The consequences of calcium: investigating intracortical reproductive signals in the American alligator for sex determination.

Identifying sex in extinct archosaurs has proven difficult due, in part, to low sample sizes, preservation biases, and methodology. While previous studies have largely focused on morphological traits, here we investigate intracortical signals of egg-shelling in extant alligators. Egg-shelling requires large mobilizations of calcium reserves. Aves utilize medullary tissue as a calcium reserve, whereas crocodylians mobilize calcium from cortical bone or osteoderms. If crocodylians derive calcium from bone cortices for egg-shelling, then egg-shelling events should be detectable in female crocodylian cortical bone. We examined mid-diaphyseal Alligator mississippiensis femoral bone cross-sections for signals of reproduction. Compaction and area of resorbed tissue were measured in femoral cross-sections from captive raised male (n = 10) and female (n = 29) A. mississippiensis of 26-27 years at age of death. This sample is more robust than previous studies, though reproductive history data is unknown. Femora from a small sample of wild caught male (n = 6) and female (n = 6) A. mississippiensis were also measured. Data were analyzed by pairwise t-tests between sex and captivity status. There was no significant difference in either compaction or resorbed tissue values between male and female alligators, regardless of habitat (wild or captive-raised). A reproductive signal was undetectable in this study and any quantifiable differences between sexes appears to be driven by size dimorphism. Cortical resorption rates in the femora of male and female alligators are reflective of normal aging processes and not indicative of egg-shelling during reproduction. Examination of younger alligators would clarify processes driving bone turnover during reproductively active years.

The dinosaurs that weren't: osteohistology supports giant ichthyosaur affinity of enigmatic large bone segments from the European Rhaetian.

Very large unidentified elongate and rounded fossil bone segments of uncertain origin recovered from different Rhaetian (Late Triassic) fossil localities across Europe have been puzzling the paleontological community since the second half of the 19th century. Different hypotheses have been proposed regarding the nature of these fossils: (1) giant amphibian bones, (2) dinosaurian or other archosaurian long bone shafts, and (3) giant ichthyosaurian jaw bone segments. We call the latter proposal the 'Giant Ichthyosaur Hypothesis' and test it using bone histology. In presumable ichthyosaur specimens from SW England (Lilstock), France (Autun), and indeterminate cortical fragments from Germany (Bonenburg), we found a combination of shared histological features in the periosteal cortex: an unusual woven-parallel complex of strictly longitudinal primary osteons set in a novel woven-fibered matrix type with intrinsic coarse collagen fibers (IFM), and a distinctive pattern of Haversian substitution in which secondary osteons often form within primary ones. The splenial and surangular of the holotype of the giant ichthyosaur Shastasaurus sikanniensis from Canada were sampled for comparison. The results of the sampling indicate a common osteohistology with the European specimens. A broad histological comparison is provided to reject alternative taxonomic affinities aside from ichthyosaurs of the very large bone segment. Most importantly, we highlight the occurrence of shared peculiar osteogenic processes in Late Triassic giant ichthyosaurs, reflecting special ossification strategies enabling fast growth and achievement of giant size and/or related to biomechanical properties akin to ossified tendons.

Lipids in the American Alligator stratum corneum provide insights into the evolution of vertebrate skin.

In terrestrial vertebrates, the outermost layer of the skin, the stratum corneum (SC), provides a durable and flexible interface with the environment and is comprised of corneocytes embedded in lipids. However, the morphology and lipid composition of the SC varies throughout evolutionary history. Because crocodilians and birds phylogenetically bracket the Archosaurian clade, lipid composition in crocodilian SC may be compared with that of birds and other vertebrates to make inferences about broader phylogenetic patterns within Archosaurs while highlighting adaptations in vertebrate skin. We identified and quantified lipid classes in the SC of the American Alligator (Alligator mississippiensis) from three skin regions varying in mobility. Our results find similarities in lipid composition between alligator and avian SC, including a high percentage of cerebrosides, a polar lipid previously found only in the SC of birds and bats. Furthermore, polar lipids were more abundant in the most mobile region of the SC. Because polar lipids bind with water to increase skin hydration and therefore its pliability under physical stress, we hypothesize that selection for lipids in Archosaurian SC was driven by the unique distribution of proteins in the SC of this clade, and cerebrosides may have served as pre-adaptations for flight.

