Central African dwarf crocodiles found in syntopy are comparably divergent to South American dwarf caimans.

Recent molecular taxonomic advancements have expanded our understanding of crocodylian diversity, revealing the existence of previously overlooked species, including the Congo dwarf crocodile (Osteolaemus osborni) in the central Congo Basin rainforests. This study explores the genomic divergence between O. osborni and its better-known relative, the true dwarf crocodile (Osteolaemus tetraspis), shedding light on their evolutionary history. Field research conducted in the northwestern Republic of the Congo uncovered a locality where both species coexist in sympatry/syntopy. Genomic analysis of sympatric individuals reveals a level of divergence comparable to that between ecologically similar South American dwarf caimans (Paleosuchus palpebrosus and Paleosuchus trigonatus), suggesting parallel speciation in the Afrotropics and Neotropics during the Middle to Late Miocene, 10-12 Ma. Comparison of the sympatric and allopatric dwarf crocodiles indicates no gene flow between the analysed sympatric individuals of O. osborni and O. tetraspis. However, a larger sample will be required to answer the question of whether or to what extent these species hybridize. This study emphasizes the need for further research on the biology and conservation status of the Congo dwarf crocodile, highlighting its significance in the unique biodiversity of the Congolian rainforests and thus its potential as a flagship species.

Chromosomal evolution, environmental heterogeneity, and migration drive spatial patterns of species richness in Calochortus (Liliaceae).

We used nuclear genomic data and statistical models to evaluate the ecological and evolutionary processes shaping spatial variation in species richness in Calochortus (Liliaceae, 74 spp.). Calochortus occupies diverse habitats in the western United States and Mexico and has a center of diversity in the California Floristic Province, marked by multiple orogenies, winter rainfall, and highly divergent climates and substrates (including serpentine). We used sequences of 294 low-copy nuclear loci to produce a time-calibrated phylogeny, estimate historical biogeography, and test hypotheses regarding drivers of present-day spatial patterns in species number. Speciation and species coexistence require reproductive isolation and ecological divergence, so we examined the roles of chromosome number, environmental heterogeneity, and migration in shaping local species richness. Six major clades-inhabiting different geographic/climatic areas, and often marked by different base chromosome numbers (n = 6 to 10)-began diverging from each other ~10.3 Mya. As predicted, local species number increased significantly with local heterogeneity in chromosome number, elevation, soil characteristics, and serpentine presence. Species richness is greatest in the Transverse/Peninsular Ranges where clades with different chromosome numbers overlap, topographic complexity provides diverse conditions over short distances, and several physiographic provinces meet allowing immigration by several clades. Recently diverged sister-species pairs generally have peri-patric distributions, and maximum geographic overlap between species increases over the first million years since divergence, suggesting that chromosomal evolution, genetic divergence leading to gametic isolation or hybrid inviability/sterility, and/or ecological divergence over small spatial scales may permit species co-occurrence.

The Chalcidoidea bush of life: evolutionary history of a massive radiation of minute wasps.

Chalcidoidea are mostly parasitoid wasps that include as many as 500 000 estimated species. Capturing phylogenetic signal from such a massive radiation can be daunting. Chalcidoidea is an excellent example of a hyperdiverse group that has remained recalcitrant to phylogenetic resolution. We combined 1007 exons obtained with Anchored Hybrid Enrichment with 1048 ultra-conserved elements (UCEs) for 433 taxa including all extant families, >95% of all subfamilies, and 356 genera chosen to represent the vast diversity of the superfamily. Going back and forth between the molecular results and our collective knowledge of morphology and biology, we detected bias in the analyses that was driven by the saturation of nucleotide data. Our final results are based on a concatenated analysis of the least saturated exons and UCE datasets (2054 loci, 284 106 sites). Our analyses support an expected sister relationship with Mymarommatoidea. Seven previously recognized families were not monophyletic, so support for a new classification is discussed. Natural history in some cases would appear to be more informative than morphology, as illustrated by the elucidation of a clade of plant gall associates and a clade of taxa with planidial first-instar larvae. The phylogeny suggests a transition from smaller soft-bodied wasps to larger and more heavily sclerotized wasps, with egg parasitism as potentially ancestral for the entire superfamily. Deep divergences in Chalcidoidea coincide with an increase in insect families in the fossil record, and an early shift to phytophagy corresponds with the beginning of the "Angiosperm Terrestrial Revolution". Our dating analyses suggest a middle Jurassic origin of 174 Ma (167.3-180.5 Ma) and a crown age of 162.2 Ma (153.9-169.8 Ma) for Chalcidoidea. During the Cretaceous, Chalcidoidea may have undergone a rapid radiation in southern Gondwana with subsequent dispersals to the Northern Hemisphere. This scenario is discussed with regard to knowledge about the host taxa of chalcid wasps, their fossil record and Earth's palaeogeographic history.

