Controlling the false-positive rate in multilocus genome scans for selection

Rapid typing of genetic variation at many regions of the genome is an efficient way to survey variability in natural populations in an effort to identify segments of the genome that have experienced recent natural selection. Following such a genome scan, individual regions may be chosen for further sequencing and a more detailed analysis of patterns of variability, often to perform a parametric test for selection and to estimate the strength of a recent selective sweep. We show here that not accounting for the ascertainment of loci in such analyses leads to false inference of natural selection when the true model is selective neutrality, because the procedure of choosing unusual loci (in comparison to the rest of the genome-scan data) selects regions of the genome with genealogies similar to those expected under models of recent directional selection. We describe a simple and efficient correction for this ascertainment bias, which restores the false-positive rate to near-nominal levels. For the parameters considered here, we find that obtaining a test with the expected distribution of P-values depends on accurately accounting both for ascertainment of regions and for demography. Finally, we use simulations to explore the utility of relying on outlier loci to detect recent selective sweeps. We find that measures of diversity and of population differentiation are more effective than summaries of the site-frequency spectrum and that sequencing larger regions (2.5 kbp) in genome-scan studies leads to more power to detect recent selective sweeps.     

INTEGRATING LANDSCAPE GENOMICS AND SPATIALLY EXPLICIT APPROACHES TO DETECT LOCI UNDER SELECTION IN CLINAL POPULATIONS

Uncovering the genetic basis of adaptation hinges on the ability to detect loci under selection. However, population genomics outlier approaches to detect selected loci may be inappropriate for clinal populations or those with unclear population structure because they require that individuals be clustered into populations. An alternate approach, landscape genomics, uses individual‐based approaches to detect loci under selection and reveal potential environmental drivers of selection. We tested four landscape genomics methods on a simulated clinal population to determine their effectiveness at identifying a locus under varying selection strengths along an environmental gradient. We found all methods produced very low type I error rates across all selection strengths, but elevated type II error rates under “weak” selection. We then applied these methods to an AFLP genome scan of an alpine plant, Campanula barbata, and identified five highly supported candidate loci associated with precipitation variables. These loci also showed spatial autocorrelation and cline patterns indicative of selection along a precipitation gradient. Our results suggest that landscape genomics in combination with other spatial analyses provides a powerful approach for identifying loci potentially under selection and explaining spatially complex interactions between species and their environment.     

Genetic structure and seed-mediated dispersal rates of an endangered shrub in a fragmented landscape: a case study for Juniperus communis in northwestern Europe

Background

Population extinction risk in a fragmented landscape is related to the differential ability of the species to spread its genes across the landscape. The impact of landscape fragmentation on plant population dynamics will therefore vary across different spatial scales. We quantified successful seed-mediated dispersal of the dioecious shrub Juniperus communis in a fragmented landscape across northwestern Europe by using amplified fragment length polymorphism (AFLP) markers. Furthermore we investigated the genetic diversity and structure on two spatial scales: across northwestern Europe and across Flanders (northern Belgium). We also studied whether seed viability and populations size were correlated with genetic diversity.

Results

Unexpectedly, estimated seed-mediated dispersal rates were quite high and ranged between 3% and 14%. No population differentiation and no spatial genetic structure were detected on the local, Flemish scale. A significant low to moderate genetic differentiation between populations was detected at the regional, northwest European scale (PhiPT = 0.10). In general, geographically nearby populations were also genetically related. High levels of within-population genetic diversity were detected but no correlation was found between any genetic diversity parameter and population size or seed viability.

Conclusions

In northwestern Europe, landscape fragmentation has lead to a weak isolation-by-distance pattern but not to genetic impoverishment of common juniper. Substantial rates of successful migration by seed-mediated gene flow indicate a high dispersal ability which could enable Juniperus communis to naturally colonize suitable habitats. However, it is not clear whether the observed levels of migration will suffice to counterbalance the effects of genetic drift in small populations on the long run.     

