogenous cells. Scale bars = 25 um.

Myrothecium variabile Y.M. Wu & T.Y. Zhang, sp. nov. FIG. 1

MycoBank MB 808618

Differs from Myrothecium cinctum and M. striatisporum by its longer conidia.

Type: China, Tibet: Xigaze, altitude 3100 m, from a forest soil, 11 Sept. 2007, Y.M. Wu

(Holotype HSAUP II 1083; isotype HMAS 196284).

Erymo ocy: The epithet refers to the variable shape of the conidia.

CoLonigs on PDA effuse, flocculent, pale yellow at initial stage, later green to

black. Mycelium superficial or immersed, hyphae branched, septate, smooth,

hyaline, 2-3 um wide. SPORODOCHIA usually not more than 1.0 mm diameter,

often confluent, at first green, later black with a white margin, without setae.

CONIDIOPHORES Straight or flexuous, branched, smooth, with apical branches

arranged penicillately. CoNIDIOGENOUS CELLS phialides colourless, smooth,

10-25 x 2-2.5 um. Conrpia unicellular, cylindrical, clavate, fusiform to

navicular (L/W=6-11.25), often slightly protuberant and truncate at the base,

smooth, pale greenish brown, in mass greenish brown to black, with distinct

longitudinal or oblique striations, 12.5-16.25 x 2.5-3.5 um.

Myrothecium spp. nov. (China) ... 405

Comments: Morphologically, Myrothecium variabile resembles M. cinctum

(Corda) Sacc. (Saccardo 1886) and M. striatisporum N.C. Preston (Preston

1948) its longitudinally or obliquely striate conidia. However, M. cinctum

(6.5-14 x 2.5-4.5 um) and M. striatisporum (7-12 x 2.5-3.5 um) have smaller

conidia and verrucose conidiophores.

Fia. 2. Myrothecium xigazense (ex holotype HSAUP II ,.0969).

Conidia, conidiophores, setae, and conidiogenous cells. Scale bars = 25 um.

Myrothecium xigazense Y.M. Wu & T.Y. Zhang, sp. nov. FIG.i2

MycoBank MB 808619

Differs from Myrothecium carmichaelii and M. jollymanii by its longer conidia.

TyPE: China, Tibet: Xigaze, altitude 3900 m, from a grassland soil, 9 Sept. 2007, Y.M. Wu

(Holotype HSAUP II 0969; isotype HMAS 196285).

EryMo_oey: in reference to the type locality.

406 ... Wu, Jiang, & al.

Co.ontEs on PDA effuse, flocculent, white at initial stage, later black, Mycelium

mostly superficial, hyphae branched, septate, smooth, hyaline, 2-3 um wide.

SPORODOCHIA usually not more than 1.2 mm diameter, often confluent,

at first green, later black with a white margin. SETAE pale brown, smooth,

septate, 220-250 um long, 4-6 um thick. CoNIDIOPHORES straight or flexuous,

branched, with apical branches arranged penicillately. CONIDIOGENOUS CELLS

phialides colourless, smooth, 8-15x2-2.5 um. ConrpiA unicellular, cylindrical

or clavate, often slightly protuberant and truncate at the base, smooth, pale

greenish brown, dark greenish in mass, 12.5-15 x 2-3 um.

Comments: Morphologically, Myrothecium xigazense resembles M. carmichaelii

Grev. (Greville 1825) and M. jollymanii N.C. Preston (Preston 1948) in

conidial colour and shape. However, both M. carmichaelii (10-11 x 1-1.3

um) and M. jollymanii (10-12 x 2-2.5 um) have shorter conidia. Additionally,

M. carmichaelii lacks setae while the setae in M. jollymanii are hyaline, not pale

brown as in M. xigazense.

Acknowledgments

The authors are grateful for pre-submission comments and suggestions provided

by Dr. Eric McKenzie, Dr. Yong Wang, and Dr. Shaun Pennycook. This project was

supported by the National Science Foundations of China (no. 30970011 & 30499340).

Literature cited

Domsch KH, Gams W, Anderson TH. 2007. Compendium of soil fungi, ed. 2. IHW-Verlag Eching.

672 p. http://dx.doi.org/10.1111/j.1365-2389.2008.01052_1.x

Greville RK. 1825. Scottish Cryptogamic Flora 3: pl. 121-180.

Preston NC. 1948. Observations on the genus Myrothecium. Il. Myrothecium gramineum Lib.

and two new species. Transactions of the British Mycological Society 31(3-4): 271-276.

http://dx.doi.org/10.1016/S0007-1536(48)80010-0

Saccardo PA. 1886. Tuberculariea dematiea, Amerospora, Myrothecium. Sylloge Fungorum 4:

750-752.

Seifert K, Morgan-Jones G, Gams W, Kendrick B. 2011. The genera of hyphomycetes. CBS

Biodiversity Series 9. 997 p. http://dx.doi.org/10.3767/003158511X617435

Tode HJ. 1790. Fungi mecklenburgensis selecti. 1. 47 p.

MY COTAXON

http://dx.doi.org/10.5248/129.407

Volume 129(2), pp. 407-413 October-December 2014

Nomenclatural novelties in the Postia caesia complex

VIKTOR PAPP

Department of Botany, Corvinus University of Budapest,

H-1118 Budapest, 44 Ménesi st, Hungary

CORRESPONDENCE TO: viktor.papp@uni-corvinus.hu

ABSTRACT - Within the genus Postia, the P. caesia complex forms a distinctive morphological

group. Based on recent molecular data, the current taxonomic status of the P. caesia complex

is discussed and the nomenclature of the related taxa is revised as well. New combinations

are: Postia subg. Cyanosporus, Postia africana, Postia amyloidea, Postia caesioflava, and Postia

coeruleivirens.

Keyworps - polypores, basidiomycetes, Oligoporus

Introduction

Based on recent molecular phylogenetic studies, the cosmopolitan polypore

genus Postia Fr. belongs to the antrodia clade, with members characterized by

brown-rot wood decay (Hibbett & Donoghue 2001, Binder et al. 2013). Within

the genus, the Postia caesia complex forms a distinctive morphological group

(Tura et al. 2008). The foremost species of this complex is P caesia (Schrad.)

P. Karst., which was described from Germany (Schrader 1794) but probably

has a circumglobal distribution (e.g., Pildain & Rajchenberg 2012, Ryvarden

& Gilbertson 1994). The main morphological features of P caesia are annual

soft white basidiocarps that become blue-grey when bruised or spontaneously

in age and small cylindrical to allantoid cyanophilous basidiospores (Niemela

2013). In addition to P caesia are similar taxa described from Europe (David

1974, 1980; Niemela et al. 2001; Pieri & Rivoire 2005) and worldwide (Corner

1989). Species delimitation in this difficult complex has not been sufficiently

clarified (Yao et al. 2005), and the generic name of this peculiar polypore group

has changed more than once, depending on the various taxonomic concepts

held in the past. The current taxonomic status of the P caesia complex is

discussed based on recent molecular data and the nomenclature of the related

taxa is revised accordingly.

408 ... Papp

Nomenclature and taxonomy of Boletus caesius

McGinty (1909) proposed a new monotypic genus (Cyanosporus) for

Polyporus caesius (Schrad.) Fr., based on its cyanophilous spores. However, the

name Cyanosporus caesius (Schrad.) McGinty was not accepted in significant

studies (e.g., Donk 1960, Jahn 1963, Lowe 1975), while the combination

introduced earlier by Murrill (1907), Tyromyces caesius, was commonly used.

After it became more evident that the species in Tyromyces P. Karst. were white-

rot-producing fungi, David (1980) placed the brown-rot taxa into Spongiporus

Murrill, Julich (1982) combined them into Postia, and Gilbertson & Ryvarden

(1985) transferred them into Oligoporus Bref. Depending on the biological,

morphological and anatomical features of the type species, these genera

also have been regarded as one genus. Postia (Fries 1874) has priority over

Oligoporus (Brefeld 1888) on the basis of wide acceptance of the validity of

Fries’s 1874 publication (Karsten 1879, 1881; Donk 1960, Jiilich 1982, Larsen

& Lombard 1986, Renvall 1992, Walker 1996, Pieri & Rivoire 1998), although

some authors (Ryvarden 1991, Ryvarden & Gilbertson 1994) have interpreted

“Postia Fr? as an invalid provisional name. The phylogenetic study by Yao et

al. (1999) also confirmed that there is no difference between Oligoporus and

Postia. Nonetheless, a subsequent phylogenetic study cited by Kotiranta et al.

(2009) supported the two genera as independent, with Oligoporus containing

species with thick-walled and distinctly cyanophilous spores. In contrast and

more recently, rDNA ITS and LSU sequence analyses by Pildain & Rajchenberg

(2012) place Postia s. lat. species in the ‘postia clade’ regardless of the spore

wall thickness. They regard the main features of the ‘postia clade’ as tetrapolar

mating, normal nuclear behavior, metachromatic generative hyphae, and the

absence of fiber hyphae in culture. In their phylogenetic study using LSU,

ITS, and combined LSU-ITS datasets, Ortiz-Santana et al. (2013) showed

that the Postia group falls into two main clades (core postia clade, sarcoporia

clade) with the ‘core postia clade’ comprising four subclades: /spongiporus,

/oligoporus, /postia sensu stricto, and /spongiporus-undosus). They accepted

Oligoporus, Postia, and Spongiporus as separate genera and also confirmed the

independence of Gilbertsonia, Rhodonia, Ryvardenia, and Taiwanofungus.

Currently, the validity of Postia Fr. is generally accepted and the genus

name is widely used (e.g., Buchanan & Ryvarden 2000, Wei & Dai 2006, Dai &

Hattori 2007, Dai et al. 2009, Hattori et al. 2010, Yuan et al. 2010, Cui & Li 2012,

Shen et al. 2014). The most recent molecular studies (Pildain & Rajchenberg

2012, Ortiz-Santana et al. 2013) place Postia caesia in Postia in the restricted

sense while within Postia the P. caesia complex forms a separate branch. The

phylogenies do not support the P. caesia group as a separate genus (as suggested

by McGinty in 1909) but do support the group as a subgenus within Postia.

Therefore the following combination is proposed:

Nomenclature of the Postia caesia complex ... 409

Postia subg. Cyanosporus (McGinty) V. Papp, comb. & stat. nov.

MycoBank MB810903

= Cyanosporus McGinty, in Lloyd, Mycol. Notes 33: 436 (1909)

Type: Boletus caesius Schrad., Spic. Fl. Germ.: 167 (1794) [= Postia caesia (Schrad.) P.

Karst. ]

Basidiocarp medium- to small-sized, whitish when young, becoming more or

less grey to blue tinted. Basidiospores narrow, small, faintly amyloid in Melzer’s

reagent. No chlamydospores in culture.

Species in the Postia caesia complex

Within the P. caesia complex, five species (P. alni, P. caesia, P. luteocaesia,

P. mediterraneocaesia, P. subcaesia) are accepted in Europe (Bernicchia 2005,

Niemela et al. 2001, Pieri & Rivoire 2005, Ryvarden & Gilbertson 1994).

However, other uncertain taxa described from Europe also probably belong to

the P caesia complex and are considered synonyms of P. caesia (e.g., Bjerkandera

ciliatula, Boletus candidus, B. coeruleus, Polyporus caesiocoloratus).

Based on the original description (Britzelmayr 1893), Polyporus

caesiocoloratus Britzelm. grew on a Picea trunk and had spores 0.7-1 um wide.

Apart from spore size, no other feature was mentioned that would distinguish

the taxon from P. caesia, suggesting that the two species are identical.

Boletus candidus Roth is also a potential synonym of P. caesia, although in

the original description Roth (1797) wrote that the pileus is white-yellow, with

white margin and has a brown-blue pore surface.

Postia luteocaesia (A. David) Julich is a rare Central European species,

which grows exclusively on Pinus; its main morphological feature besides the

typical blue greyish discoloration is the additional bright yellow colour of the

basidiocarp (Niemela et al. 2004, Ryvarden & Gilbertson 1994).

David (1974), who described Tyromyces subcaesius [= Postia subcaesia

(A. David) Julich] from deciduous trees, proved by interfertility tests that it

differs genetically from P caesia. With the exception its host preference, the

only features separating P. subcaesia from P. caesia are the narrower spores and

typically more whitish basidiocarp (Ryvarden & Gilbertson 1994).

Boletus coeruleus Schumach. (Schumacher 1803) is morphologically close

to Postia caesia but grows on Quercus. Fries (1821) regarded this species as a

variety of P. caesia, while other mycologists (e.g., Persoon 1825, Quélet 1886)

considered it a distinct species. However, Bernicchia et al. (2008) reported

P. caesia also from Quercus cerris, Q. ilex, and Q. pubescens, suggesting that the

host is not sufficient to characterize a species.

Bjerkandera ciliatula P. Karst., described from Finland, is mentioned among

the synonyms of P. caesia (Robert et al. 2014). However, Karsten (1887) described

the type basidiocarp as small (1-1.5 cm) and growing on Alnus incana. These

410... Papp

characteristics, combined with a basidiospore width of 1 um, suggest a closer

relationship with P alni than with P caesia. Postia mediterraneocaesia M. Pieri &

B. Rivoire also has a small (<25 mm long) basidiocarp, but its pileipellis hyphae

are encrusted and the basidiospores are wider (1.45-1.68 tum). It seems to be a

southern European and Mediterranean species that grows on both conifers and

hardwoods (Pieri & Rivoire 2005).

Tyromyces caesiosimulans G.F. Atk. was described from North America from

an unknown substratum. Although Atkinson (1908) wrote that the new species

resembled Polyporus caesius [= Postia caesia], it differs by having globose,

pedicellate spores. This basidiospore shape is not known in the complex, and a

type study is necessary to clarify the taxonomic position of this species.

Patouillard & Lagerheim (1892) described Polyporus caesioflavus from

Ecuador. This species is morphologically very similar to Postia caesia and

P. subcaesia (Ryvarden 1983). Polyporus caesioflavus is distinguished by

narrower (7-9/mm) pores and a glabrous pileus surface (Loguercio-Leite et

al. 2008). Carranza (1982) transferred this species to Tyromyces after studying

the type specimen and collecting specimens from Costa Rica. Carranza’s

combination, which proved to be invalid (ICN, Art 41.4), was subsequently

validated by Ryvarden (1983). Loguercio-Leite et al. (2008) later transferred

Tyromyces caesioflavus, a brown-rot species (Carranza 1982), to Oligoporus, as

O. caesioflavus (Pat.) Baltazar et al.

Corner (1989) described two Tyromyces species (T: amyloideus,

T. coeruleivirens) from Malaysia (Mt. Kinabalu, Borneo), which were studied

by Hattori (2002). Based on its olivaceous colour and allantoid (3-4.2 x 0.8-1

um) slightly amyloid spores., Hattori (2002) considered T. amyloideus only a

form of P. caesia s. lat., later accepting it as a synonym of P. caesia (Hattori et al.

2012). However, P. caesia s. str. has wider spores (1.3-1.7(-1.9) um; Ryvarden &

Gilbertson 1994, Niemela 2013), suggesting that T: amyloideus may represent a

distinct species. Tyromyces coeruleivirens also has a greenish pileus, monomitic

hyphal system, and allantoid inamyloid basidiospores (Corner 1989, Hattori

2002). Morphologically similar to T: amyloideus, T. coeruleivirens has narrower

pores (7-9/mm vs. 4-5/mm in T. amyloideus) and inamyloid, slightly longer

(4-5 x 0.8-1 um) basidiospores. Otherwise most characteristics (e.g., locality)

are highly similar, and a taxonomic revision of these two species is needed.

Ryvarden (1988), who described Oligoporus africanus from Africa

(Muramyya, Burundi), considered it as related to the O. caesius [= Postia caesia]

group based on the basidiocarp type, hyphal system, and allantoid slightly

amyloid spores. The main character that distinguishes O. africanus from

P. caesia, P. subcaesia, and P. luteocaesia is the smaller spore size (3.5-4.5 x

1-1.2 um) (Ryvarden 1988).

Nomenclature of the Postia caesia complex ... 411

Despite the fact that Oligoporus africanus, Polyporus caesioflavus, Tyromyces

amyloideus, and T. coeruleivirens have been accepted as closely related to the

Postia caesia group (‘core postia clade’), they have never been placed into Postia.

I therefore propose the following combinations:

Postia africana (Ryvarden) V. Papp, comb. nov.

MycoBank MB810904

= Oligoporus africanus Ryvarden, Mycotaxon 31(2): 407 (1988)

Postia amyloidea (Corner) V. Papp, comb. nov.

MycoBank MB810905

= Tyromyces amyloideus Corner, Beih. Nova Hedwigia 96: 160 (1989)

Postia caesioflava (Pat.) V. Papp, comb. nov.

MycoBank MB810907

= Polyporus caesioflavus Pat., Bull. Soc. mycol. Fr. 8(3): 114 (1892)

Postia coeruleivirens (Corner) V. Papp, comb. nov.

MycoBank MB810908

= Tyromyces coeruleivirens Corner, Beih. Nova Hedwigia 96: 163 (1989)

Acknowledgements

I sincerely thank Prof. Dr. Solomon P. Wasser (Haifa, Israel) and Dr. Ivan Zmitrovich

(Saint Petersburg, Russia) for pre-submission review. I am also grateful to Editor-in-

Chief Dr. Lorelei L. Norvell (Portland, USA) and Nomenclature Editor Dr. Shaun

Pennycook (Auckland, New Zealand) for the helpful remarks and suggestions. I wish

to thank Anna Szabé and Balint Dima for their valuable comments on the manuscript.

Literature cited

Atkinson GF. 1908. Notes on some new species of fungi from the United States. Annales Mycologici

6: 54-62.

Bernicchia A. 2005. Polyporaceae s. |. Fungi Europaei 10. Edizioni Candusso, Alassio.

Bernicchia A, Benni A, Venturella G, Gargano ML, Saitta A, Gorjon SP. 2008. Aphyllophoraceous

wood-inhabiting fungi on Quercus spp. in Italy. Mycotaxon 104: 425-428.

Binder M, Justo A, Riley R, Salamov A, Lopez-Giraldez FE, Sjékvist E, Copeland A, Foster

B, Sun H, Larsson E, Larsson K-H, Townsend J, Grigoriev IV, Hibbett DS. 2013.

Phylogenetic and phylogenomic overview of the Polyporales. Mycologia 105(6): 1350-1373.

http://dx.doi.org/10.3852/13-003

Brefeld JO. 1888. Untersuchungen aus dem Gesammtgebiete der Mykologie, vol. 8. 305 p.

Britzelmayr M. 1893. Materialien zur Beschreibung der Hymenomyceten 3. Botanisches

Centralblatt 54(4): 97-105.

Buchanan PK, Ryvarden L. 2000. An annotated checklist of polypore and polypore-like fungi

recorded from New Zealand. New Zealand Journal of Botany 38(2): 265-323.

Carranza J. 1982. Polypores new to Costa Rica. Mycotaxon 15: 405-408.

Corner EJH. 1989. Ad Polyporaceas V. Beihefte zur Nova Hedwigia 96: 1-218.

412 ... Papp

Cui BK, Li HJ. 2012. A new species of Postia (Basidiomycota) from Northeast China. Mycotaxon

120: 231-237. http://dx.doi.org/10.5248/120.231

Dai Y-C, Hattori T. 2007. Postia japonica (Basidiomycota), a new polypore from Japan. Mycotaxon

102: 113-118.

Dai Y-C, Yuan HS, Wang HC, Yang KE Wei YL. 2009. Polypores (Basidiomycota) from

Qin Mts. in Shaanxi Province, central China. Annales Botanica Fennica 46(1): 54-61.

http://dx.doi.org/10.5735/085.046.0105

David A. 1974. Une nouvelle espéce de Polyporaceae: Tyromyces subcaesius. Bulletin Mensuel de la

Société Linnéenne de Lyon 43: 119-126.

David A. 1980. Etude du genre Tyromyces sensu lato: répartition dans les genres Leptoporus,

Spongiporus et Tyromyces sensu stricto. Bulletin Mensuel de la Société Linnéenne de Lyon

49(1): 6-56.

Donk MA. 1960. The generic names proposed for Hymenomycetes - X. The generic names

proposed for Polyporaceae. Persoonia 1: 173-302.

Fries EM. 1821. Systema mycologicum, vol. 1. Gryphiswald.

Fries EM. 1874. Hymenomycetes europaei. Upsaliae, Typis descripsit ed. Berling.

Gilbertson RL, Ryvarden L. 1985. Some new combinations in the Polyporaceae. Mycotaxon 22:

363-365.

Hattori T. 2002. Type studies of the polypores described by E.J.H. Corner from Asia and

West Pacific Areas. IV. Species described in Tyromyces (1). Mycoscience 43: 307-315.

http://dx.doi.org/10.1007/s102670200045

Hattori T, Sotome K, Ota Y, Thi BK, Lee SS, Salleh B. 2010. Postia stellifera sp. nov., a stipitate and

terrestrial polypore from Malaysia. Mycotaxon 114: 151-161. http://dx.doi.org/10.5248/114.151

Hattori T, Yamashita S, Lee SS. 2012. Diversity and conservation of wood-inhabiting polypores

and other aphyllophoraceous fungi in Malaysia. Biodiversity Conservation 21(9): 2375-2396.

http://dx.doi.org/10.1007/s10531-012-0238-x

Hibbett DS, Donoghue MJ. 2001. Analysis of character correlations among wood decay mechanisms,

mating systems, and substrate ranges in homobasidiomycetes. Systematic Biology 50: 215-242.

Jahn H. 1963. Mitteleuropaische Porlinge (Polyporaceae s. lato) und ihr Vorkommen in Westfalen.

Westfalische Pilzbriefe 4: 1-143.

Julich W. 1982. Notes on some Basidiomycetes (Aphyllophorales and Heterobasidiomycetes).

Persoonia 11(4): 421-428.

Karsten PA. 1879. Symbolae ad mycologiam fennicam. VI. Meddelanden af Societas pro Fauna et

Flora Fennica 5: 15-46.

Karsten PA. 1881. Enumeratio boletinearum et polyporearum fennicarum, systemate novo

dispositarum. Revue Mycologique 3(9): 16-19.

Karsten PA. 1887. Symbolae ad mycologiam fennicam. XVIII. Meddelanden af Societas pro Fauna

et Flora Fennica 14: 78-84.

Kotiranta H, Saarenoksa R, Kytévuori I. 2009. Aphyllophoroid fungi of Finland - A check-list with

ecology, distribution, and threat categories. Norrlinia 19: 1-223.

Larsen MJ, Lombard FF. 1986. New combinations in the genus Postia Fr. (Polyporaceae). Mycotaxon

26: 27 122735

Loguercio-Leite C, Michels J, Baltazar JM. 2008. Austro-American lignolytic polypores

(Agaricomycetes) — new records for Southern Brazil. Mycotaxon 104: 205-213.

Lowe JL. 1975. Polyporaceae of North America. The genus Tyromyces. Mycotaxon 2(1): 1-82.

McGinty NJ. 1909. A new genus, Cyanosporus. 436, in: CG Lloyd. Mycological Notes 33.

Murrill WA. 1907. Polyporaceae, part 1. North American Flora 9(1): 1-72.

Niemela T. 2013. Polypores of the Bialowieza Forest. Bialowieza.

Nomenclature of the Postia caesia complex ... 413

Niemela T, Kinnunen J, Lindgren M, Manninen O, Miettinen O, Penttila E, Turunen O. 2001.

Novelties and records of poroid Basidiomycetes in Finland and adjacent Russia. Karstenia 41(1):

1-21.

Niemela T, Dai Y-C, Kinnunen J, Schigel DS. 2004. New and in North Europe rare polypore species

(Basidiomycota) with annual, monomitic basidiocarps. Karstenia 44(12): 67-77.

Ortiz-Santana B, Lindner DL, Miettinen O, Justo A, Hibbett DS. 2013. A phylogenetic

overview of the antrodia clade (Basidiomycota, Polyporales). Mycologia 105(6): 1391-1411.

http://dx.doi.org/10.3852/13-051

Patouillard NT, Lagerheim G de. 1892. Champignons de |’Equateur (Pugillus II). Bulletin de la

Société Mycologique de France 8(2): 113-140.

Persoon CH. 1825. Mycologia europaea, vol. 2. Erlanga.

Pieri M, Rivoire B. 1998. Postia inocybe (David Malengon) Julich f. inocybe et f. pileatus, f. nov.

Notes nomenclaturales sur le genre Postia. Bulletin de la Société Mycologique de France 114(3):

19-34.

Pieri M, Rivoire B. 2005. Postia mediterraneocaesia, une nouvelle espece de polypore découverte dans

le sud de Europe. Bulletin de la Fédération des Associations Mycologiques Méditerranéennes

28: 33-38.

Pildain MB, Rajchenberg M. 2012. ‘The phylogenetic disposition of Postia s.l.

(Polyporales, Basidiomycota) from Patagonia, Argentina. Mycologia 105(2): 357-367.

http://dx.doi.org/10.3852/12-088

Renvall P. 1992. Basidiomycetes at the timberline in Lapland 4. Postia lateritia n. sp. and its rust-

coloured relatives. Karstenia 32: 43-60.

