Evolution of thermal physiology alters the projected range of threespine stickleback under climate change.

Species distribution models (SDMs) are widely used to predict range shifts but could be unreliable under climate change scenarios because they do not account for evolution. The thermal physiology of a species is a key determinant of its range and thus incorporating thermal trait evolution into SDMs might be expected to alter projected ranges. We identified a genetic basis for physiological and behavioural traits that evolve in response to temperature change in natural populations of threespine stickleback (Gasterosteus aculeatus). Using these data, we created geographical range projections using a mechanistic niche area approach under two climate change scenarios. Under both scenarios, trait data were either static ("no evolution" models), allowed to evolve at observed evolutionary rates ("evolution" models) or allowed to evolve at a rate of evolution scaled by the trait variance that is explained by quantitative trait loci (QTL; "scaled evolution" models). We show that incorporating these traits and their evolution substantially altered the projected ranges for a widespread panmictic marine population, with over 7-fold increases in area under climate change projections when traits are allowed to evolve. Evolution-informed SDMs should improve the precision of forecasting range dynamics under climate change, and aid in their application to management and the protection of biodiversity.

Investigating the evolution and development of biological complexity under the framework of epigenetics.

Biological complexity is a key component of evolvability, yet its study has been hampered by a focus on evolutionary trends of complexification and inconsistent definitions. Here, we demonstrate the utility of bringing complexity into the framework of epigenetics to better investigate its utility as a concept in evolutionary biology. We first analyze the existing metrics of complexity and explore the link between complexity and adaptation. Although recently developed metrics allow for a unified framework, they omit developmental mechanisms. We argue that a better approach to the empirical study of complexity and its evolution includes developmental mechanisms. We then consider epigenetic mechanisms and their role in shaping developmental and evolutionary trajectories, as well as the development and organization of complexity. We argue that epigenetics itself could have emerged from complexity because of a need to self-regulate. Finally, we explore hybridization complexes and hybrid organisms as potential models for studying the association between epigenetics and complexity. Our goal is not to explain trends in biological complexity but to help develop and elucidate novel questions in the investigation of biological complexity and its evolution.

Conditional Creation and Rescue of Nipbl-Deficiency in Mice Reveals Multiple Determinants of Risk for Congenital Heart Defects.

Elucidating the causes of congenital heart defects is made difficult by the complex morphogenesis of the mammalian heart, which takes place early in development, involves contributions from multiple germ layers, and is controlled by many genes. Here, we use a conditional/invertible genetic strategy to identify the cell lineage(s) responsible for the development of heart defects in a Nipbl-deficient mouse model of Cornelia de Lange Syndrome, in which global yet subtle transcriptional dysregulation leads to development of atrial septal defects (ASDs) at high frequency. Using an approach that allows for recombinase-mediated creation or rescue of Nipbl deficiency in different lineages, we uncover complex interactions between the cardiac mesoderm, endoderm, and the rest of the embryo, whereby the risk conferred by genetic abnormality in any one lineage is modified, in a surprisingly non-additive way, by the status of others. We argue that these results are best understood in the context of a model in which the risk of heart defects is associated with the adequacy of early progenitor cell populations relative to the sizes of the structures they must eventually form.

Contemporary ancestor? Adaptive divergence from standing genetic variation in Pacific marine threespine stickleback.

