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Genomic studies of endangered species have primarily focused on describing diversity patterns and resolving phylogenetic relationships, with the overarching goal of informing conservation efforts. However, few studies have investigated genomic diversity housed in captive populations. For tigers ( Panthera tigris ), captive individuals vastly outnumber those in the wild, but their diversity remains largely unexplored. Privately owned captive tiger populations have remained an enigma in the conservation community, with some believing that these individuals are severely inbred, while others believe they may be a source of now-extinct diversity. Here, we present a large-scale genetic study of the private (non-zoo) captive tiger population in the United States, also known as "Generic" tigers. We find that the Generic tiger population has an admixture fingerprint comprising all six extant wild tiger subspecies. Of the 138 Generic individuals sequenced for the purpose of this study, no individual had ancestry from only one subspecies. We show that the Generic tiger population has a comparable amount of genetic diversity relative to most wild subspecies, few private variants, and fewer deleterious mutations. We observe inbreeding coefficients similar to wild populations, although there are some individuals within both the Generic and wild populations that are substantially inbred. Additionally, we develop a reference panel for tigers that can be used with imputation to accurately distinguish individuals and assign ancestry with ultralow coverage (0.25×) data. By providing a cost-effective alternative to whole-genome sequencing (WGS), the reference panel provides a resource to assist in tiger conservation efforts for both ex- and in situ populations., Competing Interests: Competing interests statement:The authors declare no competing interest.

Objectives: The Bengal tiger Panthera tigris tigris, is an emblematic animal for Bangladesh. Despite being the apex predator in the wild, their number is decreasing due to anthropogenic activities such as hunting, urbanization, expansion of agriculture and deforestation. By contrast, captive tigers are flourishing due to practical conservation efforts. Breeding within the small captive population can produce inbreeding depression and genetic bottlenecks, which may limit the success of conservation efforts. Despite past decades of research, a comprehensive database on genetic variation in the captive and wild Bengal tigers in Bangladesh still needs to be included. Therefore, this research aimed to investigate the White Bengal tiger genome to create a resource for future studies to understand variation underlying important functional traits., Data Description: Blood samples from Chattogram Zoo were collected for three white Bengal tigers. Genomic DNA for all collected samples were extracted using a commercial DNA extraction kit. Whole genome sequencing was performed using a DNBseq platform. We generated 77 Gb of whole-genome sequencing (WGS) data for three white Bengal tigers (Average 11X coverage/sample). The data we generated will establish a paradigm for tiger research in Bangladesh by providing a genomic resource for future functional studies on the Bengal white tiger., (© 2024. The Author(s).)

The Amur tiger is currently confronted with challenges of anthropogenic development, leading to its population becoming fragmented into two geographically isolated groups: smaller and larger ones. Small and isolated populations frequently face a greater extinction risk, yet the small tiger population's genetic status and survival potential have not been assessed. Here, a total of 210 samples of suspected Amur tiger feces were collected from this small population, and the genetic background and population survival potentials were assessed by using 14 microsatellite loci. Our results demonstrated that the mean number of alleles in all loci was 3.7 and expected heterozygosity was 0.6, indicating a comparatively lower level of population genetic diversity compared to previously reported studies on other subspecies. The genetic estimates of effective population size (Ne) and the Ne/N ratio were merely 7.6 and 0.152, respectively, representing lower values in comparison to the Amur tiger population in Sikhote-Alin (the larger group). However, multiple methods have indicated the possibility of genetic divergence within our isolated population under study. Meanwhile, the maximum kinship recorded was 0.441, and the mean inbreeding coefficient stood at 0.0868, both of which are higher than those observed in other endangered species, such as the African lion and the grey wolf. Additionally, we have identified a significant risk of future extinction if the lethal equivalents were to reach 6.26, which is higher than that of other large carnivores. Further, our simulation results indicated that an increase in the number of breeding females would enhance the prospects of this population. In summary, our findings provide a critical theoretical basis for further bailout strategies concerning Amur tigers., (© 2024. The Author(s).)

There is an urgent need to find a way to improve the genetic diversity of captive South China tiger (SCT, Panthera tigris amoyensis ), the most critically endangered taxon of living tigers, facing inbreeding depression. The genomes showed that 13 hybrid SCTs from Meihuashan were divided into two groups; one group included three individuals who had a closer relationship with pureblood SCTs than another group. The three individuals shared more that 40% of their genome with pureblood SCTs and might be potential individuals for genetic rescuing in SCTs. A large-scale genetic survey based on 319 pureblood SCTs showed that the mean microsatellite inbreeding coefficient of pureblood SCTs decreased significantly from 0.1789 to 0.0600 ( p = 0.000009) and the ratio of heterozygous loci increased significantly from 38.5% to 43.2% ( p = 0.02) after one individual of the Chongqing line joined the Suzhou line and began to breed in the mid-1980s, which is a reason why the current SCTs keep a moderate level of microsatellite heterozygosity and nucleotide diversity. However, it is important to establish a back-up population based on the three individuals through introducing one pureblood SCT into the back-up population every year. The back-up population should be an important reserve in case the pureblood SCTs are in danger in the future.

The tiger (Panthera tigris) is a charismatic megafauna species that originated and diversified in Asia and probably experienced population contraction and expansion during the Pleistocene, resulting in low genetic diversity of modern tigers. However, little is known about patterns of genomic diversity in ancient populations. Here we generated whole-genome sequences from ancient or historical (100-10,000 yr old) specimens collected across mainland Asia, including a 10,600-yr-old Russian Far East specimen (RUSA21, 8× coverage) plus six ancient mitogenomes, 14 South China tigers (0.1-12×) and three Caspian tigers (4-8×). Admixture analysis showed that RUSA21 clustered within modern Northeast Asian phylogroups and partially derived from an extinct Late Pleistocene lineage. While some of the 8,000-10,000-yr-old Russian Far East mitogenomes are basal to all tigers, one 2,000-yr-old specimen resembles present Amur tigers. Phylogenomic analyses suggested that the Caspian tiger probably dispersed from an ancestral Northeast Asian population and experienced gene flow from southern Bengal tigers. Lastly, genome-wide monophyly supported the South China tiger as a distinct subspecies, albeit with mitochondrial paraphyly, hence resolving its longstanding taxonomic controversy. The distribution of mitochondrial haplogroups corroborated by biogeographical modelling suggested that Southwest China was a Late Pleistocene refugium for a relic basal lineage. As suitable habitat returned, admixture between divergent lineages of South China tigers took place in Eastern China, promoting the evolution of other northern subspecies. Altogether, our analysis of ancient genomes sheds light on the evolutionary history of tigers and supports the existence of nine modern subspecies., (© 2023. The Author(s), under exclusive licence to Springer Nature Limited.)

Polymorphism of nine microsatellite loci in the Sikhote-Alin tiger population was analyzed in the modern and recent historical periods, using blood, excrement, and museum bone samples. Tests for excess heterozygosity to determine whether the population went through a period of low abundance and a low value of the Garza-Williamson coefficient indicated that such events were highly probable to occur in both recent and earlier history. The mean effective population size Ne of a current sample was 34.4 (95% confidence interval 17-106.8). This fact is of great interest in the contest of conservation and restoration of endangered large cat species., (© 2023. The Author(s).)