LANDSCAPE GENOMICS IN ATLANTIC SALMON (SALMO SALAR): SEARCHING FOR GENE–ENVIRONMENT INTERACTIONS DRIVING LOCAL ADAPTATION

A growing number of studies are examining the factors driving historical and contemporary evolution in wild populations. By combining surveys of genomic variation with a comprehensive assessment of environmental parameters, such studies can increase our understanding of the genomic and geographical extent of local adaptation in wild populations. We used a large‐scale landscape genomics approach to examine adaptive and neutral differentiation across 54 North American populations of Atlantic salmon representing seven previously defined genetically distinct regional groups. Over 5500 genome‐wide single nucleotide polymorphisms were genotyped in 641 individuals and 28 bulk assays of 25 pooled individuals each. Genome scans, linkage map, and 49 environmental variables were combined to conduct an innovative landscape genomic analysis. Our results provide valuable insight into the links between environmental variation and both neutral and potentially adaptive genetic divergence. In particular, we identified markers potentially under divergent selection, as well as associated selective environmental factors and biological functions with the observed adaptive divergence. Multivariate landscape genetic analysis revealed strong associations of both genetic and environmental structures. We found an enrichment of growth‐related functions among outlier markers. Climate (temperature–precipitation) and geological characteristics were significantly associated with both potentially adaptive and neutral genetic divergence and should be considered as candidate loci involved in adaptation at the regional scale in Atlantic salmon. Hence, this study significantly contributes to the improvement of tools used in modern conservation and management schemes of Atlantic salmon wild populations.     

Divergent selection and heterogeneous genomic divergence

Levels of genetic differentiation between populations can be highly variable across the genome, with divergent selection contributing to such heterogeneous genomic divergence. For example, loci under divergent selection and those tightly physically linked to them may exhibit stronger differentiation than neutral regions with weak or no linkage to such loci. Divergent selection can also increase genome‐wide neutral differentiation by reducing gene flow (e.g. by causing ecological speciation), thus promoting divergence via the stochastic effects of genetic drift. These consequences of divergent selection are being reported in recently accumulating studies that identify: (i) ‘outlier loci’ with higher levels of divergence than expected under neutrality, and (ii) a positive association between the degree of adaptive phenotypic divergence and levels of molecular genetic differentiation across population pairs [‘isolation by adaptation’ (IBA)]. The latter pattern arises because as adaptive divergence increases, gene flow is reduced (thereby promoting drift) and genetic hitchhiking increased. Here, we review and integrate these previously disconnected concepts and literatures. We find that studies generally report 5–10% of loci to be outliers. These selected regions were often dispersed across the genome, commonly exhibited replicated divergence across different population pairs, and could sometimes be associated with specific ecological variables. IBA was not infrequently observed, even at neutral loci putatively unlinked to those under divergent selection. Overall, we conclude that divergent selection makes diverse contributions to heterogeneous genomic divergence. Nonetheless, the number, size, and distribution of genomic regions affected by selection varied substantially among studies, leading us to discuss the potential role of divergent selection in the growth of regions of differentiation (i.e. genomic islands of divergence), a topic in need of future investigation.     

A genome scan to detect candidate regions influenced by local natural selection in human populations

As human populations dispersed throughout the world, they were subjected to new selective forces, which must have led to local adaptation via natural selection and hence altered patterns of genetic variation. Yet, there are very few examples known in which such local selection has clearly influenced human genetic variation. A potential approach for detecting local selection is to screen random loci across the genome; those loci that exhibit unusually large genetic distances between human populations are then potential markers of genomic regions under local selection. We investigated this approach by genotyping 332 short tandem repeat (STR) loci in Africans and Europeans and calculating the genetic differentiation for each locus. Patterns of genetic diversity at these loci were consistent with greater variation in Africa and with local selection operating on populations as they moved out of Africa. For 11 loci exhibiting the largest genetic differences, we genotyped an additional STR locus located nearby; the genetic distances for these nearby loci were significantly larger than average. These genomic regions therefore reproducibly exhibit larger genetic distances between populations than the "average" genomic region, consistent with local selection. Our results demonstrate that genome scans are a promising means of identifying candidate regions that have been subjected to local selection.     

