A population genomic analysis of species boundaries: neutral processes, adaptive divergence and introgression between two hybridizing plant species

Little is known about the nature of species boundaries between closely related plant species and about the extent of introgression as a consequence of hybridization upon secondary contact. To address these topics we analyzed genome-wide differentiation between two closely related Silene species, Silene latifolia and S. dioica , and assessed the strength of introgression in sympatry. More than 300 AFLP markers were genotyped in three allopatric and three sympatric populations of each species. Outlier analyses were performed separately for sympatric and allopatric populations. Both positive and negative outlier loci were found, indicating that divergent and balancing selection, respectively, have shaped patterns of divergence between the two species. Sympatric populations of the two species were found to be less differentiated genetically than allopatric populations, indicating that hybridization has led to gene introgression. We conclude that differentiation between S. latifolia and S. dioica has been shaped by a combination of introgression and selection. These results challenge the view that species differentiation is a genome-wide phenomenon, and instead support the idea that genomes can be porous and that species differentiation has a genic basis.     

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.     

物种形成过程中的分化基因组岛及其形成机制

理解物种形成机制是生态和进化领域的重要任务。得益于测序技术的快速发展, 越来越多研究发现分化种群(亚种、物种)间的基因组常呈现异质性分化景观, 存在分化基因组岛, 这被认为是基因流存在下的歧化选择引起的, 支持基因流存在下的成种假说。然而, 基因渐渗、祖先多态性的差异分选、连锁选择等其他进化过程也可导致分化基因组岛的形成。现有实证研究在解析分化基因组岛的形成机制时, 往往忽略了上述其他进化过程的作用。为此, 本文在辨析分化基因组岛相关概念的基础上, 总结了利用种群基因组数据鉴定分化基因组岛的方法, 对比了不同进化过程形成分化基因组岛的特征, 指出在区分不同机制时联用基因渐渗程度、绝对分化指数(d_XY)、相对节点深度(RND)、重组率等多个指标的必要性, 归纳了物种形成过程中分化基因组岛形成机制解析的研究思路, 并对未来在生殖隔离机制上的深入探索以及实证研究的整合分析等方面进行了展望。     

A rapidly evolving genomic toolkit for Drosophila heterochromatin

Heterochromatin is the enigmatic eukaryotic genome compartment found mostly at telomeres and centromeres. Conventional approaches to sequence assembly and genetic manipulation fail in this highly repetitive, gene-sparse, and recombinationally silent DNA. In contrast, genetic and molecular analyses of euchromatin-encoded proteins that bind, remodel, and propagate heterochromatin have revealed its vital role in numerous cellular and evolutionary processes. Utilizing the 12 sequenced Drosophila genomes, Levine et al¹ took a phylogenomic approach to discover new such protein “surrogates” of heterochromatin function and evolution. This paper reported over 20 new members of what was traditionally believed to be a small and static Heterochromatin Protein 1 (HP1) gene family. The newly identified HP1 proteins are structurally diverse, lineage-restricted, and expressed primarily in the male germline. The birth and death of HP1 genes follows a “revolving door” pattern, where new HP1s appear to replace old HP1s. Here, we address alternative evolutionary models that drive this constant innovation.     

Characterization of a genomic divergence island between black-and-yellow and gopher Sebastes rockfishes

Islands of high genomic divergence that contain genes of evolutionary significance may form between diverging species. The gopher rockfish, Sebastes carnatus, and black-and-yellow rockfish, S. chrysomelas, are sympatrically distributed temperate marine species inhabiting rocky reefs and kelp forests on the west coast of the United States. Prior studies documented low levels of genetic divergence between the two species, except at a single microsatellite locus that displayed high divergence, Sra.7-2. To better characterize genome wide divergence, we scored 25 additional microsatellite loci. Mean neutral divergence between species (F(ST) = 0.01) changed little from prior estimates. Sra.7-2 continued to be an extreme divergence outlier. Five novel microsatellites within ± 15 kb of Sra.7-2 were characterized. High divergence, consistently low diversity in S. chrysomelas, and linkage disequilibrium were detected at these loci, suggesting the influence of recent selection. However, coalescent modelling of divergence at neutral and Sra.7-2 regions showed that initial divergence at Sra.7-2 was ancient, likely predating divergence at neutral regions. It is therefore unlikely that Sra.7-2 divergence represents solely recent ecological divergence within one species and may represent the action of recurrent selection. Introgressive gene flow (2N(E) m) was much higher (>1) at neutral than Sra.7-2 regions (<1) despite evidence that two S. carnatus individuals have recently mixed ancestry at the Sra.7-2 region. The Sra.7-2 genomic region is likely one of several regions containing genes involved in initiating and maintaining species integrity. Completion of the final stages of speciation appears to be a slow and ongoing process for these species.     

