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. 2022 Apr 25;13:818358. doi: 10.3389/fmicb.2022.818358

Outline, Divergence Times, and Phylogenetic Analyses of Trechisporales (Agaricomycetes, Basidiomycota)

Zhan-Bo Liu 1, Ying-Da Wu 1,2, Heng Zhao 1, Ya-Ping Lian 1, Ya-Rong Wang 1, Chao-Ge Wang 1, Wei-Lin Mao 1, Yuan Yuan 1,*
PMCID: PMC9083364  PMID: 35547118

Abstract

Phylogenetic analyses inferred from the nuc rDNA ITS1-5.8S-ITS2 (ITS) data set and the combined 2-locus data set [5.8S + nuc 28S rDNA (nLSU)] of taxa of Trechisporales around the world show that Sistotremastrum family forms a monophyletic lineage within Trechisporales. Bayesian evolutionary and divergence time analyses on two data sets of 5.8S and nLSU sequences indicate an ancient divergence of Sistotremastrum family from Hydnodontaceae during the Triassic period (224.25 Mya). Sistotremastrum family is characterized by resupinate and thin basidiomata, smooth, verruculose, or odontoid-semiporoid hymenophore, a monomitic hyphal structure, and generative hyphae bearing clamp connections, the presence of cystidia and hyphidia in some species, thin-walled, smooth, inamyloid, and acyanophilous basidiospores. In addition, four new species, namely, Trechispora dentata, Trechispora dimitiella, Trechispora fragilis, and Trechispora laevispora, are described and illustrated. In addition, three new combinations, namely, Brevicellicium daweishanense, Brevicellicium xanthum, and Sertulicium limonadense, are also proposed.

Keywords: Hydnodontaceae, phylogenetic analysis, Trechispora, taxonomy, wood-rotting fungi

Introduction

Trechisporales K.H. Larss. was established by Hibbett et al. (2007). Most species in this order are corticioid fungi with smooth, grandinioid, odontioid, or hydnoid hymenophores, and others are polypores. All species have a monomitic or dimitic hyphal system with generative hyphae bearing clamp connections, and many species have rhizomorphs (mycelial cords) (Larsson, 2007).

At present, there is only an acknowledged and a named family belonging to Trechisporales, i.e., Hydnodontaceae Jülich. Hydnodontaceae contains 11 genera now, namely, Brevicellicium K.H. Larss. and Hjortstam, Dextrinocystis Gilb. and M. Blackw., Fibrodontia Parmasto, Pteridomyces Jülich, Luellia K.H. Larss. and Hjortstam, Porpomyces Jülich, Scytinopogon Singer, Subulicystidium Parmasto, Suillosporium Pouzar, Trechispora P. Karst., and Tubulicium Oberw (Larsson, 2007; Spirin et al., 2021).

Trechispora is the genus type of Trechisporales and Hydnodontaceae. It is the largest genus in this order, with more than 50 accepted species (Meiras-Ottoni et al., 2021; Zhao and Zhao, 2021). Identification keys for Trechispora species recorded in China and Brazil have been provided by some fungal taxonomists (Chikowski et al., 2020; Meiras-Ottoni et al., 2021; Zong et al., 2021). Trechispora was typified with Trechispora onusta P. Karst. [= Trechispora hymenocystis (Berk. and Broome) K.H. Larss.] (Karsten, 1890). It is characterized by the resupinate basidiomata (a few species have stipitate, flabellate, and effused–reflexed basidiomata) with smooth grandinioid, odontioid, hydnoid, or poroid hymenophores, a monomitic or dimitic hyphal structure with clamped generative hyphae and smooth to verrucose or aculeate basidiospores (Larsson, 1992; Larsson et al., 2004). Most species in Trechispora are soil-dwelling (Larsson et al., 2004). One remarkable character is the presence of ampullate septa on the subicular and especially on some hyphae of the mycelial cords. Above all, ampullate septa are only known from Scytinopogon, Trechispora, and Porpomyces mucidus (Pers.) Jülich within Trechisporales (Furtado et al., 2021; Meiras-Ottoni et al., 2021).

Larsson (2007) used the term “Sistotremastrum family” for the first time to accommodate Sistotremastrum suecicum Litsch. ex J. Erikss. and Sistotremastrum niveocremeum [= Sertulicium niveocremeum (Höhn. and Litsch.) Spirin and K.H. Larss.]. Since then, “Sistotremastrum family” has been adopted by some taxonomists (Telleria et al., 2013; Liu et al., 2019). In this work, the phylogeny of Trechisporales is carried out based on combined 5.8S + nLSU sequences. In addition, Bayesian evolutionary and divergence time analyses are also carried out to indicate the divergence time of Trechisporales, Hydnodontaceae, and Sistotremastrum family. We outline the Sistotremastrum family and discuss the difference between Hydnodontaceae and Sistotremastrum family.

During investigations on the diversity of wood-rotting fungi, seven resupinate specimens were collected from China and Malaysia. Their morphology corresponds to the concept of Trechispora. To confirm their affinity, phylogenetic analyses based on the ITS sequences are carried out. Both morphological characteristics and molecular evidence demonstrate that these seven resupinate specimens represent the four new species of Trechispora.

In addition, we downloaded the type sequences of Trechispora daweishanensis C.L. Zhao, Trechispora xantha C.L. Zhao, and Sistotremastrum limonadense G. Gruhn and P. Alvarado from GenBank. We also studied the type specimens of T. daweishanensis and T. xantha. In conclusion, T. daweishanensis and T. xantha were transferred to Brevicellicium, while S. limonadense was transferred to Sertulicium.

Materials and Methods

Morphological Studies

Macro-morphological descriptions are based on field notes and dry herbarium specimens. Microscopic structures are photographed using a Nikon Digital Sight DS-L3 (Japan) or Leica ICC50 HD (Japan) camera. Microscopic measurements are made from slide preparations of dry tissues stained with 1% Phloxine B (C20H4Br4Cl2K2O5) (Fan et al., 2021). We also use other reagents, such as Cotton Blue and Melzer’s reagent following Dai’s (2010) study. Spore measurements include both with ornamentation and without ornamentation. The following abbreviations are used: KOH = 5% potassium hydroxide; CB = Cotton Blue; CB(+) = weakly cyanophilous; CB− = acyanophilous in Cotton Blue; IKI = Melzer’s reagent; IKI− = neither amyloid nor dextrinoid in Melzer’s reagent; L = mean spore length (arithmetic average of all spores including ornamentation); W = mean spore width (arithmetic average of all spores including ornamentation); Q = a variation in the L/W ratios between the specimens studied; L′ = mean spore length (arithmetic average of all spores excluding ornamentation); W′ = mean spore width (arithmetic average of all spores excluding ornamentation); Q′ = a variation in the L′/W′ ratios between the specimens studied; n (a/b) = the number of spores (a) measured from a given number of specimens (b). When presenting spore size variation, 5% of measurements are excluded from each end of the range and these values are given in parentheses. Special color terms follow Petersen (1996). Herbarium abbreviations follow Thiers (2018). The studied specimens are deposited at the herbarium of the Institute of Microbiology, Beijing Forestry University (BJFC), and the herbarium of Southwest Forestry University (SWFC).

