A likelihood-based comparison of population histories in a parasitoid guild

Little is known about the stability of trophic relationships in complex natural communities over evolutionary timescales. Here, we use sequence data from 18 nuclear loci to reconstruct and compare the intraspecific histories of major Pleistocene refugial populations in the Middle East, the Balkans and Iberia in a guild of four Chalcid parasitoids (Cecidostiba fungosa, Cecidostiba semifascia, Hobbya stenonota and Mesopolobus amaenus) all attacking Cynipid oak galls. We develop a likelihood method to numerically estimate models of divergence between three populations from multilocus data. We investigate the power of this framework on simulated data, and—using triplet alignments of intronic loci—quantify the support for all possible divergence relationships between refugial populations in the four parasitoids. Although an East to West order of population divergence has highest support in all but one species, we cannot rule out alternative population tree topologies. Comparing the estimated times of population splits between species, we find that one species, M. amaenus, has a significantly older history than the rest of the guild and must have arrived in central Europe at least one glacial cycle prior to other guild members. This suggests that although all four species may share a common origin in the East, they expanded westwards into Europe at different times.     

Likelihood‐based inference of population history from low‐coverage de novo genome assemblies

Short‐read sequencing technologies have in principle made it feasible to draw detailed inferences about the recent history of any organism. In practice, however, this remains challenging due to the difficulty of genome assembly in most organisms and the lack of statistical methods powerful enough to discriminate between recent, nonequilibrium histories. We address both the assembly and inference challenges. We develop a bioinformatic pipeline for generating outgroup‐rooted alignments of orthologous sequence blocks from de novo low‐coverage short‐read data for a small number of genomes, and show how such sequence blocks can be used to fit explicit models of population divergence and admixture in a likelihood framework. To illustrate our approach, we reconstruct the Pleistocene history of an oak‐feeding insect (the oak gallwasp Biorhiza pallida), which, in common with many other taxa, was restricted during Pleistocene ice ages to a longitudinal series of southern refugia spanning the Western Palaearctic. Our analysis of sequence blocks sampled from a single genome from each of three major glacial refugia reveals support for an unexpected history dominated by recent admixture. Despite the fact that 80% of the genome is affected by admixture during the last glacial cycle, we are able to infer the deeper divergence history of these populations. These inferences are robust to variation in block length, mutation model and the sampling location of individual genomes within refugia. This combination of de novo assembly and numerical likelihood calculation provides a powerful framework for estimating recent population history that can be applied to any organism without the need for prior genetic resources.     

MAXIMUM-LIKELIHOOD ESTIMATION OF POPULATION DIVERGENCE TIMES AND POPULATION PHYLOGENY IN MODELS WITHOUT MUTATION

In this paper we present a method for estimating population divergence times by maximum likelihood in models without mutation. The maximum-likelihood estimator is compared to a commonly applied estimator based on Wright's F_ST statistic. Simulations suggest that the maximum-likelihood estimator is less biased and has a lower variance than the F_ST-based estimator. The maximum-likelihood estimator provides a statistical framework for the analysis of population history given genetic data. We demonstrate how maximum-likelihood estimates of the branching pattern of divergence of multiple populations may be obtained. We also describe how the method may be applied to test hypotheses such as whether populations have maintained equal population sizes. We illustrate the method by applying it to two previously published sets of human restriction fragment length polymorphism (RFLP) data.     

50,000 years of ice and seals: Impacts of the Last Glacial Maximum on Antarctic fur seals

Ice is one of the most important drivers of population dynamics in polar organisms, influencing the locations, sizes, and connectivity of populations. Antarctic fur seals, Arctocephalus gazella, are particularly interesting in this regard, as they are concomitantly reliant on both ice‐associated prey and ice‐free coastal breeding areas. We reconstructed the history of this species through the Last Glacial Maximum (LGM) using genomic sequence data from seals across their range. Population size trends and divergence events were investigated using continuous‐time size estimation analysis and divergence time estimation models. The combined results indicated that a panmictic population present prior to the LGM split into two small refugial populations during peak ice extent. Following ice decline, the western refugial population founded colonies at the South Shetlands, South Georgia, and Bouvetøya, while the eastern refugial population founded the colony on Iles Kerguelen. Postglacial population divergence times closely match geological estimates of when these coastal breeding areas became ice free. Given the predictions regarding continued future warming in polar oceans, these responses of Antarctic fur seals to past climate variation suggest it may be worthwhile giving conservation consideration to potential future breeding locations, such as areas further south along the Antarctic Peninsula, in addition to present colony areas.     

