RPB2 Loci Research Articles

Two new species of Cytospora (Diaporthales, Cytosporaceae) causing canker disease of Malusdomestica and M.sieversii in Xinjiang, China.

Apple tree canker is a serious disease caused by species of Cytospora. Xinjiang Uygur Autonomous Region is one of the most important apple-producing areas in China. However, losses due to apple Cytospora canker have seriously damaged the apple industry and affected the economic development of the apple growers in this region. In this study, we used morphological characteristics combined with multilocus phylogenetic analyses of the ITS, act, rpb2, tef1 and tub2 loci to identify isolates from apple (Malusdomestica) and wild apple (M.sieversii). As a result, C.hippophaopsis sp. nov. from M.sieversii and C.shawanensis sp. nov. from M.domestica were discovered and proposed herein. Pathogenicity tests were further conducted on 13 varieties of apple and wild apple, which confirmed C.hippophaopsis and C.shawanensis as canker pathogens. Meanwhile, C.hippophaopsis is generally more aggressive than C.shawanensis on the tested varieties of apple and wild apple.

Morphology and multigene phylogeny reveal four new Xylaria (Xylariales, Xylariaceae) species from karst region in China.

This study presents the identification of four novel Xylaria species, discovered in the karst region of China. The discovery was facilitated by a rigorous analysis that encompassed both morpho-anatomical features and multi-locus phylogenetics utilizing sequences from the ITS, rpb2, and TUB2 loci. The newly identified species are designated as Xylariajichuanii sp. nov., X.nanningensis sp. nov., X.orientalis sp. nov., and X.taiyangheensis sp. nov. The distinction of these species from their known counterparts was verified through comparison of morphological features and phylogenetic analysis. The study further provides detailed morphological descriptions, illustrative representations, and a phylogenetic tree, all of which contribute to the taxonomic positioning of these novel species.

The Origin of the Problem: Characterization of Paraguayan Septoria steviae, Causal Agent of Septoria Leaf Spot of Stevia, Based on Multilocus Sequence Analysis.

Septoria leaf spot is a significant disease affecting cultivated stevia, potentially reducing yields by > 50%. The disease is caused by Septoria steviae, first identified in 1978 in Japan as a new pathogen of stevia. Understanding the origin of S. steviae could clarify how it spread to new production areas. To investigate this, 12 isolates of Septoria sp. were obtained from stevia's native range in the Amambay forests and field plantings in Paraguay from 2018 to 2020. These isolates underwent colony morphology and molecular characterization of Actin, β-Tubulin, Calmodulin, ITS, LSU, RPB2, and TEF1α loci. GenBank sequences from S. steviae isolates collected in France, Japan, and the United States were included. Multilocus sequence phylogenetic analysis generated a maximum likelihood (ML) tree. The morphological characteristics of Paraguayan isolates were similar to those of previously reported S. steviae type cultures from Japan. The ML analysis showed that Paraguayan isolates formed a monophyletic group with S. steviae isolates from France, Japan, and the United States. During blotter tests, pycnidia and cirri of S. steviae were observed on multiple stevia seed surfaces from different sources. Further characterization confirmed viable pathogenic conidia of S. steviae. This observation suggests that S. steviae could be associated with stevia seed, possibly spreading from the center of origin to other countries. This research is the first to genetically characterize S. steviae from Paraguay and propose its potential spread mechanism from the center of origin to the rest of the world.

Botryosphaeriaceae species associated with branch dieback and decline of macadamia trees in South Africa

Botryosphaeriaceae species are important latent pathogens causing diseases on trees utilized in forestry and agriculture. In recent years, there has been an increase in the incidence and severity of branch dieback and decline on macadamia trees in South Africa, and species of Botryosphaeriaceae have been considered as a possible cause. Although botryosphaeria dieback has been well-studied in Australia, there is little information regarding these fungi on Macadamia in South Africa. The aims of this study were consequently to (i) identify species of Botryosphaeriaceae from Macadamia branches from main production regions in South Africa, (ii) compare the diversity of species between symptomatic and asymptomatic branches, as well as between different growing regions, (iii) and to consider their relative importance in causing dieback. Eight species and three putative hybrids of the Botryosphaeriaceae were identified based on a phylogenetic comparison of sequence data from the ITS rDNA, tub2, tef-1α and rpb2 loci. These included an unidentified Diplodia sp., and Lasiodiplodia sp., as well as L. gilanensis, L. theobromae, L. pseudotheobromae, Neofusicoccum kwambonambiense, N. luteum, N. parvum and three hybrid species. The unidentified species of Diplodia., Lasiodiplodia sp., L. gilanensis, and N. kwambonambiense are reported for the first time on Macadamia in South Africa. All species showed a potential to cause branch dieback symptoms, with species of Neofusicoccum identified as the most aggressive species. This study revealed a high level of diversity of Botryosphaeriaceae species and illustrates their potential as causal agents of dieback on Macadamia in South Africa.