BACKGROUND: Populations that have repeatedly colonized novel environments are useful for studying the role of ecology in adaptive divergence - particularly if some individuals persist in the ancestral habitat. Such "contemporary ancestors" can be used to demonstrate the effects of selection by comparing phenotypic and genetic divergence between the derived population and their extant ancestors. However, evolution and demography in these "contemporary ancestors" can complicate inferences about the source (standing genetic variation, de novo mutation) and pace of adaptive divergence. Marine threespine stickleback (Gasterosteus aculeatus) have colonized freshwater environments along the Pacific coast of North America, but have also persisted in the marine environment. To what extent are marine stickleback good proxies of the ancestral condition? RESULTS: We sequenced > 5800 variant loci in over 250 marine stickleback from eight locations extending from Alaska to California, and phenotyped them for platedness and body shape. Pairwise FST varied from 0.02 to 0.18. Stickleback were divided into five genetic clusters, with a single cluster comprising stickleback from Washington to Alaska. Plate number, Eda, body shape, and candidate loci showed evidence of being under selection in the marine environment. Comparisons to a freshwater population demonstrated that candidate loci for freshwater adaptation varied depending on the choice of marine populations. CONCLUSIONS: Marine stickleback are structured into phenotypically and genetically distinct populations that have been evolving as freshwater stickleback evolved. This variation complicates their usefulness as proxies of the ancestors of freshwater populations. Lessons from stickleback may be applied to other "contemporary ancestor"-derived population studies.

Body size and allometric variation in facial shape in children.

OBJECTIVES: Morphological integration, or the tendency for covariation, is commonly seen in complex traits such as the human face. The effects of growth on shape, or allometry, represent a ubiquitous but poorly understood axis of integration. We address the question of to what extent age and measures of size converge on a single pattern of allometry for human facial shape. METHODS: Our study is based on two large cross-sectional cohorts of children, one from Tanzania and the other from the United States (N = 7,173). We employ 3D facial imaging and geometric morphometrics to relate facial shape to age and anthropometric measures. RESULTS: The two populations differ significantly in facial shape, but the magnitude of this difference is small relative to the variation within each group. Allometric variation for facial shape is similar in both populations, representing a small but significant proportion of total variation in facial shape. Different measures of size are associated with overlapping but statistically distinct aspects of shape variation. Only half of the size-related variation in facial shape can be explained by the first principal component of four size measures and age while the remainder associates distinctly with individual measures. CONCLUSIONS: Allometric variation in the human face is complex and should not be regarded as a singular effect. This finding has important implications for how size is treated in studies of human facial shape and for the developmental basis for allometric variation more generally.

Speciation through the lens of biomechanics: locomotion, prey capture and reproductive isolation.

Speciation is a multifaceted process that involves numerous aspects of the biological sciences and occurs for multiple reasons. Ecology plays a major role, including both abiotic and biotic factors. Whether populations experience similar or divergent ecological environments, they often adapt to local conditions through divergence in biomechanical traits. We investigate the role of biomechanics in speciation using fish predator-prey interactions, a primary driver of fitness for both predators and prey. We highlight specific groups of fishes, or specific species, that have been particularly valuable for understanding these dynamic interactions and offer the best opportunities for future studies that link genetic architecture to biomechanics and reproductive isolation (RI). In addition to emphasizing the key biomechanical techniques that will be instrumental, we also propose that the movement towards linking biomechanics and speciation will include (i) establishing the genetic basis of biomechanical traits, (ii) testing whether similar and divergent selection lead to biomechanical divergence, and (iii) testing whether/how biomechanical traits affect RI. Future investigations that examine speciation through the lens of biomechanics will propel our understanding of this key process.

The structural and compositional transition of the meniscal roots into the fibrocartilage of the menisci.