Low‐coverage genomic data resolve the population divergence and gene flow history of an Australian rain forest fig wasp

Population divergence and gene flow are key processes in evolution and ecology. Model‐based analysis of genome‐wide data sets allows discrimination between alternative scenarios for these processes even in nonmodel taxa. We used two complementary approaches (one based on the blockwise site frequency spectrum [bSFS], the second on the pairwise sequentially Markovian coalescent [PSMC]) to infer the divergence history of a fig wasp, Pleistodontes nigriventris. Pleistodontes nigriventris and its fig tree mutualist Ficus watkinsiana are restricted to rain forest patches along the eastern coast of Australia and are separated into The Northern population is to the north of the Southern populations by two dry forest corridors (the Burdekin and St. Lawrence Gaps). We generated whole genome sequence data for two haploid males per population and used the bSFS approach to infer the timing of divergence between northern and southern populations of P. nigriventris, and to discriminate between alternative isolation with migration (IM) and instantaneous admixture (ADM) models of postdivergence gene flow. Pleistodontes nigriventris has low genetic diversity (π = 0.0008), to our knowledge one of the lowest estimates reported for a sexually reproducing arthropod. We find strongest support for an ADM model in which the two populations diverged ca. 196 kya in the late Pleistocene, with almost 25% of northern lineages introduced from the south during an admixture event ca. 57 kya. This divergence history is highly concordant with individual population demographies inferred from each pair of haploid males using PSMC. Our analysis illustrates the inferences possible with genome‐level data for small population samples of tiny, nonmodel organisms and adds to a growing body of knowledge on the population structure of Australian rain forest taxa.     

Genetic signature of recent glaciation on populations of a near-shore marine fish species (Syngnathus leptorhynchus)

Continental glaciation has played a major role in shaping the present-day phylogeography of freshwater and terrestrial species in the Northern Hemisphere. Recent work suggests that coastal glaciation during ice ages may have also had a significant impact on marine species. The bay pipefish, Syngnathus leptorhynchus , is a near-shore Pacific coast fish species with an exceptionally wide latitudinal distribution, ranging from Bahia Santa Maria, Baja California to Prince William Sound, Alaska. Survey data indicate that S. leptorhynchus is experiencing a range expansion at the northern limit of its range, consistent with colonization from southern populations. The present study uses six novel microsatellite markers and mitochondrial DNA (mtDNA) sequence data to study the present-day population genetic structure of four coastal populations of S. leptorhynchus . Deficits in mtDNA and nuclear DNA diversity in northern populations from regions glaciated during the last glacial maximum (LGM) [ c . 18 000 years before present ( bp )] suggest that these populations were effected by glacial events. Direct estimates of population divergence times derived from both isolation and isolation-with-migration models of evolution are also consistent with a postglacial phylogenetic history of populations north of the LGM. Sequence data further indicate that a population at the southern end of the species range has been separated from the three northern populations since long before the last interglacial event ( c . 130 000 years bp ), suggesting that topographical features along the Pacific coast may maintain population separation in regions unimpacted by coastal glaciation.     

