Testing for shared biogeographic history in the lower Central American freshwater fish assemblage using comparative phylogeography: concerted,independent, or multiple evolutionary responses?

A central goal of comparative phylogeography is determining whether codistributed species experienced (1) concerted evolutionary responses to past geological and climatic events, indicated by congruent spatial and temporal patterns (“concerted‐response hypothesis”); (2) independent responses, indicated by spatial incongruence (“independent‐response hypothesis”); or (3) multiple responses (“multiple‐response hypothesis”), indicated by spatial congruence but temporal incongruence (“pseudocongruence”) or spatial and temporal incongruence (“pseudoincongruence”). We tested these competing hypotheses using DNA sequence data from three livebearing fish species codistributed in the Nicaraguan depression of Central America (Alfaro cultratus, Poecilia gillii, and Xenophallus umbratilis) that we predicted might display congruent responses due to co‐occurrence in identical freshwater drainages. Spatial analyses recovered different subdivisions of genetic structure for each species, despite shared finer‐scale breaks in northwestern Costa Rica (also supported by phylogenetic results). Isolation‐with‐migration models estimated incongruent timelines of among‐region divergences, with A. cultratus and Xenophallus populations diverging over Miocene–mid‐Pleistocene while P. gillii populations diverged over mid‐late Pleistocene. Approximate Bayesian computation also lent substantial support to multiple discrete divergences over a model of simultaneous divergence across shared spatial breaks (e.g., Bayes factor [B₁₀] = 4.303 for Ψ [no. of divergences] > 1 vs. Ψ = 1). Thus, the data support phylogeographic pseudoincongruence consistent with the multiple‐response hypothesis. Model comparisons also indicated incongruence in historical demography, for example, support for intraspecific late Pleistocene population growth was unique to P. gillii, despite evidence for finer‐scale population expansions in the other taxa. Empirical tests for phylogeographic congruence indicate that multiple evolutionary responses to historical events have shaped the population structure of freshwater species codistributed within the complex landscapes in/around the Nicaraguan depression. Recent community assembly through different routes (i.e., different past distributions or colonization routes), and intrinsic ecological differences among species, has likely contributed to the unique phylogeographical patterns displayed by these Neotropical fishes.     

Fine-scale phylogeographic congruence despite demographic incongruence in two low-mobility saproxylic springtails

Evolutionary trajectories of codistributed taxa with comparable ecological preferences and dispersal abilities may be similarly impacted by historical landscape-level processes. Species' responses to changes in a shared biogeographic landscape may be purely concerted, completely independent, or classified as falling within an intermediate part of the continuum bounded by these two extremes. With sufficient molecular data, temporal contrasts of congruence among taxa with respect to these responses can be made. Such contrasts provide insights into the relative influence of ancient versus more recent climatic (and other) impacts on genetic structuring. Using phylogenetic, allele frequency, and genotypic data from two low-mobility, rotting-log-adapted (saproxylic) springtail species (Collembola) from an isolated 100-km-long section of the Great Dividing Range in southeastern Australia, we tested the concerted-response hypothesis over three timescales. Tests of phylogeographic, demographic, and contemporary population-genetic congruence were performed using an integrative approach that draws on both direct (pattern-based) and indirect (scenario-based) analyses. Our data revealed a general pattern of broad-scale similarities in species' responses to the interaction between Pleistocene climatic cycles and landscape setting, overlaid with some species-specific differences on local geographic and more recent temporal scales. This general pattern of phylogeographic congruence was accompanied by evidence for contemporaneous demographic incongruence indicating that, even at relatively small spatial scales, biogeographic context can exert an overarching influence on genetic structuring.     

Congruence, consensus, and the comparative phylogeography of codistributed species in California

Comparative phylogeography has emerged as a means of understanding the spatial patterns of genetic divergence of codistributed species. However, researchers are often frustrated because of the lack of appropriate statistical tests to assess concordancy of multiple phylogeographic trees. We develop a method for testing congruence across multiple species and synthesizing the data into a regional supertree. Nine phylogeographic data sets of species with different life histories and ecologies were statistically compared using maximum agreement subtrees (MAST) and showed a high degree of concordancy. A supertree combining the different phylogeographic trees was then computed using matrix representation with parsimony, and the groups defined by this supertree were tested against climatic data to investigate a potential mechanism driving divergence. Our data suggest that species and genetic lineages in California are shaped by climatic regimes. The supertree method in combination with MAST represents a new approach to test congruence hypotheses and detect common geographic signals in comparative phylogeography.     