Robert V, Stegehuis G, Stalpers J. 2014. The MycoBank engine and related databases.

http://www.mycobank.org [accessed April 2014].

Roth AW. 1797. Catalecta botanica, vol. 1. Bibliopolo I.G. Miilleriano, Leipzig.

Ryvarden L. 1983. Type studies in the Polyporaceae 14. Species described by N. Patouillard, either

alone or with other mycologists. Occasional papers of Farlow Herbarium 18: 1-39.

Ryvarden L. 1988. Two new polypores from Burundi in Africa. Mycotaxon 31(2): 407-409.

Ryvarden L. 1991. Genera of polypores. Nomenclature and taxonomy. Synopsis Fungorum 5:

1363;

Ryvarden L, Gilbertson RL. 1994. European polypores 2. Synopsis Fungorum 6: 394-743.

Schrader HA. 1794. Spicilegium florae germanicae. C. Ritscher, Hannover.

Schumacher HCF. 1803. Enumeratio plantarum, vol. 2. F Brummer, Kobenhavn.

Shen LL, Cui BK, Dai Y-C. 2014. A new species of Postia (Polyporales, Basidiomycota) from

China based on morphological and molecular evidence. Phytotaxa 162(3): 147-156.

http://dx.doi.org/10.11646/3529

Tura D, Spirin WA, Zmitrovich IV, Wasser SP, Nevo E. 2008. Polypores new to Israel - 1: Genera

Ceriporiopsis, Postia and Skeletocutis. Mycotaxon 103: 217-227.

Walker J. 1996. An opinion on the validity of the generic name Postia Fries 1874 (Eumycota:

Aphyllophorales). Australasian Mycological Newsletter 15(2): 23-26.

Wei YL, Dai Y-C. 2006. Three new species of Postia (Aphyllophorales, Basidiomycota) from China.

Fungal Diversity 23: 391-402.

Yao YJ, Pegler DN, Chase MW. 1999. Application of ITS (nrDNA) sequences in the phylogenetic

study of Tyromyces s.l. Mycological Research 103(2): 219-229.

Yao YJ, Pegler DN, Chase MW. 2005. Molecular variation in the Postia caesia complex. FEMS

Microbiology Letters 242(1): 109-116.

Yuan HS, Dai YC, Wei YL. 2010. Postia cana sp. nov. (Basidiomycota, Polyporales) from Shanxi

Province, northern China. Nordic Journal of Botany 28(5): 629-631.

http://dx.doi.org/10.1111/j.1756-1051.2010.00849.x

MYCOTAXON

http://dx.doi.org/10.5248/129.415

Volume 129(2), pp. 415-419 October-December 2014

The new lectotypification of Umbilicaria kisovana

(Umbilicariaceae, lichenized Ascomycota)

EvGeny A. Davypov’, LIDIA YAKOVCHENKO??, & YOSHIHITO OHMURA*

' Altai State University, Lenin Avenue, 61, Barnaul, 656049, Russia

? Institute of Biology and Soil Science FEB RAS,

Stoletiya Vladivostoka Avenue, 159, Vladivostok, 690022, Russia

° Botanical Garden-Institute FEB RAS, Makovskogo Str, 142, Vladivostok, 690024, Russia

* Department of Botany, National Museum of Nature and Science,

4-1-1 Amakubo, Tsukuba, Ibaraki, 305-0005, Japan

* CORRESPONDENCE TO: eadavydov@yandex.ru

ABSTRACT — The protologue and typification of the name Gyrophora kisovana | Umbilicaria

kisovana, an East Asian lichen-forming fungus] are discussed. The lectotype is designated

from the Asahina collection deposited in TNS.

Key worps — authorship, Japan, nomenclature, spelling, valid publication

Introduction

Umbilicaria kisovana (Umbilicariaceae, \lichenized Ascomycota) is

characterized by small brown brittle thalli lacking rhizinomorphs and

thalloconidia, sometimes bearing gyrodisc apothecia containing octosporic

asci. The distribution of the species is restricted to East Asia (Japan, Korea,

Russian Far East, and China) where it grows on siliceous rocks at elevations

between 400 and 2100 m (Sat6 1956, Wei & Jiang 1993, Davydov & Zhdanov

2010). Wei & Jiang (1993) lectotypified the species name by an illustration

published in its protologue. However, Asahina (1931) cited several syntype

specimens in the protologue, and the lectotype should be selected from those

specimens. The purpose of this study is to analyze the protologue and to

designate the correct lectotype of U. kisovana from its syntypes.

Nomenclature

Asahina (1931) published Gyrophora kisovana (the basionym of Umbilicaria

kisovana) with a description in Japanese. Although the protologue lacks a

416 ... Davydov, Yakovchenko, & Ohmura

Latin description, the name was accepted by Asahina and therefore is validly

published according to ICN (2012: Art. 36.1(a), 38.1, 39.1).

Because the protologue was in Japanese only, there has been nomenclatural

confusion. Sat6 (1940) provided a Latin description for Gyrophora kisovana

based on the specimens cited by Asahina (1931) as well as several additional

specimens, including a fertile specimen from Korea collected by Saté. In the

Japanese summary, Satd (1940) explained the reason for providing a Latin

description as, “the detailed description with illustrations of “G. kisovana

Zahlbr. was provided by Asahina (1931) but the Latin description has not been

published yet. And also, the species was not cited in the Catalogus Lichenum

Universalis. Because of such situation this species seems not to be recognized

in science. Therefore, I provide the Latin description for the species””

Satd was most likely influenced by the attempts to introduce the mandatory

use of Latin in earlier Codes. The Brussels Rules (1912) required Latin

beginning from 1908 (Art. 35). However, the Cambridge Rules (1935), which

were in effect by 1940 although probably not taken into account by Sat6, moved

the date to 1935 (Art. 38) - thus legalizing the new plant names in Asahina

(1931). Sat6 (1956: 33) again published a Latin description (as “sp. nov.’)

citing the same reason used earlier (Sat6 1940). When Zahlbruckner (1940)

transferred the species into Umbilicaria, he cited Asahina (1931) as the source

of the basionym, an authorship accepted by Llano (1950) and Wei & Jiang

(1993), among others. However, Kurokawa & Nakanishi (1971) regarded Satd

(1940) as the valid publication for Gyrophora kisovana, and proposed a new

combination on that presumed basionym [i.e., Umbilicaria kisovana (Zahlbr. ex

M. Satd) Kurok.]. Index Fungorum (www.indexfungorum.org) lists the name

as Umbilicaria kisovana (Zahlbr. ex M. Sat6) Zahlbr.

For reference purposes, we provide a translation of the complete Japanese

protologue of Gyrophora kisovana (Asahina 1931), which may be difficult to

access for most of lichenologists, along with some comments:

«..., however a species which resembled Heppia guepini is distributed in Japan. I

gave the Japanese common name for the species as “Hime-iwatake” [that means

‘small iwatake; i.e., small Umbilicaria esculenta (Miyoshi) Minks], which was

collected by me in Tamba Province [old district where was located in parts of

Kyoto and Hyogo Prefectures, western Japan] at first. And then, this species

was also collected in Nezamenotoko, along Kiso River [Nagano Prefecture]

and middle slope of Mt. Nishikoma [= Mt. Nishikoma-ga-take, Kisokoma, or

Kisokoma-ga-take, in Nagano Prefecture]. Hime-iwatake is slightly larger than

Heppia guepini, however the thallus of hime-iwatake is also monophyllous and

gregarious. Thallus is thin and frangible; upper surface is dark brown, and lower

surface is dark blackish brown; and the thallus is attaching to granite rock by

the umbilicus on the lower surface. Distinct upper and lower cortex and gonidia

layer (Pleurococcus) are developed in the thallus. Apothecia are gyrodisc type.

Umbilicaria kisovana lectotypified ... 417

Asci are clavate, 66 x 18 Um, and 8-spored. Ascospores are ellipsoid, colorless,

simple, and 16-23 x 5-7 um in size. All of my collections were sterile but a

specimen collected from Miyajima Island in Hiroshima Prefecture (leg. Atsushi

Yamamoto) bear apothecia. I sent a specimen to Dr. Zahlbruckner, and he

named it Gyrophora kisovana Zahlbr. sp. nov., but it has not yet been published.»

For the lectotype of the name Gyrophora kisovana, Wei & Jiang (1993) proposed

the photograph of the species habit as well as the drawing of the species habit,

ascus and spores that Asahina (1931) published in the protologue. However,

since specimens were cited in the protologue, the lectotype must be chosen

from this part of the original material (ICN 2012: Art. 9.12, 9.19 + Ex. 13).

The above translation shows that Asahina (1931) cited four collections in

the protologue. They were collected from Tamba Province, Nezamenotoko

along Kiso River, middle slope of Mt. Nishikoma, and Miyajima Island in Aki

Province. The protologue indicates that although the first three collections

were sterile, the last (from Miyajima Island) was fertile. TNS houses specimens

labeled “Gyrophora kisovana” that were collected in the mentioned localities

before the publication of Asahina (1931), but the specimen from Miyajima

Island collected by Atsushi Yamamoto could not be found in TNS. Among

Asahina’s specimens, the specimen collected in Mt. Nishikoma is richly fertile,

which is inconsistent with the protologue, although Dr. Asahina labeled it as

“Typus” for Gyrophora kisovana. One of the remaining gatherings was collected

at “Oninokakehashi” located in Tamba. Other specimens, separated into three

packets, were collected along Kiso River in Agematsu-machi, Kiso-gun, Nagano

Prefecture where “Nezamenotoko” is situated but it was not mentioned on the

original label. They are morphologically consistent with the protologue and

possible candidates for the lectotype. We could not find the sterile specimen

on a rock, which appeared in the figure of the protologue. Among the available

original material, we designate one of the specimens (TNS-L-27375) collected

along Kiso River as the lectotype of Gyrophora kisovana (Fie. 1).

The nomenclature of the species should be treated as follows.

Umbilicaria kisovana (Zahlbr. ex Asahina) Zahlbr., Cat. Lich. Univ. 10: 405. 1940.

= Gyrophora kisovana Zahlbr. ex Asahina, J. Jap. Bot. 7: 327. 1931.

Type: Japan. Honshu. Prov. Shinano (Pref. Nagano): along Kiso River, Nezamenotoko,

Agematsu-machi, Kiso-gun, on granite rocks, elevation about 700 m, 17 October 1927,

Y. Asahina s.n. (Lectotype designated here, TNS-L-27375; isolectotypes, TNS-L-27389,

TNS-L-27390).

OTHER SYNTYPE TRACED: Japan. Honshu. Prov. Tamba (Pref. Hyogo): Oninokakehashi,

Hikami-gun, 14 July 1927, Y. Asahina s.n. (TNS-L-27393).

Asahina did not directly explain the etymology of Gyrophora kisovana in the

protologue, but the epithet was apparently derived from the locality, Kiso.

However, some of the original labels signed by Asahina read “Gyrophora

418 ... Davydov, Yakovchenko, & Ohmura

Wnty Wit

NATIONAL MUSEUM OF NATURE AND SCIENCE (TNS)

HERBARIUM OF LICHENS TNS-L-27375

LECTOTYPE

Gyrophora kisovana Zahlbr. ex Asahina

J. Jpn. Bot. 7: 327 (1931)

JAPAN. Honshu. Prov. Shinano (Pref. Nagano): along Kiso River, Nezamenotoko,

Agematsu-machi, Kiso-gun. On granite rocks; elevation about 700 m. October 17,

1927

v2 -Coll.: Y. Asahina

Bp Det:

FiGuRE 1. Gyrophora kisovana (lectotype, TNS-L-27375).

kisoana” (TNS-L-27390) or “Gyrophora kisogawana” (TNS-L-27393). These

provisional names have never appeared in the protologue or any other

publication, however, and the validly published original orthography “kisovana”

must be accepted (ICN 2012: Art. 60.1).

Acknowledgments

We acknowledge Dr. A. Sennikov (Finnish Museum of Natural History, University

of Helsinki) for his valuable comments, Dr. S. Chabanenko (Sakhalin Botanical Garden)

Umbilicaria kisovana lectotypified ... 419

for technical help, and Prof. B. McCune (Oregon State University) for improving the

text.

Literature cited

Asahina Y. 1931. The Raiken’s soliloquy on botanical science. XL. Heppia guepini (Del.) Nyl. and

Gyrophora kisovana A. Zahlbr. Journal of Japanese Botany 7: 325-327.

Davydov EA, Zhdanov IS. 2010. Umbilicaria kisovana and U. formosana (Umbilicariaceae,

lichenized Ascomycota) from Far East, new for Russia. Botanicheskii Zhurnal [St. Petersburg]

O57 7 285:

ICN. 2012. International Code of Nomenclature for algae, fungi, and plants (Melbourne Code).

Regnum Vegetabile 154. Koeltz Scientific Books. 208 p.

http://www.iapt-taxon.org/nomen/main.php

Kurokawa S, Nakanishi S. 1971. Lichens of the Hidaka Mountains, Hokkaido. Memoir of the

National Science Museum 4: 59-71.

Llano GA. 1950. A monograph of the lichen family Umbilicariaceae in the Western Hemisphere.

Washington D.C. 281 p.

Saté M. 1940. East Asiatic lichens (II). Journal of Japanese Botany 16: 42-47.

Sat6 M. 1956. Range of the Japanese lichens (I). Bulletin of the Faculty of Liberal Arts, Ibaraki

University, Natural Science 6: 27-39.

Wei JC, Jiang YM. 1993. The Asian Umbilicariaceae (Ascomycota). Mycosystema Monographicum

Series No. 1. Beijing: International Academic Publishers. 218 p.

Zahlbruckner A. 1940. Catalogus Lichenum Universalis 10: 1-660.

MY COTAXON

http://dx.doi.org/10.5248/129.421

Volume 129(2), pp. 421-427 October-December 2014

Galerella xalapensis sp. nov. found in

an urban green area in Xalapa, Veracruz, Mexico

ViIcTOR M. BANDALA * & LETICIA MONTOYA

Instituto de Ecologia, A.C., RO. Box 63, Xalapa, Veracruz 91000, Mexico

* CORRESPONDENCE TO: victor.bandala@inecol.mx

Axsstract — While monitoring species diversity of urban green areas in Xalapa city

(Veracruz, Mexico), a new species of Galerella was found in an abandoned urban green area.

After macro- and microscopic analysis it is proposed as a new species, Galerella xalapensis,

characterized by a faintly plicate-sulcate dry pale grayish brown pileus, white to yellowish

glabrous stipe, thick-walled rusty-brown ellipsoid basidiospores, utriform cheilocystidia, and

clampless hyphae. The new taxon is described and illustrated, and a comparison with similar

species is provided. This is the first Galerella recorded from Mexico.

Key worps — Agaricales, Bolbitiaceae, biodiversity, neotropical fungi, taxonomy

Introduction

A core of agaricoid species within the Bolbitiaceae (Agaricales), related

to Bolbitius Fr., Conocybe Fayod, and Pholiotina Fayod, has been recognized

as the genus Galerella Earle. Although supraspecific relationships derived

from molecular phylogenetic analyses have not yet been fully explored, the

striking combination of morphological features of the taxa accommodated in

Galerella led to their characterization by a Coprinus-like dry and plicate-sulcate

pileus, hymeniform pileipellis, rust-colored basidiospores, and lageniform

cheilocystidia (Horak 1968, Pegler 1986, Singer 1986, Hausknecht & Contu

2003). We refer especially to the works of Horak & Hausknecht (2002),

Arnolds & Hausknecht (2003), Hausknecht & Contu (2003), and Hausknecht

et al. (2004), who not only described new species but also reevaluated the

taxonomy of the genus through type studies of earlier named species and

integrated informative taxonomic characters to recognize six species currently

in Galerella. The literature shows that Galerella species are rare, but distributed

worldwide (Fic. 1). Two widespread species include G. plicatella (Peck) Singer,

described from USA and reported from Argentina, Brazil, India, Italy, and

422 ... Bandala & Montoya

Figure. 1. Representation of the worldwide distribution of Galerella species according reports

G. plicatelloides (=). G. xalapensis (4).

Trinidad (Dennis 1953, Singer & Digilio 1951, Thomas et al. 2001, Horak &

Hausknecht 2002, Hausknecht & Contu 2003, Arnolds & Hausknecht 2003,

Hausknecht et al. 2004) and G. fibrillosa Hauskn., described from Mauritius

and recorded from Brazil (Horak & Hausknecht 2002); but known only from

their type localities are G. floriformis Hauskn. from Vanuatu, G. microphues

(Berk. & Broome) Pegler from Sri Lanka, G. nigeriensis Tkalcec et al. from

Nigeria, and G. plicatelloides Sarwal & Locq. from India (Sarwal & Locquin

1983, Pegler 1986, Hausknecht & Contu 2003, Tkalcec et al. 2010).

Recently a bolbitiaceous fungus was found that turned out to represent

Galerella, a genus previously unrecorded from Mexico. The site is in an

abandoned green area, one of multiple points previously established to monitor

species diversity of the vascular plants, macrofungi, ants, butterflies, beetles,

amphibians, and birds that inhabit urban green areas in Xalapa city (Veracruz,

Mexico), a long-term program currently being conducted by a research team

of the Instituto de Ecologia. After a thorough morphological examination,

we concluded that the specimen’s unique combination of characters (faintly

plicate-sulcate dry pale grayish brown pileus, white to pale yellowish glabrous

stipe, thick-walled ellipsoid basidiospores, utriform cheilocystidia, clampless

hyphae) confirmed it as an undescribed species of Galerella.

Materials & methods

As part of the monitoring program developed in urban green areas from Xalapa city,

once a week during June-September 2012 we visited the intra-urban point located at

1230 m alt. south east of Xalapa that corresponds to a heavily disturbed patch of forest

formerly used as an orchard. After collecting the single Galerella sample in July 2012, we

Galerella xalapensis sp. nov. (Mexico) ... 423

sampled this point of our sampling mesh daily during May—October 2013 to document

thoroughly the fruiting variation of this Galerella species. The macroscopic depiction in

PL. 1 is based on field specimens. Macroscopic characters are described from material

and microscopic characters were based on dried basidiomata. Color descriptions are

according to Kornerup & Wanscher (1967). Tissue sections were mounted in 3% KOH

or 1% Congo Red aqueous solution for microscopical examination according to Largent

et al. (1977). 110 basidiospores (55 in lateral view and 55 in frontal view) were measured

in KOH, and 35 cheilocystidia in Congo Red, basidiospore measurements correspond

to length x width in side view x width in frontal view; X = mean length & width values;

Q = the mean length/width ratio. Line drawings were made under the microscope,

with the aid of a drawing tube. The examined specimen is deposited at XAL herbarium

(Thiers B., Index Herbariorum: http://sweetgum.nybg.org/ih/).

Taxonomy

Galerella xalapensis Bandala & Montoya, sp. nov. PLATE 1

MycoBank MB 809145

Differs from other similar species of Galerella by its pale grayish brown pileus, white

glabrous stipe, narrowly utriform to utriform cheilocystidia, and absence of clamp

connections.

Ho.otyPe: Mexico. Veracruz: Xalapa, 26 July 2012, Corona 699 (XAL).

Erymo_oey: the epithet refers to Xalapa, the city of origin.

BASIDIOMA slender, delicate, moderately small-sized. PrL—Eus 48-50 mm

broad, at first probably broadly convex, becoming plane-depressed or plane-

concave, with a weakly raised knob at center, surface pale grayish brown (5D8)

irregularly interrupted by paler, almost whitish radial lines, with a darker

contrasting area rounding the center, dull, dry, not hygrophanous, faintly

plicate-sulcate (Coprinus-like) from the disk towards the margin, this latter

somewhat undulating. LAMELLAE narrowly adnate, close, ventricose, whitish

to pale grayish, with paler, weakly fimbriate edges. Stipe 150 x 1-1.5 mm,

cylindric, almost straight, white, in parts pale yellowish (2.5 Y 8/2), finely and

weakly striate, glabrous, dry, fistulose. CONTEXT (pileus) very thin (<1 mm

thick), whitish (pileus and stipe); odor and taste not distinctive.

BASIDIOSPORES 8-11.5(-12.5) x 4.5-6 x 5.5-7(-7.5) um, ellipsoid in side

view, X = 9.5 x 5.3 um, Q = 1.80 um, broadly ellipsoid, often slightly tapering

towards apex and weakly angular at base, then somewhat subhexagonal or

subamygdaliform in dorso-frontal view, X = 9.6 x 6.1 um, (Q = 1.58 um), at

times the adaxial side faintly depressed then more or less reniform, thick-

walled (<1 um thick), smooth, with central, more or less truncate germ-

pore, reddish-orange, inamyloid, not-dextrinoid. Bastp1a 16-25 x 8-11 um,

clavate to broadly clavate, 4-spored, some 3-spored, hyaline, thin-walled.

PLEUROCYSTIDIA and PILEOCYSTIDIA absent. CHEILOCYSTIDIA 24-48(-55)

x 7-20(-22) um, apex 5-13 um broad, narrowly utriform to utriform, often

424 ... Bandala & Montoya

ventricose or with a flexuous neck, occasionally sublageniform, thin-walled,

smooth, hyaline, scarce on lamellae edge. PILEIPELLIS hymeniform, composed

of clavate to broadly clavate elements 18-35(-38) x 7-12(-13) um, thin-walled,

smooth, easily collapsing, hyaline or with a pale grayish content then in mass

the layer appears irregularly pale brownish, inamyloid. HyMENOPHORAL

TRAMA regular, made of cylindrical hyphae tightly arranged, hyaline, thin-

walled. PILEUS TRAMA not recovered, collapsed. CAULOCysTIDIA 21-41 x 6-9

um, more or less similar to cheilocystidia, scarce, occasionally in small groups.

CLAMP CONNECTIONS absent.

Hasitat — Solitary on naked soil, in a small slope of the floor of a green

heavily disturbed urban forest patch, with scattered trees (Eriobotrya japonica

(Thunb.) Lindl., Fraxinus uhdei (Wenz.) Lingelsh., Psidium guajava L., and

Platanus mexicana Moric.), shrubs (Bunchosia lindeniana A. Juss. and Solanum

nudum Dunal), and herbs (Hypoxis decumbens L., Paspalum conjugatum PJ.

Bergius, Acalypha alopecuroides Jacq., Desmodium uncinatum (Jacq.) DC.,

Cyperus hermaphroditus (Jacq.) Standl., and Rivina humilis L.).

DISTRIBUTION — Known only from the type locality in Xalapa.

REMARKS — The unique combined set of characters such as the faintly plicate-

sulcate dry pale grayish brown pileus, thick-walled ellipsoid basidiospores,

narrowly utriform to utriform cheilocystidia, and clampless hyphae distinguish

Galerella xalapensis. To a certain extent, its habit resembles G. microphues,

G. plicatella, and G. fibrillosa but the Mexican species differs strikingly in

remarkable important features. Galerella microphues, a rather uncommon

species restricted to Sri Lanka, is distinguished by narrowly lageniform

cheilocystidia (22-35 x 7-9 um) with a long tapering neck (10-20 x 1.5-2 um),

pyriform (16-25 x 11-16 um) pileipellis elements, and clamped hyphae (Pegler

1986). The most frequently encountered species (per Horak 1968, Hausknecht

& Contu 2003, Hausknecht et al. 2004, Thomas et al. 2001), G. plicatella is easily

diagnosed by its brown pileus with alternating regions of pale orange, yellowish

or fulvous-brown that turns alutaceous-buff with an orange-apricot disc, pale

orange lamellae, finely pubescent orange-white stipe, ventricose or lageniform

cheilocystidia with a remarkably slender long neck and measuring 30-50 x

6-11 um, 16.5-54 x 8-16.5 um, or 20-50 x 6.5-10 um, and clamped hyphae.

Galerella fibrillosa — known only from the holotype collected on soil in

Mauritius and another specimen from Brazil gathered on a decayed twig — is

readily separated by its small fragile basidiomata (pileus 7-15 mm; stipe 30-40

x 0.5-0.8 mm), a conspicuously crenate and fibrillose pale yellow-brown pileus

with yellowish ochre or pale brown disc, rapidly deliquescent pale brown to

yellowish brown lamellae, vesiculose cheilocystidia (28-)32-55(-62) x 10-20

um with more or less distinctive papilla to lageniform with elongate cylindrical

Galerella xalapensis sp. nov. (Mexico) ... 425

Pate 1. Galerella xalapensis (holotype, XAL Corona 699). a. Basidiomata. b. basidiospores. c. elements

of pileipellis. d. caulocystidia. e. basidia. f. cheilocystidia. Bars: a = 25 mm; b = 5 um; c-f = 10 um.

426 ... Bandala & Montoya

neck, and clamped hyphae (Horak & Hausknecht 2002). The remaining three

known Galerella species also differ: G. plicatelloides and G. floriformis can

be recognized by their fertile lamellae edges (i.e., cheilocystidia absent) and

G. nigeriensis by its strongly plicate-sulcate pale yellowish brown to light orange

brown pileus, pubescent stipe, tibiiform to lageniform cheilocystidia, and

presence of hymenophysalides (Sarwal & Locquin 1983, Hausknecht & Contu

2003, Tkaléec et al. 2010).