Tiger fish (genus Datnioides) are critical ornamental and economic fish and are valuable freshwater fish worldwide, belonging to the order Lobotiformes. Currently, there are five extant species (Datnioides campbelli, D. microlepis, D. polota, D. pulcher, and D. undecimradiatus) of Datnioides in the world, usually inhabiting in south and southeast Asia. Due to the decline of wild population sizes of tiger fish and the lack of molecular research on them, in the present study, we sequenced, assembled, and characterized the complete nuclear ribosomal DNA (nrDNA) operon of all five extant tiger fish species, in order to elucidate the phylogenetic relationship among the genus Datnioides. The nrDNA sequences of five tiger fish species were 8548-9182 bp in length, encompassing complete 18S rDNA, ITS1, 5.8S rDNA, ITS2, 28S rDNA, and IGS regions. Numerous repetitive sequences were detected, substantially influencing the sequence length of different regions in each species. We employed maximum-likelihood (ML) method and Bayesian inference (BI) method to construct phylogenetic trees for Datnioides. Phylogenetic analyses indicated that each region in nrDNA operon is not sufficiently phylogenetically informative to delineate the species in Datnioides; nevertheless, the whole operon is able to delineate five tiger fish species much better, three of five species were successfully partitioned. Particularly, regardless of employed markers, it was strongly supported that D. campbelli was considerably partitioned from the other four species, possibly due to the geographical separation. In spite of the fact that discrimination of Datnioides species requires further investigation, our study provides reference genome resources for the Lobotiformes, as well as insights into the phylogenetic position of Lobotiformes and further biological conservation., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Isolation of wildlife populations represents a key conservation challenge in the twenty-first century. This may necessitate consideration of translocations to ensure population viability. We investigated the potential population and genetic trajectory of a small, isolated tiger (Panthera tigris) population in Thailand's Dong Phayayen-Khao Yai forest complex across a range of scenarios. Using an individual-based, spatially-explicit population modelling approach, we simulate population and genetic trajectories and evaluate the relative impact of translocations from a related population. Population and genetic trajectories in our study were most sensitive to sex and number of individuals translocated and translocation frequency. Translocation of females produced consistently higher population, allelic richness, and heterozygosity compared to equal numbers of males. Despite population increases, declines in allelic richness and heterozygosity across simulations were stark, with simulations predicting a mean decline of allelic richness and heterozygosity of 46.5% and 53.5% without intervention, respectively. Translocations of four females every generation or every other generation were required to prevent substantial heterozygosity declines. While translocations could increase population size, they may fail to prevent long-term loss of genetic diversity in small populations unless applied frequently. This reinforces the importance of incorporating realistic processes of genetic inheritance and gene flow in modelling small populations., (© 2023. The Author(s).)

All species of big cats, including tigers, cheetahs, leopards, lions, snow leopards, and jaguars, are protected under the Convention on the International Trade in Endangered Species (CITES). This is due in large part to population declines resulting from anthropogenic factors, especially poaching and the unregulated and illegal trade in pelts, bones, teeth and other products that are derived from these iconic species. To enhance and scale up monitoring for big cat products in this trade, we created a rapid multiplex qPCR test that can identify and differentiate DNA from tiger (Panthera tigris), cheetah (Acinonyx jubatus), leopard (Panthera pardus), lion (Panthera leo), snow leopard (Panthera uncia), and jaguar (Panthera onca) in wildlife products using melt curve analysis to identify each species by its unique melt peak temperature. Our results showed high PCR efficiency (> 90%), sensitivity (detection limit of 5 copies of DNA per PCR reaction) and specificity (no cross amplification between each of the 6 big cat species). When paired with a rapid (< 1 h) DNA extraction protocol that amplifies DNA from bone, teeth, and preserved skin, total test time is less than three hours. This test can be used as a screening method to improve our understanding of the scale and scope of the illegal trade in big cats and aid in the enforcement of international regulations that govern the trade in wildlife and wildlife products, both ultimately benefiting the conservation of these species worldwide., (© 2023. The Author(s).)

Background: Among six extant tiger subspecies, the South China tiger (Panthera tigris amoyensis) once was widely distributed but is now the rarest one and extinct in the wild. All living South China tigers are descendants of only two male and four female wild-caught tigers and they survive solely in zoos after 60 years of effective conservation efforts. Inbreeding depression and hybridization with other tiger subspecies were believed to have occurred within the small, captive South China tiger population. It is therefore urgently needed to examine the genomic landscape of existing genetic variation among the South China tigers., Results: In this study, we assembled a high-quality chromosome-level genome using long-read sequences and re-sequenced 29 high-depth genomes of the South China tigers. By combining and comparing our data with the other 40 genomes of six tiger subspecies, we identified two significantly differentiated genomic lineages among the South China tigers, which harbored some rare genetic variants introgressed from other tiger subspecies and thus maintained a moderate genetic diversity. We noticed that the South China tiger had higher F ROH values for longer runs of homozygosity (ROH > 1 Mb), an indication of recent inbreeding/founder events. We also observed that the South China tiger had the least frequent homozygous genotypes of both high- and moderate-impact deleterious mutations, and lower mutation loads than both Amur and Sumatran tigers. Altogether, our analyses indicated an effective genetic purging of deleterious mutations in homozygous states from the South China tiger, following its population contraction with a controlled increase in inbreeding based on its pedigree records., Conclusions: The identification of two unique founder/genomic lineages coupled with active genetic purging of deleterious mutations in homozygous states and the genomic resources generated in our study pave the way for a genomics-informed conservation, following the real-time monitoring and rational exchange of reproductive South China tigers among zoos., (© 2023. The Author(s).)

The South China tiger (Panthera tigris amoyensis, SCT) is the most critically endangered subspecies of tiger due to functional extinction in the wild. Inbreeding depression is observed among the captive population descended from six wild ancestors, resulting in high juvenile mortality and low reproduction. We assembled and characterized the first SCT genome and an improved Amur tiger (P. t. altaica, AT) genome named AmyTig1.0 and PanTig2.0. The two genomes are the most continuous and comprehensive among any tiger genomes yet reported at the chromosomal level. By using the two genomes and resequencing data of 15 SCT and 13 AT individuals, we investigated the genomic signature of inbreeding depression of the SCT. The results indicated that the effective population size of SCT experienced three phases of decline, ~5.0-1.0 thousand years ago, 100 years ago, and since captive breeding in 1963. We found 43 long runs of homozygosity fragments that were shared by all individuals in the SCT population and covered a total length of 20.63% in the SCT genome. We also detected a large proportion of identical-by-descent segments across the genome in the SCT population, especially on ChrB4. Deleterious nonsynonymous single nucleotide polymorphic sites and loss-of-function mutations were found across genomes with extensive potential influences, despite a proportion of these loads having been purged by inbreeding depression. Our research provides an invaluable resource for the formulation of genetic management policies for the South China tiger such as developing genome-based breeding and genetic rescue strategy., (© 2022 The Authors. Molecular Ecology Resources published by John Wiley & Sons Ltd.)