Phylogeography and paleodistribution models of a widespread birch (Betula platyphylla Suk.) across East Asia: Multiple refugia,multidirectional expansion,and heterogeneous genetic pattern

Widespread tree species cover large geographical areas and play important roles in various vegetation types. Understanding how these species responded to historical climatic changes is important for understanding community assembly mechanisms with evolutionary and conservation implications. However, the location of refugial areas and postglacial history of widespread trees in East Asia remain poorly known. We combined microsatellite data (63 populations, 1756 individuals) and ecological niche modeling to examine the range‐wide population diversity, genetic structure, and historical demography of a pioneer tree species, Asian white birch (Betula platyphylla Suk.) across East Asia. We found a north‐to‐south trend of declining genetic diversity and five clusters, corresponding to geographical regions. Different clusters were inferred to have diverged through Pleistocene climatic oscillations and have different expansion routes, leading to genetic admixture in some populations. Ecological niche models indicated that the distribution of B. platyphylla during the last glacial maximum still had a large latitude span with slight shifts toward southeast, and northern populations had more variable distribution ranges than those in the south during later climatic oscillations. Our results reflect the relatively stable distribution through the last glacial–interglacial cycles and recent multidirectional expansion of B. platyphylla, providing new hypotheses for the response pattern of widespread tree species to climate change. The gradual genetic pattern from northeast to southwest and alternative distribution dynamics possibly resulted from environmental differences caused by latitude and topographic heterogeneity.     

Pleistocene origin and population history of a neoendemic alpine butterfly

Alpine environments underwent dramatic transformation during glacial–interglacial cycles, with the consequence that geographical, ecological and demographic changes of alpine populations provided the opportunity for formation of neoendemic species. Several biogeographical models have been proposed to account for the unique history of alpine populations, with different expectations of genetic divergence and speciation. The expanding alpine archipelago model proposes that alpine populations expand spatially and demographically during glacial events, dispersing between mountain ranges. Under this model, alpine populations are unlikely to diverge in isolation due to substantial interpopulation gene flow. In contrast, the alpine archipelago refuge model proposes that gene flow during glacial phases is limited and populations expand demographically during interglacial phases, increasing genetic isolation and the likelihood of speciation. We assess these models by reconstructing the evolutionary history of Colias behrii, a morphologically and ecologically distinct alpine butterfly restricted to the California Sierra Nevada. C. behrii exhibits very low genetic diversity at mitochondrial and nuclear loci, limited population structure and evidence of population expansion. C. behrii and Rocky Mountain C. meadii share identical mitochondrial haplotypes, while in contrast, nuclear data indicate common ancestry between C. behrii and Cascades Range Colias pelidne. The conflict in gene genealogies may be a result of recent expansion in North American Colias, but an isolation with migration analysis indicates that genetic patterns in C. behrii might result from differential introgression following hybridization. Based on the timing of population expansion and gene flow between mountain ranges, the expanding alpine archipelago model is supported in C. behrii.     

Estimating population divergence time and phylogeny from single-nucleotide polymorphisms data with outgroup ascertainment bias

The inference of population divergence times and branching patterns is of fundamental importance in many population genetic analyses. Many methods have been developed for estimating population divergence times, and recently, there has been particular attention towards genome-wide single-nucleotide polymorphisms (SNP) data. However, most SNP data have been affected by an ascertainment bias caused by the SNP selection and discovery protocols. Here, we present a modification of an existing maximum likelihood method that will allow approximately unbiased inferences when ascertainment is based on a set of outgroup populations. We also present a method for estimating trees from the asymmetric dissimilarity measures arising from pairwise divergence time estimation in population genetics. We evaluate the methods by simulations and by applying them to a large SNP data set of seven East Asian populations.     

Estimating the phylogeny in mollusc Littorina saxatilis (Olivi) from enzyme data: methodological considerations

The evolutionary history of 19 populations of Littorina saxatilis (Olivi) was estimated by four different approaches. Three of these operate upon a population by population matrix of genetic distances: average linkage clustering, and two versions of the Fitch-Margoliash method. The fourth method was a maximum likelihood estimate based on differences in allele frequencies between populations. The study aims to assess how well each method estimates the phylogeny by including seven populations of the closely related species L. arcana Hannaford Ellis. The rationale behind this is that a good estimation technique should be able to separate these two monophyletic taxa.The results show that, by our criteria, the maximum likelihood method yields the best estimate and the unconstrained Fitch-Margoliash technique gives reasonable estimates. Both average-linkage clustering and the Fitch-Margoliash method with evolutionary clock perform less well. We argue that this is expected since both these techniques are based on probably unrealistic assumptions such as the overall rate of evolutionary divergence being homogeneous over phyletic lines.     