Pleistocene climatic changes drive diversification across a tropical savanna

Spatial responses of species to past climate change depend on both intrinsic traits (climatic niche breadth, dispersal rates) and the scale of climatic fluctuations across the landscape. New capabilities in generating and analysing population genomic data, along with spatial modelling, have unleashed our capacity to infer how past climate changes have shaped populations, and by extension, complex communities. Combining these approaches, we uncover lineage diversity across four codistributed lizards from the Australian Monsoonal Tropics and explore how varying climatic tolerances interact with regional climate history to generate common vs. disparate responses to late Pleistocene change. We find more divergent spatial structuring and temporal demographic responses in the drier Kimberley region compared to the more mesic and consistently suitable Top End. We hypothesize that, in general, the effects of species’ traits on sensitivity to climate fluctuation will be more evident in climatically marginal regions. If true, this points to the need in climatically marginal areas to craft more species‐(or trait)‐specific strategies for persistence under future climate change.     

Testing for genetic evidence of population expansion and contraction: an empirical analysis of microsatellite DNA variation using a hierarchical Bayesian model

The role of past climatic change in shaping the distributions of tropical rain forest vertebrates is central to long-standing hypotheses about the legacy of the Quaternary ice ages. One approach to testing such hypotheses is to use genetic data to infer the demographic history of codistributed species. Population genetic theory that relates the structure of allelic genealogies to historical changes in effective population size can be used to detect a past history of demographic expansion or contraction. The fruit bats Cynopterus sphinx and C. brachyotis (Chiroptera: Pteropodidae) exhibit markedly different distribution patterns across the Indomalayan region and therefore represent an exemplary species pair to use for such tests. The purpose of this study was to test alternative hypotheses about historical patterns of demographic expansion and contraction in C. sphinx and C. brachyotis using a coalescent-based analysis of microsatellite variation. Specifically, we used a hierarchical Bayesian model based on Markov chain Monte Carlo simulations to estimate the posterior distribution of genealogical and demographic parameters. The results revealed strong evidence for population contraction in both species. Evidence for a population contraction in C. brachyotis was expected on the basis of biogeographic considerations. However, similar evidence for population contraction in C. sphinx does not support the hypothesis that this species underwent a pronounced range expansion during the late Quaternary. Genetic evidence for population decline may reflect the consequences of habitat destruction on a more recent time scale.     

Consistent geographic structure among multiple nuclear sequences and cpDNA polymorphisms of Cardamine nipponica Franch. et Savat. (Brassicaceae)

Molecular phylogeography has inferred the history of differentiation between regions and/or among populations following the Pleistocene climatic oscillations, mostly based on the genetic structure of organelle DNA. However, such genetic structure only reflects the history of a single gene, and studies based on single-copy genes of nuclear DNA (nDNA) are required for phylogeography, although their efficiency remains unclear. To examine the utility of nDNA loci, the genetic structures of three genes from Cardamine nipponica, which is closely related to the model species Arabidopsis thaliana, were elucidated: the nDNA genes DET1, PHYA, PHYE, as well as chloroplast DNA (cpDNA). In 279 individuals collected from throughout the range of the species, strong genetic differentiation between northern and central Japan was found for all loci. This result suggested that populations in central Japan experienced a different history from those in northern Japan during the Pleistocene climatic oscillations. In addition, the evidence of refugia at the edges of the distribution, where the genetic structure was less influenced by colonization following range expansion, was shown for several loci. The specific genetic structure within the southernmost populations of northern Japan suggested that this region was also isolated during range expansion. Hence, the consistent history among loci and a more detailed history from several loci indicated that cpDNA can represent the history of vicariance and demonstrated the efficiency of single-copy nuclear genes in phylogeography.     

