DOI: 10.5061/dryad.8sf7m0cnq
title: Hyphodermella zixishanensis (Polyporales, Basidiomycota), a new species with reddish hymenial surface

description: A new corticioid fungal species, Hyphodermella zixishanensis, is described
 based on combination of morphological features and molecular evidence. The
 species is characterized by an annual growth habit, resupinate basidiomata
 with reddish, a monomitic hyphal system with generative hyphae bearing
 simple septa, IKI–, CB– and ellipsoid, colorless, thin-walled,
 smooth basidiospores. Sequences of ITS and nLSU gene regions were
 generated, and phylogenetic analyses were performed with maximum
 likelihood, maximum parsimony and Bayesian inference methods. The
 phylogenetic analyses showed that the species belongs to
 Hyphodermella and placed as sister taxon to H. aurantiaca.
description: <b>Morphological
 studies</b> The studied specimens
 were deposited at the herbarium of Southwest Forestry University (SWFC),
 Kunming, Yunnan Province, P.R. China. Macromorphological descriptions are
 based on field notes. Colour terms follow Petersen (1996).
 Micromorphological data were obtained from dried specimens, and observed
 under a light microscope following Dai (2012). The following abbreviations
 were used: KOH = 5% potassium hydroxide, CB = Cotton Blue, CB– =
 acyanophilous, IKI = Melzer’s reagent, IKI– = both inamyloid and
 indextrinoid, L = mean spore length (arithmetic average of all spores), W
 = mean spore width (arithmetic average of all spores), Q = variation in
 the L/W ratios between the specimens studied, n (a/b) = number of spores
 (a) measured from given number (b) of specimens. <b>Molecular techniques and
 phylogenetic analyses</b> CTAB rapid plant
 genome extraction kit-DN14 (Aidlab Biotechnologies Co., Ltd, Beijing) was
 used to obtain genomic DNA from dried specimens. A small piece of dried
 fungal specimen (about 30 mg) was ground to powder with liquid nitrogen.
 The powder was transferred to a 1.5 ml centrifuge tube, suspended in 0.4
 ml of lysis buffer, and incubated in a 65 °C water bath for 60 min. After
 that, 0.4 ml phenol-chloroform (24:1) was added and the suspension was
 shaken vigorously. After centrifugation at 13,000 rpm for 5 min, 0.3
 ml supernatant was transferred to a new tube and mixed with 0.45
 ml binding buffer. The mixture was then transferred to an adsorbing column
 (AC) for centrifugation at 13,000 rpm for 0.5 min. Then, 0.5 ml inhibitor
 removal fluid was added in AC for a centrifugation at 12,000 rpm for 0.5
 min. After washing twice with 0.5 ml washing buffer, the AC was
 transferred to a clean centrifuge tube, and 100 ml elution buffer was
 added to the middle of the adsorbed film to elute the DNA. ITS region was
 amplified with primer pair ITS5 and ITS4 (White et al. 1990). Nuclear LSU
 region was amplified with primer pair LR0R and LR7
 (http://www.biology.duke.edu/fungi/mycolab/primers.htm). The PCR procedure
 for ITS was as follows: initial denaturation at 95°C for 3 min, followed
 by 35 cycles at 94°C for 40 s, 58°C for 45 s and 72°C for 1 min, and a
 final extension of 72°C for 10 min. The PCR procedure for nLSU was as
 follows: initial denaturation at 94°C for 1 min, followed by 35 cycles at
 94°C for 30 s, 48°C 1 min and 72°C for 1.5 min, and a final extension of
 72°C for 10 min. The PCR products were purified and directly sequenced at
 Kunming Tsingke Biological Technology Limited Company, Kunming, Yunnan
 Province, P.R. China. All newly generated sequences were deposited at
 GenBank (Table 1). Sequences were aligned in MAFFT
 6 (Katoh and Toh 2008; http://mafft.cbrc.jp/alignment/ server/) using the
 “G-INS-I” strategy for nLSU, the “E-INS-I” strategy for ITS+nLSU, and
 manually adjusted in BioEdit (Hall 1999). Alignment datasets
 were deposited in TreeBase (submission ID
 28053). <i>Candelabrochaete africana </i>Boidin obtained from
 GenBank was used as an outgroup to root the LSU tree (Fig. 1.) following
 Justo et al. (2017) and <i>Pirex concentricus</i> was selected
 as an outgroup for phylogenetic analyses of the ITS+nLSU region (Fig. 2)
 following Zhao et al. (2017). Maximum
 parsimony analyses were applied to the ITS+nLSU and nLSU dataset
 sequences. Approaches to phylogenetic analysis followed Chen et
 al. (2016), and the tree construction procedure was performed in PAUP*
 version 4.0b10 (Swofford 2002). All characters were equally weighted and
 gaps were treated as missing data. Trees were inferred using the heuristic
 search option with TBR branch swapping and 1000 random sequence additions.
 Max-trees was set to 5000, branches of zero length were collapsed and all
 parsimonious trees were saved. Clade robustness was assessed using a
 bootstrap (BT) analysis with 1000 replicates (Felsenstein 1985).
 Descriptive tree statistics tree length (TL), consistency index (CI),
 retention index (RI), rescaled consistency index (RC), and homoplasy index
 (HI) were calculated for each maximum parsimonious tree generated.
 Sequences were also analyzed using Maximum Likelihood (ML) with RAxML-HPC2
 through the Cipres Science Gateway
 (http://www.phylo.org/sub_sections/portal). Branch support (BS) for ML
 analysis was determined by 1000 bootstrap replicate.                      
                               MrModeltest 2.3 (Nylander 2004) was used to
 determine the best-fit evolution model for each data set for Bayesian
 inference (BI). Bayesian inference was calculated with MrBayes3.1.2 (Ronquist and Huelsenbeck 2003). Four Markov chains were run for 2 runs from random starting trees for 250 thousand generations for nLSU (Fig. 1) and 100 thousand generations for ITS+nLSU (Fig. 2), and trees were sampled every 100 generations. The first one-fourth generations were discarded as burn-in. A majority rule consensus tree of all remaining trees was calculated. Branches were considered as significantly supported if they received ML bootstrap value &gt;75%, maximum parsimony bootstrap value (MP) &gt;75%, or Bayesian posterior probabilities (PP) &gt;0.95.