Discovery of facultative parthenogenesis in a new world crocodile.

Over the past two decades, there has been an astounding growth in the documentation of vertebrate facultative parthenogenesis (FP). This unusual reproductive mode has been documented in birds, non-avian reptiles-specifically lizards and snakes-and elasmobranch fishes. Part of this growth among vertebrate taxa is attributable to awareness of the phenomenon itself and advances in molecular genetics/genomics and bioinformatics, and as such our understanding has developed considerably. Nonetheless, questions remain as to its occurrence outside of these vertebrate lineages, most notably in Chelonia (turtles) and Crocodylia (crocodiles, alligators and gharials). The latter group is particularly interesting because unlike all previously documented cases of FP in vertebrates, crocodilians lack sex chromosomes and sex determination is controlled by temperature. Here, using whole-genome sequencing data, we provide, to our knowledge, the first evidence of FP in a crocodilian, the American crocodile, Crocodylus acutus. The data support terminal fusion automixis as the reproductive mechanism; a finding which suggests a common evolutionary origin of FP across reptiles, crocodilians and birds. With FP now documented in the two main branches of extant archosaurs, this discovery offers tantalizing insights into the possible reproductive capabilities of the extinct archosaurian relatives of crocodilians and birds, notably members of Pterosauria and Dinosauria.

Reconstruction of soft noncontractile tissue in the derived therizinosaur Nothronychus: The interplay of soft tissue and stress on hindlimb ossification and posture.

Nothronychus graffami was a large therizinosaur from the Upper Cretaceous of North America. Much of the skeleton is well-preserved and relatively undistorted. The synovial capsule, extracapsular, and intracapsular tendons are reconstructed in N. graffami using existing scars and comparison with the hips of extant theropods and models of extinct theropods. The iliofemoral, pubofemoral, and ischiofemoral ligaments are all modeled. Soft tissue, especially the ischiofemoral and pubofemoral ligaments reduced possible protraction/retraction of the femur at the hip while stabilizing the joint. Therefore, most hindlimb movements took place at the knee. Weight-bearing function in Nothronychus was transferred from the supra-acetabular crest to the pubic peduncle and associated labrum at rest. The femur possessed an intermediately angled neck, convergent with titanosaurs. Therefore, some lateral abduction is proposed, requiring a well-developed meniscus at the knee. Such a posture would result in considerable mediolateral stress along the femoral shaft, resulting in increased transverse ossification. The femur is considered somewhat abducted from the midline, especially when maximally pronated, as the preacetabular ala is more extensive than the post-acetabular ala. This trait would probably result in a laterally divergent femur, as in many birds, resulting in a broad-gauge trackway with wider separation between left and right footprints than observed in many theropod trackways. Limb ratios and ossification patterns suggest a slow, waddling gait. A standard digitiform pose is reconstructed as is common for theropods, but a plantigrade stance cannot be firmly rejected.

Inner workings of the alligator ankle reveal the mechanistic origins of archosaur locomotor diversity.

Major transformations in the locomotor system of archosaurs (a major clade of reptiles including birds, crocodiles, dinosaurs, and pterosaurs) were accompanied by significant modifications to ankle anatomy. How the evolution of such a complex multi-joint structure is related to shifts in ankle function and locomotor diversity across this clade remains unclear and weakly grounded in extant experimental data. Here, we used X-ray Reconstruction of Moving Morphology to reconstruct skeletal motion and quantify the sources of three-dimensional ankle mobility in the American alligator, a species that retains the ancestral archosaur ankle structure. We then applied the observed relationships between joint excursion and locomotor behaviors to predict ankle function in extinct archosaurs. High-resolution reconstructions of Alligator skeletal movement revealed previously unseen regionalized coordination among joints responsible for overall ankle rotation. Differences in joint contributions between maneuvers and steady walking parallel transitions in mobility inferred from the ankle structure of fossil taxa in lineages with more erect hind limb postures. Key ankle structures related to ankle mobility were identified in the alligator, which permitted the characterization of ancestral archosaur ankle function. Modifications of these structures provide morphological evidence for functional convergence among sublineages of bird-line and crocodylian-line archosaurs. Using the dynamic insight into the internal sources of Alligator ankle mobility and trends among locomotor modes, we trace anatomical shifts and propose a mechanistic hypothesis for the evolution of ankle structure and function across Archosauria.

