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Populus tremuloides is the widest‐ranging tree species in North America and an ecologically important component of mesic forest ecosystems displaced by the Pleistocene glaciations. Using phylogeographic analyses of genome‐wide SNPs (34,796 SNPs, 183 individuals) and ecological niche modeling, we inferred population structure, ploidy levels, admixture, and Pleistocene range dynamics of P. tremuloides, and tested several historical biogeographical hypotheses. We found three genetic lineages located mainly in coastal–Cascades (cluster 1), east‐slope Cascades–Sierra Nevadas–Northern Rockies (cluster 2), and U.S. Rocky Mountains through southern Canadian (cluster 3) regions of the P. tremuloides range, with tree graph relationships of the form ((cluster 1, cluster 2), cluster 3). Populations consisted mainly of diploids (86%) but also small numbers of triploids (12%) and tetraploids (1%), and ploidy did not adversely affect our genetic inferences. The main vector of admixture was from cluster 3 into cluster 2, with the admixture zone trending northwest through the Rocky Mountains along a recognized phenotypic cline (Utah to Idaho). Clusters 1 and 2 provided strong support for the “stable‐edge hypothesis” that unglaciated southwestern populations persisted in situ since the last glaciation. By contrast, despite a lack of clinal genetic variation, cluster 3 exhibited “trailing‐edge” dynamics from niche suitability predictions signifying complete northward postglacial expansion. Results were also consistent with the “inland dispersal hypothesis” predicting postglacial assembly of Pacific Northwestern forest ecosystems, but rejected the hypothesis that Pacific‐coastal populations were colonized during outburst flooding from glacial Lake Missoula. Overall, congruent patterns between our phylogeographic and ecological niche modeling results and fossil pollen data demonstrate complex mixtures of stable‐edge, refugial locations, and postglacial expansion within P. tremuloides. These findings confirm and refine previous genetic studies, while strongly supporting a distinct Pacific‐coastal genetic lineage of quaking aspen.

A lack of optimal gene combinations, as well as low levels of genetic diversity, is often associated with the formation of species range margins. Conservation efforts rely on predictive modelling using abiotic variables and assessments of genetic diversity to determine target species and populations for controlled breeding, germplasm conservation and assisted migration. Biotic factors such as interspecific competition and hybridization, however, are largely ignored, despite their prevalence across diverse taxa and their role as key evolutionary forces. Hybridization between species with well‐developed barriers to reproductive isolation often results in the production of offspring with lower fitness. Generation of novel allelic combinations through hybridization, however, can also generate positive fitness consequences. Despite this possibility, hybridization‐mediated introgression is often considered a threat to biodiversity as it can blur species boundaries. The contribution of hybridization towards increasing genetic diversity of populations at range margins has only recently gathered attention in conservation studies. We assessed the extent to which hybridization contributes towards range dynamics by tracking spatio‐temporal changes in the central location of a hybrid zone between two recently diverged species of pines: Pinus strobiformis and P. flexilis. By comparing geographic cline centre estimates for global admixture coefficient with morphological traits associated with reproductive output, we demonstrate a northward shift in the hybrid zone. Using a combination of spatially explicit, individual‐based simulations and linkage disequilibrium variance partitioning, we note a significant contribution of adaptive introgression towards this northward movement, despite the potential for differences in regional population size to aid hybrid zone movement. Overall, our study demonstrates that hybridization between recently diverged species can increase genetic diversity and generate novel allelic combinations. These novel combinations may allow range margin populations to track favourable climatic conditions or facilitate adaptive evolution to ongoing and future climate change.