The molecular organization of the beta-globin complex of the deer mouse, Peromyscus maniculatus

Recombinant DNA clones have been isolated that contain 80 kb of the beta-globin complex from the deer mouse, Peromyscus maniculatus. Comparisons of this complex with that from the laboratory mouse, Mus domesticus (with an order 5'-Hbby, Hbb-bhO, Hbb-bhl, Hbb-bh2, Hbb-bh3, Hbb-bl, Hbb-b2 3') highlight organizational trends in the beta-globin complex since the two species diverged. Unlike other mammals studied thus far, the deer mouse possesses three adult genes. Partial sequence analysis indicates that each of the three adult genes is intact and hence may be functional. Hybridization of one of the two Mus pseudogenes, Hbb-bh3, to genomic blots from Peromyscus reveals that it has a homologous counterpart in Peromyscus. Homologous genes to the two gamma-like Mus genes, Hbb-bhO and Hbb-bhl, are also found in Peromyscus. The strong hybridization between the Hbb-bhl genes and significant nucleotide similarity between the Hbb-bhO genes suggest that both pairs are important for the ontogeny of these mice although no known product has been identified for the Hbb-bhO genes. The presence of Hbb-bhO and Hbb-bhl in Peromyscus suggests that the duplication that created this related gene set occurred before the two lineages diverged. A single gene for Hbb-y has been isolated from Peromyscus. The adult region in Peromyscus has undergone significant divergence from the same region in Mus, having three rather than two adult genes, the acquisition of at least 15 kb of extra DNA relative to Mus, and possibly the loss of the Hbb-bh2 pseudogene. The nonadult region of the complex, in contrast, contains the same set of genes apparently distributed over the same amount of DNA as in the Mus beta- globin complex. This observation suggests that the embryonic region of the complex is more evolutionarily stable than the adult region.      

Mixed Fortunes: Ancient Expansion and Recent Decline in Population Size of a Subtropical Montane Primate,the Arunachal Macaque Macaca munzala

Quaternary glacial oscillations are known to have caused population size fluctuations in many temperate species. Species from subtropical and tropical regions are, however, considerably less studied, despite representing most of the biodiversity hotspots in the world including many highly threatened by anthropogenic activities such as hunting. These regions, consequently, pose a significant knowledge gap in terms of how their fauna have typically responded to past climatic changes. We studied an endangered primate, the Arunachal macaque Macaca munzala, from the subtropical southern edge of the Tibetan plateau, a part of the Eastern Himalaya biodiversity hotspot, also known to be highly threatened due to rampant hunting. We employed a 534 bp-long mitochondrial DNA sequence and 22 autosomal microsatellite loci to investigate the factors that have potentially shaped the demographic history of the species. Analysing the genetic data with traditional statistical methods and advance Bayesian inferential approaches, we demonstrate a limited effect of past glacial fluctuations on the demographic history of the species before the last glacial maximum, approximately 20,000 years ago. This was, however, immediately followed by a significant population expansion possibly due to warmer climatic conditions, approximately 15,000 years ago. These changes may thus represent an apparent balance between that displayed by the relatively climatically stable tropics and those of the more severe, temperate environments of the past. This study also draws attention to the possibility that a cold-tolerant species like the Arunachal macaque, which could withstand historical climate fluctuations and grow once the climate became conducive, may actually be extremely vulnerable to anthropogenic exploitation, as is perhaps indicated by its Holocene ca. 30-fold population decline, approximately 3,500 years ago. Our study thus provides a quantitative appraisal of these demographically important events, emphasising the ability to potentially infer the occurrence of two separate historical events from contemporary genetic data.     

Phylogeography recapitulates topography: very fine-scale local endemism of a saproxylic 'giant' springtail at Tallaganda in the Great Dividing Range of south-east Australia

Comparative phylogeography can reveal processes and historical events that shape the biodiversity of species and communities. As part of a comparative research program, the phylogeography of a new, endemic Australian genus and species of log-dependent (saproxylic) collembola was investigated using mitochondrial sequences, allozymes and anonymous single-copy nuclear markers. We found the genetic structure of the species corresponds with five a priori microbiogeographical regions, with population subdivision at various depths owing to palaeoclimatic influences. Closely related mtDNA haplotypes are codistributed within a single region or occur in adjacent regions, nuclear allele frequencies are more similar among more proximate populations, and interpopulation migration is rare. Based on mtDNA divergence, a late Miocene-late Pliocene coalescence is likely. The present-day distribution of genetic diversity seems to have been impacted by three major climatic events: Pliocene cooling and drying (2.5-7 million years before present, Mybp), early Pleistocene wet-dry oscillations (c. 1.2 Mybp) and the more recent glacial-interglacial cycles that have characterized the latter part of the Quaternary (<0.4 Mybp).     

