Population genomics of flat-tailed horned lizards (Phrynosoma mcallii) informs conservation and management across a fragmented Colorado Desert landscape.

Phrynosoma mcallii (flat-tailed horned lizards) is a species of conservation concern in the Colorado Desert of the United States and Mexico. We analysed ddRADseq data from 45 lizards to estimate population structure, infer phylogeny, identify migration barriers, map genetic diversity hotspots, and model demography. We identified the Colorado River as the main geographic feature contributing to population structure, with the populations west of this barrier further subdivided by the Salton Sea. Phylogenetic analysis confirms that northwestern populations are nested within southeastern populations. The best-fit demographic model indicates Pleistocene divergence across the Colorado River, with significant bidirectional gene flow, and a severe Holocene population bottleneck. These patterns suggest that management strategies should focus on maintaining genetic diversity on both sides of the Colorado River and the Salton Sea. We recommend additional lands in the United States and Mexico that should be considered for similar conservation goals as those in the Rangewide Management Strategy. We also recommend periodic rangewide genomic sampling to monitor ongoing attrition of diversity, hybridization, and changing structure due to habitat fragmentation, climate change, and other long-term impacts.

Interspecific Gene Flow and Mitochondrial Genome Capture during the Radiation of Jamaican Anolis Lizards (Squamata; Iguanidae).

Gene flow and reticulation are increasingly recognized as important processes in the diversification of many taxonomic groups. With the increasing ease of collecting genomic data and the development of multispecies coalescent network approaches, such reticulations can be accounted for when inferring phylogeny and diversification. Caribbean Anolis lizards are a classic example of an adaptive radiation in which species have independently radiated on the islands of the Greater Antilles into the same ecomorph classes. Within the Jamaican radiation at least one species, Anolis opalinus, has been documented to be polyphyletic in its mitochondrial DNA, which could be the result of an ancient reticulation event or incomplete lineage sorting (ILS). Here, we generate mtDNA and genotyping-by-sequencing (GBS) data and implement gene tree, species tree, and multispecies coalescent network methods to infer the diversification of this group. Our mtDNA gene tree recovers the same relationships previously inferred for this group, which is strikingly different from the species tree inferred from our GBS data. Posterior predictive simulations suggest that our genomic data violate commonly adopted assumptions of the multispecies coalescent model (MSCM), so we use network approaches to infer phylogenetic relationships. The inferred network topology contains a reticulation event but does not explain the mtDNA polyphyly observed in this group; however, coalescent simulations suggest that the observed mtDNA topology is likely the result of past introgression. How common a signature of gene flow and reticulation is across the radiation of Anolis is unknown; however, the reticulation events that we demonstrate here may have allowed for adaptive evolution, as has been suggested in other, more recent, adaptive radiations. [Adaptive radiation; hybridization; introgression; multispecies network coalescent; posterior predictive simulation.].

Phylogeographic and phenotypic outcomes of brown anole colonization across the Caribbean provide insight into the beginning stages of an adaptive radiation.

Some of the most important insights into the ecological and evolutionary processes of diversification and speciation have come from studies of island adaptive radiations, yet relatively little research has examined how these radiations initiate. We suggest that Anolis sagrei is a candidate for understanding the origins of the Caribbean Anolis adaptive radiation and how a colonizing anole species begins to undergo allopatric diversification, phenotypic divergence and, potentially, speciation. We undertook a genomic and morphological analysis of representative populations across the entire native range of A. sagrei, finding that the species originated in the early Pliocene, with the deepest divergence occurring between western and eastern Cuba. Lineages from these two regions subsequently colonized the northern Caribbean. We find that at the broadest scale, populations colonizing areas with fewer closely related competitors tend to evolve larger body size and more lamellae on their toepads. This trend follows expectations for post-colonization divergence from progenitors and convergence in allopatry, whereby populations freed from competition with close relatives evolve towards common morphological and ecological optima. Taken together, our results show a complex history of ancient and recent Cuban diaspora with populations on competitor-poor islands evolving away from their ancestral Cuban populations regardless of their phylogenetic relationships, thus providing insight into the original diversification of colonist anoles at the beginning of the radiation. Our research also supplies an evolutionary framework for the many studies of this increasingly important species in ecological and evolutionary research.

Comparative Evolution of an Archetypal Adaptive Radiation: Innovation and Opportunity in Anolis Lizards.