A new lignicolous fungus, Rhexoacrodictys chiangraiensis (Pleurotheciales, Ascomycota), from Northern Thailand

ABSTRACT In this study, we introduce a novel species, Rhexoacrodictys chiangraiensis, collected from a decaying palm sheath (Arecaceae) in Northern Thailand. Morphological examination and phylogenetic analysis were carried out to identify the species. Maximum likelihood (ML) and Bayesian inference (BI) analyses were performed using the combined dataset of the internal transcribed spacer (ITS), large subunit ribosomal RNA (LSU), small subunit ribosomal RNA (SSU) and the RNA polymerase II second largest subunit (rpb2) loci. Phylogenetic analysis supported the introduction of the isolate as a novel taxon as it formed a basal clade to the other species of Rhexoacrodictys. Rhexoacrodictys chiangraiensis exhibits distinct morphological characteristics compared to other Rhexoacrodictys species by having obovoid to pyriform conidia that have smooth surfaces with 2–4 transverse septa and fewer longitudinal incomplete septa. The novel taxon was compared with other species of Rhexoacrodictys, and a comprehensive description and illustration are provided.

Identification and Pathogenicity of Fusarium Fungi Associated with Dry Rot of Potato Tubers.

Dry rot of potato tubers is a harmful disease caused by species of the Fusarium genus. Studies on the composition and features of Fusarium spp. that cause the disease in Russia are limited. Thirty-one Fusarium strains belonging to the F. sambucinum species complex (FSAMSC) and F. solani species complex (FSSC) were accurately identified using multilocus phylogenetic analysis of the tef and rpb2 loci, and their physiological characteristics were studied in detail. As a result, 21 strains of F. sambucinum s. str. and 1 strain of F. venenatum within the FSAMSC were identified. Among the analyzed strains within the FSSC, one strain of F. mori, four strains of F. noneumartii, and two strains of both F. stercicola and F. vanettenii species were identified. This is the first record of F. mori on potato as a novel host plant, and the first detection of F. noneumartii and F. stercicola species in Russia. The clear optimal temperature for the growth of the strains belonging to FSAMSC was noted to be 25 °C, with a growth rate of 11.6-15.0 mm/day, whereas, for the strains belonging to FSSC, the optimal temperature range was between 25 and 30 °C, with a growth rate of 5.5-14.1 mm/day. The distinctive ability of F. sambucinum strains to grow at 5 °C has been demonstrated. All analyzed Fusarium strains were pathogenic to potato cv. Gala and caused extensive damage of the tuber tissue at an incubation temperature of 23 °C for one month. Among the fungi belonging to the FSAMSC, the F. sambucinum strains were more aggressive and caused 23.9 ± 2.2 mm of necrosis in the tubers on average compared to the F. venenatum strain-17.7 ± 1.2 mm. Among the fungi belonging to the FSSC, the F. noneumartii strains were the most aggressive and caused 32.2 ± 0.8 mm of necrosis on average. The aggressiveness of the F. mori, F. stercicola, and especially the F. vanettenii strains was significantly lower: the average sizes of damage were 17.5 ± 0.5 mm, 17.2 ± 0.2 mm, and 12.5 ± 1.7 mm, respectively. At an incubation temperature of 5 °C, only the F. sambucinum strains caused tuber necroses in the range of 6.7 ± 0.5-15.9 ± 0.8 mm.

Two new species of Parastagonospora and a new species of Phaeoseptoriella (Phaeosphaeriaceae, Pleosporales) from grasslands in Yunnan Province, China.