Origin, fate, and architecture of ecologically relevant genetic variation

Recent advances in molecular genetics combined with field manipulations are yielding new insight into the origin, evolutionary fate, and genetic architecture of phenotypic variation in natural plant populations, with two surprising implications for the evolution of plant genomes. First, genetic loci exhibiting antagonistic pleiotropy across natural environments appear rare relative to loci that are adaptive in one or more environments and neutral elsewhere. These 'conditionally neutral' alleles should sweep to fixation when they arise, yet genome comparisons find little evidence for such selective sweeps. Second, genes under biotic selection tend to be of larger effect than genes under abiotic selection. Recent theory suggests this may be a consequence of high gene flow among populations under selection for local adaptation.     

Experimental evidence for ecological selection on genome variation in the wild

Understanding natural selection's effect on genetic variation is a major goal in biology, but the genome‐scale consequences of contemporary selection are not well known. In a release and recapture field experiment we transplanted stick insects to native and novel host plants and directly measured allele frequency changes within a generation at 186 576 genetic loci. We observed substantial, genome‐wide allele frequency changes during the experiment, most of which could be attributed to random mortality (genetic drift). However, we also documented that selection affected multiple genetic loci distributed across the genome, particularly in transplants to the novel host. Host‐associated selection affecting the genome acted on both a known colour‐pattern trait as well as other (unmeasured) phenotypes. We also found evidence that selection associated with elevation affected genome variation, although our experiment was not designed to test this. Our results illustrate how genomic data can identify previously underappreciated ecological sources and phenotypic targets of selection.     

The genetic consequences of spatially varying selection in the panmictic American eel (Anguilla rostrata)

Our understanding of the genetic basis of local adaptation has recently benefited from the increased power to identify functional variants associated with environmental variables at the genome scale. However, it often remains challenging to determine whether locally adaptive alleles are actively maintained at intermediate frequencies by spatially varying selection. Here, we evaluate the extent to which this particular type of balancing selection explains the retention of adaptive genetic variation in the extreme situation of perfect panmixia, using the American eel (Anguilla rostrata) as a model. We first conducted a genome scan between two samples from opposite ends of a latitudinal environmental gradient using 454 sequencing of individually tagged cDNA libraries. Candidate SNPs were then genotyped in 992 individuals from 16 sampling sites at different life stages of the same cohort (including larvae from the Sargasso Sea, glass eels, and 1-year-old individuals) as well as in glass eels of the following cohort. Evidence for spatially varying selection was found at 13 loci showing correlations between allele frequencies and environmental variables across the entire species range. Simulations under a multiple-niche Levene's model using estimated relative fitness values among genotypes rarely predicted a stable polymorphic equilibrium at these loci. Our results suggest that some genetic-by-environment interactions detected in our study arise during the progress toward fixation of a globally advantageous allele with spatially variable effects on fitness.     

The adaptive genomic landscape of beak morphology in Darwin's finches

Beak shape in Darwin's ground finches (Geospiza) is emblematic of natural selection and adaptive radiation, yet our understanding of the genetic basis of beak shape variation, and thus the genetic target of natural selection, is still evolving. Here we reveal the genomic architecture of beak shape variation using genomewide comparisons of four closely related and hybridizing species across 13 islands subject to parallel natural selection. Pairwise contrasts among species were used to identify a large number of genomic loci that are consistently related to species differences across a complex landscape. These loci are associated with hundreds of genes that have enriched GO categories significantly associated with development. One genomic region of particular interest is a section of Chromosome 1A with many candidate genes and increased linkage. The distinct, pointed beak shape of the cactus finch is linked to an excess of intermediate frequency alleles and increased heterozygosity in significant SNPs, but not across the rest of the genome. Alleles associated with pointier beaks among species were associated with pointier‐beaked populations within each species, thus establishing a common basis for natural selection, species divergence and adaptive radiation. The adaptive genomic landscape for Darwin's finches mirrors theoretical expectations based on morphological variation. The implication that a large number of genes are actively maintained to facilitate beak variation across parallel populations with documented interspecies admixture challenges our understanding of evolutionary processes in the wild.     

