The macroevolutionary singularity of snakes.

Snakes and lizards (Squamata) represent a third of terrestrial vertebrates and exhibit spectacular innovations in locomotion, feeding, and sensory processing. However, the evolutionary drivers of this radiation remain poorly known. We infer potential causes and ultimate consequences of squamate macroevolution by combining individual-based natural history observations (>60,000 animals) with a comprehensive time-calibrated phylogeny that we anchored with genomic data (5400 loci) from 1018 species. Due to shifts in the dynamics of speciation and phenotypic evolution, snakes have transformed the trophic structure of animal communities through the recurrent origin and diversification of specialized predatory strategies. Squamate biodiversity reflects a legacy of singular events that occurred during the early history of snakes and reveals the impact of historical contingency on vertebrate biodiversity.

Microchromosome fusions underpin convergent evolution of chameleon karyotypes.

Evolutionary shifts in chromosome compositions (karyotypes) are major drivers of lineage and genomic diversification. Fusion of ancestral chromosomes is one hypothesized mechanism for the evolutionary reduction of the total chromosome number, a frequently implied karyotypic shift. Empirical tests of this hypothesis require model systems with variable karyotypes, known chromosome features, and a robust phylogeny. Here we used chameleons, diverse lizards with exceptionally variable karyotypes ($2n=20\text{-}62$), to test whether chromosomal fusions explain the repeated evolution of karyotypes with fewer chromosomes than ancestral karyotypes. Using a multidisciplinary approach including cytogenetic analyses and phylogenetic comparative methods, we found that a model of constant loss through time best explained chromosome evolution across the chameleon phylogeny. Next, we tested whether fusions of microchromosomes into macrochromosomes explained these evolutionary losses using generalized linear models. Multiple comparisons supported microchromosome fusions as the predominant agent of evolutionary loss. We further compared our results to various natural history traits and found no correlations. As such, we infer that the tendency of microchromosomes to fuse was a quality of the ancestral chameleon genome and that the genomic predisposition of ancestors is a more substantive predictor of chromosome change than the ecological, physiological, and biogeographical factors involved in their diversification.

Multiple pathways to herbivory underpinned deep divergences in ornithischian evolution.

The extent to which evolution is deterministic is a key question in biology,1,2,3,4,5,6,7,8,9 with intensive debate on how adaptation6,10,11,12,13 and constraints14,15,16 might canalize solutions to ecological challenges.4,5,6 Alternatively, unique adaptations1,9,17 and phylogenetic contingency1,3,18 may render evolution fundamentally unpredictable.3 Information from the fossil record is critical to this debate,1,2,11 but performance data for extinct taxa are limited.7 This knowledge gap is significant, as general morphology may be a poor predictor of biomechanical performance.17,19,20 High-fiber herbivory originated multiple times within ornithischian dinosaurs,21 making them an ideal clade for investigating evolutionary responses to similar ecological pressures.22 However, previous biomechanical modeling studies on ornithischian crania17,23,24,25 have not compared early-diverging taxa spanning independent acquisitions of herbivory. Here, we perform finite-element analysis on the skull of five early-diverging members of the major ornithischian clades to characterize morphofunctional pathways to herbivory. Results reveal limited functional convergence among ornithischian clades, with each instead achieving comparable performance, in terms of reconstructed patterns and magnitudes of functionally induced stress, through different adaptations of the feeding apparatus. Thyreophorans compensated for plesiomorphic low performance through increased absolute size, heterodontosaurids expanded jaw adductor muscle volume, ornithopods increased jaw system efficiency, and ceratopsians combined these approaches. These distinct solutions to the challenges of herbivory within Ornithischia underpinned the success of this diverse clade. Furthermore, the resolution of multiple solutions to equivalent problems within a single clade through macroevolutionary time demonstrates that phenotypic evolution is not necessarily predictable, instead arising from the interplay of adaptation, innovation, contingency, and constraints.1,2,3,7,8,9,18.

Middle Jurassic fossils document an early stage in salamander evolution.