A genomic view of introgression and hybrid speciation

Hybridization in plants and animals is more common and has more complex outcomes than previously realized. Genome-wide analyses of introgression in organisms ranging from oaks to sunflowers to fruit flies show that a substantial fraction of their genomes are permeable to alleles from related species. Hybridization can lead to rapid genomic changes, including chromosomal rearrangements, genome expansion, differential gene expression, and gene silencing, some of which are mediated by transposable elements. These genomic changes may lead to beneficial new phenotypes, and selection for fertility and ecological traits may in turn alter genome structure. Dramatic increases in the availability of genomic tools will produce a new understanding of the genetic nature of species and will resolve a century-old debate over the basis of hybrid vigor, while the natural recombinants found in hybrid zones will permit genetic mapping of species differences and reproductive barriers in nonmodel organisms.     

Molecular signatures of lineage‐specific adaptive evolution in a unique sea basin: the example of an anadromous goby Leucopsarion petersii

Climate changes on various time scales often shape genetic novelty and adaptive variation in many biotas. We explored molecular signatures of directional selection in populations of the ice goby Leucopsarion petersii inhabiting a unique sea basin, the Sea of Japan, where a wide variety of environments existed in the Pleistocene in relation to shifts in sea level by repeated glaciations. This species consisted of two historically allopatric lineages, the Japan Sea (JS) and Pacific Ocean (PO) lineages, and these have lived under contrasting marine environments that are expected to have imposed different selection regimes caused by past climatic and current oceanographic factors. We applied a limited genome‐scan approach using seven candidate genes for phenotypic differences between two lineages in combination with 100 anonymous microsatellite loci. Neuropeptide Y (NPY) gene, which is an important regulator of food intake and potent orexigenic agent, and three anonymous microsatellites were identified as robust outliers, that is, candidate loci potentially under directional selection, by multiple divergence‐ and diversity‐based outlier tests in comparisons focused on multiple populations of the JS vs. PO lineages. For these outlier loci, populations of the JS lineage had putative signals of selective sweeps. Additionally, real‐time quantitative PCR analysis using fish reared in a common environment showed a higher expression level for NPY gene in the JS lineage. Thus, this study succeeded in identifying candidate genomic regions under selection across populations of the JS lineage and provided evidence for lineage‐specific adaptive evolution in this unique sea basin.     

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.     

The neutral theory in the genomic era

A number of tests have been developed to detect positive selection at the molecular level. These tests are based on DNA polymorphism within and divergence between species. Applications of these tests have revealed a large collection of genes that have evolved under positive selection and some general insights into adaptive evolution. Recently, these tests have been applied on a genomic scale and have provided estimates of the frequency of adaptive substitutions and a critical test of the neutral theory.     

A simulation study on the performance of differentiation-based methods to detect selected loci using linked neutral markers

We investigated the performance of two of the most popular differentiation-based methods to detect loci under selection (dfdist/fdist and bayescan) in order to ascertain the average chromosome map distance between the detected outlier markers and the nearest loci under selection. We used a model of neutral markers genetically linked to selected loci (QTL) controlling a quantitative trait subject to divergent selection in two subpopulations connected by migration. The results are not particularly encouraging because for chromosome lengths above 0.5 morgan, at least 30% of outliers detected were positioned in chromosomes where QTL were absent, clearly denoting false positives. Outliers linked to QTL were on average closer to the nearest QTL than randomly chosen markers, but the methods showed a substantial uncertainty about the genetic association between markers and selected loci, as this association could be shown significantly only in a moderate number of replicates for most scenarios. At equal conditions, bayescan seemed to perform somewhat more efficiently than dfdist/fdist, with little difference between results for dominant and codominant markers.     