Pleistocene range dynamics in the eastern Greater Cape Floristic Region: A case study of the Little Karoo endemic Berkheya cuneata (Asteraceae)

The glacial–interglacial climate cycles of the Pleistocene played a significant role in dramatically altering species distributions across the globe. However, the climate of the Greater Cape Floristic Region is thought to have been decoupled from global fluctuations and the current Mediterranean climate remained relatively buffered during this period. Here we explore the roles of climate stability and the topographic complexity of the region on the range history of an endemic Little Karoo plant, Berkheya cuneata, using ensemble species distribution modelling and multi-locus phylogeography. The species distribution models projected onto downscaled climate simulation of the Last Glacial Maximum demonstrated a considerable range contraction and fragmentation into the western and eastern Little Karoo, separated by the Rooiberg inselberg. This population fragmentation is mirrored in the phylogeographic structuring of both chloroplast and nuclear DNA. These results suggest that sufficient climatic buffering coupled with regionally complex topography ensured the localised population persistence during Pleistocene climate cycles but these features have also promoted population vicariance in this, and likely other, Little Karoo lowland species.     

The persistence of Pliocene populations through the Pleistocene climatic cycles: evidence from the phylogeography of an Iberian lizard

Ancient climatic fluctuations have caused changes in the demography and distribution of many species. The genetic differentiation between populations of the same species and of sister species is often attributed largely to the more recent Pleistocene fluctuations. Recent interpretations, which implicate earlier episodes, have proved controversial. We address the timing of genetic divergence in the Iberian lizard Lacerta schreiberi by studying the phylogeography of the cytochrome b sequence. The species has a remarkable morphological uniformity, yet our evidence suggests that earlier events in the Pliocene initiated the main divergence between populations. This interpretation implies that the different populations survived through the Pleistocene in separate localities. This conclusion is robust to different molecular clock calibrations. The persistence of earlier differentiation through the Pleistocene has wide implications for our understanding of Pleistocene refugia in this species and, by extension, to the biogeography of the whole region.     

Extreme population subdivision throughout a continuous range: phylogeography of Batrachoseps attenuatus (Caudata: Plethodontidae) in western North America

Low-vagility species with deep evolutionary histories are key to our understanding of the biogeographical history of geologically complex areas, such as the west coast of North America. We present a detailed study of the phylogeography of the salamander Batrachoseps attenuatus (Caudata: Plethodontidae) using sequences of the mitochondrial gene cob from 178 individuals sampled from throughout the species' range. Sequences of three other mitochondrial genes (16S, cox1, nad4) and a nuclear gene (RAG-1) were used to investigate the deeper evolutionary history of the species. We found high levels of genetic diversity and deep divergences within a mostly continuous distribution, with five genetically well-differentiated and geographically structured mitochondrial DNA clades. Significant association between geographical and genetic distances within these clades suggests demographic stability, whereas Fu's FS tests suggest demographic expansions in three of them. Mantel tests identify two biogeographical barriers, the San Andreas Fault and the Sacramento-San Joaquin Delta, as important in the diversification of lineages. The timing of the main splitting events between intraspecific lineages was estimated by applying relaxed molecular clock methods combining several mutation rates and a fossil calibration. The earliest splitting events are old (Pliocene/Miocene), with more recent (Pleistocene) subdivisions in some clades. Disjunct populations distributed along the western foothills of the Sierra Nevada colonized this area relatively recently from a single refugium east of San Francisco Bay. The combination of fine-scale, comprehensive sampling with phylogenetic, historical demographic and hypothesis-based tests allowed delineation of a complex biogeographical scenario with general implications for the study of codistributed taxa.     

Mitochondrial DNA variation and reticulate evolution of the genus Abies

The sequences of three regions of mitochondrial DNA (mtDNA) of a total length of 5226 bp were used to study the phylogeography of the genus Abies. The mtDNA haplotype network, comprising 36 studied Abies taxa, consisted of two branches; the first represented all American species plus two Asian, and the second included the remaining Eurasian species. Within these clusters, the haplotypes formed nine major groups, generally corresponding to the clades of the previously obtained phylogeny based on chloroplast DNA (cpDNA), but the relationships of these groups were significantly different; species assignment to the particular mtDNA haplotype group was more in line with its geographical distribution. In addition, the mtDNA haplotype network contains cycles indicating the recombination. It is assumed that the incongruence of cpDNA and mtDNA phylogenies is caused by the introgression capture of alien mtDNA during species hybridization and thus contains information about past migrations. The cases of incongruence of mitochondrial and chloroplast DNA suggesting a migration of Abies between Asia and North America are discussed.     

