Genotyping faecal samples of Bengal tiger (Panthera tigris tigris) for population estimation: a pilot study. BMC Genet 7:48

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BMC Genetics (Impact Factor: 2.36). 02/2006; 7(1):48. DOI: 10.1186/1471-2156-7-48
Source: PubMed

ABSTRACT Bengal tiger Panthera tigris tigris the National Animal of India, is an endangered species. Estimating populations for such species is the main objective for designing conservation measures and for evaluating those that are already in place. Due to the tiger's cryptic and secretive behaviour, it is not possible to enumerate and monitor its populations through direct observations; instead indirect methods have always been used for studying tigers in the wild. DNA methods based on non-invasive sampling have not been attempted so far for tiger population studies in India. We describe here a pilot study using DNA extracted from faecal samples of tigers for the purpose of population estimation.
In this study, PCR primers were developed based on tiger-specific variations in the mitochondrial cytochrome b for reliably identifying tiger faecal samples from those of sympatric carnivores. Microsatellite markers were developed for the identification of individual tigers with a sibling Probability of Identity of 0.005 that can distinguish even closely related individuals with 99.9% certainty. The effectiveness of using field-collected tiger faecal samples for DNA analysis was evaluated by sampling, identification and subsequently genotyping samples from two protected areas in southern India.
Our results demonstrate the feasibility of using tiger faecal matter as a potential source of DNA for population estimation of tigers in protected areas in India in addition to the methods currently in use.

