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'.

High frequency of an otherwise rare phenotype in a small and isolated tiger population.

Most endangered species exist today in small populations, many of which are isolated. Evolution in such populations is largely governed by genetic drift. Empirical evidence for drift affecting striking phenotypes based on substantial genetic data are rare. Approximately 37% of tigers (Panthera tigris) in the Similipal Tiger Reserve (in eastern India) are pseudomelanistic, characterized by wide, merged stripes. Camera trap data across the tiger range revealed the presence of pseudomelanistic tigers only in Similipal. We investigated the genetic basis for pseudomelanism and examined the role of drift in driving this phenotype's frequency. Whole-genome data and pedigree-based association analyses from captive tigers revealed that pseudomelanism cosegregates with a conserved and functionally important coding alteration in Transmembrane Aminopeptidase Q (Taqpep), a gene responsible for similar traits in other felid species. Noninvasive sampling of tigers revealed a high frequency of the Taqpep p.H454Y mutation in Similipal (12 individuals, allele frequency = 0.58) and absence from all other tiger populations (395 individuals). Population genetic analyses confirmed few (minimal number) tigers in Similipal, and its genetic isolation, with poor geneflow. Pairwise FST (0.33) at the mutation site was high but not an outlier. Similipal tigers had low diversity at 81 single nucleotide polymorphisms (mean heterozygosity = 0.28, SD = 0.27). Simulations were consistent with founding events and drift as possible drivers for the observed stark difference of allele frequency. Our results highlight the role of stochastic processes in the evolution of rare phenotypes. We highlight an unusual evolutionary trajectory in a small and isolated population of an endangered species.

Empowering conservation practice with efficient and economical genotyping from poor quality samples.

Moderate- to high-density genotyping (100 + SNPs) is widely used to determine and measure individual identity, relatedness, fitness, population structure and migration in wild populations.However, these important tools are difficult to apply when high-quality genetic material is unavailable. Most genomic tools are developed for high-quality DNA sources from laboratory or medical settings. As a result, most genetic data from market or field settings is limited to easily amplified mitochondrial DNA or a few microsatellites.To enable genotyping in conservation contexts, we used next-generation sequencing of multiplex PCR products from very low-quality DNA extracted from faeces, hair and cooked samples. We demonstrated utility and wide-ranging potential application in endangered wild tigers and tracking commercial trade in Caribbean queen conch.We genotyped 100 SNPs from degraded tiger samples to identify individuals, discern close relatives and detect population differentiation. Co-occurring carnivores do not amplify (e.g. Indian wild dog/dhole) or are monomorphic (e.g. leopard). Sixty-two SNPs from conch fritters and field-collected samples were used to test relatedness and detect population structure.We provide proof of concept for a rapid, simple, cost-effective and scalable method (for both samples and number of loci), a framework that can be applied to other conservation scenarios previously limited by low-quality DNA samples. These approaches provide a critical advance for wildlife monitoring and forensics, open the door to field-ready testing, and will strengthen the use of science in policy decisions and wildlife trade.

Conservation priorities for endangered Indian tigers through a genomic lens.

Tigers have lost 93% of their historical range worldwide. India plays a vital role in the conservation of tigers since nearly 60% of all wild tigers are currently found here. However, as protected areas are small (<300 km2 on average), with only a few individuals in each, many of them may not be independently viable. It is thus important to identify and conserve genetically connected populations, as well as to maintain connectivity within them. We collected samples from wild tigers (Panthera tigris tigris) across India and used genome-wide SNPs to infer genetic connectivity. We genotyped 10,184 SNPs from 38 individuals across 17 protected areas and identified three genetically distinct clusters (corresponding to northwest, southern and central India). The northwest cluster was isolated with low variation and high relatedness. The geographically large central cluster included tigers from central, northeastern and northern India, and had the highest variation. Most genetic diversity (62%) was shared among clusters, while unique variation was highest in the central cluster (8.5%) and lowest in the northwestern one (2%). We did not detect signatures of differential selection or local adaptation. We highlight that the northwest population requires conservation attention to ensure persistence of these tigers.

A possible novel function of dominance behaviour in queen-less colonies of the primitively eusocial wasp Ropalidia marginata.

Unlike the queens of other primitively eusocial species, Ropalidia marginata queens are strikingly docile and non-aggressive individuals, never at the top of the behavioural dominance hierarchy of their colonies. Nevertheless, these queens are completely successful at suppressing worker reproduction, suggesting that they do not use aggression but employ some other mechanism (e.g. pheromones) to do so. Upon removal of the queen from a colony, a single worker, the 'potential queen', immediately begins to display highly elevated levels of aggression towards her nest mates. This individual becomes the next docile queen if the original queen is not returned. We attempt to understand the function of the temporary and amplified dominance behaviour displayed by the potential queen. We find that the dominance behaviour shown by the potential queen is unrelated to the number of her nest mates, their dominance ranks or ovarian condition. This suggests that aggression may not be used to actively suppress other workers and counter threat. Instead we find evidence that dominance behaviour is required for the potential queen's rapid ovarian development, facilitating her speedy establishment as the sole reproductive individual in the colony.