Adaptive radiation is a widely recognized pattern of evolution wherein substantial phenotypic change accompanies rapid speciation. Adaptive radiation may be triggered by environmental opportunities resulting from dispersal to new areas or via the evolution of traits, called key innovations, that allow for invasion of new niches. Species sampling is a known source of bias in many comparative analyses, yet classic adaptive radiations have not been studied comparatively with comprehensively sampled phylogenies. In this study, we use unprecedented comprehensive phylogenetic sampling of Anolis lizard species to examine comparative evolution in this well-studied adaptive radiation. We compare adaptive radiation models within Anolis and in the Anolis clade and a potential sister lineage, the Corytophanidae. We find evidence for island (i.e., opportunity) effects and no evidence for trait (i.e., key innovation) effects causing accelerated body size evolution within Anolis. However, island effects are scale dependent: when Anolis and Corytophanidae are analyzed together, no island effect is evident. We find no evidence for an island effect on speciation rate and tenuous evidence for greater speciation rate due to trait effects. These results suggest the need for precision in treatments of classic adaptive radiations such as Anolis and further refinement of the concept of adaptive radiation.

A Phylogenetic, Biogeographic, and Taxonomic study of all Extant Species of Anolis (Squamata; Iguanidae).

Anolis lizards (anoles) are textbook study organisms in evolution and ecology. Although several topics in evolutionary biology have been elucidated by the study of anoles, progress in some areas has been hampered by limited phylogenetic information on this group. Here, we present a phylogenetic analysis of all 379 extant species of Anolis, with new phylogenetic data for 139 species including new DNA data for 101 species. We use the resulting estimates as a basis for defining anole clade names under the principles of phylogenetic nomenclature and to examine the biogeographic history of anoles. Our new taxonomic treatment achieves the supposed advantages of recent subdivisions of anoles that employed ranked Linnaean-based nomenclature while avoiding the pitfalls of those approaches regarding artificial constraints imposed by ranks. Our biogeographic analyses demonstrate complexity in the dispersal history of anoles, including multiple crossings of the Isthmus of Panama, two invasions of the Caribbean, single invasions to Jamaica and Cuba, and a single evolutionary dispersal from the Caribbean to the mainland that resulted in substantial anole diversity. Our comprehensive phylogenetic estimate of anoles should prove useful for rigorous testing of many comparative evolutionary hypotheses. [Anoles; biogeography; lizards; Neotropics; phylogeny; taxonomy].

Amber fossils demonstrate deep-time stability of Caribbean lizard communities.

Whether the structure of ecological communities can exhibit stability over macroevolutionary timescales has long been debated. The similarity of independently evolved Anolis lizard communities on environmentally similar Greater Antillean islands supports the notion that community evolution is deterministic. However, a dearth of Caribbean Anolis fossils--only three have been described to date--has precluded direct investigation of the stability of anole communities through time. Here we report on an additional 17 fossil anoles in Dominican amber dating to 15-20 My before the present. Using data collected primarily by X-ray microcomputed tomography (X-ray micro-CT), we demonstrate that the main elements of Hispaniolan anole ecomorphological diversity were in place in the Miocene. Phylogenetic analysis yields results consistent with the hypothesis that the ecomorphs that evolved in the Miocene are members of the same ecomorph clades extant today. The primary axes of ecomorphological diversity in the Hispaniolan anole fauna appear to have changed little between the Miocene and the present, providing evidence for the stability of ecological communities over macroevolutionary timescales.

The genome of the green anole lizard and a comparative analysis with birds and mammals.

The evolution of the amniotic egg was one of the great evolutionary innovations in the history of life, freeing vertebrates from an obligatory connection to water and thus permitting the conquest of terrestrial environments. Among amniotes, genome sequences are available for mammals and birds, but not for non-avian reptiles. Here we report the genome sequence of the North American green anole lizard, Anolis carolinensis. We find that A. carolinensis microchromosomes are highly syntenic with chicken microchromosomes, yet do not exhibit the high GC and low repeat content that are characteristic of avian microchromosomes. Also, A. carolinensis mobile elements are very young and diverse-more so than in any other sequenced amniote genome. The GC content of this lizard genome is also unusual in its homogeneity, unlike the regionally variable GC content found in mammals and birds. We describe and assign sequence to the previously unknown A. carolinensis X chromosome. Comparative gene analysis shows that amniote egg proteins have evolved significantly more rapidly than other proteins. An anole phylogeny resolves basal branches to illuminate the history of their repeated adaptive radiations.

Homology of the enigmatic nuchal bone reveals novel reorganization of the shoulder girdle in the evolution of the turtle shell.

