How methodological changes have influenced our understanding of population structure in threatened species: insights from tiger populations across India.

Unprecedented advances in sequencing technology in the past decade allow a better understanding of genetic variation and its partitioning in natural populations. Such inference is critical to conservation: to understand species biology and identify isolated populations. We review empirical population genetics studies of Endangered Bengal tigers within India, where 60-70% of wild tigers live. We assess how changes in marker type and sampling strategy have impacted inferences by reviewing past studies, and presenting three novel analyses including a single-nucleotide polymorphism (SNP) panel, genome-wide SNP markers, and a whole-mitochondrial genome network. At a broad spatial scale, less than 100 SNPs revealed the same patterns of population clustering as whole genomes (with the exception of one additional population sampled only in the SNP panel). Mitochondrial DNA indicates a strong structure between the northeast and other regions. Two studies with more populations sampled revealed further substructure within Central India. Overall, the comparison of studies with varied marker types and sample sets allows more rigorous inference of population structure. Yet sampling of some populations is limited across all studies, and these should be the focus of future sampling efforts. We discuss challenges in our understanding of population structure, and how to further address relevant questions in conservation genetics. This article is part of the theme issue 'Celebrating 50 years since Lewontin's apportionment of human diversity'.

An environmental DNA sampling method for aye-ayes from their feeding traces.

Noninvasive sampling is an important development in population genetic monitoring of wild animals. Particularly, the collection of environmental DNA (eDNA) which can be collected without needing to encounter the target animal facilitates the genetic analysis of endangered species. One method that has been applied to these sample types is target capture and enrichment which overcomes the issue of high proportions of exogenous (nonhost) DNA from these lower quality samples. We tested whether target capture of mitochondrial DNA from sampled feeding traces of the aye-aye, an endangered lemur species would yield mitochondrial DNA sequences for population genetic monitoring. We sampled gnawed wood where aye-ayes excavate wood-boring insect larvae from trees. We designed RNA probes complementary to the aye-aye's mitochondrial genome and used these to isolate aye-aye DNA from other nontarget DNA in these samples. We successfully retrieved six near-complete mitochondrial genomes from two sites within the aye-aye's geographic range that had not been sampled previously. Our method demonstrates the application of next-generation molecular techniques to species of conservation concern. This method can likely be applied to alternative foraged remains to sample endangered species other than aye-ayes.