The extraordinary species diversity of the Neotropical freshwater fish fauna is world renown. Yet, despite rich species diversity, taxonomic and genetic resources for its Cerrado ichthyofauna remain poorly developed. We provide a reference library of 149 DNA barcodes for 39 species/lineages of Cerrado headwater stream fishes from the Brazilian Distrito Federal and nearby areas and test the utility of distance-based criteria, tree-based criteria and minibarcodes for specimen identification. Mean Kimura 2-parameter genetic distances within species to orders ranged 1·8-12·1%. However, mean intraspecific v. congeneric-interspecific distances (0·9-1·3%) overlapped extensively and distance-based barcoding failed to achieve correct identifications due to c. 4-12·1% error rates and 19·5% ambiguous identifications related to the presence of singletons. Overlap was reduced and best-match success rates improved drastically to 83·5% when Characidium barcodes representing potential misidentifications or undescribed species were removed. Tree-based monophyly criteria generally performed similarly to distance methods, correctly differentiating up to c. 85% of species/lineages despite neighbour-joining and Bayesian tree errors (random lineage-branching events, long-branch attraction). Five clusters (Ancistrus aguaboensis, Characidium spp., Eigenmannia trilineata, Hasemania hanseni and Hypostomus sp. 2) exhibited deep intraspecific divergences or para-/polyphyly and multiple Barcode Index Number assignments indicative of putative candidate species needing taxonomic re-examination. Sliding-window analyses also indicated that a 200 bp minibarcode region performed just as well at specimen identification as the entire barcode gene. Future DNA barcoding studies of Distrito Federal-Cerrado freshwater fishes will benefit from increased sampling coverage, as well as consideration of minibarcode targets for degraded samples and next-generation sequencing.

Anoura Gray, 1838 are Neotropical nectarivorous bats and the most speciose genus within the phyllostomid subfamily Glossophaginae. However, Anoura species limits remain debated, and phylogenetic relationships remain poorly known, because previous studies used limited Anoura taxon sampling or focused primarily on higher-level relationships. Here, we conduct the first phylogenomic study of Anoura by analyzing 2039 genome-wide ultraconserved elements (UCEs) sequenced for 42 individuals from 8 Anoura species/lineages plus two outgroups. Overall, our results based on UCEs resolved relationships in the genus and supported (1) the monophyly of small-bodied Anoura species (previously genus Lonchoglossa); (2) monotypic status of A. caudifer; and (3) nested positions of “A. carishina”, A. caudifer aequatoris, and A. geoffroyi peruana specimens within A. latidens, A. caudifer and A. geoffroyi, respectively (suggesting that these taxa are not distinct species). Additionally, (4) phylogenetic networks allowing reticulate edges did not explain gene tree discordance better than the species tree (without introgression), indicating that a coalescent model accounting for discordance solely through incomplete lineage sorting fit our data well. Sensitivity analyses indicated that our species tree results were not adversely affected by varying taxon sampling across loci. Tree calibration and Bayesian coalescent analyses dated the onset of diversification within Anoura to around ∼6–9 million years ago in the Miocene, with extant species diverging mainly within the past ∼4 million years. We inferred a historical biogeographical scenario for Anoura of parapatric speciation fragmenting the range of a wide-ranging ancestral lineage centered in the Central to Northern Andes, along with Pliocene–Pleistocene dispersal or founder event speciation in Amazonia and the Brazilian Atlantic forest during the last ∼2.5 million years.

Made available in DSpace on 2021-06-25T11:00:32Z (GMT). No. of bitstreams: 0 Previous issue date: 2021-01-01 This study describes two new endemic Hypostomus species from central Brazil, which were previously identified as genetically distinct lineages in a recent genomic study that recommended their testing and potential description based on morphological data. A machine learning classification procedure (random forest) was used to investigate morphological variation and identify putatively diagnostic characters for these candidate species and revealed that each is morphologically distinct. The new species Hypostomus cafuringa is characterized by small size, dark spots under a light background, deeper caudal peduncle and shorter first ray of the pectoral fin and base of the dorsal fin when compared to congeneric species from the region. H. cafuringa is known from the headwaters of the Maranhão River, upper Tocantins River basin, Distrito Federal, Brazil. The second new species, Hypostomus crulsi, is characterized by dark spots under a light background, absence of plates along the abdomen region, shorter first ray of the pelvic fin, shorter first ray of the pectoral fin and smaller body size. H. crulsi is known from the headwaters of the São Bartolomeu River, upper Paraná River basin, Distrito Federal, Brazil. The rapid conversion of natural habitats for agricultural development and the isolation of protected areas represent a serious threat to the continued existence of these two newly described endemic species, which warrant conservation assessment. Programa de Pós-Graduação em Ecologia Instituto de Ciências Biológicas Universidade de Brasília Departamento de Zoologia Instituto de Ciências Biológicas Universidade de Brasília Department of Biology Jacksonville State University Department of Biology Virginia Commonwealth University Departamento de Zoologia e Botânica Instituto de Biociências Letras e Ciências Exatas UNESP – Universidade Estadual Paulista Departamento de Zoologia e Botânica Instituto de Biociências Letras e Ciências Exatas UNESP – Universidade Estadual Paulista

