From single species to communities: microsatellite amplicon sequencing to monitor felids using Feliplex
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The current biodiversity crisis demands a shift from single-species to multispecies approaches in conservation, particularly for rare and endangered species. However, this transition requires tools optimised for multispecies research, which are currently limited.
Recent advances in high-throughput sequencing (HTS) technologies and bioinformatics have enabled efficient and robust acquisition of genetic data. Amplicon sequencing approaches, in particular, have demonstrated potential for enhancing non-invasive genetic studies of endangered species, but their application has been mostly limited to single species.
To enable multispecies genetic research, we introduce a cost-effective and robust HTS-based amplicon sequencing approach for genotyping multiple species simultaneously, designed for population monitoring, including individual identification and ascertaining patterns of population structure.
We developed Feliplex, a multiplex panel of 85 co-amplifying tetranucleotide microsatellite markers for cross-genotyping Felidae species, to demonstrate the utility of our approach. Feliplex was validated on known samples from nine Indian felid species across the genera Panthera, Prionailurus , and Felis . We applied it to invasive (blood and tissue) and non-invasive (hair and faeces) DNA extracts from 173 wild individuals obtaining respectively 70% and 56% multilocus genotyping success rates. The panel accurately identified known population clusters in tigers ( Panthera tigris , n=19) and revealed hitherto unknown genetic structure in fishing cats ( Prionailurus viverrinus , n=40).
Feliplex’s wide applicability across Felidae allows reliable multispecies genotyping from low-quality/quantity samples, while supporting cost-effective genetic studies and conservation monitoring of lesser-known species like small cats. Our approach has a broad applicability and can be adapted to develop similar multispecies panels for closely related species groups.