Tigers are endangered apex predators. They typify endangered species because they are elusive, rare, and face numerous threats across their range. Tigers also symbolize conservation. However, it is very difficult to study tigers because of their stated nature. Also, tiger conservation is a geopolitically sensitive topic, making it difficult to use the studies to propose evidence-based management that allows their recovery, especially in the context of conservation genetics. Zhang et al. (Mol. Ecol. Resour., 2022) have created very valuable and rare resources to aid the community in conserving tigers. First, they present chromosome level genome assemblies of a South China tiger and an Amur tiger. Second, they present whole genome sequences of 16 captive South China tigers. Additionally, by using the assemblies they model the demographic history of these populations, estimate inbreeding and the potential threats they face in captivity. This approach is particularly important because genetic management is now the only remaining option for South China tigers, because they are extinct in the wild. In other words, captive individuals are our only hope for some day restoring the wild populations of South China tigers., (© 2022 The Author. Molecular Ecology Resources published by John Wiley & Sons Ltd.)

The tiger, a poster child for conservation, remains an endangered apex predator. Continued survival and recovery will require a comprehensive understanding of genetic diversity and the use of such information for population management. A high-quality tiger genome assembly will be an important tool for conservation genetics, especially for the Indian tiger, the most abundant subspecies in the wild. Here, we present high-quality near-chromosomal genome assemblies of a female and a male wild Indian tiger (Panthera tigris tigris). Our assemblies had a scaffold N50 of >140 Mb, with 19  scaffolds corresponding to the 19 numbered chromosomes, containing 95% of the genome. Our assemblies also enabled detection of longer stretches of runs of homozygosity compared to previous assemblies, which will help improve estimates of genomic inbreeding. Comprehensive genome annotation identified 26,068 protein-coding genes, including several gene families involved in key morphological features such as the teeth, claws, vision, olfaction, taste, and body stripes. We also identified 301 microRNAs, 365 small nucleolar RNAs, 632 transfer RNAs, and other noncoding RNA elements, several of which are predicted to regulate key biological pathways that likely contribute to the tiger's apex predatory traits. We identify signatures of positive selection in the tiger genome that are consistent with the Panthera lineage. Our high-quality genome will enable use of noninvasive samples for comprehensive assessment of genetic diversity, thus supporting effective conservation and management of wild tiger populations., (© The Author(s) 2022. Published by Oxford University Press GigaScience.)

The Amur or Siberian tiger Panthera tigris altaica (Temminck, 1844) is currently restricted to a small region of its original geographical range in northwestern Asia and is considered 'endangered' by the IUCN Red List of Threatened Species. This solitary, territorial, and large top predator is in major need of genomic resources to inform conservation management strategies. This study formally tested if complete mitochondrial genomes of P. tigris altaica can be assembled from non-enriched metagenomic libraries generated from scat eDNA samples using the Illumina sequencing platform and open-access bioinformatics pipelines. The mitogenome of P. tigris altaica was assembled and circularized using the pipeline GetOrganelle with a coverage ranging from 322.7x to 17.6x in four different scat eDNA samples. A nearly complete mitochondrial genome (101x) was retrieved from a fifth scat eDNA sample. The complete or nearly complete mitochondrial genomes of P. tigris altaica were AT-rich and composed of 13 protein coding genes (PCGs), 22 transfer RNA genes, two ribosomal RNA genes, and a putative control region. Synteny observed in all assembled mitogenomes was identical to that reported before for P. tigris altaica and other felids. A phylogenomic analysis based on all PCGs demonstrated that the mitochondrial genomes assembled from scat eDNA reliably identify the sequenced samples as belonging to P. tigris and distinguished the same samples from closely and distantly related congeneric species. This study demonstrates that it is viable to retrieve accurate whole and nearly complete mitochondrial genomes of P. tigris altaica (and probably other felids) from scat eDNA samples without library enrichment protocols and using open-access bioinformatics workflows. This new genomic resource represents a new tool to support conservation strategies (bio-prospecting and bio-monitoring) in this iconic cat., Competing Interests: The author declares that they have no competing interests., (© 2022 Baeza.)

Diarrhea-predominant irritable bowel syndrome (IBS-D) is common among the captive South China tigers in zoos. 16S rRNA gene sequencing was performed to demonstrate the compositions and structures of the gastrointestinal microbiota of this species with IBS-D. Their healthy (F1) and mushy (F2) feces were allocated into two groups. A total of 21 and 31 fecal bacterial communities of major phyla and genera were detected, respectively. The F1 and F2 groups had five common microbiotas at the phylum level (Firmicutes, Proteobacteria, Fusobacteria, Actinobacteria, and Bacteroidetes). Among the five phyla, the abundance of Bacteroidetes in the F2 group was significantly lower than that in the F1 group. The diversity level of fecal microbiota within the mild-diarrhea stool was also significantly lower than that of the healthy counterpart. Thirty-two metabolites were correlated to four genus-level bacteria ( Bacteroides, Pseudoclavibacter, Streptococcus , and Ruminococcaceae-UCG-005 ). Due to its normal role in protein degradation and metabolism, we hypothesized that the lower abundance of Bacteroides within the F2 group could be associated with the IBS-D symptoms. Therefore, this work implied that ameliorating the daily diet with a supplement of probiotics, such as Bacteroides , could improve the gut health of this species.

The big cats (genus Panthera) represent some of the most popular and charismatic species on the planet. Although some reference genomes are available for this clade, few are at the chromosome level, inhibiting high-resolution genomic studies. We assembled genomes from 3 members of the genus, the tiger (Panthera tigris), the snow leopard (Panthera uncia), and the African leopard (Panthera pardus pardus), at chromosome or near-chromosome level. We used a combination of short- and long-read technologies, as well as proximity ligation data from Hi-C technology, to achieve high continuity and contiguity for each individual. We hope that these genomes will aid in further evolutionary and conservation research of this iconic group of mammals., (© The Author(s) 2022. Published by Oxford University Press on behalf of Genetics Society of America.)

A three-year-old male South China tiger died in the tiger enclosure of the China Tiger Park in the Meihua Mountains on December 2018 after being bitten by a tick. This tiger presented clinical symptoms like whole-body severe jaundice, hepatosplenomegaly, kidney, and lymph node hemorrhages. The Colpodella sp.-specific 18S rRNA gene was detected using nested PCR. Interestingly, the DNA isolated from the blood of the tiger was found to be 100% similar to that of the tick by NCBI BLAST analysis. However, the DNA fragments isolated from the tiger's blood were 90.1% similar to the Colpodella sp. strain human erythrocyte parasite (HEP, MH208621) and 90.4% similar to the Colpodella sp. strain Heilongjiang (HLJ, KT364261). To investigate the species of ticks and ticks-carried Colpodella parasites in this region, the species of ticks obtained from the grasses outside the tiger enclosure and the species of Colpodella carried by ticks were identified. The DNA from ticks as well as that from the tick-borne Colpodella sp. were amplified from each tick using PCR followed by amplicon sequencing. In total 402 adult ticks samples were collected, among which 22 were positive for Colpodella sp. (5.5%), and the species were further determined by morphology, DNA sequencing and phylogenetic analyses. Interestingly, one Colpodella sp. was found to have 94.2% sequence similarities to the Colpodella sp. strain HEP (MH208621). This strain was previously reported to infect a woman in Yunnan, China. In addition, three Colpodella sp. showed 87-91% sequence similarities to the Colpodella sp. strain HLJ (KT364261), which was previously reported to infect human in Heilongjiang, China. This study disclosed the possibility of zoonotic transmission of Colpodella sp. by ticks in China. Finally, it provides a basis for urgently determining and monitoring the repertoire of ticks-borne piroplasmid pathogens, with the ultimate aim of strategic control.

