For controlling illegal trade in wildlife parts and products in Southeast Asia, WII has initiated the work of establishing Wildlife Forensic Cell (WFC) for proper implementation of Indian Wildlife (Protection) Act-1972. More than 600 wildlife offences have so far been referred by various agencies such as Forest, DRI, Courts, CBI, police and others. Morphology is one of the strongest tools for identifying species from animal parts like hair, bones, claws, shells, tusks, skins, horns etc. We have standardized protocols for identifying species commonly found in trade from hair, claws, tusk, bear biles, musk pods, tiger bones, canine etc. We discuss a few specific studies and development taken place in WFC. More than 70% of the case exhibits received at forensic lab involve the identification of species form hair. Keeping this in view hair characteristic profiles (Cuticle, medulla, cross-section) have been prepared for the species commonly found in wildlife trade (n=52) using light and scanning electron microscopy (SEM). We determined medulla percentages and scale count indices for the hair of 52 species. The species could be characterized successfully based on this study. However microscopic studies suffer from observational biases and are ineffective at low sample sizes, and require corroboration with other studies. Therefore, we developed protocols for identifying species with help of electrophoresis techniques like SDS-PAGE (Sodium dodecyl sulphate polyacrylamide gel electrophoresis) and IEF (iso electric focussing) using keratin protein profiles. We were able to characterize species (Shahtoosh, pashmina, angora, brown bear, black bear, sloth bear, wild boar & domestic pig) with the help of SDS-PAGE and IEF. In addition this thin layer chromatographic (TLC) studies were carried out to identify bear biles (bear gall bladders). We worked with different solvent systems (n=15) and spraying reagents (n=5) for development and developed a protocol that was 2 times faster in separating bear bile acids & 5 times more sensitive in identifying them in comparison to existing protocols.

DNA techniques are useful for identifying species from meat and other tissues (skin, blood, bones etc.) which have not been preserved properly and precluded to use of protein based techniques. In Wildlife Forensic Cell, we have around 350 cases of meat samples which have not been preserved properly and there is a need to develop protocols for identifying species using DNA techniques. Restriction Fragment Length Polymorphism (RFLP) and DNA sequencing are among the well established techniques in wildlife forensics for identifying species. We initiated the work of standardizing these protocols. Extraction of DNA is the main challenge for forensic DNA works. Sometimes the samples are either in very bad quality and quantity or fixed in formalin which in turn may inhibits Polymerase Chain Reaction. Isolation of DNA from various tissue samples was done using different protocols viz. phenol/chloroform (PC) (n=54), DNeasy tissue kit (Qiagen, Germany) (n=100), Chelex-100 (n=100) and Bio-Robot (n=16) with necessary modifications whenever required. 28 per cent samples examined (n=270) did not yield any DNA. Quality of the DNA obtained from these samples was in the order of DNA-Bio robot > DNeasy tissue kit, (Qiagen, Germany) > PC > Chelex.  48% amplification was achieved in Polymerase Chain Reaction with mitochondrial cytochrome b universal primers of 359 bp (Meyer et al., 1995) or 146 bp (Farell et al., 2000). Eight restriction enzymes were used to digest PCR amplified products of different species (n=12) to get species-specific amplicon. Three restriction enzymes (HaeIII, HinfI and NIaII) were found suitable to differentiate tiger and leopard whereas wild and domestic pigs could be differentiated by StuI. Combination of NIaIII and StuI were able to differentiate Chinkara from other deer species. DNA sequencer will be used to sequence the amplicon of different species to complement RFLP profiles.

Under WFC’s new initiatives, we have planned to ascertain the source of origin of species in wildlife poaching cases like tiger, leopard, musk deer, bears and elephant tusk. Addl. P.C.C.F and Chief Wildlife Warden, Forest Department, Govt. of Uttaranachal has evinced interest to develop data base on population genotypes of key species of the states. Forest Officials have been asked to provide samples while undertaking post-mortem. It has been possible to obtain around 28 samples and preserved properly for further analysis.