Salamanders are an important group of living amphibians and model organisms for understanding locomotion, development, regeneration, feeding, and toxicity in tetrapods. However, their origin and early radiation remain poorly understood, with early fossil stem-salamanders so far represented by larval or incompletely known taxa. This poor record also limits understanding of the origin of Lissamphibia (i.e., frogs, salamanders, and caecilians). We report fossils from the Middle Jurassic of Scotland representing almost the entire skeleton of the enigmatic stem-salamander Marmorerpeton. We use computed tomography to visualize high-resolution three-dimensional anatomy, describing morphologies that were poorly characterized in early salamanders, including the braincase, scapulocoracoid, and lower jaw. We use these data in the context of a phylogenetic analysis intended to resolve the relationships of early and stem-salamanders, including representation of important outgroups alongside data from high-resolution imaging of extant species. Marmorerpeton is united with Karaurus, Kokartus, and others from the Middle Jurassic-Lower Cretaceous of Asia, providing evidence for an early radiation of robustly built neotenous stem-salamanders. These taxa display morphological specializations similar to the extant cryptobranchid "giant" salamanders. Our analysis also demonstrates stem-group affinities for a larger sample of Jurassic species than previously recognized, highlighting an unappreciated diversity of stem-salamanders and cautioning against the use of single species (e.g., Karaurus) as exemplars for stem-salamander anatomy. These phylogenetic findings, combined with knowledge of the near-complete skeletal anatomy of Mamorerpeton, advance our understanding of evolutionary changes on the salamander stem-lineage and provide important data on early salamanders and the origins of Batrachia and Lissamphibia.

Clade-wide variation in bite-force performance is determined primarily by size, not ecology.

Performance traits are tightly linked to the fitness of organisms. However, because studies of variation in performance traits generally focus on just one or several closely related species, we are unable to draw broader conclusions about how and why these traits vary across clades. One important performance trait related to many aspects of an animal's life history is bite-force. Here, we use a clade-wide phylogenetic comparative approach to investigate relationships between size, head dimensions and bite-force among lizards and tuatara (lepidosaurs), using the largest bite-force dataset collated to date for any taxonomic group. We test four predictions: that bite-force will be greater in larger species, and for a given body size, bite-force will be greatest in species with acrodont tooth attachment, herbivorous diets, and non-burrowing habits. We show that bite-force is strongly related to body and head size across lepidosaurs and, as predicted, larger species have the greatest bite-forces. Contrary to our other predictions, tooth attachment, diet and habit have little predictive power when accounting for size. Herbivores bite more forcefully simply because they are larger. Our results also highlight priorities for future sampling to further enhance our understanding of broader evolutionary patterns.

Comparative cranial biomechanics in two lizard species: impact of variation in cranial design.

Cranial morphology in lepidosaurs is highly disparate and characterised by the frequent loss or reduction of bony elements. In varanids and geckos, the loss of the postorbital bar is associated with changes in skull shape, but the mechanical principles underlying this variation remain poorly understood. Here, we sought to determine how the overall cranial architecture and the presence of the postorbital bar relate to the loading and deformation of the cranial bones during biting in lepidosaurs. Using computer-based simulation techniques, we compared cranial biomechanics in the varanid Varanus niloticus and the teiid Salvator merianae, two large, active foragers. The overall strain magnitude and distribution across the cranium were similar in the two species, despite lower strain gradients in V. niloticus In S. merianae, the postorbital bar is important for resistance of the cranium to feeding loads. The postorbital ligament, which in varanids partially replaces the postorbital bar, does not affect bone strain. Our results suggest that the reduction of the postorbital bar impaired neither biting performance nor the structural resistance of the cranium to feeding loads in V. niloticus Differences in bone strain between the two species might reflect demands imposed by feeding and non-feeding functions on cranial shape. Beyond variation in cranial bone strain related to species-specific morphological differences, our results reveal that similar mechanical behaviour is shared by lizards with distinct cranial shapes. Contrary to the situation in mammals, the morphology of the circumorbital region, calvaria and palate appears to be important for withstanding high feeding loads in these lizards.

Disparities in the analysis of morphological disparity.

