Phylogenetic assessment and taxonomic revision of Scytalidium (Helotiales, Leotiomycetes)

1 Phylogenetic assessment and taxonomic revision of Scytalidium (Helotiales, Leotiomycetes) Shuo-Qiu Tong1, Yi-Fan Yang1, Peng Li1, Yong-Jun Wu1, Bing-Da Sun2, Zhi-Yuan Zhang3 1 College of Life Sciences, Institute of Agro-bioengineering, Guizhou University, Guiyang 550025, China 2 China General Microbiological Culture Collection Center, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China 3 College of Eco-Environmental Engineering, Guizhou Minzu University, Guiyang 550025, China Corresponding author: Zhi-Yuan Zhang ([email protected]) Copyright: © Shuo-Qiu Tong et al. This is an open access article distributed under terms of the Creative Commons Attribution License (Attribution 4.0 International – CC BY 4.0). Abstract Members of Scytalidium are primarily saprotrophic and are known for their ability to colonize a variety of substrates, including soil, decaying plant material, and wood. During an investigation of soil microfungi in Capsicum annuum cultivation areas of China, seven Scytalidium isolates were obtained from soil samples collected in Guizhou. In this study, we revised the genus Scytalidium by combining morphological characteristics and phylogenetic analyses based on concatenated ITS–LSU sequences. The results showed that Scytalidium sensu stricto comprises 16 species, including the type species S. lignicola and four novel species proposed in this study (S. chlamydosporum, S. guizhouense, S. rodionovae, and S. tongrenense). Ten species were excluded, and six species were treated as uncertain due to the lack of available molecular sequences. This study revised the genus Scytalidium, expanded its species diversity and geographical distribution, and lays the foundation for future taxonomic research on this genus. Key words: Leotiomycetes, molecular systematics, re-classification, taxonomy, three new taxa Introduction Pesante (1957) introduced the genus Scytalidium, with Scytalidium lignicola as the type species. This genus is characterized by the possession of two types of arthroconidia: hyaline, thin-walled, cylindrical entities formed by the fragmentation of undifferentiated hyphae, and brown, thick-walled, broadly ellipsoid entities borne in an intercalary fashion (i.e., chlamydospore-like cells) (Pesante 1957). Currently, there are 34 records of Scytalidium in Index Fungorum (https:// www.indexfungorum.org, accessed July 2025). Many former Scytalidium species have been transferred to other genera, with several even serving as type species for newly established genera; for example, Acidomyces, with S. acidophilum as type (Selbmann et al. 2008); Neoscytalidium, typified by S. dimidiatum (Crous et al. 2006), later superseded by S. hyalinum as type and synonym of N. dimidiatum (Phillips et al. 2013); and Mycothermus, based on the reclassified Torula thermophila (invalidly proposed by Natvig et al. 2015; validated Academic editor: Joey Tanney Received: 9 July 2025 Accepted: 25 September 2025 Published: 14 October 2025 Citation: Tong S-Q, Yang Y-F, Li P, Wu Y-J, Sun B-D, Zhang Z-Y (2025) Phylogenetic assessment and taxonomic revision of Scytalidium (Helotiales, Leotiomycetes). IMA Fungus 16: e164608. https://doi. org/10.3897/imafungus.16.164608 IMA Fungus 16: e164608 (2025) DOI: 10.3897/imafungus.16.164608 Research Article 2 IMA Fungus 16: e164608 (2025), DOI: 10.3897/imafungus.16.164608 Shuo-Qiu Tong et al.: Scytalidium revisited by Wang et al. 2019). Additionally, S. aurantiacum var. album is recognized as a synonym of S. album (https://www.indexfungorum.org, https://www.mycobank.org). To date, there are still 29 species remaining in Scytalidium. Among them, S. fulvum, S. hepiali, S. melanoxylicola, S. nielamuense, S. verruculosum, and S. xigazense lack available molecular sequence data (Morgan-Jones et al. 1984; Li and Sun 1988; Wu and Zhang 2010; Awasthi et al. 2020). The genus Scytalidium exhibits a broad ecological distribution, being found in a wide range of environments and substrates. Several species are primarily associated with plants, including wood and timber, where they act as saprobes, cause blue staining, or decay (e.g., S. album, S. aurantiacum, S. circinatum, S. cuboideum, and S. sphaerosporum) (Klingström and Beyer 1965; Sigler and Wang 1990; Robinson et al. 2007), or as pathogens (S. lignicola on cassava (Manihot esculenta)) (Oren et al. 2001; Kang et al. 2010; Silva et al. 2013; De Medeiros et al. 2019). Some are also reported from dead plant material such as twigs (e.g., S. synnematicum) (Crous et al. 2023). Other species are mycoparasitic and known from cultivated fungi, including S. auriculariicola on Auricularia polytricha and S. ganodermophthorum on Ganoderma lucidum (Kang et al. 2010; Peng et al. 2014). Soil represents another common habitat, with several species exhibiting saprophytic lifestyles, often isolated from agricultural or forest soils (e.g., S. chlamydosporum, S. guizhouense, S. tongrenense, and S. terrigenum) (Jeong et al. 2025). Additionally, one species (S. assmuthi) has been isolated from the gut of a wood-feeding termite, suggesting an association with insects (Manawasinghe et al. 2024). Another species (S. philadelphianum) has been recovered from compressed air, suggesting occurrence in airborne or industrial environments (Crous et al. 2022). The chili pepper (Capsicum annuum) is a globally grown and consumed spice crop that is rich in vitamins. It has a long history of cultivation in China. During our investigation of soil microfungi in C. annuum cultivation areas of Guizhou, China from 2022–2024, we obtained seven isolates of the genus Scytalidium. This study has the following objectives: (1) to describe three novel Scytalidium species collected from soil; (2) to propose S. rodionovae as a new species, with strain 3C designated as the ex-type culture, to legitimize the taxon; (3) to provide a checklist that includes substrate, availability of molecular data, morphological characteristics, and country of origin; and (4) to revise the genus Scytalidium by combining morphological characteristics and phylogenetic analyses based on concatenated ITS–LSU sequences. Materials and methods Fungal isolation and morphological characterization Soil sampling was conducted annually in July from 2022–2024 across three Capsicum annuum–growing regions in Guizhou Province: Liuguang Town (116.44E, 26.99N; Xiuwen County, Guiyang), Shiban Town (106.65E, 26.46N; Huaxi District, Guiyang), and Qiaojia Town (108.44E, 28.28N; Yanhe County, Tongren). Soil samples were collected using a shovel at a depth of 0–10 cm from the soil surface, stored in sterile Ziploc bags, transported to the laboratory under refrigeration (4 °C), and processed immediately upon arrival. Fungi were isolated using the dilution method described in Tong et al. (2023). 