Patterns of girdle shape and their correlates in Australian limb-reduced skinks.

The evolution of limb reduction in squamates is a classic example of convergence, but the skeletal morphological patterns associated with it are underexplored. To provide insights on the biomechanical and developmental consequences of transitions to limb reduction, we use geometric morphometrics to examine the morphology of pectoral and pelvic girdles in 90 species of limb-reduced skinks and their fully limbed relatives. Clavicle shapes converge towards an acute anterior bend when forelimbs are lost but hindlimbs are retained-a morphology typical of sand-swimmers. This may either indicate functional adaptations to locomotion in fine substrates, or a developmental consequence of complete limb loss. The shape of limb-bearing elements of both girdles (coracoid and pelvis) instead closely mirrors limb reduction, becoming more simplified as undulation replaces limbed locomotion. Integration between girdles decreases in taxa lacking elements of the forelimbs but not hindlimbs, indicating differential selection on each girdle in response to distinct locomotory strategies. However, this pattern becomes less clear when considering phylogenetic history, perhaps because it is limited to one specific clade (Lerista). We show how the functional demands of locomotion can induce changes at different levels of organismal organization, including both external and internal structures.

An Integrative Approach Using Phylogenomics and High-Resolution X-Ray Computed Tomography for Species Delimitation in Cryptic Taxa.

Morphologically cryptic taxa have proved to be a long-standing challenge for taxonomists. Lineages that show strong genomic structuring across the landscape but are phenotypically similar pose a conundrum, with traditional morphological analyses of these cryptic lineages struggling to keep up with species delimitation advances. Micro X-ray computed tomography (CT) combined with geometric morphometric analyses provides a promising avenue for identification of morphologically cryptic taxa, given its ability to detect subtle differences in anatomical structures. However, this approach has yet to be used in combination with genomic data in a comparative analytical framework to distinguish cryptic taxa. We present an integrative approach incorporating genomic and geometric morphometric evidence to assess the species delimitation of grassland earless dragons (Tympanocryptis spp.) in north-eastern Australia. Using mitochondrial and nuclear genes (ND2 and RAG1, respectively), along with $>$8500 SNPs (nuclear single nucleotide polymorphisms), we assess the evolutionary independence of target lineages and several closely related species. We then integrate phylogenomic data with osteological cranial variation between lineages using landmark-based analyses of three-dimensional CT models. High levels of genomic differentiation between the three target lineages were uncovered, also supported by significant osteological differences. By incorporating multiple lines of evidence, we provide strong support for three undescribed cryptic lineages of Tympanocryptis in north-eastern Australia that warrant taxonomic review. Our approach demonstrates the successful application of CT with integrative taxonomic approaches for cryptic species delimitation, which is broadly applicable across vertebrates containing morphologically similar yet genetically distinct lineages. Additionally, we provide a review of recent integrative taxonomic approaches for cryptic species delimitation and an assessment of how our approach can value-add to taxonomic research.

Eocene lizard from Germany reveals amphisbaenian origins.

Amphisbaenia is a speciose clade of fossorial lizards characterized by a snake-like body and a strongly reinforced skull adapted for head-first burrowing. The evolutionary origins of amphisbaenians are controversial, with molecular data uniting them with lacertids, a clade of Old World terrestrial lizards, whereas morphology supports a grouping with snakes and other limbless squamates. Reports of fossil stem amphisbaenians have been falsified, and no fossils have previously tested these competing phylogenetic hypotheses or shed light on ancestral amphisbaenian ecology. Here we report the discovery of a new lacertid-like lizard from the Eocene Messel locality of Germany that provides the first morphological evidence for lacertid-amphisbaenian monophyly on the basis of a reinforced, akinetic skull roof and braincase, supporting the view that body elongation and limblessness in amphisbaenians and snakes evolved independently. Morphometric analysis of body shape and ecology in squamates indicates that the postcranial anatomy of the new taxon is most consistent with opportunistically burrowing habits, which in combination with cranial reinforcement indicates that head-first burrowing evolved before body elongation and may have been a crucial first step in the evolution of amphisbaenian fossoriality.