Dynamics of drift, gene flow, and selection during speciation in Silene

The mechanics of speciation with gene flow are still unclear. Disparity among genes in population differentiation (F(ST)) between diverging species is often interpreted as evidence for semipermeable species boundaries, with selection preventing "key" genes from introgressing despite ongoing gene flow. However, F(ST) can remain high before it reaches equilibrium between the lineage sorting of species divergence and the homogenizing effects of gene flow (via secondary contact). Thus, when interpreting F(ST), the dynamics of drift, gene flow, and selection need to be taken into account. We illustrate this view with a multigenic analyses of gene flow and selection in three closely related Silene species, S. latifolia, S. dioica, and S. diclinis. We report that although S. diclinis appears to have evolved in allopatry, isolation with (bidirectional) gene flow between S. latifolia and S. dioica is likely, perhaps as a result of parapatric speciation followed by more extensive sympatry. Interestingly, we detected the signatures of apparently independent instances of positive selection at the same locus in S. latifolia and S. dioica. Despite gene flow between the species, the adaptive alleles have not crossed the species boundary, suggesting that this gene has independently undergone species-specific (diversifying or parallel) selection.     

Parallel evolution? Microsatellite variation of recently isolated marine and freshwater three-spined stickleback

Variation at nine microsatellite loci, four of which are linked to phenotypic traits (spine length and lateral plate morphology) in Canadian three-spined stickleback Gasterosteus aculeatus , are used to test for selection on marine and freshwater three-spined stickleback morphs in Iceland. There are indications of strong selection on loci linked to dorsal spine length, providing another potential example of parallel divergence at the genomic level in three-spined stickleback.     

Evaluation of demographic history and neutral parameterization on the performance of FST outlier tests

F_ST outlier tests are a potentially powerful way to detect genetic loci under spatially divergent selection. Unfortunately, the extent to which these tests are robust to nonequilibrium demographic histories has been understudied. We developed a landscape genetics simulator to test the effects of isolation by distance (IBD) and range expansion on F_ST outlier methods. We evaluated the two most commonly used methods for the identification of F_ST outliers (FDIST2 and BayeScan, which assume samples are evolutionarily independent) and two recent methods (FLK and Bayenv2, which estimate and account for evolutionary nonindependence). Parameterization with a set of neutral loci (‘neutral parameterization’) always improved the performance of FLK and Bayenv2, while neutral parameterization caused FDIST2 to actually perform worse in the cases of IBD or range expansion. BayeScan was improved when the prior odds on neutrality was increased, regardless of the true odds in the data. On their best performance, however, the widely used methods had high false‐positive rates for IBD and range expansion and were outperformed by methods that accounted for evolutionary nonindependence. In addition, default settings in FDIST2 and BayeScan resulted in many false positives suggesting balancing selection. However, all methods did very well if a large set of neutral loci is available to create empirical P‐values. We conclude that in species that exhibit IBD or have undergone range expansion, many of the published F_ST outliers based on FDIST2 and BayeScan are probably false positives, but FLK and Bayenv2 show great promise for accurately identifying loci under spatially divergent selection.     

Linked selection,demography and the evolution of correlated genomic landscapes in birds and beyond

Selection has a deep impact on the distribution of genetic diversity and population differentiation along the genome (the genomic landscapes of diversity and differentiation), reducing diversity and elevating differentiation not only at the sites it targets, but also at linked neutral sites. Fuelled by the high‐throughput sequencing revolution, these genomic footprints of selection have been extensively exploited over the past decade with the aim to identify genomic regions involved in adaptation and speciation. However, while this research has shown that the genomic landscapes of diversity and differentiation are usually highly heterogeneous, it has also led to the increasing realization that this heterogeneity may evolve under processes other than adaptation or speciation. In particular, instead of being an effect of selective sweeps or barriers to gene flow, accentuated differentiation can evolve by any process reducing genetic diversity locally within the genome (Charlesworth, 1998 ), including purifying selection at linked sites (background selection). In particular, in genomic regions where recombination is infrequent, accentuated differentiation can evolve as a by‐product of diversity reductions unrelated to adaptation or speciation (Cruickshank & Hahn, 2014 ; Nachman & Payseur, 2012 ; Noor & Bennett, 2009 ). In such genomic regions, linkage extends over physically larger genome stretches, and selection affects a particularly high number of linked neutral sites. Even though the effects of selection on linked neutral diversity (linked selection) within populations are well documented (Cutter & Payseur, 2013 ), recent observations of diversity and differentiation landscapes that are highly correlated even among independent lineages suggest that the effects of long‐term linked selection may have a deeper impact on the evolution of the genomic landscapes of diversity and differentiation than previously anticipated. The study on Saxicola stonechats by Van Doren et al. ( 2017 ) reported in the current issue of Molecular Ecology lines in with a rapidly expanding body of evidence in this direction. Correlations of genomic landscapes extending from within stonechats to comparisons with Ficedula flycatchers add to recent insights into the timescales across which the effects of linked selection persist. Absent and inverted correlations of genomic landscapes in comparisons involving an island taxon, on the other hand, provide important empirical clues about the role of demographic constraints in the evolution of the genomic landscapes of diversity and differentiation.     

