Taxonomic And DNA Barcoding Of The Genus Melanoleuca In Northeast China

The genus Melanoleuca Pat is a kind of wild mushroom.The genus species is mainly typified by a tricholomatoid habit,dull color,strongly amyloid and warted spores,distinct cheilocystidia,and a lack of clamp connections.Majority of the genus are edible mushroom with delicious taste and rich nutrition.In this study,a detailed morphological study and DNA barcode study were conducted based on 44 samples which belonged to 14 Melanleuca species from northeast region of China,and the phylogenetic relationship of the genus was also reconstructed by the optimal region.First of all,based on the morphological description of 14 Melanoleuca species from northeastern China,a key for the species was provided in this study.And the features,such as fruit body,gills,spores and cystidia,were discussed about their value for the taxonomic study of Melanleuca species.Then,four fragments(ITS,nr LSU,RPB2,tef-?)were sequenced for DNA barcode using 32 specimens.ITS region was seperated ITS1 and ITS2 for further analyses.A total of six regions were used for the DNA barcode analysis.The result showed that the region ITS had obvious differences between intra-specific and inter-specific.And the NJ tree could distinguish all the species.However,it was difficult to differ all the speceis using the other five regions.Therefore,ITS was proved as a optimal DNA barcode fragment for the genus Melanoleuca.Finally,based on ITS sequences,the phylogenetic relationship was reconstructed among the 14 Melanoleuca species.The result showed that there were five clades within the genus.Clade 1 included M.grammopodia,it was characterized by large sporocarp with greyish pileus and urticiform cystidia,longitudinal striate on the surface of stipe.Clade 2 included M.brevipes,the main features were medium size pileus,and striate on the surface of stipe.Clade 3 included M.exscissa,M.substrictipes and M.verrucipes.The main characters were light color,and larger spores.Clade 4 included M.strictipes,M.graminicola,M.subalpina,M.leucopoda,M.melaleuca,M.arcuata,M.stridula and M.cognata.The main characters were colored gills,light color stipe and expanded base.Clade 5 included M.porphyropoda,which was characterized by small cap,purple stipe and absence of cystsidia.