Assembling the fungal tree of life: progress, classification, and evolution of subcellular traits

FRANÇOIS LUTZONI, FRANK KAUFF, CYMON J. COX, DAVID MCLAUGHLIN, GAIL CELIO, BRYN DENTINGER, MAHAJABEEN PADAMSEE, DAVID HIBBETT, TIMOTHY Y. JAMES, ELISABETH BALOCH, MARTIN GRUBE, VALÉRIE REEB, VALÉRIE HOFSTETTER, CONRAD SCHOCH, A. ELIZABETH ARNOLD, JOLANTA MIADLIKOWSKA, JOSEPH SPATAFORA, Materials and Methods DESIREE JOHNSON, SARAH HAMBLETON, MICHAEL CROCKETT, ROBERT SHOEMAKER, GI-HO SUNG, ROBERT LÜCKING, THORSTEN LUMBSCH, KERRY O’DONNELL, MANFRED BINDER, Taxon sampling Ð For the nucSSU + nucLSU (≈2.4kb, Fig. 1) data set, a total of 13 467 GenBank PAUL DIEDERICH, DAMIEN ERTZ, CÉCILE GUEIDAN, BENJAMIN HALL, KAREN HANSEN, RICHARD C. HARRIS, KENTARO HOSAKA, YOUNG-WOON LIM, YAJUAN LIU, BRANDON MATHENY, sequences were considered, of which 1010 unique taxa had both sequences available. Sequences that were selected HIROMI NISHIDA, DON PFISTER, JACK ROGERS, AMY ROSSMAN, IMKE SCHMITT, HARRIE SIPMAN, JEFFREY STONE, JUNTA SUGIYAMA, REBECCA YAHR, AND RYTAS VILGALYS. incorrectly due to inconsistencies in the GenBank record “Definition Line” were discarded, as were sequences whose length was < 600 base pairs or whose overlap with other taxa was < 600 base pairs. Unpublished sequences available directly from the AFTOL project and laboratories associated with this project were combined with those available from GenBank and were included in preference to GenBank data. For the nucSSU + nucLSU + RPB2 (≈4.6kb, Fig. 2) data set, RPB2 sequences for 19 taxa were obtained from the AFTOL project and laboratories associated with this study. We queried GenBank using the EPU for RPB2 data for each of the remaining taxa 100 Lobaria quercizans 100 100 Lobaria scrobiculata present in the combined nucSSU + nucLSU data set, but not available from AFTOL. One hundred forty-two taxa 100 93 100 Peltigera canina Two-locus Bayesian MCMCMC tree 100 Peltigera degenii were retrieved from GenBank, such that the nucSSU + nucLSU + RPB2 data set consisted of 161 taxa. For the Nephroma parile 62 Xanthoria elegans nucSSU + nucLSU + mitSSU + RPB2 (≈5.4kb, Fig. 3) data set, mitSSU sequences for 105 taxa were obtained nucSSU+nucLSU 99 Letrouitia domingensis 78 100 100 fallax from the AFTOL project. For each of the remaining taxa not available directly from AFTOL but present in the 558 species, 430 genera, 68 orders, 5 phyla 86 75 Teloschistes exilis Megalospora tuberculosa combined nucSSU + nucLSU data set, we queried GenBank for mitSSU using the EPU. One hundred forty-eight 100 Leptogium cyanescens Placynthium nigrum "Gyalideopsis vulgaris"? taxa were retrieved, such that the final nucSSU+nucLSU+mitSSU data set consisted of 253 unique taxa. Taxa 99 Scoliciosporum umbrinum Bayesian posterior probability 93 Lopezaria versicolor common to these three preceding data sets were combined, resulting in 107 unique taxa forming this four-locus NJ bootstrap proportion Lecanora concolor 100 59 Sphaerophorus globosus data set. Crocynia pyxinoides 51 Canoparmelia caroliniana Parmotrema tinctorum 0.1 substitutions/site Punctelia hypoleucites Phylogenetic analyses Ð Bayesian Metropolis coupled Markov Chain Monte Carlo 67 100 95 Punctelia rudecta Stereocaulon paschale (B-MCMCMC) analyses were conducted with MrBayes v3.0b4 (Huelsenbeck and Ronquist, 2001). All Cladonia caroliniana Flavoparmelia caperata B-MCMCMC analyses were conducted using four chains, and a gamma distribution, if applicable, was 100 Menegazzia terebrata Xanthoparmelia conspersa approximated with four categories. In addition to posterior probabilities (PP), phylogenetic confidence was Porpidia albocaerulescens 100 Sporostatia polyspora estimated with weighted maximum parsimony bootstrap proportions (MPBP), neighbor joining bootstrap 99 80 Sporostatia testudinea disporum Lecanoromycetidae proportions (NJBP) with maximum likelihood (ML) distance implemented using PAUP* 4.0b.10 (Swofford, 2002), Acknowledgment 100 canadensis 92 100 Sarcogyne regularis and by analyzing bootstrapped data sets with B-MCMCMC (i.e., Bayesian bootstrap proportions, BBP; Douady We thank Molly McMullen and Connie Robertson (Duke University 99 Acarospora macrospora 61 100 100 Acarospora cervina et al., 2003). Step matrices for weighted parsimony analyses were generated using stepmatrix.py (written by 75 Acarospora laqueata Herbarium) for curating specimens used for this study, Josée Bélisle Sarcogyne similis 100 Acarospora complanata F. Kauff and available upon request from FK or FL). Uninformative characters were excluded from all for graphic design of phylogenetic trees, as