DNA Extraction, Polymerase Chain Reaction Amplification, and Sequencing

Total genomic DNA from the dried specimens is extracted by a CTAB rapid plant genome extraction kit (Aidlab Biotechnologies Company Limited, Beijing, China) according to the manufacturer’s instructions with some modifications (Liu and Yuan, 2020; Du et al., 2021). The ITS regions are amplified with the primers ITS4 and ITS5 (White et al., 1990). The nLSU regions are amplified with the primers LR0R and LR7 (Vilgalys and Hester, 1990).

The polymerase chain reaction (PCR) procedure for ITS is as follows: initial denaturation at 95°C for 3 min, followed by 35 cycles at 94°C for 40 s, 58°C for 45 s, and 72°C for 1 min, and a final extension of 72°C for 10 min. The PCR procedure for nLSU was as follows: initial denaturation at 94°C for 1 min, followed by 35 cycles at 94°C for 30 s, 48°C for 1 min, and 72°C for 1.5 min, and a final extension of 72°C for 10 min (Zhao et al., 2015; Liu and Dai, 2021). The PCR products are purified and sequenced in the Beijing Genomics Institute, China, with the same primers used in the PCR reactions.

Phylogenetic Analyses

Two combined matrices, an ITS1-5.8S-ITS2 (ITS) data set and a two-gene data set (5.8S + nLSU), are used for phylogenetic analyses. Phylogenetic analyses are performed with maximum likelihood (ML), maximum parsimony (MP), and Bayesian inference (BI) methods in the ITS data set. Phylogenetic analyses are performed with ML and BI methods in the combined two-gene data set (5.8S + nLSU). Species and strain sequences are adopted partly from 28S- and ITS-based tree topologies established by Meiras-Ottoni et al. (2021) and Spirin et al. (2021). New sequences generated in this study, along with reference sequences retrieved from GenBank (Table 1), are aligned by MAFFT 7 (Katoh et al., 20191) using the “G-INS-i” strategy and manually adjusted in BioEdit (Hall, 1999). Unreliably aligned sections are removed before analyses and attempts are made to manually inspect and improve alignment. The data matrix is edited in Mesquite v3.70 software (Maddison and Maddison, 2021). The sequence alignment is deposited at TreeBase (submission ID 29141 and 29142). Sequences of Auricularia sp., Exidia recisa (Ditmar) Fr., and Exidiopsis calcea (Pers.) K. Wells are included in phylogenetic analyses. They belong to another order, Auriculariales Bromhead. The order is close to Trechisporales (Sulistyo et al., 2021). We add these three sequences in the combined two-gene data set (5.8S + nLSU) to demonstrate that Trechisporales forms a strongly supported sister clade to Auriculariales. Sequences of Hyphodontia floccosa (Bourdot and Galzin) J. Erikss. and Hyphodontia subalutacea (P. Karst.) J. Erikss. in Hymenochaetales Oberw. obtained from GenBank are used as outgroups to root trees in the 5.8S + nLSU analysis. Two sequences of Brevicellicium atlanticum Melo, Tellería, M. Dueñas and M.P. Martín obtained from GenBank are used as outgroups to root trees in the ITS analysis.

TABLE 1.

Information of taxa used in phylogenetic analyses.