The turtle shell represents a unique modification of the ancestral tetrapod body plan. The homologies of its approximately 50 bones have been the subject of debate for more than 200 years. Although most of those homologies are now firmly established, the evolutionary origin of the dorsal median nuchal bone of the carapace remains unresolved. We propose a novel hypothesis in which the nuchal is derived from the paired, laterally positioned cleithra-dorsal elements of the ancestral tetrapod pectoral girdle that are otherwise retained among extant tetrapods only in frogs. This hypothesis is supported by origin of the nuchal as paired, mesenchymal condensations likely derived from the neural crest followed by a unique two-stage pattern of ossification. Further support is drawn from the establishment of the nuchal as part of a highly conserved "muscle scaffold" wherein the cleithrum (and its evolutionary derivatives) serves as the origin of the Musculus trapezius. Identification of the nuchal as fused cleithra is congruent with its general spatial relationships to other elements of the shoulder girdle in the adult morphology of extant turtles, and it is further supported by patterns of connectivity and transformations documented by critical fossils from the turtle stem group. The cleithral derivation of the nuchal implies an anatomical reorganization of the pectoral girdle in which the dermal portion of the girdle was transformed from a continuous lateral-ventral arc into separate dorsal and ventral components. This transformation involved the reduction and eventual loss of the scapular rami of the clavicles along with the dorsal and superficial migration of the cleithra, which then fused with one another and became incorporated into the carapace.

Phylogenetic relationships within the lizard clade Xantusiidae: using trees and divergence times to address evolutionary questions at multiple levels.

Xantusiidae (night lizards) is a clade of small-bodied, cryptic lizards endemic to the New World. The clade is characterized by several features that would benefit from interpretation in a phylogenetic context, including: (1) monophyletic status of extant taxa Cricosaura, Lepidophyma, and Xantusia; (2) a species endemic to Cuba (Cricosaura typica) of disputed age; (3) origins of the parthenogenetic species of Lepidophyma; (4) pronounced micro-habitat differences accompanied by distinct morphologies in both Xantusia and Lepidophyma; and (5) placement of Xantusia riversiana, the only vertebrate species endemic to the California Channel Islands, which is highly divergent from its mainland relatives. This study incorporates extensive new character data from multiple gene regions to investigate the phylogeny of Xantusiidae using the most comprehensive taxonomic sampling available to date. Parsimony and partitioned Bayesian analyses of more than 7 kb of mitochondrial and nuclear sequence data from 11 loci all confirm that Xantusiidae is monophyletic, and comprises three well-supported clades: Cricosaura, Xantusia, and Lepidophyma. The Cuban endemic Cricosaura typica is well supported as the sister to all other xantusiids. Estimates of divergence time indicate that Cricosaura diverged from the (Lepidophyma+Xantusia) clade ≈ 81 million years ago (Ma), a time frame consistent with the separation of the Antilles from North America. Our results also confirm and extend an earlier study suggesting that parthenogenesis has arisen at least twice within Lepidophyma without hybridization, that rock-crevice ecomorphs evolved numerous times (>9) within Xantusia and Lepidophyma, and that the large-bodied Channel Island endemic X. riversiana is a distinct, early lineage that may form the sister group to the small-bodied congeners of the mainland.

Parallel and non-parallel phenotypic responses to environmental variation across Lesser Antillean anoles.

Species whose ranges encompass substantial environmental variation should experience heterogeneous selection, potentially resulting in local adaptation. Repeated covariation between phenotype and environment across ecologically similar species inhabiting similar environments provides strong evidence for adaptation. Lesser Antillean anoles present an excellent system in which to study repeated local adaptation because most species are widespread generalists occurring throughout environmentally heterogenous island landscapes. We leveraged this natural replication to test the hypothesis that intraspecific variation in phenotype (coloration and morphology) is consistently associated with environment across 9 species of bimaculatus series anoles. We measured dorsal coloration from 173 individuals from 6 species and 16 morphological traits from 883 individuals from 9 species, spanning their island ranges. We identified striking, but incomplete, parallelism in dorsal coloration associated with annual precipitation in our study species. By contrast, we observed significant patterns of morphological isolation-by-environment in only 2 species and no signal of parallel morphological evolution. Collectively, our results reveal strong divergent natural selection by environment on dorsal coloration but not morphology.

Genomic library preparation and hybridization capture of formalin-fixed tissues and allozyme supernatant for population genomics and considerations for combining capture- and RADseq-based single nucleotide polymorphism data sets.