well as Snehal Sarvate and 58 54 97 Acarospora hilaris Glypholecia scabra bootstrapped data sets analyzed with MP. Parsimony ratchet search strategies (PAUPRat; Nixon, 1999; Sikes and Acarospora bullata Lindsay Higgins for assistance in preparing appendices. We are very 98 Acarospora schleicheri Lewis, 2001, http://www.ucalgary.ca/~dsikes/software2.htm) were implemented in PAUP*. Bootstrapped data sets Acarospora clauzadeana 100 thankful to John Harer, Bill Rankin, and John Pormann for providing 63 Polysporina simplex subjected to B-MCMCMC analyses were generated with P4 0.78 (Foster, 2003). For each data partition and for the Thelocarpella gordensis access to the Duke C.S.E.M. computer cluster, and to Eric Sills Pleopsidium chlorophanum Acarosporomycetidae combined data set, a hierarchical likelihood ratio test (Modeltest 3.06; Posada and Crandall, 1998) was used to 100 Gyalecta jenensis 100 97 Gyalecta hypoleuca (Information Technology Division at North Carolina State University) and "Trapeliopsis granulosa" ? determine the appropriate model (nucleotide substitution and rate heterogeneity parameters). For each NJ analysis, 100 Coenogonium leprieurii parameter values were fixed to the optimal values calculated for the optimal model. For the RPB2 data set, each Sudhakar Pamidighantam (National Center for Supercomputing 100 100 Coenogonium luteum 50 Petractis luetkemuelleri Gyalecta ulmi codon position was subjected to a separate model in the B-MCMCMC analysis. Following the recommendation in Applications) for installing software to enable us to conduct many of the 63 Diploschistes scruposus 54 100 Diploschistes rampoddensis Reeb et al. (2004), we used NJBP (500 replicates) to detect topological conflicts among data partitions. A conflict analyses essential for this study. Thanks to Joyce Longcore, and John 100 100 Diploschistes thunbergianus 100 Graphina poitiaei was assumed to be significant if two different relationships (one monophyletic the other nonmonophyletic) for the Taylor for providing fungal strains. This publication resulted in part from 100 Stictis radiata Acarosporina microspora same set of taxa were both supported with bootstrap values ≥ 70% (Mason-Gamer and Kellogg, 1996). The 100 Placopsis argillacea the Assembling the Fungal Tree of Life (AFTOL) project, which is 75 100 75 Placopsis gelida program compat.py (written by F. Kauff and available upon request from FK or FL) was used to detect such 100 100 Trapelia involuta supported by NSF Assembling the Tree of Life (ATOL) awards 98 Trapelia placodioides topological incongruences. Taxa causing conflicts were removed, and the test was reimplemented until no conflicts 70 Ionaspis lacustris Ostropomycetidae 2 100 Maronea chilensis DEB-0228725 to JS, DEB-0228657 to DH, DEB-0228671 to DJM, and 100 100 Maronea constans were detected. Each locus in the combined data sets was subjected to this incongruence test for all possible 100 Fuscidia lygaea DEB-0228668 to FL and RV, as well as, DEB-9615542 and DEB-0133891 91 Baeomyces placophyllus pairwise comparisons prior to inclusion. 100 Coccotrema coccophorum to FL, DEB-0105194 to FL and VR, DEB-9903835 to DH, DEB 0128925 64 100 Coccotrema cucurbitula 100 Coccotrema pocillarium Results 79 Ochrolechia parella to DH and MB, DEB- 0129212 to JS, DEB-9306578 and DEB-9318232 98 100 93 Ochrolechia szatalaensis 72 97 Ochrolechia juvenalis 100 100 Pertusaria erythrella Phylogenetic relationships among fungal phyla (Fig. 1) Ð The Fungi 100 to DJM, DEB-9813304 to JR, FWF P16410 and P19052 to MG. We also 100 99 Pertusaria scaberula 100 Pertusaria amara were resolved as a clade with a 100% Bayesian posterior probability (PP) with respect to the animalian outgroup gratefully acknowledge support of the NSF Research Coordination 100 Dibaeis baeomyces 100 Thamnolia subuliformis Ostropomycetidae 1 taxa. Both the and formed clades supported by a PP of 100% and NJBP of 67% and Networks in Biological Sciences: A Phylogeny for Kingdom Fungi Umbilicaria muelhenbergii 100 Ascosphaera apis 100 Eremascus albus 93%, respectively, in the nucSSU + nucLSU analysis. Furthermore, a sister relationship between the Basidiomycota (NSF 0090301). 92 100 Paracoccidioides brasiliensis and Ascomycota (the “Dikaryomycota”) received medium support (PP = 100% and NJBP = 54%). The 91 83 Byssochlamys nivea 100 Hamigera avellanea formed a clade (PP = 100% and NJBP = 98%) sister to the “Dikaryomycota.” This clade has often 100 Penicillium viridicatum 96 61 Aphanoascus fulvescens 76 75 Auxarthron zuffianum been recovered in nuclear rDNA phylogenetic analyses (Sugiyama, 1998; Schüßler et al., 2001; Tehler et al., 2003). 