lthough it is possible that other tiger populations outside of India harboured increased genetic variation in the past, our results show that currently Indian tiger retain the majority of the species' genetic variation. This suggests subspecies-based conservation criteria are inappropriate. Despite having experienced recent demographic declines, extensive habitat loss, extant Indian wild tigers retain 76% of the mitochondrial diversity and 63% of the species' nuclear genetic diversity and are adapted to a greater diversity of habitats ,. They are thus critically important from demographic, evolutionary and ecological perspectives for future survival and recovery of the species. More than a billion people, afflicted by poverty and yet experiencing rapid economic growth live in India. That Indian tigers have managed to retain their genetic diversity in the face of such high anthropogenic pressure provides some hope for species survival in the future.
Materials and Methods
Sampling and DNA extraction
Samples were opportunistically collected from wild individuals living inside protected areas and national parks spanning all over India. We collected 71 fecal samples and two tissue samples from most of the tiger habitat in India. Tissue samples were collected from poached animals with permissions. All samples were collected in sterile vials and preserved in absolute alcohol until processed. To avoid the effects of inbreeding in our analyses, samples were collected spatially far apart within a protected area (at least 15 km apart). DNA extraction and species identification was performed by methods explained in Mukherjee et al. . Sampling information is provided in supplementary material (Table S1). We generated mitochondrial DNA data for 54 samples and STR data for 58 samples. In addition to the samples collected in this study, we used genetic data from all tiger subspecies (Indo-Chinese, Malayan, Sumatran and Siberian, in addition to Indian tigers) from Luo et al. . All genetic data for tigers outside the Indian subcontinent (as presented in Figure 1) are from Luo et al. . We only considered data from tigers of known wild origin. A total of 57 tiger mitochondrial DNA and 68 STR data have been used for comparison in this study.
Marker selection and amplification
We designed tiger-specific mitochondrial DNA primers. Polymorphic regions were ascertained using tiger mitochondrial sequences across all subspecies based on Luo et al.  (Figure S1). These primers were then standardized for fecal DNA samples. A total of nine primer sets were designed, and used to amplify1263 bp from Indian tiger fecal samples. Primers sequences are presented in supplementary information (Table S2).
We selected ten felid-specific microsatellite loci based on PCR success rate, amplicon size, number of alleles and the level of observed heterozygosity (Hobs) in Indian tigers (Table S3). All samples were genotyped at this panel of nine dinucleotide and one tetranucleotide microsatellite locus described initially in the domestic cat .
The mitochondrial regions were amplified in 10 µl volume reactions, cleaned by Exo-Sap mixture (NEB) and sequenced from both ends on an ABI 3100XL capillary sequencer. To monitor possible contamination, PCR blanks were included in all experiments.
Amplification for all the microsatellite loci was done using a multiplex approach.
A modified multiple tube approach, combined with a quality index approval was used for data quality management to account for the varying quality and quantity of DNA obtained from non-invasive sources. The complete genotyping process was repeated three times for all samples, and only those loci with quality index ≥0.75 were included in the analysis .
Genetic diversity statistics, population growth indicators (Fu's F, Tajima's D) and genetic difference (Fst) were calculated using ARLEQUIN 3.1 , assuming two populations of tigers (Indian subspecies and all other subspecies). To avoid the effects of related individuals in our analyses, we used STR data from 39 representative samples from all the areas sampl