It is important to mention that G. xalapensis may have a particular fruiting

pattern, judging by the behavior observed in the sampled site. After a single

collection appeared in July 2012, we increased our sampling frequency to once

a week during August-September 2012, during which we found no basidiomata

of G. xalapensis. Although we visited the site daily during May-—October 2013,

we never again observed specimens of this Galerella species. The question is

whether G. xalapensis represents a fungal group that fluctuates in occurrence

and abundance, not fruiting for several consecutive months or years but

remaining in place as a physiologically active mycelium, or whether it suffers

from disturbance, so that finding a specimen at the site was a matter of chance

and good fortune. A similar example is represented by G. nigeriensis, which was

collected at the edge of a heavily disturbed secondary tropical forest (Tkaléec

et al. 2010). Given that we continue to monitor the study site thoroughly and

recognizing that the specimen combines a striking set of taxonomic characters,

we decided to propose the new species here. Nonetheless, we will continue to

monitor the type locality for G. xalapensis and other agaricaceous fungi as part

of our sampling program in Xalapa’s urban green areas.

Acknowledgments

We thank INECOL for the financial seed grant to support the monitoring program

in the urban green areas in Xalapa. We appreciate the collaboration in the monitoring

of macrofungi in the field of the biologists J.C. Corona and D. Ramos, this latter also

assisted us during the microscopic study. Special thanks to E. Saavedra for elaborating

the line drawing of the basidiomata and Dr. L. Lorea for the plants information from

the site studied (all at INECOL). We acknowledge the valuable revision and comments

on the manuscript by Dr. A. Hausknecht (Austrian Mycological Society) and Dr. I.

Krisai-Greilhuber (University of Vienna). Especial thanks to Dr. S. Pennycook and Dr.

L. Norvell for their meticulous revision and improving the text.

Literature cited

Arnolds E, Hausknecht A. 2003. Notulae ad floram agaricinam neerlandicam — XLI: Conocybe and

Pholiotina. Persoonia 18: 239-252.

Dennis RWG.1953. Les Agaricales de I'lile de la Trinité: Rhodosporae-Ochrosporae. Bull. Soc. Myc.

France 69: 145-198.

Hausknecht A, Contu M. 2003. The genus Galerella. A world-wide survey. Osterr. Z. Pilzk. 12:

31-40.

Galerella xalapensis sp. nov. (Mexico) ... 427

Hausknecht A, Krisai-Greilhuber I, Voglmayr H. 2004. Type studies in North American species

of Bolbitiaceae belonging to the genera Conocybe and Pholiotina. Osterr. Z. Pilzk. 13: 153-235.

Horak E. 1968. Synopsis generum Agaricalium (Die Gattungstypen der Agaricales). Beitrage zur

Kryptogamenflora der Schweiz. Band 13. 741 p.

Horak E, Hausknecht A. 2002. Notes on extra-European taxa of Bolbitiaceae (Agaricales,

Basidiomycota). Osterr. Z. Pilzk. 11: 213-264.

Kornerup A, Wanscher JH. 1967. Methuen handbook of colour. 2nd edn. Methuen, London. 243

p. 30 pl.

Largent D, Johnson D, Watling R. 1977. How to identify mushrooms to genus III: Microscopic

features. Mad River Press, Eureka. 148 p.

Pegler DN. 1986. Agaric Flora of Sri Lanka. Kew Bulletin Additional Series 12. 519 p.

Sarwal BM, Locquin MV. 1983. Les champignons de ’Himalaya dans leurs relations avec la flore

eurasiatique. Compt. Rend. Congr. Natl. Soc. Savantes, Sec. Sci. 108: 191-201.

Singer R. 1986. The Agaricales in modern taxonomy. 4th ed. Koeltz Scientific Books: Koenigstein.

981 p. 8 pl.

Singer R, Digilio APL .1951. Prédromo de la Flora Agaricina Argentina. Lilloa 25: 5-461.

Tkaléec Z, Mesié A, Cerkez M. 2010. Galerella nigeriensis (Agaricales), a new species from tropical

Africa. Mycotaxon 114: 263-270. http://dx.doi.org/10.5248/114.263

Thomas KA, Hausknecht A, Manimohan P. 2001. Bolbitiaceae of Kerala State, India: new species

and new and noteworthy records. Osterr. Z. Pilzk. 10: 87-114.

MY COTAXON

http://dx.doi.org/10.5248/129.429

Volume 129(2), pp. 429-432 October-December 2014

A new report of Uromyces ambiens on Buxus from Pakistan

SADIQULLAH’, A. ISHAQ’, M. FiAz', N.S. AFSHAN?”, & A.N. KHALID?

"Department of Botany, Hazara University, Mansehra

*Department of Botany, & >*Centre for Undergraduate Studies, University of the Punjab,

Quaid-e-Azam Campus, Lahore, 54590, Pakistan

*CORRESPONDENCE TO: pakrust@gmail.com

ABSTRACT — The rust fungus, Uromyces ambiens, is described and illustrated based on a

specimen of Buxus wallichiana, Himalayan boxwood, from northern Pakistan. This rare

fungus is reported for the first time from this host and for the first time from Pakistan.

Key worps — dimorphic spores, Himalayas, Puccinia buxi, Shangla

Introduction

During the exploration of Uredinales of District Shangla, Khyber

Pakhtunkhwa, Pakistan, trees of Himalayan boxwood, Buxus wallichiana, were

found to be infected with a species of Uromyces. Leaves of B. wallichiana collected

in September 2013 were infected with pale yellowish spermogonial sori. Later

samples collected in November and January showed both spermogonial and

telial sori. After an extensive literature survey, we identified the rust fungus on

Buxus as Uromyces ambiens.

Another rust fungus, Puccinia buxi Sowerby, known as boxwood rust,

occurs on many species of Buxus. It is widespread throughout Europe but

rarely reported on native hosts in China and Japan. The boxwood rust has been

intercepted at ports of entry in North America over the past decades and was

discovered in Pennsylvania on recently received nursery stock of boxwood

from Greece (Yun 2014).

Himalayan boxwood is a medium sized small tree found commonly in the

western and central Himalayas at elevations ranging from 1200 to 2900 m

(Viswanath et al. 2006). The tree is native to Pakistan and India. In Pakistan, it

grows in the sub-Himalayan tract from Azad Kashmir westward to Rawalpindi,

Islamabad, Murree, Hazara, Chitral, Swat, Shangla, Rawlakot, and the Gadoon

area (Stewart 1972).

430 ... Sadiqullah & al.

Our collections of Uromyces ambiens represent a new record for Pakistan as

well as a new host record. Pakistan is the second country in which this rare rust

on Buxus occurs. In addition, this represents the only recent re-description and

illustration of this species.

SSie

PLaTE 1: Uromyces ambiens. A, B: Leaves of Buxus wallichiana showing raised spermogonial

and telial sori. C: Inflorescence of Buxus wallichiana. D: Telial sori under stereomicroscope.

E: Longitudinal section of spermogonium. F-H: Dimorphic teliospores. Scale bar = 10 um.

Uromyces ambiens on Buxus wallichiana (Pakistan) ... 431

Materials & methods

Freehand sections and scrape mounts of infected plant materials were made in

lactic acid. The plant was photographed and infected portions were observed under a

stereomicroscope. Twenty spores of each spore state were examined under a microscope

(Nikon YS 100) and measured using an ocular micrometer (Zeiss Eyepiece Screw

Micrometer). Sections and spores were photographed by Digiporo-Labomed. The

spores were illustrated with the aid of a camera lucida (Ernst Leitz Wetzlar Germany).

Taxonomy

Uromyces ambiens Cooke, Grevillea 3: 75 (1874) PLATES 1, 2

AECIA and UREDINIA not found. SPERMOGONIA subepidermal, group 1, type

1, golden brown to hyaline, 175 x 137 um. TeExta abaxial, raised, developing

as circular black boundaries around central yellow spots, scattered, naked,

chestnut brown, 2-3 x 4-5 mm. TELIOsporEs dimorphic: ovate teliospores

yellow, elongated, 1-celled, 29-40 x 48-60(-79), walls smooth, yellow; globose

to subglobose teliospores brown, 29-47 x 39-48 um, walls smooth and dark

prays

EES

ENOL LATENT MSE GPT E LOTT Ts

PLATE 2: Uromyces ambiens: teliospores. Scale bar = 10 um.

432 ... Sadiqullah & al.

brown, 2.8-5 um thick at sides, up to 3.5 um thick apically, germ pore one,

subapical to apical; pedicel hyaline, 6.5-9 x 160-205(-285) um.

MATERIAL EXAMINED: PAKISTAN, KuHYBER PAKHTUNKHWA, Shangla district,

Yukhtange, at 2200 m a.s.l., on Buxus wallichiana Baill. (Buxaceae), stages 0 + III, 2

Sep. 2013, A.N. Khalid AM-S1 (LAH 20914); 4 Nov. 2013, Sadiqullah RS 36A (LAH

20913A); 1 Jan. 2014, Sadiqullah RS 36B (LAH 20913B).

ComMENTs: Cooke (1874) originally described Uromyces ambiens on Buxus

sempervirens L. from the Indian Himalayas, and Sydow (1922) published a

revised and expanded description of the type collection. Later Butler & Bisby

(1931) reported this species on leaves of Buxus (probably sempervirens) near

Dunooltie above Dehra Dun as well as from Bashahr, near Simla, 6000 ft.

elevation. Although Spaulding (1961) listed U. ambiens as present in India and

Pakistan, we found nothing in the Pakistani literature about this rust.

Uromyces ambiens differs morphologically from Puccinia buxi. In

U. ambiens the teliospores are dimorphic (ovate or globose to subglobose),

non-septate, and thick-walled (up to 5 um thick on the sides), while PR. buxi

has only monomorphic teliospores that are oblong to clavate, one-septate, and

thin-walled.

Acknowledgements

We are sincerely thankful to Dr. Amy Rossman (Systematic Mycology and

Microbiology Laboratory, USDA-ARS, Beltsville) and Dr. Abdul Rehman Niazi

(Department of Botany, University of the Punjab, Lahore, Pakistan) for acting as

presubmission reviewers and giving valuable suggestions.

Literature cited

Butler EJ, Bisby GR. 1931. The fungi of India. The Imperial Council of Agricultural Research.

Calcutta: Government of India, Central Publication Branch. 237 p.

Cooke MC. 1874. Himalayan leaf fungi. Grevillea 3: 75-76.

Spaulding P. 1961. Foreign diseases of forest trees of the world. U.S.D.A. Agricultural Handbook

197. 361 p.

Stewart RR. 1972. An annotated catalogue of the vascular plants of West Pakistan and Kashmir.

Fakhri Printing Press, Karachi. 1028 p.

Sydow H. 1922. Uber einige wenig bekannte Uredineen aus dem Kew Herbar. Annales Mycologici

20: 54-60.

Viswanath S, Singh RP, Thapliyal RC. 2006. Seed bank dynamics of Buxus wallichiana Baillon in a

Himalayan moist temperate forest. Tropical Ecology 47: 145-148.

Yun, H.Y. Systematic Mycology and Microbiology Laboratory, ARS, USDA. . Invasive Fungi. Box

Rust or Boxwood Rust - Puccinia buxi. /sbmlweb/fungi/index.cfm (Retrieved April 15, 2014,)

MY COTAXON

http://dx.doi.org/10.5248/129.433

Volume 129(2), pp. 433-438 October-December 2014

Lophium elegans (Ascomycota),

a rare European species

GEIR MATHIASSEN*, ALFRED GRANMO, & TEPPO RAMA

Tromso University Museum, University of Tromso — ‘The Arctic University of Norway,

PO Box 6050 Langnes, NO-9037 Tromso, Norway

* CORRESPONDENCE TO: geir.mathiassen@uit.no

Asstract — Lophium elegans (Mytilinidiales, Mytilinidiaceae) is recorded for the first time

in northern Europe. It was found on Juniperus communis in two locations in Finnmark

County, Norway, which represent the northernmost finds for the species. A morphological

description of the species is given and its ecology and distribution discussed.

Key worps — host preference, junipericolous fungi

Introduction

Lophium elegans is an ascomycete known to science for about 60 years (Zogg

1954) but collected only a few times by a small number of mycologists. The

species has been found only in Europe where it has been reported from rather

high altitudes in southern France (Alpes Maritimes, Hautes Alpes), northern

Italy (South Tyrol), Switzerland, and Scotland (Zogg 1962, Kirk & Spooner

1984). The finds from southern Europe were all made between 1940 and 1960

(Zogg 1962), and the two finds from Great Britain date back to the beginning of

the 1980s (Kirk & Spooner 1984). In 2008, the species was collected from near

Dijon, in the Burgundy region of east central France (Lechat 2013).

Lophium elegans seems to be restricted to Juniperus. In Great Britain and

France the species was found on J. communis, and in Italy and Switzerland on

J. communis ssp. nana [as J. nana] (Zogg 1962, Kirk & Spooner 1984, Lechat

2013). Apart from Lophium igoschinae Chleb., found on Dryas (Rosaceae;

Chlebicki & Knudsen 2001), all Lophium species have been collected only from

coniferous substrates (Zogg 1962, Boehm et al. 2009). Zogg (1962) reported

Lophium elegans as growing on bark of small living and dead twigs of its host

tree, while Ellis & Ellis (1997) reported it on wood of J. communis.

434 ... Mathiassen, Granmo, & Rama

Here we report three records of L. elegans from Northern Norway, where the

species was found on J. communis in two different locations. These represent

the first northern European records of the species.

Materials & methods

Lophium elegans was collected during ascomycete field inventories in Finnmark,

Norway, 2010 and 2011. Geographical coordinates were taken in the field using GPS. The

collections were studied using Wild M10 and Zeiss 475052-9901 dissecting microscopes

and Leitz DMRBE and Zeiss 473028 light microscopes. Microscopic slides were

prepared from dried herbarium material and mounted in water for measurements and

photographs. Photographs were taken with Olympus UC30 and UCS0 digital cameras.

Collector abbreviations include GM (Mathiassen) and AG (Granmo). Specimens are

deposited in the herbarium of Troms University Museum, Norway (TROM).

Taxonomy

Lophium elegans H. Zogg, Ber. Schweiz. Bot. Ges. 64: 141 (1954) FIG. 1

ASCOMATA 200-500 um high, 180-340 um wide, and 90-120 um thick, seated

upright on bark or erumpent through cracks in bark, scattered or clustered,

laterally flattened and often striate, usually axe-head shaped (dolabriform),

somewhat tapering towards base, with a long slit-like ostiolum along the +

sharp upper edge, brittle and black. Asci 160-210 x 7.2-8.7 um, cylindrical,

bitunicate, short stiped, containing 8 spirally entwined spores. PARAPHYSOIDS

ca. 15 um diam., long, branched, anastomosing, septate and _ hyaline.

ASCOSPORES 210-265 um x 1.3-2 um, filiform, tapered at ends, multiseptate

with septa at intervals of mostly 5-8 um, yellowish hyaline, gradually becoming

spirally arranged within the ascus.

SPECIMENS EXAMINED: NORWAY, FINNMARK: Municipality of Alta, the valley

Eibydalen, 69°46’30"N 23°18'49"E, 217 m asl, on dead twig of Juniperus communis L.,

17.VIII.2011, G. Mathiassen & A. Granmo, GM 12472 C (TROM F-41138); 69°46’31"N

23°18'28"E, 204 m asl, on dead twig of J. communis, 17.VUI.2011, A. Granmo & G.

Mathiassen, AG 25A/2011 (TROM F-25034). Municipality of Karasjok, Basavzeguoika,

in the upper, southern part of the Anarjohka river valley, 53°20°689” N 33°22’29” E, 205

m asl, on dead twigs of J. communis, 18.VIII.2010, G. Mathiassen & A. Granmo, GM

12297 A (TROM F-25741).

Discussion

Lophium elegans was found on Juniperus communis at Basavzeguoika 205

m asl, in the Anarjohka river valley, Karasjok, in August 2010. This area is a

continental part of Northern Norway with relatively high summer temperatures

(mean of the warmest month, July, 13.1°C; DNMI 1991) and low precipitation

(interpolated annual precipitation 366 mm; Moen 1998). The species was

also found twice in 2011 in the valley Eibydalen in Alta. The Alta area is an

inner fjord district in the middle boreal zone with luxuriant woodland and

Lophium elegans in Norway ... 435

Fic. 1. Lophium elegans: a. Ascomata (GM 12297 A). b. Asci containing ascospores (GM 12472 C).

c. One ascospore (GM 12472 C). Photos: GM (a-b), TR (c).

warm summers (mean of the warmest month, July, 13.4°C; DNMI 1991). Our

first find at Basavzeguoika and the world’s northernmost finds in Eibydalen

represent the first records of L. elegans from northern Europe despite earlier

searches by the renowned Swedish ascomycete researchers Kerstin and

Lennart Holm. They were well aware of this species, but did not find it in any

of the Nordic countries (Holm & Holm 1977). The species has a disjunctive

distribution pattern on the European mainland (Fic. 2), similar to that of a

few other pyrenomycetous species, e.g., Glyphium grisonense Math., Hypoxylon

macrosporum P. Karst., and Saccardoella kanderana Math. (Mathiassen 1989,

1993; Granmo 1999; Mathiassen & Granmo 2012). Therefore, as L. elegans is

known from Scotland, it is likely that some of those species may also occur at

436 ... Mathiassen, Granmo, & Rama

Fic. 2. World distribution of Lophium elegans. The large dot represents 6 localities along the

University Museum, 2013.

high altitudes in the northern British Isles. Although L. elegans is undoubtedly

a rather rare species in Europe, we expect it to occur occasionally along the

Lophium elegans in Norway ... 437

whole Scandinavian mountain range as well as in other high altitude localities

along the Alps, e.g., in Austria and southern Germany.

The Norwegian records strengthen the impression of L. elegans as being

restricted to Juniperus, which we consider a junipericolous fungus. The three

Norwegian specimens had ascocarps growing on bark as well as on rather small

twigs. Nevertheless, one single ascocarp was observed growing on a needle,

which has never been reported earlier (cf. Kirk & Spooner 1984). Because of

this and Ellis & Ellis’ (1997) observation from naked wood, L. elegans should

be considered not as a strictly, but rather a predominantly, corticolous species.

The ascospores of L. elegans are very long and arranged in a spiral parallel

configuration in the asci, which Zogg (1954) termed “parallel-spiralig

aufgerollte” spores. However, Kirk & Spooner (1984) maintained that this

character may be exhibited only at maturity, in that ascospores in the British

collections lay parallel in young immature asci and spirally coiled in mature

asci. One Norwegian sample (AG 25A/2011) supports this observation.

Lophium elegans is morphologically rather similar to L. mytilinum (Pers.: Fr.)

Fr., which differs by its longer, thicker ascocarps (Zogg 1962). In addition,

L. mytilinum ascospores always lie parallel in, and remain shorter than, the

asci (Zogg 1962). Kirk & Spooner (1984) pointed out that L. elegans was

evidently very closely related to L. mytilinum but distinguished particularly by

the spiral coiling of the ascospores. However, molecular data indicate that the

two species are not closely related within the family Mytilinidiaceae (Boehm et

al. 2009), and that Lophium elegans and L. mytilinum might, in fact, be placed

in two separate genera in that family. According to Boehm et al. (2009) both

Lophium and Mytilinidion are polyphyletic. However, until the type species of

Mytilinidion, M. aggregatum (DC.: Fr.) Duby, and type species of other genera

in Mytilinidiaceae are included in a common phylogenetic analysis with taxa

mentioned by Boehm et al. (2009), any new nomenclatural combinations can

only be speculative.

Acknowledgments

We express our gratitude to Ove E. Eriksson and Liliane E. Petrini for reviewing the

of Lophium. Rob Barrett has kindly improved the language. Ernst Hogtun provided

technical assistance. The study was financially supported by Artsdatabanken (The

Norwegian Biodiversity Information Centre), and University of Tromsa — The Arctic

University of Norway.

Literature cited

Boehm EWA, Mugambi G, Miller AN, Huhndorf SM, Marincowitz S, Spatafora JW, Schoch CL.

2009. A molecular phylogenetic reappraisal of the Hysteriaceae, Mytilinidiaceae and Gloniaceae

(Pleosporomycetidae, Dothideomycetes) with keys to world species. Studies in Mycology 64: 49-83.

http://dx.doi.org/10.3114/sim.2009.64.03

438 ... Mathiassen, Granmo, & Rama

Chlebicki A, Knudsen H. 2001. Dryadicolous microfungi from Greenland. I. List of species. Acta

Societatis Botanicorum Poloniae 70: 291-301. http://dx.doi.org/10.5586/asbp.2001.037

DNMI (Det norske meteorologiske institutt) 1991. Temperaturnormaler 1961-1990. Forelopige

normaler. Oslo.

Ellis MB, Ellis JP. 1997. Microfungi on land plants: an identification handbook. 2nd ed. Richmond

Publishing. 868 p.

Granmo A. 1999. Morphotaxonomy and chorology of the genus Hypoxylon (Xylariaceae) in

Norway. Sommerfeltia 26. 81 p.

Holm K, Holm L. 1977. Nordic junipericolous ascomycetes. Symbolae Botanicae Upsalienses

21(3). 70 p.

Kirk PM, Spooner BM. 1984. An account of the fungi of Arran, Gigha and Kintyre. Kew Bulletin

38: 503-597. http://dx.doi.org/10.2307/4108573

Lechat C. 2013. ASCOfrance. http://www.ascofrance.fr [accessed June 2013].

Mathiassen G. 1989. Some corticolous and lignicolous Pyrenomycetes s. lat. (Ascomycetes) on Salix

in Troms, N Norway. Sommerfeltia 9. 100 p.

Mathiassen G. 1993. Corticolous and lignicolous Pyrenomycetes s. lat. (Ascomycetes) on Salix along

a mid-Scandinavian transect. Sommerfeltia 20. 180 p.

Mathiassen G, Granmo A. 2012. Sluttrapport for Artsprosjektet Sekksporesopper i Finnmark

2010-2011. Tromso Museum, Universitetsmuseet 2012, ADB 56-09, Prosjekt 70184216.

Available at: http://www.artsdatabanken.no/artArticle.aspx?m=259&amid=7030.

Moen A. 1998. Nasjonalatlas for Norge: Vegetasjon. Statens Kartverk, Honefoss. 200 p.

Zogg H. 1954. Uber eine neue Hysteriaceen-Art, Lophium elegans n. sp. Berichte der Schweizerischen

Botanischen Gesellschaft 64: 139-141.

Zogg H. 1962. Die Hysteriaceae s. str. und Lophiaceae unter besonderer Beriicksichtigung der

mitteleuropdischen Formen. Beitrage zur Kryptogamenflora der Schweiz 11(3). 190 p.

MY COTAXON

http://dx.doi.org/10.5248/129.439

Volume 129(2), pp. 439-454 October-December 2014

New and interesting Laboulbeniales

from southern and southeastern Asia

D. HAELEWATERS" & S. YAAKOP?

"Farlow Reference Library and Herbarium of Cryptogamic Botany, Harvard University

22 Divinity Avenue, Cambridge, Massachusetts 02138, U.S.A.

?Faculty of Science & Technology, School of Environmental and Natural Resource Sciences,

Universiti Kebangsaan Malaysia, Bangi 43600, Malaysia

* CORRESPONDENCE TO: dhaelewaters@fas.harvard.edu

ABSTRACT — Two new species of Laboulbenia from the Philippines are described and

illustrated: Laboulbenia erotylidarum on an erotylid beetle (Coleoptera, Erotylidae) and

Laboulbenia poplitea on Craspedophorus sp. (Coleoptera, Carabidae). In addition, we present

ten new records of Laboulbeniales from several countries in southern and southeastern Asia

on coleopteran hosts. These are Blasticomyces lispini from Borneo (Indonesia), Cantharomyces

orientalis from the Philippines, Dimeromyces rugosus on Leiochrodes sp. from Sumatra

(Indonesia), Laboulbenia anoplogenii on Clivina sp. from India, L. cafii on Remus corallicola

from Singapore, L. satanas from the Philippines, L. timurensis on Clivina inopaca from Papua

New Guinea, Monoicomyces stenusae on Silusa sp. from the Philippines, Ormomyces clivinae

on Clivina sp. from India, and Peyritschiella princeps on Philonthus tardus from Lombok

(Indonesia).

KEY worps — Ascomycota, insect-associated fungi, morphology, museum collection study,

Roland Thaxter, taxonomy

Introduction

One group of microscopic insect-associated parasitic fungi, the order

Laboulbeniales (Ascomycota, Pezizomycotina, Laboulbeniomycetes), is

perhaps the most intriguing and yet least studied of all entomogenous fungi.

Laboulbeniales are obligate parasites on invertebrate hosts, which include

insects (mainly beetles and flies), millipedes, and mites. About 80% of the

Laboulbeniales described thus far are parasitic on the order Coleoptera (Weir

& Blackwell 2005). With currently about 2,000 species in 140 genera (Rossi

& Santamaria 2012), the total number of Laboulbeniales associated with

arthropod hosts is estimated to be between 15,000 and 75,000 species (Weir &

440 ... Haelewaters & Yaakop

Hammond 1997), most of which still need to be described from wet tropical

regions (Haelewaters et al. 2014a).

