﻿Three new species of Cortinarius section Delibuti (Cortinariaceae, Agaricales) from China

﻿Abstract Three new species of CortinariussectionDelibuti, namely C.fibrillososalor, C.pseudosalor, and C.subtropicus are described as new to science based on morphological and phylogenetic evidences. Cortinariuspseudosalor is extremely morphologically similar to C.salor, but it differs from the latter by smaller coarsely verrucose basidiospores. Cortinariusfibrillososalor can be easily differentiated by its fibrillose pileus. The pileus of C.subtropicus becomes brown without lilac tint at maturity comparing with other members of section Delibuti. A combined dataset of ITS and LSU sequences was used for phylogenetic analysis. The phylogenetic reconstruction of section Delibuti revealed that these three new species clustered and formed independent lineages with full support respectively. A key to the three new species and related species of section Delibuti is provided in this work.

The research on Cortinarius has mainly been conducted in Europe and North America, while it is still lacking in East Asia (Peintner et al. 2002;Garnica et al. 2003;Peintner et al. 2004;Garnica et al. 2005;Liimatainen et al. 2014;Stefani et al. 2014;Garnica et al. 2016;Niskanen et al. 2016;Soop et al. 2019, Liimatainen et al. 2022).To date, fewer than 30 species were originally reported from China, and only two new species in sect.Delibuti were originally found in China (Yang 1998;Wei and Yao 2013;Xie et al. 2019Xie et al. , 2020Xie et al. , 2021aXie et al. , 2021bXie et al. , 2022;;Luo andBau 2021, Zhang et al. 2023;Zhou et al. 2023).With the combination of morphological observations and phylogenetic analysis, we describe three species belonging to sect.Delibuti as new to science in this study.

Specimens
The specimens were collected from central and southwestern China during 2012-2022.The vouchers are all deposited in the Mycological Herbarium of Hunan Normal University (MHHNU) and Cryptogamic Herbarium of Kunming Institute of Botany, Chinese Academy of Sciences (KUN-HKAS).Detailed information is listed in Table 1.

Morphological observation
The descriptions of macromorphological characters were based on field records and photographs.Color codes were used following Kornerup and Wanscher (1978).The size of basidiomes, as determined by pileus width, was described as small (< 5.0 cm), medium-sized (5.0-9.0 cm) or large (> 9.0 cm).Microscopic features were observed from dried specimens that were mounted with 5% aqueous KOH and stained with 1% Congo red solution under a light microscope (Motic Ltd., China).Melzer's reagent was used as an indicator of the

Phylogenetic analyses
The sequences newly generated in this study and downloaded from GenBank were used for phylogenetic analysis (Table 1).Alignment was performed by MAFFT v7.149b (Katoh and Standley 2013) and adjusted manually by MEGA5 (Tamura et al. 2011).SequenceMatrix 1.7.8 (Vaidya et al. 2011) was applied to generate multigene matrixes.GTR+I+G was selected as the best-fit model for combined matrix based on the Akaike Information Criterion (AIC) by MrModeltest 2.3 (Nylander 2004).Maximum likelihood (ML) analysis was performed using the W-IQ-TREE web service (http://iqtree.cibiv.univie.ac.at/) with 1000 ultrafast bootstrap replicates (Trifinopoulos et al. 2016).Bayesian inference (BI) was performed in MrBayes v3.2 (Ronquist et al. 2012).Four Metropolis-coupled Monte Carlo Markov chains were run for 5000000 generations, sampling every 1000 th generation.Subsequently, the sampled trees were summarized after omitting the first 25% of trees as burn-in.

Phylogenetic analyses
In the concatenated dataset (ITS+LSU), a total of 78 sequences (48 ITS, 30 LSU) from 48 samples were used for phylogenetic analyses among sect.Delibuti, sect.Subtorti, sect.Camphorati, sect.Bolares, sect.Spilomei, and sect.Anomali, of which 24 sequences (13 ITS,11 LSU) were newly yielded in this study (Table 1).The estimates of tree topology inferred from ML and Bayesian analyses were extremely similar.The ML phylogenetic tree is shown with both bootstrap values (BP) and posterior probabilities (PP) annotated near the nodes (Fig. 1).
The phylogenetic relationship of sections within the genus Cortinarius in the present study was unclear and weakly supported.In the multi-locus tree, the monophyly of sect.Delibuti was supported with well-supported values (BP = 99%, PP = 1.00), including 12 species.Section Camphorati was also monophyletic with fully supported values (BP = 100%, PP = 1.00), emcompassing 5 species.In sect.Diagnosis.Differs from the other species of sect.Delibuti from its fibrillose pileus.
Habitat, ecology and distribution.Solitary to gregarious on soil in evergreen broad-leaved forest, known from Hunan, China; July to September.