Analyses of morphological disparity have been used to characterize and investigate the evolution of variation in the anatomy, function and ecology of organisms since the 1980s. While a diversity of methods have been employed, it is unclear whether they provide equivalent insights. Here, we review the most commonly used approaches for characterizing and analysing morphological disparity, all of which have associated limitations that, if ignored, can lead to misinterpretation. We propose best practice guidelines for disparity analyses, while noting that there can be no 'one-size-fits-all' approach. The available tools should always be used in the context of a specific biological question that will determine data and method selection at every stage of the analysis.

Colobops: a juvenile rhynchocephalian reptile (Lepidosauromorpha), not a diminutive archosauromorph with an unusually strong bite.

Correctly identifying taxa at the root of major clades or the oldest clade-representatives is critical for meaningful interpretations of evolution. A small, partially crushed skull from the Late Triassic (Norian) of Connecticut, USA, originally described as an indeterminate rhynchocephalian saurian, was recently named Colobops noviportensis and reinterpreted as sister to all remaining Rhynchosauria, one of the earliest and globally distributed groups of herbivorous reptiles. It was also interpreted as having an exceptionally reinforced snout and powerful bite based on an especially large supratemporal fenestra. Here, after a re-analysis of the original scan data, we show that the skull was strongly dorsoventrally compressed post-mortem, with most bones out of life position. The cranial anatomy is consistent with that of other rhynchocephalian lepidosauromorphs, not rhynchosaurs. The 'reinforced snout' region and the 'exceptionally enlarged temporal region' are preservational artefacts and not exceptional among clevosaurid rhynchocephalians. Colobops is thus not a key taxon for understanding diapsid feeding apparatus evolution.

Reproductive phenotype predicts adult bite-force performance in sex-reversed dragons (Pogona vitticeps).

Sex-related differences in morphology and behavior are well documented, but the relative contributions of genes and environment to these traits are less well understood. Species that undergo sex reversal, such as the central bearded dragon (Pogona vitticeps), offer an opportunity to better understand sexually dimorphic traits because sexual phenotypes can exist on different chromosomal backgrounds. Reproductively female dragons with a discordant sex chromosome complement (sex reversed), at least as juveniles, exhibit traits in common with males (e.g., longer tails and greater boldness). However, the impact of sex reversal on sexually dimorphic traits in adult dragons is unknown. Here, we investigate the effect of sex reversal on bite-force performance, which may be important in resource acquisition (e.g., mates and/or food). We measured body size, head size, and bite force of the three sexual phenotypes in a colony of captive animals. Among adults, we found that males (ZZm) bite more forcefully than either chromosomally concordant females (ZWf) or sex-reversed females (ZZf), and this difference is associated with having relatively larger head dimensions. Therefore, adult sex-reversed females, despite apparently exhibiting male traits as juveniles, do not develop the larger head and enhanced bite force of adult male bearded dragons. This pattern is further illustrated in the full sample by a lack of positive allometry of bite force in sex-reversed females that is observed in males. The results reveal a close association between reproductive phenotype and bite force performance, regardless of sex chromosome complement.

Evolution of cranial shape in a continental-scale evolutionary radiation of Australian lizards.

A defining character of adaptive radiations is the evolution of a diversity of morphological forms that are associated with the use of different habitats, following the invasion of vacant niches. Island adaptive radiations have been thoroughly investigated but continental scale radiations are more poorly understood. Here, we use 52 species of Australian agamid lizards and their Asian relatives as a model group, and employ three-dimensional geometric morphometrics to characterize cranial morphology and investigate whether variation in cranial shape reflects patterns expected from the ecological process of adaptive radiation. Phylogenetic affinity, evolutionary allometry, and ecological life habit all play major roles in the evolution of cranial shape in the sampled lizards. We find a significant association between cranial shapes and life habit. Our results are in line with the expectations of an adaptive radiation, and this is the first time detailed geometric morphometric analyses have been used to understand the selective forces that drove an adaptive radiation at a continental scale.

Digital dissection of the head of the rock dove (Columba livia) using contrast-enhanced computed tomography.