Skull osteology of the Eocene amphisbaenian Spathorhynchus fossorium (Reptilia, Squamata) suggests convergent evolution and reversals of fossorial adaptations in worm lizards.

The fossorial amphisbaenians, or worm lizards, are characterized by a suite of specialized characters in the skull and postcranium, however fossil evidence suggests that at least some of these shared derived traits evolved convergently. Unfortunately the lack of detailed knowledge of many fossil taxa has rendered a more precise interpretation difficult. Here we describe the cranial anatomy of the oldest-known well-preserved amphisbaenian, Spathorhynchus fossorium, from the Eocene Green River Formation, Wyoming, USA, using high-resolution X-ray computed tomography (HRXCT). This taxon possesses one of the most strongly reinforced crania known among amphisbaenians, with many dermal bones overlapping each other internally. In contrast to modern taxa, S. fossorium has a paired orbitosphenoid, lacks a true compound bone in the mandible, and possesses a fully enclosed orbital rim. The last feature represents a highly derived structure in that the jugal establishes contact with the frontal internally, reinforcing the posterior orbital margin. S. fossorium also possesses a strongly modified Vidian canal with a previously unknown connection to the ventral surface of the parabasisphenoid. Comparison with the closely related fossil taxon Dyticonastis rensbergeri reveals that these derived traits are also shared by the latter species and potentially represent synapopmorphies of an extinct Paleogene clade of amphisbaenians. The presence of a reinforced orbital rim suggests selection against the loss of a functional eye and indicates an ecology potentially different from modern taxa. Given the currently accepted phylogenetic position of Spathorhynchus and Dyticonastis, we predict that supposedly 'unique' cranial traits traditionally linked to fossoriality such as a fused orbitosphenoid and the reduction of the eye show a more complex character history than previously assumed, including both parallel evolution and reversals to superficially primitive conditions.

Frequency-dependent and correlational selection pressures have conflicting consequences for assortative mating in a color-polymorphic lizard, Uta stansburiana.

Genetically determined polymorphisms incorporating multiple traits can persist in nature under chronic, fluctuating, and sometimes conflicting selection pressures. Balancing selection among morphs preserves equilibrium frequencies, while correlational selection maintains favorable trait combinations within each morph. Under negative frequency-dependent selection, females should mate (often disassortatively) with rare male morphotypes to produce conditionally fit offspring. Conversely, under correlational selection, females should mate assortatively to preserve coadapted gene complexes and avoid ontogenetic conflict. Using controlled breeding designs, we evaluated consequences of assortative mating patterns in color-polymorphic side-blotched lizards (Uta stansburiana), to identify conflict between these sources of selection. Females who mated disassortatively, and to conditionally high-quality males in the context of frequency-dependent selection, experienced highest fertility rates. In contrast, assortatively mated females experienced higher fetal viability rates. The trade-off between fertility and egg viability resulted in no overall fitness benefit to either assortative or disassortative mating patterns. These results suggest that ongoing conflict between correlational and frequency dependent selection in polymorphic populations may generate a trade-off between rare-morph advantage and phenotypic integration and between assortative and disassortative mating decisions. More generally, interactions among multiple sources of diversity-promoting selection can alter adaptations and dynamics predicted to arise under any of these regimes alone.

Morphological disparity opposes latitudinal diversity gradient in lacertid lizards.