Statistical phylogeography

While studies of phylogeography and speciation in the past have largely focused on the documentation or detection of significant patterns of population genetic structure, the emerging field of statistical phylogeography aims to infer the history and processes underlying that structure, and to provide objective, rather than ad hoc explanations. Methods for parameter estimation are now commonly used to make inferences about demographic past. Although these approaches are well developed statistically, they typically pay little attention to geographical history. In contrast, methods that seek to reconstruct phylogeographic history are able to consider many alternative geographical scenarios, but are primarily nonstatistical, making inferences about particular biological processes without explicit reference to stochastically derived expectations. We advocate the merging of these two traditions so that statistical phylogeographic methods can provide an accurate representation of the past, consider a diverse array of processes, and yet yield a statistical estimate of that history. We discuss various conceptual issues associated with statistical phylogeographic inferences, considering especially the stochasticity of population genetic processes and assessing the confidence of phylogeographic conclusions. To this end, we present some empirical examples that utilize a statistical phylogeographic approach, and then by contrasting results from a coalescent-based approach to those from Templeton's nested cladistic analysis (NCA), we illustrate the importance of assessing error. Because NCA does not assess error in its inferences about historical processes or contemporary gene flow, we performed a small-scale study using simulated data to examine how our conclusions might be affected by such unconsidered errors. NCA did not identify the processes used to simulate the data, confusing among deterministic processes and the stochastic sorting of gene lineages. There is as yet insufficient justification of NCA's ability to accurately infer or distinguish among alternative processes. We close with a discussion of some unresolved problems of current statistical phylogeographic methods to propose areas in need of future development.     

Landscape genetics indicate recently increased habitat fragmentation in African forest‐associated chafers

Today, indigenous forests cover less than 0.6% of South Africa's land surface and are highly fragmented. Most forest relicts are very small and typically occur in fire‐protected gorges along the eastern Great Escarpment. Yet, they hold a unique and valuable fauna with high endemism and ancient phylogenetic lineages, fostered by long‐term climatic stability and complex microclimates. Despite numerous studies on southern African vegetation cover, the current state of knowledge about the natural extension of indigenous forests is rather fragmentary. We use an integrated approach of population‐level phylogeography and climatic niche modeling of forest‐associated chafer species to assess connectivity and extent of forest habitats since the last glacial maximum. Current and past species distribution models ascertained potential fluctuations of forest distribution and supported a much wider potential current extension of forests based on climatic data. Considerable genetic admixture of mitochondrial and nuclear DNA among many populations and an increase in mean population mutation rate in Extended Bayesian Skyline Plots of all species indicated more extended or better connected forests in the recent past (<5 kya). Genetic isolation of certain populations, as revealed by population differentiation statistics (), as well as landscape connectivity statistics and habitat succession scenarios suggests considerable loss of habitat connectivity. As major anthropogenic influence is likely, conservational actions need to be considered.     

Are glacial refugia hotspots of speciation and cytonuclear discordances? Answers from the genomic phylogeography of Spanish common frogs

Subdivided Pleistocene glacial refugia, best known as “refugia within refugia”, provided opportunities for diverging populations to evolve into incipient species and/or to hybridize and merge following range shifts tracking the climatic fluctuations, potentially promoting extensive cytonuclear discordances and “ghost” mtDNA lineages. Here, we tested which of these opposing evolutionary outcomes prevails in northern Iberian areas hosting multiple historical refugia of common frogs (Rana cf. temporaria), based on a genomic phylogeography approach (mtDNA barcoding and RAD‐sequencing). We found evidence for both incipient speciation events and massive cytonuclear discordances. On the one hand, populations from northwestern Spain (Galicia and Asturias, assigned to the regional endemic R. parvipalmata), are deeply‐diverged at mitochondrial and nuclear genomes (~4 My of independent evolution), and barely admix with northeastern populations (assigned to R. temporaria sensu stricto) across a narrow hybrid zone (~25 km) located in the Cantabrian Mountains, suggesting that they represent distinct species. On the other hand, the most divergent mtDNA clade, widespread in Cantabria and the Basque country, shares its nuclear genome with other R. temporaria s. s. lineages. Patterns of population expansions and isolation‐by‐distance among these populations are consistent with past mitochondrial capture and/or drift in generating and maintaining this ghost mitochondrial lineage. This remarkable case study emphasizes the complex evolutionary history that shaped the present genetic diversity of refugial populations, and stresses the need to revisit their phylogeography by genomic approaches, in order to make informed taxonomic inferences.     