100 52 Arthroderma curreyi 71 Coccidioides posadasii It has recently been given the informal name “Symbiomycota” because most of its members form symbioses (Tehler Arachniotus ruber Eurotiomycetidae 64 98 mansonii et al., 2003), but statistical support for this clade in this (PP = 90%) and other studies, has never been achieved. 100 50 Exophiala jeanselmei 100 71 Capronia pilosella 94 97 Ceramothyrium carniolicum 100 Glyphium elatum Relationships among and within , , Introduction and objectives 90 87 100 luridum 98 100 65 Dermatocarpon miniatum Despite the numerous technological advancements 98 Verrucaria pachyderma and Glomeromycota (Fig. 1) Ð The nucSSU + nucLSU phylogeny resolves a chytrid clade 94 100 Pyrenula cruenta 88 Pyrenula pseudobufonia Chaetothyriomycetidae sister to all fungi and a paraphyletic assemblage of zygomycete lineages + Allomyces, which form a grade leading 100 Strangospora pinicola available to fungal systematists, progress in 100 Strangospora pinicola Lasallia pensylvanica to the Glomeromycota sister to the “Dikaryomycota.” Of the five orders of these groups represented Discula destructiva understanding the deepest nodes in the fungal tree Gnomoniella fraxini by more than a single taxon, only one of these is monophyletic (). The nucSSU + nucLSU analyses 100 Gnomonia setacea 97 of life will be limited without a new approach to 87 Melanconis marginalis (Fig. 2) provide a conservative estimate, which is fully resolved (though not well supported), of the relationships 59 Plagiostoma euphorbiae 86 Valsa ambiens among the earliest branching fungal lineages and provide new insight into a few critical branching events. The conducting large-scale multilocus phylogenetic studies 60 Cryptodiaporthe corni 100 Cryphonectria cubensis divergence of the two Basidiobolus spp. within the Zygomycota + Allomyces group is more consistent with the 99 100 Cryphonectria havanensis Diaporthe phaseolorum ecological and morphological traits of these fungi than the placement of the within the chytrid lineage. and phenotype-based comparative studies on Fungi. 99 Gaeumannomyces graminis var. graminis Thyridium vestutum Monoblepharella sp. (representing the Monoblepharidales) groups within the basal-most fungal clade of chytrid This novel approach will require concerted data 100 Camarops microspora 100 77 Menispora tortuosa 100 91 Neurospora crassa fungi (though this position is unsupported), that also includes the orders Chytridiales and Spizellomycetales. In acquisition by focusing sequencing efforts on specific 98 100 Sordaria macrospora 100 88 Sordaria fimicola contrast, the Blastocladiales, represented by Allomyces arbusculus, group with the remainder of the the Fungi 69 Chaetomium globosum loci and fungal taxa, by conducting phenotypic studies 100 96 Farrowia longicollea rather than with the other 78 Farrowia seminuda Chaetomium globosum on specific fungal traits, by improving interaction Cephalotheca sulfurea Relationships within Ascomycota and Basidiomycota (Figs. 1-3) 100 Astrocystis cocoes 88 Rosellinia necatrix As a consequence of the lack of complementarity among single-locus data sets, the three- and four-locus analyses among fungal systematists, and by the automation of 97 60 Xylaria acuta 65 75 Xylaria hypoxylon 65 Graphostroma platystoma included only members of the Ascomycota and Basidiomycota. The four-locus analysis resolved multiple deep data acquisition and analysis coupled with data bases 100 Hypoxylon fragiforme 100 Fasciatispora petrakii relationships within the Ascomycota and Basidiomycota that were not revealed previously or that received only Halorosellinia oceanica 97 accessible through the World Wide Web. These goals 100 Cainia graminis weak support values in previous studies. 59 Seynesia erumpens 58 Cryptosphaeria eunomia var. eunomia form the framework of the Assembling the Fungal Tree 100 Apiospora sinensis Arthrinium phaeospermum Publication 100 100 100 69 Hyponectria buxi of Life Project (AFTOL), which seeks to infer the Microdochium nivale The impact of this newly discovered phylogenetic structure on supraordinal classifications, the results and reanalysis Oxydothis frondicola 84 Cordyceps irangiensis of subcellular data, a synthesis of current knowledge regarding the evolution of septal features of fungal hyphae, phylogenetic relationships among 1500 species 73 Emericellopsis terricola 66 Cordyceps cylindrica and a preliminary reassessment of ascomal evolution will be published in the October (2004) issue of the American Torrubiella luteorostrata representing all fungal phyla based on eight loci 100 86 94 Verticillium epiphytum Journal of Botany. 