First record of the rare genus Typhrasa (Psathyrellaceae, Agaricales) from China with description of two new species

Abstract Typhrasa is a rare genus that comprises two species and that has previously been reported only from Europe and North America. The present study expands the geographical scope of the genus by describing two new species – T. polycystis and T. rugocephala – from subtropical China. The new species are supported by morphological characteristics and phylogenetic analyses (ITS, LSU and tef-1α). The new species have very similar morphological characteristics and are 98% similar in their ITS region. However, T. rugocephala has two types of long gills at the same time, rarely fusiform pleurocystidia with rostrum. Detailed descriptions, colour photos, illustrations and a key to related species are presented in this paper.


Introduction
The genus Typhrasa Örstadius & E. Larss. was established in 2015. It is characterised by a hygrophanous cap, crowded gills with white edge, small-to-medium-sized spores, large hymenial cystidia with intracellular oily drops or globules and a hymeniderm or paraderm pileipellis (Örstadius et al. 2015). The genus includes two species, T. gossypina (Bull.) Örstadius & E. Larss. and T. nanispora Örstadius, Hauskn. & E. Larss., the former being previously reported occasionally from some countries in Europe, North America and Asia within the genus Psathyrella (Fr.) Quél (Smith 1972;Kits van Waveren 1985;Knudsen and Vesterholt 2012;Örstadius et al. 2015). In addition, Psathyrella delineata (Peck) A.H. Sm., P. canadensis A.H. Sm. and P. subtenacipes A.H. Sm. are also reported to have oily drops in their cystidia (Smith 1972;Kits van Waveren 1985) and seems to be a candidate for Typhrasa. However, P. delineata and P. canadensis were combined into T. gossypina, based on the morphology (Örstadius et al. 2015). During investigations in subtropical China during 2018-2020, Typhrasa was recorded for the first time in China with two unrecorded species, which were frequently collected. Based on morphological characters and phylogenetic analyses, they are described as new species in this paper.

Morphological studies
Macromorphological characters and habitat details were noted from fresh, young to mature basidiomata (over five basidiomata for each species) in the field. The location of the collection point is marked on the map (Suppl. material 1: Fig. S1). Colour codes are from the Methuen Handbook of Colour (Kornerup and Wanscher 1978). Micromorphological characters were observed with a light microscope (Olympus BX53). Sections from dry specimens were observed in water, 5% aqueous potassium hydroxide (KOH) solution, 10% aqueous ammonia (NH 3 ·H 2 O) solution and Melzer's Reagent, separately. More than fifty basidiospores, cystidia and basidia in 5% aqueous KOH solution were measured under the microscope. Basidiospore measurements were recorded in front and profile view. The measurements and Q values are given as (a)b-c(d), in which "a" is the lowest value, "b-c" covers a minimum of 90% of the values and "d" is the highest value. "Q" represents the ratio of length to width of a spore (Bas 1969;Ge et al. 2017;Na and Bau 2019). Specimens are deposited in the Herbarium of Fungi, Jiangxi Agricultural University (HFJAU) and Herbarium of Mycology, Jilin Agricultural University (HMJAU).

Data analyses
The ITS, LSU and Tef-1α datasets were assembled following Örstadius et al. (2015) and BLAST in GenBank. Sequences from a total of 24 taxa were analysed using five data partitions (ITS, LSU, Tef 1 st , Tef 2 nd and Tef 3 rd ). The details are presented in Table 1. Sequences were aligned separately in MAFFT v.7 (Katoh and Standley 2013). The best-fit models of nucleotide evolution for ITS, LSU, Tef 1 st , Tef 2 nd and Tef 3 rd datasets (GTR+G, GTR+I, SYM, SYM and GTR+G, respectively) were obtained in MrModeltest v.2.3 (Nylander et al. 2008). Phylogenetic analysis was conducted using Bayesian Inference (BI) in MrBayes v.3.2.6 (Ronquist et al. 2012). Gaps were treated as missing data following Örstadius et al. (2015). Four Monte Carlo Markov Table 1. Sequences used in this study. Newly generated sequences are given in bold. Type material is indicated in the column Voucher.
Ecology and distribution. Saprotrophic, solitary or gregarious on soil or humus in broad-leaved forests.

Discussion
Typhrasa was established by Örstadius et al. (2015), based on the main characters of having rostrate, hymenial cystidia with oily drops. Only T. gossypina and T. nanispora Örstadius, Hauskn. & E. Larss. were reported in that study. T. gossypina, as the type species of the genus, was, therefore, separated from Psathyrella (Fr.) Quél. This species can be separated from the two new species through its longer spores up to 9.0 μm long, 5.0-6.0 μm broad in front view and pleurocystidia often with a long rostrum (Kits van Waveren 1985;Örstadius et al. 2015). T. nanispora has smaller spores, 5.0-6.0 × 3.0-4.0 μm and can be thereby easily be distinguished (Örstadius et al. 2015). T. rugocephala is very easily confused with T. polycystis, but the former has two types of long gills arranged at intervals, scanty cheilocystidia and rarely rostrum can be found in fusiform pleurocystidia. In addition, P. subtenacipes are also reported to have oily drops in their cystidia (Smith 1972) and seems to be a candidate for Typhrasa, but study of type material is needed to settle this question. Morphologically, the spores of P. subtenacipes are up to 7.8-9.5 × 5.0-5.6 μm, significantly larger than the two new species (Smith et al. 1950;Smith 1972). A key to these related species is presented below: