Stuart B. Piertney

DNA Barcoding using Sanger sequencing is a popular technique for identifying species on a per specimen basis. However for larger projects, sequencing individual voucher specimens can be time and resource intensive and moreover is associated with high levels of sequencing failure and contamination. Oxford Nanopore Sequencing Technology (ONT) has emerged as a scalable alternative, capable of generating hundreds of DNA barcodes simultaneously using the portable, benchtop MinION sequencing device. In this study we aim to compare and contrast the sequencing outcomes of Oxford Nanopore R10 Flongle flowcells verses Sanger sequencing for DNA barcoding and produce a DNA barcode reference library. We demonstrate that DNA barcodes generated using ONT outperform those produced by Sanger sequencing in terms of recovery and sequence quality with lower rates of contamination. We then produced DNA barcodes for 146 seagrass associated marine invertebrate OTUs collected from four seagrass beds in Scotland, targeting COI and 18S V4 regions. Using both markers, we show the number of recovered OTUs was higher than if each marker was used in isolation and make use of degenerate and group-specific primer pairs to improve recovery. Furthermore we demonstrate how mapping ONT reads to pre-existing DNA barcodes can be used to reduce ambiguous basecalls and improve recovery of sequences from contaminated specimens. Overall this study informs prospective users intending to carry out multimarker DNA barcode projects using Oxford Nanopore Sequencing. Furthermore, we generated the first DNA barcode reference library for seagrass beds in Scotland to support future biomonitoring of these priority habitats.

The examination of genetic structure in the deep‐ocean hadal zone has focused on divergence between tectonic trenches to understand how environment and geography may drive species divergence and promote endemism. There has been little attempt to examine localized genetic structure within trenches, partly because of logistical challenges associated with sampling at an appropriate scale, and the large effective population sizes of species that can be sampled adequately may mask underlying genetic structure. Here we examine genetic structure in the superabundant amphipod Hirondellea gigas in the Mariana Trench at depths of 8126–10,545 m. RAD sequencing was used to identify 3182 loci containing 43,408 single nucleotide polymorphisms (SNPs) across individuals after stringent pruning of loci to prevent paralogous multicopy genomic regions being erroneously merged. Principal components analysis of SNP genotypes resolved no genetic structure between sampling locations, consistent with a signature of panmixia. However, discriminant analysis of principal components identified divergence between all sites driven by 301 outlier SNPs in 169 loci and significantly associated with latitude and depth. Functional annotation of loci identified differences between singleton loci used in analysis and paralogous loci pruned from the data set and also between outlier and nonoutlier loci, all consistent with hypotheses explaining the role of transposable elements driving genome dynamics. This study challenges the traditional perspective that highly abundant amphipods within a trench form a single panmictic population. We discuss the findings in relation to eco‐evolutionary and ontogenetic processes operating in the deep sea, and highlight key challenges associated with population genetic analysis in nonmodel systems with inherent large effective population sizes and genomes.

A population's effective size (N_e) is a key parameter that shapes rates of inbreeding and loss of genetic diversity, thereby influencing evolutionary processes and population viability. However, estimating N_e, and identifying key demographic mechanisms that underlie the N_e to census population size (N) ratio, remains challenging, especially for small populations with overlapping generations and substantial environmental and demographic stochasticity and hence dynamic age‐structure. A sophisticated demographic method of estimating N_e/N, which uses Fisher's reproductive value to account for dynamic age‐structure, has been formulated. However, this method requires detailed individual‐ and population‐level data on sex‐ and age‐specific reproduction and survival, and has rarely been implemented. Here, we use the reproductive value method and detailed demographic data to estimate N_e/N for a small and apparently isolated red‐billed chough (Pyrrhocorax pyrrhocorax) population of high conservation concern. We additionally calculated two single‐sample molecular genetic estimates of N_e to corroborate the demographic estimate and examine evidence for unobserved immigration and gene flow. The demographic estimate of N_e/N was 0.21, reflecting a high total demographic variance () of 0.71. Females and males made similar overall contributions to . However, contributions varied among sex‐age classes, with greater contributions from 3 year‐old females than males, but greater contributions from ≥5 year‐old males than females. The demographic estimate of N_e was ~30, suggesting that rates of increase of inbreeding and loss of genetic variation per generation will be relatively high. Molecular genetic estimates of N_e computed from linkage disequilibrium and approximate Bayesian computation were approximately 50 and 30, respectively, providing no evidence of substantial unobserved immigration which could bias demographic estimates of N_e. Our analyses identify key sex‐age classes contributing to demographic variance and thus decreasing N_e/N in a small age‐structured population inhabiting a variable environment. They thereby demonstrate how assessments of N_e can incorporate stochastic sex‐ and age‐specific demography and elucidate key demographic processes affecting a population's evolutionary trajectory and viability. Furthermore, our analyses show that N_e for the focal chough population is critically small, implying that management to re‐establish genetic connectivity may be required to ensure population viability.

Norway rats are a globally distributed invasive species, which have colonized many islands around the world, including in the South Atlantic Ocean. We investigated the phylogeography of Norway rats across the South Atlantic Ocean and bordering continental countries. We identified haplotypes from 517 bp of the hypervariable region I of the mitochondrial D-loop and constructed a Bayesian consensus tree and median-joining network incorporating all other publicly available haplotypes via an alignment of 364 bp. Three Norway rat haplotypes are present across the islands of the South Atlantic Ocean, including multiple haplotypes separated by geographic barriers within island groups. All three haplotypes have been previously recorded from European countries. Our results support the hypothesis of rapid Norway rat colonization of South Atlantic Ocean islands by sea-faring European nations from multiple European ports of origin. This seems to have been the predominant pathway for repeated Norway rat invasions of islands, even within the same archipelago, rather than within-island dispersal across geographic barriers.

Summary: The control and eradication of invasive species is a common management strategy to protect or restore native biodiversity. On South Georgia in the Southern Ocean, the brown rat Rattus norvegicus was brought onto the island with the onset of whaling and sealing activity in the 1800s and has had a significant detrimental impact on key bird species of conservation concern. Efforts to eradicate rats from South Georgia using poisoned bait are ongoing. Despite the South Georgia rat eradication programme being the geographically largest and most ambitious eradication initiative to date, its success is facilitated by the potential that rat populations are effectively isolated by glacial barriers. This allows for localized eradication effort at manageable scales, leading to sequential eradication of individual populations with minimal risk of incursion from neighbouring areas. Here, we use the levels of population genetic divergence estimated from 299 single nucleotide polymorphism (SNP) loci and DNA sequence variation across 993 base pairs of the mitochondrial DNA cytochrome B locus to examine whether rat populations from nine glacially isolated areas on South Georgia are genetically distinct and so can be treated as independent eradication units. Bayesian clustering of individuals based on SNP similarity identified seven different genetic groups, which were confirmed using analyses based on pairwise genetic distance estimates and ordination of individuals using principal coordinate analysis. From a management perspective, these seven groups represent individual targets in baiting operations. Two mtDNA haplotypes were resolved across South Georgia, with a distinct geographical separation between the north‐western and south‐eastern populations. Approximate Bayesian computation (ABC) was used to identify that this divergence was a consequence of two separate historical colonization events. Synthesis and applications. We illustrate that molecular markers are a valuable tool in species management and pest eradication given that the spatial distribution of genetic diversity can: (i) identify demographically and genetically independent populations on which local eradication effort can be focussed, (ii) distinguish between incomplete eradication and immigration in situations where individuals remain after eradication has been attempted and (iii) identify the source of migrants when dispersal occurs over large spatial scales.

Landscape genomics promises to provide novel insights into how neutral and adaptive processes shape genome‐wide variation within and among populations. However, there has been little emphasis on examining whether individual‐based phenotype–genotype relationships derived from approaches such as genome‐wide association (GWAS) manifest themselves as a population‐level signature of selection in a landscape context. The two may prove irreconcilable as individual‐level patterns become diluted by high levels of gene flow and complex phenotypic or environmental heterogeneity. We illustrate this issue with a case study that examines the role of the highly prevalent gastrointestinal nematode Trichostrongylus tenuis in shaping genomic signatures of selection in red grouse (Lagopus lagopus scotica). Individual‐level GWAS involving 384 SNPs has previously identified five SNPs that explain variation in T. tenuis burden. Here, we examine whether these same SNPs display population‐level relationships between T. tenuis burden and genetic structure across a small‐scale landscape of 21 sites with heterogeneous parasite pressure. Moreover, we identify adaptive SNPs showing signatures of directional selection using outlier analysis and relate population‐ and individual‐level patterns of multilocus neutral and adaptive genetic structure to T. tenuis burden. The five candidate SNPs for parasite‐driven selection were neither associated with T. tenuis burden on a population level, nor under directional selection. Similarly, there was no evidence of parasite‐driven selection in SNPs identified as candidates for directional selection. We discuss these results in the context of red grouse ecology and highlight the broader consequences for the utility of landscape genomics approaches for identifying signatures of selection.