Drivers of population genetic differentiation in the wild: isolation by dispersal limitation,isolation by adaptation and isolation by colonization

Empirical population genetic studies have been dominated by a neutralist view, according to which gene flow and drift are the main forces driving population genetic structure in nature. The neutralist view in essence describes a process of isolation by dispersal limitation (IBDL) that generally leads to a pattern of isolation by distance (IBD). Recently, however, conceptual frameworks have been put forward that view local genetic adaptation as an important driver of population genetic structure. Isolation by adaptation (IBA) and monopolization (M) posit that gene flow among natural populations is reduced as a consequence of local genetic adaptation. IBA stresses that effective gene flow is reduced among habitats that show dissimilar ecological characteristics, leading to a pattern of isolation by environment. In monopolization, local genetic adaptation of initial colonizing genotypes results in a reduction in gene flow that fosters the persistence of founder effects. Here, we relate these different processes driving landscape genetic structure to patterns of IBD and isolation by environment (IBE). We propose a method to detect whether IBDL, IBA and M shape genetic differentiation in natural landscapes by studying patterns of variation at neutral and non‐neutral markers as well as at ecologically relevant traits. Finally, we reinterpret a representative number of studies from the recent literature by associating patterns to processes and identify patterns associated with local genetic adaptation to be as common as IBDL in structuring regional genetic variation of populations in the wild. Our results point to the importance of quantifying environmental gradients and incorporating ecology in the analysis of population genetics.     

History,geography and host use shape genomewide patterns of genetic variation in the redheaded pine sawfly (Neodiprion lecontei)

Divergent host use has long been suspected to drive population differentiation and speciation in plant‐feeding insects. Evaluating the contribution of divergent host use to genetic differentiation can be difficult, however, as dispersal limitation and population structure may also influence patterns of genetic variation. In this study, we use double‐digest restriction‐associated DNA (ddRAD) sequencing to test the hypothesis that divergent host use contributes to genetic differentiation among populations of the redheaded pine sawfly (Neodiprion lecontei), a widespread pest that uses multiple Pinus hosts throughout its range in eastern North America. Because this species has a broad range and specializes on host plants known to have migrated extensively during the Pleistocene, we first assess overall genetic structure using model‐based and model‐free clustering methods and identify three geographically distinct genetic clusters. Next, using a composite‐likelihood approach based on the site frequency spectrum and a novel strategy for maximizing the utility of linked RAD markers, we infer the population topology and date divergence to the Pleistocene. Based on existing knowledge of Pinus refugia, estimated demographic parameters and patterns of diversity among sawfly populations, we propose a Pleistocene divergence scenario for N. lecontei. Finally, using Mantel and partial Mantel tests, we identify a significant relationship between genetic distance and geography in all clusters, and between genetic distance and host use in two of three clusters. Overall, our results indicate that Pleistocene isolation, dispersal limitation and ecological divergence all contribute to genomewide differentiation in this species and support the hypothesis that host use is a common driver of population divergence in host‐specialized insects.     

Different processes shape the patterns of divergence in the nuclear and chloroplast genomes of a relict tree species in East Asia

Isolation by spatial distance (IBD), environment (IBE), and historical climatic instability (IBI) are three common processes assessed in phylogeographic and/or landscape genetic studies. However, the relative contributions of these three processes with respect to spatial genetic patterns have seldom been compared. Moreover, whether the relative contribution differs in different regions or when assessed using different genetic markers has rarely been reported. Lindera obtusiloba has been found to have two sister genetic clades of chloroplast (cpDNA) and nuclear microsatellite (nSSR), both of which show discontinuous distribution in northern and southern East Asia. In this study, we used the Mantel test and multiple matrix regression with randomization (MMRR) to determine the relative contributions of IBD, IBE, and IBI with respect to L. obtusiloba populations. Independent Mantel tests and MMRR calculations were conducted for two genetic data sets (cpDNA and nSSR) and for different regions (the overall species range, and northern and southern subregions of the range). We found a significant IBI pattern in nSSR divergence for all assessed regions, whereas no clear IBI pattern was detected with respect to cpDNA. In contrast, significant (or marginal) divergent IBD patterns were detected for cpDNA in all regions, whereas although a significant IBE was apparent with respect to the overall range, the effect was not detected in the two subregions. The differences identified in nSSR and cpDNA population divergence may be related to differences in the heredity and ploidy of the markers. Compared with the southern region, the northern region showed less significant correlation patterns, which may be related to the shorter population history and restricted population range. The findings of this study serve to illustrate that comparing between markers or regions can contribute to gaining a better understanding the population histories of different genomes or within different regions of a species' range.     