Populating a Continent: Phylogenomics Reveal the Timing of Australian Frog Diversification.

The Australian continent's size and isolation make it an ideal place for studying the accumulation and evolution of biodiversity. Long separated from the ancient supercontinent Gondwana, most of Australia's plants and animals are unique and endemic, including the continent's frogs. Australian frogs comprise a remarkable ecological and morphological diversity categorized into a small number of distantly related radiations. We present a phylogenomic hypothesis based on an exon-capture dataset that spans the main clades of Australian myobatrachoid, pelodryadid hyloid, and microhylid frogs. Our time-calibrated phylogenomic-scale phylogeny identifies great disparity in the relative ages of these groups which vary from Gondwanan relics to recent immigrants from Asia and include arguably the continent's oldest living vertebrate radiation. This age stratification provides insight into the colonization of⁠, and diversification on, the Australian continent through deep time, during periods of dramatic climatic and community changes. Contemporary Australian frog diversity highlights the adaptive capacity of anurans, particularly in response to heat and aridity, and explains why they are one of the continent's most visible faunas.

Biogeography and the evolution of acoustic communication in the polyploid North American grey treefrog complex.

After polyploid species are formed, interactions between diploid and polyploid lineages may generate additional diversity in novel cytotypes and phenotypes. In anurans, mate choice by acoustic communication is the primary method by which individuals identify their own species and assess suitable mates. As such, the evolution of acoustic signals is an important mechanism for contributing to reproductive isolation and diversification in this group. Here, we estimate the biogeographical history of the North American grey treefrog complex, consisting of the diploid Hyla chrysoscelis and the tetraploid Hyla versicolor, focusing specifically on the geographical origin of whole genome duplication and the expansion of lineages out of glacial refugia. We then test for lineage-specific differences in mating signals by applying comparative methods to a large acoustic data set collected over 52 years that includes >1500 individual frogs. Along with describing the overall biogeographical history and call diversity, we found evidence that the geographical origin of H. versicolor and the formation of the midwestern polyploid lineage are both associated with glacial limits, and that the southwestern polyploid lineage is associated with a shift in acoustic phenotype relative to the diploid lineage with which they share a mitochondrial lineage. In H. chrysoscelis, we see that acoustic signals are largely split by Eastern and Western lineages, but that northward expansion along either side of the Appalachian Mountains is associated with further acoustic diversification. Overall, results of this study provide substantial clarity on the evolution of grey treefrogs as it relates to their biogeography and acoustic communication.

Chromosome-Aware Phylogenomics of Assassin Bugs (Hemiptera: Reduvioidea) Elucidates Ancient Gene Conflict.

Though the phylogenetic signal of loci on sex chromosomes can differ from those on autosomes, chromosomal-level genome assemblies for nonvertebrates are still relatively scarce and conservation of chromosomal gene content across deep phylogenetic scales has therefore remained largely unexplored. We here assemble a uniquely large and diverse set of samples (17 anchored hybrid enrichment, 24 RNA-seq, and 70 whole-genome sequencing samples of variable depth) for the medically important assassin bugs (Reduvioidea). We assess the performance of genes based on multiple features (e.g., nucleotide vs. amino acid, nuclear vs. mitochondrial, and autosomal vs. X chromosomal) and employ different methods (concatenation and coalescence analyses) to reconstruct the unresolved phylogeny of this diverse (∼7,000 spp.) and old (>180 Ma) group. Our results show that genes on the X chromosome are more likely to have discordant phylogenies than those on autosomes. We find that the X chromosome conflict is driven by high gene substitution rates that impact the accuracy of phylogenetic inference. However, gene tree clustering showed strong conflict even after discounting variable third codon positions. Alternative topologies were not particularly enriched for sex chromosome loci, but spread across the genome. We conclude that binning genes to autosomal or sex chromosomes may result in a more accurate picture of the complex evolutionary history of a clade.

Rapid radiation of ant parasitic butterflies during the Miocene aridification of Africa.

Africa has undergone a progressive aridification during the last 20 My that presumably impacted organisms and fostered the evolution of life history adaptations. We test the hypothesis that shift to living in ant nests and feeding on ant brood by larvae of phyto-predaceous Lepidochrysops butterflies was an adaptive response to the aridification of Africa that facilitated the subsequent radiation of butterflies in this genus. Using anchored hybrid enrichment we constructed a time-calibrated phylogeny for Lepidochrysops and its closest, non-parasitic relatives in the Euchrysops section (Poloyommatini). We estimated ancestral areas across the phylogeny with process-based biogeographical models and diversification rates relying on time-variable and clade-heterogeneous birth-death models. The Euchrysops section originated with the emerging Miombo woodlands about 22 million years ago (Mya) and spread to drier biomes as they became available in the late Miocene. The diversification of the non-parasitic lineages decreased as aridification intensified around 10 Mya, culminating in diversity decline. In contrast, the diversification of the phyto-predaceous Lepidochrysops lineage proceeded rapidly from about 6.5 Mya when this unusual life history likely first evolved. The Miombo woodlands were the cradle for diversification of the Euchrysops section, and our findings are consistent with the hypothesis that aridification during the Miocene selected for a phyto-predaceous life history in species of Lepidochrysops, with ant nests likely providing caterpillars a safe refuge from fire and a source of food when vegetation was scarce.

Goldenrod herbariomics: Hybrid-sequence capture reveals the phylogeny of Solidago.

PREMISE: The phylogenetic relationships among the ca. 138 species of goldenrods (Solidago; Asteraceae) have been difficult to infer due to species richness, and shallow interspecific genetic divergences. This study aims to overcome these obstacles by combining extensive sampling of goldenrod herbarium specimens with the use of a custom Solidago hybrid-sequence capture probe set. METHODS: A set of tissues from herbarium samples comprising ca. 90% of Solidago species was assembled and DNA was extracted. A custom hybrid-sequence capture probe set was designed, and data from 854 nuclear regions were obtained and analyzed from 209 specimens. Maximum likelihood and coalescent approaches were used to estimate the genus phylogeny for 157 diploid samples. RESULTS: Although DNAs from older specimens were both more fragmented and produced fewer sequencing reads, there was no relationship between specimen age and our ability to obtain sufficient data at the target loci. The Solidago phylogeny was generally well-supported, with 88/155 (57%) nodes receiving ≥95% bootstrap support. Solidago was supported as monophyletic, with Chrysoma pauciflosculosa identified as sister. A clade comprising Solidago ericameriodes, Solidago odora, and Solidago chapmanii was identified as the earliest diverging Solidago lineage. The previously segregated genera Brintonia and Oligoneuron were identified as placed well within Solidago. These and other phylogenetic results were used to establish four subgenera and fifteen sections within the genus. CONCLUSIONS: The combination of expansive herbarium sampling and hybrid-sequence capture data allowed us to quickly and rigorously establish the evolutionary relationships within this difficult, species-rich group.

Phylogenomics and biogeography of arid-adapted Chlamydogobius goby fishes.

