Molecular identification of Lepiota acutesquamosa and L. cristata (Basidiomycota, Agaricales) based on ITS-rDNA barcoding from Himalayan moist temperate forests of Pakistan

Lepiota acutesquamosa and L. cristata (Basidiomycota, Agaricales) collected from Himalayan moist temperate forests of Pakistan were characterized using internal transcribed spacers (ITS) of rNDA, a fungal molecular marker. The ITS-rDNA of both species was analyzed using polymerase chain reaction (PCR) and DNA sequencing. The target region when amplified using universal fungal primers (ITSIF and ITS4) generated 650-650bp fragments. Consensus sequences of both species were submitted for initial blast analysis which revealed and confirmed the identification of both species by comparing the sequences of these respective species already present in the GenBank. Sequence of Pakistani collection of L. acutesquamosa matched 99% with sequences of same species (FJ998400) and Pakistani L. cristata matched 97% with its sequences (EU081956, U85327, AJ237628). Further, in phylogenetic analysis both species distinctly clustered with their respective groups. Morphological characters like shape, size and color of basidiomata, basidiospore size, basidial lengths, shape and size of cheilocystidia of both collections were measured and compared. Both these species have been described first time from Pakistan on morph-anatomical and molecular basis.