River capture is a geological process of potentially great importance in shaping the genetic diversity, distributions, and community composition of freshwater taxa. Using phylogeographic analyses of ddRAD-seq data from suckermouth armored catfish (Hypostomussp. 2) populations, we tested for predicted genetic effects of headwater river capture events in central Brazil, previously supported by geological and community ecological data. We analyzed 227 ddRAD tags (3829 SNP loci) across 42 samples. Molecular results strongly supported sixHypostomusgenetic clusters/lineages, with the deepest divergence ∼1.25 million years ago in the early Pleistocene between a clade from the Upper Paraná and Upper São Francisco river basins versus all other lineages. Consistent with the ‘Paraná Capture Hypothesis’, several lines of evidence supported mid-Pleistocene colonization and vicariant isolation ofHypostomuspopulations from an ancestral Upper Paraná population, including: (1) significant phylogeographic structure, with predicted phylogenetic patterns, (2) higher Paraná lineage diversity, (3) ancestral geographic locations reconstructed in the Paraná basin, and (4) non-random interdrainage dispersal and vicariance events, indicating river captures primarily into the Tocantins and Upper São Francisco basinsc.∼220,000–145,500 years ago. Phylogeographic inference was complicated by lack of lineage monophyly across loci and lineages distributed in multiple basins, the latter of which lent support to the non-mutually exclusive ‘Frequent Interdrainage Dispersal Hypothesis’. However, species tree and demographic modeling results suggested these were artefacts of incomplete sorting of alleles in large ancestral populations over a geologically recent timeframe of divergence. Qualitative and quantitative sensitivity analyses demonstrated that our downstream genetic results were robust to effects of varying ddRAD-seq assembly parameters, which heavily influenced the number of output loci. We predict that codistributed freshwater taxa in Central Brazil may not exhibit phylogeographic patterns similar toHypostomussp. 2 due to complex patterns of superimposed river capture events, or if smaller ancestral population sizes have allowed more complete lineage sorting in other taxa.

Extant conifer species may be susceptible to rapid environmental change owing to their long generation times, but could also be resilient due to high levels of standing genetic diversity. Hybridisation between closely related species can increase genetic diversity and generate novel allelic combinations capable of fuelling adaptive evolution. Our study unravelled the genetic architecture of adaptive evolution in a conifer hybrid zone formed between Pinus strobiformis and P. flexilis. Using a multifaceted approach emphasising the spatial and environmental patterns of linkage disequilibrium and ancestry enrichment, we identified recently introgressed and background genetic variants to be driving adaptive evolution along different environmental gradients. Specifically, recently introgressed variants from P. flexilis were favoured along freeze-related environmental gradients, while background variants were favoured along water availability-related gradients. We posit that such mosaics of allelic variants within conifer hybrid zones will confer upon them greater resilience to ongoing and future environmental change and can be a key resource for conservation efforts., Mitra Menon et al. study the hybrid zone of two conifer species in New Mexico, and report that adaptation to cooler environments is associated with introgressed variants, whereas drier environments are associated with background genetic variants. These results may indicate how mosaics of allelic variants could show greater resilience to ongoing and future environmental change.