Patterns of Population Differentiation and Natural Selection on the Celiac Disease Background Risk Network

Celiac disease is a common small intestinal inflammatory condition induced by wheat gluten and related proteins from rye and barley. Left untreated, the clinical presentation of CD can include failure to thrive, malnutrition, and distension in juveniles. The disease can additionally lead to vitamin deficiencies, anemia, and osteoporosis. Therefore, CD potentially negatively affected fitness in past populations utilizing wheat, barley, and rye. Previous analyses of CD risk variants have uncovered evidence for positive selection on some of these loci. These studies also suggest the possibility that risk for common autoimmune conditions such as CD may be the result of positive selection on immune related loci in the genome to fight infection. Under this evolutionary scenario, disease phenotypes may be a trade-off from positive selection on immunity. If this hypothesis is generally true, we can expect to find a signal of natural selection when we survey across the network of loci known to influence CD risk. This study examines the non-HLA autosomal network of gene loci associated with CD risk in Europe. We reject the null hypothesis of neutrality on this network of CD risk loci. Additionally, we can localize evidence of selection in time and space by adding information from the genome of the Tyrolean Iceman. While we can show significant differentiation between continental regions across the CD network, the pattern of evidence is not consistent with primarily recent (Holocene) selection across this network in Europe. Further localization of ancient selection on this network may illuminate the ecological pressures acting on the immune system during this critically interesting phase of our evolution.     

Explorative genome scan to detect candidate loci for adaptation along a gradient of altitude in the common frog (Rana temporaria)

Today, with the rapid development of population genomics, the genetic basis of adaptation can be unraveled directly at the genome level, without any prerequisites about the selectively advantageous genes or traits. For nonmodel species, it is now possible to screen many markers randomly scattered across the genome and to distinguish between the neutral genetic background and outlier loci displaying an atypical behavior (e.g., a higher differentiation between populations). This study investigated the genetic frame of adaptation to a gradient of altitude in the common frog (Rana temporaria) by means of a genome scan based on 392 amplified fragment length polymorphism markers. Using two outlier detection methods never applied to dominant data so far, we sought for loci with a genetic differentiation diverging from neutral expectations when comparing populations from different altitudes. All the detected loci were sorted out according to their most probable cause for outlier behavior and classified as false positives, outliers due to local effects, or outliers associated with altitude. Altogether, eight good candidate loci were identified as potentially involved in adaptation to altitude because they were picked out in several independent interaltitude comparisons. This result illustrated the potential of genome-wide surveys to reveal selection signatures along selection gradients, where the association between environmental variables and fitness-related traits may be complex and/or cryptic. In this article, we also underlined the need for confirmation of the selection footprints for the outlier loci. Finally, we provided some preliminary insights into the genetic basis of adaptation along an altitudinal cline in the common frog.     

Characterization of a divergent chromosome region in the willow warbler Phylloscopus trochilus using avian genomic resources