Species Collector ID (herbarium ID) GenBank accession no.
ITS nLSU
Auricularia sp. PBM 2295 DQ200918 AY634277
Brevicellicium atlanticum LISU 178566 (holotype) NR_119820 HE963774
Brevicellicium atlanticum LISU 178590 HE963775 HE963776
Brevicellicium daweishanense CLZhao 18255 (SWFC) MW302338 MW293867
Brevicellicium daweishanense CLZhao 17860 (SWFC, holotype) MW302337 MW293866
Brevicellicium exile MA-Fungi 26554 (holotype) HE963777 HE963778
Brevicellicium olivascens KHL 8571 (GB) HE963792 HE963793
Brevicellicium olivascens MA-Fungi 23496 HE963787 HE963788
Brevicellicium xanthum CLZhao 17781 (SWFC) MW302340 MW293869
Brevicellicium xanthum CLZhao 2632 (SWFC, holotype) MW302339 MW293868
Dextrinocystis calamicola He 5700 (BJFC) MK204534 MK204547
Dextrinocystis calamicola He 5693 (BJFC) MK204533 MK204546
Exidia recisa EL 15-98 (GB) AF347112 AF347112
Exidiopsis calcea MW 331 AF291280 AF291326
Fibrodontia alba TNM F24944 (holotype) KC928274 KC928275
Fibrodontia gossypina AFTOL-ID 599 DQ249274 AY646100
Hyphodontia floccosa Berglund 150-02 (GB) DQ873618 DQ873617
Hyphodontia subalutacea GEL2196 (KAS) DQ340341 DQ340362
Porpomyces mucidus Dai 12692 (BJFC) KT157833 KT157838
Porpomyces submucidus Cui 5183 (BJFC) KU509521 KT152145
Pteridomyces galzinii GB0150230 LR694188 LR694210
Pteridomyces galzinii Bernicchia 8122 (GB) MN937559 MN937559
Scytinopogon angulisporus TFB13611 – JQ684661
Scytinopogon chartaceum FLOR56185 MK458775 –
Scytinopogon pallescens He 5192 (BJFC) – MK204553
Sertulicium chilense MA-Fungi 86368 (holotype) HG315521 –
Sertulicium granuliferum He 3338 MK204552 MK204540
Sertulicium jacksonii Spirin 10425 (H) MN987943 MN987943
Sertulicium lateclavigerum LY 13467 MG913225 –
Sertulicium limonadense LIP 0001683 (holotype) MT180981 MT180978
Sertulicium limonadense He 6276 (BJFC) OK298489 * OK298947 *
Sertulicium niveocremeum KHL13727 (GB) MN937563 MN937563
Sertulicium vernale Soderholm 3886 (H, holotype) MT002311 MT664174
Sistotremastrum aculeatum Miettinen 10380.1 (H) MN991176 MW045423
Sistotremastrum aculeatum Cui 8401 (BJFC) KX081133 KX081184
Sistotremastrum aculeocrepitans KHL 16097 (URM) MN937564 MN937564
Sistotremastrum confusum KHL 16004 (URM) MN937567 MN937567
Sistotremastrum denticulatum Motato-Vásquez 894 (SP, holotype) MN954694 MW045424
Sistotremastrum fibrillosum LIP 0001413 (holotype) NR_161047 NG_075239
Sistotremastrum fibrillosum s. l. GUY13-119 (GG) MG913224 MG913210
Sistotremastrum fibrillosum s. l. KHL 16988 (MG) MN937568 MN937568
Sistotremastrum geminum Miettinen 14333 (MAN, holotype) MN937568 MN937568
Sistotremastrum induratum Spirin 8598 (H, holotype) MT002324 MT664173
Sistotremastrum mendax KHL 12022 (O, holotype) MN937570 MN937570
Sistotremastrum rigidum Motato-Vásquez 833 (SP, holotype) MN954693 MW045435
Sistotremastrum suecicum Kunttu 5959 (H) MT075859 MT002335
Sistotremastrum suecicum Miettinen 14550.1 (H) MT075860 MT002336
Sistotremastrum suecicum KHL 11849 (GB) MN937571 MN937571
Sistotremastrum vigilans Fonneland 2011-78 (O, holotype) MN937572 MN937572
Sistotremastrum vigilans Spirin 8778 (H) MN991182 MN991182
Subulicystidium tropicum He 3968 (BJFC) MK204531 MK204544
Suillosporium cystidiatum Spirin 3830 (H) MN937573 MN937573
Trechispora alnicola AFTOL-ID 665 DQ411529 AY635768
Trechispora araneosa KHL8570 (GB) AF347084 AF347084
Trechispora bambusicola CLZhao 3302 (SWFC) MW544021 MW520171
Trechispora bispora CBS 142.63 (holotype) MH858241 MH869842
Trechispora cohaerens TU 110332 UDB008249 –
Trechispora cohaerens TU 115568 UDB016421 –
Trechispora confinis KHL11064 (GB) AF347081 AF347081
Trechispora copiosa AMO456 MN701019 MN687976
Trechispora copiosa AMO422 (holotype) MN701013 MN687971
Trechispora cyatheae FR-0219442 UDB024014 UDB024014
Trechispora cyatheae FR-0219443 (holotype) UDB024015 UDB024015
Trechispora dentata Dai 22565 (BJFC) OK298491 * OM049408 *
Trechispora dimitiella Dai 21931 (BJFC) OK298492 * OK298948 *
Trechispora dimitiella Dai 21181 (BJFC) OK298493 * OK298949 *
Trechispora echinocristallina FR-0219445 (holotype) UDB024018 UDB024019
Trechispora echinocristallina FR-0219448 UDB024022 –
Trechispora echinospora MA-Fungi 82485 (holotype) JX392845 JX392846
Trechispora farinacea KHL 8793 (GB) AF347089 AF347089
Trechispora farinacea KHL 8451 (GB) AF347082 AF347082
Trechispora fimbriata CLZhao 7969 (SWFC) MW544024 MW520174
Trechispora fimbriata CLZhao 4154 (SWFC, holotype) MW544023 MW520173
Trechispora fissurata CLZhao 4571 (SWFC, holotype) MW544027 MW520177
Trechispora fissurata CLZhao 995 (SWFC) MW544026 MW520176
Trechispora fragilis Dai 20535 (BJFC) OK298494 * OK298950 *
Trechispora gelatinosa AMO1139 (holotype) MN701021 MN687978
Trechispora gelatinosa AMO824 MN701020 MN687977
Trechispora havencampii SFSU DED8300 (holotype) NR_154418 NG_059993
Trechispora hymenocystis TL 11112 (holotype) UDB000778 UDB000778
Trechispora hymenocystis KHL 8795 (GB) AF347090 AF347090
Trechispora incisa GB0090648 KU747095 KU747087
Trechispora incisa GB0090521 KU747093 –
Trechispora kavinioides KGN 981002 (GB) AF347086 AF347086
Trechispora laevispora Dai 21655 (BJFC) OK298495 * OM108710
Trechispora minispora MEXU 28300 (holotype) MK328886 MK328894
Trechispora minispora MEXU 28301 MK328886 MK328895
Trechispora mollis URM 85884 (holotype) MK514945 MH280003
Trechispora mollusca DLL2011-186 (CFMR) KJ140681 –
Trechispora mollusca DLL2010-077 (CFMR) JQ673209 –
Trechispora nivea GB0102694 KU747096 AY586720
Trechispora nivea MA-Fungi 74044 JX392832 JX392833
Trechispora papillosa AMO713 MN701022 MN687979
Trechispora papillosa AMO795 (holotype) MN701023 MN687981
Trechispora regularis KHL10881 (GB) AF347087 AF347087
Trechispora rigida URM 85754 MT406381 MH279999
Trechispora sp. AMO799 MN701008 MN687969
Trechispora sp. AMO440 MN701006 MN687967
Trechispora sp. KHL16968 (O) MH290763 MH290763
Trechispora sp. Dai 22173 (BJFC) OK298496 * OK298951 *
Trechispora sp. Dai 22174 (BJFC) OK298497 * OK298952 *
Trechispora stevensonii TU 115499 UDB016467 UDB016467
Trechispora stevensonii MA-Fungi 70669 JX392841 JX392842
Trechispora subsphaerospora KHL 8511 (GB) AF347080 AF347080
Trechispora termitophila AMO396 (holotype) MN701025 MN687983
Trechispora termitophila AMO390 MN701024 MN687982
Trechispora torrendii URM 85886 (holotype) MK515148 MH280004
Tubulicium raphidisporum He 3191 (BJFC) MK204537 MK204545
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*Newly generated sequences for this study. New species and new combinations or putatively new species are in bold.

The MP analysis is applied to the ITS data set sequences. Approaches to phylogenetic analysis follow Liu and Dai (2021), and the tree construction procedure is performed in PAUP* version 4.0 beta 10 software (Swofford, 2002). All characters are equally weighted, and gaps are treated as missing data. Trees are inferred using the heuristic search option with tree bisection and reconnection (TBR) branch swapping, and 1,000 random sequence additions maxtrees are set to 5,000, branches of zero length are collapsed, and all parsimonious trees are saved. Clade robustness is assessed using a bootstrap (BT) analysis with 1,000 replicates (Felsenstein, 1985). Descriptive tree statistics tree length (TL), consistency index (CI), retention index (RI), rescaled consistency index (RC), and homoplasy index (HI) are calculated for each maximum parsimonious tree (MPT) generated.

Maximum likelihood research is conducted with RAxML-HPC v. 8.2.3 (Stamatakis, 2014) and RAxML-HPC through the CIPRES Science Gateway (Miller et al., 20092). Statistical support values (BS) are obtained using nonparametric bootstrapping with 1,000 replicates. The BI analysis is performed with MrBayes 3.2.7a (Ronquist and Huelsenbeck, 2003). Four Markov chains are run for two runs from random starting trees for 4 million generations (ITS) and 8 million generations (5.8S + nLSU) until the split deviation frequency value reaches <0.01, and trees are sampled every 1,000 generations. The first 25% of the sampled trees are discarded as burn-in, and the remaining ones are used to reconstruct a majority rule consensus tree and to calculate Bayesian posterior probabilities (BPP) of the clades.

The optimal substitution models for the combined data set are determined using the Akaike information criterion (AIC) implemented in MrModeltest 2.3 (Posada and Crandall, 1998; Nylander, 2004) after scoring 24 models of evolution by PAUP* version 4.0 beta 10 software (Swofford, 2002). The selected model applied in the BI analyses and ML analyses is the model GTR + I + G.

Branches that received BT support for ML (BS), MP (BP), and BPP greater than 65% (BS), 70% (BP), and 0.9 (BPP) are considered as significantly supported, respectively. Additionally, the ML analysis results in the best tree, and only the ML tree is presented along with the support values from the MP and BI analyses. FigTree v1.4.4 (Rambaut, 2018) is used to visualize the resulting tree.