97 Balansia henningsiana ≈ Cordyceps khaoyaiensis ( 10 kb). Here, we report phylogenetic studies for the 93 Cordyceps sinensis 64 Cordyceps militaris 100 61 Cordyceps pseudomilitaris maximal number of species across all known fungal 73 60 Cordyceps tuberculata 100 100 Hydropisphaera erubescens 87 Cordyceps crassispora phyla for which DNA sequence data from two, three, 100 Hypomyces polyporinus 60 Hypocrea citrina 79 Halosarpheia marina 76 and four loci are available. The resulting phylogenetic 100 64 Natantispora lotica 100 97 100 Natantispora retorquens 80 75 Ascosacculus heteroguttulatus trees are based on sequences available in GenBank and 100 Corollospora maritima 92 Varicosporina ramulosa 100 Sagaaromyces abonnis unpublished sequences generated by various 99 100 100 Magnisphaera stevemossago 70 80 Ascosalsum cincinnatulum 71 Microascus trigonosporus laboratories or by the AFTOL project. This study Pleurothecium recurvatum 100 Lulworthia fucicola 100 Lulworthia grandispora Sordariomycetidae provides a phylogenetic synthesis for the Fungi and a Kirschsteiniothelia aethiops 100 Setosphaeria monoceras 87 100 94 Cochliobolus heterostrophus framework for future phylogenetic studies on fungi. 100 78 Curvularia brachyspora 90 Pyrenophora tritici-repentis 87 Ampelomyces quisqualis 55 Phoma glomerata Phaeosphaeria avenaria 62 Setomelanomma holmii 80 Byssothecium circinans Lojkania enalia 61 52 Trematosphaeria heterospora 81 Westerdykella cylindrica Preussia terricola 100 Xanthoparmelia conspersa 100 97 /67 Parmotrema tinctorum 100 Bimuria novae-zelandiae 100 Canoparmelia caroliniana Letendraea helminthicola Three-locus Bayesian MCMCMC tree 100 100/100 Punctelia hypoleucites Other presentations at this meeting 51 "Lithothelium septemseptatum" ? 98 100/100 Flavoparmelia caperata Phaeotrichum benjaminii Dothideomycetidae 1 84 Lecanora concolor nucSSU+nucLSU+RPB2 100 100 Roccella boergesenii 85/54 100 Cladonia caroliniana 78 99 Roccella fuciformis 97/90 Stereocaulon paschale associated with this poster: 75 Schismatomma decolorans 157 species, 134 genera, 34 orders, 2 phyla 100 Sphaerophorus globosus 100 Dendrographa minor 100 Xanthomendoza fallax 98 100/100 Xanthoria elegans 100 99 Dendrographa leucophaea Lecanactis abietina 88 Calicium viride 61 100 Peltigera degenii Arthonia dispersa Arthoniomycetidae 1) Valerie Hofstetter, J. Miadlikowska, C. Gueidan, 100 100/100 Peltigera canina 65 Peltula umbilicata 100 85/87 e 100 60 Lobaria quercizans a 84 Peltula obscurans Bayesian posterior probability id /73 100 Leptogium cyanescens t 73 100 e V. Reeb, A. E. Arnold, F. Kauff, C. Cox, R. Yahr Lempholemma polyanthes c 81 90/96 Placynthium nigrum y Piedraia hortae NJ bootstrap proportion / MP bootstrap proportion 100 100 Sporostatia testudinea m 95 ro Myriangium duriaei 95 Sporostatia polyspora o 100/100 n & F. Lutzoni. What do protein-coding genes 88 a Microxyphium citri Rhizocarpon disporum c e Raciborskiomyces longisetosum Dothideomycetidae 2 0.1 substitutions/site Porpidia albocaerulescens L (ATP6, EF 1-alpha and RNA polymerase II) bring Dothidea ribesia 100 Coenogonium leprieurii 100 99 Stylodothis puccinioides 100 100/100 Coenogonium luteum 51 86 94 Dothidea sambuci 67 Petractis luetkemuelleri 58/ 100 Diploschistes scruposus to molecular systematics of ? Tuesday 100 91 Sydowia polyspora Delphinella strobiligena 99 100/100 Graphina poitiaei 100 89 67/ Discosphaerina fagi 88 100 Gyalecta hypoleuca August 17, Poster Session 1: ÊSystematics and 100 100 100/100 Gyalecta jenensis Botryosphaeria ribis Dothideomycetidae 3 74/84 61/90 Gyalecta ulmi 100 Cudonia circinans 100 100 Acarosporina microspora Evolution, 18:00-19:00. 100 95 Spathularia flavida 100 99 95/86 Stictis radiata 71 Coccomyces dentatus Placopsis perrugosa 100 100 e 99 a Tryblidiopsis pinastri Ionaspis lacustris d ti "Ostropa barbara" ? e 100 Baeomyces placophyllus c Rhytisma acerinum 100 Dibaeis baeomyces y 83 100 m 82 o 2) Frank Kauff, Cymon Cox & François Lutzoni. 83 "Trypethelium" ? 100 100/100 Thamnolia subuliformis p o 83 "Chloroscypha" ? 98 91/92 100 Pertusaria amara tr s Lachnum bicolor 65/70 Ochrolechia juvenalis O Bioinformatics of AFTOL (Assembling the Fungal Lachnum virgineum 100 Lasallia pensylvanica Leveillula taurica 99 100/100 Umbilicaria muelhenbergii 91 /55 100 Fuscidia lygaea Tree of Life). ÊTuesday August 17, Poster Session 1: Microsphaera friestii var. dahurica Maronea chilensis 100 Uncinula mori 100/100 91 100 Strangospora pinicola Phyllactinia moricola 100/100 Strangospora pinicola Systematics and Evolution, 18:00-19:00. 