3 IMA Fungus 16: e164608 (2025), DOI: 10.3897/imafungus.16.164608 Shuo-Qiu Tong et al.: Scytalidium revisited After obtaining pure colonies on potato dextrose agar (PDA; Coolaber, China), they were transferred onto fresh PDA and synthetic nutrient-poor agar (SNA; Coolaber, China) media, followed by incubation at 25 °C in darkness for 14 d to observe macroscopic and morphological characteristics of the colonies. Color names and codes adhered to the Methuen Handbook of Colour (Kornerup and Wanscher 1978). For light microscopic observations, slides were prepared from cultures grown on PDA, and 25% lactic acid was used as the mounting fluid. Morphological features were observed and recorded using a Zeiss Axio Imager A2 microscope and a Zeiss AxioCam MRc color digital camera (Carl Zeiss Ltd., München, Germany). Structural measurements were carried out using Digimizer software (v6.4.5), with a minimum of 50 measurements taken for each structure, such as conidia. The ex-type strains for the novel species were deposited at the China General Microbiological Culture Collection Center (CGMCC), China. Additionally, all living cultures were stored in a metabolically inactive state (i.e., kept in sterile 30% glycerol in a –80 °C freezer) and were deposited in the College of Eco-Environmental Engineering, Guizhou Minzu University, and at the Institute of Agro-bioengineering, Guizhou University, China. Dried culture (at 50 °C) specimens were deposited at the Fungarium (HMAS), Institute of Microbiology, Chinese Academy of Sciences (CAS). MycoBank numbers were registered for new names, including novel species and combinations. DNA extraction, PCR amplification, and sequencing Scraped fungal mycelia were used for DNA extraction with a BioTeke Fungal Genomic DNA Extraction Kit (DP2032, BioTeke, Beijing, China), following the manufacturer’s protocol. Polymerase chain reactions (PCR) were performed using internal transcribed spacer (ITS) and large nuclear ribosomal subunit rDNA (LSU) regions, which were amplified using primer pairs ITS1/ITS4 (White et al. 1990) and LR0R/LR5 (Vilgalys and Hester 1990), respectively. PCR was carried out in a 25 μL reaction volume containing 12.5 μL of 2× Power Taq PCR MasterMix (a premixed and ready-to-use solution including 0.1 units/μL Taq DNA Polymerase, 500 μM dNTP mixture each [dATP, dCTP, dGTP, dTTP], 20 mM Tris-HCl pH 8.3, 100 mM KCl, 3 mM MgCl2, stabilizer, and enhancer), 1 μL of each primer, 1 μL genomic DNA extract, and 8.5 μL double-distilled water. The PCR products were purified and sequenced at Shanghai Sangon Biological Engineering Technology & Services Co., Shanghai, China. Raw forward and reverse sequences were assembled using Lasergene software (version 6.0, DNASTAR). All newly generated sequences have been deposited in the GenBank database (Table 1). Phylogenetic analyses Based on BLASTn search results and recently published data (Table 1), reference sequences were downloaded and aligned using MAFFT v7.037 (Katoh and Standley 2013) and further refined with MEGA 6.06 (Tamura et al. 2013). Phylogenetic analyses of combined aligned ITS and LSU sequences were performed with Bayesian and maximum-likelihood algorithms. Maximum likelihood (ML) analysis was carried out using IQ-TREE v1.6.11 (Nguyen et al. 2015) with 10,000 bootstrap tests and the ultrafast algorithm (Minh et al. 2013). The best evolutionary models for phylogenetic analyses were selected independently for each locus using 4 IMA Fungus 16: e164608 (2025), DOI: 10.3897/imafungus.16.164608 Shuo-Qiu Tong et al.: Scytalidium revisited Table 1. Strains/vouchers used in this study, with information on the GenBank accessions of the sequences. Species Strain/voucher GenBank accession no. References ITS LSU Albonectria rigidiuscula CBS 133754 MW827602 MW827641 Crous et al. 2021 Bisifusarium dimerum CBS 108944 T JQ434586 JQ434514 Ropars et al. 2012 Caliciopsis calicioides 211 JX968549 NA Assefa et al. 2014 Caliciopsis moriondi CBS 146717 MN156540 NA Migliorini et al. 2020 Caliciopsis orientalis CBS 138.64 KP881690 MH870024 Wood et al. 2016; Vu et al. 2019 Caliciopsis pinea CBS 139.64 KP881691 NA Wood et al. 2016 Cyanonectria multiformsporum CGMCC 3.20774 T OL897004 OL897046 Zhang et al. 2023a Fusarium callistephi CBS 187.53 T MH857158 MH868694 Vu et al. 2019 Fusarium hoodiae CBS 132474 T MH866022 MH877470 Vu et al. 2019 Fusarium oxysporum LC13766 MW016600 NA Wang et al. 2022 Geejayessia celtidicola CBS 125502 T HM626657 HM626669 Schroers et al. 2011 Geniculospora grandis CBS 261.84 MH861735 MH873440 Vu et al. 2019 Hyaloscypha hepaticicola CBS 652.89 T MH862193 MH873881 Vu et al. 2019 Hypoxylon florendophyticum GUCC 193025.1 T ON791190 ON791224 Zhang et al. 2023b Hypoxylon hinnuleum MUCL 3621 T MK287537 MK287549 Sir et al. 2019 Hypoxylon investiens CBS 118183 KC968925 KY610450 Kuhnert et al. 2014; Wendt et al. 2018 Hypoxylon lateripigmentum MUCL 53304 T KC968933 KY610486 Kuhnert et al. 2014; Wendt et al. 2018 Hypoxylon lignicola MFLUCC 16-0926 T MK828609 MK835808 Luo et al. 2019 Lasiobelonium lonicerae FC-2270 AB481284 AB481319 Hosoya et al. 2010 Luteonectria albida CBS 102683 MW827615 MH874402 Crous et al. 2021; Vu et al. 2019 Monochaetia dimorphospora NBRC 9980 T LC146750 LC146750 Liu et al. 2019 Monochaetia hanzhongensis CFCC 54451 T OK339776 OK339747 Jiang et al. 2023 Monochaetia quercus CBS 144034 T MH554171 MH554365 Liu et al. 2019 Mycofalcella calcarata CCM F-10289 KC834065 KC834033 Baschien et al. 2013 Mycothermus thermophiloides CBS 183.81 T LT993603 LT993603 Wang et al. 2019 Mycothermus thermophilus CBS 625.91 T LT993604 LT993604 Wang et al. 2019 Neocosmospora regularis CBS 230.34 T LR583763 LR583967 Sandoval-Denis et al. 2019 Neocosmospora silvicola CBS 123846 T LR583766 LR583971 Sandoval-Denis et al. 2019 Neodevriesia cladophorae CGMCC 3.17901 T KU578112 KU578114 Wang et al. 2017 Neodevriesia knoxdaviesii CBS 122898 T MH863254 MH874778 Vu et al. 2019 Neodevriesia metrosideri CBS 145084 T NR_161141 NG_066296 Crous 2018 Neodevriesia strelitziae CBS 122379 T MH863206 EU436763 Vu et al. 2019; Arzanlou and Crous 2008 Neoscytalidium hyalinum CBS 145.78 eiT MH861121 DQ377922 Vu et al. 2019; Crous et al. 2006 Neoscytalidium hylocereum PSU-HP01 T LC590859 NA Wonglom et al. 2023 Neoscytalidium novaehollandiae CBS 122071 T MH863173 MH874720 Vu et al. 2019 Nothofusarium devonianum NRRL 22134 T MW827632 MW827673 Crous et al. 2021 Rectifusarium ventricosum CBS 748.79 T HQ897816 KM231658 Gräfenhan et al. 2011; Lombard et al. 2015 Remersonia tenuis CBS 784.85 T LT993609 LT993609 Wang et al. 2019 Remersonia thermophila CBS 643.91 LT993610 LT993610 Wang et al. 2019 Scytalidium album 173 MF992676 MF966378 Pavlov et al. 2018 Scytalidium album CBS 373.65 MH858618 MH870258 Vu et al. 2019 Scytalidium album CBS 372.65 T MH858617 MH858617 Vu et al. 2019 Scytalidium assmuthi PYCC 9837 T OR415883 OR415885 Manawasinghe et al. 2024 Scytalidium aurantiacum CBS 374.65 T MH858619 MH870259 Vu et al. 2019 Scytalidium auriculariicola YBI-3 T GU591724 NA Peng et al. 2014 Scytalidium candidum 3C T MF992675 MG018250 Pavlov et al. 2018 5 IMA Fungus 16: e164608 (2025), DOI: 10.3897/imafungus.16.164608 Shuo-Qiu Tong et al.: Scytalidium revisited Species Strain/voucher GenBank accession no. References ITS LSU Scytalidium chlamydosporum CGMCC 3.28993 T PV890025 PV890032 This study Scytalidium chlamydosporum SQT11 PV890026 PV890033 This study Scytalidium chlamydosporum SQT12 PV890027 PV890034 This study Scytalidium chinense H1091 T HQ213805 HQ221579 Geng et al. 2016 Scytalidium circinatum CBS 654.89 T NR_160180 MH873882 Vu et al. 2019 Scytalidium circinatum SM14-27-6-5 MN905823 NA Held et al. 2020 Scytalidium cuboideum KACC 41223 T GQ272628 NA Kang et al. 2010 Scytalidium cuboideum UAMH 3792 GQ503338 NA Kang et al. 2010 Scytalidium cuboideum CBS 241.62 MH858144 MH858144 Vu et al. 2019 Scytalidium flavobrunneum CBS 244.59 T MH857854 MH857854 Vu et al. 2019 Scytalidium ganodermophthorum TPML 97003 GQ272622 NA Kang et al. 2010 Scytalidium ganodermophthorum UAMH 10320 T GQ272617 NA Kang et al. 2010 