Divergence Time Estimation

Divergence time is estimated with the BEAST v2.6.5 software package (Bouckaert et al., 2019) with 5.8S and nLSU sequences representing all main lineages in Basidiomycota (Table 2). Sequences of the species are adopted partly from the topology established by Wang et al. (2021). Neurospora crassa Shear and B.O. Dodge from Ascomycota are designated as outgroup taxon (Wang et al., 2021). A BEAST XML input file is generated with BEATUti v2. The estimation of rates of evolutionary changes at nuclear acids is using ModelTest 3.7 with the GTR substitution model (Posada and Crandall, 1998). A log-normal distribution is employed for molecular clock analysis (Drummond and Rambaut, 2007). A Yule speciation model is selected as prior assuming a constant speciation rate per lineage. Three fossil fungi, viz. Paleopyrenomycites devonicus (Taylor et al., 1999, 2005), Archaeomarasmius leggetti (Hibbett et al., 1995, 1997), and Quatsinoporites cranhamii (Smith et al., 2004; Berbee and Taylor, 2010) are taken from Wang et al.’s (2021) study. An XML file is conducted for 10 billion generations, producing log files and trees files. The log file is analyzed in Tracer 1,3 and a maximum clade credibility (MCC) tree is interpreted in TreeAnnotator by trees file, removing the first 10% of the sampled trees as burn-in, and viewed in FigTree v1.4.2.

TABLE 2.

Information of taxa used in molecular clock analysis.

Species Specimen no. ITS nLSU
Amylocorticium cebennense HHB-2808 GU187505 GU187561
Anomoloma myceliosum MJL-4413 GU187500 GU187559
Athelia arachnoidea CBS 418.72 GU187504 GU187557
Auricularia heimuer Xiaoheimao LT716074 KY418890
Auricularia sp. PBM 2295 DQ200918 AY634277
Australovuilleminia coccinea MG75 HM046875 HM046931
Boletopsis leucomelaena AFTOL-ID 1527 DQ484064 DQ154112
Bondarzewia montana AFTOL-ID 452 DQ200923 DQ234539
Brevicellicium atlanticum LISU 178566 NR_119820 HE963774
Brevicellicium atlanticum LISU 178590 HE963775 HE963776
Brevicellicium daweishanense CLZhao 17860 MW302337 MW293866
Brevicellicium daweishanense CLZhao 18255 MW302338 MW293867
Brevicellicium exile MA-Fungi 26554 HE963777 HE963778
Brevicellicium olivascens KHL8571 HE963792 HE963793
Brevicellicium olivascens MA-Fungi 23496 HE963787 HE963788
Brevicellicium xanthum CLZhao 17781 MW302340 MW293869
Brevicellicium xanthum CLZhao 2632 MW302339 MW293868
Bridgeoporus sinensis Cui 10013 KY131832 KY131891
Calocera cornea AFTOL-ID 438 AY789083 AY701526
Coltricia perennis Cui 10319 KU360687 KU360653
Coltriciella dependens Dai 10944 KY693737 KY693757
Corticium roseum MG43 GU590877 AY463401
Craterocolla cerasi TUB 020203 KF061265 KF061265
Cryptococcus humicola AFTOL-ID 1552 DQ645516 DQ645514
Dacryopinax spathularia AFTOL-ID 454 AY854070 AY701525
Dextrinocystis calamicola He 5700 MK204534 MK204547
Dextrinocystis calamicola He5693 MK204533 MK204546
Exidia recisa EL 15-98 AF347112 AF347112
Exidiopsis calcea MW 331 AF291280 AF291326
Fasciodontia brasiliensis MSK-F 7245a MK575201 MK598734
Fasciodontia bugellensis MSK-F 5548 MK575204 MK598736
Fibrodontia alba TNMF 24944 KC928274 KC928275
Fibrodontia gossypina AFTOL-ID 599 DQ249274 AY646100
Fomitiporia hartigii MUCL 53551 JX093789 JX093833
Fomitiporia mediterranea AFTOL 688 AY854080 AY684157
Gloeophyllum sepiarium Wilcox-3BB HM536091 HM536061
Gloeophyllum striatum ARIZAN 027866 HM536092 HM536063
Grifola frondosa AFTOL-ID 701 AY854084 AY629318
Gymnopilus picreus ZRL2015011 LT716066 KY418882
Hymenochaete rubiginosa He1049 JQ716407 JQ279667
Hyphodontia densispora LWZ 20170908-5 MT319426 MT319160
Hyphodontia zhixiangii LWZ 20170818-13 MT319420 MT319151
Jaapia argillacea CBS 252.74 GU187524 GU187581
Gomphidius roseus MB 95-038 DQ534570 DQ534669
Kneiffiella barba-jovis KHL 11730 DQ873609 DQ873610
Kneiffiella subalutacea LWZ 20170816-9 MT319407 MT319139
Lepiota cristata ZRL20151133 LT716026 KY418841
Leptosporomyces raunkiaeri HHB-7628 GU187528 GU187588
Leucophellinus hobsonii Cui 6468 KT203288 KT203309
Lyomyces macrosporus LWZ20170817-2 MT319459 MT319194
Multiclavula mucida AFTOL-ID 1130 DQ521417 AY885163
Neoantrodiella gypsea Cui 10372 KT203290 MT319396
Neoantrodiella thujae Dai 5065 KT203293 MT319397
Neurospora crassa OR74A HQ271348 AF286411
Nigrofomes melanoporus JV 1704/39 MF629835 MF629831
Nigrofomes sinomelanoporus Cui 5277 MF629836 MT319398
Porodaedalea chinensis Cui 10252 KX673606 MH152358
Porpomyces mucidus Dai 12692 KT157833 KT157838
Porpomyces submucidus Cui 5183 KU509521 KT152145
Pteridomyces galzinii GB0150230 LR694188 LR694210
Pteridomyces galzinii Bernicchia8122 MN937559 MN937559
Ramaria rubella AFTOL-ID 724 AY854078 AY645057
Rigidoporus corticola ZRL20151459 LT716075 KY418899
Rigidoporus ginkgonis Cui 5555 KT203295 KT203316
Scytinopogon angulisporus TFB13611 – JQ684661
Scytinopogon pallescens He 5192 – MK204553
Sertulicium chilense MA-Fungi 86368 HG315521 –
Sertulicium granuliferum He 3338 MK204552 MK204540
Sertulicium jacksonii Spirin 10425 MN987943 MN987943
Sertulicium lateclavigerum LY 13467 MG913225 –
Sertulicium limonadense LIP 0001683 MT180981 MT180978
Sertulicium niveocremeum KHL13727 MN937563 MN937563
Sertulicium vernale Soderholm 3886 MT002311 MT664174
Sistotremastrum aculeatum Cui 8401 KX081133 KX081184
Sistotremastrum aculeatum Miettinen 10380.1 MN991176 MW045423
Sistotremastrum aculeocrepitans KHL 16097 MN937564 MN937564
Sistotremastrum confusum KHL 16004 MN937567 MN937567
Sistotremastrum denticulatum MV894 MN954694 MW045424
Sistotremastrum fibrillosum LIP 0001413 NR_161047 NG_075239
Sistotremastrum fibrillosum s. l. GUY13-119 MG913224 MG913210
Sistotremastrum fibrillosum s. l. KHL 16988 MN937568 MN937568
Sistotremastrum geminum Miettinen 14333 MN937568 MN937568
Sistotremastrum induratum Spirin 8598 MT002324 MT664173
Sistotremastrum mendax KHL12022 MN937570 MN937570
Sistotremastrum rigidum MV833 MN954693 MW045435
Sistotremastrum suecicum Kunttu 5959 MT075859 MT002335
Sistotremastrum suecicum Miettinen 14550.1 MT075860 MT002336
Sistotremastrum suecicum KHL 11849 (GB) MN937571 MN937571
Sistotremastrum vigilans Fonneland 2011-78 MN937572 MN937572
Sistotremastrum vigilans Spirin 8778 MN991182 MN991182
Subulicystidium tropicum He3968 MK204531 MK204544
Suillosporium cystidiatum VS3830 MN937573 MN937573
Suillus pictus AFTOL 717 AY854069 AY684154
Thelephora ganbajun ZRL20151295 LT716082 KY418908
Trametes versicolor ZRL20151477 LT716079 KY418903
Trechispora hymenocystis KHL8795 AF347090 AF347090
Tremellodendron sp. PBM2324 DQ411526 –
Tubulicium raphidisporum He 3191 MK204537 MK204545
Ustilago maydis AFTOL 505 AY854090 AF453938
Xylodon heterocystidiatus LWZ 20171015-33 MT319518 MT319264
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Results

