ORIGINAL ARTICLE

The phylogeny and taxonomy of genera Cystodermaand Cystodermella (Agaricales) based on nuclear ITSand LSU sequences

Irja Saar & Kadri Põldmaa & Urmas Kõljalg

Received: 18 March 2008 /Revised: 10 November 2008 /Accepted: 1 December 2008 / Published online: 9 January 2009# German Mycological Society and Springer-Verlag 2009

Abstract Species delimitation in Cystoderma and Cysto-dermella was evaluated based on ITS and LSU rDNAsequences as well as morphological data. Two species ofCystoderma are synonymised with C. carcharias and threespecies with C. jasonis, distinguishing the synonymisedtaxa as varieties of these accepted species. Analyses ofpartial LSU rDNA sequences revealed Cystoderma andCystodermella as distinct monophyletic genera, withRipartitella representing a well-supported sister group ofthe latter. Phaeolepiota aurea represents either an unsup-ported sister group or member of Cystoderma in thephylogenies based on LSU and ITS sequences rDNA data,respectively. The tribe Cystodermateae sensu Singer did notappear monophyletic according to analyses of LSUsequences. On the basis of these data, the phylogeneticrelationships among most of the analyzed genera could notbe resolved unequivocally.

Keywords Cystoderma Fayod . ITS and LSU rDNA .

Cystoderma and Cystodermella . Phaeolepiota aurea .

Species delimitation

Introduction

The agaric genus Cystoderma Fayod (1889) comprisesspecies with epithelial pileipellis, hyaline, smooth, amyloidor inamyloid basidiospores, lamellae regularly attached tothe stipe, and persistent or evanescent annulus. The firstmonograph of the genus Cystoderma by Smith and Singer(1945) assigned taxa into two sections: Granulosa (withinamyloid basidiospores) and Amianthina (with amyloidbasidiospores); the latter was renamed Cystoderma bySinger (1962). Furthermore, Heinemann and Thoen (1973)introduced two sections: Superba (basidiospores amyloidonly at the suprahilar zone) and Cinnabarina (basidiosporesinamyloid, with cheilocystidia). Harmaja (2002) evaluatedthe taxonomic significance of the basidiospore variationwithin Cystoderma and considered the results of a study onthe nuclear DNA content of selected species (Saar andKullman 2000). Likewise, he considered the results of thephylogenetic analysis by Moncalvo et al. (2002) anddivided Cystoderma into two genera: Cystoderma s. str.containing species that possess amyloid basidiospores andCystodermella, a new genus, encompassing species withinamyloid basidiospores. According to Kirk et al. (2008),the genus Cystoderma contains approximately 35 andCystodermella 12 species, with C. amianthinum (Scop.:Fr.) Fayod and C. granulosa (Batsch: Fr.) Harmaja as therespective type species.

Singer (1986) assigned seven genera to the tribeCystodermateae: Cystoderma, Dissoderma (A.H. Sm. &Singer) Singer, Horakia Oberw., Phaeolepiota Maire ex

Mycol Progress (2009) 8:59–73DOI 10.1007/s11557-008-0578-9

Taxonomical novelties: Cystoderma carcharias (Pers.: Fr.) Fayod var.fallax (A.H. Sm. & Singer) I. Saar, C. jasonis (Cooke & Massee)Harmaja var. lilacipes (Harmaja) I. Saar, C. jasonis (Cooke & Massee)Harmaja var. niveum (Harmaja) I. Saar, C. jasonis (Cooke & Massee)Harmaja var. saarenoksae (Harmaja) I. Saar, Cystodermella granulosa(Batsch: Fr.) Harmaja var. ambrosii (Bres.) I. Saar, C. papallactae(I. Saar & Laessøe) I. Saar.

I. Saar (*) :U. KõljalgInstitute of Ecology and Earth Sciences, University of Tartu,40 Lai St.,51005 Tartu, Estoniae-mail: irja.saar@ut.ee

K. Põldmaa :U. KõljalgNatural History Museum, University of Tartu,Vanemuise 46,51014 Tartu, Estonia

Konrad & Maubl., Pseudobaeospora Singer, RipartitellaSinger and Squamanita Imbach. Kühner (1980) regardedthree—Cystoderma, Phaeolepiota, Squamanita—as mem-bers of this tribe, but assigned Pseudobaeospora in the tribeTricholomateae. The Cystodermateae was regarded as amember of the Agaricaceae by Singer and the Tricholoma-taceae by Kühner. Jülich (1981) considered Cystodermateaea member of the Agaricales, raising it to the rank of familyunder the name Squamanitaceae (nom. rej., Greuter et al.1994) and assigning Phaeolepiota in the family Cortinar-iaceae. Bon (1999) assigned tribes Cystodermateae andDermolomateae Bon into the family Dermolomataceae(Bon) Bon (nom. rej., Greuter et al. 1994) under theTricholomatales. The latter tribe embraced the generaDermoloma J.E. Lange ex Herink and CamarophyllopsisHerink (Bon 1999).

The morphology-based exclusion of the tribe Cystoder-mateae from the Agaricaceae and transfer to the Tricholo-mataceae (Heinemann and Thoen 1973; Bas 1988) wassupported later by phylogenetic analyses of nucLSU sequen-ces (Johnson and Vilgalys 1998). In further large-scalestudies of agaricoid fungi, the phylogenetic relationships ofCystodermateae remained unresolved (Moncalvo et al.2000, 2002). The former study included two representativesfrom two genera (Cystodermella and Ripartitella) that did notform a clade. The latter study added five species representingfour genera: Cystoderma (including the type species ofCystoderma and Cystodermella), Phaeolepiota, Ripartitella,Squamanita, dispersed across various clades of the trees. Therepresentatives of the genera Cystoderma and Ripartitellaformed the sister lineage to the clade comprising members ofthe Agaricaceae. Neither molecular analysis combined withconidiogenesis (Walther et al. 2005) nor basidiospore ultra-structure (Garnica et al. 2007) could resolve the phylogeneticplacement of these taxa.

The recent multi-locus analysis of the Agaricales(Matheny et al. 2006) was the first to resolve the positionof Cystoderma, together with Cyathus and Crucibulum, as amoderately supported sister group of the Agaricaceae in theAgaricoid clade. The results of the study by Garnica et al.(2007) support a close relationship between the generaCystoderma and Phaeolepiota, which are combined withCrucibulum and Cyathus into a well-supported lineage.Unfortunately, both studies included only the type speciesof Cystoderma. Morphological studies on species occurringin Northern Europe and other temperate areas have beenpublished by Saar (2003, 2008a, b).