Scytalidium ganodermophthorum H123 GQ272620 NA Kang et al. 2010 Scytalidium guizhouense CGMCC 3.28992 T PV890023 PV890030 This study Scytalidium guizhouense SQT09 PV890024 PV890031 This study Scytalidium indonesiacum CBS 259.81 T MH861338 MH873098 Vu et al. 2019 Scytalidium infestans CBS 161.91 T MH862246 MH862246 Vu et al. 2019 Scytalidium japonicum CBS 494.88 T MH873833 MH873833 Vu et al. 2019 Scytalidium japonicum CBS 125804 MH863771 NA Vu et al. 2019 Scytalidium lignicola UAMH 1502 T NR_121314 NA Schoch et al. 2014 Scytalidium lignicola CBS 125602 MH863583 NA Vu et al. 2019 Scytalidium multiseptatum CBS 693.70 MH859908 MH871702 Vu et al. 2019 Scytalidium multiseptatum CBS 241.68 MH859124 MH870836 Vu et al. 2019 Scytalidium parasiticum AAX0113 T KF925449 NA Goh et al. 2015 Scytalidium philadelphianum CPC 40793 T ON811538 NA Crous et al. 2022 Scytalidium sphaerosporum CBS 187.69 GQ272623 MG018251 Kang et al. 2010; Pavlov et al. 2018 Scytalidium sphaerosporum ATCC 34392 T NR_145360 NA Kang et al. 2010 Scytalidium sphaerosporum KACC 41222 GQ272626 NA Kang et al. 2010 Scytalidium synnematicum CCMB 207/13 T OQ430525 OQ430526 Crous et al. 2023 Scytalidium terminale CBS 171.40 T MH856079 NA Vu et al. 2019 Scytalidium terrigenum KNUF-23-236 T LC859329 LC859330 Jeong et al. 2025 Scytalidium tibetense H1127 T HQ213808 HQ221582 Geng et al. 2016 Scytalidium tongrenense CGMCC 3.28994 T PV890028 PV890035 This study Scytalidium tongrenense SQT14 PV890029 PV890036 This study Scytalidium tuberculatum H1195 T HQ213809 HQ221583 Geng et al. 2016 Scytalidium uredinicola CBS 578.75 MH860954 NA Vu et al. 2019 Setofusarium setosum CBS 635.92 T MW827634 MW827675 Crous et al. 2021 Trichopeziza sulphurea KUS-F52218 JN033398 JN086701 Han et al. 2014 Tricladium obesum CCM F-14598 KC834068 KC834035 Baschien et al. 2013 Geoglossum azoricum AMI-SPL1247 OQ618223 OQ618224 Crous et al. 2023 Geoglossum dunense TUR-A 199830 T KP744516 KP744517 Loizides et al. 2015 Notes: T: ex-type; eiT: ex-isotype; Scytalidium vaccinii is a synonym of Hyaloscypha hepaticicola, and CBS 652.89 is the type strain of S. vaccinii. CBS: Westerdijk Fungal Biodiversity Institute, Utrecht, Netherlands; CGMCC: China General Microbiological Culture Collection Center, Beijing, China; CPC: Collection of Pedro Crous housed at CBS; MFLUCC: Mae Fah Luang University Culture Collection, Chiang Rai, Thailand; GUCC: Culture Collection of the Department of Plant Pathology, Agriculture College, Guizhou University, China; ATCC: American Type Culture Collection, Manassas, USA; KACC: Korean Agricultural Culture Collection, Republic of Korea; UAMH: University of Alberta Mold Herbarium and Culture Collection, Edmonton, Canada; NRRL: Agricultural Research Service Culture Collection, National Center for Agricultural Utilization Research, USA; NBRC: Biological Resource Center, National Institute of Technology and Evaluation, Tokyo, Japan; MUCL: Université Catholique de Louvain, Louvain-la-Neuve, Belgium. Other acronyms represent personal collections; NA, not available. DNA sequences for the new isolates were in bold. 6 IMA Fungus 16: e164608 (2025), DOI: 10.3897/imafungus.16.164608 Shuo-Qiu Tong et al.: Scytalidium revisited ModelFinder (Kalyaanamoorthy et al. 2017) under the corrected Akaike Information Criterion (AICc). Bayesian analyses were performed with MrBayes v3.2 (Ronquist et al. 2012). The Markov Chain Monte Carlo (MCMC) method was used to perform 5 × 107 simulations with a sampling frequency of 10³ generations and a 25% burn-in. After the analysis was finished, Tracer v1.5 (Drummond and Rambaut 2007) was used to determine burn-in and confirm that both runs had converged. Results Phylogenetic analyses Phylogenetic trees were generated to determine the class-level placement of the Scytalidium isolates (Analysis 1) and to resolve the phylogenetic relationships among species within Scytalidium sensu stricto (Analysis 2) (Figs 1, 2). Geoglossum azoricum (AMI-SPL1247) and G. dunense (TUR-A 199830) were used as the outgroups in Analysis 1, whereas Tricladium obesum (CCM F-14598) and Mycofalcella calcarata (CCM F-10289) were used as the outgroups in Analysis 2. The concatenated sequences of Analyses 1 and 2 included 87 and 33 taxa, respectively, and consisted of 1,347 characters (ITS: 542 bp and LSU: 805 bp) and 1,400 characters (ITS: 581 bp and LSU: 819 bp), respectively, with gaps. Analysis 1. The phylogenetic tree showed that 16 species of Scytalidium, including the type species S. lignicola and the three new species proposed here, formed a highly supported clade (98% BS/1 PP) within Leotiomycetes (Fig. 1). In addition, multiple species or strains of Scytalidium were dispersed across Sordariomycetes, Dothideomycetes, and Eurotiomycetes (Fig. 1). These findings strongly suggest that Scytalidium is polyphyletic, with several species placed outside the main clade containing the type species, including taxa in three other classes (Dothideomycetes, Eurotiomycetes, and Sordariomycetes). Analysis 2. The phylogenetic tree showed that Scytalidium formed a strongly supported clade (100/1), which can be broadly divided into five clades (Fig. 2). Clade I comprised S. album, S. assmuthi, S. aurantiacum, S. rodionovae, S. circinatum, S. terrigenum, and S. tongrenense with high support (89/0.99). Clade II comprised S. ganodermophthorum and S. synnematicum with high support (91/0.98). Clade III comprised only S. guizhouense. Clade IV comprised S. chlamydosporum, S. cuboideum, and S. sphaerosporum with high support (80/0.98). Clade V comprised S. auriculariicola, S. lignicola, and S. philadelphianum with high support (88/0.99). Additionally, the novel species proposed in this study each formed distinct subclades with strong statistical support (100/1). Taxonomy Scytalidium Pesante Type species. Scytalidium lignicola Pesante. Notes. In this study, the phylogeny of Scytalidium was reconstructed using concatenated ITS and LSU sequences from (i) the ex-type strain of S. lignicola (type species of the genus), (ii) ex-type strains of 24 additional Scytalidium species, and (iii) ex-type strains of phylogenetically related fungi in Leotiomycetes, Sordariomycetes, Dothideomycetes, and Eurotiomycetes, as identified through 7 IMA Fungus 16: e164608 (2025), DOI: 10.3897/imafungus.16.164608 Shuo-Qiu Tong et al.: Scytalidium revisited Figure 1. Phylogenetic tree inferred from a maximum likelihood analysis based on a concatenated alignment of ITS and LSU sequences from 87 isolates representing Scytalidium, related taxa, and outgroup taxa. Numbers at branches indicate support values (IQ-TREE-BS/BI-PP) above 70%/0.90. The new species are printed in bold blue, and the taxa transferred out of Scytalidium in bold black. Strains with a type status are indicated with “T.” The tree is rooted to Geoglossum azoricum (AMI-SPL1247) and G. dunense (TUR-A 199830). 