This paper presents ten new records for different southern and southeastern

Asian countries. Additionally, we report thalli in the genus Laboulbenia that

did not fit any existing description and therefore represent two new species.

Materials & methods

Over 23,000 insect specimens in the collection of Invertebrate Zoology at the

American Museum of Natural History (AMNH) were screened for the presence of

entomopathogenic fungi using a Nikon SMZ-U stereomicroscope. Almost six percent

of these insects were found infested with Laboulbeniales (Haelewaters unpublished).

Although the vast majority of the entomological collection at AMNH is the result of

samplings in North America, a few infected hosts from Asia were found. Individual

thalli were removed from the host’s integument with a Minuten Pin (BioQuip #1208SA)

and embedded in Amann solution or PVA Mounting Medium (BioQuip #6371A). Cover

slips were ringed with transparent nail varnish. All microscopic slides are deposited at

the Farlow Herbarium, Harvard University (FH).

Additional fungal material was found in unidentified mountings from Roland

Thaxter, whose complete collection is deposited at the Farlow Herbarium, Harvard

University (FH).

Morphological analyses, measurements, and photographs were made using an

Olympus BX40 light microscope with Olympus XC50 digital camera and MicroSuite

Special Edition software 3.1 (Soft Imaging Solutions GmbH). Images were optimized

(with LEVELs and BRIGHTNESS/CONTRAST tools) and cropped in Adobe Photoshop CS

Version 8.0 (San Jose, California).

Taxonomy

Blasticomyces lispini (Thaxt.) II. Tav., Mycol. Memoir 9: 155 (1985) PLATE 1A

SPECIMEN EXAMINED: INDONESIA, BORNEO, CENTRAL KALIMANTAN Prov, Tanjung

Puting National Park, on “staphylinid close to Lispinus” [sic] (Staphylinidae, Osoriinae),

1926, leg. Eric Mjéberg, Thaxter 3571, slide FH 00313486 (4 thalli collected from elytra).

PREVIOUS RECORDS: On Nacaeus impressicollis [as Lispinus] and various Lispinus sp.

from Indonesia (Java), Taiwan, Sri Lanka, the Philippines, China, Malaysian Borneo,

and the Solomon Islands (Thaxter 1915, 1931, Majewski & Sugiyama 1986).

Three species are included in Blasticomyces 1.1. Tav. All three have been

previously observed on the island of Borneo: B. denigratus T. Majewski &

K. Sugiy. on Lispinus coarcticollis from Sabah (Malaysia), B. fastigiatus (Thaxt.)

LI. Tav. on Cercyon sp. from Sarawak (Malaysia), and B. lispini on Lispinus spp.

from Sarawak and Sabah (Malaysia) (Thaxter 1931, Majewski & Sugiyama

1986). Our specimens undoubtedly represent B. lispini, based on (1) no

blackening of the basal parts of the receptacle, (2) receptacle consisting of

superposed, flattened cells, usually undivided (a few cells may be divided by

two vertical septa), and (3) a subconical and rather short perithecium. One

Laboulbenia spp. nov. (Philippines) ... 441

Tai

re,

A B

PiaTE 1. A. Blasticomyces lispini (FH 00313486). B. Cantharomyces orientalis (FH 00313465).

C. Laboulbenia anoplogenii (FH 00313461). Scale bars = 50 um.

studied thallus differs from the original description of B. lispini in displaying

shallow blackening of the outer margins of receptacle basal cells 2-4.

Some Lispinus species have been transferred to Nacaeus. Nacaeus (Lispinus)

impressicollis is a cosmopolitan species; it is broadly distributed in Australia,

Japan (Naomi 1997), Central and South America, many Caribbean islands,

Atlantic islands, and the Indo-Pacific biogeographic region (Irmler 2003).

Cantharomyces orientalis Speg., Anales Mus. Nac. Hist. Nat. Buenos Aires 27: 43

(1915) PLATE 1B

SPECIMENS EXAMINED: THE PHILIPPINES, LUZON ISLAND, METROPOLITAN MANILA,

Manila, on “aleocharid” [sic] (Staphylinidae, Aleocharinae), Dec 1911, no collector,

Thaxter 2421, slide FH 00313465 (2 thalli).

PREVIOUS RECORDS: On Carpelimus (syn. Trogophloeus) and related genera

(Staphylinidae, Oxytelinae); two records were collected from Bledius (Staphylinidae,

Oxytelinae). Cantharomyces orientalis is reported from Finland, Russia (Huldén 1983),

Germany (Scheloske 1969), Italy (Spegazzini 1915), Poland (Siemaszko & Siemaszko

1928, 1932, Majewski 1994), the Netherlands [as C. thaxteri] (Middelhoek 1949),

Spain (Santamaria 1989), Sweden, Switzerland (Huldén 1985), Belgium (De Kesel &

Haghebaert 1991), Great Britain (Weir & Beakes 1993), Greece (Castaldo et al. 2004),

Czech Republic, Slovakia (Rossi et al. 2010), and Algeria [as C. abbreviatus] (Maire

1920).

All 29 Cantharomyces species are characterized by a receptacle consisting of

three superposed cells and a compound antheridium subtending a simple or

variably branched primary appendage, and a perithecium having four to five

442 ... Haelewaters & Yaakop

cells in each vertical row of outer wall cells (Haelewaters & De Kesel 2013). Hosts

are mainly Staphylinidae (subfamily Oxytelinae, tribes Blediini, Euphaniini, and

Oxyteliini; subfamily Aleocharinae, tribes Aleocharini, and Oxypodini), next to

Dryopidae, Limnichidae, and Hydrophilidae.

Cantharomyces orientalis is a very variable species, leading to confusion

among all Cantharomyces species occurring on Carpelimus s.l. (see Santamaria

2003). The Philippine thalli have the primary appendage broken above the

suprabasal cell (see Majewski 1990) but otherwise correspond to the description

for C. orientalis with cell I hyaline and contrasting with the yellowish amber

toned cells II (isodiametric to trapezoidal) and III (subequal to cell II but

less high), the squarish basal cell of the primary appendage with the lateral

antheridium, cell VI hyaline and elongated (<40 um), and the perithecium 68 x

37 um, symmetrical, and broadest near basal third.

The Philippine finding of C. orientalis represents the first record of this

species for Asia.

Dimeromyces rugosus Thaxt., Proc. Amer. Acad. Arts 55: 245 (1920) PLATE 3A

SPECIMENS EXAMINED: INDONESIA, SUMATRA, WEsT SUMATRA PRov., Mount

Singgalang, on Leiochrodes sp. (Tenebrionidae, Diaperinae), Nov-Dec 1925, leg. Edward

Jacobson, Thaxter 3466, slide FH 00313485 (4 male + 4 female thalli from the legs).

PREVIOUS RECORDS: Known from material mentioned in the original description

(Thaxter 1920), on Leiochrodes medianus from the Solomon Islands and on L. minutus

from Malaysian Borneo. Afterwards recorded only in Taiwan, on Leiochrodes spp.

(Terada 1976).

Dimeromyces Thaxt. is a large genus with 109 species (Kirk et al. 2008),

parasitizing Acarini, Blattodea, Dermaptera, Thysanoptera, Coleoptera, and

Diptera (Santamaria 2003). Its perithecial wall showing conspicuous transverse

striae above the hyaline base easily distinguishes D. rugosus from other species.

In addition, two tiny papillae are borne on opposite sides of the perithecial

apex.

Laboulbenia anoplogenii Thaxt., Proc. Amer. Acad. Arts 35: 156 (1899) | PLATE 1¢

= Laboulbenia stenolophi Speg., Redia 10: 65 (1914)

SPECIMENS EXAMINED: INDIA, WesT BENGAL STATE, Kanchrapara, on Clivina

sp. (Carabidae, Scaritinae), 5 Aug 1944, leg. Mont A. Cazier, D. Haelew. 305, in coll.

American Museum of Natural History, slides FH 00313459 (4 thalli from right elytron),

FH 00313460 (1 thallus from right metatrochanter), FH 00313461 (3 thalli from left

mesocoxa + mesotrochanter), FH 00313462 (2 thalli from right elytron), and FH

00313463 (1 thallus from right mesofemur).

PREVIOUS RECORDS: Described from Anoplogenius cyanescens [as A. circumcinctus]

(Carabidae, Harpalinae). In Asia, beetles in Abacetus, Chlaeminus (Carabidae,

Pterostichinae), Egadroma, Harpalus, Platynus [as Colpodes in Juan & Chien 1995],

and Stenolophus (Carabidae, Harpalinae) are known to host this parasite (Santamaria

Laboulbenia spp. nov. (Philippines) ... 443

et al. 1991, Lee et al. 2002, Shen et al. 2006). Despite some confusion (see below),

L. anoplogenii has a global distribution but has not yet been reported from South

America (Santamaria et al. 1991).

Laboulbenia anoplogenii has been reported from India only once, from

Bembidion (Carabidae, Trechinae) (Kaur et al. 1993). The same authors also

report L. stenolophi Speg. and L. egens Speg. [misidentified as L. tachyis Thaxt.]

from Bembidion, thereby adding the latter as a new host genus for all three

parasite species. However, we suggest that this material should be re-examined.

Santamaria (1989, 1998) considers Laboulbenia stenolophi a synonym of

L. anoplogenii. Other authors discriminate two species based on morphological

characters (e.g., Terada 2001, Terada et al. 2004). Terada (2001) maintains that

Anoplogenius is the only host genus for L. anoplogenii, and that previous records

of L. anoplogenii from other hosts represent misidentifications. However, we

observed subdivisions of cell IV, supposedly characteristic for L. anoplogenii

on Anoplogenius (sensu Terada 2001) on three Roland Thaxter slides — from

Abacetus spp.: FH 00313469 (from Sri Lanka) and FH 00313470 (from the

Bengal region) and from Stenolophus sp.: FH 00313471 (from Massachusetts,

USA). Thaxter’s slide FH 00313468 (from Stenolophus sp. collected in Nantes,

France) shows two mature thalli without transverse septa dividing cell IV. We

share Santamaria’s (1989, 1998) opinion that L. anoplogenii and L. stenolophi are

synonyms, taking into account some degree of variability of the subdivisions of

cell IV.

The species-rich genus Clivina is known as a host to several species of

Laboulbeniales: Dixomyces clivinae (Thaxt.) I.I. Tav., D. pallescens (Thaxt.) LI.

Tav., Laboulbenia clivinalis Thaxt., L. schizogenii Thaxt., L. timurensis T. Majewski

& K. Sugiy., Ormomyces clivinae, and Peyritschiella clivinae Thaxt. Clivina may

also be an accidental host for L. anoplogenii (“nebenwirt”; Scheloske 1969),

since it occupies the same habitat as the parasite’s typical hosts (Santamaria

1998). The first record of L. anoplogenii on Clivina, an obvious consequence of

an accidental infection (S. Santamaria pers. comm.), was found in the Natural

Park of El Fondo, Spain, on a single specimen of C. ypsilon (Balazuc et al. 1983,

Santamaria 1989).

Laboulbenia cafii Thaxt., Proc. Amer. Acad. Arts 35: 162 (1899)

NEW RECORD: SINGAPORE, East REGION, Changi, on Remus corallicola

(Staphylinidae, Staphylininae), no date, leg. Malcolm Cameron, D. Haelew. 303, in coll.

American Museum of Natural History, slide FH 00313440 (1 thallus collected from left

metafemur).

PREVIOUS RECORDS: Laboulbenia cafii is known from America, Europe, Asia, and

Oceania (Santamaria 1998). In Asia, it has been reported from Hong Kong (Thaxter

1908), India (Balazuc collection, in Santamaria et al. 1991), and Japan (Sugiyama 1973).

Hosts are species belonging to the genus Cafius and related genera.

444 ... Haelewaters & Yaakop

Laboulbenia erotylidarum Haelew., sp. nov. PLATE 2A,B

MycoBank MB808169

Differs from all other Laboulbenia species on Erotylidae by its blackening of the basal

outermost part of the outer appendage and the posterior margin of the perithecium

immersed in the receptacle for about 3/4 of its length.

Type: The Philippines, Mindanao Island, Zamboanga Peninsula administrative region,

on “erotylid” [sic] (family Erotylidae), no date, no collector, Thaxter 3051 (Holotype,

FH 00313483 (slide, 1 mature thallus); isotype, FH 00313484 (slide, 1 mature thallus

collected from the elytron)).

Erymo.tocy: Named after the host family.

THALLuS 212-265 um long from foot to perithecial tip, pale greyish-brown in

color with only the distal 3/4 of cell I and perithecium darker. Cetts I, II, IU,

and IV of similar length, 2-2.5x longer than broad. Cell II with subparallel

margins. CELL V wedge-shaped, slightly protruding between the perithecium

and the insertion cell; located in the inner-upper corner of cell IV, which is

about twice as long as cell V. Septum IV-V oblique, strongly curved. INSERTION

CELL oblique, very dark, marking a strong constriction on both sides above

cells IV and V, located at the distal quarter of the perithecium but separated

from it. OUTER APPENDAGE consisting of up to four hyaline branches of

gradually more elongate cells, resulting from up to two successive dichotomies,

the first of which occurring above the large, irregular basal cell; the basal cells,

the outermost suprabasal, and the following cells of the outermost branch

are externally blackened. INNER APPENDAGE consisting of a basal cell that

is broader than long, carrying two suprabasal cells, the inner of which gives

rise to a single branch, the outer one rounded, large, giving rise to two simple

branches; all branches composed of gradually more elongate cells. ANTHERIDIA

not observed. Longest outer appendage 205 um, longest inner appendage 220 um.

CELL VI trapezoidal, 1.5x longer than broad. PERITHECIUM 111-118 x 38-42 um,

fusiform, hardly inflated; the apex asymmetrical, with very prominent and

rounded posterior lips, each of which bearing a conspicuous, rounded papilla;

pre-ostiolar spots black, sub-opaque, both more or less merging by a pre-apical

shading; the ostiole hyaline. AscosporEs 64-72 x 4.7-5.6 um.

Six species of Laboulbenia are previously reported from Erotylidae: L. parvula

Thaxt. (but see discussion below); L. scaphidomorphi Speg. on Scaphidomorphus

bosci from Panama (type), Bolivia, Brazil, Ecuador, Paraguay, and Peru

(Spegazzini 1915, Barragan et al. 2013); L. nesitidis Balazuc on Nesitis sexnotata

from Peninsular Malaysia (Balazuc 1975); L. encaustis K. Sugiy. & T. Majewski

on Encaustis praenobilis from Ecuador (Sugiyama & Majewski 1987); L. skelleyi

W. Rossi & Bergonzo on Pselaphacus spp. from Brazil, Costa Rica, and Ecuador

(Rossi & Bergonzo 2008; Barragan et al. 2013); and L. mycotreti W. Rossi on

Mycotretus spp. from Ecuador (Rossi 2011).

Laboulbenia spp. nov. (Philippines) ... 445

Piate 2. Laboulbenia erotylidarum. A. Habitus (isotype, FH 00313484). B. Detail of inner

appendage with three (1-3) branches and outer appendage with four (a-d) branches (holotype,

FH 00313483). Laboulbenia poplitea. C. Habitus, with the outer appendage more than doubling

the full thallus length (holotype, FH 00313480). D. Detail of the perithecial apex, with the focus

on one papilla (arrow). E. Detail of the single antheridium (arrow) (isotype, FH 00313481).

Scale bars: A, C = 100 um; B = 50 um; D, E = 25 um.

Unlike L. erotylidarum, both L. scaphidomorphi and L. encaustis have the

insertion cell joined to the posterior margin of the perithecium (Spegazzini

1915, Sugiyama & Majewski 1987). Laboulbenia mycotreti differs from the new

species by its stout habitus, the unbranched outer and inner appendages, and

the exceptional position of cell VI (Rossi 2011).

446 ... Haelewaters & Yaakop

Laboulbenia erotylidarum, L. nesitidis, and L. skelleyi share having cell

V protrude between the perithecium and the insertion cell. Laboulbenia

skelleyi, however, differs in its unbranched outer appendage, a bifurcate inner

appendage, and a much shorter and stouter general appearance (Rossi &

Bergonzo 2008). Laboulbenia nesitidis is distinguished by a perithecium that

is free for more than half of its length, a different branching pattern, and a

different pigmentation of the appendages (Balazuc 1975).

Colla (1926) reported L. parvula also from Erotylidae but provided no

description or illustration. We agree with Rossi & Bergonzo (2008) that the

record of L. parvula on Brachysphaenus bimaculatus (Colla 1926) is doubtful.

This parasite was described and is known from Carabidae (subfamilies

Harpalinae and Trechinae). Laboulbenia parvula obviously differs from

L. erotylidarum in its largely free perithecium, a cell VI that is broader than

long, and the differently structured appendages, the outermost with more or

less darkened lower cells (mainly at their posterior margins).

We retain the species name written on the slide labels by Roland Thaxter.

Laboulbenia poplitea Haelew., sp. nov. PLATE 2C-E

MycoBank MB808170

Differs from other Laboulbenia species by its kinked cell II posterior margin, its

perithecium covering most of cell III, and cells III and IV posterior margins parallel to

the perithecial anterior margin.

Type: The Philippines, Luzon Island, Metropolitan Manila, Manila, on Craspedophorus

sp. (Carabidae, Harpalinae, Panagaeini), Jan 1912, no collector, Thaxter 2549 (Holotype,

FH 00313480 (slide, 4 mature thalli; isotype, FH 00313481 (slide, 3 mature thalli)).

EryMo.oey: From the Latin noun popliteus = back of the knee joint, referring to the

angulated posterior margin of cell II.

THALLUS 195-212 um long from foot to perithecial tip, light brown in color

with darker cell VI and perithecium. Cetts I and II forming a pedicel; the

former rectangular, 44 x 20 um; the latter abruptly broadening upwards, distally

up to 41 um wide, its posterior margin kinked and longer (38-52 um) than the

(straight) anterior margin (30-44 um). CELL III rectangular, 47-56 um long,

hardly to distinguish from the body of the perithecium. CELL V wedge-shaped,

distal half separated from perithecial wall, located in inner-upper corner of

CELL IV, which is elongated, 39-44 um long, on average 2.4x longer than cell

V. INSERTION CELL dark and thick, marking a constriction above cells IV and

V, located at height of preostiolar spots of perithecium, but separated from it.

OUTER APPENDAGE simple, long and slender, up to 320 um in length, consisting

of a rectangular basal cell of about 19 um long, followed by one to three cells

of similar length; the other cells forming the appendage more elongate. INNER

APPENDAGE consisting of a small, subtriangular basal cell, reaching only 2/3

of the length of the basal cell of the outer appendage and bearing distally one

Laboulbenia spp. nov. (Philippines) ... 447

elongate cell that produces a slender antheridium. With age, the antheridium

degenerates and elongates as a simple evanescent branch. Ceti VI broader

than long, wedge-shaped, its anterior side measuring 14-17 um. PERITHECIUM

95-106 x 37-42 um, asymmetrical, the anterior margin nearly straight, the

posterior margin strongly convex, broadest at 1/3, tapering upwards; the

apex asymmetrical, ending in three rounded lips, the anterior being distinctly

shorter; papillae conspicuous on anterior and posterior lips; the ostiole hyaline,

very contrasting to the pre-ostiolar spots, the anterior of which is more reduced

to a longitudinal stria. AscosporREs 51-63 x 3.2-4.7 um.

Laboulbenia poplitea is recognized by the following characteristics: 1) the

kinked posterior margin of cell I, giving it the look of the back of the knee joint,

2) cell III covered by the body of the perithecium, and 3) the posterior margins

of cells III and IV parallel to the anterior perithecial margin. In addition, the

outer appendage is unbranched and can become very long, contributing to a

total thallus length (from foot to tip of the outer appendage) up to 0.5 mm.

In many Laboulbenia species damaged appendages are known to regenerate

atypically. Yet, among all examined specimens of L. poplitea, we found no

thalli with abnormal regeneration; two thalli with a normally regenerated

— unbranched — outer appendage were observed.

Laboulbenia poplitea bears a superficial resemblance to L. erecta Thaxt.,

parasitic on Platynus spp. [as Colpodes] (Carabidae, Harpalinae, Platynini)

from Mexico, which, however, has a straight cell II, an inner appendage with

the suprabasal cell producing two antheridia, and an outer appendage always

branched above the suprabasal cell and with sometimes an additional branch

arising from the basal cell anterior side (Thaxter 1899, Haelewaters, pers. obs.).

In addition and in contrast to L. poplitea, the perithecium of L. erecta does not

cover cell III but has an almost symmetrical profile.

Craspedophorus is a senior synonym of Brachyonychus, which Roland Thaxter

wrote [misspelled as “Brachionychus”| on the slide labels. Craspedophorus,

an Old World genus, is very speciose. Only three species of Laboulbeniales

have been reported on this host genus: Laboulbenia brachyonychi Thaxt.,

L. proliferans Thaxt., and L. taiwaniana Terada et al.

Laboulbenia proliferans parasitizes carabid beetles representing several

subfamilies and tribes (e.g., Brachinini, Licinini, Callistini, Panagaeini) from

Europe, Asia, Africa, and Oceania (Arndt & Santamaria 2004) but differs

from L. poplitea in many respects, e.g. by the proliferation of cell V (Thaxter

1893). Also L. brachyonychi (Thaxter 1899) is very different morphologically,

with a wholly free slender perithecium, the androstichum (cells II, IV, and V)

forming a dark pigmented stalk separated from the perithecium, and both the

outer and inner appendage simple and elongate (the inner one may be branched

448 ... Haelewaters & Yaakop

PLaTE 3. A. Dimeromyces rugosus (FH 00313485), mature male thallus (m) and female thallus

(f), showing transverse striae at the perithecial wall (area between dashed lines) and two

papillae at opposite sides of the perithecial apex (arrow). B. Monoicomyces stenusae (FH 2535).

C. Laboulbenia satanas (FH 00313478). D. Laboulbenia timurensis, arrow showing the non-

pigmented, lens-shaped insertion cell, above which occur subdivisions of the appendage basal cells

(FH 00313449). E. Ormomyces clivinae, showing ascospores oozing out of the perithecium two by

two (FH 00313445). Scale bars: A = 50 um; B-E = 100 um.

one or twice). More recently, Terada et al. (2008) described L. taiwaniana on

Craspedophorus formosanus from Taiwan, which is distinguished by the black

septum between the basal and suprabasal cell of the inner appendage, next to

the elongate perithecium and cell V subequal in length to cell IV.

Laboulbenia spp. nov. (Philippines) ... 449

Laboulbenia satanas Balazuc, Rev. Mycol. 37: 261 (1973) PLATE 3C

SPECIMENS EXAMINED: THE PHILIPPINES, Luzon IsLaAnp, Lacuna PRov., Mount

Makiling, on “gyrinid beetle” (Gyrinidae), no date, leg. Charles E Baker, Thaxter 3244,

slides FH 00313475 (12 thalli), FH 00313476 (4 thalli), FH 00313477 (26 thalli), FH

00313478 (11 thalli), and FH 00313479 (14 thalli).

PREVIOUS RECORD: The only previous report is from the Philippines (Balazuc 1973), on

Orectochilus discus (Gyrinidae).

Some new Laboulbenia species that Thaxter intended to characterize in the

never-finished sixth volume of his monumental monograph have now been

described by other scientists (Haelewaters & Rossi 2014). Laboulbenia satanas

parasitizing Gyrinidae is one. Thaxter had provisionally named “his” new

species L. “auriculata,” but Balazuc (1973) formally described the same species

as L. satanas.

The combination of cell V forming an elongated lobe along the posterior

margin of the perithecium and the horn-like outgrowths at the perithecial apex

is unlike any other species in the genus. Laboulbenia bicornis Thaxt. shares the

slender (olive-)brown horn-like processes, yet these processes are differently

shaped in both species (see Thaxter 1908, Plate LXVII, Figs. 1-2). In addition,

L. bicornis is much more slender and elongated (780-950 um) than L. satanas

(250-390 um). Laboulbenia fallax Thaxt. (on Gyretes spp.) and L. rotundata

Thaxt. (on Dineutes spinosus) also parasitize members of the Gyrinidae. Both

species share with L. satanas the unusual development of cell V, but lack

any differentiation at the perithecial tip (except for two minute tooth-like

projections in L. rotundata).

Laboulbenia timurensis T. Majewski & K. Sugiy., Trans. Mycol. Soc. Japan 27: 436

(1987) PLATE 3D

SPECIMENS EXAMINED: PAPUA NEW GUINEA, MILNE Bay Prov., Modeway Bay, on

Clivina inopaca (Carabidae, Scaritinae), 2 Feb 1956, leg. Leonard J. Brass, det. Philip J.

Darlington, D. Haelew. 308, in coll. American Museum of Natural History, slides FH

00313447 (4 thalli from left elytron), FH 00313448 (1 thallus from right mesotibia), FH

00313449 (3 thalli from left metatibia), FH 00313450 (6 thalli from legs), FH 00313451

(1 thallus from left mesotibia), FH 00313452 (2 thalli from right elytron), and FH

00313453 (4 thalli from right elytron).