The aims of this study were as follows:

(1) To test current concepts of species delimitation inCystoderma and Cystodermella

(2) To test whether Cystoderma and Cystodermella sensuHarmaja represent distinct monophyletic genera

(3) To test which are the sister groups of these two genera(4) To test the monophyly of the tribe Cystodermateae

sensu Singer (1986)

Materials and methods

Taxon sampling

The aim of this study was to include as many species from thegenera Cystoderma and Cystodermella as possible. Unfortu-nately, many species are represented only by type specimenstoo old for DNA extraction. The fungal specimens used arelisted in Table 1. The abbreviations of fungaria are accordingto the Index Herbariorum (Holmgren and Holmgren 1998).The sequences of the nuclear ribosomal DNA internaltranscribed spacer (ITS) and/or partial large subunit (LSU)regions were generated for 23 species from the tribeCystodermataceae, including the type specimens of sixspecies (Table 1). In the analyses of LSU data, only onesequence per species was used. Additional LSU sequences ofrepresentatives from the Agaricales sensu Singer (1986) wereproduced or taken from GenBank.

Molecular methods

Genomic DNA was extracted with a CTAB procedure(Gardes and Bruns 1993) with the following modifications:1% β-mercaptoethanol in 600 μl CTAB, tissues werecrushed and incubated at 65°C for 30 min, DNA eluted in40 μl of ddH2O or with High pure PCR templatepreparation kit (Roche Applied Science, Mannheim, Ger-many) following the protocol of the manufacturer. TheCTAB extraction was followed by DNA purification withGeneClean®III kit (Qbiogene, Heidelberg, Germany) orUltraClean™15 kit (Mo Bio Lab., Carlsbad, USA),according to the manufacturer’s instructions. PCR amplifi-cation was accomplished with the primer pairs ITS1F -ITS4B (Gardes and Bruns 1993) and LR0R - LR7 (http://www.biology.duke.edu/fungi/mycolab/primers.htm) for thenuclear ITS (ITS1-5.8S-ITS2) and partial LSU regions,respectively. PCR was performed using puReTaq Ready-To-Go™ PCR Beads (GE Healthcare, Freiburg, Germany)with 0.8 μM of each primer and 5 μl of DNA solution on aTechne Genius thermocycler. The amplification programwas as follows: an initial denaturation at 95°C for 3 min,followed by 35 cycles at 95°C for 30 sec, at 55°C for30 sec, at 72°C for 1 min (increasing time 2 sec per cycle),and a final extension at 72°C for 10 min. PCR productswere purified with QIAquick PCR purification kit (Qiagen,Hilden, Germany), UltraClean™ PCR Clean-up kit (Mo

60 Mycol Progress (2009) 8:59–73

Table 1 Material sequenced for this study

Species Fungarium code a Origin EMBL/ UNITE sequenceaccession nos.

ITS LSU

Armillaria ostoyae TAA 172737 Estonia AM946413Calocybe gambosa TAA 179639 Estonia AM946414Catathelasma imperiale TAA 176551 Canada, Newfoundland AM946417Chamaemyces fracidus TAA 172045 Sweden AM946419Crinipellis scabella TAA 146345 Estonia AM946420Cystoderma amianthinum TAA 182001 Norway AM946479/

UDB003119AM946423

Cystoderma amianthinum TU 101287 Estonia AM946480/UDB003120

AM946424

Cystoderma amianthinum C 4489 Denmark, Greenland AM946478/UDB003121

Cystoderma andinum C 57998 it Ecuador AM946481/UDB003122

AM946425

Cystoderma andinum C 58476 Ecuador AM946482/UDB003123

AM946426

Cystoderma carcharias var. carcharias TAA 172011 Sweden AM946483/UDB003124

AM946428

Cystoderma carcharias var. carcharias TAA 182011 Estonia AM946484/UDB003125

Cystoderma carcharias var. fallax H (IK 92-1309) Finland AM946485/UDB003126

AM946430

Cystoderma carcharias var. fallax TAA 145380 France AM946486/UDB003127

Cystoderma carcharias var. fallax MICH 28586 USA, Arizona AM946487/UDB003128

Cystoderma carcharias var. fallax H ht (as C. intermedium) Finland AM946488/UDB003129

Cystoderma clastotrichum PDD 75555 New Zealand AM946489/UDB003130

AM946432

Cystoderma clastotrichum PDD 82706 New Zealand AM946433Cystoderma clastotrichum PDD 83705 New Zealand AM946490/

UDB003131AM946434

Cystoderma japonicum BR 79022-64 ht Japan AM946491/UDB003132

AM946435

Cystoderma jasonis var. jasonis TAA 182013 Estonia AM946492/UDB003133

AM946436

Cystoderma jasonis var. jasonis TAA 147393 Norway AM946493/UDB003134

Cystoderma jasonis var. jasonis C 40762 Denmark AM946494/UDB003135

Cystoderma jasonis var. lilacipes TAA 147362 Finland AM946495/UDB003136

AM946437

Cystoderma jasonis var. lilacipes TAA 147382 Finland AM946496/UDB003137

Cystoderma jasonis var. niveum H (RS 52585) Finland AM946497/UDB003138

AM946438

Cystoderma jasonis var. niveum O 153866 Finland AM946498/UDB003139

Cystoderma jasonis var. saarenoksae TAA 147361 Finland AM946499/UDB003140

AM946440

Cystoderma jasonis var. saarenoksae H (RT 17.11.1982) Finland AM946500/UDB003141

Cystoderma subvinaceum WU 19742 Austria AM946501/ AM946441

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Table 1 (continued)

Species Fungarium code a Origin EMBL/ UNITE sequenceaccession nos.ITS LSU

UDB003142Cystoderma subvinaceum WU 10567 Austria AM946502/

UDB003143Cystoderma superbum REG (Oct 1976) Germany AM946503/

UDB003144AM946443

Cystoderma superbum BR 22288-75 Belgium AM946504/UDB003145

AM946442

Cystoderma tuomikoskii H ht Finland AM946505/UDB003146

AM946444

Cystoderma tuomikoskii H (SH 88/56; as C. arcticum) Norway AM946506/UDB003147

AM946427

Cystoderma tuomikoskii O 153775 (as C. arcticum) Norway AM946507/UDB003148

Cystoderma tuomikoskii C 34722 (as C. arcticum) USA, Alaska AM946508/UDB003149

Cystoderma tuomikoskii C 2509 (as C. arcticum) Denmark, Greenland AM946509/UDB003150