The progressive aridification of the Australian continent from âˆ¼ 20 million years ago posed severe challenges for the persistence of its resident biota. A key question involves the role of refugial habitats - specifically, their ability to mediate the effects of habitat loss and fragmentation, and their potential to shape opportunities for allopatric speciation. With freshwater species, for example, the patchiness, or absence, of water will constrain distributions. However, aridity may not necessarily isolate populations if disjunct refugia experience frequent hydrological connections. To investigate this potential dichotomy, we explored the evolutionary history of the Chlamydogobius gobies (Gobiiformes: Gobiidae), an arid-adapted genus of six small, benthic fish species that exploit all types of waterbodies (i.e. desert springs, waterholes and bore-fed wetlands, coastal estuarine creeks and mangroves) across parts of central and northern Australia. We used Anchored Phylogenomics to generate a highly resolved phylogeny of the group from sequence data for 260 nuclear loci. Buttressed by companion allozyme and mtDNA datasets, our molecular findings infer the diversification of Chlamydogobius in arid Australia, and provide a phylogenetic structure that cannot be simply explained by invoking allopatric speciation events reflecting current geographic proximity. Our findings are generally consistent with the existing morphological delimitation of species, with one exception: at the shallowest nodes of phylogenetic reconstruction, the molecular data do not fully support the current dichotomous delineation of C. japalpa from C. eremius in Kati Thanda-Lake Eyre-associated waterbodies. Together these findings illustrate the ability of structural (hydrological) connections to generate patterns of connectivity and isolation for an ecologically moderate disperser in response to ongoing habitat aridification. Finally, we explore the implications of these results for the immediate management of threatened (C. gloveri) and critically endangered (C. micropterus, C. squamigenus) congeners.

The population genetics of speciation by cascade reinforcement.

Species interactions drive diverse evolutionary outcomes. Speciation by cascade reinforcement represents one example of how species interactions can contribute to the proliferation of species. This process occurs when the divergence of mating traits in response to selection against interspecific hybridization incidentally leads to reproductive isolation among populations of the same species. Here, we investigated the population genetic outcomes of cascade reinforcement in North American chorus frogs (Hylidae: Pseudacris). Specifically, we estimated the frequency of hybridization among three taxa, assessed genetic structure within the focal species, P. feriarum, and ascertained the directionality of gene flow within P. feriarum across replicated contact zones via coalescent modeling. Through field observations and preliminary experimental crosses, we assessed whether hybridization is possible under natural and laboratory conditions. We found that hybridization occurs among P. feriarum and two conspecifics at a low rate in multiple contact zones, and that gene flow within the former species is unidirectional from allopatry into sympatry with these other species in three of four contact zones studied. We found evidence of substantial genetic structuring within P. feriarum including a divergent western allopatric cluster, a behaviorally-distinct sympatric South Carolina cluster, and several genetically-overlapping clusters from the remainder of the distribution. Furthermore, we found sub-structuring between reinforced and nonreinforced populations in the two most intensely-sampled contact zones. Our literature review indicated that P. feriarum hybridizes with at least five heterospecifics at the periphery of its range providing a mechanism for further intraspecific diversification. This work strengthens the evidence for cascade reinforcement in this clade, revealing the geographic and genetic landscape upon which this process can contribute to the proliferation of species.

The use of Anchored Hybrid Enrichment data to resolve higher-level phylogenetic relationships: A proof-of-concept applied to Asterales (Eudicotyledoneae; Angiosperms).