During our investigation of microfungi on grasslands in Yunnan Province, China, three new fungal taxa associated with grasses were collected. Morphological observations and phylogenetic analyses of the combined SSU, LSU, ITS, tef1-α, and rpb2 loci based on maximum likelihood and Bayesian inference were used to reveal the taxonomic placement of these fungal taxa. This study introduces Parastagonosporayunnanensis, Para.zhaotongensis, Phaeoseptoriellapoaceicola. Parastagonosporayunnanensis is characterized by ampulliform or globose to subglobose conidiogenous cells, with conidia that are cylindrical to subcylindrical, 0-1-septate, rounded at the apex and slightly truncate at the base. Parastagonosporazhaotongensis features similar globose to subglobose conidiogenous cells but with 0-3-septate, cylindrical to subcylindrical conidia. Phaeoseptoriellapoaceicola is distinguished by its globose to subglobose conidiogenous cells and phragmosporous conidia that are initially hyaline, turn pale yellowish at maturity, and are 7-septate, cylindrical to subcylindrical, either straight or slightly curved. These discoveries underscore the significance of exploring and accurately identifying fungal taxa within Ascomycota, highlighting the species richness and potential for new species discoveries in grass-based habitats. The findings from this study expand our understanding of the taxonomy and phylogeny of grassland-associated Ascomycota, providing a foundation for further ecological and taxonomic studies of these fungi within their natural environments.

Taxonomic and phylogenetic characterisations of six species of Pleosporales (in Didymosphaeriaceae, Roussoellaceae and Nigrogranaceae) from China.

Pleosporales comprise a diverse group of fungi with a global distribution and significant ecological importance. A survey on Pleosporales (in Didymosphaeriaceae, Roussoellaceae and Nigrogranaceae) in Guizhou Province, China, was conducted. Specimens were identified, based on morphological characteristics and phylogenetic analyses using a dataset composed of ITS, LSU, SSU, tef1 and rpb2 loci. Maximum Likelihood (ML) and Bayesian analyses were performed. As a result, three new species (Neokalmusiakarka, Nigrogranaschinifolium and N.trachycarpus) have been discovered, along with two new records for China (Roussoellaneopustulans and R.doimaesalongensis) and a known species (Roussoellapseudohysterioides). Morphologically similar species and phylogenetically close taxa are compared and discussed. This study provides detailed information and descriptions of all newly-identified taxa.

Morphological and molecular characterization of Fusarium incarnatum as a causal disease agent of pepper (Capsicum annuum) fruit rot

AbstractChilli pepper (Capsicum annuum L.) is one of the most important commercially cultivated and consumed vegetables in Turkey. During a disease survey, typical symptoms of fruit rot were observed on mature chilli pepper fruits in several surveyed fields and on dried pepper fruits obtained from local retailers/bazaars in Hatay Province, Turkey. Disease incidence varied from 15 to 45% of the plants in the inspected fields. Following standard isolation procedures, 40 fungal isolates were isolated, purified and single‐spore cultures were obtained from surface‐disinfected, rotted dried pepper tissue. Of these isolates, six fungal isolates with dense, cottony white aerial mycelia that became beige with age, were isolated on a potato sucrose agar. All isolates were found to be pathogenic on artificially inoculated chilli pepper fruit. Based on morphological characteristics, the isolates were initially identified as Fusarium incarnatum (Desm.) Sacc. 1886. Morphological identification of F. incarnatum isolates was further confirmed by MALDI‐TOF and molecular analyses using the sequences of the internal transcribed spacer (ITS), partial translation elongation factor‐1α (TEF‐1α) and second largest subunit of RNA polymerase II (RPB2) loci. Phylogenetic analysis based on ITS, TEF‐1α, RPB2 and concatenation of TEF‐1α, RPB2 loci sequences performed with several isolates of Fusarium spp. confirmed that representative fungal isolates (MKUZF1 and MKUZF4) belong to F. incarnatum. To our knowledge, this is the first report of F. incarnatum causing fruit rot in chilli peppers grown in Turkey.

Paramicrosphaeropsis eriobotryae sp. nov., a new agent of loquat canker in Iran

AbstractLoquat (Eriobotrya japonica) is a subtropical evergreen plant, important in terms of both horticulture and green space in the urban environment. In recent years, symptoms of branch and trunk canker, dieback and decline have been observed in the main cultivation areas of this tree in Iran. To study the aetiology of loquat canker in Fars Province, branches and trunks of loquat trees with disease symptoms were sampled from 2018 to 2019. Several isolates of an unknown Paramicrosphaeropsis species (Didymellaceae, Pleosporales, Dothideomycetes) were recovered from the trunks and branches of infected loquat trees. Phylogenetic analysis of the isolates based on a multigene phylogeny of the ribosomal internal transcribed spacer (ITS), partial β‐tubulin gene (tub2) and the second‐largest subunit of RNA polymerase II (rpb2) loci revealed that the isolates belonged to a new taxon, Paramicrosphaeropsis eriobotryae sp. nov. Pathogenicity tests revealed that this species was able to cause canker, trunk flaking, dieback, decline, leaf yellowing, defoliation and growth reduction in loquat trees. A host range evaluation on detached branches of some important horticultural trees occurring adjacent to loquat tree orchards of Fars Province showed that P. eriobotryae might have a broader host range, as it was also able to infect almond and olive trees.