Cystodermella adnatifolia TAA 175640 Estonia AM946510/UDB003151

AM946421

Cystodermella adnatifolia TAA 147895 Latvia AM946511/UDB003152

Cystodermella cinnabarina TAA 147423 Estonia AM946512/UDB003153

AM946429

Cystodermella cinnabarina TAA 147394 Sweden AM946513/UDB003154

Cystodermella cinnabarina TAA 147387 Estonia AM946514/UDB003155

Cystodermella cinnabarina TAA 175189 Russia, NorthernCaucasus

AM946515/UDB003156

Cystodermella granulosa var. ambrosii H (TU 23.08.74) Finland AM946516/UDB003157

AM946422

Cystodermella granulosa var. ambrosii S F5083 Sweden AM946517/UDB003158

Cystodermella granulosa var.granulosa

TAA 147491 Estonia AM946518/UDB003159

AM946431

Cystodermella granulosa var.granulosa

TAA 147492 Norway AM946519/UDB003160

Cystodermella granulosa var.granulosa

C 2604 Denmark, Greenland AM946520/UDB003161

Cystodermella papallactae C 58002 it Ecuador AM946521/UDB003162

AM946439

Cystolepiota seminuda TAA 185602 Estonia AM946445Delicatula integrella H (IK 92-055) Finland AM946446Flammulina fennae TAA 128242 Estonia AM946448Gerhardtia incarnatobrunnea H (US 1593) Finland AM946449Gymnopus fusipes C 42389 Denmark AM946450Hygrocybe coccinea TAA 176130 Estonia AM946460Laccaria laccata TAA 176097 Estonia AM946452Lepiota aspera TAA 185687 Estonia AM946447Lepista nuda TAA 182753 Estonia AM946453Leucoagaricus leucothites TAA 176064 Estonia AM946454Leucocortinarius bulbiger H (IK 98-1345) Finland AM946455Macrolepiota procera TAA 147834 Estonia AM946456Megacollybia platyphylla C 45901 Denmark AM946457

62 Mycol Progress (2009) 8:59–73

Bio Lab., Carlsbad, USA) or Exo-SAP (GE Healthcare,Freiburg, Germany) according to the manufacturer’sinstructions. The sequences were performed on an ALFex-pressII (GE Healthcare, Freiburg, Germany) automatedsequencer or by MWG-Biotech AG (Ebersberg, Germany)and Macrogen Inc. (Seoul, Korea) using primers ITS1 andITS4 (White et al. 1990) or CTB6 (http://plantbio.berkeley.edu/~bruns/tour/primers.html), LR3R, LR5 (http://www.biology.duke.edu/fungi/mycolab/primers.htm) for ITS andLSU regions, respectively. In the first case cycle sequencingwas performed using Thermo Sequenase Primer CycleSequencing kit (GE Healthcare, Freiburg, Germany),according to the manufacturer’s instructions, using0.2 μM Cy5 primers.

Alignments and phylogenetic analyses

Sequence fragments were inspected and assembled usingSequencher 4.6 (Gene Codes, Ann Arbor, USA). DNAsequences were submitted to the European MolecularBiology Laboratory (EMBL) and UNITE (Kõljalg et al.2005) databases under accession numbers given in Table 1.Alignments were performed using L-INS-i strategy asimplemented in MAFFT v6.240 (Katoh et al. 2002,2005). Minor manual adjustments were performed withSe-Al 2.0a11 (Rambaut 1996).

Phylogenetic analyses were performed separately for theLSU and ITS datasets due to incomplete ITS data and

alignment problems between distantly related genera. Theinitial analyses of the LSU dataset were based on a moreexhaustive data matrix that included representatives of themain groups of the euagaric clade (Moncalvo et al. 2002).

The ITS dataset was divided into two subsets: Cystodermaand Cystodermella corresponding to the respective twomajor, distantly related clades of the ingroup obtained inLSU analyses. In the LSU dataset Boletus edulis Bull. waschosen to represent the outgroup. For ITS datasets ofCystoderma and Cystodermella, the species pairs Cyathusstriatus (Huds.) Willd. and Crucibulum laeve (Huds.)Kambly or Ripartitella alba Halling & Franco-Mol. and R.brasiliensis (Speg.) Singer, respectively, were chosen asoutgroups. Maximum parsimony (MP) analyses were con-ducted in PAUP* 4.0b10 (Swofford 2002) using 1000heuristic searches with random taxon addition sequences,TBR branch swapping, Maxtrees set to 10,000, the restric-tion to save 100 trees in each replicate applied, followed byadditional swapping of the resulting trees. The confidence ofbranching was assessed using bootstrap re-sampling (bs):1,000 replicates, each with 10 random taxon additionsequences and MulTrees off. All characters were treated asunordered, equally weighted, treating gaps as missing data.

Bayesian inference of phylogeny was performed withMrBayes 3.1.2 (Huelsenbeck and Ronquist 2001; Ronquistand Huelsenbeck 2003) applying the GTR + I + G substi-tution model for the LSU and ITS_Cystoderma datasets,and the HKY + G DNA model for ITS_Cystodermella

Table 1 (continued)

Species Fungarium code a Origin EMBL/ UNITE sequenceaccession nos.ITS LSU

Melanoleuca melaleuca H (PS 7265) Finland AM946458Phaeolepiota aurea TAA 146976 Estonia AM946522/

UDB003163Phaeolepiota aurea C 27851 Denmark AM946523/

UDB003164AM946459

Resupinatus applicatus TAA 171569 Estonia AM946461Rhodocollybia maculata TAA 147123 Estonia AM946462Ripartitella alba NY (RH 7182) it Costa Rica AM946525/

UDB003165AM946464

Ripartitella brasiliensis NY (EFM 499) Colombia AM946524/UDB003166

AM946465

Ripartites tricholoma TAA 171619 Estonia AM946466Rugosomyces onychinus TAA 146671 Sweden AM946467Strobilurus esculentus TAA 176630 Estonia AM946468Tricholoma equestre TAA 176069 Estonia AM946471Xerula pudens H (IK 94-1816) France AM946474

a BR National Botanic Garden of Belgium, C University of Copenhagen, H University of Helsinki, MICH University of Michigan, NY New YorkBotanical Garden, O Botanical Museum, PDD Landcare Research REG University of Regensburg, S Swedish Museum of Natural History, TAAEstonian University of Life Sciences, TU University of Tartu, WU University of Vienna; collectors: EFM A.E. Franco-Molano, IK I. Kytövuori,PS P. Salo, RH R.E. Halling, RS R. Saarenoksa, RT R. Tuomikoski, SH S. Huhtinen, TU T. Ulvinen, US U. Söderholmht holotypeit isotype

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dataset, all chosen by the AIK information criterion in theprogram MrModeltest 2.2 (Nylander 2004). The Markovchains Monte Carlo (MCMC) were run with 2 M gen-erations for both ITS datasets and 5 M generations for theLSU dataset, applying default values of other prior settings.According to the specified generations, the first 200 K/500 K generations without a stable likelihood score werediscarded, leaving 36,002/ 90,002 trees for calculating theconsensus trees and Bayesian posterior probability (pp)values.