0.07 Lasiobelonium lonicerae FC-2270 Scytalidium lignicola UAMH 1502 T Remersonia thermophila CBS 643.91 Scytalidium tuberculatum H1195 T Hypoxylon hinnuleum MUCL 3621 T Monochaetia quercus CBS 144034 T Mycothermus thermophilus CBS 625.91 T Scytalidium sphaerosporum CBS 187.69 Scytalidium cuboideum CBS 241.62 Scytalidium lignicola CBS 125602 Scytalidium japonicum CBS 494.88 T Neodevriesia strelitziae CBS 122379 T Neodevriesia metrosideri CBS 145084 T Scytalidium terminale CBS 171.40 T Geoglossum azoricum AMI-SPL1247 Scytalidium aurantiacum CBS 374.65 T Geejayessia celtidicola CBS 125502 T SQT11 CGMCC 3.28993 T Setofusarium setosum CBS 635.92 T Neodevriesia knoxdaviesii CBS 122898 T Hypoxylon lateripigmentum MUCL 53304 T Scytalidium parasiticum AAX0113 T Scytalidium cuboideum UAMH 676 IT Fusarium oxysporum LC13766 Scytalidium circinatum SM14-27-6-5 Scytalidium multiseptatum CBS 241.68 Scytalidium album CBS 373.65 SQT09 Scytalidium auriculariicola YBI-3 T Neoscytalidium hyalinum CBS 145.78 eiT Scytalidium tibetense H1127 T 3C T Scytalidium rodionovae sp. nov. Scytalidium japonicum CBS 125804 Trichopeziza sulphurea KUS-F52218 Fusarium callistephi CBS 187.53 T Mycothermus thermophiloides CBS 183.81 T Scytalidium chinense H1091 T Scytalidium assmuthi PYCC9837 T Scytalidium sphaerosporum ATCC 34392 T Hypoxylon florendophyticum GUCC 193025.1 T Geniculospora grandis CBS 261.84 Scytalidium terrigenum KNUF-23-236 T Neoscytalidium novaehollandiae CBS 122071 T Albonectria rigidiuscula CBS 133754 Scytalidium ganodermophthorum H123 Cyanonectria multiformsporum CGMCC 3.20774 T Neoscytalidium hylocereum PSU-HP01 T Monochaetia dimorphospora NBRC 9980 T Scytalidium ganodermophthorum UAMH 10320 T Tricladium obesum CCM F-14598 Scytalidium flavobrunneum CBS 244.59 T Hypoxylon lignicola MFLUCC 16-0926 T Scytalidium cuboideum UAMH 3792 Scytalidium infestans CBS 161.91 T Remersonia tenuis CBS 784.85 T Scytalidium album CBS 372.65 T Fusarium hoodiae CBS 132474 T Scytalidium ganodermophthorum TPML 97003 SQT12 Scytalidium philadelphianum CPC 40793 T CGMCC 3.28992 T Hypoxylon investiens CBS 118183 Caliciopsis pinea CBS 139.64 Caliciopsis calicioides 211 Neodevriesia cladophorae CGMCC 3.17901 T Scytalidium indonesiacum CBS 259.81 T Scytalidium sphaerosporum KACC 41222 Scytalidium album 173 Scytalidium synnematicum CCMB207/13 T Nothofusarium devonianum NRRL 22134 T Scytalidium circinatum CBS 654.89 T Geoglossum dunense TUR-A 199830 T Hyaloscypha hepaticicola CBS 652.89 T Scytalidium uredinicola CBS 578.75 Scytalidium multiseptatum CBS 693.70 Neocosmospora silvicola CBS 123846 T Caliciopsis moriondiCBS 146717 SQT14 Caliciopsis orientalis CBS 138.64 Neocosmospora regularis CBS 230.34 T CGMCC 3.28994 T Mycofalcella calcarata CCM F-10289 Luteonectria albida CBS 102683 Monochaetia hanzhongensis CFCC 54451 T 88/0.92 89/0.83 100/1 100/1 98/0.98 100/1 100/1 100/1 100/1 100/0.99 100/1 100/1 100/1 100/1 95/0.95 100/1 96/0.95 86/0.97 100/1 94/- 96/0.99 99/1 100/1 100/1 100/1 100/1 100/0.95 98/0.71 100/1 81/0.74 98/1 100/1 100/0.99 84/- 82/- 100/1 100/1 100/1 92/- 100/1 100/1 97/0.94 93/0.95 100/1 98/1 99/0.99 100/1 77/- 100/1 82/- 72100/1 93/0.86 99/0.96 100/1 98/1 88/0.88 99/0.97 100/1 78/0.91 100/ 91/- 72/- 100/1 84/0.99 81/0.91 97/1 96/0.89 100/1 97/0.96 100/1 75/0.92 ×4 ×2 ×2 ×2 ×2 Scytalidium tongrenense sp. nov. Scytalidium guizhouense sp. nov. Scytalidium chlamydosporum sp. nov. Rectifusarium ventricosum CBS 748.79 T Bisifusarium dimerum CBS 108944 T Scytalidium s. str. Tricladiaceae Leotiomycetes Sordariomycetes Dothideomycetes Eurotiomycetes 8 IMA Fungus 16: e164608 (2025), DOI: 10.3897/imafungus.16.164608 Shuo-Qiu Tong et al.: Scytalidium revisited preliminary phylogenetic analyses. The results showed that Scytalidium is a polyphyletic genus spread across at least four classes in Ascomycota (Fig. 1). We formally propose to restrict Scytalidium sensu stricto to a monophyletic group of 16 species, with the exclusion of all phylogenetically divergent lineages currently classified in Scytalidium. Accepted species 1. Scytalidium album L. Beyer & Klingström, Svensk bot. Tidskr. 59: 35 (1965) Description and illustration. Klingström and Beyer (1965). Notes. Scytalidium album was initially isolated from Norway spruce wood damaged by root rot, and it induces blue staining in timber (Klingström and Beyer 1965). Scytalidium album is phylogenetically closely related to S. assmuthi, S. aurantiacum, S. rodionovae, S. circinatum, S. terrigenum, and S. tongrenense Figure 2. Phylogenetic tree inferred from a maximum likelihood analysis based on a concatenated alignment of ITS and LSU sequences from 33 isolates representing Scytalidium and outgroup taxa. Numbers at branches indicate support values (IQ-TREE-BS/BI-PP) above 70%/0.90. The new species are printed in blue. Strains with a type status are indicated with “T.” The tree is rooted to Tricladium obesum (CCM F-14598) and Mycofalcella calcarata (CCM F-10289). 0.04 173 SQT09 UAMH 1502 T CBS 374.65 T Mycofalcella calcarata CCM F-10289 CGMCC 3.28992 T Tricladium obesum CCM F-14598 SM14-27-6-5 UAMH 10320 T CGMCC 3.28993 T CPC 40793 T CBS 125602 CGMCC 3.28994 T YBI-3 T CBS 654.89 T KACC 41222 ATCC 34392 T SQT14 TPML 97003 CBS 241.62 H123 UAMH 3792 CBS 373.65 PYCC9837 T 3C T CBS 372.65 T CBS 187.69 CCMB207/13 T SQT11 SQT12 UAMH 676 IT AAX0113 T (ex-type of Scytalidium parasiticum) KNUF-23-236 T 97/0.99 100/1 83/0.9 89/0.99 100/1 96/0.97 100/1 100/1 91/0.99 89/0.99 88/0.99 99/0.99 93/0.99 91/0.98 100/1 100/1 71/- 100/1 100/1 100/1 100/1 100/1 80/0.98 Scytalidium tongrenense sp. nov. Scytalidium guizhouense sp. nov. Scytalidium chlamydosporum sp. nov. Scytalidium terrigenum Scytalidium aurantiacum Scytalidium album Scytalidium circinatum Scytalidium assmuthi Scytalidium rodionovae sp. nov. Scytalidium ganodermophthorum Scytalidium synnematicum Scytalidium sphaerosporum Scytalidium cuboideum Scytalidium lignicola Scytalidium auriculariicola Scytalidium philadelphianum Clade Ⅰ Clade Ⅱ Clade Ⅲ Clade Ⅳ Clade Ⅴ 9 IMA Fungus 16: e164608 (2025), DOI: 10.3897/imafungus.16.164608 Shuo-Qiu Tong et al.: Scytalidium revisited (Figs 1, 2). Morphologically, S. album is distinguished from S. assmuthi, S. rodionovae, and S. tongrenense by its unknown sexual morph and simultaneous production of arthroconidia and chlamydospore-like cells (Klingström and Beyer 1965; Pavlov et al. 2018; Manawasinghe et al. 2024). Scytalidium album differs from S. terrigenum by its hyaline arthroconidia (Jeong et al. 2025) and from S. circinatum by the shape and size of its chlamydospore-like cells (globose or ellipsoidal, 6.4–14.4 × 4.8–9.6 μm in S. album vs. globose, lobed, or irregular, 4–9 × 3–9 μm in S. circinatum) (Sigler and Wang 1990). Furthermore, S. album differs from S. aurantiacum by its secretion of a pale-yellow pigment (Klingström and Beyer 1965). Additionally, they can be distinguished by their low sequence similarities. Based on a pairwise comparison of ITS and LSU, S. album (ex-type CBS 372.65) differs from S. assmuthi (ex-type PYCC 9837) by 7.4% (34/460 bp, four gaps) in the ITS and 4.2% (23/536 bp, one gap) in the LSU; from S. aurantiacum (ex-type CBS 374.65) by 2.9% (15/521 bp, no gap) in the ITS and 2.5% (14/553 bp, no gap) in the LSU; from S. rodionovae (ex-type 3C) by 8.6% (49/571 bp, five gaps) in the ITS and 3.7% (22/591 bp, no gap) in the LSU; from S. circinatum (ex-type CBS 654.89) by 6.8% (49/721 bp, five gaps) in the ITS and 5.4% (67/1,234 bp, five gaps) in the LSU; from S. terrigenum (extype KNUF-23-236) by 3.9% (23/580 bp, one gap) in the ITS and 3.3% (19/561 bp, four gaps) in the LSU; and from S. tongrenense (ex-type CGMCC 3.28994) by 6.6% (36/544 bp, six gaps) in the ITS and 2.4% (14/575 bp, one gap) in the LSU. This species demonstrates notable inhibitory effects against various wood-decay fungi, revealing promising biocontrol potential (Klingström and Beyer 1965; Klingstrom and Johansson 1973; Cease et al. 1989). Furthermore, it produces diverse, unique metabolites (such as sorbicillinoid analogs) that exhibit significant inhibitory activity against cancer cells and Aspergillus niger, indicating considerable medicinal potential (El-Elimat et al. 2015). 