GENETIC ISOLATION BY ENVIRONMENT OR DISTANCE: WHICH PATTERN OF GENE FLOW IS MOST COMMON?

Gene flow among populations can enhance local adaptation if it introduces new genetic variants available for selection, but strong gene flow can also stall adaptation by swamping locally beneficial genes. These outcomes can depend on population size, genetic variation, and the environmental context. Gene flow patterns may align with geographic distance (IBD—isolation by distance), whereby immigration rates are inversely proportional to the distance between populations. Alternatively gene flow may follow patterns of isolation by environment (IBE), whereby gene flow rates are higher among similar environments. Finally, gene flow may be highest among dissimilar environments (counter‐gradient gene flow), the classic “gene‐swamping” scenario. Here we survey relevant studies to determine the prevalence of each pattern across environmental gradients. Of 70 studies, we found evidence of IBD in 20.0%, IBE in 37.1%, and both patterns in 37.1%. In addition, 10.0% of studies exhibited counter‐gradient gene flow. In total, 74.3% showed significant IBE patterns. This predominant IBE pattern of gene flow may have arisen directly through natural selection or reflect other adaptive and nonadaptive processes leading to nonrandom gene flow. It also precludes gene swamping as a widespread phenomenon. Implications for evolutionary processes and management under rapidly changing environments (e.g., climate change) are discussed.     

Isolation by distance,resistance and/or clusters? Lessons learned from a forest‐dwelling carnivore inhabiting a heterogeneous landscape

Landscape genetics provides a valuable framework to understand how landscape features influence gene flow and to disentangle the factors that lead to discrete and/or clinal population structure. Here, we attempt to differentiate between these processes in a forest‐dwelling small carnivore [European pine marten (Martes martes)]. Specifically, we used complementary analytical approaches to quantify the spatially explicit genetic structure and diversity and analyse patterns of gene flow for 140 individuals genotyped at 15 microsatellite loci. We first used spatially explicit and nonspatial Bayesian clustering algorithms to partition the sample into discrete clusters and evaluate hypotheses of ‘isolation by barriers’ (IBB). We further characterized the relationships between genetic distance and geographical (‘isolation by distance’, IBD) and ecological distances (‘isolation by resistance’, IBR) obtained from optimized landscape models. Using a reciprocal causal modelling approach, we competed the IBD, IBR and IBB hypotheses with each other to unravel factors driving population genetic structure. Additionally, we further assessed spatially explicit indices of genetic diversity using sGD across potentially overlapping genetic neighbourhoods that matched the inferred population structure. Our results revealed a complex spatial genetic cline that appears to be driven jointly by IBD and partial barriers to gene flow (IBB) associated with poor habitat and interspecific competition. Habitat loss and fragmentation, in synergy with past overharvesting and possible interspecific competition with sympatric stone marten (Martes foina), are likely the main factors responsible for the spatial genetic structure we observed. These results emphasize the need for a more thorough evaluation of discrete and clinal hypotheses governing gene flow in landscape genetic studies, and the potential influence of different limiting factors affecting genetic structure at different spatial scales.     

Habitat use,but not gene flow,is influenced by human activities in two ecotypes of Egyptian fruit bat (Rousettus aegyptiacus)

Understanding the ecological, behavioural and evolutionary response of organisms to changing environments is of primary importance in a human‐altered world. It is crucial to elucidate how human activities alter gene flow and what are the consequences for the genetic structure of a species. We studied two lineages of the Egyptian fruit bat (Rousettus aegyptiacus) throughout the contact zone between mesic and arid Ecozones in the Middle East to evaluate the species' response to the growing proportion of human‐altered habitats in the desert. We integrated population genetics, morphometrics and movement ecology to analyse population structure, morphological variation and habitat use from GPS‐ or radio‐tagged individuals from both desert and Mediterranean areas. We classified the spatial distribution and environmental stratification by describing physical–geographical conditions and land cover. We analysed this information to estimate patch occupancy and used an isolation‐by‐resistance approach to model gene flow patterns. Our results suggest that lineages from desert and Mediterranean habitats, despite their admixture, are isolated by environment and by adaptation supporting their classification as ecotypes. We found a positive effect of human‐altered habitats on patch occupancy and habitat use of fruit bats by increasing the availability of roosting and foraging areas. While this commensalism promotes the distribution of fruit bats throughout the Middle East, gene flow between colonies has not been altered by human activities. This discrepancy between habitat use and gene flow patterns may, therefore, be explained by the breeding system of the species and modifications of natal dispersal patterns.     

