Recovering trace reptile DNA from the illegal wildlife trade

Abstract

Purpose: The Illegal Wildlife Trade (IWT), aided by improved global transport, and the expansion of the internet, has facilitated the international demand for exotic reptiles. The risks associated with trafficking of live reptiles requires robust forensic techniques for detecting housed or transported animals. Detection of species of high IWT demand can be challenging due to the illicit nature of the trade, particularly when a specimen is missing. The ability to detect trace DNA in empty holdings and transport containers can be pivotal as a source of evidence. Methods: Vivaria, containing either a corn snake (Pantherophis guttatus) or boa (Boa constrictor), were set up and monitored for 24 h simulating reptile holdings. Once removed, Diamond Nucleic Acid Dye™ (DD) was sprayed on experimental glass and plastic tiles recovered from within the vivaria, and trace DNA was visualized using a Polilight. Trace DNA was amplified using a novel reptile target specific qPCR assay and sequenced to identify both species. Results: Movement patterns and scale imprints associated with reptile contact were visible on experimental tiles after DD-staining. Successful qPCR amplification and subsequent bi-directional Sanger sequencing confirmed the presence of both the species in the respective vivaria. DNA recovered from glass tiles had significantly greater amplification success than plastic tiles. Conclusions: DD revealed valuable information about the presence, and movement, of reptiles in the absence of a specimen. Successful amplification of trace reptile DNA demonstrated that this approach could offer an effective tool for biosecurity staff to rapidly identify live reptiles in the IWT.