Additionally, two constraint analyses were performed onthe LSU dataset to test the probability of two speciesforming a monophyletic clade together with the species ofCystoderma. The likelihood values of resulting constrainedtrees with the lowest −ln L were compared to the bestunconstrained MP tree using the Shimodaira–Hasegawa test(Shimodaira and Hasegawa 1999) in PAUP* with aresampling estimated log-likelihood (RELL) bootstrap with1,000 replicates.

The pairwise base differences for ITS datasets werecalculated with the default setting in PAUP* 4.0b10.

Results

Analyses of the LSU dataset

The LSU dataset comprises 68 taxa and has an alignedlength of 1,137 characters with 733 constant, 142 parsimo-ny-uninformative, and 262 parsimony-informative charac-ters. Parsimony analysis of this dataset resulted in 76equally most parsimonious trees with the best tree depictedin Fig. 1 (length=1,164, CI=0.349, RI=0.495, −ln L=9,422.79845). In the strict consensus many of the deeperunsupported branches were collapsed. The consensus treeobtained from Bayesian analysis were generally concordantwith the strict consensus of the MP trees; their majordifferences are outlined below.

Members of the tribe Cystodermateae sensu Singer didnot constitute a monophyletic group. The genus Cystodermasensu Singer did not appear monophyletic in any analysesbased on partial LSU sequences. The two major subclades,well-supported by bootstrap and/or Bayesian analyses,correspond more or less to the genera established byHarmaja (2002): 1) Cystoderma (excluding C. superbum;94% bs, 100% pp) and 2) Cystodermella (84% bs). Theformer corresponds to the section Cystoderma and the latterto the section Granulosa sensu Singer (1986). Phaeolepiotaaurea formed an unsupported sister group to the Cystodermaclade. The larger clade, comprising all these taxa, wassupported (99% pp) only in the Bayesian tree.

The genus Ripartitella formed the sister group to theclade comprising representatives from the genus Cystoder-

mella (93% bs, 99% pp) in both LSU analyses. Pseudo-baeospora pyrifera was designated a sister group ofTricholoma and Lepista (95% pp) by Bayesian analysisbut by MP analysis its relationship remained unresolved.The representatives of the genus Squamanita formed anunsupported clade in both LSU analyses.

Two separate analyses were run, the first constrainingrepresentatives of Cystoderma including C. superbum toform a monophyletic group, and the second includedadditionally Phaeolepiota. The unconstrained tree with thebest likelihood value was assigned the best likelihood valueby the Shimodaira–Hasegawa test (−ln L=9,422.79845).The hypotheses about the monophyly of Cystodermaincluding C. superbum or including both C. superbum andPhaeolepiota were considered equal to the topology of thetree with the best likelihood value (P=1.000).

Analyses of the ITS datasets

The ITS_Cystoderma dataset comprises 36 taxa and has analigned length of 751 characters with 440 constant, 59parsimony-uninformative, and 252 parsimony-informative.Parsimony analysis of this dataset resulted in two equallymost parsimonious trees (length=652, CI=0.716, RI=0.866, −ln L=3,712.55193); one of which is depicted inFig. 2.

Monophyly of the ingroup is not supported in thisanalysis. Thirteen well-supported clades are distinguishedamong the species of Cystoderma, incorporating alsoPhaeolepiota aurea. Ten of these clades represent distinctspecies as delimited on the grounds of morphological andmolecular characters. These are: 1) C. carcharias, includingC. fallax and C. intermedium; 2) C. tuomikoskii, includingC. arcticum; 3) C. amianthinum; 4) C. andinum; 5) C.subvinaceum; 6) C. japonicum; 7) C. clastotrichum; 8) C.jasonis, including C. lilacipes, C. niveum and C. saarenok-sae; 9) C. superbum; 10) Phaeolepiota aurea. Theirtaxonomy is discussed below.

The ITS_Cystodermella dataset comprises 14 taxa andhas an aligned length of 619 characters with 433 constant,27 parsimony-uninformative, and 159 parsimony-informa-tive. Parsimony analysis of this dataset resulted in one mostparsimonious tree (length=233, CI=0.944, RI=0.966, −lnL=1,867.26609) depicted as a phylogram (Fig. 3). Strongsupport was assigned to the ingroup as well as most of itssubclades. Four species were distinguished: C. adnatifolia,C. granulosa, C. papallactae, and C. cinnabarina. Cys-todermella cinnabarina, the only species with cystidiaincluded in this study, formed a sister group to the rest ofthe species. These results concur with the current speciesconcepts of the genus Cystodermella. Only C. ambrosii didnot segregate from C. granulosa based on the gene datainvestigated.

64 Mycol Progress (2009) 8:59–73

Taxonomy

Twenty species of Cystoderma sensu Singer were incorpo-rated into the study. Analyses of ITS and LSU datasupported the distinction of eleven species of Cystodermasensu Harmaja and four species of Cystodermella Harmaja.In making decisions on species delimitation, we evaluatedthe information on the phylogenetic relationships obtainedin the various analyses of the studied material as well as thevariation of ITS sequences based on pairwise base differ-ences. Combining species was considered when thevariability of ITS sequences between two taxa was lessthan 1% (p=0.01). Additionally, morphological charactersof the species, discussed in more detail by Saar (2003),

were considered. The different types of evidence resulted inthe following new combinations.

Genus Cystoderma Fayod

Cystoderma amianthinum (Scop.: Fr.) Fayod, Ann. Sci.Nat., Bot., Sér. VII 9: 351. 1889.

≡Agaricus amianthinus Scop., Fl. carn., Ed. 2, 2: 434. 1772.≡ Agaricus granulosus var. amianthinus (Scop.) Fr.,

Epicr.: 18. 1838.≡ Lepiota granulosa var. amianthina (Scop.: Fr.) P.

Kumm., Führer Pilzk.: 136. 1871.≡ Lepiota amianthina (Scop.: Fr.) P. Karst., Hattsvamp.:

15. 1879.

Fig. 1 The best tree of 76equally parsimonious trees (−lnL=9,422.79845, length=1,164,CI=0.349, RI=0.495) obtainedin parsimony analysis of LSUsequence data. Bootstrap sup-port (≥70%) and posterior prob-abilities (≥95%) are shownabove and below the branches(bs/pp), respectively. Speciesaccepted in the paper are markedwith scale bars and the typespecimens with asterisks (*)

Mycol Progress (2009) 8:59–73 65

≡ Armillaria amianthina (Scop.: Fr.) Kauffman, Pap.Michigan Acad. Sci. 2: 60. 1923.