Population differentiation and evolution in the common guillemot Uria aalge

Common (Uria aalge) and Brünnich's guillemots (U. Iomvia) are colonial seabirds that nest in temperate to arctic oceans throughout the Northern hemisphere. They are very similar in the characteristics of ecology, demography and life history that are thought to determine the extent of differentiation among populations, yet geographic variation in morphology is notably greater in common guillemots. Despite evidence of strong natal philopatry, previous analyses of allozymes and the mitochondrial cytochrome b gene revealed little genetic differentiation among North Atlantic colonies of Brünnich's guillemots. To determine if the more extensive morphological variability in common guillemots reflects greater genetic variability, we sequenced part of the cytochrome b gene for 160 common guillemots from 10 colonies distributed throughout the Northern hemisphere. Genotype frequencies and phylogenetic relationships among genotypes both indicated that Atlantic and Pacific populations are genetically distinct. Genetic divergence among genotypes suggested that differentiation of these populations has resulted from separation by Pleistocene glaciers and the Bering Landbridge, as well as by currently unsuitable breeding habitat in the Arctic Ocean. Cytochrome b genotype frequencies also differed among Atlantic colonies, and appeared to define a cline similar to that described for morphological characters. Analyses of sequence variation suggested that this variation probably results from secondary contact between two refugial populations from the Pleistocene glaciations, rather than from isolation by distance or selection. In contrast, the Atlantic population of Brünnich's guillemots appears to have arisen through recent expansion of a single homogeneous refugial population.     

Molecular phylogeography of white‐lipped tree viper (Trimeresurus; Viperidae)

The white‐lipped tree viper (Trimeresurus albolabris) is one of the most common venomous snakes with medicine importance in South East Asia. To explore the genetic diversity, population structure and evolutionary history of Trimeresurus albolabris, we collected 98 samples from 27 localities covering its entire distribution. Two mitochondrial gene fragments (cyt‐b and ND‐4) and two nuclear genes (RAG‐1 and NT‐3) were sequenced and analysed. Bayesian inference and maximum‐likelihood methods were employed to reconstruct the phylogenetic relationships among populations based on the two mitochondrial fragments, and the median‐joining networks were depicted using nuclear genes. Divergence date and ancestral area were estimated, and the population demographic history was inferred. Both phylogenetic analyses consistently uncovered that Trimeresurus albolabris was monophyletics, with five geographically structured lineages. Divergence date and ancestral area estimation indicated that T. albolabris originated in northern Thailand and eastern Myanmar at c. 7.15 Ma. Population dynamics analyses showed the southern China lineage has experienced population expansion and contraction, but the others have not. Both the interglacial expansion and the highly heterogeneous habitats resulting from the uplift of the Plateau played a joint role in shaping the present distribution and population structure. The evolutionary history of T. albolabris can be explained by a pattern of two direction dispersal: first from North to South, and then from West to East.     

Population genomic analyses reveal possible drivers of population divergence

Recent advances in sequencing technology and efficiency enable new and improved methods to investigate how populations diverge and species evolve. Fungi have relatively small and simple genomes and can often be cultured in the laboratory. Fungal populations can thus be sequenced for a relatively low cost, which makes them ideal for population genomic analyses. In several recent population genomic studies, wild populations of fungal model organisms and human pathogens have been analysed, for example Neurospora crassa (Ellison et al. 2011 ), Saccharomyces uvarum (Almeida et al. 2014 ), Coccidioides spp. (Neafsey et al. 2010 ) and Cryptococcus gatti (Engelthaler et al. 2014 ). In this issue of Molecular Ecology, Branco et al. ( 2015 ) apply population genomic tools to understand population divergence and adaptation in a symbiotic (mycorrhizal) fungus. This study exemplifies the possibilities of diving deeper into the genomic features involved in population divergence and speciation, also for nonmodel organisms, and how molecular and analytical tools will improve our understanding of the patterns and mechanisms that underlie adaptation to habitats, population divergence and dispersal limitation of fungi.     