2. Scytalidium assmuthi G. Mane, R. Avchar, R. Morey & Rohit Sharma, Fungal Diversity 130: 86 (2024) Description and illustration. Manawasinghe et al. (2024). Notes. Scytalidium assmuthi was introduced to accommodate an isolate obtained from the gut of the termite Odontotermes assmuthi feeding on wood logs from the northern Western Ghat (India) (Manawasinghe et al. 2024). Scytalidium assmuthi is phylogenetically closely related to S. album, S. aurantiacum, S. rodionovae, S. circinatum, S. terrigenum, and S. tongrenense (Figs 1, 2). The distinctions between S. assmuthi and S. album are provided in the notes for S. album. Scytalidium assmuthi is distinguished from S. aurantiacum, S. rodionovae, S. circinatum, S. terrigenum, and S. tongrenense by its unknown sexual morph and production of only chlamydospore-like cells (Klingström and Beyer 1965; Sigler and Wang 1990; Pavlov et al. 2018; Manawasinghe et al. 2024; Jeong et al. 2025). Furthermore, based on a pairwise comparison of ITS, S. assmuthi (ex-type PYCC 9837) differs from S. aurantiacum (ex-type CBS 374.65) by 8.2% (37/453 bp, 37 gaps) in the ITS and 9.9% (87/879 bp, 62 gaps) in the LSU; from S. rodionovae (ex-type 3C) by 3.7% (17/461 bp, four gaps) in the ITS and 0.5% (4/830 bp, one gap) in the LSU; from S. circinatum (ex-type CBS 654.89) by 3.3% (15/459 bp, two gaps) in the ITS and 0.9% (5/573 bp, one gap) in the LSU; from S. terrigenum 16 IMA Fungus 16: e164608 (2025), DOI: 10.3897/imafungus.16.164608 Shuo-Qiu Tong et al.: Scytalidium revisited 4.5–6.5 μm (avg. 10.5 × 5.4 μm, n = 30). Fertile hyphae borne laterally on simple conidiophores, fragmenting into arthroconidia. Arthroconidia hyaline to brown, smooth, cuboidal to oblong or cylindrical, 3–7.5 × 2–3 µm (avg. 5.3 × 2.7 μm, n = 30). Sexual morph unknown. Geographical distribution. Guizhou Province, China. GenBank numbers. SQT08–SQT09, ITS: PV890023–PV890024, LSU: PV890030–PV890031. Notes. Phylogenetic analysis showed that two new isolates (CGMCC 3.28992 and SQT09) clustered in a single subclade with strong support (100/1) (Figs 1, 2). Morphologically, S. guizhouense differs from other species in Scytalidium by its production of conidia, arthroconidia, and chlamydospore-like cells, and an unknown sexual morph (see notes of key). Scytalidium guizhouense was isolated from soil in a pepper cultivation area and is likely to have a saprophytic lifestyle. 10. Scytalidium lignicola Pesante, Ann. Sperim. Agrar. 11 (suppl.): 265 (1957) Description and illustration. Pesante (1957) Notes. Scytalidium lignicola is phylogenetically closely related to S. auriculariicola and S. philadelphianum (Figs 1, 2). The distinctions between S. lignicola and S. auriculariicola are provided in the notes for S. auriculariicola. Morphologically, S. lignicola differs from S. philadelphianum by not producing conidia and by having chlamydospore-like cells swollen up to 7 µm wide (Pesante 1957; Crous et al. 2022). Furthermore, based on a pairwise comparison of ITS, S. lignicola (ex-type UAMH 1502) differs from S. philadelphianum (ex-type CPC 40793) in 5.8% (34/580 bp, 14 gaps) in the ITS. Scytalidium lignicola acts as a saprotrophic opportunist in wood, soil, and compost but can shift to a pathogenic mode of life, causing infections in humans (Dickinson et al. 1983; De Gannes et al. 2013). Notably, as a soil-borne pathogen, Scytalidium lignicola often causes cassava black root rot, which is difficult to control and results in significant losses in cassava production (Silva et al. 2013). 11. Scytalidium philadelphianum Crous & Jurjević, Fungal Syst. Evol. 10: 84 (2022) Description and illustration. Crous et al. (2022). Notes. Scytalidium philadelphianum was initially isolated from compressed air in a factory located in Philadelphia, Pennsylvania, USA (Crous et al. 2022). Scytalidium philadelphianum is phylogenetically closely related to S. auriculariicola and S. lignicola (Figs 1, 2). The distinctions between S. philadelphianum and S. auriculariicola are provided in the notes for S. auriculariicola, while the distinctions between S. philadelphianum and S. lignicola are provided in the notes for S. lignicola. Notably, Ringhofer et al. (2025) obtained the strain UTHSCSA DI24-300 from urine and intervertebral disc aspirate cultures of a 3-year-old male Belgian Malinois presenting with progressive hindlimb weakness. The isolate was identified as S. philadelphianum based on 100% similarity of its ITS with that of ex-type strain CPC 40793. 17 IMA Fungus 16: e164608 (2025), DOI: 10.3897/imafungus.16.164608 Shuo-Qiu Tong et al.: Scytalidium revisited 12. Scytalidium rodionovae S.Q. Tong & Zhi.Y. Zhang, sp. nov. MycoBank No: 860661 Type. RUSSIA • Moscow, isolated from a rotting rope in the 1970s, living culture 3C. Etymology. It was initially isolated by Rodionova et al. Description and illustration. Pavlov et al. (2018). Notes. Scytalidium rodionovae is phylogenetically closely related to S. album, S. assmuthi, S. aurantiacum, S. circinatum, S. terrigenum, and S. tongrenense (Figs 1, 2). The distinctions between S. rodionovae and S. album, S. assmuthi, and S. aurantiacum are detailed in the respective notes for S. album, S. assmuthi, and S. aurantiacum. Scytalidium rodionovae is distinguished from S. circinatum, S. terrigenum, and S. tongrenense by its known sexual morph, hyaline, ovoid, and septate chlamydospore-like cells (Sigler and Wang 1990; Jeong et al. 2025). Additionally, based on a pairwise comparison of ITS and LSU, S. rodionovae (ex-type 3C) differs from S. circinatum (ex-type CBS 654.89) by 4% (23/571 bp, four gaps) in the ITS and 1.1% (7/628 bp, no gap) in the LSU; from S. terrigenum (ex-type KNUF-23-236) by 7.8% (44/559 bp, six gaps) in the ITS and 3.2% (42/1299 bp, two gaps) in the LSU; from S. tongrenense (ex-type CGMCC 3.28994) by 9.7% (53/548 bp, 11 gaps) in the ITS and 3.7% (34/909 bp, one gap) in the LSU. Rodionova et al. (1974) identified strain 3C as Geotrichum candidum based only on morphological characters. Pavlov et al. (2018) integrated morphological characteristics and phylogenetic analyses based on molecular data derived from 3C to reclassify G. candidum into Scytalidium, naming it S. candidum. However, they overlooked the fact that 3C does not belong to G. candidum. Here, we propose the new species Scytalidium rodionovae to accommodate the species for which 3C is the ex-type. To date, the genome sequencing of strain 3C has been completed (Polev et al. 2014). Studies have shown that this strain secretes a variety of enzymes capable of efficiently degrading cellulose, exhibits broad pH adaptability, and holds potential application value in lignocellulosic biomass conversion and green industrial processes (Lapin et al. 2002; Borisova et al. 2015). 13. Scytalidium sphaerosporum Sigler & Kang, Mycologia 102(5): 1179 (2010) Description and illustration. Kang et al. (2010). Notes. Scytalidium sphaerosporum was initially isolated from wood chips of pine (Pinus sylvestris) in Sweden (Kang et al. 2010). Scytalidium sphaerosporum is phylogenetically closely related to S. chlamydosporum and S. cuboideum (Figs 1, 2). However, the distinctions between S. sphaerosporum and S. chlamydosporum are provided in the notes for S. chlamydosporum, while the distinctions between S. sphaerosporum and S. cuboideum are provided in the notes for S. cuboideum. Scytalidium sphaerosporum is commonly found in wood chips of both coniferous and broad-leaved trees, as well as in preservative-treated timber, and has no pathogenic relationship with Ganoderma (Kang et al. 2010; Goh et al. 2015). 