Tigers ( Panthera tigris ) are flagship big cats and attract extensive public attention due to their charismatic features and endangered status. Despite this, little is known about their prehistoric lineages and detailed evolutionary histories. Through palaeogenomic analyses, we identified a Pleistocene tiger from northeastern China, dated to beyond the limits of radiocarbon dating (greater than 43 500 years ago). We used a simulated dataset and different reads processing pipelines to test the validity of our results and confirmed that, in both mitochondrial and nuclear phylogenies, this ancient individual belongs to a previously unknown lineage that diverged prior to modern tiger diversification. Based on the mitochondrial genome, the divergence time of this ancient lineage was estimated to be approximately 268 ka (95% CI: 187-353 ka), doubling the known age of tigers' maternal ancestor to around 125 ka (95% CI: 88-168 ka). Furthermore, by combining our findings with putative mechanisms underlying the discordant mito-nuclear phylogenetic placement for the South China tigers, we proposed a more complex scenario of tiger evolution that would otherwise be missed using data from modern tigers only. Our study provides the first glimpses of the genetic antiquity of tigers and demonstrates the utility of aDNA-based investigation for further understanding tiger evolution.

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'.

Background: Genetics driven interventions if adopted in conservation breeding projects may enhance the overall success by prioritizing breeding among genetically most competent individuals and delaying or completely diminishing the ill effects of inbreeding., Methods and Results: In the present study, we investigated genetic make-up of 15 tigers housed at five different captive facilities of West Bengal in India and report the moderate level of genetic variation. We identified five tigers based on individual genetic attributes that may be prioritized for future breeding or animal exchange programmes. The occurrence of first and second order related individuals in captivity require management attention and they should be paired considering their immediate genetic background., Conclusion: Considering tiger as a case study, we highlight the use of genetic assessment and necessity to validate the studbook records in formulating adaptive management strategies for long-term conservation and management of species of interest., (© 2022. The Author(s), under exclusive licence to Springer Nature B.V.)

We report an outbreak of severe acute respiratory syndrome coronavirus 2 involving 3 Malayan tigers (Panthera tigris jacksoni) at a zoo in Tennessee, USA. Investigation identified naturally occurring tiger-to-tiger transmission; genetic sequence change occurred with viral passage. We provide epidemiologic, environmental, and genomic sequencing data for animal and human infections.

Identification of genetic structure within wildlife populations have implications in their conservation and management. Accurately inferring population genetic structure requires whole-genome data across the geographical range of the species, which can be resource-intensive. A cheaper strategy is to employ a subset of markers that can efficiently recapitulate the population genetic structure inferred by the whole genome data. Such ancestry informative markers (AIMs), have rarely been developed for endangered species such as tigers utilizing single nucleotide polymorphisms (SNPs). Here, we first identify the population structure of the Indian tiger using whole-genome sequences and then develop an AIMs panel with a minimum number of SNPs that can recapitulate this structure. We identified four population clusters of Indian tigers with North-East, North-West, and South Indian tigers forming three separate groups, and Terai and Central Indian tigers forming a single cluster. To evaluate the robustness of our AIMs, we applied it to a separate dataset of tigers from across India. Out of 92 SNPs present in our AIMs panel, 49 were present in the new dataset. These 49 SNPs were sufficient to recapitulate the population genetic structure obtained from the whole genome data. To the best of our knowledge, this is the first-ever SNP-based AIMs panel for big cats, which can be used as a cost-effective alternative to whole-genome sequencing for detecting the biogeographical origin of Indian tigers. Our study can be used as a guideline for developing an AIMs panel for the management of other endangered species where obtaining whole genome sequences are difficult., (© 2021. The Author(s), under exclusive licence to The Genetics Society.)

Postnatal colonization and development of the gut microbiota is linked to health and growth. A comprehensive understanding of the postnatal compositional changes and development of the microbial community is helpful to understand the gut health and improve the survival rate of South China tiger cubs (Panthera tigris amoyensis). Fecal samples from three tiger cubs were collected on the day of birth in 2018 (June 17-21 [G0], July 18 [G1], July 31 [G2], and August 7 [G3]). The 16S rRNA genes of the fecal microflora were sequenced. Results showed that 38 phyla, 58 classes, 134 orders, 272 families, and 636 genera of bacteria from 3,059 operational taxonomic units were identified from 12 fecal samples. The diversity and abundance of species of group G0 were significantly higher (p < 0.05 or 0.01) than those of groups G2 and G3. The predominant phylum was Proteobacteria in groups G0 and G1 (38.85% and 48%, respectively) and Firmicutes in groups G2 and G3 (71.42% and 75.29%, respectively). At the phylum level, the abundance of Deinococcus-Thermus was significantly decreased in groups G1, G2, and G3 as compared to group G0 (p < 0.05), while that of Firmicutes was significantly increased in groups G2 and G3 (p < 0.05). At the genus level, the abundance of Faecalibacterium, Ralstonia, and unidentified Rickettsiales was significantly decreased in groups G1, G2, and G3 as compared with group G0 (p < 0.05), while that of Pseudomonas was significantly decreased in groups G2 and G3 (p < 0.05). The composition and structure of fecal microbiota of South China tiger cubs changed after birth., (© 2021 The Author(s) Published by S. Karger AG, Basel.)

Background: Immune cells play important roles in mediating immune response and host defense against invading pathogens. However, insights into the molecular mechanisms governing circulating immune cell diversity among multiple species are limited., Methods: In this study, we compared the single-cell transcriptomes of immune cells from 12 species. Distinct molecular profiles were characterized for different immune cell types, including T cells, B cells, natural killer cells, monocytes, and dendritic cells., Results: Our data revealed the heterogeneity and compositions of circulating immune cells among 12 different species. Additionally, we explored the conserved and divergent cellular crosstalks and genetic regulatory networks among vertebrate immune cells. Notably, the ligand and receptor pair VIM-CD44 was highly conserved among the immune cells., Conclusions: This study is the first to provide a comprehensive analysis of the cross-species single-cell transcriptome atlas for peripheral blood mononuclear cells (PBMCs). This research should advance our understanding of the cellular taxonomy and fundamental functions of PBMCs, with important implications in evolutionary biology, developmental biology, and immune system disorders., (© 2022 The Authors. Clinical and Translational Medicine published by John Wiley & Sons Australia, Ltd on behalf of Shanghai Institute of Clinical Bioinformatics.)