The meniscal roots, or insertional ligaments, firmly attach the menisci to tibial plateau. These strong attachments anchor the menisci and allow for the generation of hoop stress in the tissue. The meniscal roots have a ligament-like structure that transitions into the fibrocartilagenous structure of the meniscal body. The purpose of this study was to carry out a complete analysis of the structure and tissue organization from the body of the meniscus through the transition region and into the insertional roots. Serial sections were obtained from the meniscal roots into the meniscal body in fixed juvenile bovine menisci. Sections were stained for collagen and proteoglycans (PG) using fast green and safranin-o staining protocols. Unstained sections were imaged used a backlit stereo microscope. Optical projection tomography (OPT) was employed to evaluate the three-dimensional collagen architecture of the root-meniscus transition in lapine menisci. Tie-fibres were observed in the sections of the ligaments furthest from the bovine meniscal body. Blood vessels were observed to be surrounded by these tie-fibres and a PG-rich region within the ligaments. Near the tibial insertion, the roots contained large ligament-like collagen fascicles. In sections approaching the meniscus, there was an increase in tie-fibre size and density. Small tie-fibres extended into the ligament from the epiligamentous structure in the outermost sections of the meniscal roots, while large tie-fibre bundles were apparent at the meniscus transition. The staining pattern indicates that the root may continue into the outer portion of the meniscus where it then blends with the more fibrocartilage-like inner portions of the tissue. In unstained sections it was observed that the femoral side of the epiligamentous structure surrounding the root becomes more fibrous and thickens in the inferior inner portion of the posterior medial root. This thickening changes the shape of the root to more closely resemble the meniscus wedge shape. These observations support the concept of root continuity with the outer portion of the meniscus, thereby connecting with the hoop-like structure of the peripheral meniscus. OPT identified continuous collagen organization from the root into the meniscal body in longitudinal sections. In the radial direction, the morphology of the root continues into the meniscal body consistent with the serially sectioned bovine menisci. Blood vessels were prevalent on the periphery of the root. These blood vessels then arborized to cover the anterior femoral surface of the meniscus. This is the first study of the structural transition between the insertional ligaments (roots) and the fibrocartilagenous body of the menisci. These new structural details are important to understanding the meniscal load-bearing mechanism in the knee.

The shape of things to come in the study of the origin of species?

Perhaps Darwin would agree that speciation is no longer the mystery of mysteries that it used to be. It is now generally accepted that evolution by natural selection can contribute to ecological adaptation, resulting in the evolution of reproductive barriers and, hence, to the evolution of new species (Schluter & Conte 2009; Meyer 2011; Nosil 2012). From genes that encode silencing proteins that cause infertility in hybrid mice (Mihola et al. 2009), to segregation distorters linked to speciation in fruit flies (Phadnis & Orr 2009), or pollinator-mediated selection on flower colour alleles driving reinforcement in Texan wildflowers (Hopkins & Rausher 2012), characterization of the genes that drive speciation is providing clues to the origin of species (Nosil & Schluter 2011). It is becoming apparent that, while recent work continues to overturn historical ideas about sympatric speciation (e.g. Barluenga et al. 2006), ecological circumstances strongly influence patterns of genomic divergence, and ultimately the establishment of reproductive isolation when gene flow is present (Elmer & Meyer 2011). Less clear, however, are the genetic mechanisms that cause speciation, particularly when ongoing gene flow is occurring. Now, in this issue, Franchini et al. (2014) employ a classic genetic mapping approach augmented with new genomic tools to elucidate the genomic architecture of ecologically divergent body shapes in a pair of sympatric crater lake cichlid fishes. From over 450 segregating SNPs in an F2 cross, 72 SNPs were linked to 11 QTL associated with external morphology measured by means of traditional and geometric morphometrics. Annotation of two highly supported QTL further pointed to genes that might contribute to ecological divergence in body shape in Midas cichlids, overall supporting the hypothesis that genomic regions of large phenotypic effect may be contributing to early-stage divergence in Midas cichlids.

An evaluation of meniscal collagenous structure using optical projection tomography.