Spatial genetic structuring in a widespread wetland plant on a plateau: Effects of elevation-driven geographic isolation and environmental heterogeneity

1. Highlands are ideal research areas for improving our understanding of the influence of ecological factors on the diversity and spatial patterns of natural species. Elevation-driven physical and environmental isolation greatly affect the evolution of plants. The mechanisms and essential drivers underlying these processes may differ among research scales, habitats and landscapes. Wetlands are important elements of the Qinghai–Tibetan Plateau, which is the highest plateau in the world, and these habitats harbour high aquatic organismal diversity. However, how the environments shape the genetic variation and structure of hydrophilous plants is poorly understood.
2. Using microsatellite markers and a chloroplast fragment, we quantified the genetic diversity and spatial genetic pattern of Stuckenia filiformis, one of the most widespread aquatic plants on the plateau. The relative contributions of geography, climate and local conditions to intra- and interpopulation variation were estimated. The results showed that intrapopulation genetic variation of the plant is moderate to high and not constrained by high-altitude environments. Topographical isolation mainly contributes to the genetic structure of S. filiformis, as inferred by simple sequence repeats and chloroplast DNA data. Significant effects of environmental variables on the spatial genetic patterns of this freshwater species were also suggested by landscape genetic analysis.
3. Infrequent long-distance dispersal, sexual recruitment and annual growth are probably important for the maintenance and distribution of this variation. Our findings imply a combined effect of geography and elevation-driven environmental heterogeneity on the evolution of aquatic organisms in highlands.

     

Exploring seascape genetics and kinship in the reef sponge Stylissa carteri in the Red Sea

A main goal of population geneticists is to study patterns of gene flow to gain a better understanding of the population structure in a given organism. To date most efforts have been focused on studying gene flow at either broad scales to identify barriers to gene flow and isolation by distance or at fine spatial scales in order to gain inferences regarding reproduction and local dispersal. Few studies have measured connectivity at multiple spatial scales and have utilized novel tools to test the influence of both environment and geography on shaping gene flow in an organism. Here a seascape genetics approach was used to gain insight regarding geographic and ecological barriers to gene flow of a common reef sponge, Stylissa carteri in the Red Sea. Furthermore, a small‐scale (<1 km) analysis was also conducted to infer reproductive potential in this organism. At the broad scale, we found that sponge connectivity is not structured by geography alone, but rather, genetic isolation in the southern Red Sea correlates strongly with environmental heterogeneity. At the scale of a 50‐m transect, spatial autocorrelation analyses and estimates of full‐siblings revealed that there is no deviation from random mating. However, at slightly larger scales (100–200 m) encompassing multiple transects at a given site, a greater proportion of full‐siblings was found within sites versus among sites in a given location suggesting that mating and/or dispersal are constrained to some extent at this spatial scale. This study adds to the growing body of literature suggesting that environmental and ecological variables play a major role in the genetic structure of marine invertebrate populations.     

Evolutionary insights into Rhinolophus episcopus (Chiroptera,Rhinolophidae) in China: Isolation by distance,environment, or sensory system?

Evolutionary processes can be influenced by several factors, such as geographic isolation, environmental selection, and sensory variation. For most nocturnal bats, echolocation is the primary sensory system used to prey and communicate, and plays important roles in chiropteran diversification and evolution. Understanding the relative contribution of geography, the environment, and this sensory system to population genetic divergence can elucidate the processes involved in bat incipient speciation and evolution. In this study, we collected spatial and environmental information, echolocation calls, as well as the previously published genetic data (six microsatellite loci and the mitochondrial cytochrome b gene) of widely distributed Rhinolophus episcopus populations to test three hypotheses for nuclear and mitochondrial divergence (isolation by distance, isolation by environment, and isolation by sensory variation) and unveil the factors that drive intraspecific genetic differentiation. The moderate level of nuclear differentiation was correlated with geographic/spatial distance and acoustic variation, whereas the relatively high level of mitochondrial differentiation was mainly associated with acoustic divergence. No significant correlation was observed between genetic divergence and environmental variables. Among the three factors, acoustic divergence explained the highest percentage of both nuclear and mitochondrial divergence. Thus, our results indicate that sensory variation may have played important roles in driving population isolation early in bat speciation, which is consistent with the hypothesis of isolation by sensory variation. Our study emphasizes the need to consider more factors, especially sensory traits, and combine multiple statistical methods in landscape genetic studies to test their potential contributions to driving population divergence.     

