Genotyping‐in‐Thousands by sequencing reveals marked population structure in Western Rattlesnakes to inform conservation status

Abstract

Delineation of units below the species level is critical for prioritizing conservation actions for species at‐risk. Genetic studies play an important role in characterizing patterns of population connectivity and diversity to inform the designation of conservation units, especially for populations that are geographically isolated. The northernmost range margin of Western Rattlesnakes ( Crotalus oreganus ) occurs in British Columbia, Canada, where it is federally classified as threatened and restricted to five geographic regions. In these areas, Western Rattlesnakes hibernate (den) communally, raising questions about connectivity within and between den complexes. At present, Western Rattlesnake conservation efforts are hindered by a complete lack of information on genetic structure and degree of isolation at multiple scales, from the den to the regional level. To fill this knowledge gap, we used Genotyping‐in‐Thousands by sequencing (GT‐seq) to genotype an optimized panel of 362 single nucleotide polymorphisms (SNPs) from individual samples ( n = 461) collected across the snake's distribution in western Canada and neighboring Washington (USA). Hierarchical STRUCTURE analyses found evidence for population structure within and among the five geographic regions in BC, as well as in Washington. Within these regions, 11 genetically distinct complexes of dens were identified, with some regions having multiple complexes. No significant pattern of isolation‐by‐distance and generally low levels of migration were detected among den complexes across regions. Additionally, snakes within dens generally were more related than those among den complexes within a region, indicating limited movement. Overall, our results suggest that the single, recognized designatable unit for Western Rattlesnakes in Canada should be re‐assessed to proactively focus conservation efforts on preserving total genetic variation detected range‐wide. More broadly, our study demonstrates a novel application of GT‐seq for investigating patterns of diversity in wild populations at multiple scales to better inform conservation management.

Genotyping-in-thousands by sequencing (GT-seq) of non-invasive fecal and degraded samples: a new panel to enable ongoing monitoring of Canadian polar bear populations

“…ddRADseq), as long as there are loci in common. However, differences in genotyping calling methods between GT-seq and RADseq datasets may contribute to genotype discordance among datasets (Schmidt et al, 2020). To address this possibility, we called genotypes from our GT-seq data using both the original calling pipeline for GT-seq and a BCFTOOLS workflow previously used to call genotypes from ddRADseq data (Jensen et al, 2020).…”

“…We used the GT-seq pipeline available on GitHub (https://github.com/GTseq/GTseq-Pipeline) for filtering to a minimum depth of 10 and genotype calling, as suggested by Campbell et al (2015). However, we were also interested in examining genotype discordance between different SNP calling models, as some discrepancy has been found between the GT-seq pipeline and other workflows used to call RADseq data (Schmidt et al, 2020). As BCFTOOLS (Li, 2011) is a common genotyping tool and was used to call genotypes for our original ddRADseq dataset (Jensen et al, 2020), we used this same workflow to process our raw GT-seq sequencing reads.…”

“…The technique uses suites of multiplexed, species-specific primers, which may also mitigate some of the challenges presented by exogenous DNA and degraded host DNA in non-invasive fecal samples. GT-seq has been successfully applied to minimally invasive cloacal swab DNA samples collected from Western Rattlesnakes (Crotalus oreganus ) with low rates of genotyping error and discordance relative to RADseq (Schmidt et al, 2020). Natesh et al (2019) also found high genotyping success using GT-seq of non-invasive scat samples for Indian Bengal Tigers (Panthera tigris tigris ).…”

RapidRat: Development, validation and application of a genotyping-by-sequencing panel for rapid biosecurity and invasive species management

“…Originally demonstrated in steelhead trout [24], GT-seq has been subsequently applied to the study of other fish species, including redband trout [66], brook trout [67], Pacific lamprey [68], and walleye [69]. More recently, the efficacy of using GT-seq for genotyping minimally-invasive samples has been demonstrated [46], and successfully applied to further understanding of the molecular ecology and conservation status of the at-risk Western rattlesnake [70]. To the best of our knowledge, our study is the first to demonstrate use of GT-seq to inform invasive species management to help mitigate biodiversity loss.…”

Corrigendum

“…In the recent article by Schmidt et al (2020), the authors inadvertently neglected to acknowledge Daniel Beck, Joey Chase, and J. D. Brooks (Central Washington University), who contributed samples that were invaluable to this study. The updated Acknowledgements section is as follows:…”