18 IMA Fungus 16: e164608 (2025), DOI: 10.3897/imafungus.16.164608 Shuo-Qiu Tong et al.: Scytalidium revisited 14. Scytalidium synnematicum G.G. Barreto & Gusmão, Persoonia 50: 287 (2023) Description and illustration. Crous et al. (2023). Notes. Scytalidium synnematicum was isolated by Crous et al. (2023) as a saprobe from dead twigs of an unidentified plant collected in Amazonas, Brazil. Scytalidium synnematicum is phylogenetically closely related to S. ganodermophthorum (Figs 1, 2). However, the distinctions between S. synnematicum and S. ganodermophthorum are provided in the notes for S. ganodermophthorum. To date, S. synnematicum is the only species within the genus Scytalidium that produces synnemata (Crous et al. 2023). 15. Scytalidium terrigenum Y.S. Jeong, S.Yeol Lee & H.Y. Jung, Mycobiology 53(3): 297 (2025) Description and illustration. Jeong et al. (2025). Notes. Scytalidium terrigenum was isolated and named by Jeong et al. (2025) from a soil sample collected in Chungcheongnam-do, South Korea. Scytalidium terrigenum is phylogenetically closely related to S. album, S. assmuthi, S. aurantiacum, S. rodionovae, S. circinatum, and S. tongrenense (Figs 1, 2). However, the distinctions between S. terrigenum and S. album, S. assmuthi, S. aurantiacum, S. rodionovae, and S. circinatum are provided in their respective notes. Additionally, S. terrigenum differs from S. tongrenense by its production of hyaline to brown arthroconidia and oval chlamydospore-like cells (Jeong et al. 2025). Furthermore, based on a pairwise comparison of ITS and LSU, S. terrigenum (ex-type KNUF-23-236) differs from S. tongrenense (ex-type CGMCC 3.28994) by 5.5% (30/544 bp, seven gaps) in the ITS and 2.4% (22/895 bp, five gaps) in the LSU. Notably, S. terrigenum is capable of growing in acidic (pH 4) and low-temperature (10 °C) environments (Jeong et al. 2025). 16. Scytalidium tongrenense S.Q. Tong & Zhi.Y. Zhang, sp. nov. MycoBank No: 859965 Fig. 5 Type. CHINA • Guizhou, Tongren, Yanhe County, Qiaojia Town, 28.28°N, 108.44°E, soil, 5 July 2023, Shuo-Qiu Tong (holotype HMAS 354098, dried culture; ex-type CGMCC 3.28994, ibid., SQT13) Etymology. The epithet refers to the type location. Description. Culture characteristics (14 days at 25 °C): Colonies on PDA attaining 17–34 mm diam., flat, margin fimbriate, floral white (1A2) to white (1A1). Reverse floral white (1A2) to white (1A1). Colonies on SNA attaining 66– 68 mm diam., flat, margin entire, pale gray (30B2). Reverse pale gray (30B2). Chlamydospore-like cells yellow-brown to dark brown, oblong to globose, ellipsoidal, subcylindrical, guttulate, or irregular, 0–1-septate, thick-walled, smooth, catenate, 5.5–12 × 4.5–6 µm (avg. 8.1 × 5.2 μm, n = 30). Arthroconidia unknown. Sexual morph unknown. Geographical distribution. Guizhou Province, China. 19 IMA Fungus 16: e164608 (2025), DOI: 10.3897/imafungus.16.164608 Shuo-Qiu Tong et al.: Scytalidium revisited Additional material examined. CHINA • Guizhou, Guiyang, Huaxi District, Shiban Town, 26.46°N, 106.65°E, soil, 8 July 2022, Shuo-Qiu Tong, SQT14. GenBank numbers. SQT13–SQT14, ITS: PV890028–PV890029, LSU: PV890035–PV890036. Notes. Phylogenetic analysis showed that two new isolates (SQT13 and SQT14) clustered in a single subclade with high supported value (100/1) and are sister to S. album, S. assmuthi, S. aurantiacum, S. rodionovae, S. circinatum, and S. terrigenum (Figs 1, 2). However, the distinctions between S. tongrenense and S. album, S. assmuthi, S. aurantiacum, S. rodionovae, S. circinatum, and S. terrigenum are provided in their respective notes. Scytalidium tongrenense was isolated from soil in a pepper cultivation area and is likely to have a saprophytic lifestyle. Neocosmospora E.F. Sm., Bull. U.S.D.A. 17: 45. 1899. Neocosmospora xizangensis (Y.H. Geng & T.Y. Zhang) S.Q. Tong & Zhi.Y. Zhang, comb. nov. MycoBank No: 860664 Basionym. Scytalidium tuberculatum Y.H. Geng & T.Y. Zhang, Mycosystema 35(11): 1313. 2016. Description and illustration. Geng et al. (2016). Notes. Scytalidium tuberculatum was introduced by Geng et al. (2016). In this study, the ex-type strain (H1195) of S. tuberculatum nested in the genus Figure 5. Scytalidium tongrenense (from holotype HMAS 354098) a, b upper and reverse views of cultures on PDA and SNA after 14 d at 25 °C c–e chlamydospore-like cells. Scale bars: 10 μm (c); 20 μm (d, e). 20 IMA Fungus 16: e164608 (2025), DOI: 10.3897/imafungus.16.164608 Shuo-Qiu Tong et al.: Scytalidium revisited Neocosmospora (Fig. 1). However, the name Neocosmospora tuberculata is already occupied (Lee et al. 2025). Therefore, we transferred S. tuberculatum to the genus Neocosmospora as Neocosmospora xizangensis. Mycothermus D.O. Natvig, J.W. Taylor, A. Tsang, M.I. Hutch. & A.J. Powell ex X. Wei Wang, Houbraken & D.O. Natvig, Stud. Mycol. 93: 107. 2018. Mycothermus thermophilus (Cooney & R. Emers.) X. Wei Wang, Houbraken & D.O. Natvig, Stud. Mycol. 93: 107. 2018. Mycothermus thermophilus (Cooney & R. Emers.) D.O. Natvig et al., Mycologia 107: 321. 2015, nom. inval. Humicola insolens Cooney & R. Emers., Thermophilic Fungi: 72. 1964. Humicola grisea var. thermoides Cooney & R. Emers., Thermophilic Fungi: 72. 1964. Scytalidium indonesiacum Hedger, Samson & Basuki, Trans. Brit. Mycol. Soc. 78(2): 365. 1982. Basionym. Torula thermophila Cooney & R. Emers., Thermophilic Fungi: 92. 1964. Synonyms. Scytalidium thermophilum (Cooney & R. Emers.) Austwick, New Zealand J. Agric. Res. 19: 29. 1976. Description and illustration. Wang et al. (2019). Notes. Wang et al. (2019) validated Mycothermus thermophilus. In this study, the ex-type strain (CBS 259.81) of S. indonesiacum nested in the genus Mycothermus (Fig. 1). Although the clade containing S. indonesiacum (CBS 259.81) and M. thermophilus (CBS 625.91) received low statistical support (80/-), they exhibit minimal morphological and sequence divergence. The ITS and LSU sequence similarities between S. indonesiacum (CBS 259.81) and M. thermophilus (CBS 625.91) were 99% and 99.5%, respectively. Therefore, we synonymize S. indonesiacum under M. thermophilus. Hypoxylon Bull., Histoire des champignons de la France: 168. 1791. Hypoxylon terminale (G.V. Rao & de Hoog) S.Q. Tong & Zhi.Y. Zhang, comb. nov. MycoBank No: 860666 Synonyms. Scytalidium terminale G.V. Rao & de Hoog, Persoonia 8(2): 203. 1975. Description and illustration. Rao and Hoog (1975). Notes. Scytalidium terminale was introduced by Rao and Hoog (1975). In this study, the ex-type strain (CBS 171.40) of S. terminale nested in the genus Hypoxylon (Fig. 1). Therefore, we transferred S. terminale to the genus Scytalidium as H. terminale. Monochaetia (Sacc.) Allesch., Rabenhorst’s Kryptogamen-Flora, Pilze - Fungi Imperfecti Ed. 2, 1(7): 665. 1902. Monochaetia dimorphospora T. Yokoy., Trans. Brit. Mycol. Soc. 65(3): 500. 1975. Synonyms. Scytalidium flavobrunneum (J.H. Mill., Giddens & A.A. Foster) Sigler, Mycotaxon 4(2): 400. 1976. 