Populus tremuloides is the widest‐ranging tree species in North America and an ecologically important component of mesic forest ecosystems displaced by the Pleistocene glaciations. Using phylogeographic analyses of genome‐wide SNPs (34,796 SNPs, 183 individuals) and ecological niche modeling, we inferred population structure, ploidy levels, admixture, and Pleistocene range dynamics of P. tremuloides, and tested several historical biogeographical hypotheses. We found three genetic lineages located mainly in coastal–Cascades (cluster 1), east‐slope Cascades–Sierra Nevadas–Northern Rockies (cluster 2), and U.S. Rocky Mountains through southern Canadian (cluster 3) regions of the P. tremuloides range, with tree graph relationships of the form ((cluster 1, cluster 2), cluster 3). Populations consisted mainly of diploids (86%) but also small numbers of triploids (12%) and tetraploids (1%), and ploidy did not adversely affect our genetic inferences. The main vector of admixture was from cluster 3 into cluster 2, with the admixture zone trending northwest through the Rocky Mountains along a recognized phenotypic cline (Utah to Idaho). Clusters 1 and 2 provided strong support for the “stable‐edge hypothesis” that unglaciated southwestern populations persisted in situ since the last glaciation. By contrast, despite a lack of clinal genetic variation, cluster 3 exhibited “trailing‐edge” dynamics from niche suitability predictions signifying complete northward postglacial expansion. Results were also consistent with the “inland dispersal hypothesis” predicting postglacial assembly of Pacific Northwestern forest ecosystems, but rejected the hypothesis that Pacific‐coastal populations were colonized during outburst flooding from glacial Lake Missoula. Overall, congruent patterns between our phylogeographic and ecological niche modeling results and fossil pollen data demonstrate complex mixtures of stable‐edge, refugial locations, and postglacial expansion within P. tremuloides. These findings confirm and refine previous genetic studies, while strongly supporting a distinct Pacific‐coastal genetic lineage of quaking aspen.

We describe a new species of Enyalius endemic to the Brazilian Cerrado, based on morphological and molecular data sets. In the face of uncertain taxonomy among museum specimens of Enyalius, we used a novel analytical approach based on Gaussian mixture modeling for species assignments. We also used a machine-learning classification procedure (random forests) to investigate morphological variation and identify species diagnostic characters. Phylogenetic and species delimitation analyses supported the distinction of the new species from its congeners. The new species is characterized by the fewest ventral scales and smallest snout–vent length in the genus. Moreover, we infer that this species diverged from its closest relative, E. bilineatus, in the late Miocene, presumably after colonization of Cerrado gallery forests by an Atlantic Forest ancestor, followed by ecological or geographical speciation linked to shrinkage or fragmentation of gallery forests associated with global cooling and increased aridity. Rapid conversion of natural habitats, the isolation of protected areas, and recent changes to the Brazilian Forest Code pose serious threats to the conservation of the new species described herein, and other gallery forest inhabitants.

Interactions between extrinsic factors, such as disruptive selection and intrinsic factors, such as genetic incompatibilities among loci, often contribute to the maintenance of species boundaries. The relative roles of these factors in the establishment of reproductive isolation can be examined using species pairs characterized by gene flow throughout their divergence history. We investigated the process of speciation and the maintenance of species boundaries between Pinus strobiformis and Pinus flexilis. Utilizing ecological niche modelling, demographic modelling and genomic cline analyses, we illustrated a divergence history with continuous gene flow. Our results supported an abundance of advanced generation hybrids and a lack of loci exhibiting steep transition in allele frequency across the hybrid zone. Additionally, we found evidence for climate-associated variation in the hybrid index and niche divergence between parental species and the hybrid zone. These results are consistent with extrinsic factors, such as climate, being an important isolating mechanism. A build-up of intrinsic incompatibilities and of coadapted gene complexes is also apparent, although these appear to be in the earliest stages of development. This supports previous work in coniferous species demonstrating the importance of extrinsic factors in facilitating speciation. Overall, our findings lend support to the hypothesis that varying strength and direction of selection pressures across the long lifespans of conifers, in combination with their other life history traits, delays the evolution of strong intrinsic incompatibilities.

