Population genetic assessment of marine megafauna using seawater environmental DNA: a case study of whale sharks from the Ningaloo Coast World Heritage Area.

There is a growing need for scalable and cost-effective genetic monitoring tools to support the management of species and populations of conservation importance, driven by the increasing biodiversity loss, need for long-term population data and limitations of invasive genetic sampling. Whale sharks, along with other Endangered, elusive and highly migratory species, spend the majority of their lives in offshore ocean waters. This behaviour poses logistical and ethical challenges to invasive genetic sampling of the species, which is traditionally done via tissue biopsy. Here, we develop a genetic toolkit to study populations of whale sharks from seawater environmental DNA using short segments of nuclear DNA (100 to 300bp) containing two or more SNPs called microhaplotypes. Amplifying these markers from seawater collected in 1L bottles behind sharks showed that we can reliably genotype sharks from water samples taken immediately behind the animal. Moreover, we observed a strong relationship between population-level allele frequencies and estimates of genetic diversity between eDNA and tissue-derived samples, demonstrating the capability of eDNA in capturing population-level genetic information with high fidelity. We also analysed a tissue dataset of 72 sharks that attended the Ningaloo Coast World Heritage Area over six-years to showcase the utility of our SNP panel to study temporal processes in these populations. Our data revealed patterns of genetic variation through time that were consistent with whole genome techniques. Contributor estimation from mock environmental samples using various combinations of tissue-derived sequence data to estimate abundance of animals in a mixed DNA sample was shown to accurately identify the number of contributors in mixtures containing upto 10 individuals, beyond which biases were too large. Our findings illustrate the viability of using microhaplotype markers from seawater eDNA as a tool to study conservation genetics of whale sharks, with the potential for broader expansion of eDNA-based genetic assessments to other marine megafauna and aquatic species.