21 IMA Fungus 16: e164608 (2025), DOI: 10.3897/imafungus.16.164608 Shuo-Qiu Tong et al.: Scytalidium revisited Description and illustration. Yokoyama (1975). Notes. Monochaetia dimorphospora was introduced by Yokoyama (1975). In this study, the ex-type strain (CBS 244.59) of S. flavobrunneum nested in the genus Monochaetia and is closely related to M. dimorphospora (ex-type NBRC 9980) (Fig. 1). Additionally, S. flavobrunneum and M. dimorphospora exhibit minimal morphological and sequence divergence. The ITS-LSU sequence similarities between S. flavobrunneum (CBS 244.59) and M. dimorphospora (NBRC 9980) were 99.9%. Therefore, we synonymize S. flavobrunneum under M. dimorphospora. Neodevriesia Quaedvl. & Crous, Persoonia 33: 24. 2014. Neodevriesia infestans (Iwatsu, Udagawa & Hatai) S.Q. Tong & Zhi.Y. Zhang, comb. nov. MycoBank No: 860667 Basionym. Scytalidium infestans Iwatsu, Udagawa & Hatai, Trans. Mycol. Soc. Japan 31(3): 389. 1990. Synonyms. Neodevriesia cladophorae M.M. Wang & W. Li, Mycologia 109(6): 967. 2018. Description and illustration. Iwatsu et al. (1990). Notes. Scytalidium infestans was introduced by Iwatsu et al. (1990). Neodevriesia cladophorae was introduced by Wang et al. (2017). In this study, the ex-type strain (CBS 161.91) of S. infestans nested in the genus Neodevriesia and was closely related to N. cladophorae (ex-type CGMCC 3.17901) (Fig. 1). Additionally, both S. infestans and N. cladophorae share similar conidia and chlamydospore-like cell morphology and dimensions (Iwatsu et al. 1990; Wang et al. 2017), along with highly conserved sequences. The ITS and LSU sequence similarities between S. infestans (CBS 161.91) and N. cladophorae (CGMCC 3.17901) were 99.6% and 100%, respectively. Since S. infestans was published prior to N. cladophorae, we transfer S. infestans to Neodevriesia as Neodevriesia infestans, with N. cladophorae being synonymized under it. Caliciopsis Peck, Rep. (Annual) New York State Mus. Nat. Hist. 33: 32. 1880. Caliciopsis uredinicola (Kuhlman, J.W. Carmich. & T. Mill.) Zhi.Y. Zhang, comb. nov. MycoBank No: 860668 Basionym. Scytalidium uredinicola Kuhlman, J.W. Carmich. & T. Mill., Mycologia 68(6): 1189. 1976. Description and illustration. Kuhlman et al. (1976). Notes. Scytalidium uredinicola was introduced by Kuhlman et al. (1976). In this study, the ex-type strain (CBS 578.75) of S. uredinicola nested in the genus Caliciopsis and was closely related to C. pinea (CBS 139.64) (Fig. 1). However, they can be distinguished by their low sequence similarities. The ITS sequence similarities between S. uredinicola (CBS 578.75) and C. pinea (CBS 139.64) were 97.1%. Therefore, we transfer S. uredinicola to Caliciopsis as Caliciopsis uredinicola. 22 IMA Fungus 16: e164608 (2025), DOI: 10.3897/imafungus.16.164608 Shuo-Qiu Tong et al.: Scytalidium revisited Excluded species 1. Scytalidium multiseptatum Hol.-Jech., Česká Mykol. 44(2): 101. 1990. Description and illustration. Holubová-Jechová (1990). Notes. In the phylogenetic tree (Fig. 1), S. multiseptatum (CBS 693.70 and CBS 241.68) formed a distinct clade within Tricladiaceae. Due to the lack of molecular sequence data from the ex-type strain (CBS 136.91) of S. multiseptatum, we excluded this species from Scytalidium, but further data are required to clarify its taxonomic status. 2. Scytalidium tibetense Y.H. Geng & T.Y. Zhang, Mycosystema 35(11): 1312. 2016. Description and illustration. Geng et al. (2016). Notes. In the phylogenetic tree (Fig. 1), S. tibetense (ex-type H1127) nested in the genus Fusarium. Fusarium is one of the most species-rich genera in Sordariomycetes, and its species identification requires a combination of multiple approaches. Therefore, we excluded S. tibetense from Scytalidium, but further studies are needed to determine its precise taxonomic status. 3. Scytalidium japonicum Udagawa, K. Tominaga & Hamaoka, Mycotaxon 25(1): 281 (1986) Description and illustration. Udagawa et al. (1986). Notes. In the phylogenetic tree (Fig. 1), the two strains of S. japonicum, CBS 494.88 (ex-type) and CBS 125804, clearly separated into two distinct clades. The CBS 494.88 formed an independent subclade within Dothideomycetes, and further studies are required to clarify its taxonomic status. Additionally, the CBS 125804 was embedded within Neoscytalidium and showed a close relationship with CBS 145.78 (ex-type of N. dimidiatum), sharing 99.6% ITS-LSU sequence similarity. However, due to the lack of morphological characterization for CBS 125804, we refrain from assigning it to N. dimidiatum at this time. 4. Scytalidium chinense Y.H. Geng & T.Y. Zhang, Mycosystema 35(11): 1311 (2016) Description and illustration. Geng et al. (2016). Notes. In the phylogenetic tree (Fig. 1), the H1091 (ex-type of Scytalidium chinense) formed an independent subclade within Dothideomycetes, and further studies are required to clarify its taxonomic status. Uncertain species During our compilation of literature and molecular data on Scytalidium, we identified several species for which no sequence data were available. Phylogenetic 23 IMA Fungus 16: e164608 (2025), DOI: 10.3897/imafungus.16.164608 Shuo-Qiu Tong et al.: Scytalidium revisited analyses revealed that multiple morphologically defined species originally classified in Scytalidium do not belong to this genus. Consequently, we designate these species as incertae sedis. 1. Scytalidium fulvum Morgan-Jones & Gintis, Mycologia 76(2): 214. 1984. Description and illustration. Morgan-Jones et al. (1984). 2. Scytalidium hepiali C. Lan Li, Acta Mycol. Sin. 7(1): 24. 1988. Description and illustration. Li and Sun (1988). 3. Scytalidium melanoxylicola N. Awasthi, A. Dubey, S. Bhardwaj & A.N. Rai, Kavaka 55: 108. 2020. Description and illustration. Awasthi et al. (2020). 4. Scytalidium nielamuense Y.M. Wu & T.Y. Zhang, Mycotaxon 114: 205. 2011. Description and illustration. Wu and Zhang (2010). 5. Scytalidium verruculosum Y.M. Wu & T.Y. Zhang, Mycotaxon 114: 207. 2011. Description and illustration. Wu and Zhang (2010). 6. Scytalidium xigazense Y.M. Wu & T.Y. Zhang, Mycotaxon 114: 209. 2011. Description and illustration. Wu and Zhang (2010). Discussion To date, the taxonomic status of Scytalidium remains unresolved. Based on large-scale molecular data from Leotiomycetes, Ekanayaka et al. (2019) placed Scytalidium within Helotiales under Hyaloscyphaceae in their phylogenetic analysis. However, their sampling only included sequences from S. vaccinii (a synonym of Hyaloscypha hepaticicola) (CBS 652.89), while omitting the type species S. lignicola. In the same year, Johnston et al. (2019) focused on type species within Leotiomycetes and conducted a phylogenetic analysis using 15 concatenated genes, indicating that Scytalidium occupies a basal position within Helotiales, albeit with low resolution. Their data, however, were derived from strain IHIA52 (i.e., DSM 105466 in GenBank, identified as S. lignicola), yet the ITS sequence (MG815782) of this strain shows a 10.1% divergence (60/593, 28 gaps) compared to the ITS sequence of the type strain of S. lignicola (UAMH 24 IMA Fungus 16: e164608 (2025), DOI: 10.3897/imafungus.16.164608 Shuo-Qiu Tong et al.: Scytalidium revisited 1502; AY762623). Furthermore, after Ellis (1971) defined Scytalidium as fungi with darkly pigmented conidia, Sigler and Carmichael (1976) included several additional species based on morphological characteristics, though these species are not necessarily phylogenetically related. This has resulted in an overly broad concept of the genus, which urgently requires revision. Scytalidium is characterized by two asexual morphs: one with dematiaceous intercalary conidia and another with hyaline, bacilliform arthroconidia (Pesante 1957; Ellis 1971). The arthroconidia are smooth, occasionally verrucose in