Morphological and Phylogenetic Characterisations Reveal Four New Species in Leptosphaeriaceae (Pleosporales, Dothideomycetes).

Leptosphaeriaceae is a widely distributed fungal family with diverse lifestyles. The family includes several genera that can be distinguished by morphology and molecular phylogenetic analysis. During our investigation of saprobic fungi on grasslands in Yunnan Province, China, four fungal taxa belonging to Leptosphaeriaceae associated with grasses were collected. Morphological observations and phylogenetic analyses of the combined SSU, LSU, ITS, tub2, and rpb2 loci based on maximum likelihood and Bayesian inference were used to reveal the taxonomic placement of these fungal taxa. This study introduces four new taxa, viz. Leptosphaeria yunnanensis, L. zhaotongensis, Paraleptosphaeria kunmingensis, and Plenodomus zhaotongensis. Colour photo plates, full descriptions, and a phylogenetic tree to show the placement of the new taxa are provided.

Plant-Associated Novel Didymellaceous Taxa in the South China Botanical Garden (Guangzhou, China)

The South China Botanical Garden (SCBG), one of the largest and oldest botanical gardens in China, conserves important plant germplasms of endangered species. Therefore, ensuring tree health and studying the associated mycobiome of the phyllosphere is essential to maintaining its visual aesthetics. During a survey of plant-associated microfungal species in SCBG, we collected several coelomycetous taxa. Phylogenetic relationships were evaluated based on the analyses of ITS, LSU, RPB2, and β-tubulin loci. The morphological features of the new collections were compared with those of existing species, emphasizing close phylogenetic affinities. Based on the morphological comparisons and multi-locus phylogeny, we introduce three new species. These are Ectophoma phoenicis sp. nov., Remotididymella fici-microcarpae sp. nov., and Stagonosporopsis pedicularis-striatae sp. nov. In addition, we describe a new host record for Allophoma tropica in the Didymellaceae. Detailed descriptions and illustrations are provided along with notes comparing allied species.

Russulaceae of the Pakaraima Mountains of Guyana. IV. New species forming a distinct lineage of Lactarius subg. Plinthogalus

ABSTRACT The Neotropics have recently emerged as an important region for studies of tropical ectomycorrhizal (ECM) fungi. Specific neotropical areas with high ECM host tree densities have ECM fungal diversities rivaling those of higher-latitude forests. Some forests of the Guiana Shield are dominated by endemic ECM trees of the Fabaceae, including species of Dicymbe (subfam. Detarioideae), Aldina (subfam. Papilionoideae), and Pakaraimaea (Cistaceae). One of the most species-rich ECM fungal families present in each of these systems is Russulaceae. Long-term sampling in forests in Guyana’s Pakaraima Mountains has revealed a number of species of the Russulaceae genera Lactarius, Lactifluus, and Russula. In this study, we document a previously unknown, distinct lineage of Lactarius subg. Plinthogalus containing eight species from the Guiana Shield. Here, we describe five of these species from Guyana as new to science: Lactarius humiphilus, Lactarius mycenoides, Lactarius guyanensis, Lactarius dicymbophilus, and Lactarius aurantiolamellatus. Morphological descriptions, habit, habitat, and known distribution are provided for each new species. Sequence data for the barcode internal transcribed spacer (ITS) locus are provided for types and most other collections of the new taxa, and a molecular phylogenetic analysis based on the ITS, 28S, and RPB2 (second-largest subunit of the RNA polymerase II) loci across the genus Lactarius corroborates their morphology-based infrageneric placement. The discovery of this lineage changes our insights into the biogeography and evolutionary history of Lactarius subg. Plinthogalus.

New species and records of Neomassaria, Oxydothis and Roussoella (Pezizomycotina, Ascomycota) associated with palm and bamboo from China.