Theoretical models of the influence of genomic architecture on the dynamics of speciation

A long‐standing problem in evolutionary biology has been determining whether and how gradual, incremental changes at the gene level can account for rapid speciation and bursts of adaptive radiation. Using genome‐scale computer simulations, we extend previous theory showing how gradual adaptive change can generate nonlinear population transitions, resulting in the rapid formation of new, reproductively isolated species. We show that these transitions occur via a mechanism rooted in a basic property of biological heredity: the organization of genes in genomes. Genomic organization of genes facilitates two processes: (i) the build‐up of statistical associations among large numbers of genes and (ii) the action of divergent selection on persistent combinations of alleles. When a population has accumulated a critical amount of standing, divergently selected variation, the combination of these two processes allows many mutations of small effect to act synergistically and precipitously split one population into two discontinuous, reproductively isolated groups. Periods of allopatry, chromosomal linkage among loci, and large‐effect alleles can facilitate this process under some conditions, but are not required for it. Our results complement and extend existing theory on alternative stable states during population divergence, distinct phases of speciation and the rapid emergence of multilocus barriers to gene flow. The results are thus a step towards aligning population genomic theory with modern empirical studies.     

Correcting for bias in estimation of quantitative trait loci effects

Estimates of quantitative trait loci (QTL) effects derived from complete genome scans are biased, if no assumptions are made about the distribution of QTL effects. Bias should be reduced if estimates are derived by maximum likelihood, with the QTL effects sampled from a known distribution. The parameters of the distributions of QTL effects for nine economic traits in dairy cattle were estimated from a daughter design analysis of the Israeli Holstein population including 490 marker-by-sire contrasts. A separate gamma distribution was derived for each trait. Estimates for both the α and β parameters and their SE decreased as a function of heritability. The maximum likelihood estimates derived for the individual QTL effects using the gamma distributions for each trait were regressed relative to the least squares estimates, but the regression factor decreased as a function of the least squares estimate. On simulated data, the mean of least squares estimates for effects with nominal 1% significance was more than twice the simulated values, while the mean of the maximum likelihood estimates was slightly lower than the mean of the simulated values. The coefficient of determination for the maximum likelihood estimates was five-fold the corresponding value for the least squares estimates.     

Contrasting patterns of quantitative and neutral genetic variation in locally adapted populations of the natterjack toad, Bufo calamita

The relative importance of natural selection and genetic drift in determining patterns of phenotypic diversity observed in nature is still unclear. The natterjack toad (Bufo calamita) is one of a few amphibian species capable of breeding in saline ponds, even though water salinity represents a considerable stress for them. Results from two common-garden experiments showed a pattern of geographic variation in embryonic salinity tolerance among populations from either fresh or brackish environments, consistent with the hypothesis of local adaptation. Full-sib analysis showed increased variation in survival among sibships within population for all populations as osmotic stress was increased (broad-sense heritability increased as salinity raised). Nevertheless, toads native to the brackish water environment had the highest overall survival under brackish conditions. Levels of population genetic differentiation for salinity tolerance were higher than those of neutral genetic differentiation, the latter obtained through the analysis of eight microsatellite loci. Microsatellite markers also revealed little population differentiation, lack of an isolation-by-distance pattern, and moderate gene flow connecting the populations. Therefore, environmental stress tolerance appears to have evolved in absence of geographic isolation, and consequently we reject the null hypothesis of neutral differentiation.