Syntopic frogs reveal different patterns of interaction with the landscape: A comparative landscape genetic study of Pelophylax nigromaculatus and Fejervarya limnocharis from central China

Amphibians are often considered excellent environmental indicator species. Natural and man‐made landscape features are known to form effective genetic barriers to amphibian populations; however, amphibians with different characteristics may have different species–landscape interaction patterns. We conducted a comparative landscape genetic analysis of two closely related syntopic frog species from central China, Pelophylax nigromaculatus (PN) and Fejervarya limnocharis (FL). These two species differ in several key life history traits; PN has a larger body size and larger clutch size, and reaches sexual maturity later than FL. Microsatellite DNA data were collected and analyzed using conventional (F_ST, isolation by distance (IBD), AMOVA) and recently developed (Bayesian assignment test, isolation by resistance) landscape genetic methods. As predicted, a higher level of population structure in FL (F_ST′ = 0.401) than in PN (F_ST′ = 0.354) was detected, in addition to FL displaying strong IBD patterns (r = .861) unlike PN (r = .073). A general north–south break in FL populations was detected, consistent with the IBD pattern, while PN exhibited clustering of northern‐ and southern‐most populations, suggestive of altered dispersal patterns. Species‐specific resistant landscape features were also identified, with roads and land cover the main cause of resistance to FL, and elevation the main influence on PN. These different species–landscape interactions can be explained mostly by their life history traits, revealing that closely related and ecologically similar species have different responses to the same landscape features. Comparative landscape genetic studies are important in detecting such differences and refining generalizations about amphibians in monitoring environmental changes.     

Isolation by environment

The interactions between organisms and their environments can shape distributions of spatial genetic variation, resulting in patterns of isolation by environment (IBE) in which genetic and environmental distances are positively correlated, independent of geographic distance. IBE represents one of the most important patterns that results from the ways in which landscape heterogeneity influences gene flow and population connectivity, but it has only recently been examined in studies of ecological and landscape genetics. Nevertheless, the study of IBE presents valuable opportunities to investigate how spatial heterogeneity in ecological processes, agents of selection and environmental variables contributes to genetic divergence in nature. New and increasingly sophisticated studies of IBE in natural systems are poised to make significant contributions to our understanding of the role of ecology in genetic divergence and of modes of differentiation both within and between species. Here, we describe the underlying ecological processes that can generate patterns of IBE, examine its implications for a wide variety of disciplines and outline several areas of future research that can answer pressing questions about the ecological basis of genetic diversity.     

Environmental gradients predict the genetic population structure of a coral reef fish in the Red Sea

The relatively recent fields of terrestrial landscape and marine seascape genetics seek to identify the influence of biophysical habitat features on the spatial genetic structure of populations or individuals. Over the last few years, there has been accumulating evidence for the effect of environmental heterogeneity on patterns of gene flow and connectivity in marine systems. Here, we investigate the population genetic patterns of an anemonefish, Amphiprion bicinctus, along the Saudi Arabian coast of the Red Sea. We collected nearly one thousand samples from 19 locations, spanning approximately 1500 km, and genotyped them at 38 microsatellite loci. Patterns of gene flow appeared to follow a stepping‐stone model along the northern and central Red Sea, which was disrupted by a distinct genetic break at a latitude of approximately 19°N. The Red Sea is characterized by pronounced environmental gradients along its axis, roughly separating the northern and central from the southern basin. Using mean chlorophyll‐a concentrations as a proxy for this gradient, we ran tests of isolation by distance (IBD, R² = 0.52) and isolation by environment (IBE, R² = 0.64), as well as combined models using partial Mantel tests and multiple matrix regression with randomization (MMRR). We found that genetic structure across our sampling sites may be best explained by a combined model of IBD and IBE (Mantel: R² = 0.71, MMRR: R² = 0.86). Our results highlight the potential key role of environmental patchiness in shaping patterns of gene flow in species with pelagic larval dispersal. We support growing calls for the integration of biophysical habitat characteristics into future studies of population genetic structure.     