MITOCHONDRIAL GENE GENEALOGY AND GENE FLOW AMONG ISLAND AND MAINLAND POPULATIONS OF A SEDENTARY SONGBIRD,THE GREY-CROWNED BABBLER (POMATOSTOMUS TEMPORALIS)

Distinguishing between ongoing gene flow and purely historical association of populations can be difficult without data on times of population separation and effective population sizes. To help discriminate between these two scenarios, I examined mitochondrial DNA sequence diversity in three geographically close populations of the grey-crowned babbler (Pomatostomus temporalis) separated by water barriers of known age in the Northern Territory, Australia, using the polymerase chain reaction (PCR), direct sequencing, and genealogical methods of inference. PCR primers were designed to obtain sequences from region I, a highly variable segment of the control region. Sequence diversity in all populations was consistent with neutrality. In the population on Melville Island, a Pleistocene land-bridge island, sequence variability is as high as on the mainland and consists of two mitochondrial lineages differing by 2%. Phylogenetic analyses of the sequence variation observed among 44 individuals suggest that the number of times lineages in one population trace back to ancestors of a different population (between-population coalescent events) was too high to be compatible with a model of population divergence solely by drift since rising of the water barriers, implying instead recent or ongoing gene flow across water barriers. Similar estimates of F_st, the fraction of genetic diversity apportioned among populations, were obtained when calculated using the divergence times of alleles and when estimated from Nm values derived from trees and ranging from 0.29-0.55. Both the phylogenies and patterns of allelic divergence suggest that the population on Melville Island exchanges migrants with both continental populations, although statistical tests indicated that some alternative phylogenies implying restricted gene flow among the populations could not be discounted.     

Demographic inferences using short‐read genomic data in an approximate Bayesian computation framework: in silico evaluation of power,biases and proof of concept in Atlantic walrus

Approximate Bayesian computation (ABC) is a powerful tool for model‐based inference of demographic histories from large genetic data sets. For most organisms, its implementation has been hampered by the lack of sufficient genetic data. Genotyping‐by‐sequencing (GBS) provides cheap genome‐scale data to fill this gap, but its potential has not fully been exploited. Here, we explored power, precision and biases of a coalescent‐based ABC approach where GBS data were modelled with either a population mutation parameter (θ) or a fixed site (FS) approach, allowing single or several segregating sites per locus. With simulated data ranging from 500 to 50 000 loci, a variety of demographic models could be reliably inferred across a range of timescales and migration scenarios. Posterior estimates were informative with 1000 loci for migration and split time in simple population divergence models. In more complex models, posterior distributions were wide and almost reverted to the uninformative prior even with 50 000 loci. ABC parameter estimates, however, were generally more accurate than an alternative composite‐likelihood method. Bottleneck scenarios proved particularly difficult, and only recent bottlenecks without recovery could be reliably detected and dated. Notably, minor‐allele‐frequency filters – usual practice for GBS data – negatively affected nearly all estimates. With this in mind, we used a combination of FS and θ approaches on empirical GBS data generated from the Atlantic walrus (Odobenus rosmarus rosmarus), collectively providing support for a population split before the last glacial maximum followed by asymmetrical migration and a high Arctic bottleneck. Overall, this study evaluates the potential and limitations of GBS data in an ABC‐coalescence framework and proposes a best‐practice approach.     