Phylogenetic Analyses

The concatenated 5.8S + nLSU data set contains 50 5.8S and 50 nLSU sequences from 52 fungal specimens representing 35 taxa in Trechisporales. The data set has an aligned length of 1,528 characters, of which 1,126 are constant, 89 are variable but parsimony-uninformative, and 313 are parsimony-informative. The average standard deviation (SD) of split frequencies is 0.005271 (BI). Three new combinations, namely, Brevicellicium daweishanense, Brevicellicium xanthum, and Sertulicium limonadense, are proposed based on the examination of type materials and phylogenetic analyses of type sequences (Figure 1).

FIGURE 1.

FIGURE 1

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Phylogeny of Trechisporales generated by maximum likelihood (ML) analyses based on combined 5.8S + nLSU sequences. Branches are labelled with ML bootstrap (BT) >65%, and Bayesian posterior probabilities (BPP) >0.90, respectively. New combinations, the sequence origin from holotype and the type status of the species in the genus are indicated in bold.

The ITS data set contains sequences from 58 fungal specimens representing 36 Trechispora taxa (4 new species and another 32 Trechispora taxa). The data set has an aligned length of 753 characters, of which 284 are constant, 72 are variable but parsimony-uninformative, and 397 are parsimony-informative. MP analysis yields 13 equally parsimonious trees (TL = 2,318, CI = 0.398, RI = 0.638, RC = 0.254, and HI = 0.602). The average SD of split frequencies in BI analyses is 0.006959 (BI). The phylogenetic tree (Figure 2) reveals four new and independent lineages represented by our specimens, indicating that they are phylogenetically distinct from the species currently known in the genus. In addition, another taxon (Dai 22173 and Dai 22174) is treated as Trechispora sp.

FIGURE 2.

FIGURE 2

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Phylogeny of Trechispora generated by ML analyses based on combined ITS sequences. Branches are labelled with ML BT >65%, Parsimony Bootstrap Proportions >70%, and BPP >0.90, respectively. Putatively new species, the sequence origin from the holotype, and the type status of the species in the genus are indicated in bold.

The combined data set for the molecular clock analysis includes 100 collections, of which 47 belonged to Trechisporales. This data set results in a concatenated alignment of 1,588 characters with GTR as the best-fit evolutionary model. The MCC tree is used to study divergence time. The tree shows that Trechisporales occurs in a mean stem age of 270.85 Mya with a 95% highest posterior density (HPD) of 234.1–307.93 Mya (Figure 3). The tree also shows that the Sistotremastrum family and Hydnodontaceae occur in a mean stem age of 224.25 Mya [posterior probabilities (PP) = 0.8] with a 95% HPD of 182.47–266.75 Mya.

FIGURE 3.

FIGURE 3

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Maximum clade credibility (MCC) chronogram and estimated divergence times of all main lineages in Basidiomycota inferred from the combined data set of 5.8S and LSU regions. The estimated divergence times of 95% highest posterior density (HPD) for all clades are indicated as node bars. The colored dots refer to the positions of the mean stem age of Sistotremastrum family, Hydnodontaceae, Trechisporales, and Hymenochaetales. The BPP above 0.8 and the mean divergence times of clades are labelled above and below the branches, respectively, at the nodes.

Taxonomy

Sistotremastrum family

“Type genus”: Sistotremastrum J. Erikss.

Habitat: It grows on rotten angiosperm and gymnosperm wood.

Basidioma are resupinate, thin, pruinose, or waxy. Hymenophores are smooth, verruculose, or odontioid-semiporoid. The hyphal structure is monomitic; generative hyphae bear clamp connections, CB(+). Cystidia and hyphidia are present in some species. Basidia are clavate or cylindrical, often with a median constriction, mostly with 2–4 or 4–6 sterigmata, and rarely with 6–8 sterigmata. Basidiospores are narrowly ellipsoid, ovoid, or cylindrical, thin-walled (but the wall is distinct), smooth, inamyloid, and acyanophilous.

Notes: Sistotremastrum family accommodates the genera Sistotremastrum and Sertulicium in the order Trechisporales based on its distinct lineage in the phylogenetic analysis. The combined phylogeny of two-gene data (Figure 1) demonstrates that Sistotremastrum family forms a supported sister clade to Hydnodontaceae. Basidia of most species in the Sistotremastrum family have more than four sterigmata, and basidiospores are smooth, while basidia of species in Hydnodontaceae have four sterigmata and their basidiospores are smooth to verrucose or aculeate. In addition, ampullate septa are only present in Scytinopogon, Trechispora, and P. mucidus in Hydnodontaceae.

Trechispora dentata Z.B. Liu and Yuan Yuan, sp. November Figure 4

FIGURE 4.

FIGURE 4

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Trechispora dentata (holotype, Dai 22565). (A) A basidioma, (B) hyphae from subiculum, (C) hyphae from trama, (D) hyphae with ampullate septa (black arrow), (E) basidia and basidioles, and (F) basidiospores. Photo by Ya-Ping Lian and Zhan-Bo Liu.

MycoBank number: MB 842865.

Type: China, Yunnan province, Sipsongpanna, Mengla County, XiShuangBanNa Tropical Botanical Garden, on soil, in southwestern China, ca. E 101° 25′, N 21° 41′, alt. 570 m. The vegetation is a natural tropical forest. 4 July 2021, Y.C. Dai 22565 (holotype BJFC 037139).

Etymology: Dentata (Lat.): It refers to the species having a dentate hymenophore.

Basidioma: They are annual, resupinate, soft when fresh, fragile when dry, easily separable from the substratum, up to 2.5-cm long, 2-cm wide, and less than 1-mm thick at the center; hymenial surface irpicoid, white when fresh, becoming cream (4A2/3) when dry; margin indistinct and fimbriate, mycelial cords absent; pores or aculei 3–4/mm; hymenophore lacerate to dentate; subiculum very thin to almost absent; tubes or aculei concolorous with a hymenial surface, less than 1 mm long.

Hyphal structure: Hyphal system is monomitic; generative hyphae bear clamp connections; ampullate septa occasionally present in subiculum and trama, up to 5-μm wide; all hyphae IKI−, CB− are unchanged in KOH; rhomboidal calcium oxalate crystals are scattered.