Cortinarius pseudosalor
Diagnosis.This species differs from other species in sect.Delibuti for its high morphological similarity with C. salor, but having smaller coarsely verrucose basidiospores.
Habitat, ecology and distribution.Solitary to gregarious on soil in coniferous and broad-leaved mixed forest or evergreen broad-leaved forest, known from Hunan and Hubei, China; August.
Notes.Cortinarius pseudosalor is easily misidentified as C. salor for their high morphological similarity, except the former has smaller coarsely verrucose basidiospores.Besides, C. pseudosalor distributed in Central China under coniferous and broad-leaved mixed forest or evergreen broad-leaved forest at alt. 1000-1413 m.Etymology.Subtropicus (Latin) refers to subtropical distribution range of the species.
Diagnosis.Differs from the other species of sect.Delibuti species in having an epicutis pileipellis that can be easily separated from the context of the pileus.
Habitat, ecology and distribution.Solitary to gregarious on soil in under evergreen broad-leaved forest, on the ground, known from Hunan, China; July.Notes.Cortinarius subtropicus has an epicutis pileipellis that can be easily separated from the context of the pileus.Besides, pileus become brown without lilac or dark olive tint at maturity comparing with other members of section Delibuti.(Kibby and Tortelli 2021;Xie et al. 2021a).Meanwhile, C. salor is characterized by its lilaceous lamellae all the time and the narrow distribution in European woodlands, while C. pseudosalor and C. subsalor occurs in Asia (Xie et al. 2021a).Cortinarius fibrillososalor with violaceous to whitish mauve tint differ from other species in this section in the appearance of fibrils on the pileus and its broadly globose to long ellipsoid basidiospores (Kibby and Tortelli 2021;Xie et al. 2021a).Cortinarius subtropicus was found in the subtropical monsoon climate region of the Hunan and Guangxi provinces distributed from 1625 m to 1900 m.Cortinarius tibeticisalor was only distributed in Tibetan Plateau and was usually olivaceous to brown at maturity, while olivaceous species in sect.Delibuti mainly occurred in the South Pacific (Soop et al. 2019;Xie et al. 2021a).
Phylogenetic analysis was first applied to the taxonomic study of Cortinarius with ITS (Liu et al. 1997).Later, phylogenetic relationships were inferred mainly based on ITS, LSU sequences, and rpb1, rpb2 were also confirmed to help elucidate the relationships of species in Cortinarius (Frøslev et al. 2005;Soop et al. 2019;Xie et al. 2022).Species delimitation could be justified by the combination of ITS and LSU sequences (Nilsen et al. 2021;Zhou et al. 2023), a two-locus dataset (ITS and LSU) was used for the research of three new species and their similar species in the present study.However, it needs more sequence data and DNA markers for recognising higher taxonomic rank such as subgenus or genus.In section rank, a two-locus dataset (ITS, LSU) and four-locus dataset (ITS, LSU, rpb1 and rpb2) were first employed for phylogenetic analyses, and the latter provided a higher BP value and clearer tree structure, although with limited rpb1 and rpb2 (Soop et al. 2019).Besides, the first phylogenomic study based on shallow whole genome sequencing was conducted for Cortinariaceae revision (Liimatainen et al. 2022).
amyloidity of basidiospores.In the description of basidiospores, the abbreviation [n/m/p] represents that the measurements were made on n basidiospores from m basidiomes of p collections.At least twenty matured basidiospores and basidia from each of the basidiomes were measured.The range (a)b-c(d) stands for the dimensions of basidiospores in which b-c contains a minimum of 90% of the measured values, while a and d indicate the extreme values.In addition, a Q value shows the ratio of length to width of basidiospores, and a Qm value shows the average Q ± standard deviation.A JSM-6380LV scanning electron microscope (JEOL Ltd., Tokyo, Japan) was used for the observation of ornamentations of basidiospores.

Figure 1 .
Figure 1.Phylogenetic tree of Cortinarius sect.Delibuti inferred from a combined matrix of ITS and LSU through maximum likelihood and Bayesian inference.Bayesian posterior probabilities (PP) > 0.90 and bootstrap values (BP) >85% are reported at the nodes (PP/BP); "-" indicates that the support value was less than the respective threshold.The three newly described species are highlighted in bold.Aus: Australia; EA: East Asia; Eur: Europe; NAm: North America; NZ: New Zealand.
Pseudosalor (Latin) refers to the species morphologically similar to Cortinarius salor.

Table 1 .
List of sequences of Cortinarius used for phylogenetic analyses.The sequences newly generated in this study are in bold, and all type specimens are highlighted with an asterisk.