Anchored Hybrid Enrichment (AHE) is a tool for capturing orthologous regions of the nuclear genome shared in low or single copy across lineages. Despite the increasing number of studies using this method, its usefulness to estimate relationships at deeper taxonomic levels in plants has not been fully explored. Here we present a proof of concept about the performance of nuclear loci obtained with AHE to infer phylogenetic relationships and explore the use of gene sampling schemes to estimate divergence times in Asterales. We recovered low-copy nuclear loci using the AHE method from herbarium material and silica-preserved samples. Maximum likelihood, Bayesian inference, and coalescence approaches were used to reconstruct phylogenomic relationships. Dating analyses were conducted under a multispecies coalescent approach by jointly inferring species tree and divergence times with random gene sampling schemes and multiple calibrations. We recovered 403 low-copy nuclear loci for 63 species representing nine out of eleven families of Asterales. Phylogenetic hypotheses were congruent among the applied methods and previously published results. Analyses with concatenated datasets were strongly supported, but coalescence-based analyses showed low support for the phylogenetic position of families Argophyllaceae and Alseuosmiaceae. Estimated family ages were congruent among gene sampling schemes, with the mean age for Asterales around 130 Myr. Our study documents the usefulness of AHE for resolving phylogenetic relationships at deep phylogenetic levels in Asterales. Observed phylogenetic inconsistencies were possibly due to the non-inclusion of families Phellinceae and Pentaphragmataceae. Random gene sampling schemes produced consistent age estimates with coalescence and species tree relaxed clock approaches.

Phylogenomics of Anguis and Pseudopus (Squamata, Anguidae) indicates Balkan-Apennine mitochondrial capture associated with the Messinian event.

A dated phylogenetic hypothesis on the evolutionary history of the extant taxa of the Western Palearctic lizards Anguis and Pseudopus is revised using genome-wide nuclear DNA and mitogenomes. We found overall concordance between nuclear and mitochondrial DNA phylogenies, with one significant exception - the Apennine A. veronensis. In mitochondrial DNA, this species forms a common clade with the earliest diverging lineage, the southern Balkan endemic A. cephallonica, while it clusters together with A. fragilis in nuclear DNA. The nuclear phylogeny conforms to the morphology, which is relatively similar between A. veronensis and A. fragilis. The most plausible explanation for the mitonuclear discordance is ancient mitochondrial capture from the Balkan ancestor of A. cephallonica to the Apennine population of the A. fragilis-veronensis ancestor. We hypothesize that this capture occurred only in a geographically restricted population. The dating of this presumed mitochondrial introgression and capture coincides with the Messinian event, when the Balkan and Apennine Peninsulas were presumably largely connected. The dated nuclear phylogenomic reconstruction estimated the divergence of A. cephallonica around 12 Mya, while the sister clade representing the A. fragilis species complex consisting of the sister species A. fragilis-A. veronensis and A. colchica-A. graeca further diversified around 7 Mya. The depth of nuclear divergence among the evolutionary lineages of Pseudopus (0.5-1.2 Mya) supports their subspecies status.

Lack of host phylogenetic structure in the gut bacterial communities of New Zealand cicadas and their interspecific hybrids.

Host-microbe interactions are intimately linked to eukaryotic evolution, particularly in sap-sucking insects that often rely on obligate microbial symbionts for nutrient provisioning. Cicadas (Cicadidae: Auchenorrhyncha) specialize on xylem fluid and derive many essential amino acids and vitamins from intracellular bacteria or fungi (Hodgkinia, Sulcia, and Ophiocordyceps) that are propagated via transmission from mothers to offspring. Despite the beneficial role of these non-gut symbionts in nutrient provisioning, the role of beneficial microbiota within the gut remains unclear. Here, we investigate the relative abundance and impact of host phylogeny and ecology on gut microbial diversity in cicadas using 16S ribosomal RNA gene amplicon sequencing data from 197 wild-collected cicadas and new mitochondrial genomes across 38 New Zealand cicada species, including natural hybrids between one pair of two species. We find low abundance and a lack of phylogenetic structure and hybrid effects but a significant role of elevation in explaining variation in gut microbiota.

Phylogenomics of trophically diverse cichlids disentangles processes driving adaptive radiation and repeated trophic transitions.