Several micro fungi were gathered from bamboo and palm in Guizhou Province, China. In morphology, these taxa resemble Neomassaria, Roussoella and Oxydothis. Multi-gene phylogenetic analyses based on combined ITS, LSU, SSU, rpb2 and tef1 loci confirmed that two are new geographical records for China, (viz. Roussoellasiamensis, Neomassariafabacearum), while two of them are new to science (viz. Oxydothisfortunei sp. nov. and Roussoellabambusarum sp. nov.). The stromata of Roussoellabambusarum are similar to those of R.thailandica, but its ascospores are larger. In addition, multi-gene phylogenetic analyses show that Oxydothisfortunei is closely related to O.inaequalis, but the J- ascus subapical ring as well as the ascospores of O.inaequalis are smaller. Morphological descriptions and illustrations of all species are provided.

Contribution to a modern treatment of Graphidaceae biodiversity in South Africa: genera of tribe Graphideae with hyaline ascospores

AbstractAdditions and corrections are provided for the South African species of Graphidaceae tribe Graphideae with hyaline ascospores. Allographa oldayana I. Medeiros sp. nov. is described as new to science based on morphological, chemical and molecular data. The new species is characterized by lirellae with striate labia and a complete thalline margin, a completely carbonized excipulum, large, muriform ascospores, and the presence of hirtifructic acid. Allographa consanguinea (Müll. Arg.) Lücking, A. leptospora (Vain.) Lücking & Kalb, Diorygma aff. minisporum Kalb et al., Graphis crebra Vain., Gr. dupaxana Vain., Gr. furcata Fée, Gr. handelii Zahlbr., Gr. longula Kremp., Gr. pinicola Zahlbr., Gr. proserpens Vain, Gr. subhiascens (Müll. Arg.) Lücking and Platythecium sp. are reported as new records for South Africa. Allographa striatula (Ach.) Lücking & Kalb, Graphis analoga Nyl. and Gr. scripta (L.) Ach. are shown to be misapplied names that should be removed from the South African checklist. The new combination Mangoldia bylii (Vain.) I. Medeiros comb. nov. (bas. Graphis bylii Vain) is made; this represents an earlier name for M. atronitens (A. W. Archer) Lücking et al. Taxonomic notes are provided for Graphis bylii var. lividula Vain. and Gr. denudans Vain., species that are known only from their South African holotypes. Phylogenetic analyses that include new DNA sequence data from the nrLSU, mtSSU and RPB2 loci confirm the generic placements of several species for which molecular data were lacking: Allographa consanguinea, Glyphis atrofusca (Müll. Arg.) Lücking, Graphis crebra and Gr. subhiascens.

Multigene phylogeny, morphology, and pathogenicity trials reveal novel Cytospora species involved in perennial canker disease of apple trees in Iran

In recent years, canker and die-back diseases have become a growing threat for the productivity and longevity of apple orchards in Iran. In this study, 131 Cytospora isolates were recovered from symptomatic tissues of apple trees in apple orchards of Iran. Multigene phylogenetic inference based on combined sequence data of ITS, act, rpb2, and tef1-α loci, supplemented with morphological characteristics and pathogenicity assay revealed four novel Cytospora species which were described as C. avicennae, C. azerbaijanica, C. ershadii, and C. iranica, and four known species, namely C. chrysosperma, C. parasitica, C. paratranslucens, and C. sorbicola. Also, C. sorbicola is newly reported on apple trees worldwide. Koch's postulates were fulfilled to confirm that all eight species in this study were pathogenic on apple trees in Iran, among which C. sorbicola was the most intensive species. The results of this study further highlight rich diversity among Cytospora species occurring on apple trees, revealing several novel Cytospora species on this host. The host range, geographical distribution, and economic significance of novel species on apple industry remains to be studied.

Taxonomic and Phylogenetic Characterizations Reveal Four New Species, Two New Asexual Morph Reports, and Six New Country Records of Bambusicolous Roussoella from China.

During the ongoing investigation of bambusicolous ascomycetous fungi in Yunnan, China, 24 specimens belonging to the family Roussoellaceae were collected and identified based on morphological features and phylogenetic support. Maximum-likelihood (ML) analyses and Bayesian analyses were generated based on the combined data set of ITS, LSU, tef1, and rpb2 loci. The phylogenetic analyses revealed four novel lineages in Roussoella s. str.; thus, we introduced four new species viz., Roussoella multiloculate sp. nov., R. papillate sp. nov., R. sinensis sp. nov., and R. uniloculata sp. nov. Their morphological characters were compared with the known Roussoella taxa, which lack sequence data in the GenBank. Asexual morphs of R. kunmingensis and R. padinae were recorded from dead bamboo culms in China (from the natural substrates) for the first time. Neoroussoella bambusae, Roussoella japanensis, R. nitidula, R. padinae, R. scabrispora, and R. tuberculate were also reported as the first records from China. All new taxa are described and illustrated in detail. Plates are provided for new reports.