Increasing habitat fragmentation leads to wild populations becoming small, isolated, and threatened by inbreeding depression. However, small populations may be able to purge recessive deleterious alleles as they become expressed in homozygotes, thus reducing inbreeding depression and increasing population viability. We used whole-genome sequences from 57 tigers to estimate individual inbreeding and mutation load in a small-isolated and two large-connected populations in India. As expected, the small-isolated population had substantially higher average genomic inbreeding ( F ROH = 0.57) than the large-connected ( F ROH = 0.35 and F ROH = 0.46) populations. The small-isolated population had the lowest loss-of-function mutation load, likely due to purging of highly deleterious recessive mutations. The large populations had lower missense mutation loads than the small-isolated population, but were not identical, possibly due to different demographic histories. While the number of the loss-of-function alleles in the small-isolated population was lower, these alleles were at higher frequencies and homozygosity than in the large populations. Together, our data and analyses provide evidence of 1) high mutation load, 2) purging, and 3) the highest predicted inbreeding depression, despite purging, in the small-isolated population. Frequency distributions of damaging and neutral alleles uncover genomic evidence that purifying selection has removed part of the mutation load across Indian tiger populations. These results provide genomic evidence for purifying selection in both small and large populations, but also suggest that the remaining deleterious alleles may have inbreeding-associated fitness costs. We suggest that genetic rescue from sources selected based on genome-wide differentiation could offset any possible impacts of inbreeding depression., Competing Interests: The authors declare no competing interest., (Copyright © 2021 the Author(s). Published by PNAS.)

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 F ST (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., Competing Interests: The authors declare no competing interest., (Copyright © 2021 the Author(s). Published by PNAS.)

Deforestation and agricultural intensification have resulted in an alarming change in the global land cover over the past 300 years, posing a threat to species conservation. Dhole is a monophyletic, social canid and, being an endangered and highly forest-dependent species, is more prone to the loss of favorable habitat in the Anthropocene. We determined the genetic differentiation and demographic history of dhole across the tiger reserves of Maharashtra using the microsatellite data of 305 individuals. Simulation-based analyses revealed a 77-85% decline in the major dhole sub-populations. Protected areas have provided refuge to the historically declining dhole population resulting in clustering with strong genetic structure in the remnant dhole population. The historical population decline coincides with the extreme events in the landscape over the past 300 years. The study highlights the pattern of genetic differentiation and diversity of a highly forest-dependent species which can be associated with the loss of forest cover outside tiger reserves. It also warrants attention to develop conservation plans for the remnant surviving population of dholes in India., (© 2021. The Author(s).)

Background: Currently, Tigers (the top predator of an ecosystem) are on the list of endangered species. Thus the need is to understand the tiger's population genomics to design their conservation strategies., Objective: We analyzed the molecular evolution of tiger diversity using NADH dehydrogenase subunit 4 (ND4), a significant electron transport chain component., Methods: We have analyzed nucleotide composition and distribution pattern of ND genes, molecular evolution, evolutionary conservation pattern and conserved blocks of NADH, phylogenomics of ND4, and estimating species divergence, etc., using different bioinformatics tools and software, and MATLAB programming and computing environment., Results: The nucleotide composition and distribution pattern of ND genes in the tiger genome demonstrated an increase in the number of adenine (A) and a lower trend of A+T content in some place of the distribution analysis. However, the observed distributions were not significant (P > 0.05). Evolutionary conservation analysis showed three highly align blocks (186 to 198, 406 to 416, and 527 to 545). On mapping the molecular evolution of ND4 among model species (n = 30), we observed its presence in a broader range of species. ND4 based molecular evolution of tiger diversity and time divergence for a tiger (20 different other species) shows that genus Panthera originated more or less at a similar time., Conclusions: The nucleotide composition and nucleotide distribution pattern of tiger ND genes showed the evolutionary pattern and origin of tiger and Panthera lineage concerning the molecular clock, which will help to understand their adaptive evolution.

A fifth of all known species are currently classified as threatened in the wild: the rate of biodiversity loss is rapid, continuous, and mostly due to anthropogenic activities. To slow down this decline, the accurate estimation of demographic parameters for threatened species is critical. With this aim, zoo institutions play an important role, giving access to data on zoo-housed animals, which aids researchers working on species life-history traits and intrinsic factors influencing the fitness of both sexes, such as age. While tigers (Panthera tigris) are particularly threatened in their natural environment, few of their demographic parameters have been determined because of their solitary and elusive nature as well as low population density. Using individual-based information for more than 9200 tigers (from 1938 to 2018) recorded in the International Tiger Studbook 2018, we aimed to determine sub-species and sex-specific variability of survival and reproductive parameters with age. No significant sex-difference in actuarial senescence (i.e., decline of survival probabilities with age) was observed but males tended to have a higher juvenile mortality and a faster senescence than females. Reproductive senescence (i.e., decline of reproductive parameters with age) was more pronounced in females than males. Moreover, we observed sub-species-specific variation in mortality and reproductive patterns, pointing out the necessity to consider them independently for conservation goals. Our findings can provide meaningful improvements to the husbandry of zoo-housed tigers, emphasizing the importance of adult breeding females of 7-9 years-old to control zoo-housed population size, but also providing accurate demographic estimates, crucial to set up effective conservation plans., (© 2021 Wiley Periodicals LLC.)

Species conservation can be improved by knowledge of evolutionary and genetic history. Tigers are among the most charismatic of endangered species and garner significant conservation attention. However, their evolutionary history and genomic variation remain poorly known, especially for Indian tigers. With 70% of the world's wild tigers living in India, such knowledge is critical. We re-sequenced 65 individual tiger genomes representing most extant subspecies with a specific focus on tigers from India. As suggested by earlier studies, we found strong genetic differentiation between the putative tiger subspecies. Despite high total genomic diversity in India, individual tigers host longer runs of homozygosity, potentially suggesting recent inbreeding or founding events, possibly due to small and fragmented protected areas. We suggest the impacts of ongoing connectivity loss on inbreeding and persistence of Indian tigers be closely monitored. Surprisingly, demographic models suggest recent divergence (within the last 20,000 years) between subspecies and strong population bottlenecks. Amur tiger genomes revealed the strongest signals of selection related to metabolic adaptation to cold, whereas Sumatran tigers show evidence of weak selection for genes involved in body size regulation. We recommend detailed investigation of local adaptation in Amur and Sumatran tigers prior to initiating genetic rescue., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

Accurate species and sex identification of non-invasive and forensic samples of the tiger and leopard is still confusing when using the allele-specific methods. We designed allele-specific methods with penultimate nucleotide mismatch in a nested manner for the exact identification and double-checking of forensic samples. The mismatch design is a novel concept in species and sex identification, making the allele-specific targeting precise. We developed three sets of markers, a 365 bp outer and a 98 bp inner marker for nested tiger species identification assay, 136 bp leopard specific marker, and carnivore sex identification markers. We validated the method with tissue/blood forensic samples of various felids and herbivorous available in our lab and on known fecal samples from Vandalur Zoo. We also collected 37 scat samples at diverse stages of deterioration from the Mudumalai Tiger Reserve, Tamil Nadu, India. The 365 bp targeted markers resulted in 70.2% (n = 22; 22/37) amplification success, while the 98 bp FAM-labelled marker amplified 89% (n = 33; 33/37) scat samples independently. The 136 bp leopard markers answered four scat samples (11%) unrequited by the tiger specific markers. We evaluated species and the sex identification with these markers in another 190 non-invasive samples provided by the Mudumalai Tiger Reserve authorities. Among which 56.3% (n = 107) of samples were recognized as tiger (64 male and 43 female) and 38.9% (n = 74) as leopard (41 male and 33 female). The method supersedes any other previous methods in this regard by its high accuracy and simplicity.

