Javan tiger DNA sequencing 79
polymorphism (SNP) panels (Khan et al., 2022), low-depth sequencing (Fuentes-Pardo & Ruzzante, 2017), multiplex PCR panels (Natesh et al., 2019) and pooled sequencing (Fuentes-Pardo & Ruzzante, 2017) are low-cost alternatives that overcome the limitations of mitochondrial DNA from non-invasive samples. At present, the need for com- putational infrastructure and expertise, high start-up costs, high import costs of reagents and lack of access to high-throughput sequencers are major barriers to using these alternatives in many tropical countries (Khan & Tyagi, 2021). In this study, we confirmthat the hair sample collected in
West Java nests within the clade of Sundaland tigers, but we are unable to assign it to a
subspecies.This is partially because there is no database of extinct tiger lineages. Although Sanger sequencing techniques are commonly used in several lower- andmiddle-income countries, the lack of databases remains a challenge. There are several specimens of the extinct Javan, Bali and Caspian Panthera tigris virgata tigers in museums (Yamaguchi et al., 2013), but genetic resources from these lineages are lacking. This has limited our ability to explore the possibility of shared haplotypes in extant tigers and to determine the similarities and differences between extinct and extant lineages. Wilting et al. (2015), for example, de- monstrated the inability of short DNA sequences to resolve tiger subspecies, whereas whole-genome sequencing studies (Liu et al., 2018; Armstrong et al., 2021) have been success- ful in doing so, at least for extant lineages. We highlight some of the challenges in studies of big cat
genetics (Wirdateti et al., 2024): (1) There are few good- quality samples of big cats and use of non-invasive samples is the norm. Because of the abundance of mitochondrial DNAcompared to nuclearDNAin cells, it has been common practice to analyse mitochondrial sequences. However, Numts are common in big cats and can lead tomisrepresen- tation in analyses. (2) Despite the availability of methods for analysing whole genomes fromnon-invasive samples such as shed hair and faeces (Khan et al., 2020;Tyagi et al., 2022; Khan 2023), the lack of sequencing facilities, computational infrastructure,DNA enrichment reagents or SNP panels ren- der these tools inaccessible in some countries (Khan&Tyagi, 2021). (3) There is a need to develop expertise in next- generation sequencing in biodiversity-rich tropical countries to support the discovery, conservation and management of the species in these regions. (4) Big cats and their parts are often subjects of forensic investigation in cases of trafficking or livestock depredation or in cases such as that ofWirdateti et al. (2024). All such investigations could be affected by Numts if their analyses are only restricted to the mitogenome. There is a need for cost-effective SNP panels to be developed for ancestry determination and population assignment (Khan et al., 2022). (5) Databases such as GenBank, and re- search journals, need to insist and verify that the sequences they report are archived properly and made available, as the
lack of genetic data hampers further studies in this field. For example, despite there being numerous genetic studies of ti- gers, these sequences are difficult to retrieve or use because of improper annotations or a lack of archiving; e.g. the se- quences from Wilting et al. (2015) need better annotation, and the genome assemblies from Armstrong et al. (2021) and Zhang et al. (2023) need to be released, along with those of many other studies. Making these data available would facilitate progress in the science of large felid genetics.
Author contributions Study conceptualization, analysis, writing: AK; coordination of author communication: SGA; laboratory experi- ments: YY; supervision: WW; revision: all authors.
Acknowledgements This research received no specific grant from any funding agency or commercial or not-for-profit sectors.
Conflicts of interest None.
Ethical standards This research abided by the Oryx guidelines on ethical standards.
Data availability The sequences generated will be submitted to the National Center for Biotechnology Information.
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Oryx, 2025, 59(1), 75–80 © The Author(s), 2025. Published by Cambridge University Press on behalf of Fauna & Flora International doi:10.1017/S0030605324001297
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