While global variation in taxonomic diversity is strongly linked to latitude, the extent to which morphological disparity follows geographical gradients is less well known. We estimated patterns of lineage diversification, morphological disparity and rates of phenotypic evolution in the Old World lizard family Lacertidae, which displays a nearly inverse latitudinal diversity gradient with decreasing species richness towards the tropics. We found that lacertids exhibit relatively constant rates of lineage accumulation over time, although the majority of morphological variation appears to have originated during recent divergence events, resulting in increased partitioning of disparity within subclades. Among subclades, tropical arboreal taxa exhibited the fastest rates of shape change while temperate European taxa were the slowest, resulting in an inverse relationship between latitudinal diversity and rates of phenotypic evolution. This pattern demonstrates a compelling counterexample to the ecological opportunity theory of diversification, suggesting an uncoupling of the processes generating species diversity and morphological differentiation across spatial scales.

Comparative skull osteology of Amphisbaena arda and Amphisbaena vermicularis (Squamata: Amphisbaenidae).

The skull anatomy of amphisbaenians directly influences their capacity to burrow and is crucial for the study of their systematics, which ultimately contributes to our comprehension of their evolution and ecology. In this study, we employed three-dimensional X-ray computed tomography to provide a detailed description and comprehensive comparison of the skull anatomy of two amphisbaenian species with similar external morphology, Amphisbaena arda and Amphisbaena vermicularis. Our findings revealed some differences between the species, especially in the sagittal crest of the parietal bone, the ascendant process, and the transverse occipital crest of the occipital complex. We also found intraspecific variation within A. vermicularis, with some specimens displaying morphology that differed from their conspecifics but not from A. arda. The observed intraspecific variation within A. vermicularis cannot be attributed to soil features because all specimens came from the same locality. Specimen size and soil type may play a role in the observed differences between A. arda and A. vermicularis, as the single A. arda specimen is the largest of our sample and soil type and texture differ between the collection sites of the two species.

Integration of molecules and new fossils supports a Triassic origin for Lepidosauria (lizards, snakes, and tuatara).

BACKGROUND: Lepidosauria (lizards, snakes, tuatara) is a globally distributed and ecologically important group of over 9,000 reptile species. The earliest fossil records are currently restricted to the Late Triassic and often dated to 227 million years ago (Mya). As these early records include taxa that are relatively derived in their morphology (e.g. Brachyrhinodon), an earlier unknown history of Lepidosauria is implied. However, molecular age estimates for Lepidosauria have been problematic; dates for the most recent common ancestor of all lepidosaurs range between approximately 226 and 289 Mya whereas estimates for crown-group Squamata (lizards and snakes) vary more dramatically: 179 to 294 Mya. This uncertainty restricts inferences regarding the patterns of diversification and evolution of Lepidosauria as a whole. RESULTS: Here we report on a rhynchocephalian fossil from the Middle Triassic of Germany (Vellberg) that represents the oldest known record of a lepidosaur from anywhere in the world. Reliably dated to 238-240 Mya, this material is about 12 million years older than previously known lepidosaur records and is older than some but not all molecular clock estimates for the origin of lepidosaurs. Using RAG1 sequence data from 76 extant taxa and the new fossil specimens two of several calibrations, we estimate that the most recent common ancestor of Lepidosauria lived at least 242 Mya (238-249.5), and crown-group Squamata originated around 193 Mya (176-213). CONCLUSION: A Early/Middle Triassic date for the origin of Lepidosauria disagrees with previous estimates deep within the Permian and suggests the group evolved as part of the faunal recovery after the end-Permain mass extinction as the climate became more humid. Our origin time for crown-group Squamata coincides with shifts towards warmer climates and dramatic changes in fauna and flora. Most major subclades within Squamata originated in the Cretaceous postdating major continental fragmentation. The Vellberg fossil locality is expected to become an important resource for providing a more balanced picture of the Triassic and for bridging gaps in the fossil record of several other major vertebrate groups.

Molecular, morphological and osteological differentiation of a new species of microhylid frog of the genus Stumpffia from northwestern Madagascar.