The zoos manage small populations of endangered big cat species like tiger, lion, and leopard for display, research, and conservation breeding. Genetic management of these populations is essential to ensure long term survival and conservation utility. Here we propose a simple and cost effective microsatellite based protocol for the genetic management of captive big cats. We sampled 36 big cat individuals from Seoul Grand Park Zoo (Republic of Korea) and amplified 33 published microsatellite loci. Overall, allelic richness and gene diversity was found highest for leopards, followed by lions and tigers. Twelve of the thirty-three markers showed a high degree of polymorphism across all target species. These microsatellites provide a high degree of discrimination for tiger (1.45 × 10 -8 ), lion (1.54 × 10 -10 ), and leopard (1.88 × 10 -12 ) and thus can be adopted for the genetic characterization of big cats in accredited zoos globally. During captive breeding, zoo authorities rely on pedigree records maintained in studbooks to ensure mating of genetically fit unrelated individuals. Several studies have reported errors in studbook records of big cat species. Microsatellites are simple and cost effective tool for DNA fingerprinting, estimation of genetic diversity, and paternity assessment. Our unified microsatellite panel (12-plex) for big cats is efficient and can easily be adopted by zoo authorities for regular population management.

Antagonistic coevolution between host and parasite drives species evolution. However, most of the studies only focus on parasitism adaptation and do not explore the coevolution mechanisms from the perspective of both host and parasite. Here, through the de novo sequencing and assembly of the genomes of giant panda roundworm, red panda roundworm, and lion roundworm parasitic on tiger, we investigated the genomic mechanisms of coevolution between nonmodel mammals and their parasitic roundworms and those of roundworm parasitism in general. The genome-wide phylogeny revealed that these parasitic roundworms have not phylogenetically coevolved with their hosts. The CTSZ and prolyl 4-hydroxylase subunit beta (P4HB) immunoregulatory proteins played a central role in protein interaction between mammals and parasitic roundworms. The gene tree comparison identified that seven pairs of interactive proteins had consistent phylogenetic topology, suggesting their coevolution during host-parasite interaction. These coevolutionary proteins were particularly relevant to immune response. In addition, we found that the roundworms of both pandas exhibited higher proportions of metallopeptidase genes, and some positively selected genes were highly related to their larvae's fast development. Our findings provide novel insights into the genetic mechanisms of coevolution between nonmodel mammals and parasites and offer the valuable genomic resources for scientific ascariasis prevention in both pandas., (© The Author(s) 2020. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

The availability of completed and draft genome assemblies of tiger, leopard, and other felids provides an opportunity to gain comparative insights on their unique evolutionary adaptations. However, genome-wide comparative analyses are susceptible to errors in genome sequences and thus require accurate genome assemblies for reliable evolutionary insights. In this study, while analyzing the tiger genome, we found almost one million erroneous substitutions in the coding and non-coding region of the genome affecting 4,472 genes, hence, biasing the current understanding of tiger evolution. Moreover, these errors produced several misleading observations in previous studies. Thus, to gain insights into the tiger evolution, we corrected the erroneous bases in the genome assembly and gene set of tiger using 'SeqBug' approach developed in this study. We sequenced the first Bengal tiger genome and transcriptome from India to validate these corrections. A comprehensive evolutionary analysis was performed using 10,920 orthologs from nine mammalian species including the corrected gene sets of tiger and leopard and using five different methods at three hierarchical levels, i.e. felids, Panthera, and tiger. The unique genetic changes in tiger revealed that the genes showing signatures of adaptation in tiger were enriched in development and neuronal functioning. Specifically, the genes belonging to the Notch signalling pathway, which is among the most conserved pathways involved in embryonic and neuronal development, were found to have significantly diverged in tiger in comparison to the other mammals. Our findings suggest the role of adaptive evolution in neuronal functions and development processes, which correlates well with the presence of exceptional traits such as sensory perception, strong neuro-muscular coordination, and hypercarnivorous behaviour in tiger.

The South China tiger (Panthera tigris amoyensis) is endemic to China and also the most critically endangered subspecies of living tigers. It is considered extinct in the wild and only about 150 individuals survive in captivity to date, whose genetic heritage, however, is ambiguous and controversial. Here, we conducted an explicit genetic assessment of 92 studbook-registered South China tigers from 14 captive facilities using a subspecies-diagnostic system in the context of comparison with other voucher specimens to evaluate the genetic ancestry and level of distinctiveness of the last surviving P. t. amoyensis. Three mtDNA haplotypes were identified from South China tigers sampled in this study, including a unique P. t. amoyensis AMO1 haplotype not found in other subspecies, a COR1 haplotype that is widespread in Indochinese tigers (P. t. corbetti), and an ALT haplotype that is characteristic of Amur tigers (P. t. altaica). Bayesian STRUCTURE analysis and parentage verification confirmed the verified subspecies ancestry (VSA) as the South China tiger in 74 individuals. Genetic introgression from other tigers was detected in 18 tigers, and subsequent exclusion of these and their offspring from the breeding program is recommended. Both STRUCTURE clustering and microsatellite-based phylogenetic analyses demonstrated a close genetic association of the VSA South China tigers to Indochinese tigers, an issue that could only be elucidated by analysis of historical South China tiger specimens with wild origin. Our results also indicated a moderate level of genetic diversity in the captive South China tiger population, suggesting a potential for genetic restoration., (© The American Genetic Association 2019. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Natural populations display a variety of spatial arrangements, each potentially with a distinctive impact on genetic diversity and genetic differentiation among subpopulations. Although the spatial arrangement of populations can lead to intricate migration networks, theoretical developments have focused mainly on a small subset of such networks, emphasizing the island-migration and stepping-stone models. In this study, we investigate all small network motifs: the set of all possible migration networks among populations subdivided into at most four subpopulations. For each motif, we use coalescent theory to derive expectations for three quantities that describe genetic variation: nucleotide diversity, FST, and half-time to equilibrium diversity. We describe the impact of network properties on these quantities, finding that motifs with a high mean node degree have the largest nucleotide diversity and the longest time to equilibrium, whereas motifs with low density have the largest FST. In addition, we show that the motifs whose pattern of variation is most strongly influenced by loss of a connection or a subpopulation are those that can be split easily into disconnected components. We illustrate our results using two example data sets-sky island birds of genus Sholicola and Indian tigers-identifying disturbance scenarios that produce the greatest reduction in genetic diversity; for tigers, we also compare the benefits of two assisted gene flow scenarios. Our results have consequences for understanding the effect of geography on genetic diversity, and they can assist in designing strategies to alter population migration networks toward maximizing genetic variation in the context of conservation of endangered species., (© The Author(s) 2019. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.)