Genome scans have made it possible to find outlier markers thought to have been influenced by divergent selection in almost any wild population. However, the lack of genomic information in nonmodel species often makes it difficult to associate these markers with certain genes or chromosome regions. Furthermore, the extent of linkage disequilibrium (LD) in the genome will determine the density of markers required to identify the genes under selection. In this study, we investigated a chromosome region in the willow warbler Phylloscopus trochilus surrounding a single marker previously identified in a genome scan. We first located the marker in the assembled genome of another species, the zebra finch Taeniopygia guttata, and amplified surrounding sequences in Fennoscandian willow warblers. Within an investigated chromosome region of 7.3 Mb as mapped to the zebra finch genome, we observed elevated genetic differentiation between a southern and a northern population across a 2.5-Mb interval comprising numerous coding genes. Within the southern and northern populations, higher values of LD were mostly found between SNPs within the same locus, but extended across distantly situated loci when the analyses were restricted to sampling sites showing intermediate allele frequencies of southern and northern alleles. Our study shows that cross-species genome information is a useful resource to obtain candidate sequences adjacent to outlier markers in nonmodel species.     

Interpreting the genomic landscape of speciation: a road map for finding barriers to gene flow

Speciation, the evolution of reproductive isolation among populations, is continuous, complex, and involves multiple, interacting barriers. Until it is complete, the effects of this process vary along the genome and can lead to a heterogeneous genomic landscape with peaks and troughs of differentiation and divergence. When gene flow occurs during speciation, barriers restricting gene flow locally in the genome lead to patterns of heterogeneity. However, genomic heterogeneity can also be produced or modified by variation in factors such as background selection and selective sweeps, recombination and mutation rate variation, and heterogeneous gene density. Extracting the effects of gene flow, divergent selection and reproductive isolation from such modifying factors presents a major challenge to speciation genomics. We argue one of the principal aims of the field is to identify the barrier loci involved in limiting gene flow. We first summarize the expected signatures of selection at barrier loci, at the genomic regions linked to them and across the entire genome. We then discuss the modifying factors that complicate the interpretation of the observed genomic landscape. Finally, we end with a road map for future speciation research: a proposal for how to account for these modifying factors and to progress towards understanding the nature of barrier loci. Despite the difficulties of interpreting empirical data, we argue that the availability of promising technical and analytical methods will shed further light on the important roles that gene flow and divergent selection have in shaping the genomic landscape of speciation.     

EST‐SSR‐based landscape genetics of Pseudotaxus chienii,a tertiary relict conifer endemic to China

Pseudotaxus chienii, belonging to the monotypic genus Pseudotaxus (Taxaceae), is a relict conifer endemic to China. Its populations are usually small and patchily distributed, having a low capacity of natural regeneration. To gain a clearer understanding of how landscape variables affect the local adaptation of P. chienii, we applied EST‐SSR markers in conjunction with landscape genetics methods: (a) to examine the population genetic pattern and spatial genetic structure; (b) to perform genome scan and selection scan to identify outlier loci and the associated landscape variables; and (c) to model the ecological niche under climate change. As a result, P. chienii was found to have a moderate level of genetic variation and a high level of genetic differentiation. Its populations displayed a significant positive relationship between the genetic and geographical distance (i.e., “isolation by distance” pattern) and a strong fine‐scale spatial genetic structure within 2 km. A putatively adaptive locus EMS6 (functionally annotated to cellulose synthase A catalytic subunit 7) was identified, which was found significantly associated with soil Cu, K, and Pb content and the combined effects of temperature and precipitation. Moreover, P. chienii was predicted to experience significant range contractions in future climate change scenarios. Our results highlight the potential of specific soil metal content and climate variables as the driving force of adaptive genetic differentiation in P. chienii. The data would also be useful to develop a conservation action plan for P. chienii.     

Riverscape genomics of a threatened fish across a hydroclimatically heterogeneous river basin