PREVIOUS RECORDS: Described on Clivina sp. (epippiata group) from Indonesia

(Borneo); afterwards recorded only once, on Clivina yanoi from Taiwan (Majewski &

Sugiyama 1986, Terada et al. 2004).

The studied thalli correspond morphologically with the figures of L. timurensis

in Terada et al. (2004). We agree with their suggestion that “the division [into

a mass of small cells] seems to be in the appendage above the insertion cell,”

which is undivided and lens-shaped. The parasites were found on both the legs

and elytra of the host.

450 ... Haelewaters & Yaakop

Monoicomyces stenusae Thaxt., Proc. Amer. Acad. Arts 51: 30 (1915) PLATE 3B

EXAMINED SPECIMEN: THE PHILIPPINES, Luzon ISLAND, METROPOLITAN MANILA,

Manila, on Silusa sp. (Carabidae, Scaritinae), Dec 1911, no collector, Thaxter 2417, slide

FH 00313466 (1 thallus collected from the elytron).

PREVIOUS RECORDS: Described from Neosilusa ceylonica [as Stenusa in Thaxter 1915; as

Silusa in Thaxter 1931] (Staphylinidae, Aleocharinae) from Indonesia (Java). Afterwards

reported only once, on “Silusa kamerunensis” from Cameroon (Thaxter 1915, 1931).

Majewski (1988) found Monoicomyces plagiusae Thaxt. in Japan and suggested

that M. plagiusae and M. stenusae are synonyms. Before Majewski (1988),

Monoicomyces plagiusae was known only from the type locality in Indonesia

(Sumatra) (Thaxter 1931).

A single thallus mounted by Thaxter was studied. The available specimen

corresponds most to the description for Monoicomyces stenusae. The two

secondary axes on either side of the suprabasal cell are proliferated, consisting

of three cells, the latter of which gives rise to two antheridia and a perithecium.

Thaxter’s (1915) original description mentions that the third cell is “usually

terminated by two antheridia.” Also, our measurements differ significantly

from Thaxter’s measurements of thalli from Java (Indonesia) and Cameroon;

especially the cells VI (up to 114 x 32 um) and the perithecia (144-156 x

41-47 um) are considerably larger than reported by Thaxter himself.

Considering Thaxter’s (1915, 1931) suggestion of prominent morphological

variation within this species, we think it is safe to identify the Philippine thallus

as M. stenusae, until more material becomes available. Moreover, this thallus

might represent another form, restricted to the elytra, whereas Thaxter (1915,

1931) mentions only smaller, compact forms taken from the legs and “more

developed” ones from the abdomen.

Ormomyces clivinae (Thaxt.) I.I. Tav., Mycol. Memoir 9: 266 (1985) PLATE 3E

NEW RECORD FROM INp1A: INDIA, WEsT BENGAL STATE, Kanchrapara, on Clivina

sp. (Carabidae, Scaritinae), 9 Jul 1944, leg. Mont A. Cazier, D. Haelew. 306, in coll.

American Museum of Natural History, slides FH 00313443 (2 thalli from right elytron),

FH 00313444 (2 thalli from distal tip abdomen), and FH 00313445 (4 thalli from right

elytron).

PREVIOUS RECORDS: Recorded on Clivina spp. from Indonesia (type), Madagascar,

Sierra Leone, and Cameroon (Balazuc 1982, Rossi 1982, Tavares 1985).

Ormomyces I.I. Tav. was erected to accommodate a single species, O. clivinae,

which Thaxter (1915) had earlier placed in Misgomyces. The species is related

to Ecteinomyces trichopterophilus Thaxt., which shares the uniseriate receptacle.

However, E. trichopterophilus differs from O. clivinae in its cell III having a

corner cell (which may give rise to a secondary branch), a perithecium with long

Laboulbenia spp. nov. (Philippines) ... 451

narrow neck, and in having different hosts (Ptiliidae, Acrotrichinae: Acrotrichis

spp., Baeocrara variolosa) (Tavares 1985, Haelewaters et al. 2014b). Notable is

that E. trichopterophilus has been collected in North and South America and

Europe (Haelewaters et al. 2014b), while O. clivinae has so far been recorded

only in southeastern Asia and Africa. Future collections will elucidate whether

or not the geographical distributions of the two species overlap.

A thorough literature review of all Laboulbeniales reported from India

yielded only 32 species (Thaxter 1896, 1899, 1900, 1901, 1902, 1908, 1915,

1926; Batra 1963; Kaur et al. 1993; Kaur & Mukerji 1995, 1996a,b; Pathak &

Mukerji 1997; present paper). However, since India is considered one of the

17 “megadiverse” countries in the world, with the Western Ghats mountain

range recognized as a biodiversity hotspot (Myers et al. 2000), we anticipate

that many more species of Laboulbeniales will be found in this country.

Peyritschiella princeps (Thaxt.) I.1. Tav., Mycol. Memoir 9: 270 (1985)

SPECIMENS EXAMINED: INDONESIA, LomBok, West Nusa TENGGARA PROV.

Sapit village, on Philonthus tardus (Staphylinidae, Staphylininae), Apr 1896, leg. Hans

Fruhstorfer, D. Haelew. 304, in coll. American Museum of Natural History, slides FH

00313441 (1 thallus from pronotum) and FH 00313442 (1 thallus from left-hand side

tergite).

PREVIOUS RECORDS: Described as Dichomyces princeps on Philonthus sordidus

(Staphylinidae, Staphylininae), Massachusetts, U.S.A. Peyritschiella princeps is known

from all continents except Antarctica, attacking beetles in the genera Quediomacrus,

Spatulonthus, and Philonthus (Santamaria et al. 1991).

Acknowledgments

Thanks are due to Lee Herman, Curator Emeritus at the American Museum of

Natural History (Invertebrate Zoology) for help and support. We thank Walter Rossi

and Sergi Santamaria for reviewing the manuscript and Rosanne Healy for assistance

with the plates. Lastly, Shaun Pennycook is acknowledged for valuable suggestions on

nomenclature and improvements of the manuscript.

The authors acknowledge the following grants: a Collection Study Grant and the

Theodore Roosevelt Memorial Grant from the American Museum of Natural History

(DH) and the Industry Grant-2013-030 from Universiti Kebangsaan Malaysia (SY).

Literature cited

Arndt E, Santamaria S. 2004. Laboulbeniales (Ascomycota) of the Canary Islands. Vieraea 32:

107-115.

Balazuc J. 1973. Recherches sur les Laboulbéniomycetes. I. Trois especes nouvelles et une malconnue.

Revue de Mycologie 37(5): 253-262.

Balazuc J. 1982. Laboulbeniales (Ascomycetes) de Madagascar, des Comores et des Mascareignes.

Bulletin Mensuel de la Société linnéenne de Lyon 51(1): 209-219.

Balazuc J, Espadaler X, Girbal J. 1983. Laboulbenials (Ascomicets) Iberiques, II. Noves aportacions.

Collectanea Botanica (Barcelona) 14: 39-42.

452 ... Haelewaters & Yaakop

Barragan A, Bernardi M, Rossi W. 2013. New records of Laboulbenia (Fungi, Ascomycota) from

Ecuador and other countries. Webbia 68(1): 25-34.

http://dx.doi.org/10.1080/00837792.2013.779816

Batra SWT. 1963. Some Laboulbeniaceae (Ascomycetes) on insects from India and Indonesia.

American Journal of Botany 50(10): 986-992. http://dx.doi.org/10.2307/2439905

Blackwell M. 2011. The Fungi: 1, 2, 3 ... 5.1 million species? American Journal of Botany 98(3):

426-438. http://dx.doi.org/10.3732/ajb.1000298

Castaldo D, Rossi W, Sabatini F. 2004. Contribution to the knowledge of the Laboulbeniales from

Greece. Plant Biosystems 138(3): 261-269. http://dx.doi.org/10.1080/11263500400006969

De Kesel A, Haghebaert G. 1991. Laboulbeniales (Ascomycetes) of Belgian Staphylinidae

(Coleoptera). Bulletin de la Société Royale Belge d’Entomologie 127: 253-270.

Haelewaters D, De Kesel A. 2013. A new species of Cantharomyces (Laboulbeniales, Ascomycota)

from the Netherlands. Mycotaxon 123: 467-472. http://dx.doi.org/10.5248/123.467

Haelewaters D, Rossi W. 2015 (in press). Three new species of Laboulbenia from Roland Thaxter’s

backlog of slides and a brief review of Laboulbeniales associated with Chrysomelidae. Mycologia

107(1). http://dx.doi.org/10.3852/14-022

Haelewaters D, Schilthuizen M, Pfister DH. 2014a. On Diphymyces (Laboulbeniales, Ascomycota) in

Malaysian Borneo. Plant Ecology and Evolution 147: 93-100.

http://dx.doi.org/10.5091/plecevo.2014.912

Haelewaters D, Vorst O, De Kesel A. 2014b. New and interesting Laboulbeniales (Fungi, Ascomycota)

from the Netherlands. Nova Hedwigia 98(1-2): 113-125.

http://dx.doi.org/10.1127/0029-5035/2013/0150

Huldén L. 1983. Laboulbeniales (Ascomycetes) of Finland and adjacent parts of the U.S.S.R.

Karstenia 23(2): 31-136.

Huldén L. 1985. Floristic notes on Palearctic Laboulbeniales (Ascomycetes). Karstenia 25(1): 1-16.

Irmler U. 2003. Taxonomy and distribution of the Neotropical species of the genera Tannea

Blackwelder, 1952 and Nacaeus Blackwelder, 1942 with remarks on the genus Lispinus

(Coleoptera: Staphylinidae). Bulletin de Institut Royal des Sciences Naturelles de Belgique 73:

85-134.

Juan L-Y, Chien C-Y. 1995. Study on the Laboulbeniales (Ascomycetes) of Taiwan. Biological

Bulletin of National Taiwan Normal University 30(1): 11-22.

Kaur S, Mukerji KG. 1995. Studies on Indian Laboulbeniales IV: Three species of Laboulbenia.

Mycoscience 36: 311-314. http://dx.doi.org/10.1007/BF02268606

Kaur S, Mukerji KG. 1996a. Studies on Indian Laboulbeniales II. Three unrecorded species. Nova

Hedwigia 62 (1-2): 151-156.

Kaur S, Mukerji KG. 1996b. Studies on Indian Laboulbeniales UI. Three unrecorded dioecious

genera. Mycoscience 37: 61-64. http://dx.doi.org/10.1007/BF02461458

Kaur S, Pathak A, Mukerji KG. 1993. Studies on Indian Laboulbeniomycetes 1. Three unrecorded

species of the genus Laboulbenia Mont. et Robin. Cryptogamic Botany 3(4): 357-360.

Kirk PM, Cannon PF, Minter DW, Stalpers JA. 2008. Ainsworth and Bisby’s Dictionary of the Fungi

(10th Edition). CSIRO Publishing. 771 p.

Lee Y-B, Kim K-T, Lim C-K. 2002. Interesting species of the Laboulbeniales from Upo Swamp.

Mycobiology30(3): 128-132. http://dx.doi.org/10.4489/MYCO.2002.30.3.128

Maire R. 1920. Troisieme contribution a étude des Laboulbéniales de l'Afrique du Nord. Bulletin de

la Société d'Histoire Naturelle de l'Afrique du Nord 11(8): 123-138.

Majewski T. 1990. Rare and new Laboulbeniales from Poland. X. Acta Mycologica 23 (2): 97-108.

Majewski T. 1994. The Laboulbeniales of Poland. Polish Botanical Studies 7: 1-466.

Laboulbenia spp. nov. (Philippines) ... 453

Majewski T, Sugiyama K. 1986. Notes on the Laboulbeniomycetes (Ascomycotina) of Borneo IV.

Transactions of the Mycological Society of Japan 27: 425-439.

Middelhoek A. 1949. Laboulbeniaceae in Nederland III. Nederlands Kruidkundig Archief 56:

249-260.

Myers N, Mittermeier RA, Mittermeier CG, da Fonseca GAB, Kent J. 2000. Biodiversity hotspots

for conservation priorities. Nature 403: 853-858. http://dx.doi.org/10.1038/35002501

Naomi S-I. 1997. A revision of the genus Nacaeus Blackwelder from Japan. Japanese Journal of

Entomology 65(1): 127-142.

Pathak A, Mukerji KG. 1997. Studies on Indian Laboulbeniales, VIII. Two new species.

Phytomorphology 47(3): 333-337.

Rossi W. 1982. Laboulbeniali della Sierra Leone (Ascomycetes). Accademia Nazionale dei Lincei,

Quaderno 255: 9-22 + Plates I-IV.

Rossi W. 2011. New species of Laboulbenia from Ecuador, with evidence for host switch in the

Laboulbeniales. Mycologia 103(1): 184-194. http://dx.doi.org/10.3852/10-117

Rossi W, Bergonzo E. 2008. New and interesting Laboulbeniales from Brazil. Aliso 26: 1-8.

Rossi W, Santamaria S. 2012. Rodaucea, a new genus of the Laboulbeniales. Mycologia 104(3):

785-788. http://dx.doi.org/10.3852/11-337

Rossi W, Maca J, Vavra J. 2010. New records of Laboulbeniales (Ascomycota) from the Czech

Republic and Slovakia. Polish Botanical Journal 55(2): 343-351.

Santamaria S. 1989. El orden Laboulbeniales (Fungi, Ascomycotina) en la Peninsula Ibérica e Islas

Baleares. Edicions Especials de la Societat Catalana de Micologia 3: 1-396.

Santamaria S. 1998. Laboulbeniales, 1. Laboulbenia. Flora Mycologica Iberica 4: 1-186.

Santamaria S. 2003. Laboulbeniales, Il. Acompsomyces-Ilyomyces. Flora Mycologica Iberica 5:

1-344.

Santamaria S, Balazuc J, Tavares II. 1991. Distribution of the European Laboulbeniales (Fungi,

Ascomycotina). An annotated list. Treballs de l'Institut Botanic de Barcelona 14: 1-123.

Scheloske H-W. 1969. Beitrage zur Biologie, Okologie und Systematik der Laboulbeniales

(Ascomycetes) unter besondere _ Beriicksichtigung des Parasit-Wirt-Verhaltnisses.

Parasitologische Schriftenreihe 19: 1-176.

Shen Y-H, Ye D-H, Li T-H, Song B, Zhang A-L, Tian M-Y. 2006. Laboulbeniales. Flora Fungorum

Sinicorum 28: 1-294, Plates I-II.

Siemaszko J, Siemaszko W. 1928. Owadorosty polskie i palearktyczne. Polskie Pismo

Entomologiczne 6: 188-211, Plate VII.

Siemaszko J, Siemaszko W. 1932. Owadorosty polskie i palearktyczne, II. Polskie Pismo

Entomologiczne 10: 149-188, Plates VII-X.

Spegazzini C. 1915. Segunda contribution al conocimiento de las Laboulbeniales italianas. Anales

del Museo Nacional de Historia Naturel de Buenos Aires 27: 37-74.

Sugiyama K. 1973. Species and genera of the Laboulbeniales (Ascomycetes) in Japan. Academia

Scientific Book Inc. Tokyo. 97 p. + 27 Plates

Tavares II. 1985. Laboulbeniales (Fungi, Ascomycetes). Mycologia Memoir 9: 1-627.

Terada K. 1976. Some species of the Laboulbeniales from Taiwan. Transactions of the Mycological

Society of Japan 17: 23-34.

Terada K. 2000. New records of the carabidicolous Laboulbeniales (Ascomycetes) of Japan (III).

Mycoscience 41: 39-48. http://dx.doi.org/10.1007/BF02464384

Terada K. 2001. Notes on Laboulbenia stenolophi and Laboulbenia anoplogenii (Ascomycetes,

Laboulbeniales). Mycoscience 42: 1-9. http://dx.doi.org/10.1007/BF02463969

Terada K, Hsu M-H, Wu W-J. 2004. Notes on the carabidicolous Laboulbeniales (Ascomycetes) of

Taiwan I. Botanical Bulletin of Academia Sinica 45(1): 87-94.

454 ... Haelewaters & Yaakop

Terada K, Hsu M-H, Wu W-J. 2008. A new species of genus Laboulbenia (Laboulbeniales) on

Craspedophorus formosanus (Coleoptera, Carabidae) from Taiwan, with a note on Laboulbenia

asiatica. Aliso 26: 23-27.

Thaxter R. 1893. New species of Laboulbeniaceae from various localities. Proceedings of the

American Academy of Arts and Sciences 28: 156-188. http://dx.doi.org/10.2307/20020515

Thaxter R. 1896. Contribution towards a monograph of the Laboulbeniaceae. Memoirs of the

American Academy of Arts and Sciences 12(3): 187-429.

Thaxter R. 1899. Diagnosis of new species of Laboulbeniaceae. I. Proceedings of the American

Academy of Arts and Sciences 35(9): 153-209. http://dx.doi.org/10.2307/25129915

Thaxter R. 1900. Preliminary diagnosis of new species of Laboulbeniaceae. II. Proceedings of the

American Academy of arts and Sciences 35(21): 409-450. http://dx.doi.org/10.2307/25129954

Thaxter R. 1901. Preliminary diagnosis of new species of Laboulbeniaceae. IV. Proceedings of the

American Academy of Arts and Sciences 37(2): 21-45. http://dx.doi.org/10.2307/20021631

Thaxter R. 1902. Preliminary diagnosis of new species of Laboulbeniaceae. V. Proceedings

of the National Academy of Sciences of the United States of America 38(2): 9-57.

http://dx.doi.org/10.2307/20021736

Thaxter R. 1908. Contribution towards a monograph of the Laboulbeniaceae. Part II. Memoirs

of the American Academy of Arts and Sciences 13: 217-469, Plates XXVHI-LXXI.

http://dx.doi.org/10.2307/25058090

Thaxter R. 1915. New Indo-Malayan Laboulbeniales. Proceedings of the American Academy of

Arts and Sciences 51(1): 3-51. http://dx.doi.org/10.2307/20025560

Thaxter R. 1920. New Dimorphomyceteae. Proceedings of the American Academy of Arts and

Sciences 55(6): 211-282. http://dx.doi.org/10.2307/20025798

Thaxter R. 1926. Contribution towards a monograph of the Laboulbeniaceae. Part IV.

Memoirs of the American Academy of Arts and Sciences 15(4): 427-580, Plates I-XXIV.

http://dx.doi.org/10.2307/25058132

Thaxter R. 1931. Contribution towards a monograph of the Laboulbeniaceae. Part V.

Memoirs of the American Academy of Arts and Sciences 16: 1-435, Plates I-LX.

http://dx.doi.org/10.2307/25058136

Weir A, Beakes GW. 1993. New British Laboulbeniales. Mycological Research 97(9): 1045-1055.

http://dx.doi.org/10.1016/S0953-7562(09)80505-4

Weir A, Blackwell M. 2005. Fungal biotrophic parasites of insects and other arthropods. 119-145,

in FE Vega, M Blackwell (eds.), Insect-fungal associations: Ecology and evolution. Oxford

University Press, New York.

Weir A, Hammond PM. 1997. Laboulbeniales on beetles: Host utilization patterns and species

richness of the parasites. Biodiversity and Conservation 6: 701-719.

http://dx.doi.org/10.1023/A:1018318320019

MY COTAXON

http://dx.doi.org/10.5248/129.455

Volume 129(2), pp. 455-458 October-December 2014

Dictydiaethalium dictyosporangium sp. nov. from China

Bo ZHANG? & Yu Li'*

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

Jilin Agricultural University, 2888 Xincheng Street, Changchun City, P. R. China

* CORRESPONDENCE TO: yuli966@126.com

AsstraAct — Dictydiaethalium dictyosporangium, described as new, is characterized by a

branched pseudocapillitium and spores, which are 10-12 um in diam. and ornamented with

long ridges that sometimes form an incomplete banded reticulum. The new species, which

was collected on the bark of a dead log in Henan Province, is described and illustrated by

light and scanning electron micrographs. A key to three known Dictydiaethalium species is

also provided. The holotype is deposited in the Herbarium of Mycological Institute of Jilin

Agricultural University (HMJAU) in Changchun, China.

KEY WORDS — myxomycetes, taxonomy, Dictydiaethaliaceae

Introduction

Dictydiaethalium was established by Rostafinsky in 1873. Two species are

currently recognised in the genus (Kirk et al. 2008), of which only one has been

reported from China (Li & Li 1989). A specimen found on the bark of a dead

2013 differed from the two named species and is described here as new.

Materials & methods

Fruiting bodies and microscopic structures were examined by light and scanning

electron microscopes (Zhang & Li 2013). Permanent slides were prepared according

to Robbrecht (1974) by spreading a capillitium in a drop of 94% alcohol, determining

color after one minute, and then mounting in Hoyer’s medium (Martin & Alexopoulos

1969). Color terms are given according to Flora of British Fungi (Royal Botanic

Garden Edinburgh 1969). Morphological observations and measurements were made

using a stereomicroscope (20X) and optical microscope (400x and 1000x). About ten

pseudoaethalia, pseudocapillitia, and 20 spores were measured using a Leica DM100

microscope, and photographs were taken with a Canon G15 camera. For SEM images,

pseudoaethalia attached to a holder were coated with gold using a Hitachi E-1010 sputter

456 ... Zhang & Li

Pirate 1. Dictydiaethalium dictyosporangium (Isotype, HMJAU10256): 1-2. Fruiting

bodies; 3. Pseudocapillitium and spores (400x); 4. Pseudocapillitium and spores (1000).

Scale bars: 1 = 0.5 mm; 2 = 0.2 mm.

coating unit and examined with a Hitachi S-4800 scanning electron microscope at 10

kV at Changchun Institute of Applied Chemistry, Chinese Academy of Sciences. The

specimens are deposited in the Herbarium of Mycological Institute, Jilin Agricultural

University, Changchun, China (HMJAU).

Taxonomy

Dictydiaethalium dictyosporangium B. Zhang & Yu Li, sp. nov. PLATEs 1-2

MycoBank MB807341

Differs from Dictydiaethalium dictyosporum by its branched pseudocapillitia and from

D. plumbeum by its spores with long ridges sometimes forming an incomplete banded

reticulum.

TypE—China, Henan province, Sanmenxia city, Shiziping village, on the bark surface of

a dead log, 22 September 2013, Zhang Bo 100802 (Holotype, HMJAU10244; isotype,

HMJAU10256).

ETYMOLOGY—dictyosporangium (Latin) = with a branched pseudocapillitium.

SPOROPHORES a pseudoaethalium, at maturity simulating an aethalium,

effused, depressed, spreading over 22 mm, olivaceous to gray olivaceous,

pulvinate, irregular in outline, more or less circular, extending up to 2.2 cm,

up to 1.1 cm thick. HyPOTHALLUs shining, membranous, abundantly developed

Dictydiaethalium dictyosporangium sp. nov. (China) ... 457

A _ : ~

AccV SpotMaan De }—— Ace Spot Moan

1OORV 3.0 2439 SEP WO TS . 10.0R¥ 3.0 4806x) )SE

\ ;

PLATE 2. Dictydiaethalium dictyosporangium (Isotype, HMJAU10256), scanning electron

micrographs: 1-2. Partial pseudocapillitium and spores; 3. Branched pseudocapillitium;

4. Spores ornamented with long ridges, arranged in an incomplete banded reticulum.

and surrounding the pseudoaethalium. PrERipIum single, membranous,

translucent, slender and evanescent at the base, olivaceous brown in

transmitted light, smooth, persistent. COLUMELLA absent. CAPILLITIUM

absent. PSEUDOCAPILLITIUM filiform, flat, thick on one side, 3-5 um wide,

smooth expect for the thickened part which bears a row of warts, branched

and anastomosed, running down to the base of pseudoaethalia, yellowish

green to olivaceous green, pale yellow by transmitted light. Sporgs free,

bright yellowish green in mass, pale yellow to colourless by transmitted light,

10-12 um diam., ornamented with long ridges sometimes forming an

incomplete banded reticulum

Comments: Dictydiaethalium was previously reported with two species:

D. dictyosporum Nann.-Bremek. (Nannenga-Bremekamp 1966) and D. plumbeum

(Schumach.) Rostaf. (Lister 1894). Dictydiaethalium dictyosporum has

minutely banded-reticulate spores (ca. 9-12 um diam.) and unbranched and

thicker (ca. 8 um wide) pseudocapillitia, while D. plumbeum is diagnosed by

spinulose or warted spores (ca. 8-12 um diam.) and narrower (ca. 2 um wide)

pseudocapillitia.

458 ... Zhang & Li

Key to species of Dictydiaethalium

1. Spores reticulate, completely or incompletely banded ....................0000. 2

1. Spores spinulose or warted, not reticulate ....................004. D. plumbeum

2. Pseudocapillitia connected by threads ..................... D. dictyosporangium

2. Pseudocapillitia 16F COMME CIE”, ..2.%15 .24b estes thos ebb elleey D. dictyosporum

Acknowledgments

We express our deep appreciation to Professors A.J.S. Whalley and Shuanglin Chen

for their valuable revisions and kind help. This research was funded by grants from the

National Basic Research Program of China (3140010180).