In recent years, the Kiss1 gene has been reported in a number of vertebrate species, and a substantial dataset has been acquired to demonstrate the critical role of kisspeptins in the reproductive system; yet limited information is available for carnivores. In the present study, we identified and characterized feline Kiss1 by isolating and cloning its full-length cDNA in the domestic cat hypothalamus and caracal testis, using the method of rapid amplification of cDNA ends. Additionally, we isolated and cloned the 3' end of Kiss1 cDNA, containing kisspeptin-10 (Kp10), from the ovaries of a clouded leopard and Siberian tiger. Nucleotide sequencing revealed that domestic cat Kiss1 cDNA is of 711 base pairs and caracal Kiss1 cDNA is of 792 base pairs, both having an open reading frame of 450 base pairs, encoding a precursor protein Kiss1 of 149 amino acids. The core sequence of the feline kisspeptin Kp10 was found to be identical in all species analyzed here and is highly conserved in other vertebrate species. Using an anti-Kp10 antibody, we found the immunoreactive kisspeptin to be localized in the periventricular and infundibular nuclei of the cat hypothalamus. The results show that kisspeptin is highly conserved among different feline families, and its immunoreactive distribution in the hypothalamus may indicate its physiological function in the domestic cat.

Of all the big cats, or perhaps of all the endangered wildlife, the tiger may be both the most charismatic and most well-recognized flagship species in the world. The rapidly changing field of molecular genetics, particularly advances in genome sequencing technologies, has provided new tools to reconstruct what characterizes a tiger. Here we review how applications of molecular genomic tools have been used to depict the tiger's ancestral roots, phylogenetic hierarchy, demographic history, morphological diversity, and genetic patterns of diversification on both temporal and geographical scales. Tiger conservation, stabilization, and management are important areas that benefit from use of these genome resources for developing survival strategies for this charismatic megafauna both in situ and ex situ.

No other species attracts more international resources, public attention, and protracted controversies over its intraspecific taxonomy than the tiger (Panthera tigris) [1, 2]. Today, fewer than 4,000 free-ranging tigers survive, covering only 7% of their historical range, and debates persist over whether they comprise six, five, or two subspecies [3-6]. The lack of consensus over the number of tiger subspecies has partially hindered the global effort to recover the species from the brink of extinction, as both captive breeding and landscape intervention of wild populations increasingly require an explicit delineation of the conservation management units [7]. The recent coalescence to a late Pleistocene bottleneck (circa 110 kya) [5, 8, 9] poses challenges for detecting tiger subspecific morphological traits, suggesting that elucidating intraspecific evolution in the tiger requires analyses at the genomic scale. Here, we present whole-genome sequencing analyses from 32 voucher specimens that resolve six statistically robust monophyletic clades corresponding to extant subspecies, including the recently recognized Malayan tiger (P. tigris jacksoni). The intersubspecies gene flow is very low, corroborating the recognized phylogeographic units. We identified multiple genomic regions that are candidates for identifying the adaptive divergence of subspecies. The body-size-related gene ADH7 appears to have been strongly selected in the Sumatran tiger, perhaps in association with adaptation to the tropical Sunda Islands. The identified genomic signatures provide a solid basis for recognizing appropriate conservation management units in the tiger and can benefit global conservation strategic planning for this charismatic megafauna icon., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Tiger (Panthera tigris) populations are in danger across their entire range due to habitat loss, poaching and the demand for tiger parts. The Bengal tiger (Panthera tigris tigris) is an endangered apex predator with a population size estimated to be less than 200 in Nepal. In spite of strict wildlife protection laws, illegal trade of tiger parts is increasing; and Nepal has become one of the major sources and transit routes for poached wildlife parts. Identification of wildlife parts is often challenging for law enforcement officials due to inadequate training and lack of available tools. Here, we describe a molecular forensic approach to gain insight into illegally trafficked tiger parts seized across Nepal. We created Nepal's first comprehensive reference genetic database of wild tigers through the Nepal Tiger Genome Project (2011-2013). This database has nuclear DNA microsatellite genotype and sex profiles, including geo-spatial information, of over 60% (n = 120) of the wild tigers of Nepal. We analyzed 15 putative cases of confiscated poached tiger parts and all were confirmed to be of tiger. Ten samples were identified as male and five were female. We determined probable geo-source location for 9 of the 14 samples with 6-8 nuclear DNA microsatellite loci using inferences from four different statistical assignment methods. Six samples were assigned to Bardia National Park and one of these was an exact match to a female tiger previously profiled in our fecal DNA reference database. Two tiger samples were assigned to Shuklaphanta Wildlife Reserve and one to Chitwan National Park. We are unable to definitively assign five tiger samples which could be offspring dispersers or might have come from tiger population outside of Nepal. Our study revealed that the western region, particularly Bardia National Park, is a poaching hotspot for illegal tiger trade in Nepal. We present feasibility of using molecular forensic based evidence to incriminate criminals in a court of law in the fight against wildlife crime., Competing Interests: The authors have declared that no competing interests exist.

Background: The Amur tiger is one of the most endangered species in the world, and the healthy population of captive Amur tigers assists the recovery of the wild population. Gut microbes have been shown to be important for human disease and health, but little research exists regarding the microbiome of Amur tigers in captivity., Methods: In this study, we used an integrated approach of 16S rRNA gene sequencing combined with ultra-high-performance liquid chromatography-mass spectrometry (UHPLC-MS)-based metabolomics to analyze the effects of Fenbendazole and Ivermectin Tablets on the gut microbiota and fecal metabolic phenotype of the Amur tiger., Results: The relative abundances of the bacterial genera Collinsella, Clostridium XI and Megamonas were decreased, whereas those of Escherichia and Clostridium sensu stricto were increased in experimental Amur tigers compared with those in normal controls. Meanwhile, distinct changes in the fecal metabolic phenotype of the experimental Amur tigers were also found, including lower levels of acrylic acid, acetoacetate and catechol and higher amounts of 5,6-dihydrouracil, adenine hydrochloride hydrate and galactitol. Moreover, the differentially abundant gut microbes were substantially associated with the altered fecal metabolites, especially the bacteria in the Firmicutes and Actinomycetes, which were involved in the metabolism of 5,6-dihydrouracil, 6-phospho-d-gluconate and 1-methylnicotinamide., Conclusion: Our results indicate for the first time that Fenbendazole and Ivermectin Tablets not only disturb the gut microbiota at the abundance level but also alter the metabolic homeostasis of the Amur tiger., (Copyright © 2018 Elsevier Inc. All rights reserved.)