Cichlid fishes of the tribe Tropheini are a striking case of adaptive radiation, exemplifying multiple trophic transitions between herbivory and carnivory occurring in sympatry with other established cichlid lineages. Tropheini evolved highly specialized eco-morphologies to exploit similar trophic niches in different ways repeatedly and rapidly. To better understand the evolutionary history and trophic adaptations of this lineage, we generated a dataset of 532 targeted loci from 21 out of the 22 described Tropheini species. We resolved the Tropheini into seven monophyletic genera and discovered one to be polyphyletic. The polyphyletic genus, Petrochromis, represents three convergent origins of the algae grazing trophic specialization. This repeated evolution of grazing may have been facilitated by adaptive introgression as we found evidence for gene flow among algae grazing genera. We also found evidence of gene flow among algae browsing genera, but gene flow was restricted between herbivorous and carnivorous genera. Furthermore, we observed no evidence supporting a hybrid origin of this radiation. Our molecular evolutionary analyses suggest that opsin genes likely evolved in response to selection pressures associated with trophic ecology in the Tropheini. We found surprisingly little evidence of positive selection in coding regions of jaw-shaping genes in this trophically diverse lineage. This suggests low degrees of freedom for further change in these genes, and possibly a larger role for regulatory variation in driving jaw adaptations. Our study emphasizes Tropheini cichlids as an important model for studying the evolution of trophic specialization and its role in speciation.

Detecting and Removing Sample Contamination in Phylogenomic Data: An Example and its Implications for Cicadidae Phylogeny (Insecta: Hemiptera).

Contamination of a genetic sample with DNA from one or more nontarget species is a continuing concern of molecular phylogenetic studies, both Sanger sequencing studies and next-generation sequencing studies. We developed an automated pipeline for identifying and excluding likely cross-contaminated loci based on the detection of bimodal distributions of patristic distances across gene trees. When contamination occurs between samples within a data set, a comparison between a contaminated sample and its contaminant taxon will yield bimodal distributions with one peak close to zero patristic distance. This new method does not rely on a priori knowledge of taxon relatedness nor does it determine the causes(s) of the contamination. Exclusion of putatively contaminated loci from a data set generated for the insect family Cicadidae showed that these sequences were affecting some topological patterns and branch supports, although the effects were sometimes subtle, with some contamination-influenced relationships exhibiting strong bootstrap support. Long tip branches and outlier values for one anchored phylogenomic pipeline statistic (AvgNHomologs) were correlated with the presence of contamination. While the anchored hybrid enrichment markers used here, which target hemipteroid taxa, proved effective in resolving deep and shallow level Cicadidae relationships in aggregate, individual markers contained inadequate phylogenetic signal, in part probably due to short length. The cleaned data set, consisting of 429 loci, from 90 genera representing 44 of 56 current Cicadidae tribes, supported three of the four sampled Cicadidae subfamilies in concatenated-matrix maximum likelihood (ML) and multispecies coalescent-based species tree analyses, with the fourth subfamily weakly supported in the ML trees. No well-supported patterns from previous family-level Sanger sequencing studies of Cicadidae phylogeny were contradicted. One taxon (Aragualna plenalinea) did not fall with its current subfamily in the genetic tree, and this genus and its tribe Aragualnini is reclassified to Tibicininae following morphological re-examination. Only subtle differences were observed in trees after the removal of loci for which divergent base frequencies were detected. Greater success may be achieved by increased taxon sampling and developing a probe set targeting a more recent common ancestor and longer loci. Searches for contamination are an essential step in phylogenomic analyses of all kinds and our pipeline is an effective solution. [Auchenorrhyncha; base-composition bias; Cicadidae; Cicadoidea; Hemiptera; phylogenetic conflict.].

Localized Phylogenetic Discordance Among Nuclear Loci Due to Incomplete Lineage Sorting and Introgression in the Family of Cotton and Cacao (Malvaceae).