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    • "PCR products were checked on an ethidium bromide-stained 1.5% agarose gel and sent for direct fragment analysis using an ABI Prism 3730xl DNA Analyzer at the University of Illinois DNA Core Sequencing Facility. Two sex chromosome genes, Zinc-finger and Amelogenin, using primers specifically designed for felids (Pilgrim et al. 2005; Bhagavatula and Singh 2006) were amplified separately using the same PCR conditions used for microsatellite loci, checked, and sent for fragment analysis. "
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    ABSTRACT: Landscape fragmentation is often a major cause of species extinction as it can affect a wide variety of ecological processes. The impact of fragmentation varies among species depending on many factors, including their life-history traits and dispersal abilities. Felids are one of the groups most threatened by fragmented landscapes because of their large home ranges, territorial behavior, and low population densities. Here, we model the impacts of habitat fragmentation on patterns of genetic diversity in the guigna (Leopardus guigna), a small felid that is closely associated with the heavily human-impacted temperate rainforests of southern South America. We assessed genetic variation in 1798 base pairs of mitochondrial DNA sequences, 15 microsatellite loci, and 2 sex chromosome genes and estimated genetic diversity, kinship, inbreeding, and dispersal in 38 individuals from landscapes with differing degrees of fragmentation on Chiloé Island in southern Chile. Increased fragmentation was associated with reduced genetic diversity, but not with increased kinship or inbreeding. However, in fragmented landscapes, there was a weaker negative correlation between pairwise kinship and geographic distance, suggesting increased dispersal distances. These results highlight the importance of biological corridors to maximize connectivity in fragmented landscapes and contribute to our understanding of the broader genetic consequences of habitat fragmentation, especially for forest-specialist carnivores. © The American Genetic Association 2015. All rights reserved. For permissions, please e-mail:
    Journal of Heredity 01/2015; 2015:522-536. DOI:10.1093/jhered/esv025 · 1.97 Impact Factor
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    • "leo persica) (Singh et al., 2002), South China tiger (P. t. amoyensis) (Zhang et al., 2006), puma (Puma concolor) (Kurushima et al., 2006) and Bengal tiger (Bhagvatula and Singh, 2006; Sharma et al., 2008) among the Felidae. The use of species-specific microsatellites is limited by the time and expense involved and the difficulty in isolating these short tandem repeats and their flanking regions from the genomes of the target organisms. "
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    ABSTRACT: Comparison of genetic diversity indices of heterologous and species-specific microsatellite loci within a species may provide a panel of appropriate markers for genetic studies, but few studies have carried out such comparisons. We examined and compared the genetic characteristics of tiger-specific and heterologous loci in eight captive Bengal tigers. The mean polymorphic information content (PIC) value of the tiger-specific microsatellite loci (n = 15) was 0.447, and the number of alleles was from 2 to 4 per locus. In comparison, the heterologous microsatellite loci (n = 15) had a mean PIC value of 0.539, and the number of alleles per locus was three to five. Our findings indicate that the heterologous markers have a higher frequency (n = 11) of polymorphic microsatellite loci and number of alleles per locus compared with tiger-specific loci. We pooled the highly polymorphic (PIC > 0.5) tiger-specific loci (n = 5) and heterologous microsatellite loci (n = 11) except one and noted a higher mean observed heterozygosity and PIC values of 0.668 and 0.575, respectively, compared with the heterologous and tiger-specific loci taken alone. Using a locus selection criterion of PIC > 0.5, we recommend a combined panel of 16 highly polymorphic loci for genetic studies of the wild population of the Bengal tigers and suggest that either a combination of tiger-specific and heterologous microsatellite primers or heterologous primers be used in genetic studies related to the ecology, biology, socio-biology and behavior of Bengal tigers as >13 loci are needed in such studies. INTRODUCTION The tiger (Panthera tigris) once had the widest geographical distribution among cat species, extending from almost 10° south of the equator (Bali and Java) to beyond 60° north (the Russian Far East) and through more than100°
    The Central African journal of medicine 02/2014; 13(8):936-943. DOI:10.5897/AJB12.2977
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    • "Most of the studies undertaken so far on the Bengal tiger (Bhagavatula and Singh 2006; Mondol et al. 2009b; Reddy et al. 2012; Sharma et al. 2013) fail to provide detailed information on locus-specific genetic characteristics (polymorphic information content [PIC] and probability of identity [P ID ]) and genotyping profile characteristics (stutter, allele to peak height etc.). Besides, information of these studies have been from fecal DNA, except for a few loci, which have been studied using high-quality DNA (Bhagavatula and Singh 2006; Mondol et al. 2009b). Thus, selection of the best loci for use in population genetics and forensic studies and minimizing genotyping errors has hitherto been precluded. "
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    ABSTRACT: In India, six landscapes and source populations that are important for long-term conservation of Bengal tigers (Panthera tigris tigris) have been identified. Except for a few studies, nothing is known regarding the genetic structure and extent of gene flow among most of the tiger populations across India as the majority of them are small, fragmented and isolated. Thus, individual-based relationships are required to understand the species ecology and biology for planning effective conservation and genetics-based individual identification has been widely used. But this needs screening and describing characteristics of microsatellite loci from DNA from good-quality sources so that the required number of loci can be selected and the genotyping error rate minimized. In the studies so far conducted on the Bengal tiger, a very small number of loci (n = 35) have been tested with high-quality source of DNA, and information on locus-specific characteristics is lacking. The use of such characteristics has been strongly recommended in the literature to minimize the error rate and by the International Society for Forensic Genetics (ISFG) for forensic purposes. Therefore, we describe for the first time locus-specific genetic and genotyping profile characteristics, crucial for population genetic studies, using high-quality source of DNA of the Bengal tiger. We screened 39 heterologous microsatellite loci (Sumatran tiger, domestic cat, Asiatic lion and snow leopard) in captive individuals (n = 8), of which 21 loci are being reported for the first time in the Bengal tiger, providing an additional choice for selection. The mean relatedness coefficient (R = -0.143) indicates that the selected tigers were unrelated. Thirty-four loci were polymorphic, with the number of alleles ranging from 2 to 7 per locus, and the remaining five loci were monomorphic. Based on the PIC values (> 0.500), and other characteristics, we suggest that 16 loci (3 to 7 alleles) be used for genetic and forensic study purposes. The probabilities of matching genotypes of unrelated individuals (3.692 × 10(-19)) and siblings (4.003 × 10(-6)) are within the values needed for undertaking studies in population genetics, relatedness, sociobiology and forensics.
    SpringerPlus 01/2014; 3:4. DOI:10.1186/2193-1801-3-4
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