53 98 Golovinomyces orontii 84 100 Acarospora canadensis Sphaerotheca cucurbitae 100 93/100 Sarcogyne regularis Chaetomella acutiseta 100 92/100 Acarospora macrospora 57 Chlorociboria aeruginosa 100 /99 100 Acarospora laqueata 87/98 Acarospora cervina Leotia lubrica Eurotiomycetes 51 80/100 3) ÊCymon Cox & Frank Kauff. ÊA data management 100 Botryotinia fuckeliana Glypholecia scabra 62 e Sarcogyne similis a 100 Monilinia fructicola /69 d Pleopsidium chlorophanum ti e Crinula caliciiformis 100 c framework for the AFTOL project.ÊSaturday, 100 Acarospora complanata y Neofabraea malicorticis 1 100/71 100/99 Acarospora hilaris m Biatoridium monasteriense 55 100 100 ro 100 Acarospora schleicheri o 100 79/100 100/98 p e August 21, Discussion session 3: ÊPhylogenetic 100 97 Sarcosagium campestre 100 s a 100/100 Acarospora clauzadeana o d 100 96 r ti Thelocarpon laureri 100/100 Acarospora bullata a e c c 100 Trichoglossum hirsutum Polysporina simplex A y methods 15:00-17:50. m 100 97 Trichoglossum hirsutum 100 Hamigera avellanea io Leotiomycetes 2 t e 96 Geoglossum nigritum 100 100/100 Penicillium freii ro a u d 100/100 ti Calicium viride Byssochlamys nivea E e c Sarea resinae 83 Capronia mansonii y Leotiomycetes 55 100 m 55 /99 Capronia pilosella o 86 Morchella esculenta 100 ri 4) François Lutzoni et al. Where are we in 100 100/100 Exophiala jeanselmei y Morchella esculenta 100 100/95 th Dermatocarpon luridum o 77 t 100 Morchella elata /100 e 100 Pyrenula pseudobufonia a assembling the fungal tree of life, classifying the 96 59 Disciotis venosa 100/100 Pyrenula cruenta h Verpa conica C 100 Lempholemma polyanthes Gyromitra californica 100 70 81 100/100 100 Peltula umbilicata Lichinomycetes Fungi and understanding the evolution of their Gyromitra melaleucoides 51 100/100 100 Peltula obscurans Barssia oregonensis 97 Biatoridium monasteriense subcellular traits? Ê Saturday, August 21, 71 Helvella compressa Thelocarpon laureri 66 56 Aleuria aurantia Trichoglossum hirsutum 84 100 62 Cheilymenia stercorea 96 Rhytisma acerinum Discussion session 3: ÊPhylogenetic methods 100 91 Otidea onotica 99 "Ostropa barbara" ? 98 100 71 "Anthracobia" ? 100 100 84/ 100 Sclerotinia sclerotiorum Leotia lubrica 72 100 "Pithya" ? /88 89/70 70/ 15:00-17:50. 96 Cudonia circinans 100 Sarcoscypha coccinea Dendrographa minor 70 Peziza succosa 100 100 97 100/100 Lecanactis abietina Arthoniomycetidae 97 Sarcosphaera crassa /69 100 Curvularia brachyspora Dothideomycetidae 94 Peziza proteana 98 Hydropisphaera erubescens 67 100 Peziza quelepidotia 100 Cordyceps militaris 100 /80 66 100 Ascobolus carbonarius 98 100 Hypocrea citrina 98 58/58 e 100 Ascobolus crenulatus /69 Microascus trigonosporus a d Saccharomyces cerevisiae 100 Neurospora crassa ti 82 69 e 100 100/100 c 100 Saccharomyces cerevisiae Sordaria macrospora y 100 57/ 100 Cryptodiaporthe corni m Ascomycota 100 Saccharomyces cerevisiae 100 o 100 100 /62 Discula destructiva ri 98 100 Candida albicans 100/100 a 100 /52 68/96 Diaporthe phaseolorum rd 100 100 Candida albicans o Seynesia erumpens S Arxula adeninivorans Ascomycota Morchella esculenta vitellina 100 100 Pezizomycetes Interpretation of 100 100/100 Morchella elata /63 67 98 Taphrina communis Peziza quelepidotia 100 85 Taphrina wiesneri PEZIZOMYCOTINA 100 Candida albicans 97 support values 100 Protomyces inouyei 69 100 100/100 Candida albicans SACCHAROMYCOTINA 69 Saitoella complicata /72 100/100 100 Saccharomyces cerevisiae 100 Schizosaccharomyces pombe 100/100 Saccharomyces cerevisiae 100 Schizosaccharomyces pombe 100 Taphrina communis 100 Taphrina wiesneri Pneumocystis carinii 99/100 TAPHRINOMYCOTINA Piriformospora indica cantharelloid ? 100/100 Protomyces inouyei Plicaturopsis crispa Saitoella complicata 99 Cryptoporus volvatus 54 100 Fistulina antarctica 85 99 Ganoderma australe Posterior 91 Fistulina hepatica Lentinus tigrinus 100 Porodisculus pendulus 93 95 98 74 99 Polyporus melanopus id Fistulina pallida ro 100 Polyporus squamosus o 89 Panellus serotinus Lenzites betulina p Probabilities ly o Typhula phacorrhiza 99 100 Pycnoporus cinnabarinus p Ossicaulis lignatilis Trametes versicolor re o 98 Halocyphina villosa Dentocorticium sulphurellum c 100 83 Nia vibrissa Albatrellus syringae 100 97 92 Calathella mangrovei 54 98 Oligoporus rennyi 66 100 At Cyphellopsis anomala 97 Oligoporus lacteus Crucibulum laeve Antrodia carbonica Chondrostereum purpureum 51 89 Auriporia aurea ≥ 70 73 Ceriporia purpurea least 95% < 95% 58 Physalacria maipoensis Phlebia radiata 55 Henningsomyces candidus 100 57 60 Fomitopsis pinicola Limnoperdon incarnatum 79 55/ Piptoporus betulinus one BP