Ecological, Morphological and Molecular Study of Swertia Chirayita (ROXB.EX FLEM) Karst of Nepal

Abstract

Swertia. chirayita is the most valuable and highly demanded species in national and International trade. In the present study, attempts have been made to conduct morphological, ecological and molecular study on S. chirayita. Morphological and ecological studies were conducted in Pharche VDC of Kaski district while for the molecular study, samples from four districts namely Sankhuwasabha, Terhathum, Kaski and Lalitpur were considered. The objectives of ecological and morphological studies were to explore quantitative information on the ecological status of S. chirayita in the study sites of ( Parche VDC, Kaski ) as well as the morphological variation of the plant population with respect to various altitudinal and geographical gradients. S. chirayita was found to occur between 1650m to 2700m but highest number of plants were found between 1900-2650m. The species was mostly concentrated on moist slopes than in dry and rocky places with density value ranging from 1.7-3.2 plant/ m 2 . Swertia chirayita was observed in south east, north east and south west face moist open habitats and under canopy of Acer and Quercus mixed forest. Swertia chirayita was also observed beyond the study sites at different altitudinal range but the number was very low. The regular grazing and movement of livestock were observed through out the study area but the higher pressure was seen in field site there i.e. Gorejure danda. The highest biomass content in the fresh sample of the plant of Swertia chirayita was found to be 69% while for air dry sample was found only 15.06% in the site at 2650m altitude in Gorujure danda..Correlation between morphological parameters shows that all morphological parameters except numbers of branches are significant correlated with altitude (p=0.01) within the range of studied area. The overall objective of the molecular study was to detect DNA sequence polymorphism in the nrDNA ITS sequences of Swertia chirayita samples collected from various geographical regions of eastern, central and western Nepal using Polymerase Chain Reaction (PCR)-Restriction Fragments Length Polymorphism (RFLP) and DNA sequencing based molecular marker tools. Preliminary studies employing two main marker systems viz. 1) PCR-RFLP on the PCR amplified ITS sequences and 2) DNA sequencing of nuclear ribosomal DNA ITS region for the generation of DNA barcodes for S. chirayita were performed. As an initial step, Polymerase Chain Reaction (PCR) amplification of Nuclear Ribosomal DNA Internal Transcribed Spacers (ITS) sequences for the samples collected from Kaski, Sankhuwasabha, Terhathum, and Lalitpur districts were amplified using specific primers. Restriction analysis patterns on amplified ITS sequences were generated using three different Restriction Endonucleases (REs) viz. Bst UI, EcoRI and XhoI. Of the three REs used in the study, enzyme EcoRI was revealed to be a non-cutter. Whereas with XhoI, single cut in the ITS sequences of all the S. chirayita genotypes resulted into two fragments in the agarose gels. With BstUI, all genotypes (except three genotypes from Sankhuwasabha) of S. chirayita were restricted into two fragments indicating single site in the ITS sequence.