Targeted sequence capture is a promising approach for large-scale phylogenomics. However, rapid evolutionary radiations pose significant challenges for phylogenetic inference (e.g. incomplete lineages sorting (ILS), phylogenetic noise), and the ability of targeted nuclear loci to resolve species trees despite such issues remains poorly studied. We test the utility of targeted sequence capture for inferring phylogenetic relationships in rapid, recent angiosperm radiations, focusing on Burmeistera bellflowers (Campanulaceae), which diversified into ~130 species over less than 3 million years. We compared phylogenies estimated from supercontig (exons plus flanking sequences), exon-only, and flanking-only datasets with 506–546 loci (~4.7 million bases) for 46 Burmeistera species/lineages and 10 outgroup taxa. Nuclear loci resolved backbone nodes and many congruent internal relationships with high support in concatenation and coalescent-based species tree analyses, and inferences were largely robust to effects of missing taxa and base composition biases. Nevertheless, species trees were incongruent between datasets, and gene trees exhibited remarkably high levels of conflict (~4–60% congruence, ~40–99% conflict) not simply driven by poor gene tree resolution. Higher gene tree heterogeneity at shorter branches suggests an important role of ILS, as expected for rapid radiations. Phylogenetic informativeness analyses also suggest this incongruence has resulted from low resolving power at short internal branches, consistent with ILS, and homoplasy at deeper nodes, with exons exhibiting much greater risk of incorrect topologies due to homoplasy than other datasets. Our findings suggest that targeted sequence capture is feasible for resolving rapid, recent angiosperm radiations, and that results based on supercontig alignments containing nuclear exons and flanking sequences have higher phylogenetic utility and accuracy than either alone. We use our results to make practical recommendations for future target capture-based studies of Burmeistera and other rapid angiosperm radiations, including that such studies should analyze supercontigs to maximize the phylogenetic information content of loci.

Most Neotropical frog and freshwater fish species sampled to date show phylogeographic breaks along the Pacific coast of the Isthmus of Panama, with lineages in Costa Rica and western Panama isolated from central Panama. We examine temporal patterns of diversification of taxa across this &lsquo, western Panama isthmus&rsquo, (WPI) break to test hypotheses about the origin of species geographical distributions and genetic structuring in this region. We tested for synchronous diversification of four codistributed frog taxon-pairs and three fish taxon-pairs sharing the WPI break using hierarchical approximate Bayesian computation with model averaging based on mitochondrial DNA sequences. We also estimated lineage divergence times using full-Bayesian models. Several of our results supported synchronous divergences within the frog and freshwater fish assemblages, however, Bayes factor support was equivocal for or against synchronous or asynchronous diversification. Nevertheless, we infer that frog populations were likely isolated by one or multiple Pliocene&ndash, Pleistocene events more recently than predicted by previous models, while fish genetic diversity was structured by Pleistocene events. By integrating our results with external information from geology and elevational sea level modeling, we discuss the implications of our findings for understanding the biogeographical scenario of the diversification of Panamanian frogs and fishes. Consistent with the &lsquo, Bermingham/Martin model&rsquo, (Molecular Ecology 1998, 7, 499&ndash, 517), we conclude that the regional fish assemblage was fractured by processes shaping isthmian landscapes during the Pleistocene glaciations, including drainage basin isolation during lowered sea levels.