Evolution and molecular basis of a novel allosteric property of crocodilian hemoglobin.

The extraordinary breath-hold diving capacity of crocodilians has been ascribed to a unique mode of allosterically regulating hemoglobin (Hb)-oxygenation in circulating red blood cells. We investigated the origin and mechanistic basis of this novel biochemical phenomenon by performing directed mutagenesis experiments on resurrected ancestral Hbs. Comparisons of Hb function between the common ancestor of archosaurs (the group that includes crocodilians and birds) and the last common ancestor of modern crocodilians revealed that regulation of Hb-O2 affinity via allosteric binding of bicarbonate ions represents a croc-specific innovation that evolved in combination with the loss of allosteric regulation by ATP binding. Mutagenesis experiments revealed that evolution of the novel allosteric function in crocodilians and the concomitant loss of ancestral function were not mechanistically coupled and were caused by different sets of substitutions. The gain of bicarbonate sensitivity in crocodilian Hb involved the direct effect of few amino acid substitutions at key sites in combination with indirect effects of numerous other substitutions at structurally disparate sites. Such indirect interaction effects suggest that evolution of the novel protein function was conditional on neutral mutations that produced no adaptive benefit when they first arose but that contributed to a permissive background for subsequent function-altering mutations at other sites. Due to the context dependence of causative substitutions, the unique allosteric properties of crocodilian Hb cannot be easily transplanted into divergent homologs of other species.

Walking-and Running and Jumping-with Dinosaurs and Their Cousins, Viewed Through the Lens of Evolutionary Biomechanics.

Archosauria diversified throughout the Triassic Period before experiencing two mass extinctions near its end ∼201 Mya, leaving only the crocodile-lineage (Crocodylomorpha) and bird-lineage (Dinosauria) as survivors; along with the pterosaurian flying reptiles. About 50 years ago, the "locomotor superiority hypothesis" (LSH) proposed that dinosaurs ultimately dominated by the Early Jurassic Period because their locomotion was superior to other archosaurs'. This idea has been debated continuously since, with taxonomic and morphological analyses suggesting dinosaurs were "lucky" rather than surviving due to being biologically superior. However, the LSH has never been tested biomechanically. Here we present integration of experimental data from locomotion in extant archosaurs with inverse and predictive simulations of the same behaviours using musculoskeletal models, showing that we can reliably predict how extant archosaurs walk, run and jump. These simulations have been guiding predictive simulations of extinct archosaurs to estimate how they moved, and we show our progress in that endeavour. The musculoskeletal models used in these simulations can also be used for simpler analyses of form and function such as muscle moment arms, which inform us about more basic biomechanical similarities and differences between archosaurs. Placing all these data into an evolutionary and biomechanical context, we take a fresh look at the LSH as part of a critical review of competing hypotheses for why dinosaurs (and a few other archosaur clades) survived the Late Triassic extinctions. Early dinosaurs had some quantifiable differences in locomotor function and performance vs. some other archosaurs, but other derived dinosaurian features (e.g., metabolic or growth rates, ventilatory abilities) are not necessarily mutually exclusive from the LSH; or maybe even an opportunistic replacement hypothesis; in explaining dinosaurs' success.

Architecture of the bronchial tree in Cuvier's dwarf caiman (Paleosuchus palpebrosus).