Literature cited

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

10th ed. CAB International, Wallingford. 771 p.

Li Y, Li HZ. 1989. Myxomycetes from China. I: a checklist of myxomycetes from China. Mycotaxon

35(2): 429-436.

Lister A. 1894. A monograph of the Mycetozoa. British Museum, London. 224 p.

Martin GM, Alexopoulos CJ. 1969. The myxomycetes. University of Iowa Press. Iowa. 561 p.

Nannenga-Bremekamp NE. 1966. Notes on myxomycetes X. Some new species of Licea, Reticularia,

Cribraria, Dictydiaethalium, Trichia and Metatrichia. Proceedings of the Koninklijke

Nederlandse Akademie van Wetenschappen, Series C, 69(3): 336-349.

Robbrecht E. 1974. The genus Arcyria Wiggers in Belgium. Bulletin du Jardin Botanique National

de Belgique 44: 303-353. http://dx.doi.org/10.2307/3667676

Rostafinsky JT. 1873. Versuch eines Systems der Mycetozoen. Strassburg. 21 p.

Royal Botanic Garden Edinburgh. 1969. Flora of British fungi: colour identification chart.

Edinburgh, H.M. Stationery Office. 6 p.

Zhang B, Li Y. 2013 [“2012”]. Myxomycetes from China 16: Arcyodes incarnata and Licea retiformis,

newly recorded for China. Mycotaxon 122: 157-160. http://dx.doi.org/10.5248/122.157

MY COTAXON

http://dx.doi.org/10.5248/129.459

Volume 129(2), pp. 459-463 October-December 2014

Lophodermium quadrisporum sp. nov.

(Rhytismataceae) on Rhododendron faberi subsp. prattii

DAN-DAN Lu’, YAN-PING TANG’, LEI-HONG WANG’,

SHI-JUAN WANG?, & YING-REN LIN”

' School of Life Science &? School of Forestry & Landscape Architecture,

Anhui Agricultural University, West Changjiang Road 130, Hefei, Anhui 230036, China

*CORRESPONDENCE TO: yingrenlin@yahoo.com

AsBstTRAcT — Lophodermium quadrisporum, a new fungus found on fallen leaves of

Rhododendron faberi subsp. prattii from Sichuan Province, China, is described, illustrated,

and discussed. This taxon is distinguished from its closest relatives by the occurrence of both

4- and 8-spored asci. The type specimen is deposited in the Reference Collection of Forest

Fungi of Anhui Agricultural University, China (AAUF).

Key worps — foliicolous fungus, morphological character, taxonomy

Introduction

Lophodermium Chevall. is the largest genus of Rhytismataceae (Rhytismatales,

Leotiomycetes, Ascomycota; Kirk et al. 2008) and _ is distributed worldwide

(Cannon & Minter 1986, Lin et al. 2012). IndexFungorum (2014) lists 342

epithets for the genus, among which SpeciesFungorum (2014) accepts 172

Lophodermium species.

Here, a fungus on an ericaceous plant from Sichuan Province is described

and designated as a new species of Lophodermium.

Materials & methods

Material with mature ascomata was selected from the specimen eventually designated

as the holotype and described macroscopically under the dissecting microscope at

10-50 x magnification. After rehydration in water for ca 10 min, the fruitbodies were sliced

into 8—20 um thick sections with a freezing microtome (YD-202, China). Microscopic

features were examined in water, 5% KOH, Melzer’s reagent, cotton blue in water, or

lactophenol-cotton blue. Sections were mounted in 0.7% (w/v) cotton blue in water to

observe the outlines of ascomata in vertical section. Gelatinous sheaths surrounding

460 ... Lu & al.

ascospores and paraphyses were observed in water or 0.1% (w/v) lactophenol-cotton

blue. The color of internal structures and ascospore contents were determined in water.

For each specimen, at least 30 asci, ascospores, and paraphyses were measured in

5% KOH solution. Line and point integrated illustrations of the fruitbody external shape

and internal structures were prepared using the Panasoianic XSJ-2 microscope drawing

device.

Taxonomy

Lophodermium quadrisporum D.D. Lu & Y.R. Lin, sp. nov. FIGS 1-5

MycoBAnk MB 808815

Differs from Lophodermium rufum by its elliptical to lunate ascomata, grey lips,

branched and swollen or circinate paraphyses, and 4- and 8-spored asci.

Type: China, Sichuan, Ya’an, alt. ca 3550 m, on fallen leaves of Rhododendron faberi

subsp. prattii (Franch.) D.E Chamb, (Ericaceae), 21 June 2009, Y.G. Liu & Y.R. Lin 2765

(Holotype, AAUF 68873).

EryMoLoey: quadrisporum (Latin = 4-spored), referring to one of the two types of the

asci, containing four ascospores.

Co.tonies only epiphyllous, forming subround to irregular, yellow-white to

grayish-yellow bleached areas 3-5 mm diam., which tend to coalesce into

larger irregular shapes.

ZONE LINES frequent, dark brown to black, with various width, clearly

defined or sometimes diffused, entirely or partly surrounding the paler areas.

CONIDIOMATA not observed.

Ascomarta developing on the upper side of leaves, scattered in the bleached

spots. In surface view, ascomata 680-1440 x 220-540 um, elliptical or lunate,

straight or curved to one side, black-brown to black, slightly shiny, ends rounded,

obtuse or slightly acute, with a clearly marked outline, moderately rising the

substratum surface, opening by a single longitudinal split nearly extend to the

edge of the ascomata. Lips grey, sometimes valgus. In median vertical section,

ascomata subcuticular. COVERING STROMA well developed, 45-60 um thick

near the opening, gradually thinner towards the edge, connecting to the basal

stroma, mainly composed of textura angularis-epidermoidea with dark brown

to black thick-walled cells 3-7 um diam. Lip cells well developed, subhyaline,

4-7 x 2-3 um, thin-walled, cylindrical, 0—4-septate, more or less radially

arranged. BASAL STROMA Dlack-brown, comprised of 2-4 layers of thick-

walled, angular to aliform cells 3-6 um diam. SUBHYMENIUM well developed,

18—25 um thick, flat, consisting of colorless thin-walled textura angularis and

intricata. PARAPHYSES exceeding height of asci by 20-30 um, 1.5-1.8 um wide,

thin-walled, with septa which seem to only occur in the lower part of

the paraphyses, filiform, oftenbranched, sometimes circinate or gradually

swollen to 3—4 um above, covered with a ca 1 um thick gelatinous matrix. Asci

Lophodermium quadrisporum sp. nov. (China) ... 461

IF Ae

ERIS ‘¥ P pots spn

DOSNT AAI EP A ar aD

BS AORCH We Saeed ee caena = Sea eG

GLEE GIIESS Chey ele eas” very

ys s

Fics 1-5. Lophodermium quadrisporum (Holotype, AAUF 68873) on Rhododendron faberi subsp.

prattii. 1. Habit on a leaf. 2. Detail of ascomata and zone lines. 3. Ascoma in median vertical section.

4, Portion of ascoma in median vertical section. 5. Paraphyses, asci, and ascospores.

462 ... Lu & al.

ripening sequentially, thin-walled, cylindrical-clavate, short-stalked, apex

subacute or subtruncate-conical, without circumapical thickening, not bluing

in iodine, discharging spores through a small apical hole, of two-types: ca 30%

A-asci: 4-spored, 105-140 x 12-19 um; ca 70% B-asci: 8-spored, 110-160 x

15-20 um. 5-7-spored asci caused by spore abortion not observed. A-asci

ascospores 95-125 x 3-4.5 um, wide-filiform, often curved into a snaky

shape, tapered towards the acute base, containing bright oil drops and dull-

colored granules in the nodular upper half, aseptate, covered by a 0.8-2 um

thick gelatinous sheath. B-asci ascospores 100-145 x 2—2.5 um, filiform, slight

tapered to the base, arranged in a fascicle, colorless, continuous, smooth-

walled, covered with a gelatinous sheath ca 0.8 um thick.

HOsT SPECIES, HABITAT AND DISTRIBUTION: Producing ascomata on fallen

leaves of Rhododendron faberi subsp. prattii. Known only from the type locality,

Yaan, Sichuan Province, China.

ComMENTS — Lophodermium quadrisporum is very similar to L. rufum

Y.R. Lin & K. Lion Rhododendron maculiferum subsp. anhweiense (E.H. Wilson)

D.F. Chamb. in the way ascomata are embedded, dehiscence mechanism, and

ascal shape. However, L. rufum produces elliptical ascomata, reddish-brown

lips, heavily carbonized tissues in outer layer of the covering stroma, nearly

colorless textura angularis between the covering and basal stroma, uniformly

8-spored asci, and unbranched paraphyses (Xu et al. 2001).

Lophodermium pachychilum Y.R. Lin & Z.S. Xu is distinguished from the

new species by absent zone lines, wide-elliptical intra-epidermal ascomata

dehiscing by an elongate-elliptical split, a covering stroma composed of textura

angularis with pale thin-walled cells near the edge, a subhymenium consisting

of textura porrecta, and paraphyses agglutinated to form an epithecium above

the asci (Xu et al. 2001).

In addition, Lophodermium quadrisporum resembles Lophomerum ponticum

Minter on Rhododendron ponticum L. morphologically, but L. ponticum has

smaller ascomata (only 600 um long), much shorter and narrower asci (65-105 x

9-15 um), shorter paraphyses just as long as or slightly longer than the asci, and

shorter ascospores (45—80 um) aseptate or 1—3-septate (Minter 1980).

Acknowledgements

We are grateful to Dr Z. Wang (Yale University, USA) and Dr M. Ye (Hefei University

of Technology, China) for serving as pre-submission reviewers and to Dr Y.G. Liu

for the field investigation. This study was supported by the National Natural Science

Foundation of China (No. 31270065, 31170019).

Lophodermium quadrisporum sp. nov. (China) ... 463

Literature cited

Cannon PF, Minter DW. 1986. The Rhytismataceae of the Indian subcontinent. Mycological Papers

155.1223;

IndexFungorum. 2014. [www.indexfungorum.org (viewed online on 8 May 2014)].

Kirk PM, Cannon PF, Minter DW, Stalpers JA. 2008. Dictionary of the fungi, 10 ed. CAB

International. Wallingford. 771 p.

Lin YR, Liu HY, Hou CL, Wang SJ. 2012. Flora fungorum sinicorum, vol. 40, Rhytismatales

(in Chinese). Science Press. Beijing. 261 p.

Minter DW. 1980. Two species of Lophomerum on Rhododendron leaves. Transactions of the British

Mycological Society 74(1): 201-204. http://dx.doi.org/10.1016/S0007-1536(80)80030-1

SpeciesFungorum. 2014. [www.speciesfungorum.org (viewed online on 8 May 2014)].

Xu ZS, Li K, Lin YR, Xie YS. 2001. Two new species of Lophodermium Chev. on Anhwei

rhododendron. Journal of Anhui Agricultural University 28(4): 358-361.

MY COTAXON

http://dx.doi.org/10.5248/129.465

Volume 129(2), pp. 465-471 October-December 2014

Diploschistes xinjiangensis, a new saxicolous lichen

from northwest China

ABDULLA ABBAS™*, SHOU-YU GUO”,

GULIBAHAER ABABAIKELI’, ADILJIAN ABDULLA’, & HUERNISA XAHIDIN'*

* Arid land Lichen Research Center of Western China, College of Life Science and Technology,

Xinjiang University, Urumqi 830046, P. R. China

* State Key Laboratory of Mycology, Institute of Microbiology, Chinese Academy of Sciences,

Beijing 100101, PB R. China

* . .

CORRESPONDENCE TO: “ abdula209@sina.com * guosy@im.ac.cn

ABsTRACT — Diploschistes xinjiangensis from Xinjiang in northwest China is described as

new to science. The species is characterized by the thick bluish gray thallus, asci with eight

large ellipsoid ascospores, and the presence of diploschistesic and lecanoric acids. It grows on

rock in the semiarid region at elevations of ca. 1700 m. ITS rDNA sequence analyses support

the taxonomic distinctness of this species.

Key worps — Asia, biodiversity, Graphidaceae, Ostropales, taxonomy

Introduction

Diploschistes Norman includes crustose lichens with a blackish pseudo-

parenchymatous proper exciple, lateral paraphyses, and a_ trebouxioid

photobiont (Lumbsch 1989; Lumbsch & Mangold 2007). The genus is widely

distributed in arid and semiarid regions worldwide, with approximately

35-43 species known (Pérez-Vargas et al. 2012; Fernandez-Brime et al. 2013).

Diploschistes species occur mostly on rocks, some on soil, and a few rarely on

wood or bark (Lumbsch & Mangold 2007). The genus exhibits a remarkable

variability in ascomatal morphology, varying from perithecioid to urceolate

and lecanoroid (Lumbsch 1989; Lumbsch & Mangold 2007; Mangold et al.

2009). Despite this variation, the genus is currently regarded as monophyletic

and accommodated within the Graphidaceae based on molecular studies

(Martin et al. 2003; Frisch et al. 2006; Fernandez-Brime et al. 2013).

*'These authors contributed equally to this work.

466 ... Abbas, Guo & al.

The lichen biota of northwest China is rich, with more than 670 species and

127 genera so far reported (Abbas & Wu 1998; Guo 2005). Nevertheless, new

species continue to be discovered in this region, and knowledge of its lichen

diversity remains incomplete. Of the eight Diploschistes species recorded in

China (Wei 1991), three were described from the northwest region (Guo 2005).

An additional Diploschistes species recently collected by the first author in

Xinjiang, northwest China, is named here as D. xinjiangensis. We present a

taxonomic account based on its morphological and chemical characters and

assess the phylogenetic affinities of the new species from analyses of nrDNA

ITS sequences obtained from GenBank and two samples of the type specimen.

Materials & methods

The lichen specimens were collected from South Mountain in Urumdi, Xinjiang,

China, and are deposited at the Herbarium Mycologicum Academiae Sinicae-Lichenes

(HMAS-L) and the Lichen Section of Botanical Herbarium, Xinjiang University

(XJU). The morphology was examined using a Zeiss Stemi SV 11 stereomicroscope.

For microscopical examination, sections were cut by hand using a razor blade and

were mounted and observed in water. Anatomical structures and hymenial characters

were studied with a Zeiss Axioskop 2 plus light microscope and photographed using

a Nikon Digital Camera D50. Chemical constituents were identified by thin-layer

chromatography using solvent system C as outlined by Orange et al. (2010).

DNA EXTRACTION, AMPLIFICATION AND SEQUENCING. Two thallus fragments with

ascomata were sampled from the type specimen for DNA extraction. The DNA was

extracted using the DNAsecure Plant DNA Kit (Tiangen, China) following the

manufacturer's protocol. Amplification of the ITS region followed the methods described

in Martin et al. (2003) with modification. The whole ITS region (ITS1, 5.88, and ITS2)

of the nrDNA repeat tandem was targeted for the Polymerase Chain Reaction using the

primers ITS1 with ITS4 (White et al. 1990) directly. The amplification was performed in

a 25 uL volume containing 0.75 units of TransStart Taq Polymerase (Tiangen, China),

2.5 uL of its buffer, 0.5 uL of a 5 uM solution of the primers, 2 uL of 2.5 mM for each

dNTP solution, and 1 uL of genomic DNA. Thermocycling protocols: 95°C for 3 min

linked to 35 cycles at 94°C for 30 s, 54°C for 30 s, and 72°C for 1 min, with a final

extension of 72°C for 10 min. PCR products were screened on 1% agarose gels stained

with ethidium bromide. The PCR products were sequenced by Genewiz Inc. (Beijing).

Two newly obtained sequences were submitted to GenBank (see Fic. 1 for accession

numbers). The beginning and end of the ITS1 and ITS2 spacers were determined

by comparison with sequences available from GenBank (e.g., AJ458286, labelled as

Diploschistes gypsaceus, but actually D. rampoddensis according to Martin et al. 2003:

28). We excluded the 3’ end of the 18S gene (SSU), and the 5’ end of the 26S gene (LSU)

from the analyses. Our specimen sequences were aligned with the most similar taxa

represented by ITS sequences in GenBank (Fic. 1). The representative taxa were selected

based on their morphological characters, the results of Blast searches of sequence data,

and the literature (Martin et al. 2003; Fernandez-Brime et al. 2013).

Diploschistes xinjiangensis sp. nov. (China) ... 467

PHYLOGENETIC ANALYSIS AND SEQUENCE COMPARISONS. The ITS sequences of our

two samples and the 16 other reference sequences (including Thelotrema lepadinum

as outgroup) were aligned both by ClustalW and Muscle in MEGA 5 (Tamura et al.

2011). The alignment matrix was realigned by StatAlign for reliable measurement of the

accuracy of the results (Novak et al. 2008). The final matrix (submitted to TreeBase with

accession number $15164) can be obtained from the corresponding authors.

The evolutionary history was inferred both by using the Maximum Likelihood

method (ML) based on the Kimura 2-parameter model in MEGAS and using Bayesian

inference (PP) based on GTR model with rates = Invgamma. The analyses involved 18

nucleotide sequences. Absolute distances were also calculated in MEGAS, using the

number of base differences between sequence pairs, with all gaps removed from each

sequence pair.

Results & discussion

Phylogenetic analysis

The entire ITS region was successfully sequenced for the 2 type samples.

The sequence lengths for both samples = 499 bp for the entire ITS1 + 5.88 +

KC166981 Diploschistes neutrophilus SFB 44

KC166982 Diploschistes neutrophilus SFB 63

KC166983 Diploschistes neutrophilus SFB 118

KC166979 Diploschistes diacapsis SFB 2

KC166978 Diploschistes diacapsis SFB 1

KC167021 Diploschistes scruposus SFB 69

KC167020 Diploschistes scruposus SFB 66

KC167004 Diploschistes muscorum SFB 3

KC167008 Diploschistes muscorum SFB 140

KC167007 Diploschistes muscorum SFB 125

KJ000011 D. sp. nov. voucher Abbas 21s1

KJ000012 D. sp. nov. voucher Abbas 21s2

36/0.46 | 4J458286 Diploschistes rampoddensis Aptroot 39679

KC166993 Diploschistes rampoddensis SFB 101

92/YA" KC166992 Diploschistes rampoddensis SFB 68

KC167010 Diploschistes ocellatus SFB 16

AF227960 Diploschistes ocellatus AWA 1024

AF546077 Thelotrema lepadinum

92/0.97

————=!

0.05

FiGurRE 1. Phylogenetic relationships inferred from ITS sequences of Diploschistes xinjiangensis,

D. ocellatus, and species in the D. scruposus group (with Thelotrema lepadinum as outgroup).

Support is indicated for branches characterized by bootstrap frequencies exceeding 50% under

the Maximum Likelihood method, and posterior probabilities >0.95 from Bayesian Inference.

The tree is drawn to scale, with branch lengths measured in the number of substitutions per site.

Specimen collectors: AWA = Alan W Archer (Australia); Aptroot = André Aptroot (Netherlands);

SFB = Samantha Fernandez-Brime (Spain).

468 ... Abbas, Guo & al.

ITS2 region. The partial sequences containing the SSU 3’ end and LSU 5’ end

are included in the data submitted to GenBank (578 bp total). Some positions

(especially in the ITS2 region) in our sample sequences were difficult to align

with the reference sequences and were excluded from the matrix. There were

a total of 467 positions in the final dataset. All positions were used in the

phylogenetic analyses.

The ITS sequences indicate that D. xinjiangensis probably belongs to the

D. scruposus-group (sensu Martin et al. 2003) with close affinities to the

pantropical species, D. rampoddensis (93% identity and 2% gap) and

D. neutrophilus (93% identity and 3% gap). The evolutionary history was inferred

as the tree with the highest log likelihood (—1582.0654). In the phylogenetic

analyses, there was very strong support for the monophyly of D. xinjiangensis

(ML = 96%; PP = 0.96) and D. rampoddensis (ML = 86%; PP = 0.96), but

only weak support (ML = 56%) for the relationship between D. xinjiangensis

and a sister clade including D. diacapsis, D. muscorum, D. neutrophilus, and

D. scruposus (Fic. 1).

TABLE 1. Absolute distances for alignment sequences of ITS region (gaps ignored in

pairwise comparisons) between Diploschistes xinjiangensis and related species in

the D. scruposus group. Infraspecific distances are indicated with bold font.

1. D. xinjiangensis S1

Kj000011

2. D. xinjiangensis S2 2

KJo00012

cr re

eerie! = SD

Once «=H

eects 2s

Ks tf oN ie OS, RY

Oe C166978 ee eee Oe ce

C6702 pa gee eee

eo RC67020 scan) etalk pik

PP RCI67008 ee een

gs as it 16, NG 29:7 m= 9 Ole, BG ed re oa

15. D. muscorum

KC167007 Ve 17s ° 236-1238" 207 2 eT 8. 8 5 i) 1 3

Diploschistes xinjiangensis sp. nov. (China) ... 469

Absolute distances for the aligned sequences of the ITS region also support

the separation of a new species. In our sequence matrix, distances between

infraspecific samples are <5, while distances between species are =5 (TABLE 1).

SNS,

led,

FiGuRE 2. Diploschistes xinjiangensis (holotype). A: general habit; B: Asci and ascospores.

Scale bars: A = 5 mm; B = 20 um.

Taxonomy

Diploschistes xinjiangensis A. Abbas & S.Y. Guo, sp. nov. FIGURE 2

MycoBAnk MB 807412

Differs from Diploschistes rampoddensis by its thick bluish gray thallus, its 8-spored asci,

and its long broad ascospores.

Type: China. Xinjiang: Urumqi Co., South Mountain, Aketa, 43°22’N 86°48’E, alt.

1750 m, 3 Aug. 2011, A. Abbas 11821 (Holotype, HMAS-L; isotype, XJU; GenBank

KJ000011, KJ000012).

Erymo.oey: ‘The specific epithet xinjiangensis refers to the province where the type

specimen was collected.

THALLUS saxicolous, crustose, rimose-areolate, bluish grey to grayish white,

thick, <1.5 mm thick. UPPER suRFACE dull, without pruinose. MEDULLA

white, amyloid (I+ blue). PHoToBIONT trebouxioid with cells <12 um diam.

PROTHALLUS not visible. VEGETATIVE PROPAGULES absent. ASCOMATA

apothecia, exposed. Disc urceolate, without pruinose, orbicular, 0.8-2.0 mm

diam. PROPER EXCIPLE dark brown, 60-100 um thick. Hymentum hyaline,

150-180 um high, not inspersed. HyPOTHECIUM yellowish brown, 15-30 um

thick. PARAPHYSES 1-2 um thick, simple, apices not thickened. Asctr cylindrical,

100-130 x 20-20 um, 8-spored. Ascosporgs ellipsoid, brown, muriform, with

3-5 transverse and 1-2 longitudinal septa, 24-33(-39) x 12-18 um. PYCNIDIA

unknown.

470 ... Abbas, Guo & al.

SPOT TESTS —K+ yellow, C+ and KC+ red, PD-.

SECONDARY METABOLITES —Diploschistesic and lecanoric acids detected

CIE):

EcoLocy — Diploschistes xinjiangensis grows on rock. It is known only

from the type locality at an elevation of 1700-1750 m in northwest China.

ADDITIONAL SPECIMENS EXAMINED — CHINA. XINJIANG: Urumdi, South Mountain,

Aketa, alt. 1700 m, 27 Aug. 2007, A. Abbas 7858, 7859 (XJU).

Comments — Diploschistes xinjiangensis is characterised by the thick, bluish

gray thallus, the 8-spored asci, and the large ellipsoid spores. In morphology

and habitat, this species is very similar to the 8-spored specimens of

D. rampoddensis. However, D. rampoddensis has a thin grayish yellow thallus,

shorter (18-24 um) narrower (7-12 um) ascospores, and a pantropical

distribution and lacks diploschistesic acid, (Lumbsch 1993; Pant & Upreti 1993;

Fernandez-Brime et al. 2013).

Previous studies that have used DNA sequence data for species recognition

in lichens required both monophyly in single-locus ITS phylogenies and

diagnostic morphological differences (e.g., Han et al. 2013). Our morphological

and molecular data for D. xinjiangensis satisfy these criteria.

Acknowledgements

The authors thank Dr. André Aptroot (ABL Herbarium, Soest, The Netherlands),

Prof. Liu-Fu Han (Hebei Normal University, Shijiazhuang, China), and Prof. John A.

Elix (Australian National University, Canberra, Australia) for presubmission review.

S.Y. Guo was awarded grants by the National Natural Science Foundation of China

(nos. 30770012, 31370067) and A. Abbas was awarded grants by the Natural Science

Foundation of China (Nos. 30960003, 31150003). The valuable assistance given by

colleagues is gratefully acknowledged.

Literature cited

Abbas A, Wu JN. 1998. Lichens of Xinjiang. Sci-Tec & Hygiene Publishing House of Xinjiang,

Urumdi.