BACKGROUND: The collagenous structure of menisci is a complex network of circumferentially oriented fascicles and interwoven radially oriented tie-fibres. To date, examination of this micro- architecture has been limited to two-dimensional imaging techniques. The purpose of this study was to evaluate the ability of the three-dimensional imaging technique; optical projection tomography (OPT), to visualize the collagenous structure of the meniscus. If successful, this technique would be the first to visualize the macroscopic orientation of collagen fascicles in 3-D in the meniscus and could further refine load bearing mechanisms in the tissue. OPT is an imaging technique capable of imaging samples on the meso-scale (1-10 mm) at a micro-scale resolution. The technique, similar to computed tomography, takes two-dimensional images of objects from incremental angles around the object and reconstructs them using a back projection algorithm to determine three-dimensional structure. METHODS: Bovine meniscal samples were imaged from four locations (outer main body, femoral surface, tibial surface and inner main body) to determine the variation in collagen orientation throughout the tissue. Bovine stifles (n = 2) were obtained from a local abattoir and the menisci carefully dissected. Menisci were fixed in methanol and subsequently cut using a custom cutting jig (n = 4 samples per meniscus). Samples were then mounted in agarose, dehydrated in methanol and subsequently cleared using benzyl alcohol benzyl benzoate (BABB) and imaged using OPT. RESULTS: Results indicate circumferential, radial and oblique collagenous orientations at the contact surfaces and in the inner third of the main body of the meniscus. Imaging identified fascicles ranging from 80-420 µm in diameter. Transition zones where fascicles were found to have a woven or braided appearance were also identified. The outer-third of the main body was composed of fascicles oriented predominantly in the circumferential direction. Blood vessels were also visualized using this technique, as their elastin content fluoresces more brightly than collagen at the 425 nm wavelength used by the OPT scanner. CONCLUSIONS: OPT was capable of imaging the collagenous structure, as well as blood vessels in the bovine meniscus. Collagenous structure variability, including transition zones between structural regions not previously described in the meniscus, was identified using this novel technique.

The generation of variation and the developmental basis for evolutionary novelty.

Organisms exhibit an incredible diversity of form, a fact that makes the evolution of novelty seemingly self-evident. However, despite the "obvious" case for novelty, defining this concept in evolutionary terms is highly problematic, so much so that some have suggested discarding it altogether. Approaches to this problem tend to take either an adaptation- or development-based perspective, but we argue here that an exclusive focus on either of these misses the original intent of the novelty concept and undermines its practical utility. We propose instead that for a feature to be novel, it must have evolved both by a transition between adaptive peaks on the fitness landscape and that this transition must have overcome a previous developmental constraint. This definition focuses novelty on the explanation of apparently difficult or low-probability evolutionary transitions and highlights how the integration of developmental and functional considerations are necessary to evolutionary explanation. It further reinforces that novelty is a central concern not just of evolutionary developmental biology (i.e., "evo-devo") but of evolutionary biology more generally. We explore this definition of novelty in light of four examples that range from the obvious to subtle.

Epigenetic integration of the developing brain and face.

The integration of the brain and face and to what extent this relationship constrains or enables evolutionary change in the craniofacial complex is an issue of long-standing interest in vertebrate evolution. To investigate brain-face integration, we studied the covariation between the forebrain and midface at gestational days 10-10.5 in four strains of laboratory mice. We found that phenotypic variation in the forebrain is highly correlated with that of the face during face formation such that variation in the size of the forebrain correlates with the degree of prognathism and orientation of the facial prominences. This suggests strongly that the integration of the brain and face is relevant to the etiology of midfacial malformations such as orofacial clefts. This axis of integration also has important implications for the evolutionary developmental biology of the mammalian craniofacial complex.

Sexually mediated phenotypic variation within and between sexes as a continuum structured by ecology: The mosaic nature of skeletal variation across body regions in Threespine stickleback (Gasterosteus aculeatus L.).

Ecological character displacement between the sexes, and sexual selection, integrate into a convergent set of factors that produce sexual variation. Ecologically modulated, sexually mediated variation within and between sexes may be a major contributor to the amount of total variation that selection can act on in species. Threespine stickleback (Gasterosteus aculeatus) display rapid adaptive responses and sexual variation in many phenotypic traits. We examined phenotypic variation in the skull, pectoral and pelvic girdles of threespine stickleback from two freshwater and two coastal marine sites on the Sunshine Coast of British Columbia, Canada, using an approach that avoids a priori assumptions about bimodal patterns of variation. We quantified shape and size of the cranial, pectoral and pelvic regions of sticklebacks in marine and freshwater habitats using 3D geometric morphometrics and an index of sexually mediated variation. We show that the expression of phenotypic variation is structured in part by the effects of both habitat marine vs freshwater and the effects of individual sites within each habitat. Relative size exerts variable influence, and patterns of phenotypic variation associated with sex vary among body regions. This fine-grained quantification of sexually mediated variation in the context of habitat difference and different anatomical structures indicates a complex relationship between genetically inferred sex and environmental factors, demonstrating that the interplay between shared genetic background and sexually mediated, ecologically based selective pressures structures the phenotypic expression of complex traits.