We imaged the lungs of five Cuvier's dwarf caiman (Paleosuchus palpebrosus) via computed tomography (CT) and micro-computed tomography (µCT) and compared these data to the lungs of the American alligator (Alligator mississippiensis). These data demonstrate anatomical commonalities between the lungs of P. palpebrosus and A. mississippiensis, and a few notable differences. The structural similarities are (a) a proximally narrow, distally widened, hook-shaped primary bronchus; (b) a cervical ventral bronchus that branches of the primary bronchus and immediately makes a hairpin turn toward the apex of the lung; (c) a sequential series of dorsobronchi arising from the primary bronchus caudal to the cervical ventral bronchus; (d) intraspecifically highly variable medial sequence of secondary airways; (e) sac-like laterobronchi; and (f) grossly dead-ended caudal group bronchi in the caudal and ventral aspects of the lung. The primary differences between the two taxa are in the overall number of large bronchi (fewer in P. palpebrosus), and the number of branches that contribute to the cardiac regions. Imaging data of both a live and deceased specimen under varying states (postprandial, fasting, total lung capacity, open to atmosphere) indicate that the caudal margin and position of the lungs shift craniocaudally relative to the vertebral column. These imaging data suggest that the smooth thoracic ceiling may be correlated to visceral movement during ventilation, but this hypothesis warrants validation. These results provide the scaffolding for future comparisons between crocodilians, for generating preliminary reconstructions of the ancestral crocodilian bronchial tree, and establishing new hypotheses of bronchial homology across Archosauria.

2D and 3D visualizations of archosaur jaw muscle mechanics, ontogeny and phylogeny using ternary diagrams and 3D modeling.

Comparing patterns of performance and kinematics across behavior, development and phylogeny is crucial to understand the evolution of complex musculoskeletal systems such as the feeding apparatus. However, conveying 3D spatial data of muscle orientation throughout a feeding cycle, ontogenetic pathway or phylogenetic lineage is essential to understanding the function and evolution of the skull in vertebrates. Here, we detail the use of ternary plots for displaying and comparing the 3D orientation of muscle data. First, we illustrate changes in 3D jaw muscle resultants during jaw closing taxa the American alligator (Alligator mississippiensis). Second, we show changes in 3D muscle resultants of jaw muscles across an ontogenetic series of alligators. Third, we compare 3D resultants of jaw muscles of avian-line dinosaurs, including extant (Struthio camelus, Gallus gallus, Psittacus erithacus) and extinct (Tyrannosaurus rex) species to outline the reorganization of jaw muscles that occurred along the line to modern birds. Finally, we compare 3D resultants of jaw muscles of the hard-biting species in our sample (A. mississippiensis, T. rex, P. erithacus) to illustrate how disparate jaw muscle resultants are employed in convergent behaviors in archosaurs. Our findings show that these visualizations of 3D components of jaw muscles are immensely helpful towards identifying patterns of cranial performance, growth and diversity. These tools will prove useful for testing other hypotheses in functional morphology, comparative biomechanics, ecomorphology and organismal evolution.

Ontogenetic changes in limb posture, kinematics, forces and joint moments in American alligators (Alligator mississippiensis).

As animals increase in size, common patterns of morphological and physiological scaling may require them to perform behaviors such as locomotion while experiencing a reduced capacity to generate muscle force and an increased risk of tissue failure. Large mammals are known to manage increased mechanical demands by using more upright limb posture. However, the presence of such size-dependent changes in limb posture has rarely been tested in animals that use non-parasagittal limb kinematics. Here, we used juvenile to subadult American alligators (total length 0.46-1.27 m, body mass 0.3-5.6 kg) and examined their limb kinematics, forces, joint moments and center of mass (CoM) to test for ontogenetic shifts in posture and limb mechanics. Larger alligators typically walked with a more adducted humerus and femur and a more extended knee. Normalized peak joint moments reflected these postural patterns, with shoulder and hip moments imposed by the ground reaction force showing relatively greater magnitudes in the smallest individuals. Thus, as larger alligators use more upright posture, they incur relatively smaller joint moments than smaller alligators, which could reduce the forces that the shoulder and hip adductors of larger alligators must generate. The CoM shifted nonlinearly from juveniles through subadults. The more anteriorly positioned CoM in small alligators, together with their compliant hindlimbs, contributes to their higher forelimb and lower hindlimb normalized peak vertical forces in comparison to larger alligators. Future studies of alligators that approach maximal adult sizes could give further insight into how animals with non-parasagittal limb posture modulate locomotor patterns as they increase in mass and experience changes in the CoM.