Panellus stypticus 74 Neolentiporus maculatissimus 88 Tricholoma inamoenum 100 Laetiporus sulphureus 65 93 Tricholoma myomyces 81 Phaeolus schweinitzii 50 Sparassis brevipes id Leucopaxillus albissimus 100 o id Figure 2. Phylogenetic relationships among 100 97 Sparassis crispa r ro Lentinula lateritia 93/82 o o p h 69 Humidicutis marginata Grifola frondosa ly p o le 157 ascomycete and basidiomycete taxa 53 83 Meripilus giganteus p e Hygrophorus sordidus h Pleurotus tuberregium /77 Sarcodon imbricatus t 91 "Athelia bombacina" ? based on combined evidence from nucSSU, ≥ 61 Agaricus sylvaticus 51 70% High Medium 100 /51 Nia vibrissa Laccaria pumila 61 Chondrostereum purpureum nucLSU, and RPB2. This phylogeny resulted 78 Cortinarius bolaris 99 Typhula phacorrhiza 62 Cortinarius iodes Lentinula lateritia Crepidotus variabilis 100 from a 50% majority rule consensus of 99 54 Amanita muscaria s c Inocybe geophylla 95 Inocybe geophylla ri a g 20 000 trees sampled with Bayesian 53 Hygrophorus eburneus 86 82 Entoloma strictius a u 70% Lichenomphalia umbellifera Plicaturopsis crispa e Amanita muscaria 93 Boletus satanas MCMCMC. 100 te cavipes le Cyathus striatus o ≥ Entoloma strictius 50 Coniophora arida b > BP Medium Low Albatrellus skamanius The resulting posterior probabilities (PP) Hygrocybe conica Amylostereum laevigatum Laccaria amethystina 100 Echinodontium tinctorium are shown above internal branches. NJ Pluteus petasatus euagarics 95/100 50% 98 97 100 Peniophora nuda 100 Calostoma cinnabarinum 97 97/99 Scytinostroma alutum 100 Basidiomycota 67/61 bootstrap proportions (NJBP) with ML 100 Calostoma ravenelii /60 98 Auriscalpium vulgare 91 100 / 65 Scleroderma citrinum 64/60 Russula compacta id 100 id to distance (1000 bootstrap replicates) are Boletus satanas 83 Bondarzewia montana o e l a Heterobasidion annosum u h Xerocomus chrysenteron s c s o 97 96 Chroogomphus vinicolor Stereum hirsutum ru n shown below internal branches before the 100 Coltricia perennis e < 50% Low No 92 82 Gomphidius glutinosus 85 m /73 y 100 Suillus cavipes Hyphodontia alutaria h slash sign, and weighted MP bootstrap 100 100 Gautieria otthii 100 Suillus sinuspaulianus 100 gomphoid-phalloid 98 Rhizopogon subcaerulescens 74/ Ramaria formosa support /100 100 Botryobasidium isabellinum proportions (MPBP) are shown below 100 Coniophora arida cantharelloid 100 /74 Sistotrema eximum 50 100 56 Coniophora puteana Dacrymyces chrysospermus internal branches after the slash sign. Coniophora marmorata Dacrymycetales 62 96 Coniophora olivacea 56 Tapinella panuoides Pseudomerulius aureus

Tapinella atrotomentosa bolete 64 Repetobasidium mirificum hymenochaetoid 2 Jaapia argillaceae Lopharia mirabilis polyporoid 2 "Hyphoderma praetermissum" ? 100 Hyphodontia alutaria 100 Resinicium bicolor Hydnochaete olivacea 98 Hyphodontia paradoxa 71 Inonotus hispidus Trichaptum abietinum

Coltricia perennis hymenochaetoid 1 100 Geastrum saccatum 100/75 Xanthoparmelia conspersa 56 74 Sphaerobolus stellatus 100/82 /63 Parmotrema tinctorum Ramaria stricta 100/100 Canoparmelia caroliniana Four-locus Bayesian MCMCMC tree 100/100 100/96 97 51 Gautieria otthii Punctelia hypoleucites 100/82 100/100 Flavoparmelia caperata 100 Gomphus floccosus Ramaria formosa 100/90 100/100 Stereocaulon paschale 100 nucSSU+nucLSU+mitSSU+RPB2 100/77 Cladonia caroliniana Kavinia himantia gomphoid-phalloid 100/ Sphaerophorus globosus Gloeophyllum sepiarium 61 103 species, 94 genera, 30 orders, 2 phyla 100/100 Xanthoria elegans Gloeophyllum trabeum 94/ 100/100 Xanthomendoza fallax 67 100/ Veluticeps berkeleyi Calicium viride ae id 83 Vuilleminia comedens Gloeophyllum 100/97 Lobaria quercizans t 99/ e Dendrocorticium roseocarneum corticioid 2 100/83 70/88 Peltigera degenii yc /68 m 100 Scytinostroma alutum 100/63 Placynthium nigrum ro o 59 100 Scytinostroma caudisporum Porpidia albocaerulescens n Bayesian posterior probability/Bayesian bootstrap proportion 100/51 ca Scytinostroma renisporum Lecanoromycetes 100/100 Sporostatia polyspora 100 e Scytinostroma eurasiaticogalactinum 100/100 Sporostatia testudinea L 99 70 Peniophora nuda NJ bootstrap proportion/MP bootstrap proportion Umbilicaria muehlenbergii 100/100 Coenogonium leprieurii 70 100 Russula compacta Eurotiomycetes 100/77 100/100 Coenogonium luteum 90 100 Russula exalbicans 97/58 Petractis luetkemuelleri Amylostereum laevigatum 0.1 substitutions/site 100/75 74/ 63 100/66 Diploschistes scruposus Echinodontium tinctorium 78/ 100/100 Gyalecta jenensis 100 100/91 Auriscalpium vulgare 94/87 Gyalecta ulmi 54 88/66 100 Bondarzewia berkeleyi 100/62 100/100 Acarosporina microspora 62 93 