Modularity in the skull and cranial vasculature of laboratory mice: implications for the evolution of complex phenotypes.

The generation of coordinated morphological change over time results from the interconnectedness of evolution and development. The modular architecture of development results in varying degrees of integration and independence among parts of the phenotype, and facilitates the production of phenotypic variation in complex anatomical units composed of multiple tissue types. Here we use geometric morphometrics to investigate modularity in the arterial Circle of Willis (CW) and skull of the CD-1 laboratory mouse. We contrast a hypothesis of tight integration between these tissues with a hypothesis of more modular organization, to determine the level at which natural selection works to generate coordinated change. We report a complex pattern of covariation that indicates that the skull and CW are highly integrated and developmentally linked. Further, we report higher levels of fluctuating asymmetry in the CW than in the skull, suggesting a greater potential for lability in this tissue. These results suggest that epigenetic interactions or genetic influences on regional development are more important determinants of covariation structure than the factors that produce covariation within individual tissues or organ systems.

Heritability of DNA methylation in threespine stickleback (Gasterosteus aculeatus).

Epigenetic mechanisms underlying phenotypic change are hypothesized to contribute to population persistence and adaptation in the face of environmental change. To date, few studies have explored the heritability of intergenerationally stable methylation levels in natural populations, and little is known about the relative contribution of cis- and trans-regulatory changes to methylation variation. Here, we explore the heritability of DNA methylation, and conduct methylation quantitative trait loci (meQTLs) analysis to investigate the genetic architecture underlying methylation variation between marine and freshwater ecotypes of threespine stickleback (Gasterosteus aculeatus). We quantitatively measured genome-wide DNA methylation in fin tissue using reduced representation bisulfite sequencing of F1 and F2 crosses, and their marine and freshwater source populations. We identified cytosines (CpG sites) that exhibited stable methylation levels across generations. We found that additive genetic variance explained an average of 24-35% of the methylation variance, with a number of CpG sites possibly autonomous from genetic control. We also detected both cis- and trans-meQTLs, with only trans-meQTLs overlapping with previously identified genomic regions of high differentiation between marine and freshwater ecotypes. Finally, we identified the genetic architecture underlying two key CpG sites that were differentially methylated between ecotypes. These findings demonstrate a potential role for DNA methylation in facilitating adaptation to divergent environments and improve our understanding of the heritable basis of population epigenomic variation.

Linking ecomechanical models and functional traits to understand phenotypic diversity.

Physical principles and laws determine the set of possible organismal phenotypes. Constraints arising from development, the environment, and evolutionary history then yield workable, integrated phenotypes. We propose a theoretical and practical framework that considers the role of changing environments. This 'ecomechanical approach' integrates functional organismal traits with the ecological variables. This approach informs our ability to predict species shifts in survival and distribution and provides critical insights into phenotypic diversity. We outline how to use the ecomechanical paradigm using drag-induced bending in trees as an example. Our approach can be incorporated into existing research and help build interdisciplinary bridges. Finally, we identify key factors needed for mass data collection, analysis, and the dissemination of models relevant to this framework.

Micro-computed tomography-based phenotypic approaches in embryology: procedural artifacts on assessments of embryonic craniofacial growth and development.

