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Abstract All species of big cats, including tigers, cheetahs, leopards, lions, snow leopards, and jaguars, are protected under the Convention on the International Trade in Endangered Species (CITES). This is due in large part to population declines resulting from anthropogenic factors, especially poaching and the unregulated and illegal trade in pelts, bones, teeth and other products that are derived from these iconic species. To enhance and scale up monitoring for big cat products in this trade, we created a rapid multiplex qPCR test that can identify and differentiate DNA from tiger (Panthera tigris), cheetah (Acinonyx jubatus), leopard (Panthera pardus), lion (Panthera leo), snow leopard (Panthera uncia), and jaguar (Panthera onca) in wildlife products using melt curve analysis to identify each species by its unique melt peak temperature. Our results showed high PCR efficiency (> 90%), sensitivity (detection limit of 5 copies of DNA per PCR reaction) and specificity (no cross amplification between each of the 6 big cat species). When paired with a rapid (

The ability to reliably genotype individual bald eagle (Haliaeetus leucocephalus) and golden eagle (Aquila chrysaetos) samples has proven to be a valuable forensic tool for investigations of illegal trafficking of eagle feathers and body parts. Twenty STR loci were assembled into three optimized multiplex STR panels. The three STR multiplex PCRs include loci ranging from dinucleotide to pentanucleotide motifs, and both monomorphic and polymorphic amplification. The number of alleles for bald eagles ranged 1–5 (14 polymorphic loci), and the number of alleles for golden eagles ranged from 1 to 11 (16 polymorphic loci). Bald and golden eagles shared alleles at 5 loci, the other 15 loci yielding alleles characteristic of each species. These multiplex panels will be useful in wildlife law enforcement investigations in which eagle feathers and body parts can be identified to species and individuals matched to link a crime scene and eagle victim to a suspect.

One of the biggest threats to sustainability of shark populations is overexploitation due to high global market demand for fins, meat, skin, and cartilage. Current laws designed to regulate the worldwide trade in certain shark species have not stemmed illegal and unreported trade. In January 2020, United States Fish and Wildlife Service (USFWS) wildlife inspectors seized an illegal shipment of shark fins at the Miami airport during an inspection of an in-transit shipment from South America to Asia. The shipment contained over 5000 fins, placing it among the largest seizures in the US to date. We utilized a combination of morphological and genetic analyses to evaluate species diversity of the sharks targeted for this illegal shipment. Morphological identification of pectoral (n = 2435) and dorsal (n = 1301) fins revealed over 50 % were obtained from CITES Appendix II species, including silky shark (Carcharhinus falciformis), great hammerhead (Sphyrna mokarran), and scalloped hammerhead (Sphyrna lewini). A random subset of dorsal fins (n = 149) was sampled from each of the CITES species and the tiger shark (Galeocerdo cuvier) fins (n = 49), to confirm our morphological identifications using genetic data. In addition, a subset of morphologically unidentified fins (n = 192) was genetically identified to estimate species diversity of the fins not identified by morphological analysis. The genetic results revealed 11 additional species for a total of 14 species identified within the shipment. The data from the genetic and morphological analysis of this case will be used to inform ongoing investigations of the origin of this illegal shipment, as well as future investigations of shark fisheries undergoing exploitation for the shark fin trade.

The majority of DNA casework processed by forensic laboratories focuses on human samples, but material from canids (dogs, wolves, coyotes) can also be encountered. Undomesticated canids can be the center of forensic investigations in the U.S. since some species are endangered. As many wolf species are similar morphologically, identification in the field by wildlife officers is not always straight-forward, making molecular based-approaches ideal. While some published methods using mitochondrial DNA targets can discriminate among Canis species, they are either not compatible with highly degraded samples or cannot differentiate closely related sub-species. Although some U.S. laboratories regularly perform veterinary/wildlife casework including canid identifications, their validated methods and reference genetic databases are not publicly available. We aimed to assess the utility of alternative regions in the mitochondrial genome for discriminating among canid species, including the complete genome. To achieve this, we utilized a commercial hybridization capture panel composed of biotinylated RNA “baits” designed for the domestic dog to enrich canid mitochondrial genomes for next-generation sequencing. We used this panel to successfully sequence complete mitochondrial genomes for 51 samples, representing four U.S. forensically relevant canids (coyote, wolf, Mexican wolf, dog). While the complete mitochondrial genome permitted discrimination, we also assessed previously published mitochondrial DNA targets (n, 5) for resolution and identified four alternate ∼200 bp fragments from ND1, ND5, COI and CYTB genes that could help resolve canids. The utility of these alternate regions should be assessed in future studies using forensic-type samples representing canids from diverse geographic areas, prior to casework implementation.

Wildlife forensic analyses are frequently concerned with taxonomic identification, and very often employ amplification and Sanger sequencing of informative regions of the genome to achieve this. The materials submitted to wildlife forensic laboratories for taxonomic identification span a wide scope, from plant and animal parts in trade to assemblages of incidental biota at crime scenes. As these analyses take place within the context of legal proceedings, the wildlife forensic community is subject to unique requirements and considerations. These requirements and considerations are quite different from those of human forensic DNA, and have driven standardization in this field. While there has been extensive debate over appropriate DNA-based methods for taxonomic identification of a wide variety of biota in research settings, there has been little discussion on the issues associated with this approach in the high scrutiny environment of forensic science. This review outlines: key procedural and biological factors that may impact the accuracy of interpretation and reporting taxonomic identifications; resulting conventions employed by the wildlife forensics community; and implications for the use of emergent DNA sequencing technologies in taxonomic identification of wildlife in casework.