Taxonomy and Phylogeny of Novel and Extant Taxa in Pleosporales Associated with Mangifera indica from Yunnan, China (Series I).

Pleosporales is the largest fungal order with a worldwide distribution in terrestrial and aquatic environments. During investigations of saprobic fungi associated with mango (Mangifera indica) in Baoshan and Honghe, Yunnan, China, fungal taxa belonging to pleosporales were collected. Morphological examinations and phylogenetic analyses of ITS, LSU, SSU, rpb2 and tef1-α loci were used to identify the fungal taxa. A new genus, Mangifericomes; four new species, namely Mangifericomes hongheensis, Neomassaria hongheensis, Paramonodictys hongheensis, and Paramonodictys yunnanensis; and six new host and country records, namely Byssosphaeria siamensis, Crassiparies quadrisporus, Paradictyoarthrinium aquatica, Phaeoseptum mali, Torula fici, and Vaginatispora amygdali, are introduced. Photoplates, full descriptions, and phylogenetic trees to show the placement of new and known taxa are provided.

Supplementary files

Fig. S1. Multi-locus phylogenetic tree (benA, CaM and RPB2 loci) of Aspergillus section Candidi comprising isolates from GenBank (Supplementary Table S3) and those included in Table 1. Best scoring Maximum Likelihood tree inferred in the IQ-Tree is shown, bootstrap values are appended to nodes, only support values higher than 70 % are shown. The ex-type strains are designated with a superscripted T and bold print.<br />Table S1. Comparison of micromorphological characters between species of Aspergillus section Candidi and statistical significances.<br />Table S2. Delimitation of isolates into populations by BPP v. 4.3.<br />Table S3.<br /> Aspergillus isolates from the section Candidi used in the analysis of ecology together with strains listed in Table 1.

Supplementary files

Fig. S1. Multilocus phylogeny of Aspergillus series Nigri based on three loci (benA, CaM and RPB2) without species A. chiangmaiensis, A. pseudopiperis and A. pseudotubingensis. Best-scoring Maximum Likelihood (ML) tree inferred in the IQ-TREE is shown; ultrafast bootstrap support values (ML bs) are appended to nodes; only support values ≥95 % are shown; the ex-type strains are designated with a bold print; the information on geographic origin and isolation source is plotted on the tree (see the legend). <br />Fig. S2. Comparison of single-gene genealogies based on the benA, CaM and RPB2 loci and created by three different phylogenetic methods (only one isolate per unique multilocus haplotype is included in each phylogeny). Best-scoring single-locus maximum likelihood (ML) trees are shown; ML ultrafast bootstrap support values (ML bs), maximum parsimony bootstrap support values (MP bs) and Bayesian inference posterior probabilities (BI pp) are appended to nodes. Only support values ≥95 %, ≥70 % and ≥0.95, respectively, are shown. A dash indicates lower statistical support for a specific node, or the absence of a node in the phylogeny, while an asterisk indicates full support. The ex-type strains are designated with a bold print. Alignment characteristics, partitioning schemes and substitution models are listed in Supplementary Table S1.<br />Fig. S3. Best scoring maximum likelihood tree inferred in IQ-TREE based on ITS rDNA region sequences of series Nigri members (Table 1). It is apparent from the tree that a limited variability present in this locus can only differentiate the three main lineages, and more detailed differentiation is not possible. The ITS sequence of the A. vadensis ex-type was excluded from the analysis because it contains numerous errors and unresolved positions (accession number AY585549). Alignment characteristics, partitioning schemes and substitution models are listed in Supplementary Table S1.<br />Table S1. Characteristics of alignments, partition-merging results and best substitution model for each partition according to the Bayesian information criterion.<br />Table S2. Assignment of strains into populations for analysis in DELINEATE based on BPP v. 4.3.<br />Table S3. Aspergillus series Nigri genomes analyzed in this study and their basic characteristics.<br />Table S4. Distribution of 200 orthologous genes into ten separate STACEY analyses.<br />Table S5. List of unique multilocus haplotypes.<br />Table S6. Maximum sequence dissimilarity between isolates of the same Aspergillus species whose species limits have been delimited using methods based on a multispecies coalescent model; only species represented by isolates from at least three countries were included.