We describe a new small-sized microhylid frog of the genus Stumpffia and provide a preliminary survey of Stumpffia diversity in northern and northwestern Madagascar based on molecular and morphological data. Analysis of 68 previously published and 142 newly generated DNA sequences of a fragment of the mitochondrial 16S rRNA gene revealed a large genetic diversity within Stumpffia, with numerous strongly differentiated lineages occurring across Madagascar. The anal. ysis confirmed a clade formed exclusively by Stumpffia species occurring in north/northwestern Madagascar and contain. ing almost all species from this area. Based on DNA sequences of the RAG1 gene, we further assess that most of the deep mitochondrial lineages do not share haplotypes in this nuclear marker, even in the few cases of sympatry, which we interpret as confirmation of their status as independent evolutionary lineages. Micro-computed tomography revealed subtle differences in phalangeal formulae among some of these candidate species, providing additional taxonomic characters for future revisions of the genus. The newly described species Stumpffia analamaina sp. nov. occurs 27 km north-west of Antsohihy and is at present the southernmost confirmed Stumpffia of the north/northwestern clade. It is mainly characterized by a snout-vent length of 10-12 mm, advertisement calls consisting of a series of short melodious chirps, manus with four fingers and pes with five toes, with only slight length reduction of first finger and toe. It furthermore does not share haplotypes with any other Stumpffia species in 16S or RAG1, and has relatively longer hands and feet compared with other small-sized Stumpffia of the north/northwestern clade such as S. madagascariensis and S. pygmaea. The new species lives in an area of dry forest where it was found in leaf litter along a small stream bordered by degraded gallery vegetation. Due to the limited distributions of almost all species in the north/northwestern Stumpffia clade, the rareness of appropriate microhabitat around the type locality of S. analamaina sp. nov. and the rampant habitat destruction in this part of Madagascar, we propose a threat status of Critically Endangered for this new species.

2023 BMC Ecology and Evolution image competition: the winning images.

In 2023, researchers from around the world entered the BMC Ecology and Evolution photography competition. As a result, we received a spectacular collection of photographs that capture the wonder of nature, those looking to understand it and glimpses into long lost worlds. This editorial celebrates the winning images selected by the Editor of BMC Ecology and Evolution and senior members of the journal's editorial board.

2022 BMC Ecology and Evolution image competition: the winning images.

In 2022, researchers from around the world entered the BMC Ecology and Evolution photography competition. The contest produced a spectacular collection of photographs that capture the wonder of the natural world and the growing need to protect it as the human impact on the planet intensifies. This editorial celebrates the winning images selected by the Editor of BMC Ecology and Evolution and senior members of the journal's editorial board.

A farewell to arms and legs: a review of limb reduction in squamates.

Elongated snake-like bodies associated with limb reduction have evolved multiple times throughout vertebrate history. Limb-reduced squamates (lizards and snakes) account for the vast majority of these morphological transformations, and thus have great potential for revealing macroevolutionary transitions and modes of body-shape transformation. Here we present a comprehensive review on limb reduction, in which we examine and discuss research on these dramatic morphological transitions. Historically, there have been several approaches to the study of squamate limb reduction: (i) definitions of general anatomical principles of snake-like body shapes, expressed as varying relationships between body parts and morphometric measurements; (ii) framing of limb reduction from an evolutionary perspective using morphological comparisons; (iii) defining developmental mechanisms involved in the ontogeny of limb-reduced forms, and their genetic basis; (iv) reconstructions of the evolutionary history of limb-reduced lineages using phylogenetic comparative methods; (v) studies of functional and biomechanical aspects of limb-reduced body shapes; and (vi) studies of ecological and biogeographical correlates of limb reduction. For each of these approaches, we highlight their importance in advancing our understanding, as well as their weaknesses and limitations. Lastly, we provide suggestions to stimulate further studies, in which we underscore the necessity of widening the scope of analyses, and of bringing together different perspectives in order to understand better these morphological transitions and their evolution. In particular, we emphasise the importance of investigating and comparing the internal morphology of limb-reduced lizards in contrast to external morphology, which will be the first step in gaining a deeper insight into body-shape variation.

