Carnivore diet analysis based on next‐generation sequencing: application to the leopard cat (Prionailurus bengalensis) in Pakistan

Diet analysis is a prerequisite to fully understand the biology of a species and the functioning of ecosystems. For carnivores, traditional diet analyses mostly rely upon the morphological identification of undigested remains in the faeces. Here, we developed a methodology for carnivore diet analyses based on the next‐generation sequencing. We applied this approach to the analysis of the vertebrate component of leopard cat diet in two ecologically distinct regions in northern Pakistan. Despite being a relatively common species with a wide distribution in Asia, little is known about this elusive predator. We analysed a total of 38 leopard cat faeces. After a classical DNA extraction, the DNA extracts were amplified using primers for vertebrates targeting about 100 bp of the mitochondrial 12S rRNA gene, with and without a blocking oligonucleotide specific to the predator sequence. The amplification products were then sequenced on a next‐generation sequencer. We identified a total of 18 prey taxa, including eight mammals, eight birds, one amphibian and one fish. In general, our results confirmed that the leopard cat has a very eclectic diet and feeds mainly on rodents and particularly on the Muridae family. The DNA‐based approach we propose here represents a valuable complement to current conventional methods. It can be applied to other carnivore species with only a slight adjustment relating to the design of the blocking oligonucleotide. It is robust and simple to implement and allows the possibility of very large‐scale analyses.