Subiculum: Generative hyphae hyaline, thin- to thick-walled, frequently branched, loosely interwoven, 2–4 μm in diameter.

Tubes or aculei: Generative hyphae in trama hyaline, thin- to thick-walled, frequently branched, loosely interwoven, 2–3 μm in diameter; cystidia and cystidioles are absent; basidia are clavate or barrel-shaped, hyaline, bearing four sterigmata and a basal clamp connection, 10–15 × 4–5 μm; basidioles are similar to basidia in shape but slightly shorter.

Basidiospores: They are ellipsoid, hyaline, thick-walled, aculeate, occasionally with one guttule, IKI−, CB−, (4−)4.1–5 × (3−)3.2–4(−4.1) μm (including ornamentation), L = 4.46 μm, W = 3.66 μm, Q = 1.22 (n = 60/1); (2.2−)2.6–3.7(−3.8) × 2–2.5 μm (excluding ornamentation), L′ = 3.17 μm, W′ = 2.23 μm, and Q′ = 1.42 (n = 60/1).

Notes: T. dentata was discovered in the Yunnan Province of China. Phylogenetically, T. dentata is close to Trechispora regularis (Murrill) Liberta with strong support (96% BS, 96% BP, 1.00 BPP; Figure 2). However, T. regularis is strictly poroid (Liberta, 1973), and basidiospores of T. dentata are smaller than that of T. regularis [4.1–5 × 3.2–4 μm vs. 4–5.5 × 3.5–5 μm in T. regularis (including ornamentation); Liberta, 1973].

Trechispora dimitiella Z.B. Liu and Yuan, sp. November Figure 5

FIGURE 5.

FIGURE 5

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Trechispora dimitiella (holotype, Dai 21931). (A) A basidioma, (B) hyphae with ampullate septa from subiculum (black arrow), (C) hyphae from tubes, (D) basidia, (E) basidioles, and (F) basidiospores. Photo by Ya-Ping Lian and Zhan-Bo Liu.

MycoBank number: MB 842866.

Type: China, Hainan Province, Haikou, Jinniuling Park, on a rotten leaf, in southwestern China, ca. E 110° 19′, N 20° 1′, alt. 17 m. The vegetation is a plantation in tropical China. 7 November 2020, Y.C. Dai 21931 (holotype BJFC 035830).

Etymology: Dimitiella (Lat.): It refers to the species having a dimitic hyphal system.

Basidioma: They are annual, resupinate, soft when fresh, fragile when dry, easily separable from the substratum, up to 6-cm long, 4-cm wide, and approximately 3-mm thick at the center; the hymenial surface is poroid, pore surface white to cream (4A2/3) when fresh, becoming white to buff-yellow (4A4) when dry; margin indistinct, often with emerging mycelial cords; pores angular, 5–6/mm; dissepiments thin, lacerate; subiculum up to 1 mm thick; tubes concolorous with a poroid surface, up to 2 mm long.

Hyphal structure: Hyphal system is dimitic; generative hyphae bear clamp connections; ampullate septa occasionally present in subiculum and trama, up to 4.5 μm wide; all hyphae IKI−, CB− are unchanged in KOH; rhomboidal calcium oxalate crystals are scattered.

Subiculum: Generative hyphae hyaline, thin-walled, rarely branched, 2–3 μm in diameter; skeletal hyphae thick-walled with a wide lumen, unbranched, loosely interwoven, 2–4 μm diameter.

Tubes: Generative hyphae hyaline, thin-walled, rarely branched, 1.5–2.5 μm in diameter; skeletal hyphae thick-walled with a wide lumen, unbranched, loosely interwoven, 2–3 μm in diameter; cystidia and cystidioles are absent; basidia are barrel-shaped, hyaline, bearing four sterigmata and a basal clamp connection, 9.5–12 × 4–5 μm; basidioles are similar to basidia in shape but slightly shorter.

Basidiospores: They are ellipsoid, hyaline, thick-walled, aculeate, IKI−, CB−, (3.5−)3.6–4(−4.2) × (2.5–)2.7–3.1(−3.2) μm (including ornamentation), L = 3.84 μm, W = 2.92 μm, Q = 1.31–1.33 (n = 60/2); (2.6−)2.7–3.4(−3.7) × 2–2.6(−2.9) μm (excluding ornamentation), L′ = 3.04 μm, W′ = 2.18 μm, and Q′ = 1.38–1.4 (n = 60/2).

Additional specimen examined (paratypes): China, Yunnan Province, Jinghong, Primeval Forest Park, on soil, 7 July 2021, Y.C. Dai 22601 (BJFC), Dai 22602 (BJFC). Malaysia, Selangor, Kota Damansara, Community Forest Reserve, on rotten angiosperm wood, 7 December 2019, Y.C. Dai 21181 (BJFC 032835).

Notes: T. dimitiella was discovered in China and Malaysia. Most species in Trechispora are corticioid fungi with a monomitic hyphal structure, but T. dimitiella is different. Morphologically, T. dimitiella and Trechispora brasiliensis (Corner) K.H. Larss. share the poroid hymenophore with a dimitic hyphal system and aculeate basidiospores. However, the basidiospores of T. dimitiella are smaller than that of T. brasiliensis [3.6–4 × 2.7–3.1 μm vs. 4–4.5 × 3–4 μm in T. brasiliensis (including ornamentation), Larsson, 1992]. Phylogenetically, T. dimitiella is close to Trechispora incisa K.H Larss. (80% BS, 0.99 BPP; Figure 2), but T. dimitiella can be easily distinguished from T. incisa due to its poroid hymenophore with a dimitic hyphal system because T. incisa has arachnoid to farinose or minutely granulose hymenophore with a monomitic hyphal system (Larsson, 1996).

Trechispora fragilis Z.B. Liu and Yuan Yuan, sp. November Figure 6

FIGURE 6.

FIGURE 6

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Trechispora fragilis (holotype, Dai 20535). (A) A basidioma, (B) hyphae with ampullate septa from subiculum (black arrows), (C) hyphae from aculei, (D) hymenium with basidioles, (E) basidia, and (F) basidiospores. Photo by Ya-Ping Lian and Zhan-Bo Liu.

MycoBank number: MB 842867.

Type: China, Yunnan Province, Sipsongpanna, Mengla County, XiShuangBanNa Tropical Botanical Garden, on the ground of the forest, in southwestern China, ca. E 101° 25′, N 21° 41′, alt. 570 m. The vegetation is a natural tropical forest. 18 August 2019, Y.C. Dai 20535 (holotype BJFC 032203).

Etymology: Fragilis (Lat.): It refers to the species having fragile basidiocarps.

Basidioma: They are annual, resupinate, soft when fresh, fragile when dry, easily separable from the substratum, up to 3 cm long, 2 cm wide, and less than 1 mm thick at the center; the hymenial surface is odontoid, white when fresh, becoming cream (4A2/3) to buff-yellow (4A4) when dry; margin is indistinct and fimbriate, often with emerging mycelial cords; aculei sparse, 4–6/mm; subiculum very thin to almost absent; aculei concolorous with a hymenial surface, less than 1 mm long.