The rock dove (or common pigeon), Columba livia, is an important model organism in biological studies, including research focusing on head muscle anatomy, feeding kinematics, and cranial kinesis. However, no integrated computer-based biomechanical model of the pigeon head has yet been attempted. As an initial step towards achieving this goal, we present the first three-dimensional digital dissection of the pigeon head based on a contrast-enhanced computed tomographic dataset achieved using iodine potassium iodide as a staining agent. Our datasets enable us to visualize the skeletal and muscular anatomy, brain and cranial nerves, and major sense organs of the pigeon, including very small and fragile features, as well as maintaining the three-dimensional topology of anatomical structures. This work updates and supplements earlier anatomical work on this widely used laboratory organism. We resolve several key points of disagreement arising from previous descriptions of pigeon anatomy, including the precise arrangement of the external adductor muscles and their relationship to the posterior adductor. Examination of the eye muscles highlights differences between avian taxa and shows that pigeon eye muscles are more similar to those of a tinamou than they are to those of a house sparrow. Furthermore, we present our three-dimensional data as publicly accessible files for further research and education purposes. Digital dissection permits exceptional visualisation and will be a valuable resource for further investigations into the head anatomy of other bird species, as well as efforts to reconstruct soft tissues in fossil archosaurs.

Exceptional Disparity in Australian Agamid Lizards is a Possible Result of Arrival into Vacant Niche.

Australia provides abundant examples of continental-scale evolutionary radiations. The collision of two continental shelves around 30 Ma facilitated an influx of squamates and the subsequent squamate radiations resulted in high taxonomic diversity. The morphological disparity seen in these major squamate groups, however, remains underexplored. Here, we examine the major cranial proportions of over 1,000 specimens using 2D linear measurements to explicitly quantify the morphological disparity of Australian agamid lizards (Amphibolurinae) and compare it to that of agamid, acrodont, and iguanian clades from other parts of the world. Our results indicate the Australian Amphibolurinae have exceptionally high cranial disparity, and we suggest that this is linked to the relaxed selective environment that greeted the founders of Amphibolurinae when they first arrived in Australia. Anat Rec, 302:1536-1543, 2019. © 2019 American Association for Anatomy.

Changes in ontogenetic patterns facilitate diversification in skull shape of Australian agamid lizards.

BACKGROUND: Morphological diversity among closely related animals can be the result of differing growth patterns. The Australian radiation of agamid lizards (Amphibolurinae) exhibits great ecological and morphological diversity, which they have achieved on a continent-wide scale, in a relatively short period of time (30 million years). Amphibolurines therefore make an ideal study group for examining ontogenetic allometry. We used two-dimensional landmark-based geometric morphometric methods to characterise the postnatal growth patterns in cranial shape of 18 species of amphibolurine lizards and investigate the associations between cranial morphology, and life habit and phylogeny. RESULTS: For most amphibolurine species, juveniles share a similar cranial phenotype, but by adulthood crania are more disparate in shape and occupy different sub-spaces of the total shape space. To achieve this disparity, crania do not follow a common post-natal growth pattern; there are differences among species in both the direction and magnitude of change in morphospace. We found that these growth patterns among the amphibolurines are significantly associated with ecological life habits. The clade Ctenophorus includes species that undergo small magnitudes of shape change during growth. They have dorsoventrally deep, blunt-snouted skulls (associated with terrestrial lifestyles), and also dorsoventrally shallow skulls (associated with saxicolous lifestyles). The sister clade to Ctenophorus, which includes the bearded dragon (Pogona), frill-neck lizard (Chlamydosaurus), and long-nosed dragon (Gowidon), exhibit broad and robust post-orbital regions and differing snout lengths (mainly associated with scansorial lifestyles). CONCLUSIONS: Australian agamids show great variability in the timing of development and divergence of growth trajectories which results in a diversity of adult cranial shapes. Phylogenetic signal in cranial morphology appears to be largely overwritten by signals that reflect life habit. This knowledge about growth patterns and skull shape diversity in agamid lizards will be valuable for placing phylogenetic, functional and ecological studies in a morphological context.