Until recently many historical museum specimens were largely inaccessible to genomic inquiry, but high-throughput sequencing (HTS) approaches have allowed researchers to successfully sequence genomic DNA from dried and fluid-preserved museum specimens. In addition to preserved specimens, many museums contain large series of allozyme supernatant samples, but the amenability of these samples to HTS has not yet been assessed. Here, we compared the performance of a target-capture approach using alternative sources of genomic DNA from 10 specimens of spring salamanders (Plethodontidae: Gyrinophilus porphyriticus) collected between 1985 and 1990: allozyme supernatants, allozyme homogenate pellets and formalin-fixed tissues. We designed capture probes based on double-digest restriction-site associated sequencing (RADseq) derived loci from frozen blood samples available for seven of the specimens and assessed the success and consistency of capture and RADseq approaches. This study design enabled direct comparisons of data quality and potential biases among the different data sets for phylogenomic and population genomic analyses. We found that in phylogenetic analyses, all enrichment types for a given specimen clustered together. In principal component space all capture-based samples clustered together, but RADseq samples did not cluster with corresponding capture-based samples. Single nucleotide polymorphism calls were on average 18.3% different between enrichment types for a given individual, but these discrepancies were primarily due to differences in heterozygous/homozygous single nucleotide polymorphism calls. We demonstrate that both allozyme supernatant and formalin-fixed samples can be successfully used for population genomic analyses and we discuss ways to identify and reduce biases associated with combining capture and RADseq data.

DNA barcoding of the National Museum of Natural History reptile tissue holdings raises concerns about the use of natural history collections and the responsibilities of scientists in the molecular age.

Natural history collections are essential to a wide variety of studies in biology because they maintain large collections of specimens and associated data, including genetic material (e.g., tissues) for DNA sequence data, yet they are currently under-funded and collection staff have high workloads. With the advent of aggregate databases and advances in sequencing technologies, there is an increased demand on collection staff for access to tissue samples and associated data. Scientists are rapidly developing large DNA barcode libraries, DNA sequences of specific genes for species across the tree of life, in order to document and conserve biodiversity. In doing so, mistakes are made. For instance, inconsistent taxonomic information is commonly taken from different lending institutions and deposited in data repositories, such as the Barcode of Life Database (BOLD) and GenBank, despite explicit disclaimers regarding the need for taxonomic verification by the lending institutions. Such errors can have profound effects on subsequent research based on these mis-labelled sequences in data repositories. Here, we present the production of a large DNA barcode library of reptiles from the National Museum of Natural History tissue holdings. The library contains 2,758 sequences (2,205 COI and 553 16S) from 2260 specimens (four crocodilians, 37 turtles, and 2,219 lizards, including snakes), representing 583 named species, from 52 countries. In generating this library, we noticed several common mistakes made by scientists depositing DNA barcode data in public repositories (e.g., BOLD and GenBank). Our goal is to raise awareness of these concerns and offer advice to avoid such mistakes in the future to maintain accurate DNA barcode libraries to properly document Earth's biodiversity.

Phylogenetic relationships of the Dactyloa clade of Anolis lizards based on nuclear and mitochondrial DNA sequence data.

The Dactyloa clade, one of two major subgroups of mainland Anolis lizards, is distributed from Costa Rica to Peru, including the Amazon region and the southern Lesser Antilles. We estimated the phylogenetic relationships within Dactyloa based on mitochondrial (ND2, five transfer-RNAs, COI) and nuclear (RAG1) gene regions using likelihood and Bayesian methods under different partition strategies. In addition, we tested the monophyly of five previously recognized groups within Dactyloa. The data strongly support the monophyly of Dactyloa and five major clades: eastern, latifrons, Phenacosaurus, roquet and western, each of which exhibits a coherent geographic range. Relationships among the five major clades are less clear: support for basal nodes within Dactyloa is weak and some contradictory relationships are supported by different datasets and/or phylogenetic methods. Of the previously recognized subgroups within Dactyloa, only the roquet series consistently passed the topology tests applied. The monophyly of the aequatorialis, latifrons (as traditionally circumscribed) and punctatus series was strongly rejected, and the monophyly of Phenacosaurus (as traditionally circumscribed) yielded mixed results. The results of the phylogenetic analyses suggest the need for a revised taxonomy and have implications for the biogeography and tempo of the Dactyloa radiation.

Evolutionary drivers of sexual signal variation in Amazon Slender Anoles.