Fernandez-Brime S, Llimona X, Lutzoni F, Gaya E. 2013. Phylogenetic study of Diploschistes

(lichen-forming Ascomycota: Ostropales: Graphidaceae), based on morphological, chemical,

and molecular data. Taxon 62(2): 267-280. http://dx.doi.org/10.12705/622.10

Frisch A, Kalb K, Grube M. 2006. Contributions towards a new systematics of the lichen family

Thelotremataceae. Bibliotheca Lichenologica 92. 556 p.

http://dx.doi.org/10.1017/s0024282906219066

Guo SY. 2005. Lichens. 31-82, in: WY Zhuang (ed.). Fungi of northwestern China. Ithaca, New

York, Mycotaxon Ltd.

Han LF, Zhang YY, Guo SY. 2013. Peltigera wulingensis, a new lichen (Ascomycota) from north

China. Lichenologist 45(3): 329-336. http://dx.doi.org/10.1017/s00242829 12000837

Lumbsch HT. 1989. Die holarktischen Vertreter der Flechtengattung Diploschistes

(Thelotremataceae). Journal of the Hattori Botanical Laboratory 66: 133-196.

Diploschistes xinjiangensis sp. nov. (China) ... 471

Lumbsch HT, Aptroot A. 1993, Studien tiber die Flechtengattung Diploschistes Il. Nova Hedwigia

Db: 237-239,

Lumbsch HT, Mangold A. 2007. Diploschistes elixii (Ostropales: Thelotremataceae), an

overlooked terricolous species from Western Australia. Lichenologist 39: 459-462.

http://dx.doi.org/10.1017/s0024282907007049

Mangold A, Elix JA, Lumbsch HT. 2009. Diploschistes. Flora of Australia 57(Lichens 5): 227-239.

Martin MP, LaGreca S, Lumbsch HT. 2003. Molecular phylogeny of Diploschistes inferred from ITS

sequence data. Lichenologist 35: 27-32. http://dx.doi.org/10.1006/lich.2002.0427

Novak A, Miklés I, Lyngso R, Hein J. 2008. StatAlign: an extendable software package for joint

Bayesian estimation of alignments and evolutionary trees. Bioinformatics 24(20): 2403-2404.

http://dx.doi.org/10.1093/bioinformatics/btn457

Orange A, James PW, White FJ. 2010. Microchemical methods for the identification of lichens, 2nd

edn. British Lichen Society, London.

Pant G, Upreti DK. 1993. The lichen genus Diploschistes in India and Nepal. Lichenologist 25:

33-50. http://dx.doi.org/10.1017/s0024282993000040

Pérez-Vargas I, Hernandez-Padron C, Pérez de Paz PL, Elix JA. 2012. A new saxicolous species

of Diploschistes (Thelotremataceae) from the Canary Islands. Lichenologist 44(1): 67-71.

http://dx.doi.org/10.1017/s0024282911000612

Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S. 2011. MEGAS: molecular

evolutionary genetics analysis using Maximum Likelihood, Evolutionary Distance, and

Maximum Parsimony Methods. Molecular Biology and Evolution 28: 2731-2739. http://dx.doi.

org/10.1093/molbev/msr121

Wei JC. 1991. An enumeration of lichens in China. International Academic Publishers, Beijing.

White TJ, Bruns TD, Lee SB, Taylor JW, 1990. Amplification and direct sequencing of fungal

ribosomal RNA genes for phylogenetics. 315-322, in: MA Innis et al. (eds). PCR Protocols - a

Guide to Methods and Applications. San Diego, Academic Press.

MYCOTAXON

http://dx.doi.org/10.5248/129.473

Volume 129(2), pp. 473-476 October-December 2014

Uncispora hainanensis sp. nov. isolated from decayed leaves

JIAN- YING Lr*, MIN QIAO”, JIE PENG !, WEN YUN QIAN’,

GUANG-ZHU YANG’, & ZE-FEN Yu!*

‘Laboratory for Conservation and Utilization of Bio-resources,

Key Laboratory for Microbial Resources of the Ministry of Education,

Yunnan University, Kunming, Yunnan, 650091, P. R. China

?Horticultural Research Institute of Yunnan Academy of Agricultural Science

*CORRESPONDENCE TO: zfyuqm@hotmail.com

ABSTRACT—A new species, Uncispora hainanensis, is described and illustrated. It is

characterized by narrowly obclavate or cylindrical, slightly curved at the apical cell, mostly

3-septate, subhyaline or pale brown conidia.

Key worps —fungal diversity, aquatic hyphomycetes, taxonomy

Introduction

Saprobic dematiaceous hyphomycetes are highly diverse on plant material

in China, where many new genera and species have recently been discovered

(Zhang et al. 2009, 2011; Ma et al. 2012a,b,c). During our ongoing survey of

anamorphic fungi in Hainan Province in southwest China, one new species

with morphological characteristics of Uncispora was collected.

Uncispora was introduced by Sinclair & Morgan-Jones (1979) based on

U. harroldiae R.C. Sinclair & Morgan-Jones. The genus is characterized by

brown, macronematous, synnematous or fasciculate conidiophores, and

subhyaline to pale brown, obclavate conidia that are curved or hooked at the

apex and truncate at the base. Uncispora is similar to Sporidesmium Link (Ellis,

1971) in its holoblastic, monoblastic, integrated, non-cicatrized conidiogenous

cells and broadly truncate conidia but differs by its occasionally branched

and synnematous conidiophores and its unique conidia with curved or

hooked terminal cells. There are only two species published, U. harroldiae and

U. sinensis (Yang et al. 2011). Here, we propose a third species.

* Jian- Ying Li & Min Qiao contributed equally to this work

A7A ... Li, Qiao, & al.

Materials & methods

A culture was isolated from submerged decayed leaves in a river in the Wuzhishan

National Nature Reserve. A piece of 3-4 x 6-7 cm decayed leaf was placed on the

surface of CMA (20 g cornmeal, 18 g agar, 40 mg streptomycin, 30 mg ampicillin, 1000

mL distilled water) for ten days; single conidia were isolated using a sterilized toothpick

while viewing with a Olympus BX51 microscope and cultivated on CMA in Petri plates.

Morphological observations were made from CMA after incubation at 25°C for one

week. Conidial size and septation and conidiophore size were determined by measuring

more than 50 elements. Pure cultures and a permanent slide were deposited in the

herbarium of the Laboratory for Conservation and Utilization of Bio-resources, Yunnan

University, Kunming, Yunnan, P.R. China (YMF).

Taxonomy

Uncispora hainanensis Jian Y. Li & Z.F. Yu, sp. nov. PLATE 1

MycoBAnk MB 808816

Differs from Uncispora spp. by its slightly curved apical cell and smaller conidia.

Type: PR China, Hainan Province, Wuzhishan National Nature Reserve, 18°54’N

109°40’E, elev. 754 m, isolated from decayed leaves, Dec. 2011, G. Z. Yang (Holotype,

YMF1.04038; ex-type culture, YMF1.040381).

EryMo toy: hainanensis refers to the province in which the species was found.

Co.onligs pale brown to brown, growing moderately slowly on CMA, attaining

25 mm diam. after 7 days at 25°C. Vegetative hyphae hyaline to pale brown,

branched and septate smooth, commonly 1-30 um wide, aerial mycelium

sparse, hyaline, septate, and branched. CoONIDIOPHORES macronematous,

synnematous, or sometimes in a fascicle of a few, very rarely single, arising

terminally or laterally on hyphae, erect, frequently branched, pale brown to

mid brown, smooth and septate, 55-62 x 2.5-3 um. CONIDIOGENOUS CELLS

determinate, integrated, terminal, cylindrical, monoblastic, or extending

percurrently to produce several terminal and subterminal conidia in fascicles.

CONIDIA narrow clavate to cylindrical (19-)29-52 x 2-3 um, tapering and

curved at the apex, truncate at base, 1-5-septate, mostly 3-septate, subhyaline

to pale brown, smooth, sometimes verruculose at the lower cell .

Comments: U. hainanensis is distinguished from the other two Uncispora

species by its conidia with a curved terminal cell. Conidia of U. harroldiae

are larger (62-78 x 4-5 um) with terminal cells that are mostly hooked. In

U. sinensis the conidial apices only sometimes hook and its 0-7 septate conidia

are much larger (67-89 x 2-3.5 um). Moreover, in U. sinensis the conidia are

much more densely arranged on the conidiophores than in U. hainanensis.

Uncispora hainanensis sp. nov. (China) ... 475

PiaTE 1. Uncispora hainanensis (holotype, YMF 1.04038). A. Conidia. B, C. Conidiophores bearing

conidia in clusters. D. Conidiophore. Scale bars: A, D = 10 um; B, C = 20 um.

Acknowledgements

This work was jointly financed by National Basic Research Program of China

((973’Program: 2013CB127506) and National Natural Science Foundation Program

of PR China (31160008, 31260007, 31360130), Grants from the Young Academic and

Technical Leader Raising Foundation of Yunnan Province (2010CI020). We are very

grateful to Prof. X.G. Zhang and Dr. R.F. Castafieda-Ruiz for critically reviewing the

manuscript and providing precious suggestions on this paper.

Literature cited

Sinclair RC, Morgan-Jones G. 1979. Notes on hyphomycetes. XXVI. Uncispora harroldii [sic] gen.

et sp. nov. Mycotaxon 8: 140-143.

Ellis MB. 1971. Dematiaceous hyphomycetes. Commonwealth Mycological Institute, Kew, England.

Ma LG, Ma J, Zhang YD, Zhang XG. 2012a. Spadicoides camelliae and Diplococcium livistonae, two

new hyphomycetes on dead branches from Fujian Province, China. Mycoscience 53: 25-30.

http://dx.doi.org/10.1007/s10267-011-0138-z

476 ... Li, Qiao, & al.

Ma J, Zhang YD, Ma LG, Ren SC, Castafeda-Ruiz RE, Zhang XG. 2012b. Three new

species of Solicorynespora from Hainan, China. Mycological Progress 11: 639-645.

http://dx.doi.org/10.1007/s11557-011-0775-9

Ma J, Zhang YD, Ma LG, Castafieda-Ruiz REF, Zhang XG. 2012c. Three new species of Sporidesmiella

from southern China. Mycoscience 53: 187-193. http://dx.doi.org/10.1007/s10267-011-0152-1

Yang GZ, Lu J, Yu ZF, Zhang KQ, Qiao M. 2011. Uncispora sinensis, a new species from China.

Mycotaxon 116: 171-174. http://dx.doi.org/10.5248/116.171

Zhang K, Ma LG, Zhang XG. 2009. New species and records of Shrungabeeja from southern China.

Mycologia 101(4): 573-578. http://dx.doi.org/10.3852/09-006

Zhang YD, Ma J, Wang Y, Ma LG, Castafeda-Ruiz RF, Zhang XG.. 2011. New species and

record of Pseudoacrodictys from southern China. Mycological Progress 10: 261-265.

http://dx.doi.org/10.1007/s11557-010-0696-z

MY COTAXON

http://dx.doi.org/10.5248/129.477

Volume 129(2), pp. 477-484 October-December 2014

Lactarius vesterholtii, a new species from India

KANAD DaAs* & DYUTIPARNA CHAKRABORTY

Botanical Survey of India, Cryptogamic Unit,

PO. Botanic Garden, Howrah 711103, India

* CORRESPONDENCE TO: daskanadbsi@gmail.com

ABSTRACT — Lactarius vesterholtii is proposed as new to science. It is characterized by a

greyish brown dry subvelutinous pileus, very crowded light yellow lamellae turning orange

when bruised, white copious latex becoming reddish orange in contact with the lamellae, a

light yellow spore print, amyloid basidiospores ornamented with high ridges that forms an

incomplete reticulum, the absence of pleuro- and cheilomacrocystidia, a trichopalisade to

lamprotrichopalisade-type pileipellis, and a palisade to lampropalisade-type stipitipellis. The

new species is compared with allied Asian and extralimital taxa. A key to the Indian taxa of

L. subg. Plinthogali is also provided.

Key worps — macrofungi, Plinthogali, Russulaceae, Sikkim, taxonomy

Introduction

Lactarius subg. Plinthogali (Burl.) Hesler & A.H. Sm., one of the well

established groups amongst the common milkcaps [Lactarius Pers. and

Lactifluus (Pers.) Roussel], is characterized by a pileus that is dry, velutinous

to velvety, and white to buff, brownish to grey or blackish; distant to crowded

lamellae; a stipe that is mostly concolorous with the pileus; context or latex

in contact with the context mostly changing pinkish to reddish; a spore print

that is buff, pale yellow, ochraceous, or darker but never white; basidiospores

ornamented mostly with high ridges or warts either arranged in a parallel/

zebroid pattern or aligned and connected in a broken to complete reticulum;

macrocystidia mostly absent; and a pileipellis that is a palisade, trichopalisade,

or trichoderm (Basso 1999, Das & Sharma 2004, Stubbe et al. 2008). A closely

related group, Lactifluus subg. Gerardii (A.H. Sm. & Hesler) Stubbe [= Lactarius

subg. Gerardii], can be distinguished by a white spore print and more rounded

cellular elements in a palisade-type pileipellis (Stubbe et al. 2010, Verbeken

& Nuytinck 2013). Ten taxa are known from Lactarius subg. Plinthogali

478 ... Das & Chakraborty

in India: L. subvernalis var. himalayensis Atri et al., L. subvernalis var. albo-

ochraceus Hesler & A.H. Smith, L. crenulatus K. Das & Verbeken, L. fuliginosus

(Fr.) Fr. [= L. fumosus Peck], L. picinus Fr., L. montoyae K. Das & J.R. Sharma,

L. lignyotellus A.H. Sm. & Hesler, L. croceigalus K. Das & Verbeken, L. lignyotus

Fr. var. lignyotus, and L. lignyotus var. canadensis A.H. Sm. & Hesler (Atri et al.

1993, 1994; Das & Sharma 2005; Das & Verbeken 2012).

A number of interesting ectomycorrhizal mushrooms were collected during

a recent macrofungal exploration of the North district of Sikkim Himalaya.

Detailed examination of the collections resulted in the discovery ofa new species

within Lactarius subg. Plinthogali, proposed here as Lactarius vesterholtii.

Materials & methods

Macromorphological features were recorded from the fresh basidiomata in the

field and base camp. Macrochemical reactions with KOH, FeSO, and Guaiacol were

noted. Basidiomata were dried with a field drier. Fresh and dry basidiomata were

photographed using Nikon D300s, Olympus C-5060, Leica DFC550, and Nikon-DS-Nil

cameras. Color codes and terms used are mostly from Kornerup & Wanscher (1978).

Micromorphological characters were recorded with the help of compound microscope

(Nikon Eclipse Ni-U) from free hand sections of dry samples mounted in a mixture of

5 % KOH and Phloxine, 30 % glycerol, and Melzer’s reagent. Drawings were made with

a drawing tube attached to an Olympus CX41 microscope (1000x). Basidium length

excludes length of sterigmata. Basidiospore measurements and features were noted in

side view from twenty basidiospores. Spore measurements and length/width ratios (Q)

are presented as minimum-mean-maximum. Herbarium name follows Holmgren et

al. (1990).

Taxonomy

Lactarius vesterholtii K. Das & D. Chakr., sp. nov. PLATES 1, 2

MycoBaAnk MB 808854

Differs from Lactarius montoyae by its very crowded lamellae and smaller reticulate

basidiospores and from L. croceigalus by its very crowded light yellow lamellae that turn

light orange to orange when bruised, its unbranched pleuropseudocystidia, and its lack

of pileopseudocystidia.

Type: India. Sikkim: North District, Bansoi, 27°41'56.2”N 88°34'10.6’E, alt. 2323 m,

31.VII.2013, K. Das, KD-13-84 (Holotype, CAL).

EryMoLoey: in recognition of the late Dr. Jan Vesterholt (Denmark) for his contribution

to the family Russulaceae

PiLEus 58-75 mm diam., convex when young, becoming planoconvex to

shallowly depressed or slightly infundibuliform, never umbonate; surface rough

PiaTE 1. Lactarius vesterholtii (KD 13-84, holotype). A. Basidiospores; B. Cheiloleptocystidia;

C. Pleuropseudocystidia; D. Cross-section through stipitipellis; E. Cross-section through pileipellis;

E Basidia. Scale bars = 10 um.

Lactarius vesterholtii sp. nov. (India) ... 479

480 ... Das & Chakraborty

to subvelutinous, mostly radially wrinkled, dry to moist, never viscid, light

brown (6D4) to greyish brown (6D3) or sometimes pale yellow (4A3) to greyish

yellow (4B3); margin decurved to incurved, irregularly wavy or interrupted at

maturity. LAMELLAE narrowly adnate to subdecurrent, very crowded (23/cm at

pileus-margin), brittle, light yellow (4A4), turning light orange (5A5) to orange

(5A7) when bruised; edge smooth but, minutely hairy (under stereo-zoom

microscope), concolorous; lamellulae abundant, in 7-8 series. STIPE 55-75 x

8-13 mm, central, more or less cylindrical with tapering base; surface more or

less smooth, yellowish white (4A2) at apex, towards base greyish orange (5B3)

to brownish orange (5C4) or concolorous to pileus, then paler to pale yellow

(4A3) near base and finally white at base, turning light orange (5A5) when

bruised. CONTEXT white, becoming pale yellow (4A3), light orange (5A4) or

darker when exposed, turning pale orange (5A3) with FeSO,, reddish brown

(8D7) with guaiacol and yellowish white (1A2) to pale yellow (1A3) with KOH.

LATEX abundant, white, unchanging when kept isolated, turning yellowish

white with KOH but, on contact with the cut lamellae becoming pinkish white

(7A2) after 3-4 minutes, then finally (or after drying) reddish orange (7A6).

Opor indistinctive. TasTE slightly sour initially, then acrid. SpoRE PRINT light

yellow (4A4).

BASIDIOSPORES 7.0-7.5-8.5 X 6.5-7.1-8.0 um (n = 20, Q = 1.00-1.05-1.14),

globose to subglobose; ornamentation amyloid, composed of high ridges

or wings (up to 2.3 um high) with remarkably and irregularly crenulate to

lobed margin which aligned or connected to form broken to incomplete or

almost complete reticulum, few high isolated subspinoid warts and isolated

to jointed conical to irregular numerous minute warts (often in convoluted

pattern) between high ridges; plage mostly distally amyloid. Basip1a 31-53

x 8-12 um, 4-spored, narrowly clavate to clavate, subcylindric to subclavate

or ventricose; sterigmata 5-8 x 1.5-2 um. PLEUROMACROCYSTIDIA absent.

PLEUROPSEUDOCYSTIDIA 4.5-7.5 tum wide, fairly common, cylindrical to

subcylindrical with somewhat tortuous base, content slightly dense. LAMELLA-

EDGE Sterile. CHEILOMACROCYSTIDIA absent. CHEILOLEPTOCYSTIDIA 17-58 x

5-8 um, abundant, cylindrical to subfusoid with rounded or subcapitate apex,

sometimes tortuous towards apex, emergent to 35 um, often septate, hyaline

in KOH, never with brown pigmentation. CHEILOPSEUDOCYSTIDIA absent.

SUBHYMENIUM layer 13-23 um thick, pseudoparenchymatous, composed of

irregular cells of 7-14.3 x 4-11 um. HYMENOPHORAL TRAMA with some lactifers.

PLaTE 2. Lactarius vesterholtii (KD 13-84, holotype). A, B. Fresh basidiomata; C. Lamellar edge

under stereo-zoom microscope; D. White latex turning reddish orange on cut lamellae; E. Lamellar

edge, cross-section; F. Sphaerocyst nest in stipe trama; G. Pileipellis, radial section; H. Basidiospore

(SEM). Scale bars: C = 0.5 mm; E-G = 50 um; H = 5 um.

Lactarius vesterholtii sp. nov. (India) ... 481

pele is ha a ss

ae yaeee am SS

( Ne g

I Ry y. HL

ry J . .

- a i, ae Ve

482 ... Das & Chakraborty

PILEIPELLIS 84-120 um, two layered (supra- and subpellis), trichopalisade to

lamprotrichopalisade, pileopseudocystidia absent. SUPRAPELLIS composed of

thick-walled hair-like erect often septate hyphal elements; terminal cells 13-53 x

4-7.5 um, cylindrical, ventricose, subfusoid, clavate or irregular with rounded

to subcapitate apex, brown to grey pigmented, wall 0.8-1.3 um thick. SUBPELLIS

composed of somewhat compact oval, ellipsoid to elongate or irregular thick-

walled (up to 1.5 um thick) cells with brown to grey pigmentation. PILEUS

TRAMA composed nests of sphaerocytes and surrounding branched septate

hyphae (4-8 um wide). STIPITIPELLIS up to 110 um thick, two layered, palisade

to lampropalisade. SUPRAPELLIS composed of thick-walled hair-like erect often

septate hyphal elements; terminal cells 8-60 x 4-7 um, brown-pigmented,

mostly cylindrical with subfusoid to subcapitate or rounded apex. SUBPELLIS

mostly of thick-walled isodiametric brown pigmented cells. STIPE TRAMA

composed of numerous nests of sphaerocysts and hyphae.

ECOLOGY & DISTRIBUTION — Gregarious to caespitose, in temperate mixed

(broad-leaf & coniferous) forest under Lithocarpus pachyphyllus (Kurz) Rehder.

July. Rare.

Notes — The combination of features, such as the brown to greyish brown

subvelutinous pileus, white latex turning pinkish white to reddish orange,

globose to subglobose highly ornamented winged spores, absence of

macrocystidia, and the trichopalisade pileipellis, places this species in subg.

Plinthogali. Its very crowded lamellae, emarginate lamellar edge, sour to acrid

taste, spores with combination of high wings and numerous minute warts,

abundant cheiloleptocystidia, and lack of pileocystidia makes it distinct among

the known taxa of this subgenus.

Lactarius montoyae (reported from India and Thailand), L. croceigalus

(reported from India), and L. crassiusculus H.T. Le & Stubbe (reported from

Thailand) are some Asian species that invite comparison with L. vesterholtii.

Lactarius montoyae is distinguished by its distant (4/cm) lamellae, unchanging

latex, ochraceous to orange-yellow spore print, and larger (7.5-10 x 7.3-9.3 um)

non-reticulate spores (Das & Sharma 2004, Le et al. 2007). Lactarius croceigalus

can be separated by its distant (6/cm) lamellae, which are unchanging on bruising,

the possession of pileopseudocystidia, and branched pleuropseudocystidia

(Das & Verbeken 2012). Lactarius crassiusculus has close lamellae, abundant

cheilomacrocystidia, a palisade pileipellis, and trichoderm-type stipitipellis

(Le et al. 2007).

Lactarius acris (Bolton) Gray and L. pterosporus Romagn. are two superficially

similar European species. However, L. acris is characterized by distinctly

viscid to sticky pileus, latex turning coral-pink even when isolated from

Lactarius vesterholtii sp. nov. (India) ... 483

lamellar tissues, and an ixotrichoderm- or ixotrichopalisade-type pileipellis;

L. pterosporus can be separated by a stipe that is never concolorous with the

pileus, a mild (never acrid) taste, and spiral to parallel spore-ornamentation.

Including Lactarius vesterholtii, Lactarius subg. Plinthogaliis now represented

by 11 taxa from India. A key is provided below for their easy identification.