Widespread evidence for incipient ecological speciation: a meta‐analysis of isolation‐by‐ecology

Ecologically mediated selection has increasingly become recognised as an important driver of speciation. The correlation between neutral genetic differentiation and environmental or phenotypic divergence among populations, to which we collectively refer to as isolation‐by‐ecology (IBE), is an indicator of ecological speciation. In a meta‐analysis framework, we determined the strength and commonality of IBE in nature. On the basis of 106 studies, we calculated a mean effect size of IBE with and without controlling for spatial autocorrelation among populations. Effect sizes were 0.34 (95% CI 0.24–0.42) and 0.26 (95% CI 0.13–0.37), respectively, indicating that an average of 5% of the neutral genetic differentiation among populations was explained purely by ecological contrast. Importantly, spatial autocorrelation reduced IBE correlations for environmental variables, but not for phenotypes. Through simulation, we showed how the influence of isolation‐by‐distance and spatial autocorrelation of ecological variables can result in false positives or underestimated correlations if not accounted for in the IBE model. Collectively, this meta‐analysis showed that ecologically induced genetic divergence is pervasive across time‐scales and taxa, and largely independent of the choice of molecular marker. We discuss the importance of these results in the context of adaptation and ecological speciation and suggest future research avenues.     

Investigating population differentiation in a major African agricultural pest: evidence from geometric morphometrics and connectivity suggests high invasion potential

The distribution, spatial pattern and population dynamics of a species can be influenced by differences in the environment across its range. Spatial variation in climatic conditions can cause local populations to undergo disruptive selection and ultimately result in local adaptation. However, local adaptation can be constrained by gene flow and may favour resident individuals over migrants—both are factors critical to the assessment of invasion potential. The Natal fruit fly (Ceratitis rosa) is a major agricultural pest in Africa with a history of island invasions, although its range is largely restricted to south east Africa. Across Africa, C. rosa is genetically structured into two clusters (R1 and R2), with these clusters occurring sympatrically in the north of South Africa. The spatial distribution of these genotypic clusters remains unexamined despite their importance for understanding the pest's invasion potential. Here, C. rosa, sampled from 22 South African locations, were genotyped at 11 polymorphic microsatellite loci and assessed morphologically using geometric morphometric wing shape analyses to investigate patterns of population structure and determine connectedness of pest‐occupied sites. Our results show little to no intraspecific (population) differentiation, high population connectivity, high effective population sizes and only one morphological type (R2) within South Africa. The absence of the R1 morphotype at sites where it was previously found may be a consequence of differences in thermal niches of the two morphotypes. Overall, our results suggest high invasion potential of this species, that area‐wide pest management should be undertaken on a country‐wide scale, and that border control is critical to preventing further invasions.     

Genomics of the divergence continuum in an African plant biodiversity hotspot,I: drivers of population divergence in Restio capensis (Restionaceae)

Understanding the drivers of population divergence, speciation and species persistence is of great interest to molecular ecology, especially for species‐rich radiations inhabiting the world's biodiversity hotspots. The toolbox of population genomics holds great promise for addressing these key issues, especially if genomic data are analysed within a spatially and ecologically explicit context. We have studied the earliest stages of the divergence continuum in the Restionaceae, a species‐rich and ecologically important plant family of the Cape Floristic Region (CFR) of South Africa, using the widespread CFR endemic Restio capensis (L.) H.P. Linder & C.R. Hardy as an example. We studied diverging populations of this morphotaxon for plastid DNA sequences and >14 400 nuclear DNA polymorphisms from Restriction site Associated DNA (RAD) sequencing and analysed the results jointly with spatial, climatic and phytogeographic data, using a Bayesian generalized linear mixed modelling (GLMM) approach. The results indicate that population divergence across the extreme environmental mosaic of the CFR is mostly driven by isolation by environment (IBE) rather than isolation by distance (IBD) for both neutral and non‐neutral markers, consistent with genome hitchhiking or coupling effects during early stages of divergence. Mixed modelling of plastid DNA and single divergent outlier loci from a Bayesian genome scan confirmed the predominant role of climate and pointed to additional drivers of divergence, such as drift and ecological agents of selection captured by phytogeographic zones. Our study demonstrates the usefulness of population genomics for disentangling the effects of IBD and IBE along the divergence continuum often found in species radiations across heterogeneous ecological landscapes.     