HISTORICAL DEMOGRAPHY AND PRESENT DAY POPULATION STRUCTURE OF THE GREENFINCH,CARDUEUS CHLORIS—AN ANALYSIS OF mtDNA CONTROL-REGION SEQUENCES

Genetic variability within and among 10 geographically distinct populations of Greenfinches (Carduelis chloris) was assayed by directly sequencing a 637 BP part of the mtDNA control region from 194 individuals. Thirteen variable positions defined 18 haplotypes with a maximum sequence divergence of 0.8%. Haplotype (h = 0.28–0.77) and nucleotide (π = 0.058–0.17%) diversities within populations were low, and decreased with increasing latitude (h:r_s = –0.81; π: r_s = –0.89). The distribution of pairwise nucleotide differences fit better with expectations of a “sudden expansion” than of an “equilibrium” model, and the estimates of long term effective population sizes were considerably lower than current census estimates, especially in northern European samples. Selection is an unlikely cause of observed patterns because the distribution of variability conformed to expectations of neutral infinite alleles model and haplotype diversity across populations was positively correlated with heterozygosity (H_E) in nuclear genes (r_s = 0.74, P < 0.05). Hence, a recent bottleneck, followed by serial bottlenecking during the process of post-Pleistocene recolonization of northern Europe, together with recent population expansion provide a plausible explanation for the low genetic diversity in the north. Genetic distances among populations showed a clear pattern of isolation-by-distance, and 14% of the haplotypic variation was among populations, the rest being distributed among individuals within populations. In accordance with allozyme and morphological data, a hierarchical analysis of nucleotide diversity recognized southern European populations as distinct from northern European ones. However, the magnitude of divergence in mtDNA, allozymes and morphology were highly dissimilar (morphology > mtDNA > allozymes).     

Phylogenomics at the tips: inferring lineages and their demographic history in a tropical lizard,Carlia amax

High‐throughput sequencing approaches offer opportunities to better understand the evolutionary processes driving diversification, particularly in nonmodel organisms. In particular, the 100–1000's of loci that can now be sequenced are providing unprecedented power in population, speciation and phylogenetic studies. Here, we apply an exon capture approach to generate >99% complete sequence and SNP data across >2000 loci from a tropical skink, Carlia amax, and exploit these data to identify divergent lineages and infer their relationships and demographic histories. This is especially relevant to low‐dispersal tropical taxa that often have cryptic diversity and spatially dynamic histories. For C. amax, clustering of nuclear SNPs and coalescent‐based species delimitation analyses identify four divergent lineages, one fewer than predicted based on geographically coherent mtDNA clades (>9.4% sequence divergence). Three of these lineages are widespread and parapatric on the mainland, whereas the most divergent is restricted to islands off the northeast Northern Territory. Tests for population expansion reject an equilibrium isolation‐by‐distance model for two of the three widespread lineages and infer refugial expansion sources in the relatively mesic northeast Top End and northwest Kimberley. The latter is already recognized as a hotspot of endemism, but our results also suggest that a stronger focus on the northeast Top End, and adjacent islands is warranted. More generally, our results show how genome‐reduction methods such as exon capture can yield insights into the pattern and dynamics of biodiversity across complex landscapes with as yet poorly understood biogeographic history and how exon data can link between population and phylogenetic questions.     

Efficient,Noninvasive Genetic Sampling for Monitoring Reintroduced Wolves

ABSTRACT Traditional methods of monitoring gray wolves (Canis lupus) are expensive and invasive and require extensive efforts to capture individual animals. Noninvasive genetic sampling (NGS) is an alternative method that can provide data to answer management questions and complement already-existing methods. In a 2-year study, we tested this approach for Idaho gray wolves in areas of known high and low wolf density. To focus sampling efforts across a large study area and increase our chances of detecting reproductive packs, we visited 964 areas with landscape characteristics similar to known wolf rendezvous sites. We collected scat or hair samples from 20% of sites and identified 122 wolves, using 8–9 microsatellite loci. We used the minimum count of wolves to accurately detect known differences in wolf density. Maximum likelihood and Bayesian single-session population estimators performed similarly and accurately estimated the population size, compared with a radiotelemetry population estimate, in both years, and an average of 1.7 captures per individual were necessary for achieving accurate population estimates. Subsampling scenarios revealed that both scat and hair samples were important for achieving accurate population estimates, but visiting 75% and 50% of the sites still gave reasonable estimates and reduced costs. Our research provides managers with an efficient and accurate method for monitoring high-density and low-density wolf populations in remote areas.     