Neutron scanning reveals unexpected complexity in the enamel thickness of an herbivorous Jurassic reptile.

Eilenodontines are one of the oldest radiation of herbivorous lepidosaurs (snakes, lizards and tuatara) characterized by batteries of wide teeth with thick enamel that bear mammal-like wear facets. Unlike most reptiles, eilenodontines have limited tooth replacement, making dental longevity particularly important to them. We use both X-ray and neutron computed tomography to examine a fossil tooth from the eilenodontine Eilenodon (Late Jurassic, USA). Of the two approaches, neutron tomography was more successful and facilitated measurements of enamel thickness and distribution. We find the enamel thickness to be regionally variable, thin near the cusp tip (0.10 mm) but thicker around the base (0.15-0.30 mm) and notably greater than that of other rhynchocephalians such as the extant Sphenodon (0.08-0.14 mm). The thick enamel in Eilenodon would permit greater loading, extend tooth lifespan and facilitate the establishment of wear facets that have sharp edges for orally processing plant material such as horsetails (Equisetum). The shape of the enamel dentine junction indicates that tooth development in Eilenodon and Sphenodon involved similar folding of the epithelium but different ameloblast activity.

The biomechanical role of the chondrocranium and sutures in a lizard cranium.

The role of soft tissues in skull biomechanics remains poorly understood. Not least, the chondrocranium, the portion of the braincase which persists as cartilage with varying degrees of mineralization. It also remains commonplace to overlook the biomechanical role of sutures despite evidence that they alter strain distribution. Here, we examine the role of both the sutures and the chondrocranium in the South American tegu lizard Salvator merianae We use multi-body dynamics analysis (MDA) to provide realistic loading conditions for anterior and posterior unilateral biting and a detailed finite element model to examine strain magnitude and distribution. We find that strains within the chondrocranium are greatest during anterior biting and are primarily tensile; also that strain within the cranium is not greatly reduced by the presence of the chondrocranium unless it is given the same material properties as bone. This result contradicts previous suggestions that the anterior portion (the nasal septum) acts as a supporting structure. Inclusion of sutures to the cranium model not only increases overall strain magnitudes but also leads to a more complex distribution of tension and compression rather than that of a beam under sagittal bending.

Bite force in the horned frog (Ceratophrys cranwelli) with implications for extinct giant frogs.

Of the nearly 6,800 extant frog species, most have weak jaws that play only a minor role in prey capture. South American horned frogs (Ceratophrys) are a notable exception. Aggressive and able to consume vertebrates their own size, these "hopping heads" use a vice-like grip of their jaws to restrain and immobilize prey. Using a longitudinal experimental design, we quantified the ontogenetic profile of bite-force performance in post-metamorphic Ceratophrys cranwelli. Regression slopes indicate positive allometric scaling of bite force with reference to head and body size, results that concur with scaling patterns across a diversity of taxa, including fish and amniotes (lizards, tuatara, turtles, crocodylians, rodents). Our recovered scaling relationship suggests that exceptionally large individuals of a congener (C. aurita) and extinct giant frogs (Beelzebufo ampinga, Late Cretaceous of Madagascar) probably could bite with forces of 500 to 2200 N, comparable to medium to large-sized mammalian carnivores.

Sesamoid bones in tuatara (Sphenodon punctatus) investigated with X-ray microtomography, and implications for sesamoid evolution in Lepidosauria.