Epitype: Estonia, Jõgeva Co., Saare Comm., Pedassaare,58°39.181′N 26°52.256′E, in pine forest, 26 Oct 2006, leg.I. Saar (TU 101287).

This species is characterised by a yellowish-buff pileus,white lamellae and amyloid, ellipsoid to oblong basidio-spores (5–6×3–3.5 μm). Horak (1968: 206) selected thelectotype from the Fayod’s collection (G).

Cystoderma andinum I. Saar & Laessøe, Mycotaxon 96:124. 2006.

Type studied: Ecuador, Prov. Napo, just below and E ofthe Papallacta pass, páramo El Virgen, 00°19′66″S 78°12′

06″W, among unidentified cushion plants in open páramo,alt. ca. 3800 m, 7 Apr 2002, T. Læssøe TL-9246 (QCA,holotype; C 57998, isotype).

This species is characterised by a greyish-orange pileusand strongly amyloid, broadly ellipsoid to ellipsoid basi-diospores (6.0–7.5×4.5–5.5 μm).

Cystoderma chocoanum Franco-Mol., Mycologia 85(4): 673. 1993.

Type studied: Colombia, Dept. Chocó, Municipio deIstmina, vereda Pie de La Pepe, selva degradada, alt.70 m, 10 May 1991, leg. A.E. Franco-Molano 629, P.Arbaláez & G. Henao (COL, holotype; NY 775586,isotype).

Fig. 2 Phylogeny of the genusCystoderma inferred from un-weighted parsimony analysis ofITS sequence data. One of thetwo best trees (−ln L=3,712.55193, length=652, CI=0.716, RI=0.866). Bootstrapsupport (≥70%) and posteriorprobabilities (≥95%) are shownabove and below the branches(bs/pp), respectively. Speciesaccepted in the paper are markedwith scale bars and the typespecimens with asterisks (*)

66 Mycol Progress (2009) 8:59–73

This species described from Colombia is characterisedby a brown pileus, a persistent membranous flaring ring onstipe and amyloid, ellipsoid to oblong basidiospores(5.4–6.3×2.7–3.6 μm).

Cystoderma clastotrichum (G. Stev.) E. Horak, NewZealand J. Bot. 9 (3): 411. 1971.

≡ Baespora clastotricha G. Stev., Kew Bull. 19 (1): 56.1964.

Type studied: New Zealand, Keith George Park, Wel-lington, rooting in moss on fallen wood, 15 Jun 1949, leg.G. Stevenson Cone 653 (K (M) 157434).

This species described from Nothofagus forest in NewZealand is characterised by ochraceous yellow pileus andamyloid ellipsoid basidiospores (3.5–4.5×2.5–3 μm).Arthrospores are present in the context under the pilei-pellis.

Cystoderma carcharias (Pers.: Fr.) Fayod, Ann. Sci.Nat., Ser. 7, 9: 351. 1889.

≡ Agaricus carcharias Pers., Tent. Disp. Meth. Fung. 18.1797.

≡ Agaricus granulosus var. carcharias (Pers.) Fr., Epicr.:18. 1838.

≡ Lepiota granulosa var. carcharias (Pers.) P. Kumm.,Führer Pilzk.: 136. 1871.

≡ Lepiota carcharias (Pers.: Fr.) P. Karst., Hattsvampar:14. 1879.

≡ Cystoderma carcharias (Pers.: Fr.) Fayod, Ann. Sci.Nat., Ser. 7, 9: 351. 1889.

=Cystoderma fallax A.H. Sm. & Singer, Pap. MichiganAcad. Sci. 30: 116. 1945.

=Cystoderma intermedium Harmaja, Karstenia 19: 27.1979.

Fig. 3 The only best tree infer-ring the phylogeny of the genusCystodermella from unweightedparsimony analysis of ITS se-quence data (−ln L=1,867.26609, length=233, CI=0.944, RI= 0.966). Bootstrapsupport (≥70%) and posteriorprobabilities (≥95%) are shownabove and below the branches(bs/pp), respectively. The typespecimens are marked withasterisks (*)

Mycol Progress (2009) 8:59–73 67

This species is characterised by a persistent membranousflaring ring on stipe and amyloid, broadly ellipsoid toellipsoid basidiospores (4–5.5×3–4 μm).

Cystoderma carcharias (Pers.: Fr.) Fayod var. fallax (A.H. Sm. & Singer) I. Saar, comb. nova

Basionym: Cystoderma fallax A.H. Sm. & Singer, Pap.Michigan Acad. Sci. 30: 116. 1945.

Type studied: USA, Clallam Co., Washington, LakeMills, Olympic National Park, 48°0.13″ N, 123°35.92″ W,under mixed alder and conifers, 25 Sep 1941, leg. A.H.Smith 17272 (MICH 10535).

Synonym: Cystoderma intermedium Harmaja, Karstenia19: 27. 1979.

Type studied: Finland, Etelä-Häme, Janakkala, Kiri,Takkusaare forest road, in spruce forest, 8 Oct 1977, M.Korhonen 2094 & R. Tuomikoski (H).

This variety differs from the type variety by itsferrugineous pileus and stipe, and sphaerocysts which turndark rusty cinnamon in KOH. C. intermedium (collected asC. fallax) was described from Finland and afterwardsrecorded in Norway (Sivertsen et al. 1994).

Cystoderma carcharias (Pers.: Fr.) Fayod var. carcha-rias

Epitype: Sweden, Gotland, 10 km N from Nyneshamn,in pine forest, 3 Oct 1998, leg. M. Vaasma (TAA 172011).

The type variety has pale grayish-pink or vinaceouspileus and stipe; the sphaerocysts remain hyaline in KOH.Singer (1961: 13) selected the lectotype from the Persoon’scollection (L 910.250-1473), which agrees with Fries’sconcept of this species.

Cystoderma japonicum Thoen & Hongo, Trans. Mycol.Soc. Japan 26 (1): 23. 1985.

≡ Cystodermella japonica (Thoen & Hongo) Harmaja,Karstenia 42: 46.2002.

Type studied: Japan, Shibahara, Otsu-city, in forest, onheaps of rice hulls, alt. 100 m, 7 Oct 1980, leg. T. Hongo6221 (BR 79022-64).

This species is characterised by yellow-ochre to orange-yellow pileus, a persistent membranous ring on stipe andinamyloid ellipsoid basidiospores (4–5×2.5–3.5 μm).

Cystoderma jasonis (Cooke & Massee) Harmaja, Kar-stenia 18: 29. 1978.

≡ Agaricus jasonis Cooke & Massee, Cooke Grevillea16 (79): 77. 1888.

=Lepiota amianthina var. longispora Kühner, Bull. Soc.Mycol. France 52: 204. 1936 (nom. inval., Art. 36.1).

