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Shallow shotgun sequencing of the microbiome recapitulates 16S amplicon results and provides functional insights
Stothart, Mason; McLoughlin, Philip; Poissant, Jocelyn — 2022-09-28 <p>Prevailing 16S rRNA gene-amplicon methods for characterizing the bacterial microbiome of wildlife are economical, but result in coarse taxonomic classifications, are subject to primer and 16S copy number biases, and do not allow for direct estimation of microbiome functional potential. While deep shotgun metagenomic sequencing can overcome many of these limitations, it is prohibitively expensive for large sample sets. We evaluated the ability of shallow shotgun metagenomic sequencing to characterize taxonomic and functional patterns in the fecal microbiome of a model population of feral horses (Sable Island, Canada). Since 2007, this unmanaged population has been the subject of an individual-based, long-term ecological study. Using deep shotgun metagenomic sequencing, we determined the sequencing depth required to accurately characterize the horse microbiome. In comparing conventional versus high-throughput shotgun metagenomic library preparation techniques, we validate the use of more cost-effective lab methods. Finally, we characterize similarities between 16S amplicon and shallow shotgun characterization of the microbiome and demonstrate that the latter recapitulates biological patterns first described in a published amplicon dataset. Unlike amplicon data, we further demonstrate how shallow shotgun metagenomic data provide useful insights about microbiome functional potential which support previously hypothesized diet effects in this study system.</p>
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Bacterial dispersal and drift drive microbiome diversity patterns within a population of feral hindgut fermenters
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Stothart, Mason; Greuel, Ruth; Gavriliuc, Stefan; Henry, Astrid; Wilson, Alastair; McLoughlin, Philip; Poissant, Jocelyn — 2020-11-20 <p>Studies of microbiome variation in wildlife often emphasize host physiology and diet as proximate selective pressures acting on host-associated microbiota. In contrast, microbial dispersal and ecological drift are more rarely considered. Using amplicon sequencing, we characterized the bacterial microbiome of adult female (<i>n</i> = 86) Sable Island horses (Nova Scotia, Canada) as part of a detailed individual-based study of this feral population. Using data on sampling date, horse location, age, parental status, and local habitat variables, we contrasted the ability of spatiotemporal, life history, and environmental factors to explain microbiome diversity among Sable Island horses. We extended inferences made from these analyses with both phylogeny-informed and phylogeny-independent null modeling approaches to identify deviations from stochastic expectations. Phylogeny-informed diversity measures were correlated with spatial and local habitat variables, but null modelling results suggested that heterogeneity in ecological drift, rather than differential selective pressures acting on the microbiome, was responsible for these correlations. Conversely, phylogeny-independent diversity measures were best explained by host spatial and social structure, suggesting that taxonomic composition of the microbiome was shaped most strongly by bacterial dispersal. Parental status was important but correlated with measures of β-dispersion rather than β-diversity (mares without foals had lower alpha diversity and more variable microbiomes than mares with foals). Our results suggest that between host microbiome variation within the Sable Island horse population is driven more strongly by bacterial dispersal and ecological drift than by differential selective pressures. These results emphasize the need to consider alternative ecological processes in the study of microbiomes.</p> https://creativecommons.org/publicdomain/zero/1.0/
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Data from: Estimating genome-wide heterozygosity: effects of demographic history and marker type
Miller, Joshua M.; Malenfant, René M.; David, Patrice; Davis, Corey S.; Poissant, Jocelyn; Hogg, John T.; Festa-Bianchet, Marco; Coltman, David W. — 2013-09-17 Heterozygosity–fitness correlations (HFCs) are often used to link individual genetic variation to differences in fitness. However, most studies examining HFCs find weak or no correlations. Here, we derive broad theoretical predictions about how many loci are needed to adequately measure genomic heterozygosity assuming different levels of identity disequilibrium (ID), a proxy for inbreeding. We then evaluate the expected ability to detect HFCs using an empirical data set of 200 microsatellites and 412 single nucleotide polymorphisms (SNPs) genotyped in two populations of bighorn sheep (Ovis canadensis), with different demographic histories. In both populations, heterozygosity was significantly correlated across marker types, although the strength of the correlation was weaker in a native population compared with one founded via translocation and later supplemented with additional individuals. Despite being bi-allelic, SNPs had similar correlations to genome-wide heterozygosity as microsatellites in both populations. For both marker types, this association became stronger and less variable as more markers were considered. Both populations had significant levels of ID; however, estimates were an order of magnitude lower in the native population. As with heterozygosity, SNPs performed similarly to microsatellites, and precision and accuracy of the estimates of ID increased as more loci were considered. Although dependent on the demographic history of the population considered, these results illustrate that genome-wide heterozygosity, and therefore HFCs, are best measured by a large number of markers, a feat now more realistically accomplished with SNPs than microsatellites.
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Data from: Genomic consequences of genetic rescue in an insular population of bighorn sheep (Ovis canadensis)
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Miller, Joshua M.; Poissant, Jocelyn; Hogg, John T.; Coltman, David W. — 2011-12-01 Genetic rescue is a management intervention whereby a small population is supplemented with individuals from other populations in an attempt to reverse the effects of inbreeding and increased genetic load. One such rescue was recently documented in the population of bighorn sheep (Ovis canadensis) within the National Bison Range wildlife refuge (Montana, USA). Here we examine the locus-specific effects of rescue in this population using a newly developed genome-wide set of 195 microsatellite loci and first-generation linkage map. We found that the rate of introgression varied among loci, and that 111 loci, 57% of those examined, deviated from patterns of neutral inheritance. The most common deviation was an excess of homozygous genotypes relative to neutral expectations, indicative of directional selection. As in previous study of this rescue, individuals with more introduced alleles had higher lifetime reproductive success and longevity. In addition, we found 30 loci, spread throughout the genome, which seem to have individual effects on these life history traits. Though the potential for outbreeding depression is a major concern when translocating individuals between populations, we found no evidence of such effects in this population.