Comparative phylogeography of three endemic rodents from the Albertine Rift, east central Africa

The major aim of this study was to compare the phylogeographic patterns of codistributed rodents from the fragmented montane rainforests of the Albertine Rift region of east central Africa. We sampled individuals of three endemic rodent species, Hylomyscus denniae, Hybomys lunaris and Lophuromys woosnami from four localities in the Albertine Rift. We analysed mitochondrial DNA sequence variation from fragments of the cytochrome b and control region genes and found significant phylogeographic structuring for the three taxa examined. The recovered phylogenies suggest that climatic fluctuations and volcanic activity of the Virunga Volcanoes chain have caused the fragmentation of rainforest habitat during the past 2 million years. This fragmentation has played a major role in the diversification of the montane endemic rodents of the region. Estimation of the divergence times within each species suggests a separation of the major clades occurring during the mid to late Pleistocene.     

Past tree range dynamics in the Iberian Peninsula inferred through phylogeography and palaeodistribution modelling: A review

The forests in the Iberian Peninsula have been strongly influenced by past climatic changes, but reconstructing their historical distributions and dynamics is very difficult due to the complex climatic characteristics and relief of the region. Research disciplines such as phylogeography and species distribution modelling can describe the past range dynamics of individual tree species in relatively great detail and help elucidate how these species have reacted to climatic changes. Here we review phylogeographical and modelling studies from species representative of the major Iberian forest types and attempt to extract general trends from the diversity of individual species histories in the Peninsula. To date most studies focus on geographical range dynamics during the Pleistocene, but an increasing body of evidence shows that some species have also retained the genetic imprints of much more ancient processes. Many widespread Iberian species show a deep and often remarkably clear-cut divide between populations from the Mediterranean and from the Atlantic regions of the Iberian Peninsula, suggesting that both areas have independently sustained viable populations over extended periods. In fact, phylogeographical studies commonly find that species had several glacial refugia across the Iberian Peninsula. On the other hand, distribution models help identifying further suitable areas that could have sustained so far undetected refugia. Such studies are of interest for species conservation, because refugium populations are high-priority targets due to their long-term persistence and unique evolutionary trajectory. Overall, we conclude that palaeoecology, phylogeography and species distribution modelling have a great potential to inform each other because of their complementary perspectives and results. A true integration of these approaches is therefore fundamental for further progress in our understanding of past Iberian environments and the organisms they harboured.     

The roles of climate,geography and natural selection as drivers of genetic and phenotypic differentiation in a widespread amphibian Hyla annectans (Anura: Hylidae)

The role of geological events and Pleistocene climatic fluctuations as drivers of current patterns of genetic variation in extant species has been a topic of continued interest among evolutionary biologists. Nevertheless, comprehensive studies of widely distributed species are still rare, especially from Asia. Using geographically extensive sampling of many individuals and a large number of nuclear single nucleotide polymorphisms (SNPs), we studied the phylogeography and historical demography of Hyla annectans populations in southern China. Thirty‐five sampled populations were grouped into seven clearly defined genetic clusters that closely match phenotype‐based subspecies classification. These lineages diverged 2.32–5.23 million years ago (Ma), a timing that closely aligns with the rapid and drastic uplifting of the Qinghai‐Tibet Plateau and adjacent southwest China. Demographic analyses and species distribution models indicate that different populations of this species have responded differently to past climatic changes. In the Hengduan Mountains, most populations experienced a bottleneck, whereas the populations located outside of the Hengduan Mountains have gradually declined in size since the end of the last glaciation. In addition, the levels of phenotypic and genetic divergence were strongly correlated across major clades. These results highlight the combined effects of geological events and past climatic fluctuations, as well as natural selection, as drivers of contemporary patterns of genetic and phenotypic variation in a widely distributed anuran in Asia.