=C. amianthinum (Scop.) Fayod var. longisporumKühner, Bull. Mens. Soc. Linn. Lyon 38 (6): 184. 1969.

=C. longisporum (Kühner) Heinem. & Thoen, Bull.Trimestriel Soc. Mycol. France 89: 16. 1973 (nom. inval.,Art. 36.1, 43.1).

=C. longisporum (Kühner) Heinem. & Thoen exArnolds, Bibliotheca Mycologica 90: 323. 1982.

=C. lilacipes Harmaja, Karstenia 18: 29. 1978.=C. niveum Harmaja, Karstenia 25: 41. 1985.=C. saarenoksae Harmaja, Karstenia 25: 41. 1985.This species is distinguished by amyloid, oblong to

fusoid spores (5.5–8×3–4.5 μm). Four varieties can bedistinguished according to basidiocarp colour and presenceof arthrospores in the context below the pileipellis.

Cystoderma jasonis (Cooke & Massee) Harmaja var.lilacipes (Harmaja) I. Saar, comb. nova

Basionym: Cystoderma lilacipes Harmaja, Karstenia 18:29. 1978.

Type studied: Finland, Etelä-Karjala, Vehkalahti, NWpart of Turkia, in Larix plantation, on acid rocky outcrops,1 Oct 1977, leg. L. Fagerström (H).

Synonym: Cystoderma longisporum (Kühner) Heinem.& Thoen ex Arnolds var. purpurascens Heinem. & Thoen.ex Arnolds, Bibliotheca Mycologica 90: 324. 1982.

Cystoderma longisporum var. purpurascens Heinem. &Thoen., Bull. Trimestriel Soc. Mycol. France 89: 18. 1973(nom. inval., Art. 36.1, 43.1).

This variety has violet or purple apex of the stipe and pale todark brown hygrophanous pileus. It has been reported inNordiccountries, where it usually grows on acidic rock outcropsamong mosses (Saar 2008a), Belgium (Heinemann andThoen 1973), as well as in Québec, Canada (Huhtinen 1987).

Cystoderma jasonis (Cooke & Massee) Harmaja var.niveum (Harmaja) I. Saar, comb. nova

Basionym: Cystoderma niveum Harmaja, Karstenia 25:41. 1985.

Type studied: Finland, Uusimaa, Espoo, Tapiola, Suvi-kumpu, low, acid, forested, human influenced rock out-crops, among mosses such as Dicranum scoparium andPolytrichum juniperinum; associated with Cantharellulaumbonata, Cystoderma jasonis, C. lilacipes, C. saarenok-sae, Laccaria cf. proxima, alt. 20 m, 14 Nov 1981, leg. H.Harmaja H6003345 (H).

In the white variety of this species, arthrospores areabsent and sphaerocysts hyaline in KOH. It has been foundin Nordic countries, usually as growing on acidic rockoutcrops among mosses (Saar 2008a). It has also beenrecorded in Belgium by Heinemann and Thoen (1973, as awhite variety of C. longisporum).

Cystoderma jasonis (Cooke & Massee) Harmaja var.saarenoksae (Harmaja) I. Saar, comb. nova

Basionym: Cystoderma saarenoksae Harmaja, Karstenia25: 41. 1985.

Type studied: Finland, Uusimaa, Helsinki, Vanhakau-punki, Annala, siliceous rock outcrops, on or among themosses Polytrichum juniperinum, P. piliferum, Dicranumscoparium etc., associated with e.g. Betula sp., Pinussylvestris, Quercus robur, Sorbus aucuparia, Callunavulgaris and Cystoderma lilacipes, 13 Oct 1980, leg. R.Saarenoksa 72380 (H).

68 Mycol Progress (2009) 8:59–73

This variety has yellowish-brown pileus and stipe;arthrospores are absent. It has been found only in Finland,usually as growing on acidic rock outcrops among mosses(Saar 2008a). The basidiocarps of type specimen weregrowing among those of C. lilacipes.

Cystoderma jasonis (Cooke & Massee) Harmaja var.jasonis

Type studied: Scotland, Carlisle, leg. Dr. Carlyle (K (M)140607, holotype; NY 12556, isotype).

Synonym: C. amianthinum (Scop.) Fayod var. long-isporum Kühner, Bull. Mens. Soc. Linn. Lyon 38 (6): 184.1969.

Type studied: France, Savoie, Forêt du Praz, St-Bon, inforest, on rotten wood, 29 Jul 1927, leg. R. Kühner (G53645).

This type variety is characterised by yellowish-brownpileus and stipe; arthrospores are present in the contextunder the pileipellis.

Cystoderma subvinaceum A. H. Sm., in A.H. Sm. &Singer, Pap. Michigan Acad. Sci. 30: 120. 1945.

Type studied: USA, Ontario, Nipissing Co., Sand Point,Lake Timagami, 10 Sep 1936, A.H. Smith 4755 (MICH10539).

This species is characterised by vinaceous buff pileusand amyloid ellipsoid basidiospores (4–4.5×2.5–3 μm); thepileipellis colours olivaceus grey in KOH.

Cystoderma superbum Huijsman, Fungus 26 (1–4): 39.1956.

Type studied: Switzerland, Canton of Luzern, nearSchüpfheim, on sawdust, probably of Picea, 8 Sep 1955,leg. A.W. Huijsman (L 626716, holotype; MICH 10540,isotype).

This species is characterised by the purplish colour of itsbasidiocarps; inamyloid or amyloid only at suprahilar zone,ellipsoid basidiospores (4–5×3–3.5 μm).

Cystoderma tuomikoskii Harmaja, Karstenia 19: 27.1979.

Type studied: Finland, Koillismaa, Kuusamo, Juuma, 23Aug 1974, leg. R. Tuomikoskii (H).

=Cystoderma arcticum Harmaja, Karstenia 24: 31. 1984.Type studied: Norway, Svalbard, Isfjorden, Long-

yearbyen, 78°13′ N, 15°40′ E, 22 Aug 1966, leg. E.Kankainen (Ohenoja) (TUR 69527).

This species is similar to C. carcharias var. fallax; bothare characterised by a persistent membranous flaring ring.The spores, however, are considerably larger (5.5–8 (10)×4–5 (6) μm). The holotype of C. tuomikoskii was collectedin a boreal spruce forest in Finland, and C. arcticum in anarctic meadow in Svalbard (as C. granulosum, Ohenoja1971). Harmaja (1984) considered these two species closelyrelated. He distinguished C. arcticum by darker pileus,somewhat smaller spores, considering it as the onlyCystoderma species restricted to the arctic zone. C.

tuomikoskii has been reported from Alaska by Laursenand Ammirati (1982) and by Miller (1993, as C. arcticum),and from the Russian Arctic by Karatygin et al. (1999, as C.arcticum and C. tuomikoskii).