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Data from: Individual quality and age but not environmental or social conditions modulate costs of reproduction in a capital breeder
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Debeffe, Lucie; Poissant, Jocelyn; McLoughlin, Philip D. — 2018-05-08 Costs associated with reproduction are widely known to play a role in the evolution of reproductive tactics with consequences to population and eco-evolutionary dynamics. Evaluating these costs as they pertain to species in the wild remains an important goal of evolutionary ecology. Individual heterogeneity, including differences in individual quality (i.e., among-individual differences in traits associated with survival and reproduction) or state, and variation in environmental and social conditions can modulate the costs of reproduction; however, few studies have considered effects of these factors simultaneously. Taking advantage of a detailed, long-term dataset for a population of feral horses (Sable Island, Nova-Scotia, Canada), we address the question of how intrinsic (quality, age), environmental (winter severity, location), and social conditions (group size, composition, sex ratio, density) influence the costs of reproduction on subsequent reproduction. Individual quality was measured using a multivariate analysis on a combination of four static and dynamic traits expected to depict heterogeneity in individual performance. Female quality and age interacted with reproductive status of the previous year to determine current reproductive effort, while no effect of social or environmental covariates was found. High quality females showed higher probabilities of giving birth and weaning their foal regardless of their reproductive status the previous year, while those of lower quality showed lower probabilities of producing foals in successive years. Middle-aged (prime) females had the highest probability of giving birth when they had not reproduced the year before but no such relationship with age was found among females that had reproduced the previous year, indicating that prime-aged females bear higher costs of reproduction. We show that individual quality and age were key factors modulating the costs of reproduction in a capital breeder but that environmental or social conditions were not, highlighting the importance of considering multiple factors when studying costs of reproduction.
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Data from: Targeted genome-wide SNP genotyping in feral horses using non-invasive fecal swabs
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Gavriliuc, Stefan; Reza, Salman; Jeong, Chanwoori; McLoughlin, Philip; Poissant, Jocelyn — 2022-04-19 <p>The development of high-throughput sequencing has prompted a transition in wildlife genetics from using microsatellites toward sets of Single Nucleotide Polymorphisms (SNPs). However, genotyping large numbers of targeted SNPs using non-invasive samples remains challenging due to relatively large DNA input requirements. Recently, target enrichment has emerged as a promising approach requiring little template DNA. We assessed the efficacy of Tecan Genomics’ Allegro Targeted Genotyping (ATG) for generating genome-wide SNP data in feral horses using DNA isolated from fecal swabs. Total and host-specific DNA were quantified for 989 samples collected as part of a long-term individual-based study of feral horses on Sable Island, Nova Scotia, Canada, using dsDNA fluorescence and a host-specific qPCR assay, respectively. Forty-eight samples representing 44 individuals containing at least 10ng of host DNA (ATG’s recommended minimum input) were genotyped using a custom multiplex panel targeting 279 SNPs. Genotyping accuracy and consistency were assessed by contrasting ATG genotypes with those obtained from the same individuals with SNP microarrays, and from multiple samples from the same horse, respectively. 62% of swabs yielded the minimum recommended amount of host DNA for ATG. Ignoring samples that failed to amplify, ATG recovered an average of 86.7% targeted sites per sample, while genotype concordance between ATG and SNP microarrays was 98.5%. The repeatability of genotypes from the same individual approached unity with an average of 99.9%. This study demonstrates the suitability of ATG for genome-wide, non-invasive targeted SNP genotyping, and will facilitate further ecological and conservation genetics research in equids and related species.</p>
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Data from: Temporal dynamics of linkage disequilibrium in two populations of bighorn sheep
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Miller, Joshua M.; Poissant, Jocelyn; Malenfant, René M.; Hogg, John T.; Coltman, David W. — 2016-07-14 Linkage disequilibrium (LD) is the nonrandom association of alleles at two markers. Patterns of LD have biological implications as well as practical ones when designing association studies or conservation programs aimed at identifying the genetic basis of fitness differences within and among populations. However, the temporal dynamics of LD in wild populations has received little empirical attention. In this study, we examined the overall extent of LD, the effect of sample size on the accuracy and precision of LD estimates, and the temporal dynamics of LD in two populations of bighorn sheep (Ovis canadensis) with different demographic histories. Using over 200 microsatellite loci, we assessed two metrics of multi-allelic LD, D′, and χ′2. We found that both populations exhibited high levels of LD, although the extent was much shorter in a native population than one that was founded via translocation, experienced a prolonged bottleneck post founding, followed by recent admixture. In addition, we observed significant variation in LD in relation to the sample size used, with small sample sizes leading to depressed estimates of the extent of LD but inflated estimates of background levels of LD. In contrast, there was not much variation in LD among yearly cross-sections within either population once sample size was accounted for. Lack of pronounced interannual variability suggests that researchers may not have to worry about interannual variation when estimating LD in a population and can instead focus on obtaining the largest sample size possible.

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(Author: Poissant, Jocelyn)

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