Ontogenetic origins of cranial convergence between the extinct marsupial thylacine and placental gray wolf.

Phenotypic convergence, describing the independent evolution of similar characteristics, offers unique insights into how natural selection influences developmental and molecular processes to generate shared adaptations. The extinct marsupial thylacine and placental gray wolf represent one of the most extraordinary cases of convergent evolution in mammals, sharing striking cranial similarities despite 160 million years of independent evolution. We digitally reconstructed their cranial ontogeny from birth to adulthood to examine how and when convergence arises through patterns of allometry, mosaicism, modularity, and integration. We find the thylacine and wolf crania develop along nearly parallel growth trajectories, despite lineage-specific constraints and heterochrony in timing of ossification. These constraints were found to enforce distinct cranial modularity and integration patterns during development, which were unable to explain their adult convergence. Instead, we identify a developmental origin for their convergent cranial morphologies through patterns of mosaic evolution, occurring within bone groups sharing conserved embryonic tissue origins. Interestingly, these patterns are accompanied by homoplasy in gene regulatory networks associated with neural crest cells, critical for skull patterning. Together, our findings establish empirical links between adaptive phenotypic and genotypic convergence and provides a digital resource for further investigations into the developmental basis of mammalian evolution.

Postnatal development in a marsupial model, the fat-tailed dunnart (Sminthopsis crassicaudata; Dasyuromorphia: Dasyuridae).

Marsupials exhibit unique biological features that provide fascinating insights into many aspects of mammalian development. These include their distinctive mode of reproduction, altricial stage at birth, and the associated heterochrony that is required for their crawl to the pouch and teat attachment. Marsupials are also an invaluable resource for mammalian comparative biology, forming a distinct lineage from the extant placental and egg-laying monotreme mammals. Despite their unique biology, marsupial resources are lagging behind those available for placentals. The fat-tailed dunnart (Sminthopsis crassicaudata) is a laboratory based marsupial model, with simple and robust husbandry requirements and a short reproductive cycle making it amenable to experimental manipulations. Here we present a detailed staging series for the fat-tailed dunnart, focusing on their accelerated development of the forelimbs and jaws. This study provides the first skeletal developmental series on S. crassicaudata and provides a fundamental resource for future studies exploring mammalian diversification, development and evolution.

Author Correction: High-throughput microCT scanning of small specimens: preparation, packing, parameters and post-processing.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

High-throughput microCT scanning of small specimens: preparation, packing, parameters and post-processing.

High-resolution X-ray microcomputed tomography, or microCT (µCT), enables the digital imaging of whole objects in three dimensions. The power of µCT to visualize internal features without disarticulation makes it particularly valuable for the study of museum collections, which house millions of physical specimens documenting the spatio-temporal patterns of life. Despite the potential for comparative analyses, most µCT studies include limited numbers of museum specimens, due to the challenges of digitizing numerous individuals within a project scope. Here we describe a method for high-throughput µCT scanning of hundreds of small (< 2 cm) specimens in a single container, followed by individual labelling and archival storage. We also explore the effects of various packing materials and multiple specimens per capsule to minimize sample movement that can degrade image quality, and hence µCT investment. We demonstrate this protocol on vertebrate fossils from Queensland Museum, Australia, as part of an effort to track community responses to climate change over evolutionary time. This system can be easily modified for other types of wet and dry material amenable to X-ray attenuation, including geological, botanical and zoological samples, providing greater access to large-scale phenotypic data and adding value to global collections.

Integration of Bayesian molecular clock methods and fossil-based soft bounds reveals early Cenozoic origin of African lacertid lizards.