BACKGROUND: Growing demand for three dimensional (3D) digital images of embryos for purposes of phenotypic assessment drives implementation of new histological and imaging techniques. Among these micro-computed tomography (microCT) has recently been utilized as an effective and practical method for generating images at resolutions permitting 3D quantitative analysis of gross morphological attributes of developing tissues and organs in embryonic mice. However, histological processing in preparation for microCT scanning induces changes in organ size and shape. Establishing normative expectations for experimentally induced changes in size and shape will be an important feature of 3D microCT-based phenotypic assessments, especially if quantifying differences in the values of those parameters between comparison sets of developing embryos is a primary aim. Toward that end, we assessed the nature and degree of morphological artifacts attending microCT scanning following use of common fixatives, using a two dimensional (2D) landmark geometric morphometric approach to track the accumulation of distortions affecting the embryonic head from the native, uterine state through to fixation and subsequent scanning. RESULTS: Bouin's fixation reduced average centroid sizes of embryonic mouse crania by approximately 30% and substantially altered the morphometric shape, as measured by the shift in Procrustes distance, from the unfixed state, after the data were normalized for naturally occurring shape variation. Subsequent microCT scanning produced negligible changes in size but did appear to reduce or even reverse fixation-induced random shape changes. Mixtures of paraformaldehyde + glutaraldehyde reduced average centroid sizes by 2-3%. Changes in craniofacial shape progressively increased post-fixation. CONCLUSIONS: The degree to which artifacts are introduced in the generation of random craniofacial shape variation relates to the degree of specimen dehydration during the initial fixation. Fixation methods that better maintain original craniofacial dimensions at reduced levels of dehydration and tissue shrinkage lead to the progressive accumulation of random shape variation during handling and data acquisition. In general, to the degree that embryonic organ size and shape factor into microCT-based phenotypic assessments, procedurally induced artifacts associated with fixation and scanning will influence results. Experimental designs will need to address these significant effects, either by employing alternative methods that minimize artifacts in the region of focus or in the interpretation of statistical patterns.

Ecomorphological associations of scapulocoracoid form in Greater Antillean Anolis lizards.

External morphological metrics have featured prominently in comparative studies examining the morphological convergence that characterizes anoline ecomorphs. To what degree the appendicular-skeletal morphology of Greater Antillean island Anolis lizards tracks their diversity and ecological adaptation, however, remains relatively unexplored. Here we employ computed tomographic scanning techniques to visualize in situ the scapulocoracoid of ecomorph representatives (trunk-ground, trunk-crown, crown-giant, twig) from three islands (Jamaica, Hispaniola, and Puerto Rico), and compare its three-dimensional geometry using qualitative-descriptive and quantitative-morphometric techniques. In contrast to our previous, similarly-conducted study of the pelvic girdle of these same species, the form of the scapulocoracoid varies markedly both within and between species, with much of the variation relating to phylogenetic relationship, specimen size, and assigned ecomorph category. Morphometric variation that correlates with size and/or phylogenetic signal varies between species and cannot be eliminated from the data set without markedly reducing its overall variability. The discovered patterns of skeletal variation are consistent with the demands of locomotor mechanics imposed by the structural configuration of the microhabitat of these ecomorphs. Most pertinently the ecomorphs differ in the anteroposterior length of the coracoid, the dorsoventral height of the scapulocoracoid, the dorsoventral height of the scapula in relation to the height of the suprascapula, and the relative positioning of the borders of the scapulocoracoid fenestra. In the examined ecomorph categories these skeletal differences likely relate to microhabitat usage by permitting different degrees of tilting and displacement of the scapulocoracoid in the parasagittal plane and influencing the sizes of muscle origins and the vectors of their actions. These differences relate to the amount of humeral adduction applied during its protraction, and to the structural stability of the shoulder girdle during acrobatic maneuvers, thus influencing the perch diameter that can be effectively negotiated, a critical factor in the microhabitat structure of Anolis ecomorphs.

Quantifying two-dimensional and three-dimensional stereoscopic learning in anatomy using electroencephalography.