Hyphal structure: Hyphal system monomitic; generative hyphae bear clamp connections; ampullate septa occasionally present in subiculum and aculei, up to 7 μm wide; all hyphae IKI−, CB− are unchanged in KOH; rhomboidal calcium oxalate crystals are scattered.

Subiculum: Generative hyphae hyaline, thin- to thick-walled, frequently branched, loosely interwoven, 1.5–4 μm in diameter.

Aculei: Generative hyphae in trama hyaline, thin- to thick-walled, frequently branched, loosely interwoven, 1.5–3 μm in diameter; cystidia and cystidioles are absent; basidia are clavate shaped, hyaline, bearing four sterigmata, and a basal clamp connection, 12–14 × 3.5–4 μm; basidioles are similar to basidia in shape but slightly shorter.

Basidiospores: Ellipsoid, hyaline, thick-walled, aculeate, IKI−, CB−, (3.2−)3.8–4(−4.2) × (2.4−)2.5–3 μm (including ornamentation), L = 3.53 μm, W = 2.79 μm, Q = 1.27 (n = 60/1); (2.6−)2.8–3.7(−4) × (1.9−)2–2.7(−3.1) μm (excluding ornamentation), L′ = 3.16 μm, W′ = 2.26 μm, and Q′ = 1.40 (n = 60/1).

Notes: T. fragilis was discovered in the Yunnan Province of China. Phylogenetically, T. fragilis groups with Trechispora termitophila Meiras-Ottoni and Gibertoni and Trechispora havencampii (Desjardin and B.A. Perry) Meiras-Ottoni and Gibertoni (69% BS, 0.92 BPP; Figure 2). T. termitophila can be easily distinguished from T. fragilis due to its coralloid basidioma. In addition, the basidiospores of T. fragilis are smaller than that of T. termitophila [6.5–7.5 μm vs. 4.5–5 μm in T. termitophila (including ornamentation), Meiras-Ottoni et al., 2021]. T. havencampii can also be easily distinguished from T. fragilis due to its coralloid basidioma. In addition, basidiospores of T. fragilis are smaller than that of T. havencampii [3.8–4 × 2.5–3 μm vs. 5.2–6.5 × 3.5–4.2 μm in T. havencampii (including ornamentation), Desjardin and Perry, 2015].

Trechispora laevispora Z.B. Liu, Y.D. Wu and Yuan Yuan, sp. November Figure 7

FIGURE 7.

FIGURE 7

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Trechispora laevispora (holotype, Dai 21655). (A) A basidioma, (B,C) hyphae from subiculum, (D) subicular hyphae with ampullate septa (black arrow) and a piece of hymenium, (E) basidia and basidioles, and (F) basidiospores. Photo by Ya-Ping Lian and Zhan-Bo Liu.

MycoBank number: MB 842868.

Type: China, Inner Mongolia Autonomous Region, Arxan, Bailang Feng Scenic Spot, on the charred trunk of Larix, in southwestern China, ca. E 119° 56′, N 47° 10′, alt. 1,511 m. The vegetation is a natural boreal forest. 25 August 2020, Y.C. Dai 21655 (holotype BJFC 035556).

Etymology: Laevispora (Lat.): It refers to the species having smooth basidiospores.

Basidioma: They are annual, resupinate, soft when fresh and dry, up to 8 cm long, 3 cm wide, and less than 1 mm thick at the center; the hymenial surface is smooth, white when fresh and dry; margin is indistinct and fimbriate, often with emerging mycelial cords; subiculum very thin to almost absent.

Hyphal structure: Hyphal system monomitic; generative hyphae bear clamp connections; ampullate septa frequently present in subiculum and hymenium, up to 7 μm wide; all hyphae IKI−, CB− are unchanged in KOH; rhomboidal calcium oxalate crystals are abundant.

Subiculum: Generative hyphae hyaline, thin-walled, frequently branched, loosely interwoven, 1.5–3 μm in diameter.

Hymenium: Generative hyphae in subhymenium hyaline, thin-walled, frequently branched, 1.5–3 μm in diameter; cystidia and cystidioles are absent; basidia are clavate shaped, hyaline, bearing four sterigmata and a basal clamp connection, 11.5–15 × 4–5 μm; basidioles are similar to basidia in shape but slightly shorter.

Basidiospores: Ellipsoid, hyaline, thin-walled, smooth, IKI−, CB−, (2.5−) 2.6–3.2(−3.3) × (1.8−)1.9–2.2(−2.5) μm, L = 2.92 μm, W = 2.04 μm, and Q = 1.43 (n = 60/1).

Notes: T. laevispora was discovered in the Inner Mongolia Autonomous Region of China. Phylogenetically, T. laevispora groups with Trechispora cohaerens (Schwein.) Jülich and Stalpers with strong support (94% BS, 96% BP, 1.00 BPP; Figure 2). Both species share a smooth hymenophore, a monomitic hyphal system with smooth basidiospores. However, basidiospores of T. cohaerens are thick-walled and larger than that of T. laevispora (3.5–4 × 2.2–2.5 μm in T. cohaerens; Larsson, 1992).

B. daweishanense (C.L. Zhao) Z.B. Liu and Yuan Yuan, comb. November

MycoBank number: MB 842869.

Basionym: T. daweishanensis C.L. Zhao, Phytotaxa 479(2): 153 (2021).

Type: China. Yunnan Province, Honghe, Pingbian County, Daweishan National Nature Reserve, on the fallen branch of angiosperms, 1 August 2019, CLZhao 17860 (holotype SWFC).

Description: See Zong et al. (2021, as T. daweishanensis).

B. xanthum (C.L. Zhao) Z.B. Liu and Yuan Yuan, comb. November

MycoBank number: MB 842870.

Basionym: T. xantha C.L. Zhao, Phytotaxa 479(2): 155 (2021).

Type: China. Yunnan Province, Yuxi, Xinping County, Mopanshan National Forestry Park, on the trunk of Albizia julibrissin, 20 August 2017, CLZhao 2632 (holotype SWFC).

Description: See Zong et al. (2021, as T. xantha).

Notes: Zong et al. (2021) described T. daweishanensis and T. xantha as new species. However, in our phylogeny, they belong to the genus Brevicellicium (98% BS, 1.00 BPP; Figure 1). The type specimens of abovementioned species are studied [CLZhao 17860 (SWFC); CLZhao 2632 (SWFC)]. We do not observe ampullate hyphae from type materials as mentioned by Zong et al. (2021). We suppose that Zong et al. (2021) confused basidioles with ampullate hyphae (ampullate septa on some generative hyphae), which are remarkable characters of Trechispora. In fact, T. daweishanensis and T. xantha have a smooth hymenophore, a monomitic hyphal structure with clamped generative hyphae, and the absence of ampullate septa. They fit Brevicellicium well. Herein, we combine these two species in Brevicellicium based on morphological and phylogenetic evidence (Figure 1).

S. limonadense (G. Gruhn and P. Alvarado) Z.B. Liu and Yuan Yuan, comb. November

MycoBank number: MB 842871.

Basionym: S. limonadense G. Gruhn and P. Alvarado, Phytotaxa 498(1): 36 (2021).

Type: French Guiana. On the bark of an unidentified dead trunk lying on the ground, October 22, 2013, LIP 0001683 (holotype).

Description: See Gruhn and Alvarado (2021, as S. limonadense).