Temperature and precipitation,but not geographic distance,explain genetic relatedness among populations in the perennial grass Festuca rubra

Aims Knowledge of genetic structure of natural populations and its determinants may provide key insights into the ability of species to adapt to novel environments. In many genetic studies, the effects of climate could not be disentangled from the effects of geographic proximity. We aimed to understand the effects of temperature and moisture on genetic diversity of populations and separate these effects from the effects of geographic distance. We also wanted to explore the patterns of distribution of genetic diversity in the system and assess the degree of clonality within the populations. We also checked for possible genome size variation in the system.     

The role of geography and ecology in shaping repeated patterns of morphological and genetic differentiation between European minnows (Phoxinus phoxinus) from the Pyrenees and the Alps

Neutral and selective processes can drive repeated patterns of evolution in different groups of populations experiencing similar ecological gradients. In this paper, we used a combination of nuclear and mitochondrial DNA markers, as well as geometric morphometrics, to investigate repeated patterns of morphological and genetic divergence of European minnows in two mountain ranges: the Pyrenees and the Alps. European minnows (Phoxinus phoxinus) are cyprinid fish inhabiting most freshwater bodies in Europe, including those in different mountain ranges that could act as major geographical barriers to gene flow. We explored patterns of P. phoxinus phenotypic and genetic diversification along a gradient of altitude common to the two mountain ranges, and tested for isolation by distance (IBD), isolation by environment (IBE) and isolation by adaptation (IBA). The results indicated that populations from the Pyrenees and the Alps belong to two well differentiated, reciprocally monophyletic mtDNA lineages. Substantial genetic differentiation due to geographical isolation within and between populations from the Pyrenees and the Alps was also found using rapidly evolving AFLPs markers (isolation by distance or IBD), as well as morphological differences between mountain ranges. Also, morphology varied strongly with elevation and so did genetic differentiation to a lower extent. Despite moderate evidence for IBE and IBA, and therefore of repeated evolution, substantial population heterogeneity was found at the genetic level, suggesting that selection and population specific genetic drift act in concert to affect genetic divergence.     

Effects of sample size,number of markers,and allelic richness on the detection of spatial genetic pattern

The influence of study design on the ability to detect the effects of landscape pattern on gene flow is one of the most pressing methodological gaps in landscape genetic research. To investigate the effect of study design on landscape genetics inference, we used a spatially‐explicit, individual‐based program to simulate gene flow in a spatially continuous population inhabiting a landscape with gradual spatial changes in resistance to movement. We simulated a wide range of combinations of number of loci, number of alleles per locus and number of individuals sampled from the population. We assessed how these three aspects of study design influenced the statistical power to successfully identify the generating process among competing hypotheses of isolation‐by‐distance, isolation‐by‐barrier, and isolation‐by‐landscape resistance using a causal modelling approach with partial Mantel tests. We modelled the statistical power to identify the generating process as a response surface for equilibrium and non‐equilibrium conditions after introduction of isolation‐by‐landscape resistance. All three variables (loci, alleles and sampled individuals) affect the power of causal modelling, but to different degrees. Stronger partial Mantel r correlations between landscape distances and genetic distances were found when more loci were used and when loci were more variable, which makes comparisons of effect size between studies difficult. Number of individuals did not affect the accuracy through mean equilibrium partial Mantel r, but larger samples decreased the uncertainty (increasing the precision) of equilibrium partial Mantel r estimates. We conclude that amplifying more (and more variable) loci is likely to increase the power of landscape genetic inferences more than increasing number of individuals.     

Incipient allochronic speciation by climatic disruption of the reproductive period

Disruptive selection of life-cycle timing may cause temporal isolation directly and, ultimately, allochronic speciation. Despite the fact that segregation of the reproductive period among related species has been broadly observed across taxa, it remains controversial whether temporal isolation can function as the primary process of speciation. In the Japanese winter geometrid moth Inurois punctigera, allochronic divergence has resulted from climatic disruption of the reproductive period. In habitats with severe midwinter, two sympatric groups of moth reproduce allochronically in early and late winter. These groups are genetically diverging sister lineages and now co-occur allochronically throughout Japan. By contrast, in habitats with milder midwinter these lineages form a continuous adult period and gene flow has been facilitated between the lineages. These results, together with the fact that there is no difference in larval host use, indicate that temporal isolation has been the sole mechanism for allochronic isolation in colder habitats and that allochrony is not a by-product of other adaptations. Thus, the allochronic divergence of sympatric I. punctigera populations represents an incipient speciation process driven by midwinter disruption of the reproductive period.