Genus Cystodermella Harmaja

Cystodermella adnatifolia (Peck) Harmaja, Karstenia 42(2): 45. 2002.

≡ Lepiota adnatifolia Peck, Bull. New York State Mus.Nat. Hist. 54, Bot. 5: 947. 1902.

≡ Armillaria adnatifolia (Peck) Kauffman, Pap. Michi-gan Acad. Sci. 2: 60. 1923.

≡ Cystoderma granulosum (Batsch: Fr.) Fayod var.adnatifolium (Peck) A.H. Sm. & Singer, Pap. MichiganAcad. Sci. 30: 90. 1945.

≡ Cystoderma adnatifolium (Peck) Harmaja, Karstenia14: 122. 1974.

Type studied: USA, New York, Hague, under pine, 23Sept 1900, C.H. Peck (NYS F-79, syntype).

This species is characterised by a pileus in shades oforange and inamyloid, ellipsoid to oblong basidiospores(4–5×2.5–3 μm).

Cystodermella cinnabarina (Alb. & Schwein.: Fr.)Harmaja, Karstenia 42 (2): 45. 2002.

≡ Agaricus granulosus Batsch var. cinnabarinus Alb. &Schwein., Consp. Fung.: 147. 1805.

≡ Agaricus cinnabarinus (Alb. & Schwein.) Fr., Syst.mycol. 3 (Index): 12. 1832.

≡ Lepiota granulosa var. cinnabarina (Alb. & Schwein.:Fr.) Gillet, Hyménomycètes: 71. 1874.

≡ Lepiota cinnabarina (Alb. & Schwein.: Fr.) P. Karst.,Hattsvamp.: 14. 1879.

≡ Armillaria cinnabarina (Alb. & Schwein.: Fr.) Kauff-man, Pap. Michigan Acad. Sci. 2: 60. 1923.

≡ Cystoderma cinnabarinum (Alb. & Schwein.: Fr.)Fayod, Ann. Sci. Nat., Bot., Sér. VII 9: 351. 1889.

Type: unknown.=Agaricus terreyi Berk. & Broome [as ‘terreii’], Ann.

Mag. Nat. Hist., Ser. IV, 6: 462. 1870.=Lepiota terreyi (Berk. & Broome) P. Karst., Hatts-

vamp.: 14. 1879.=Cystoderma terreyi (Berk. & Broome) Harmaja [as

‘terreii’], Karstenia 18: 29. 1978.Type studied: England?, Torres. [illeg.], 29 Oct 1870,

leg. M. Terrey (K (M) 140617, isotype).Epitype: Estonia, Tartu Co., Mäksa Comm., Kaagvere, in

pine forest, on moss, 10 Oct 1999, leg. I. Saar (TAA 147423).This species is characterised by a pileus in shades of

orange, inamyloid, ellipsoid to oblong basidiospores (4–5×2–2.5 μm) and distinctive spear-like cheilo- and pleurocystidia.

Cystodermella granulosa (Batsch: Fr.) Harmaja, Kar-stenia 42 (2): 46. 2002.

Mycol Progress (2009) 8:59–73 69

≡ Agaricus granulosus Batsch, Elench. Fung.: 79. 1783.≡ Agaricus granulosus Batsch: Fr., Syst. mycol. 1: 24. 1821.≡ Lepiota granulosa (Batsch: Fr.) S.F. Gray, Nat. Arr. Br.

Pl. 1: 602. 1821.≡ Lepiota granulosa (Batsch: Fr.) P. Kumm., Führer

Pilzk.: 136. 1871.≡ Armillaria granulosa (Batsch: Fr.) Kauffman, Pap.

Michigan Acad. Sci. 2: 60. 1923.≡ Cystoderma granulosum (Batsch: Fr.) Fayod, Ann.

Sci. Nat., Bot., Sér. VII 9: 351. 1889.=Cystodermella ambrosii (Bres.) Harmaja, Karstenia 42

(2): 45. 2002.The type species of the genus Cystodermella is charac-

terised by brownish or whitish pileus and stipe, as well asby inamyloid oblong basidiospores (3.5–5×2–3 μm).

Cystodermella granulosa (Batsch: Fr.) Harmaja var.ambrosii (Bres.) I. Saar, comb. nova

Basionym: Armillaria ambrosii Bres., Fungi Tridentini1: 27. 1881.

Synonym: Cystoderma ambrosii (Bres.) Singer, Ann.Mycol. 41: 170. 1943.

Cystoderma ambrosii (Bres.) A.H. Sm. & Singer, Pap.Michigan Acad. Sci. 30: 81. 1945 (superfluous name, Art.52.1).

Cystoderma granulosum (Batsch: Fr.) var. albidum(Peck) Heinem. & Thoen., Bull. Trimestriel Soc. Mycol.France 89: 31. 1973.

Epitype: Finland, Koillismaa, Juuma, at the margin ofeutrophic fen, in tussock, 23 Aug 1974, leg. T. Ulvinen (H).

The white variety of this species is distinguished bysphaerocysts that are hyaline in KOH. There is no typematerial by Bresadola in BPI, M nor S herbaria. Hence,because there is no known lectotype, the iconotype(Bresadola 1881: pl. 31) is considered here as the holotype.

Cystodermella granulosa (Batsch: Fr.) Harmaja var.granulosa

Epitype: Estonia, Tartu Co., Mäksa Comm., Metsanurga,in mixed pine-spruce forest, on moss, 6 Aug 2000, leg. I.Saar (TAA 147491).

The type variety is characterised by a pileus in shades ofbrown. There is no type specimen. Singer (1961) proposedthat the form b of Fries (1821) with its respective referenceto Batsch (1783: tab. 6, 24) should be regarded as thelectotype of the species.

Cystodermella papallactae (I. Saar & Laessøe) I. Saar,comb. nova

≡ Cystoderma papallactae I. Saar & Laessøe, Myco-taxon 96: 126. 2006.

Type studied: Ecuador, Prov. Napo, just below and E ofthe Papallacta pass, páramo El Virgen (00°19′66″S 78°12′06″W), in moss under big Polylepis tree, alt. ca. 3800 m, 7Apr 2002, leg. T. Læssøe TL-9250 (QCA, holotype; C58002, isotype).

This species, collected in Ecuador at high elevations(Saar and Læssøe 2006), is characterised by brownish-orange pileus and large inamyloid, ellipsoid to oblongbasidiospores (5.0–6.0×3.0–4.0 μm).