Interactions between extrinsic factors, such as disruptive selection, and intrinsic factors, such as genetic incompatibilities among loci, can contribute to the maintenance of species boundaries. The relative roles of these factors in the establishment of reproductive isolation can be examined using species pairs characterized by gene flow throughout their divergence history. We investigated the process of speciation and the maintenance of species boundaries between Pinus strobiformis and P.flexilis. Utilizing ecological niche modeling, demographic modeling, and genomic cline analyses, we illustrated a history of divergence with continuous gene flow between these species. We found an abundance of advanced generation hybrids and a lack of loci exhibiting large allele frequency differences across the hybrid zone. Additionally, we found evidence for climate-associated variation in the hybrid index and niche divergence between parental species and the hybrid zone. Our results are consistent with extrinsic factors, such as climate, being an important isolating mechanism for these species. A buildup of intrinsic incompatibilities and of co-adapted gene complexes is also apparent in our results, although these appear to be in the earliest stages of development. This supports previous work in coniferous species demonstrating the importance of extrinsic factors in creating and enforcing species boundaries. Overall, we lend support to the hypothesis that varying strengths and directions of selection pressures across the long lifespans of conifers, in combination with their life history strategies, delay the evolution of strong intrinsic incompatibilities.

Made available in DSpace on 2018-11-26T17:40:21Z (GMT). No. of bitstreams: 0 Previous issue date: 2017-08-24 Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) NASA National Science Foundation Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Brigham Young University Fulton Supercomputing Lab Background: Estimating the variability in isolation times across co-distributed taxon pairs that may have experienced the same allopatric isolating mechanism is a core goal of comparative phylogeography. The use of hierarchical Approximate Bayesian Computation (ABC) and coalescent models to infer temporal dynamics of lineage co-diversification has been a contentious topic in recent years. Key issues that remain unresolved include the choice of an appropriate prior on the number of co-divergence events (psi), as well as the optimal strategies for data summarization. Methods: Through simulation-based cross validation we explore the impact of the strategy for sorting summary statistics and the choice of prior on psi on the estimation of co-divergence variability. We also introduce a new setting (beta) that can potentially improve estimation of psi by enforcing a minimal temporal difference between pulses of co-divergence. We apply this new method to three empirical datasets: one dataset each of co-distributed taxon pairs of Panamanian frogs and freshwater fishes, and a large set of Neotropical butterfly sister-taxon pairs. Results: We demonstrate that the choice of prior on. has little impact on inference, but that sorting summary statistics yields substantially more reliable estimates of co-divergence variability despite violations of assumptions about exchangeability. We find the implementation of beta improves estimation of psi, with improvement being most dramatic given larger numbers of taxon pairs. We find equivocal support for synchronous co-divergence for both of the Panamanian groups, but we find considerable support for asynchronous divergence among the Neotropical butterflies. Conclusions: Our simulation experiments demonstrate that using sorted summary statistics results in improved estimates of the variability in divergence times, whereas the choice of hyperprior on psi has negligible effect. Additionally, we demonstrate that estimating the number of pulses of co-divergence across co-distributed taxon-pairs is improved by applying a flexible buffering regime over divergence times. This improves the correlation between. and the true variability in isolation times and allows for more meaningful interpretation of this hyperparameter. This will allow for more accurate identification of the number of temporally distinct pulses of co-divergence that generated the diversification pattern of a given regional assemblage of sister-taxon-pairs. CUNY City Coll, Dept Biol, New York, NY 10031 USA CUNY, Grad Ctr, New York, NY 10016 USA Univ Brasilia, Dept Zool, BR-70910900 Brasilia, DF, Brazil Univ Estadual Paulista, IBiLCE, Dept Zool & Bot, BR-15054000 Sao Jose Do Rio Preto, SP, Brazil Univ Estadual Paulista, IBiLCE, Dept Zool & Bot, BR-15054000 Sao Jose Do Rio Preto, SP, Brazil FAPESP: 2013/50297-0 NASA: DOB 1343578 National Science Foundation: DEB-1253710 National Science Foundation: CNS-0958379 National Science Foundation: CNS-0855217 CNPq: 314724/2014-1