Notes: Gruhn and Alvarado (2021) described S. limonadense as a new species. However, at the same time, Spirin et al. (2021) segregated the species around S. niveocremeum (Höhn. and Litsch.) J. Erikss. into the new genus Sertulicium. In our phylogeny, S. limonadense groups with Sertulicium granuliferum (Hallenb.) Spirin and Volobuev Sertulicium lateclavigerum (Boidin and Gilles) Spirin and Viner (Figure 1). We did not study specimens, but S. limonadense is characterized by smooth to tuberculate hymenophore and basidia have 6–8 sterigmata (Gruhn and Alvarado, 2021) and fits Sertulicium better. Hence, we transfer S. limonadense to Sertulicium.

Discussion

Larsson (2007) showed that S. suecicum and S. niveocremeum (= S. niveocremeum) formed a strongly supported sister clade (94% BS, 1.00 BPP) to Hydnodontaceae within Trechisporales. However, in his phylogenetic analysis of 5.8S + nLSU, there were a few species in Hydnodontaceae and Sistotremastrum to establish a new family for S. suecicum and S. niveocremeum. Hence, Larsson (2007) named this clade Sistotremastrum family. The same strongly supported topology was recovered by Telleria et al. (2013); Gruhn et al. (2018), and Meiras-Ottoni et al. (2021) by the nLSU phylogenetic analysis. Spirin et al. (2021) presented a comprehensive study of Sistotremastrum and Sertulicium with 17 species. They used the nLSU region to perform phylogenetic analyses of 16 species in the two genera (Figure 1 in Spirin et al., 2021), except for Sertulicium chilense (Telleria, M. Dueñas and M.P. Martín) Spirin and Volobuev because the nLSU sequences of S. chilense were absent. However, they were not able to generate high support values for the node connecting Sistotremastrum and Sertulicium (87% BS, 0.87 BPP, Figure 1 in Spirin et al., 2021). As a result, they gave up establishing a new family too.

ITS1-5.8S-ITS2 is an important marker used for the barcoding of fungal species (Liu et al., 2021; Wangsawat et al., 2021). However, the difficulty in aligning ITS sequences for fungi in Trechisporales is evident because it is a data set covering taxa in distinct taxonomic levels (Larsson, 2007). Therefore, it is not a good idea to run combined analyses of ITS + nLSU, so we use the most stable and conservative portion of ITS (5.8S) and nLSU to our phylogenetic analyses of Sistotremastrum and Sertulicium (5.8S + nLSU) (Figure 1). We add S. chilense and S. limonadense to phylogenetic analyses. Our results of the Sistotremastrum are the same as phylogenetic analyses by Spirin et al. (2021, Figure 1). However, our phylogenetic analyses of Sertulicium are a bit different from that by Spirin et al. (2021, Figure 1) because the data sets used in both studies are different. Above all, we generate high support values for the node connecting Sistotremastrum and Sertulicium from ML analysis (93% BS) based on 5.8S and nLSU sequences; however, BI fails to provide support for the node (0.76 BPP).

Divergence time is estimated with 5.8S and nLSU sequences representing all main lineages in Basidiomycota (Figure 3). The MCC tree shows that Basidiomycota occurs in a mean stem age of 509.57 Mya. Trechisporales occurs in a mean stem age of 270.85 Mya. The tree also shows that the Sistotremastrum family and Hydnodontaceae occur in a mean stem age of 224.25 Mya (PP = 0.8). Zhao et al. (2017) indicate that the divergence times of Basidiomycota are 530 Mya (the mean stem age). He et al. (2019) indicate that the divergence times of Trechisporales and Hydnodontaceae are 259 Mya (the mean stem age). Our experimental results agree with them. In this paper, we update the divergence times of Trechisporales and Hydnodontaceae and define the divergence time of the Sistotremastrum family.

Bayesian phylogenetic inference fails to provide support for the node of Sistotremastrum and Sertulicium, so we use the term “Sistotremastrum family” for the two genera without a formal description of the new family. In the future, we will sequence additional DNA regions or whole genomes, for a more robust phylogenetic analysis.

At present, there are only two species in the Sistotremastrum family ever been recorded from China, i.e., Sistotremastrum aculeatum Miettinen and Viner (Cui 8401) and S. granuliferum (He 3338; CLZhao 5531, 9771). Recently, we collected a specimen from the Yunnan Province of China (He 6276), and its morphological and DNA data demonstrated the specimen is S. limonadense. The species is a new record in China, and we have uploaded ITS and nLSU sequences of the specimen (He 6276) to GenBank. Above all, we study all the Chinese specimens of species in the Sistotremastrum family seriously, and their morphology fits the descriptions of Gruhn and Alvarado (2021) and Spirin et al. (2021). We also collected a specimen from the Hainan Province of China (Dai 17696). The ITS (OK298490) region is different from Sistotremastrum fibrillosum G. Gruhn and P. Alvarado by 6%, and morphologically it is similar to S. fibrillosum. However, we only have a single specimen, so for the time being we regard Dai 17696 as Sistotremastrum sp.

In this article, we use the whole ITS region in analyses of Trechispora to visualize the genetic distances among new taxa and those already described. T. dentata, T. dimitiella, T. fragilis, and T. laevispora are described as new to science based on morphological characteristics and molecular evidence (Figure 2). Most of these new species are found in subtropical or tropical Asia and conform to the phenomenon that subtropical or tropical Asia harbors high taxonomic diversity for all wood-decaying fungi (Dai, 2012; Cui et al., 2019). We also collected two resupinate specimens (Dai 22173 and Dai 22174) from the Hainan Province of China. The morphology of the two specimens corresponds to the concept of Trechispora and forms a distinct lineage within the Trechispora clade (100% BS, 1.00 BPP; Figure 2). However, these specimens are sterile, so we regard Dai 22173 and Dai 22174 as Trechispora spp. temporarily here.

Molecular phylogenetic analyses in the present study show that Brevicellicium forms a monophyletic clade in which all Brevicellicium species are included (98% BS, 1.00 BPP; Figure 1). However, when we add sequences of T. xantha and T. daweishanensis, we find sequences of a two-species cluster with Brevicellicium with high support (100% BS, 1.00 BPP; Figure 1). We request and examine type specimens from Zhao and find T. xantha and T. daweishanensis corresponding to the concept of Brevicellicium and they should be transferred to the genus Brevicellicium (see the notes of B. daweishanense).

Data Availability Statement

The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found in the article/supplementary material.

Author Contributions

Z-BL: design of the research, performance of the research, and writing and revising this manuscript. Z-BL, HZ, Y-PL, Y-RW, C-GW, and W-LM: data analysis and interpretation. Z-BL, YY, and Y-DW: a collection of the materials. All authors contributed to the article and approved the submitted version.

Conflict of Interest

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Publisher’s Note

All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.

Acknowledgments

We thank Prof. Dr. Chang-Lin Zhao (SWFC, China) and Prof. Yu-Cheng Dai for allowing us to study their specimens.

Footnotes

Funding

The research is supported by the National Natural Science Foundation of China (Project Nos. 31870007 and 32011540380).

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Associated Data

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Data Availability Statement

The datasets presented in this study can be found in online repositories. The names of the repository/repositories and accession number(s) can be found in the article/supplementary material.

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