Key to the identification of taxa of Lactarius subg. Plinthogali in India

I*Spote-ornamentation i. S7piih. 2.00 220s op ket Sleek Soe tenes oe eee ESS 2

LMSpOre-OMnaiMmen tariOuy> |S 2E7 MH ies he Pe ahaa oie ae og IM Uh aie dae lle wiacuers Rim et 6

Z.. PICUROMACHOCYSTIGIA, PLOSCIE mm. Fs. ca.o ete aa a te Ps a as os a 3

PE PlEULOMIAGrOCVSUCTasd SOM ericractss teavacsna tent gee tava cee tars yee tars cee tena pelattana GARTER & A

3. Latex does not fade to yellowish after turning carrot red; wide spore range

(6.4-9.7 x 5.6-8 um); basidia longer (40.3-68.4 x 7-16 um)

Pegi bath sata ts oda d nntalg lin zp tog etre Pokey reyes L. subvernalis var. himalayensis

. Latex fades to yellowish after turning carrot red; spore range narrow

(6.9-7.7 x 5.4-7.2 um); basidia shorter (37-48 x 8-11 um)

I Di Jad ne Idea Ve SOR oe PIGS, ch Jaen L. subvernalis var. albo-ochraceus

4 Pileus simall:(2030 min): stipe marrow nv. o'4 .% aig had tly Rly Rae dacs L. crenulatus

4, Pileusilarges( ss 0niii)s Stipe wide 17.53 casts cation tsb dative uae a waabie asada oases 5

5. Pileus yellow-brown to whitish; spore print pinkish buff; context salmon when

CXDOSEC I Rate a oct oeaed a ieta enn ola lana cvalins stolons a ad ies apt opens ae ae L. fuliginosus

5. Pileus dark grey-brown to blackish brown; spore print bright ocher; context pale

red dishi-browarsvieriex posed ilo. Ua, Geta dand 5 mlsins ogee Ls aol chk eats a L. picinus

6. Spore ornamentation never reticulate, of parallel wings >2.5 um high... L. montoyae

6. Spore ornamentation a partial to incomplete reticulum, always <2.5 um high .... 7

7. Latex white, unchanging and not staining lamellae ................. L. lignyotellus

7. Latex white, turning lamellae pinkish to yellowish pink or saffron .............. 8

8. Lamellar edge emarginate, unchanging or becoming orange after bruising;

pileopseudocystidia present or absent; spore-ornamentation <2.3 um high ... 9

8. Lamellar edge emarginate or marginate, but not with the above combination of

CH ATACTSIS: 5 Regn ce ace eegh eee eeys bere beer aes cme ars Lara ee eae beats ae 10

9. Lamellae distant, unchanging after bruising; context unchanging with guaiacol;

spore print ochraceous; pileopseudocystidia present ............. L. croceigalus

9. Lamellae very crowded, becoming orange after bruising; context reddish brown with

guaiacol; spore print light yellow; pileopseudocystidia absent ..... L. vesterholtii

10. Lamellae emarginate, lamellar edge concolorous with the surface; stipe stuffed;

found under Quercus, Rhododendron association ...... L. lignyotus var. lignyotus

10. Lamellae marginate, lamellar edge brownish black; stipe always hollow; found

under: Abies sPieed ASSOCIATION o. | inne Scones erence «es L. lignyotus var. canadensis

484 ... Das & Chakraborty

Acknowledgements

The authors are grateful to Dr. P. Singh, the Director, Botanical Survey of India (BSD),

Kolkata (India) and Department of Forest, Environment and Wild Life Management,

Government of Sikkim for providing all kinds of facilities during this study. They are

thankful to Dr. N.S. Atri and Dr. K.B. Vrinda for kindly reviewing the manuscript and

providing valuable suggestions. Thanks are also due to Dr. Md. N. Aziz (BSI, Cryptogamic

Unit, Howrah), Dr. B.S. Kholia (BSI, SHRC, Gangtok) for helping the senior author in

many ways. Assistance during macrofungal exploration to the North district of Sikkim

rendered by A. Parihar (BSI, Cryptogamic Unit, Howrah) and S. Pradhan (BSI, SHRC,

Gangtok) is also duly acknowledged.

Literature cited

Atri NS, Saini SS, Saini MK, Gupta AK. 1993. Systematic studies on Russulaceous fungi - the genus

Lactarius Pers. Journal of the Indian Botanical Society 72: 155-158.

Atri NS, Saini MK, Saini SS. 1994. Indian Russulaceae Roze - a check list. 81-93, in: TA Sharma et

al. (eds). Current Researches in Plant Sciences. Bishen Singh Mahendra Pal Singh, Dehradun,

India.

Basso MT. 1999. Lactarius Pers. Fungi Europaei 7. Mykoflora, I-Alassio, Italy. 845 p.

Das K, Sharma JR. 2004. Lactarius in Kumaon Himalaya 2: New and interesting species of subgenus

Plinthogali. Mycotaxon 89(2): 289-296.

Das K, Sharma JR. 2005. Russulaceae of Kumaon Himalaya. Botanical Survey of India, Kolkata.

Das K, Verbeken A. 2012. New species of Lactarius subg. Plinthogalus and new records of Lactifluus

subg. Gerardii (Russulaceae) from Sikkim, India. Taiwania 57(1): 37-48.

Heilmann-Clausen J, Verbeken A, Vesterholt J. 1998. The genus Lactarius. The Danish Mycological

Society.

Holmgren PK, Holmgren NH, Barnett LC. 1990. Index Herbariorum. Part 1: herbaria of the world,

8 ed. Regnum Vegetabile 120.

Kornerup A, Wanscher JH. 1978. Methuen handbook of colour, 3rd edition. Eyre Methuen,

London.

Le HT, Stubbe D, Verbeken A, Nuytinck J, Lumyong S, Desjardin DE. 2007. Lactarius in Northern

Thailand: 2. Lactarius subgenus Plinthogali. Fungal Diversity 27: 61-94.

Stubbe D, Nuytinck J, Verbeken A. 2008. Lactarius subgenus Plinthogalus of Malaysia. Fungal

Diversity 32: 125-156.

Stubbe D, NuytinckJ, Verbeken A. 2010. Critical assessment of the Lactarius gerardii species complex

(Russulales). Fungal Biology 114: 271-283. http://dx.doi.org/10.1016/j.funbio.2010.01.008

Verbeken A, Nuytinck J. 2013. Not every milkcap is a Lactarius. Scripta Botanica Belgica 51:

162-168.

MYCOTAXON

http://dx.doi.org/10.5248/129.485

Volume 129(2), pp. 485 October-December 2014

Regional annotated mycobiotas new to the Mycotaxon website

ABSTRACT — Mycotaxon is pleased to announce the posting of a new species distribution

list by Barbosa, Machiner, Barbosa and Gusmao, who review the fungi from Serra da

Jibdia, Brazil. This brings to 113 the number of free access mycobiotas now available on the

Mycotaxon website: http://www.mycotaxon.com/resources/weblists.html

SOUTH AMERICA

Brazil

FLAVIA R. BARBOSA, MONIQUE MACHINER, GLEYSON CRISTIANO K.

BARBOSA, & Lufs FE. P. GusmAo. A checklist of the fungi recorded

from Serra da Jibdia, Bahia state, Brazil. 33 p.

ABSTRACT—Serra da Jibdia is one of the priority areas for the conservation

of biodiversity located in the semiarid Caatinga biome of Northeast Brazil.

Two hundred and fifty-nine species of fungi (mitosporic and meiosporic

ascomycetes and basidiomycetes) were reported from Serra da Jibdia and

are presented here. These species are distributed in 155 genera among which

Dictyochaeta was the most highly represented genus with nine species.

Inventory of the Serra da Jibdia region has thus far contributed to the

description of one new genus and seven new species to science; six species

are considered rare while 121 are new records of fungi. These data were

based on literature records.

MYCOTAXON

http://dx.doi.org/10.5248/129.487

Volume 129(2), pp. 487-492 October-December 2014

BOOK REVIEWS AND NOTICES

ELsE C. VELLINGA, Book Review Editor

861 Keeler Avenue, Berkeley CA 94708 U.S.A.

CORRESPONDENCE TO: bookreviews@mycotaxon.com

Mushrooms of the Midwest. By M. Kuo & A.S. Methven. 2014. University of Illinois

Press, 1325 South Oak Street, Champaign IL 61820-6903. <www.press.uillinois.edu>.

ISBN 978-0-252-07976-4, 440 p., 833 color photos, paper. Price $39.95.

Slightly over a century ago, North American mushroomers wishing to identify

a mushroom from their local field or forest had their work cut out for them.

Throughout the nineteenth century, identifications depended on local experts,

oft-inaccessible scientific papers (Peck’s New York botanical reports spring

to mind), or sparingly illustrated British or Scandinavian tomes. Species

concepts were almost invariably Europe based. The continent was too vast

and mycologists too few to provide anything other than a general nudge in the

right direction, so that the first order of business was usually amassing a sizable

library before coming anywhere near the ‘right’ name. ‘Iron Guts’ McIlvaine

(1902), motivated by an earnest desire to share the edible (and warn against the

not-so-edible) fungal bounty, was arguably the first to publish a volume devoted

solely to North American mushroom identification. Despite its decided eastern

bias (and a disconcerting tendency to refer to plants and stems), it did offer the

American public a means to key out mushrooms anywhere on the continent.

With the introduction of his 1918 THE AGARICACEAE OF MICHIGAN,

Kauffman presented one of the first regional mushroom guides. What began as

a ‘series of surveys initiated in the summer of 1906’ developed into a ‘manual

of considerable size’ accessible to serious scientists. Kauffman’s formidable and

prolific student, A.H. Smith, refined that Michigan agaric manual into shorter

and more accessible guides for the entire continent. Since Smith, the continent

has welcomed a virtual cornucopia of general field guides, but mushroomers

still need a more regional focus to identify their endemic and uncommon

mushrooms with any accuracy.

488 ... Vellinga, BOOK REVIEW EDITOR

Kauffman and Smith aside, there have been relatively few guides devoted

to the central U.S.A. There are a fair number devoted to edible fungi—witness

the lively and colorful books by Kuo (2007) and McFarland & Mueller (2009).

But my library contains only three more comprehensive guides: Stubbs (1971),

covering ~60 central Midwest species; the excellent Horn & al. (1993) keying

out 150 Kansas Mushrooms; and the excellent 1989 guide to mushrooms of the

midcontinental United States by Huffman & al. (see the review of the expanded

2008 revised edition, MyCOTAXON 112: 498-499).

This new guide to 557 taxa, therefore, is both overdue and welcome.

MUSHROOMS OF THE MIDWEST covers 12 states extending “from the cold

conifer bogs of northern Michigan to the steamy oak forests of Missouri” or

from Ohio in the east to Kansas, Nebraska and the Dakotas in the west. Its

authors are well known to legions of mushroom aficionados —- Michael Kuo

as principal developer of MushroomExpert.com and enthusiastic researcher

of all things morel and Andy Methven, Eastern Illinois University professor,

Clavariadelphus and Lactarius expert, and welcome North American foray

instructor.

At 8x10” and 440 pages, MusHROoMs is not a pocket field guide. However

the stiff paper cover reinforced at the spine and semi-gloss paper confer a

hardiness appropriate for foray ground and picnic table. This durability does

present some drawbacks: the color photos are neither as sharp nor as brilliant

as those printed on glossy paper (or viewed on computer screen). The book

is divided into 7 parts: introduction, collection and preservation techniques,

microscope use, keys, mushroom descriptions, the phylogenetic scheme

followed, and a combination glossary and index (with the taxa indexed both

by genus and species).

The first three pages outline the effort needed to determine species, in

particular noting the hours it took Smith and others to collect and describe

them, the many contributions by ‘citizen’ mycologists, and the strides made

through DNA sequence analyses. The next 8 pages briefly outline the equipment

and procedures for mushroom identification and study followed by a 6-page

guide on using a microscope.

There follow 58 pages of keys: a key to the major groups is followed by keys to

the pink-spored gills; pale-spored gills 1&2; Amanita, Lactarius, Russula, dark-

spored gills 1&2; terrestrial pored; lignicolous pored; chanterelles & trumpets;

toothed fungi; puftballs & earthstars; cup mushrooms; morels, false morels,

saddles; clubs & corals; and finally ‘miscellaneous fungi’ [e.g., Hypomyces, birds

nests, stink horns, witch’s butters].

Pages 80-396 cover the fungal taxa. Scientific names stand in easy-to-read

upper case black font with the authorities fully spelled out in red upper case.

Mycotaxon 129 Book Reviews ... 489

At least one colour photo—usually two or three—illustrates each species. Entry

subheadings include ecology, fruitbody description, spore print color, chemical

reactions, microscopic features (always spores, but often other diagnostic

features such as pileipellis, cystidia, clamp connections, paraphyses, etc.), and

comments.

At first riffle, I was perplexed by what seemed to be a disorganized hodge-

podge—ascomycetes inter-mixed with basidios, gilled with pored, conks with

puffballs—that obscured the basic organizational scheme: the alphabet. Kuo &

Methven, however, explain their rationale:

We chose this arrangement because contemporary studies have been

shifting long-held assumptions about relationships between mushrooms

(for example, the polypore Bondarzewia berkeleyi ... is more closely related

to Russula ballouii ... than it is to most other polypores), so that a strictly

taxonomic arrangement (phylum, class, order, family, genus, species)

would not only seem odd to many readers but also probably be subject to

change before the book was actually printed... Contemporary studies also

make a traditional guidebook arrangement based on physical features like

spore print color, the presence of gills or pores, and so on seem antiquated.

Once this pragmatic decision is accepted, there really is little difficulty in finding

a species using the key. Such an arrangement, which is a strong incentive to

USE the key, will no doubt frustrate the key-by-photo crowd but is more likely

to produce an accurate identification.

The authors do not mention whether the photographs are vouchered by dried

specimens, which we hope is the case. I have not taken the time to scrutinize

each photo and description, but the photos appear to be accurately identified,

although in one or two instances, I wondered why a newer name has not been

used [not a Lactifluus to be found]! Nonetheless the nomenclature is more up-

to-date than in most field guides, particularly given that we must adapt to an

exponentially increasing number of name changes. The authors, who follow the

taxonomic classification of Kirk & al. (2008), explain their (and our) quandary

satisfactorily in their concluding chapter, “The evolutionary picture:”

By the time the manuscript we're writing right now is sent to the printer,

some of the information included in the tables below will have changed—

and by the time you are reading this, even more changes may have

occurred. The DNA revolution in mycology has only just begun...The

tables below include only the genera represented in this book...

Overall, this is an excellent addition to the field guide pantheon, and a very

welcome addition to my library.

Horn B, Kay R, Abel D. 1993. A guide to Kansas mushrooms. University Press of

Kansas, Lawrence. 297 p., 150 spp.

490 ... Vellinga, BOOK REVIEW EDITOR

Huffman DM, Tiffany LH, Knaphus G. 2008. Mushrooms & other fungi of the

midcontinental United States [2™ ed.]. University of Iowa Press. 384 p., 300

pl., 284 spp.

Kauffman CH. 1918. The Agaricaceae of Michigan. Michigan Geological and

Biological Survey, pub. 26-B. 924 p. +172 pl.

Kirk PM, Cannon PE Minter DW, Stalpers JA. 2008. Ainsworth & Bisby’s

dictionary of the fungi (10 ed.). CAB International, Wallingford.771 p.

Kuo M. 2007. 100 edible mushrooms. University of Michigan Press. 327 p.

McFarland J, Mueller GM. 2009. Edible wild mushrooms of Illinois and

surrounding states. University of Illinois Press. 232 p., 292 pl.

McIlvaine C, Macadam RK. 1902. One thousand American fungi (rev.). [1973

reissued: Something Else Press, Inc.], West Glover VT. 729 p., 182 pl., ~1000 spp.

Stubbs AH. 1971. Wild mushrooms of the central Midwest. 135 p., ~60 plates

LORELEI L. NORVELL

Pacific Northwest Mycology Service

Portland OR 97229 U.S.A.

IInorvell@pnw-ms.com

A field guide to Tasmanian fungi. By G. Gates & D. Ratkowsky. 2014.

Tasmanian Field Naturalists Club, P.O. Box 68, Hobart, Tasmania 7001, Australia

<http://tasfieldnats.weebly.com/bookshop.html>. ISBN 978-0-9578529-2-1, 254 p.,

colour photos (603 spp.), paper. Price Au$39.95.

Last week's post brought an unexpected jewel to our mailbox. Having the good

fortune of being able to visit Australia twice, I have over the years added four

beautifully illustrated guides to the venerable Cleland (1934-35) and scholarly

four-volume FUNGI OF AUSTRALIA (1996-2003), including Fuhrer & Robinson

(1992), Grey & Grey (1995), Bougher & Syme (1998), and Fuhrer (2005). One

glance at this new field guide to 603 Tasmanian fungi, however, tells me my

next sojourn down-under will be to that fungal paradise south of the mainland.

The authors bring considerable experience to their task, over the past 15

years having made more than 1000 forays collecting and documenting fungi in

Tasmania. Genevieve Gates received her doctorate for her study of macrofungi

on wood, litter, and soil in a Tasmanian eucalypt forest and is coauthor with

Noordeloos of the ENTOLOMATACEAE OF TASMANIA (2012); when not foraying

for mushrooms, Dr. David Ratkowsky serves as a statistician with the Tasmanian

Institute of Agriculture (University of Tasmania).

The book is compact (‘A5’ or 6x84”) and designed for field use. The paper

cover is durable and the semi-gloss papers should withstand moisture yet are

smooth enough so that every photo is crisp, clear, and brilliantly colored. From

the magnificent front cover image of Aseroé rubra to “The Thumbs’ on the back,

the photos are stunning, virtually all with a focus razor-sharp enough to catch

the smallest detail. One minor carp is that the narrow inside margins make it

Mycotaxon 129 Book Reviews ... 491

necessary to hold the book open with both hands to read all of the text, with

some left-hand index letters obscured.

The essential introductory section precedes keys to the gilled genera

followed by a table of key features (spore print, substrate, stipe, comments)

of the gilled genera. Species descriptions include gilled basidiomycetes

(alphabetical by genus) followed by non-gilled basidiomycetes in artificial

groupings—bird’s nests, boletes, chanterelles, coral fungi & clubs, earthballs,

earthstars, jelly fungi, leather fungi, polypores, puffballs, resupinates, spine

fungi, stinkhorns—and concluding with ascomycetes (also alphabetized by

genus). The volume ends with a bibliography, a glossary, the index, and five

pages of Tasmania spectacular enough to entice anyone to the island, although

the guide will definitely be useful on the main continent to the north.

The introduction covers the different biological strategies—ectomycorrhizal,

saprotrophic, pathogenic, parasitic, coprophilous, symbiotic (lichens)—and

notes the important diagnostic macrofeatures used in the book—substrate,

spore print colour, veils, gills & gill-like structures [with 3 % pages of photos of

gill types], color changes with bruising, odours. With respect to nomenclature,

the authors explain:

We have used ... INDEX FUNGORUM as our source of the most up-to-date names

for genera and species. Sometimes we did not adopt the proposed name but

used an older synonym because that is the more familiar name or fitted better

in the context of the book. The alternative name is given in brackets. For most

unknown species we have used a ‘tag name’ and this is in inverted commaas....

The abbreviation ‘aff’ (affinis - similar to) means that we are not absolutely sure

it is this species but it has similar characters, ‘cf? (compare with) is a stronger

similarity, ‘ined’ means it is in the process of being published and ‘nom. prov.

(provisional name) means that it should be given that name when it is published.

The rapid rise of molecular work is resulting in name changes of fungi. This is

not a problem as these changes can be tracked using INDEX FUNGORUM, which

gives the current name and synonyms and also the article in which the change

of name is proposed.

The description pages each offer 2-3 species with generic names at top. At least

one photo (often with detail insets) illustrates each species, accompanied by

the scientific name, a short description with comments, and a fruiting season

diagram. Descriptions are very brief, but the photos (taken in the natural

habitat) are sufficiently clear to permit easy identification. Of particular note

is the unusually large and welcome number of Entoloma species (16+ pages), a

genus usually given short shrift in general field guides. It helps to have an expert

author at hand.

This is a honey of a field guide, which I hope to take along the next time |

have an opportunity to fly to Tasmania. Buy two, though, if for no other reason

than to keep one set of pictures pristine.

492 ... Vellinga, BOOK REVIEW EDITOR

Bougher NL, Syme K. 1998. Fungi of Southern Australia. University of Western

Australia Press, Perth. 391 p., watercolor drawings, 125 spp.

Cleland JB. 1934-1935. Toadstools and mushrooms and other larger fungi

of South Australia, Parts I and II. [Photolitho reprint 1976, A.B. James,

Government Printer, South Australia]. 362 p., 77 b&w figs.

Fuhrer B. 2005. A field guide to Australian fungi. Bloomings Books Pty Ltd,

Melbourne. 360 p., 548 colour pl., >500 spp.

Fuhrer B, Robinson R. 1992. Rainforest fungi of Tasmania and south-east

Australia. CSIRO, East Melbourne. 95 p., ~109 pl.

FUNGI OF AUSTRALIA. 1996. Volume la, Introduction—Classification (eds. AE

Orchard, C Grgurinovic, K Mallet), 413 p.; Volume 1B, introduction—fungi

in the environment (eds. AE Orchard, K Mallet, C Grgurinovic), 405 p.; 1997.

May TW, Wood AE. Volume 2A, Catalogue and bibliography of Australian

macrofungi 1. Basidiomycota p.p. , 348 p.; 2003. May TW, Milne J, Shingles S,

Jones RH. Volume 2B, Catalogue and bibliography of Australian macrofungi

2. Basidiomycota p.p. & Myxomycota p.p. , 452 p. CSIRO Publishing.

Grey P, Grey E. 2005. Fungi down under—the Fungimap guide to Australian

Fungi. Fungimap, Royal Botanic Gardens Melbourne. 146 p., over 200 pl.

~150 spp., distribution maps, colour chart.

LORELEI L. NORVELL

Pacific Northwest Mycology Service

Portland OR 97229 U.S.A.

IInorvell@pnw-ms.com

Book ANNOUNCEMENT

California mushrooms. The comprehensive identification guide. By D.E. Desjardin,

M.G. Wood & F.A. Stevens, 2014. Timber Press Inc., 133 S.W. Second Avenue, Suite

450, Portland OR 97204-3527, <timberpress.com>. ISBN 978-1-60469-353-9. 560 p.,

over 700 colour photos. Price $60

MYCOTAXON

http://dx.doi.org/10.5248/129.493

Volume 129(2), pp. 493-494 October-December 2014

NOMENCLATURAL NOVELTIES AND TYPIFICATIONS

PROPOSED IN MYCOTAXON 129(2)

Coccomyces alienus Y.R. Lin & Xiao Y. Wang, p. 298

Cytospora rostrata C.M. Tian & X.L. Fan, p. 307

Dictydiaethalium dictyosporangium B. Zhang & Yu Li, p. 456

Diploschistes xinjiangensis A. Abbas & S.Y. Guo, p. 469

Entoloma discoloratum Largent, p. 343

Entoloma guttulatum Largent, p. 354

Entoloma hymenidermum Largent, p. 334

Entoloma kewarra Largent, p. 345

Entoloma pamelae Largent, p. 348

Entoloma rugosiviscosum Largent, p. 351

Entoloma violaceotinctum Largent, p. 339

Galerella xalapensis Bandala & Montoya, p. 3

Graphium variabile J.J. Xu & T.Y. Zhang, p. 398

Graphium wuweiense J.H. Kong & TY. Zhang, p. 399

Laboulbenia erotylidarum Haelew., p. 444

Laboulbenia poplitea Haelew., p. 446

Lactarius vesterholtii K. Das & D. Chakr., p. 478

Leptocorticium indicum Samita, Sanyal & Dhingra, p. 361

Leptostroma magnum Y.R. Lin & Lan Zhang, p. 284

Lophodermium quadrisporum D.D. Lu & Y.R. Lin, p. 460

Myrothecium variabile Y.M. Wu & 'T.Y. Zhang, p. 404

Myrothecium xigazense Y.M. Wu & T.Y. Zhang, p. 405

Periconia guangdongensis J.J. Xu & T.Y. Zhang, p.400

Podosphaera girardiniae Z.M. Cao & L.C. Bai, p. 366

Postia subg. Cyanosporus (McGinty) V. Papp, p. 409

Postia africana (Ryvarden) V. Papp, p. 411

Postia amyloidea (Corner) V. Papp, p. 411

Postia caesioflava (Pat.) V. Papp, p. 411

494 ... MYCOTAXON 129(2)

Postia coeruleivirens (Corner) V. Papp, p. 411

Pseudocercospora styracigena Y.L. Guo & B.J. Li, p. 229

Psilocybe cinnamomea J.F. Liang, Yang K. Li & Ye Yuan, p. 216

Rhizoglomus Sieverd., G.A. Silva & Oehl, p. 377

Rhizoglomus aggregatum (N.C. Schenck & G.S. Sm.) Sieverd.,

G.A. Silva & Oehl, p. 378

Rhizoglomus antarcticum (Cabello) Sieverd., G.A. Silva & Oehl, p. 378

Rhizoglomus arabicum (Btaszk., Symanczik & Al-Yahyaei) Sieverd.,

G.A. Silva & Oehl, p. 380

Rhizoglomus clarum (T.H. Nicolson & N.C. Schenck) Sieverd.,

G.A. Silva & Oehl, p. 380

Rhizoglomus custos (C. Cano & Dalpé) Sieverd., G.A. Silva & Oehl, p. 380

Rhizoglomus fasciculatum (Thaxt.) Sieverd., G.A. Silva & Oehl, p. 380

Rhizoglomus intraradices (N.C. Schenck & G.S. Sm.) Sieverd.,

G.A. Silva & Oehl, p. 380

Rhizoglomus invermaium (1.R. Hall) Sieverd., G.A. Silva & Oehl, p. 381

Rhizoglomus irregulare (Blaszk., Wubet, Renker & Buscot) Sieverd.,

G.A. Silva & Oehl, p. 381

Rhizoglomus manihotis (R.H. Howeler, Sieverd. & N.C. Schenck) Sieverd.,

G.A. Silva & Oehl, p. 381

Rhizoglomus microaggregatum (Koske, Gemma & P.D. Olexia) Sieverd.,

G.A. Silva & Oehl, p. 381

Rhizoglomus natalense (Blaszk., Chwat & B.T. Goto) Sieverd.,

G.A. Silva & Oehl, p.382

Rhizoglomus proliferum (Dalpé & Declerck) Sieverd., G.A. Silva & Oehl, p. 382

Sedelnikovaea S.Y. Kondr., M.H. Jeong & Hur, p. 274

Sedelnikovaea baicalensis (Zahlbr.) S.Y. Kondr., M.H. Jeong & Hur, p. 277

Stemonitis mediterraneensis H.H. Dogan & Ero§glu, p. 294

Umbilicaria kisovana (Zahlbr. ex Asahina) Zahlbr. 1940 (lectotypified), p. 417

Uncispora hainanensis Jian Y. Li & Z.F. Yu, p. 474