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Alegre, RS, Brazil Royal Museum Cent Africa, Tervuren, Belgium Univ Nacl Autonoma Mexico, Mexico City, DF, Mexico Univ Innsbruck, Inst Ecol, Innsbruck, Austria Hasselt Univ, Hasselt, Belgium Univ Fed Pernambuco, Recife, PE, Brazil Univ Fed Espirito Santo, Vitoria, ES, Brazil Univ Nacl Misiones, Felix De Azara, Argentina Albany Museum, Grahamstown, South Africa Natl Museum, Bloemfontein, South Africa Senckenberg Nat Hist Sammlungen, Dresden, Germany Univ Estadual Paulista, Botucatu, SP, Brazil Stephen F Austin State Univ, Nacogdoches, TX 75962 USA Landesmuseum, Leonding, Austria Univ Camerino, Camerino, Italy Univ Fed ABC, Sao Bernardo, Brazil Univ Estadual Londrina, Museu Zool, Londrina, Parana, Brazil Senckenberg Res Inst, Frankfurt, Germany Museo Civ Storia Nat, Carmagnola, Italy Polskiej Akad Nauk, Muzeum & Inst Zool, Warsaw, Poland Museo Civ Storia Nat, Genoa, Italy Russian Acad Sci, Inst Zool, St Petersburg, Russia Univ Estadual Paulista, Sao Vicente, Brazil Sorbonne Univ, ISYEB, 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We investigate phylogeographic patterns and delimit species boundaries within Amatitlania, a genus of Central American cichlid fishes. Phylogenetic analyses of mitochondrial DNA sequences from 318 individuals spanning the geographical ranges of all three currently recognized Amatitlania species strongly supported one major clade, with a relatively diverged subclade corresponding to A. kanna samples from eastern Costa Rica and Panama. Gene trees and networks revealed marked incongruences between phylogeographic structure and morpho-species taxonomy as a result of species-level polyphyly. Bayes factor comparisons of species delimitation models accounting for incomplete lineage sorting under the multispecies coalescent decisively supported the recognition of two distinct species within Amatitlania corresponding to Amatitlania nigrofasciata and A. kanna lineages. The only clearly genetically and morphologically diagnosable species was A. kanna. These results strongly suggest that incomplete lineage sorting provides the best explanation for the polyphyly of A. kanna, whereas the polyphyly of A. siquia is likely a result of an imperfect taxonomy. Additional insights from coalescent-dating, network, and historical demographic analyses suggested that the two species of Amatitlania diversified only since the early Pleistocene, and that A. nigrofasciata experienced population expansions from approximately 200 000 years ago in the mid-late Pleistocene onward. We discuss implications of our results for the taxonomy and evolutionary history of Amatitlania and, more broadly, of Central American freshwater fishes.

Accurately delimiting species is fundamentally important for understanding species diversity and distributions and devising effective strategies to conserve biodiversity. However, species delimitation is problematic in many taxa, including ‘non-adaptive radiations’ containing morphologically cryptic lineages. Fortunately, coalescent-based species delimitation methods hold promise for objectively estimating species limits in such radiations, using multilocus genetic data. Using coalescent-based approaches, we delimit species and infer evolutionary relationships in a morphologically conserved group of Central American freshwater fishes, the Poecilia sphenops species complex. Phylogenetic analyses of multiple genetic markers (sequences of two mitochondrial DNA genes and five nuclear loci) from 10/15 species and genetic lineages recognized in the group support the P. sphenops species complex as monophyletic with respect to outgroups, with eight mitochondrial ‘major-lineages’ diverged by ≥2% pairwise genetic distances. From general mixed Yule-coalescent models, we discovered (conservatively) 10 species within our concatenated mitochondrial DNA dataset, 9 of which were strongly supported by subsequent multilocus Bayesian species delimitation and species tree analyses. Results suggested species-level diversity is underestimated or overestimated by at least ~15% in different lineages in the complex. Nonparametric statistics and coalescent simulations indicate genealogical discordance among our gene tree results has mainly derived from interspecific hybridization in the nuclear genome. However, mitochondrial DNA show little evidence for introgression, and our species delimitation results appear robust to effects of this process. Overall, our findings support the utility of combining multiple lines of genetic evidence and broad phylogeographical sampling to discover and validate species using coalescent-based methods. Our study also highlights the importance of testing for hybridization versus incomplete lineage sorting, which aids inference of not only species limits but also evolutionary processes influencing genetic diversity.