Advances in computer visualization enabling both 2D and 3D representation have generated tools to aid perception of spatial relationships and provide a new forum for instructional design. A key knowledge gap is the lack of understanding of how the brain neurobiologically processes and learns from spatially presented content, and new quantitative variables are required to address this gap. The objective of this study was to apply quantitative neural measures derived from electroencephalography (EEG) to examine stereopsis in anatomy learning by comparing mean amplitude changes in N250 (related to object recognition) and reward positivity (related to responding to feedback) event related to potential components using a reinforcement-based learning paradigm. Health sciences students (n = 61) learned to identify and localize neuroanatomical structures using 2D, 3D, or a combination of models while EEG and behavioral (accuracy) data were recorded. Participants learning using 3D models had a greater object recognition (N250 amplitude) compared to those who learned from 2D models. Based on neurological results, interleaved learning incorporating both 2D and 3D models provided an advantage in learning, retention, and transfer activities represented by decreased reward positivity amplitude. Behavioral data did not have the same sensitivity as neural data for distinguishing differences in learning with and without stereopsis in these learning activities. Measuring neural activity reveals new insights in applied settings for educators to consider when incorporating stereoscopic models in the design of learning interventions.

Spatial packing, cranial base angulation, and craniofacial shape variation in the mammalian skull: testing a new model using mice.

The hypothesis that variation in craniofacial shape within and among species is influenced by spatial packing has a long history in comparative anatomy, particularly in terms of primates. This study develops and tests three alternative models of spatial packing to address how and to what extent the cranial base angle is influenced by variation in brain and facial size. The models are tested using mouse strains with different mutations affecting craniofacial growth. Although mice have distinctive crania with small brains, long faces, and retroflexed cranial bases, the results of the study indicate that the mouse cranial base flexes to accommodate larger brain size relative to cranial base length. In addition, the mouse cranial base also extends, but to a lesser degree, to accommodate larger face size relative to cranial base length. In addition, interactions between brain size, face size, and the widths and lengths of the components of the cranial base account for a large percentage of variation in cranial base angle. The results illustrate the degree to which the cranial base is centrally embedded within the covariation structure of the craniofacial complex as a whole.

What is bred in the bone: Ecomorphological associations of pelvic girdle form in greater Antillean Anolis lizards.

Ecological niche partitioning of Anolis lizards of the Greater Antillean islands has been the focus of many comparative studies, and much is known about external morphological convergence that characterizes anole ecomorphs. Their internal anatomy, however, has rarely been explored in an ecomorphological context, and it remains unknown to what degree skeletal morphology tracks the diversity and ecological adaptation of these lizards. Herein, we employ CT scanning techniques to visualise the skeleton of the pelvic girdle in situ, and 3D geometric morphometrics to compare the form of the ilium, ischium, and pubis within and between ecomorphs. We examine 26 species of anoles representing four ecomorphs (trunk-ground, trunk-crown, crown-giant, twig) from three islands (Jamaica, Hispaniola, and Puerto Rico). The subtle variations in pelvic girdle morphology discovered are directly associable with all three parameters that we set out to focus on: phylogenetic relationship, specimen size, and assigned ecomorph category. Morphometric variation that correlates with size and/or phylogenetic signal varies between species and cannot be eliminated from the data set without markedly reducing its overall variability. The discovered patterns of skeletal variation are consistent with the demands of locomotor mechanics pertinent to the structural configuration of the microhabitat of three of the four ecomorphs, with the fourth having no discernible distinctive features. This manifests itself chiefly in the relative anteroposterior extent and anteroventral inclination of the ilium and pubis, which differ between ecomorphs and are postulated to reflect optimization of the direction of muscle vectors of the femoral protractors and retractors. Our investigation of the form of the pelvic girdle of anoles allows us to generalize our findings to entire ecomorph categories within a broad phylogenetic and biogeographic context. Differences in the form and configuration of the postcranial skeleton are directly related to ecological patterns.