BACKGROUND: Although current molecular clock methods offer greater flexibility in modelling evolutionary events, calibration of the clock with dates from the fossil record is still problematic for many groups. Here we implement several new approaches in molecular dating to estimate the evolutionary ages of Lacertidae, an Old World family of lizards with a poor fossil record and uncertain phylogeny. Four different models of rate variation are tested in a new program for Bayesian phylogenetic analysis called TreeTime, based on a combination of mitochondrial and nuclear gene sequences. We incorporate paleontological uncertainty into divergence estimates by expressing multiple calibration dates as a range of probabilistic distributions. We also test the reliability of our proposed calibrations by exploring effects of individual priors on posterior estimates. RESULTS: According to the most reliable model, as indicated by Bayes factor comparison, modern lacertids arose shortly after the K/T transition and entered Africa about 45 million years ago, with the majority of their African radiation occurring in the Eocene and Oligocene. Our findings indicate much earlier origins for these clades than previously reported, and we discuss our results in light of paleogeographic trends during the Cenozoic. CONCLUSION: This study represents the first attempt to estimate evolutionary ages of a specific group of reptiles exhibiting uncertain phylogenetic relationships, molecular rate variation and a poor fossil record. Our results emphasize the sensitivity of molecular divergence dates to fossil calibrations, and support the use of combined molecular data sets and multiple, well-spaced dates from the fossil record as minimum node constraints. The bioinformatics program used here, TreeTime, is publicly available, and we recommend its use for molecular dating of taxa faced with similar challenges.

Integrating phylogeography and high-resolution X-ray CT reveals five new cryptic species and multiple hybrid zones among Australian earless dragons.

Cryptic lineages, comprising species complexes with deep genetic structuring across the landscape but without distinct morphological differences, impose substantial difficulties for systematists and taxonomists in determining true species diversity. Here, we present an integrative approach that combines data from phylogeography and geometric morphometric analyses of three-dimensional cranial models to revisit the uncertain taxonomy of earless dragons from southern and central Australia that at one time or another have been included under the name Tympanocryptis lineata. Our approach finds strong support for seven previously described species, and more importantly, five undescribed Tympanocryptis taxa for which we provide a taxonomic treatment. We also find evidence of introgression and hybridization in three discrete contact zones between lineages, supported by mitochondrial and nuclear genes, as well as morphological analyses. With a sampling design that includes at least five individuals for each genetic lineage with corresponding X-ray microcomputed tomography scans, we perform comparative evolutionary analyses to show that there is a significant phylogenetic signal in Tympanocryptis cranial shape. Our results demonstrate the importance of using multiple specimens in each genetic lineage, particularly in cases of potential hybridization, and that geometric morphometrics, when used in an integrative framework, is a powerful tool in species delimitation across cryptic lineages. Our results lay the groundwork for future evolutionary studies in this widespread group across multiple environmental types and identify several species of immediate conservation concern with a focus on T. petersi sp. nov. We suggest that this species has undergone significant population declines and warrants a full conservation assessment.

Letting the 'cat' out of the bag: pouch young development of the extinct Tasmanian tiger revealed by X-ray computed tomography.

The Tasmanian tiger or thylacine (Thylacinus cynocephalus) was an iconic Australian marsupial predator that was hunted to extinction in the early 1900s. Despite sharing striking similarities with canids, they failed to evolve many of the specialized anatomical features that characterize carnivorous placental mammals. These evolutionary limitations are thought to arise from functional constraints associated with the marsupial mode of reproduction, in which otherwise highly altricial young use their well-developed forelimbs to climb to the pouch and mouth to suckle. Here we present the first three-dimensional digital developmental series of the thylacine throughout its pouch life using X-ray computed tomography on all known ethanol-preserved specimens. Based on detailed skeletal measurements, we refine the species growth curve to improve age estimates for the individuals. Comparison of allometric growth trends in the appendicular skeleton (fore- and hindlimbs) with that of other placental and marsupial mammals revealed that despite their unique adult morphologies, thylacines retained a generalized early marsupial ontogeny. Our approach also revealed mislabelled specimens that possessed large epipubic bones (vestigial in thylacine) and differing vertebral numbers. All of our generated CT models are publicly available, preserving their developmental morphology and providing a novel digital resource for future studies of this unique marsupial.