Phenotypic variation among populations, as seen in the signaling traits of many species, provides an opportunity to test whether similar factors generate repeated phenotypic patterns in different parts of a species' range. We investigated whether genetic divergence, abiotic gradients, and sympatry with closely related species explain variation in the dewlap colors of Amazon Slender Anoles, Anolis fuscoauratus. To this aim, we characterized dewlap diversity in the field with respect to population genetic structure and evolutionary relationships, assessed whether dewlap phenotypes are associated with climate or landscape variables, and tested for nonrandom associations in the distributions of A. fuscoauratus phenotypes and sympatric Anolis species. We found that dewlap colors vary among but not within sites in A. fuscoauratus. Regional genetic clusters included multiple phenotypes, while populations with similar dewlaps were often distantly related. Phenotypes did not segregate in environmental space, providing no support for optimized signal transmission at a local scale. Instead, we found a negative association between certain phenotypes and sympatric Anolis species with similar dewlap color attributes, suggesting that interactions with closely related species promoted dewlap divergence among A. fuscoauratus populations. Amazon Slender Anoles emerge as a promising system to address questions about parallel trait evolution and the contribution of signaling traits to speciation.

Niche lability in the evolution of a Caribbean lizard community.

Niche conservatism--the tendency for closely related species to be ecologically similar--is widespread. However, most studies compare closely related taxa that occur in allopatry; in sympatry, the stabilizing forces that promote niche conservatism, and thus inhibit niche shifts, may be countered by natural selection favouring ecological divergence to minimize the intensity of interspecific interactions. Consequently, the relative importance of niche conservatism versus niche divergence in determining community structure has received little attention. Here, we examine a tropical lizard community in which species have a long evolutionary history of ecological interaction. We find that evolutionary divergence overcomes niche conservatism: closely related species are no more ecologically similar than expected by random divergence and some distantly related species are ecologically similar, leading to a community in which the relationship between ecological similarity and phylogenetic relatedness is very weak. Despite this lack of niche conservatism, the ecological structuring of the community has a phylogenetic component: niche complementarity only occurs among distantly related species, which suggests that the strength of ecological interactions among species may be related to phylogeny, but it is not necessarily the most closely related species that interact most strongly.

A new cryptic species of fringe-toed lizards from southwestern Arizona with a revised taxonomy of the Uma notata species complex (Squamata: Phrynosomatidae).

Fringe-toed lizards (Uma) are among the most specialized lizards in North America, adapted to insular windblown sand habitats in the hyper-arid southwestern deserts, with allopatric distributions, subtle morphological variation, and an unstable taxonomic history. We analyzed a morphological dataset of 40 characters for 65 specimens and a molecular dataset of 2,286 bases from three mitochondrial loci for 92 individuals and interpreted these data alongside published analyses of multi-locus genetic data with the goal of revising the taxonomy of the Uma notata (Baird 1858) species complex. We confirmed that fringe-toed lizards from the Mohawk Dunes in southwestern Arizona (U. sp.) constitute a cryptic species sister to the rest of the complex that can be diagnosed with DNA barcoding and geography, so we describe and name this species Uma thurmanae sp. nov. We also confirmed the evolutionary distinctiveness of U. inornata (Cope 1895), an endangered species endemic to Coachella Valley in southern California. We designate a lectotype for the taxon U. "rufopunctata", but we put its name in quotation marks to reflect its uncertain taxonomic status with respect to its neighboring species U. cowlesi and U. notata.

Phylogeny of hoplocercine lizards (Squamata: Iguania) with estimates of relative divergence times.

Hoplocercine lizards form a clade of 11 currently recognized species traditionally placed in three genera (Enyalioides, Hoplocercus, and Morunasaurus) that occur in the lowlands on both sides of the Andes between Panama and the Brazilian Cerrado. We analyze 11 mitochondrial and two nuclear loci using probabilistic methods and different partitioning strategies to (1) infer the phylogenetic relationships among species of Hoplocercinae, (2) examine amounts of inter- and intraspecific sequence divergence, (3) address monophyly of four species, (4) test previous phylogenetic hypotheses, and (5) estimate divergence times. Our preferred hypothesis places H. spinosus as the sister taxon to all other species of hoplocercines, with M. annularis nested within Enyalioides. Species with multiple samples are monophyletic except for Enyalioides oshaughnessyi, which is paraphyletic relative to an undescribed species of Enyalioides. All previously published phylogenetic hypotheses for hoplocercines are rejected. Monophyly of Enyalioides cannot be rejected and, consequently, the position of Morunasaurus remains unclear. The most recent common ancestor of Hoplocercinae probably occurred east of the Andes; western taxa included in our analyses originated from at least two separate colonizations whether pre- or post-dating vicariance resulting from uplift of the Andes.