Bondarzewia montana Lichinomycetes 100/92 Stictis radiata 100/66 e 70 a ae Heterobasidion annosum Placopsis perrugosa d d 50 100/56 Baeomyces placophyllus ti ti Hericium coralloides 93/ ce e 85 100/79 y yc Hericium erinaceum Ionaspis lacustris m 54 o m Laxitextum bicolor 100/100 Thamnolia subuliformis p ro o o 100/96 Dibaeis baeomyces r p 100 Albatrellus skamanius 100/100 st s 90/86 Pertusaria amara O o e 100 Polyporoletus sublividus ar a Acarospora complanata c id Xenasma rimicola 100/ 100/100 A et 100/100 Acarospora schleicheri yc 83 95 Dentipellis separans m 100/100 Capronia pilosella io e Stereum gausapatum Leotiomycetes 100/100 r a 100/100 Capronia mansonii y id Acanthophysium cerussatum 100/100 th t 100/100 Dermatocarpon luridum to ce Aleurobotrys botryosus Sordariomycetes e y 99/ 99/98 100/100 Pyrenula pseudobufonia a m h o 55 Megalocystidium leucoxanthum 100/61 100/100 Pyrenula cruenta C ti ro Stereum hirsutum 100/100 Penicillium freii u E Xylobolus frustulatus 100/100 100/100 Hamigera avellanea Xylobolus subpileatus 95/ 100/100 Byssochlamys nivea 50 83 Amylostereum areolatum russuloid 91/ Lempholemma polyanthes Laetisaria fuciformis corticioid 1 Trichoglossum hirsutum 54 74/ "Ostropa barbara" ? 86 Auricularia auricula-judae Exidia glandulosa 96/76 54/ Sclerotinia sclerotiorum 97 66 81 Exidiopsis calcea 93/ Cudonia circinans 84 Leotia lubrica Pseudohydnum gelatinosum auriculariales 100/98 Hydropisphaera erubescens e 99/59 a 89 Physalacria inflata /58 100/100 d 57/51 Cordyceps militaris ti Trametes suaveolens e 100 93/ /100 Hypocrea citrina c e PEZIZOMYCOTINA y a Trametes hirsuta Neurospora crassa id 100 100/71 100/100 m t ae Trametes versicolor io e d 100/100 Sordaria macrospora r yc i 72/ 100/100 96/ a et Pycnoporus cinnabarinus Diaporthe phaseolorum rd m c 100/100 o eo y 95 Datronia mollis 96/55 Seynesia erumpens S d m 50 i io 69 "Trypethelium sp." ? th n Polyporus squamosus 79/51 o o 100/100 94/54 D h Panus rudis Curvularia brachyspora rt 69 62 Ganoderma australe Ascomycota /88 Lecanactis abietina A Peziza quelepidotia 96 66 Ganoderma boninense 100/69 84 Lentinus tigrinus 100/100 Morchella esculenta Pezizomycetes 97/53 100/100 Morchella elata Lenzites betulina SACCHAROMYCOTINA 62 Meripilus giganteus Saccharomyces cerevisiae Taphrina wiesneri TAPHRINOMYCOTINA Dentocorticium sulphurellum Schizosaccharomyces pombe Tyromyces chioneus 97/65 Polyporus melanopus id 100 Oligoporus lacteus 100/84 ro Polyporus squamosus o 100 Oligoporus leucomallelus 67/ /80 p 73 Lentinus tigrinus ly 56 o 85 Oligoporus rennyi 100/92 83/60 Cryptoporus volvatus p re Cryptoporus volvatus /77 Lenzites betulina o 60 Daedaleopsis confragosa Pycnoporus cinnabarinus c 86 Albatrellus syringae 100/100 Fomitopsis pinicola "Athelia arachnoidea" ? 100/59 /100 Piptoporus betulinus /52 95 Ceriporiopsis subvermispora Neolentiporus maculatissimus Laetiporus sulphureus 86 Phlebia albomellea 98/ 97/57 94/ /68 Phaeolus schweinitzii 92 Ceriporia viridans Sparassis crispa 100 Gloeoporus taxicola 100 65 51/ Auriporia aurea 78 Ceriporia purpurea 84/ Ceriporia purpurea Leptoporus mollis 100/59 Oligoporus lacteus id 100/100 ro 65 Grifola frondosa Oligoporus rennyi o 55/ p Wolfiporia cocos 73/100 ly 70 Antrodia carbonica o p 100 Sparassis brevipes 98/ Meripilus giganteus 88 Sparassis crispa 100/57 "Athelia bombacina" ? 86/ Dacryobolus sudans /70 Nia vibrissa s 100/68 Lentinula lateritia ic 90 Laetiporus sulphureus ar Abortiporus biennis 99/ Plicaturopsis crispa ag 77 100/ u 81 Neolentiporus maculatissimus Entoloma strictius e Boletus satanas 60 Phlebia chrysocrea 100/88 bolete Phlebia radiata 62/ 51/ Suillus cavipes 91/72 Amylostereum laevigatum 91 Phlebiopsis gigantea 100/94 99/83 Echinodontium tinctorium 92 58 Pulcherricium caeruleum 100/90 88/61 96 Peniophora nuda 52 Phanerochaete chrysosporium 71/ Auriscalpium vulgare 51 Basidiomycota 68/ 98/55 Phanerochaete sordida 62/ Russula compacta /Hymenomycetes 93/52 100 Fomitopsis pinicola Albatrellus skamanius id /Homobasidiomycetes 100/90 o Piptoporus betulinus 81/ Bondarzewia montana l id id u o o Antrodia carbonica 61/ Heterobasidion annosum ss ll t u re ae Auriporia aurea Stereum hirsutum r a h th c 100/84 Botryobasidium isabellinum n o 98 Junghuhnia nitida a en 75/98 Sistotrema eximum c Mycoacia meridionalis ym Phanerochaete sanguinea Coltricia perennis h 100/100 Gautieria otthii Polyporus melanopus polyporoid 1 gomphoid-phalloid /100 Ramaria formosa 99 Bankera fuligineoalba