The economically important cotton and cacao family (Malvaceae sensu lato) have long been recognized as a monophyletic group. However, the relationships among some subfamilies are still unclear as discordant phylogenetic hypotheses keep arising when different sources of molecular data are analyzed. Phylogenetic discordance has previously been hypothesized to be the result of both introgression and incomplete lineage sorting (ILS), but the extent and source of discordance have not yet been evaluated in the context of loci derived from massive sequencing strategies and for a wide representation of the family. Furthermore, no formal methods have been applied to evaluate if the detected phylogenetic discordance among phylogenomic datasets influences phylogenetic dating estimates of the concordant relationships. The objective of this research was to generate a phylogenetic hypothesis of Malvaceae from nuclear genes, specifically we aimed to (1) investigate the presence of major discordance among hundreds of nuclear gene histories of Malvaceae; (2) evaluate the potential source of discordance; and (3) examine whether discordance and loci heterogeneity influence on time estimates of the origin and diversification of subfamilies. Our study is based on a comprehensive dataset representing 96 genera of the nine subfamilies and 268 nuclear loci. Both concatenated and coalescence-based approaches were followed for phylogenetic inference. Using branch lengths and topology, we located the placement of introgression events to directly evaluate whether discordance is due to introgression rather than ILS. To estimate divergence times, concordance and molecular rate were considered. We filtered loci based on congruence with the species tree and then obtained the molecular rate of each locus to distribute them into three different sets corresponding to shared molecular rate ranges. Bayesian dating was performed for each of the different sets of loci with the same parameters and calibrations. Phylogenomic discordance was detected between methods, as well as gene histories. At deep coalescent times, we found discordance in the position of five subclades probably due to ILS and a relatively small proportion of introgression. Divergence time estimation with each set of loci generated overlapping clade ages, indicating that, even with different molecular rate and gene histories, calibrations generally provide a strong prior.

Investigating the utility of Anchored Hybrid Enrichment data to investigate the relationships among the Killifishes (Actinopterygii: Cyprinodontiformes), a globally distributed group of fishes.

The Killifishes (Cyprinodontiformes) are a diverse and well-known group of fishes that contains sixteen families inclusive of Anablepidae, Aphaniidae Aplocheilidae, Cubanichthyidae, Cyprinodontidae, Fluviphylacidae, Fundulidae, Goodeidae, Nothobranchiidae, Orestiidae, Pantanodontidae, Poeciliidae, Procatopodidae, Profundulidae, Rivulidae, and Valenciidae and more than 1,200 species that are globally distributed in tropical and temperate, freshwater and estuarine habitats. The evolutionary relationships among the families within the group, based on different molecular and morphological data sets, have remained uncertain. Therefore, the objective of this study was to use a targeted approach, anchored hybrid enrichment, to investigate the phylogenetic relationships among the families within the Cyprindontiformes. This study included more than 100 individuals, representing all sixteen families within the Cyprinodontiformes, including many recently diagnosed families. We recovered an average of 244 loci per individual. These data were submitted to phylogenetic analyses (RaxML and ASTRAL) and although we recovered many of the same relationships as in previous studies of the group, several novel sets of relationships for other families also were recovered. In addition, two well-established clades (Suborders Cyprinodontoidei and Aplocheilodei) were recovered as monophyletic and are in agreement with most previous studies. We also assessed the degree of gene tree discordance in our dataset to evaluate support for alternative topological hypotheses for interfamilial relationships within the Cyprinodontiformes using a variety of different analyses. The results from this study will provide a robust, historical framework needed to investigate a plethora of biogeographic, taxonomic, ecological, and physiological questions for this group of fishes.

Candidate-species delimitation in Desmognathus salamanders reveals gene flow across lineage boundaries, confounding phylogenetic estimation and clarifying hybrid zones.