With fewer than 200 tigers (Panthera tigris tigris) left in Nepal, that are generally confined to five protected areas across the Terai Arc Landscape, genetic studies are needed to provide crucial information on diversity and connectivity for devising an effective country-wide tiger conservation strategy. As part of the Nepal Tiger Genome Project, we studied landscape change, genetic variation, population structure, and gene flow of tigers across the Terai Arc Landscape by conducting Nepal's first comprehensive and systematic scat-based, non-invasive genetic survey. Of the 770 scat samples collected opportunistically from five protected areas and six presumed corridors, 412 were tiger (57%). Out of ten microsatellite loci, we retain eight markers that were used in identifying 78 individual tigers. We used this dataset to examine population structure, genetic variation, contemporary gene flow, and potential population bottlenecks of tigers in Nepal. We detected three genetic clusters consistent with three demographic sub-populations and found moderate levels of genetic variation (He = 0.61, AR = 3.51) and genetic differentiation (FST = 0.14) across the landscape. We detected 3-7 migrants, confirming the potential for dispersal-mediated gene flow across the landscape. We found evidence of a bottleneck signature likely caused by large-scale land-use change documented in the last two centuries in the Terai forest. Securing tiger habitat including functional forest corridors is essential to enhance gene flow across the landscape and ensure long-term tiger survival. This requires cooperation among multiple stakeholders and careful conservation planning to prevent detrimental effects of anthropogenic activities on tigers.

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 km 2 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.

Despite massive global conservation strategies, tiger populations continued to decline until recently, mainly due to habitat loss, human-animal conflicts, and poaching. These factors are known to affect the genetic characteristics of tiger populations and decrease local effective population sizes. The Terai Arc Landscape (TAL) at the foothills of the Himalaya is one of the 42 source sites of tigers around the globe. Therefore, information on how landscape features and anthropogenic factors affect the fine-scale spatial genetic structure and variation of tigers in TAL is needed to develop proper management strategies for achieving long-term conservation goals. We document, for the first time, the genetic characteristics of this tiger population by genotyping 71 tiger samples using 13 microsatellite markers from the western region of TAL (WTAL) of 1800 km2. Specifically, we aimed to estimate the genetic variability, population structure, and gene flow. The microsatellite markers indicated that the levels of allelic diversity (MNA = 6.6) and genetic variation (Ho = 0.50, HE = 0.64) were slightly lower than those reported previously in other Bengal tiger populations. We observed moderate gene flow and significant genetic differentiation (FST= 0.060) and identified the presence of cryptic genetic structure using Bayesian and non-Bayesian approaches. There was low and significantly asymmetric migration between the two main subpopulations of the Rajaji Tiger Reserve and the Corbett Tiger Reserve in WTAL. Sibship relationships indicated that the functionality of the corridor between these subpopulations may be retained if the quality of the habitat does not deteriorate. However, we found that gene flow is not adequate in view of changing land use matrices. We discuss the need to maintain connectivity by implementing the measures that have been suggested previously to minimize the level of human disturbance, including relocation of villages and industries, prevention of encroachment, and banning sand and boulder mining in the corridors.

The Siberian tiger, Panthera tigris altaica, is an endangered species, and much more work is needed to protect this species, which is still vulnerable to extinction. Conservation efforts may be supported by the genetic assessment of wild populations, for which highly specific microsatellite markers are required. However, only a limited amount of genetic sequence data is available for this species. To identify the genes involved in the lung transcriptome and to develop additional simple sequence repeat (SSR) markers for the Siberian tiger, we used high-throughput RNA-Seq to characterize the Siberian tiger transcriptome in lung tissue (designated 'PTA-lung') and a pooled tissue sample (designated 'PTA'). Approximately 47.5 % (33,187/69,836) of the lung transcriptome was annotated in four public databases (Nr, Swiss-Prot, KEGG, and COG). The annotated genes formed a potential pool for gene identification in the tiger. An analysis of the genes differentially expressed in the PTA lung, and PTA samples revealed that the tiger may have suffered a series of diseases before death. In total, 1062 non-redundant SSRs were identified in the Siberian tiger transcriptome. Forty-three primer pairs were randomly selected for amplification reactions, and 26 of the 43 pairs were also used to evaluate the levels of genetic polymorphism. Fourteen primer pairs (32.56 %) amplified products that were polymorphic in size in P. tigris altaica. In conclusion, the transcriptome sequences will provide a valuable genomic resource for genetic research, and these new SSR markers comprise a reasonable number of loci for the genetic analysis of wild and captive populations of P. tigris altaica.

Over the past century, the endangered Amur tiger (Panthera tigris altaica) has experienced a severe contraction in demography and geographic range because of habitat loss, poaching, and prey depletion. In its historical home in Northeast China, there appears to be a single tiger population that includes tigers in Southwest Primorye and Northeast China; however, the current demographic status of this population is uncertain. Information on the abundance, distribution and genetic diversity of this population for assessing the efficacy of conservation interventions are scarce. We used noninvasive genetic detection data from scats, capture-recapture models and an accumulation curve method to estimate the abundance of Amur tigers in Northeast China. We identified 11 individual tigers (6 females and 5 males) using 10 microsatellite loci in three nature reserves between April 2013 and May 2015. These tigers are confined primarily to a Hunchun Nature Reserve along the border with Russia, with an estimated population abundance of 9-11 tigers during the winter of 2014-2015. They showed a low level of genetic diversity. The mean number of alleles per locus was 2.60 and expected and observed heterozygosity were 0.42 and 0.49, respectively. We also documented long-distance dispersal (~270 km) of a male Amur tiger to Huangnihe Nature Reserve from the border, suggesting that the expansion of neighboring Russian populations may eventually help sustain Chinese populations. However, the small and isolated population recorded by this study demonstrate that there is an urgent need for more intensive regional management to create a tiger-permeable landscape and increased genetic connectivity with other populations.

Background: Non-invasive sampling has opened avenues for the genetic study of elusive species, which has contributed significantly to their conservation. Where field based identity of non-invasive sample is ambiguous (e.g. carnivore scats), it is essential to establish identity of the species through molecular approaches. A cost effective procedure to ascertain species identity is to use species specific primers (SSP) for PCR amplification and subsequent resolution through agarose gel electrophoresis. However, SSPs if ill designed can often cross amplify non-target sympatric species. Herein we report the problem of cross amplification with currently published SSPs, which have been used in several recent scientific articles on tigers (Panthera tigris) and leopards (Panthera pardus) in India. Since these papers form pioneering research on which future work will be based, an early rectification is required so as to not propagate this error further., Results: We conclusively show cross amplification of three of the four SSPs, in sympatric non-target species like tiger SSP amplifying leopard and striped hyena (Hyaena hyaena), and leopard SSP amplifying tiger, lion (Panthera leo persica) and clouded leopard (Neofelis nebulosa), with the same product size. We develop and test a non-cross-amplifying leopard specific primer pair within the mitochondrial cytochrome b region. We also standardize a duplex PCR method to screen tiger and leopard samples simultaneously in one PCR reaction to reduce cost and time., Conclusions: These findings suggest the importance of an often overlooked preliminary protocol of conclusive identification of species from non-invasive samples. The cross amplification of published primers in conspecifics suggests the need to revisit inferences drawn by earlier work.