Sarcodon imbricatus thelephoroid 100 Botryobasidium isabellinum 100 Botryobasidium subcoronatum 100 Clavulina cinerea Figure 3. Phylogenetic relationships among 103 ascomycete and basidiomycete species based on combined evidence from nucSSU, 100 84 98 Clavulina cristata 51 100 Multiclavula mucida nucLSU, mitSSU rDNA, and RPB2. This phylogeny resulted from a 50% majority rule consensus of 69 000 trees sampled with 100 92 100 Hydnum rufescens 99 70 Hydnum albidum Bayesian MCMCMC. The resulting posterior probabilities (PP) are shown above internal branches before the slash sign. One 68 Sistotrema eximum cantharelloid 100 Trichosporon debeurmannianum hundred Bayesian MCMCMC analyses were conducted on bootstrapped versions of this four-locus data set. Bayesian bootstrap 100 94 Trichosporon dermatis 50 Cryptococcus curvatus proportions (BBP) ≥50% are presented above internal branches after the slash sign. NJ bootstrap proportions (NJBP) with ML 97 100 Cryptococcus neoformans Filobasidiella depauperata 100 100 distance (1000 bootstrap replicates) are shown below internal branches before the slash sign. Weighted MP bootstrap proportions 100 Trichosporon porosum 100 93 Cryptococcus laurentii (MPBP) are shown below branches after the slash sign. Tremella foliacea Tremellomycetidae Basidiomycota 100 Dacrymyces chrysospermus 98 Calocera cornea Dacrymycetales 100 Ustilago hordei 100 100 Ustilago maydis 100 100 69 Malassezia furfur 93 97 100 Exobasidium vaccinii 83 Tilletia caries 100 Gymnosporangium juniperi-virginianae 100 Uromyces appendiculatus 96 Gymnosporangium libocedri 99 Chrysomyxa arctostaphyli 98 Endocronartium harknessii 100 Pucciniastrum goeppertianum 61 92 Melampsora epitea 100 100 98 Melampsora euphorbiae 100 100 98 Melampsora occidentalis 100 Caeoma torreyae 100 Insolibasidium deformans Glomeromycota 100 Naohidea sebacea 100 Rhodotorula cassiicola 100 100 Rhodotorula nymphaeae 100 Rhodotorula samaneae Urediniomycetes 65 89 Acaulospora longula Diversisporales 100 caledonium 98 Glomus clarum Glomerales 93 Sporodiniella umbellata 100 Syzygites megalocarpus 66 Amylomyces rouxii 100 Rhizopus stolonifer 64 67 Rhizopus microsporus var. oligosporus 50 100 Benjaminiella poitrasii 82 Cokeromyces recurvatus Kirkomyces cordense 94 Blakeslea trispora 100 Poitrasia circinans 100 Choanephora cucurbitarum Gilbertella persicaria 56 100 Backusella circina 88 Mucor recurvus var. indicus 100 Pilobolus umbonatus 76 Utharomyces epallocaulus Actinomucor elegans 79 Helicostylum elegans Pilaira anomala 67 67 Hyphomucor assamensis 95 Mucor mucedo Thamnidium elegans Zygorhynchus heterogamus 91 Backusella ctenidia 100 78 Ellisomyces anomalus 74 75 Mucor circinelloides 74 Chaetocladium jonesii 95 50 Mucor racemosus Chaetocladium brefeldii 66 90 Mycotypha africana 89 Mycotypha microspora Dicranophora fulva Parasitella parasitica Pirella circinans 100 Apophysomyces elegans 100 Saksenaea vasiformis 63 Circinella umbellata 100 Phascolomyces articulosus 100 Zychaea mexicana 81 100 99 Fennellomyces linderi 100 97 Thamnostylum piriforme 100 Thermomucor indicae-seudaticae 100 100 Rhizomucor pusillus 54 100 Absidia blakesleeana 100 100 Absidia corymbifera 97 Dichotomocladium elegans 66 100 Absidia repens 100 94 Chlamydoabsidia padenii 100 95 Cunninghamella echinulata 53 96 100 Gongronella butleri 100 Hesseltinella vesiculosa 100 Phycomyces blakesleeanus F 100 Spinellus fusiger U 67 100 Syncephalastrum monosporum var. pluriproliferum 100 Syncephalastrum racemosum N Protomycocladus faisalabadensis G 100 Radiomyces spectabilis Mucorales 2 I Zygomycota Conidiobolus coronatus 2 Umbelopsis isabellina 100 Mucorales 1 100 Umbelopsis ramanniana 72 Dissophora decumbens 100 Mortierella verticillata Mortierellales 96 Echinosporangium transversale Basidiobolus haptosporus 100 Entomophthorales 1 99 Basidiobolus ranarum 94 Coemansia reversa Kickxellales 69 Piptocephalis corymbifera Zoopagales Allomyces arbusculus Blastocladiales Batrachochytrium dendrobatidis 90 100 Cladochytrium replicatum Chytridiales 100 95 Polychytrium aggregatum Figure 1. Two-locus (nucSSU + nucLSU) Bayesian MCMCMC fungal tree of life depicting phylogenetic relationships among 558 taxa in 430 genera, 68 orders and five phyla. 56 61 "Monoblepharella" ? Monoblepharidales This phylogeny resulted from a 50% majority rule consensus of 24 000 trees sampled with Bayesian MCMCMC. The resulting posterior probabilities (PP) are shown above Chytridiomycota Spizellomyces punctatus Spizellomycetales 100 Antipathes galapagensis Outgroup 100 61 Suberites ficus internal branches. NJ bootstrap proportions (NJBP) with ML distances (1000 bootstrap replicates) are shown below internal branches. Species names are colored according to 94 Ichthyophonus hoferi Salpingoeca infusionum their respective phylum.