Sesamoids bones are small intra-tendinous (or ligamentous) ossifications found near joints and are often variable between individuals. Related bones, lunulae, are found within the menisci of certain joints. Several studies have described sesamoids and lunulae in lizards and their close relatives (Squamata) as potentially useful characters in phylogenetic analysis, but their status in the extant outgroup to Squamata, tuatara (Sphenodon), remains unclear. Sphenodon is the only living rhynchocephalian, but museum specimens are valuable and difficult to replace. Here, we use non-destructive X-ray microtomography to investigate the distribution of sesamoids and lunulae in 19 Sphenodon specimens and trace the evolution of these bones in Lepidosauria (Rhynchocephalia + Squamata). We find adult Sphenodon to possess a sesamoid and lunula complement different from any known squamate, but also some variation within Sphenodon specimens. The penultimate phalangeal sesamoids and tibial lunula appear to mineralize prior to skeletal maturity, followed by mineralization of a sesamoid between metatarsal I and the astragalocalcaneum (MTI-AC), the palmar sesamoids, and tibiofemoral lunulae around attainment of skeletal maturity. The tibial patella, ulnar, and plantar sesamoids mineralize late in maturity or variably. Ancestral state reconstruction indicates that the ulnar patella and tibiofemoral lunulae are synapomophies of Squamata, and the palmar sesamoid, tibial patella, tibial lunula, and MTI-AC may be synapomorphies of Lepidosauria. J. Morphol. 278:62-72, 2017. ©© 2016 Wiley Periodicals,Inc.

Anatomy, morphology and evolution of the patella in squamate lizards and tuatara (Sphenodon punctatus).

The patella (kneecap) is the largest and best-known of the sesamoid bones, postulated to confer biomechanical advantages including increasing joint leverage and reinforcing the tendon against compression. It has evolved several times independently in amniotes, but despite apparently widespread occurrence in lizards, the patella remains poorly characterised in this group and is, as yet, completely undescribed in their nearest extant relative Sphenodon (Rhynchocephalia). Through radiography, osteological and fossil studies we examined patellar presence in diverse lizard and lepidosauromorph taxa, and using computed tomography, dissection and histology we investigated in greater depth the anatomy and morphology of the patella in 16 lizard species and 19 Sphenodon specimens. We have found the first unambiguous evidence of a mineralised patella in Sphenodon, which appears similar to the patella of lizards and shares several gross and microscopic anatomical features. Although there may be a common mature morphology, the squamate patella exhibits a great deal of variability in development (whether from a cartilage anlage or not, and in the number of mineralised centres) and composition (bone, mineralised cartilage or fibrotendinous tissue). Unlike in mammals and birds, the patella in certain lizards and Sphenodon appears to be a polymorphic trait. We have also explored the evolution of the patella through ancestral state reconstruction, finding that the patella is ancestral for lizards and possibly Lepidosauria as a whole. Clear evidence of the patella in rhynchocephalian or stem lepidosaurian fossil taxa would clarify the evolutionary origin(s) of the patella, but due to the small size of this bone and the opportunity for degradation or loss we could not definitively conclude presence or absence in the fossils examined. The pattern of evolution in lepidosaurs is unclear but our data suggest that the emergence of this sesamoid may be related to the evolution of secondary ossification centres and/or changes in knee joint conformation, where enhancement of extensor muscle leverage would be more beneficial.

Reliable quantification of bite-force performance requires use of appropriate biting substrate and standardization of bite out-lever.

Bite-force performance is an ecologically important measure of whole-organism performance that shapes dietary breadth and feeding strategies and, in some taxa, determines reproductive success. It also is a metric that is crucial to testing and evaluating biomechanical models. We reviewed nearly 100 published studies of a range of taxa that incorporate direct in vivo measurements of bite force. Problematically, methods of data collection and processing vary considerably among studies. In particular, there is little consensus on the appropriate substrate to use on the biting surface of force transducers. In addition, the bite out-lever, defined as the distance from the fulcrum (i.e. jaw joint) to the position along the jawline at which the jaws engage the transducer, is rarely taken into account. We examined the effect of bite substrate and bite out-lever on bite-force estimates in a diverse sample of lizards. Results indicate that both variables have a significant impact on the accuracy of measurements. Maximum bite force is significantly greater using leather as the biting substrate compared with a metal substrate. Less-forceful bites on metal are likely due to inhibitory feedback from mechanoreceptors that prevent damage to the feeding apparatus. Standardization of bite out-lever affected which trial produced maximum performance for a given individual. Indeed, maximum bite force is usually underestimated without standardization because it is expected to be greatest at the minimum out-lever (i.e. back of the jaws), which in studies is rarely targeted with success. We assert that future studies should use a pliable substrate, such as leather, and use appropriate standardization for bite out-lever.