age, mid or dark brown, cylindrical, oblong, doliform, or broadly ellipsoidal, and often 0-septate. Fission arthroconidia of a second type are hyaline or pale to mid-brown, smooth, cylindrical, 0-septate, and truncate at each end. Based on the original descriptions of 16 species within Scytalidium sensu stricto (Pesante 1957; Sigler and Wang 1990; Kang et al. 2010; Peng et al. 2014; Goh et al. 2015; Pavlov et al. 2018; Crous et al. 2022; Jeong et al. 2025), we propose that the characteristics of this genus are: ascomata cleistothecial, initially subhyaline, becoming dark brown at maturity with a wall of textura epidermoidea, without appendages, globose; asci subglobose to globose, quickly evanescent, fouror eight-spored; ascospores smooth, hyaline, subglobose to globose; conidia solitary or catenate, hyaline to dark brown; chlamydospore-like cells solitary or catenate, hyaline to dark brown, 0–1-septate, oblong to globose, subcylindrical, guttulate, or irregular; arthroconidia hyaline to light yellow, cuboidal to oblong or cylindrical. Traditional fungal taxonomy has predominantly relied on morphological characteristics for species classification and description (Hawksworth 2001). However, phenotypic convergence frequently obscures phylogenetic relationships, rendering morphology-based identification unreliable for certain taxa (Bickford et al. 2007; Otálora et al. 2017). Integrated taxonomy, which uses multiple methods to identify and characterize fungal species, is currently being embraced by the scientific community (Zhang et al. 2024). At present, increasing numbers of studies combine morphological characteristics and molecular phylogenetic reconstruction for identifying fungal species. This approach effectively reduces the taxonomic misclassification of phenotypically similar but phylogenetically distinct lineages. For instance, relying solely on morphological characteristics makes it challenging to accurately identify acremonium-like species (Hou et al. 2023) and members of Scytalidium studied here. It is noteworthy that although Vu et al. (2019) demonstrated the high efficacy of ITS and LSU in discriminating filamentous fungal species, according to the species delimitation criteria proposed by Jeewon and Hyde (2016), phylogenetic analyses should incorporate ITS and at least one protein-coding gene. However, since most species of Scytalidium are represented only by ITS and/or LSU (Table 1), this study, like recent studies on Scytalidium, had to rely on a combination of morphological characteristics and phylogenetic analyses based on concatenated ITS and LSU sequences for species identification (Crous et al. 2023; Jeong et al. 2025). This is a limitation, and future work will incorporate secondary barcodes or whole-genome data for phylogenetic reconstruction to further elucidate the taxonomic relationships within Scytalidium sensu stricto and sensu lato. Ecologically, Scytalidium has been reported to be associated with plants (wood, plant pathogens, dead branches), basidiomata, soil environments, and insects (Robinson et al. 2007; Kang et al. 2010; Peng et al. 2014; De Medeiros 25 IMA Fungus 16: e164608 (2025), DOI: 10.3897/imafungus.16.164608 Shuo-Qiu Tong et al.: Scytalidium revisited et al. 2019; Crous et al. 2023; Jeong et al. 2025). On one hand, they can cause plant diseases in certain crops, posing ecological threats to agriculture and forestry (Silva et al. 2013). On the other hand, due to their pigment production and various bioactive compounds, they demonstrate potential applications in wood and textile dyeing, as well as other fields (Lapin et al. 2002; Borisova et al. 2015; Hinsch et al. 2022). Additionally, numerous studies have shown that some species of Scytalidium, or species previously placed within the genus, are significant human pathogens capable of causing dermatomycoses, respiratory infections, and abscesses (Dickinson et al. 1983; Costa et al. 1988; Moore 1992; Machouart et al. 2013). The primary pathogenic species are S. hyalinum and S. dimidiatum (which have been merged and are now recognized as N. hyalinum; Phillips et al. 2013). Given that Scytalidium is a morphological genus, early identifications of clinical isolates based solely on morphological characteristics are questionable. With the incorporation of molecular data, almost all clinical isolates have been identified as either S. hyalinum or S. dimidiatum (Machouart et al. 2013; Enriquez-Mendez and Gonzalez 2025). Notably, Ringhofer et al. (2025) reported a case of disseminated fungal infection in a dog caused by S. philadelphianum. In this study, this species is placed within Scytalidium sensu stricto. In summary, this study has resolved the taxonomic status of most species within Scytalidium, although several taxa, including S. chinense and S. japonicum, remain to be conclusively classified. This will not only facilitate future taxonomic studies of Scytalidium and related taxa but also contribute to rapid diagnosis, prevention, and control measures in agricultural, forestry, and clinical settings. A key to accepted species in Scytalidium 1 Sexual and asexual morphs produced ......................................................... 2 – Only asexual morph produced ...................................................................... 3 2 Ascospores subglobose or globose, > 3 µm in diameter ...........................4 – Ascospores subglobose or globose, 3 µm in diameter ................................ ...................................................................................Scytalidium rodionovae 3 Arthroconidia not produced from synnemata .............................................5 – Arthroconidia produced from synnemata.........Scytalidium synnematicum 4 Ascomata 45–165 μm ............................Scytalidium ganodermophthorum – Ascomata 25–50 μm .......................................Scytalidium sphaerosporum 5 Production of only arthroconidia or chlamydospore-like cells ................... 6 – Simultaneous production of arthroconidia and chlamydospore-like cells ...7 6 Only arthroconidia produced ...................................Scytalidium cuboideum – Only chlamydospore-like cells produced .....................................................8 7 Arthroconidia hyaline to brown ....................................................................9 – Arthroconidia hyaline or color unknown ....................................................10 8 Chlamydospore-like cells aseptate ............................Scytalidium assmuthi – Chlamydospore-like cells 0–1-septate ......................................................11 9 Chlamydospore-like cells oval .................................Scytalidium terrigenum – Chlamydospore-like cells oblong or ellipsoidal to subglobose .................... ................................................................................ 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Mycosphere: Journal of Fungal Biology 15(1): 654–763. https://doi. org/10.5943/mycosphere/15/1/5 Supplementary material 1 Analysis 1 and Analysis 2 Authors: Shuo-Qiu Tong, Yi-Fan Yang, Peng Li, Yong-Jun Wu, Bing-Da Sun, Zhi-Yuan Zhang Data type: zip Copyright notice: This dataset is made available under the Open Database License (http://opendatacommons.org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow users to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited. Link: https://doi.org/10.3897/imafungus.16.164608.suppl1