Understanding how natural selection generates and maintains adaptive genetic diversity in heterogeneous environments is key to predicting the evolutionary response of populations to rapid environmental change. Detecting selection in complex spatial environments remains challenging, especially for threatened species where the effects of strong genetic drift may overwhelm signatures of selection. We carried out a basinwide riverscape genomic analysis in the threatened southern pygmy perch (Nannoperca australis), an ecological specialist with low dispersal potential. High‐resolution environmental data and 5162 high‐quality filtered SNPs were used to clarify spatial population structure and to assess footprints of selection associated with a steep hydroclimatic gradient and with human disturbance across the naturally and anthropogenically fragmented Murray–Darling Basin (Australia). Our approach included F_ST outlier tests to define neutral loci, and a combination of spatially explicit genotype–environment association analyses to identify candidate adaptive loci while controlling for the effects of landscape structure and shared population history. We found low levels of genetic diversity and strong neutral population structure consistent with expectations based on spatial stream hierarchy and life history. In contrast, variables related to precipitation and temperature appeared as the most important environmental surrogates for putatively adaptive genetic variation at both regional and local scales. Human disturbance also influenced the variation in candidate loci for adaptation, but only at a local scale. Our study contributes to understanding of adaptive evolution along naturally and anthropogenically fragmented ecosystems. It also offers a tangible example of the potential contributions of landscape genomics for informing in situ and ex situ conservation management of biodiversity.     

Evidence for evolutionary change associated with the recent range expansion of the British butterfly, Aricia agestis, in response to climate change

Poleward range expansions are widespread responses to recent climate change and are crucial for the future persistence of many species. However, evolutionary change in traits such as colonization history and habitat preference may also be necessary to track environmental change across a fragmented landscape. Understanding the likelihood and speed of such adaptive change is important in determining the rate of species extinction with ongoing climate change. We conducted an amplified fragment length polymorphism (AFLP)‐based genome scan across the recently expanded UK range of the Brown Argus butterfly, Aricia agestis, and used outlier‐based (DFDIST and BayeScan) and association‐based (Isolation‐By‐Adaptation) statistical approaches to identify signatures of evolutionary change associated with range expansion and habitat use. We present evidence for (i) limited effects of range expansion on population genetic structure and (ii) strong signatures of selection at approximately 5% AFLP loci associated with both the poleward range expansion of A. agestis and differences in habitat use across long‐established and recently colonized sites. Patterns of allele frequency variation at these candidate loci suggest that adaptation to new habitats at the range margin has involved selection on genetic variation in habitat use found across the long‐established part of the range. Our results suggest that evolutionary change is likely to affect species’ responses to climate change and that genetic variation in ecological traits across species’ distributions should be maximized to facilitate range shifts across a fragmented landscape, particularly in species that show strong associations with particular habitats.     

Divergent and linked selection shape patterns of genomic differentiation between European and North American Atlantic salmon (Salmo salar)

As populations diverge many processes can shape genomic patterns of differentiation. Regions of high differentiation can arise due to divergent selection acting on selected loci, genetic hitchhiking of nearby loci, or through repeated selection against deleterious alleles (linked background selection); this divergence may then be further elevated in regions of reduced recombination. Atlantic salmon (Salmo salar) from Europe and North America diverged >600,000 years ago and despite some evidence of secondary contact, the majority of genetic data indicate substantial divergence between lineages. This deep divergence with potential gene flow provides an opportunity to investigate the role of different mechanisms that shape the genomic landscape during early speciation. Here, using 184,295 single nucleotide polymorphisms (SNPs) and 80 populations, we investigate the genomic landscape of differentiation across the Atlantic Ocean with a focus on highly differentiated regions and the processes shaping them. We found evidence of high (mean F_ST = 0.26) and heterogeneous genomic differentiation between continents. Genomic regions associated with high trans‐Atlantic differentiation ranged in size from single loci (SNPs) within important genes to large regions (1–3 Mbp ) on four chromosomes (Ssa06, Ssa13, Ssa16 and Ssa19). These regions showed signatures consistent with selection, including high linkage disequilibrium, despite no significant reduction in recombination. Genes and functional enrichment of processes associated with differentiated regions may highlight continental differences in ocean navigation and parasite resistance. Our results provide insight into potential mechanisms underlying differences between continents, and evidence of near‐fixed and potentially adaptive trans‐Atlantic differences concurrent with a background of high genome‐wide differentiation supports subspecies designation in Atlantic salmon.