Discussion

Polyphyletic genus Cystoderma sensu Singer

The genus Cystoderma sensu Singer (1986) appearedpolyphyletic in all analyses based on partial LSU rDNAsequences (Fig. 1). The genus Cystodermella, segregatedfrom Cystoderma by Harmaja (2002), was supported in allanalyses based on partial LSU and ITS sequences. In thisrespect, our results concur with those obtained in theanalyses of LSU data in recent studies (Johnson andVilgalys 1998; Moncalvo et al. 2002) that incorporatedsome members of the group treated here. In both studies,Cystodermella granulosa (as Cystoderma) appeared closelyrelated with Ripartitella. Cystoderma chocoanum formed asister group of the Cystodermella-Ripartitella clade(Johnson and Vilgalys 1998). In the analyses by Moncalvoet al. (2002) Cystoderma chocoanum and C. amianthinumformed a well-supported clade, with Floccularia albola-naripes as their sister group.

Cystoderma sensu Harmaja and its closest relatives

In the Bayesian analysis of the LSU data, the genusCystoderma sensu Harmaja (2002) appears monophyleticwith the inclusion of C. superbum as its basal lineage.Testing the constraint hypothesis, however, could not rejectthe inclusion of C. superbum from the Cystoderma clade.Heinemann and Thoen (1973) regarded it as the singlespecies of the section Superba, delimited by spores that areamyloid only at the suprahilar zone. Singer (1986) treatedthis species as a member of the section Cystoderma,corresponding to Cystoderma by Harmaja (2002). Peglerand Young (1971) demonstrated that in C. superbum thewhole spore is weakly grey amyloid except for thesuprahilar region, which is strongly amyloid. Consideringthe results of the phylogenetic analyses, C. superbum isregarded as a member of the genus Cystoderma. Phaeole-piota aurea forms the unsupported sister group of Cysto-derma, with the Phaeolepiota-Cystoderma clade beingsupported only by the posterior probability values in theLSU rDNA analyses. Phylogenies based on ITS rDNAdata, however, include P. aurea in Cystoderma. As thisrelationship does not receive considerable support, evalua-tion of the combination of Phaeolepiota in Cystoderma asproposed by Kühner and Romagnesi (1953) will be left forfuture studies.

70 Mycol Progress (2009) 8:59–73

Members of Phaeolepiota and Cystoderma share similarfeatures of the epithelial covering of pileus and stipe. Achemotaxonomical study by Benedict et al. (1972) showsthat the distribution patterns of urea and ten chromato-graphically measured compounds supports a close relation-ship between these two genera.

The recent study on the phylogeny of Agaricales byGarnica et al. (2007) support a close relationship betweenthe monotypic genus Phaeolepiota and the type species ofthe Cystoderma (C. amianthinum), which form a signifi-cantly supported clade with the Crucibulum-Cyathuslineage. The multi-gene phylogeny presented by Mathenyet al. (2006), likewise involving only the type species ofCystoderma, reveals the Cystoderma-Crucibulum-Cyathuslineage as a moderately supported sister group of theAgaricaceae in the Agaricoid clade. Walther et al. (2005)reported the similarity of the modes of conidiogenesisbetween Cystoderma and the representatives of Agarica-ceae, referring to a close relationship between these taxa. Incontrast to the majority of the studied Agaricaceae, theconidia of Cystoderma did not usually swell. Results of theanalyses of LSU data obtained in the present study, basedon more extensive species sampling, do not support a closerelationship between the species of Cystoderma and theCyathus-Crucibulum lineage.

Cystodermella Harmaja and its closest relatives

The genus Cystodermella appeared monophyletic in allanalyses, receiving considerable support except for theBayesian analysis of LSU data. MP analysis of the LSUdata supported the sister group relationship betweenCystodermella and Ripartitella, concordantly to the resultof an earlier phylogenetic study (Moncalvo et al. 2002).Likewise, the culture studies by Pantidou et al. (1983) and(Ovrebo 1988) indicated a close relationship betweenCystodermella and Ripartitella. The representatives of bothgenera shared similar conidiogenesis: conidiophores werenot differentiated; arthroconidia were formed by thefragmentation of hyphae. However, the conidia of Cys-toderma are also formed in the same way (Walther et al.2005). Ripartitella differs from Cystodermella by echinu-late spores, which are similarly inamyloid, also in havingtrichodermium instead of epithelium in pileus and stipe(Singer 1947: 85).

Morphological delineation of Cystoderma versusCystodermella

In infrageneric taxonomy of the group, the spore wallamyloidity has been used extensively as a definingcharacteristic. The results presented in this study revealthat amyloidity of spores, considered consistent within the

treated genera (Harmaja 2002), varies in Cystoderma.Harmaja transferred Cystoderma japonicum to Cystoder-mella, because it produces inamyloid basidiospores. Ourresults, however, show that it belongs to Cystoderma.Based on the first author’s experience, determining the stateof this characteristic is complicated because most spores arecomparatively small and the colouring reaction is weak.Therefore this characteristic alone cannot be considered as areliable feature to distinguish the species.

Based on the studied species, the genus Cystoderma canbe defined by the formation of amyloid or inamyloidbasidiospores, by having a persistent membranous ring onupper part of the stipe (Cystoderma japonicum). In contrast,species of Cystodermella are characterised by inamyloidspores, evanescent floccose-scaly ring, and the presence ofcystidia in some cases.

Further studies are needed to understand the relation-ships between species not treated in this study. Consideringonly morphology can result in misleading generic relation-ships as recently exemplified by relocating Cystodermellaluteohemisphaerica (Dennis) Harmaja into the genusCystolepiota Singer (Saar and Læssøe 2008).

Tribe Cystodermateae

The tribe Cystodermateae sensu Singer (1986) appearedparaphyletic according to the analyses of LSU rDNAsequences. The genus Pseudobaeospora appeared to beclosely related to the genera Tricholoma and Lepista,concordant to Kühner (1980) who included it in the tribeTricholomateae.

The phylogenetic relationships among the genera in-volved could not be unequivocally resolved based onpartial LSU sequences only. Lack of support in thebackbone of the trees based on LSU data indicate the needfor further studies on the tribe Cystodermateae implyingphylogenetic analyses of multiple genes.

Acknowledgements The authors are indebted to the directors andcurators of the herbaria BR, C, H, K, MICH, NY, O, PDD, REG, S, TAA,TU andWU for the loan of specimens. Sincere thanks are due to ThomasLæssøe and Rasmus Kjøller for their comprehensive help during a stay inCopenhagen. We thank our colleagues, Kuulo Kalamees, Erast Parmastoand Robert Szava-Kovats, for their helpful comments and suggestions.This study was supported by the Estonian Science Foundation grant no.6606 and 6939, and the first author’s work in Copenhagen by a grant fromthe European Commission’s (FP 6) Integrated Infrastructure Initiativeprogram SYNTHESYS (DK-TAF 527).

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