A pilot study-genetic diversity and population structure of snow leopards of Gilgit-Baltistan, Pakistan, using molecular techniques.

BACKGROUND: The Hindu Kush and Karakoram mountain ranges in Pakistan's northern areas are a natural habitat of the snow leopard (Panthera uncia syn. Uncia uncia) but the ecological studies on this animal are scarce since it is human shy by nature and lives in difficult mountainous tracts. The pilot study is conducted to exploit the genetic diversity and population structure of the snow leopard in this selected natural habitat of the member of the wildcat family in Pakistan. METHOD: About 50 putative scat samples of snow leopard from five localities of Gilgit-Baltistan (Pakistan) along with a control sample of zoo maintained male snow leopard were collected for comparison. Significant quality and quantity of genomic DNA was extracted from scat samples using combined Zhang-phenol-chloroform method and successful amplification of cytochrome c oxidase I gene (190 bp) using mini-barcode primers, seven simple sequence repeats (SSR) markers and Y-linked AMELY gene (200 bp) was done. RESULTS: Cytochrome c oxidase I gene sequencing suggested that 33/50 (66%) scat samples were of snow leopard. AMELY primer suggested that out of 33 amplified samples, 21 (63.63%) scats were from male and 12 (36.36%) from female leopards. Through successful amplification of DNA of 25 out of 33 (75.75%) scat samples using SSR markers, a total of 68 alleles on seven SSR loci were identified, showing low heterozygosity, while high gene flow between population. DISCUSSION: The low gene flow rate among the population results in low genetic diversity causing decreased diversification. This affects the adaptability to climatic changes, thus ultimately resulting in decreased population size of the species.

Illegal product manufacturing and exportation from Pakistan: Revealing the factuality of highly processed wildlife skin samples via DNA mini-barcoding.

Illegal trade is a major threat to the biodiversity and the efforts initiated for the conservation of wildlife. The shortcomings of the traditional taxonomic identification methods have been coped by a revolutionary and emerging technique, the "DNA (Deoxyribonucleic Acid) barcoding". Here we report a case of trader who was allegedly making footwear for a famous international celebrity from wild animal cutis. The samples confiscated during a raid on a footwear manufacturing industry by KP Wildlife department in August, 2016, were received by Bioresource Research Centre (BRC) for molecular identification on 1st September, 2016. The study costed about USD 88 from processing to the identification of the samples. The samples identified via DNA mini-barcoding by targeting cytochrome oxidase I (COI) gene belong to Gazella bennettii and Bos taurus. Such studies are helpful for credible investigations that only lead to effective prosecution and control of illegal wildlife trade ultimately helping in conservation of wild animals.

DNA Mini-barcoding for wildlife trade control: a case study on identification of highly processed animal materials.

Although DNA barcoding is an efficient tool for species identification, however, its efficiency is uncertain for samples having degraded DNA and incomplete isolation/amplification of COI gene fragment (>500 bp). DNA mini-barcoding is a solution to this problem because small DNA fragment of COI genes is used for species identification. Twelve highly processed, chemically treated and finished animal skin (coats, tanned skins) and fur (mufflers) samples, received from the Sindh Wildlife Department, Pakistan, were subjected to DNA mini-barcoding. Eight mufflers belonged to Vulpes vulpes, one coat to Ursus thibetanus, one tanned skin to Lutra sumatrana, and one muffler to Vulpes sp. Origin of only one coat sample remained unidentified, success rate of 92% indicative of the fact that the mini barcoding technique can be used as a substitute of conventional barcoding where full length barcode (∼650 bp Folmer region) cannot be generated.