Endemic species may have complex histories: within‐refugium phylogeography of an endangered Iberian vole

Glacial refugia protected and promoted biodiversity during the Pleistocene, not only at a broader scale, but also for many endemics that contracted and expanded their ranges within refugial areas. Understanding the evolutionary history of refugial endemics is especially important in the case of endangered species to recognize the origins of their genetic structure and thus produce better informed conservation practices. The Iberian Peninsula is an important European glacial refugium, rich in endemics of conservation concern, including small mammals, such as the Cabrera vole (Microtus cabrerae). This near‐threatened rodent is characterized by an unusual suite of genetic, life history and ecological traits, being restricted to isolated geographic nuclei in fast‐disappearing Mediterranean subhumid herbaceous habitats. To reconstruct the evolutionary history of the Cabrera vole, we studied sequence variation at mitochondrial, autosomal and sex‐linked loci, using invasive and noninvasive samples. Despite low overall mitochondrial and nuclear nucleotide diversities, we observed two main well‐supported mitochondrial lineages, west and east. Phylogeographic modelling in the context of the Cabrera vole's detailed fossil record supports a demographic scenario of isolation of two populations during the Last Glacial Maximum from a single focus in the southern part of the Iberian Peninsula. In addition, our data suggest subsequent divergence within the east, and secondary contact and introgression of the expanding western population, during the late Holocene. This work emphasizes that refugial endemics may have a phylogeographic history as rich as that of more widespread species, and conservation of such endemics includes the preservation of that genetic legacy.     

Pleistocene genetic connectivity in a widespread,open‐habitat‐adapted mosquito in the Indo‐Oriental region

Aim The environmental effect of Pleistocene climatic change in the Indo‐Oriental region has resulted in allopatric fragmentation and the generation of diversity in forest‐associated species. The aim of this study was to determine the extent to which Pleistocene climatic change has resulted in the fragmentation and speciation of an open‐habitat‐adapted mosquito, Anopheles vagus s.l., across its range. Location Anopheles vagus s.l. was sampled across the Indo‐Oriental region. Methods We generated 116 mitochondrial cytochrome c oxidase subunit I (COI) and 121 nuclear internal transcribed spacer 2 (ITS2) DNA sequences from 18 populations. Relationships between mitochondrial haplotypes were reconstructed using minimum spanning networks, and population structure was examined using analyses of molecular variance. The population history, including lineage divergence times, population expansion and gene flow, was inferred using beast and the isolation with migration (IM) model. Results There was no evidence to support the presence of the endemic Philippines species, A. limosus; instead, Philippine populations were closely related to, and derived from, A. vagus on the eastern Southeast Asian mainland. The most distinct populations were those from Java and East Timor, which differed from all other populations by all individuals having a 4‐bp insertion in the ITS2 sequence. The corresponding mitochondrial haplotypes had an estimated divergence time of 2.6 Ma [95% confidence interval (CI) 1.9–3.6 Ma]. Haplotype networks and analysis of molecular variance for COI supported western (Sri Lanka, India and Myanmar) and eastern (Thailand, Singapore, Cambodia, Vietnam and the Philippines) population groupings. This grouping structure results from the divergence of an eastern and a western mitochondrial lineage, estimated to have occurred 0.37 Ma (95% CI 0.26–0.55 Ma). Subsequent migration from the east to the west (0.16 Ma) is inferred to have created an admixture zone in Myanmar and Thailand. Main conclusions With the possible exception of populations from Java and East Timor, A. vagus appears to be one widespread genetically diverse taxon across its extensive range. The abundance of grassland during long interglacial periods may have facilitated population connectivity and range expansion across the Oriental and western Australasian regions.