DOI: 10.5061/dryad.ffbg79d43
title: A new species of Lepiota with epithelial pileal covering

description: A new species, Lepiota keralensis, was collected from Kerala, India and
 was studied using morphological and molecular (ITS, LSU and rpb2 gene)
 characters. The species is characterized by medium-sized basidiomata with
 innate squarrose brown coloured squamules on a white background, hyaline,
 dextrinoid, non-metachromatic, smooth-walled basidiospores, absence of
 cheilocystidia and pleurocystidia, and globose to ellipsoid cells on
 pileal covering. Morphological as well as phylogenetic analyses confirm
 the placement of the new species in Lepiota sect. Echinatae. The
 discovery of the new species adds to the existing data on tropical
 lepiotas.
description: <em>DNA extraction, PCR amplification and
 sequencing</em> DNA extraction was done using the
 RED Extract-N-Amp kit by Sigma-Aldrich Company. ITS1F and ITS4R were the
 primers used for the amplification of the ITS gene region, LR5 and LROR
 were used for LSU gene amplification and bRPB26-F and bRPB2-7.1R primers
 were used for the<em> rpb2</em> gene. (White et al.1990,
 Vilgalys and Hester 1990, Matheny 2005). A quality check for PCR products
 was carried out by gel electrophoresis (2% agarose gel) after PCR
 amplification. PCR amplicons were purified using QIAGEN QIAquick PCR
 Purification Kit. Purified samples were taken for sequencing. The
 sequencing reaction was set-up in Applied Biosystems™ MiniAmp™ Plus
 Thermal cycler using Big Dye™ Terminator V3.1 kit. Forward and reverse DNA
 sequencing reactions of PCR amplicons were carried out with respective
 primers (ITS1F, ITS4R, LROR, LR5, bRPB26-F and bRPB2-7.1R) using BDT v3.1
 Cycle sequencing kit on ABI 3730xl Genetic Analyzer. Sequencing was done
 by Barcode Biosciences Private Limited, Bangalore, India. All the
 sequences generated were deposited in GenBank (www.ncbi.nlm.nih.gov) and the accession numbers of the sequences are given in Table 1. <em>Sequence alignment and Phylogenetic analysis</em> A comparison of the newly generated sequences with already available sequences in GenBank was done using BLASTN software (https://blast.ncbi.nlm.nih.gov/Blast.cgi). The data matrix (combined ITS, LSU and <em>rpb2 </em>sequences) was constructed to include newly generated sequences, sequences retrieved from GenBank (based on closest hits) and sequences of taxa from previous studies (Hou and Ge 2020, Niazi et al. 2021). The final data set used for analysis comprised of sequences with high sequence similarity index, and included 183 sequences from 111 taxa with representatives from each section in <em>Lepiota</em>, other closely allied genera like<em> Cystolepiota</em>,<em> Echinoderma</em>, <em>Leucoagaricus</em>, <em>Leucocoprinus</em>, <em>Macrolepiota</em>, <em>Pseudolepiota</em>. <em>Agaricus</em> was set as the outgroup. Another data matrix with only ITS sequences of all taxa considered in the combined matrix (sequences from 110 taxa) was also assembled. The data sets were aligned with MAFFT web tool (https://mafft.cbrc.jp/alignment/server). The MAFFT aligned data sets were manually aligned using MEGA X64 (Kumar et al. 2018). Maximum Likelihood (ML) analysis was conducted using IQTREE (IQTREE Web Server: Fast and accurate phylogenetic trees under maximum likelihood (univie.ac.at)) (Nguyen et al. 2015) for both the datasets using GTR model of nucleotide substitution with Gamma rate heterogeneity (as calculated by IQTREE), sand ultrafast bootstrap analysis with 1000 bootstrap alignments (Hoang et al. 2018). The phylogenetic trees obtained were processed with FigTree v1.2.3 (Rambaut 2014).
description: # A new species of Lepiota with epithelial pileal covering
 [https://doi.org/10.5061/dryad.ffbg79d43](https://doi.org/10.5061/dryad.ffbg79d43) ## Description of the data and file structure It is the combined data set (ITS,LSU & rpb2 sequence) used for the phylogenetic analysis of new species of lepiota. ### Files and variables #### File: its\_lsu\_rpb2\_-\_last\_150824\_1.fas **Description:** The final data set used for analysis comprised of sequences with high sequence similarity index, and included 183 sequences from 111 taxa with representatives from each section in *Lepiota*, other closely allied genera like *Cystolepiota*, *Echinoderma*, *Leucoagaricus*, *Leucocoprinus*, *Macrolepiota*, *Pseudolepiota*. *Agaricus* was set as the outgroup. ## Code/software The data sets were aligned with MAFFT web tool ([https://mafft.cbrc.jp/alignment/server](https://mafft.cbrc.jp/alignment/server)). The MAFFT aligned data sets were manually aligned using MEGA X64. Maximum Likelihood (ML) analysis was conducted using IQTREE (IQTREE Web Server: Fast and accurate phylogenetic trees under maximum likelihood (univie.ac.at)).The phylogenetic trees obtained were processed with FigTree v1.2.3. ## Access information Other publicly accessible locations of the data: * Genbank and Mycobank