Dusky Salamanders (genus Desmognathus) currently comprise only 22 described, extant species. However, recent mitochondrial and nuclear estimates indicate the presence of up to 49 candidate species based on ecogeographic sampling. Previous studies also suggest a complex history of hybridization between these lineages. Studies in other groups suggest that disregarding admixture may affect both phylogenetic inference and clustering-based species delimitation. With a dataset comprising 233 Anchored Hybrid Enrichment (AHE) loci sequenced for 896 Desmognathus specimens from all 49 candidate species, we test three hypotheses regarding (i) species-level diversity, (ii) hybridization and admixture, and (iii) misleading phylogenetic inference. Using phylogenetic and population-clustering analyses considering gene flow, we find support for at least 47 candidate species in the phylogenomic dataset, some of which are newly characterized here while others represent combinations of previously named lineages that are collapsed in the current dataset. Within these, we observe significant phylogeographic structure, with up to 64 total geographic genetic lineages, many of which hybridize either narrowly at contact zones or extensively across ecological gradients. We find strong support for both recent admixture between terminal lineages and ancient hybridization across internal branches. This signal appears to distort concatenated phylogenetic inference, wherein more heavily admixed terminal specimens occupy apparently artifactual early-diverging topological positions, occasionally to the extent of forming false clades of intermediate hybrids. Additional geographic and genetic sampling and more robust computational approaches will be needed to clarify taxonomy, and to reconstruct a network topology to display evolutionary relationships in a manner that is consistent with their complex history of reticulation.

The Complex History of Genome Duplication and Hybridization in North American Gray Treefrogs.

Polyploid speciation has played an important role in evolutionary history across the tree of life, yet there remain large gaps in our understanding of how polyploid species form and persist. Although systematic studies have been conducted in numerous polyploid complexes, recent advances in sequencing technology have demonstrated that conclusions from data-limited studies may be spurious and misleading. The North American gray treefrog complex, consisting of the diploid Hyla chrysoscelis and the tetraploid H. versicolor, has long been used as a model system in a variety of biological fields, yet all taxonomic studies to date were conducted with only a few loci from nuclear and mitochondrial genomes. Here, we utilized anchored hybrid enrichment and high-throughput sequencing to capture hundreds of loci along with whole mitochondrial genomes to investigate the evolutionary history of this complex. We used several phylogenetic and population genetic methods, including coalescent simulations and testing of polyploid speciation models with approximate Bayesian computation, to determine that H. versicolor was most likely formed via autopolyploidization from a now extinct lineage of H. chrysoscelis. We also uncovered evidence of significant hybridization between diploids and tetraploids where they co-occur, and show that historical hybridization between these groups led to the re-formation of distinct polyploid lineages following the initial whole-genome duplication event. Our study indicates that a wide variety of methods and explicit model testing of polyploid histories can greatly facilitate efforts to uncover the evolutionary history of polyploid complexes.

Sage Insights Into the Phylogeny of Salvia: Dealing With Sources of Discordance Within and Across Genomes.

Next-generation sequencing technologies have facilitated new phylogenomic approaches to help clarify previously intractable relationships while simultaneously highlighting the pervasive nature of incongruence within and among genomes that can complicate definitive taxonomic conclusions. Salvia L., with ∼1,000 species, makes up nearly 15% of the species diversity in the mint family and has attracted great interest from biologists across subdisciplines. Despite the great progress that has been achieved in discerning the placement of Salvia within Lamiaceae and in clarifying its infrageneric relationships through plastid, nuclear ribosomal, and nuclear single-copy genes, the incomplete resolution has left open major questions regarding the phylogenetic relationships among and within the subgenera, as well as to what extent the infrageneric relationships differ across genomes. We expanded a previously published anchored hybrid enrichment dataset of 35 exemplars of Salvia to 179 terminals. We also reconstructed nearly complete plastomes for these samples from off-target reads. We used these data to examine the concordance and discordance among the nuclear loci and between the nuclear and plastid genomes in detail, elucidating both broad-scale and species-level relationships within Salvia. We found that despite the widespread gene tree discordance, nuclear phylogenies reconstructed using concatenated, coalescent, and network-based approaches recover a common backbone topology. Moreover, all subgenera, except for Audibertia, are strongly supported as monophyletic in all analyses. The plastome genealogy is largely resolved and is congruent with the nuclear backbone. However, multiple analyses suggest that incomplete lineage sorting does not fully explain the gene tree discordance. Instead, horizontal gene flow has been important in both the deep and more recent history of Salvia. Our results provide a robust species tree of Salvia across phylogenetic scales and genomes. Future comparative analyses in the genus will need to account for the impacts of hybridization/introgression and incomplete lineage sorting in topology and divergence time estimation.