Scaling of axial muscle architecture in juvenile Alligator mississippiensis reveals an enhanced performance capacity of accessory breathing mechanisms.

Quantitative functional anatomy of amniote thoracic and abdominal regions is crucial to understanding constraints on and adaptations for facilitating simultaneous breathing and locomotion. Crocodilians have diverse locomotor modes and variable breathing mechanics facilitated by basal and derived (accessory) muscles. However, the inherent flexibility of these systems is not well studied, and the functional specialisation of the crocodilian trunk is yet to be investigated. Increases in body size and trunk stiffness would be expected to cause a disproportionate increase in muscle force demands and therefore constrain the basal costal aspiration mechanism, necessitating changes in respiratory mechanics. Here, we describe the anatomy of the trunk muscles, their properties that determine muscle performance (mass, length and physiological cross-sectional area [PCSA]) and investigate their scaling in juvenile Alligator mississippiensis spanning an order of magnitude in body mass (359 g-5.5 kg). Comparatively, the expiratory muscles (transversus abdominis, rectus abdominis, iliocostalis), which compress the trunk, have greater relative PCSA being specialised for greater force-generating capacity, while the inspiratory muscles (diaphragmaticus, truncocaudalis ischiotruncus, ischiopubis), which create negative internal pressure, have greater relative fascicle lengths, being adapted for greater working range and contraction velocity. Fascicle lengths of the accessory diaphragmaticus scaled with positive allometry in the alligators examined, enhancing contractile capacity, in line with this muscle's ability to modulate both tidal volume and breathing frequency in response to energetic demand during terrestrial locomotion. The iliocostalis, an accessory expiratory muscle, also demonstrated positive allometry in fascicle lengths and mass. All accessory muscles of the infrapubic abdominal wall demonstrated positive allometry in PCSA, which would enhance their force-generating capacity. Conversely, the basal tetrapod expiratory pump (transversus abdominis) scaled isometrically, which may indicate a decreased reliance on this muscle with ontogeny. Collectively, these findings would support existing anecdotal evidence that crocodilians shift their breathing mechanics as they increase in size. Furthermore, the functional specialisation of the diaphragmaticus and compliance of the body wall in the lumbar region against which it works may contribute to low-cost breathing in crocodilians.

Alligators employ intermetatarsal reconfiguration to modulate plantigrade ground contact.

Feet must mediate substrate interactions across an animal's entire range of limb poses used in life. Metatarsals, the 'bones of the sole', are the dominant pedal skeletal elements for most tetrapods. In plantigrade species that walk on the entirety of their sole, such as living crocodylians, intermetatarsal mobility offers the potential for a continuum of reconfiguration within the foot itself. Alligator hindlimbs are capable of postural extremes from a belly sprawl to a high walk to sharp turns - how does the foot morphology dynamically accommodate these diverse demands? We implemented a hybrid combination of marker-based and markerless X-ray reconstruction of moving morphology (XROMM) to measure 3D metatarsal kinematics in three juvenile American alligators (Alligator mississippiensis) across their locomotor and maneuvering repertoire on a motorized treadmill and flat-surfaced arena. We found that alligators adaptively conformed their metatarsals to the ground, maintaining plantigrade contact throughout a spectrum of limb placements with non-planar feet. Deformation of the metatarsus as a whole occurred through variable abduction (twofold range of spread) and differential metatarsal pitching (45 deg arc of skew). Internally, metatarsals also underwent up to 65 deg of long-axis rotation. Such reorientation, which correlated with skew, was constrained by the overlapping arrangement of the obliquely expanded metatarsal bases. Such a proximally overlapping metatarsal morphology is shared by fossil archosaurs and archosaur relatives. In these extinct taxa, we suggest that intermetatarsal mobility likely played a significant role in maintaining ground contact across plantigrade postural extremes.

Evidence of integumentary scale diversity in the late Jurassic Sauropod Diplodocus sp. from the Mother's Day Quarry, Montana.

The life appearance of dinosaurs is a hotly debated topic in the world of paleontology, especially when it comes to dinosaur integument. In the case of sauropods, however, the topic is harder to properly discuss due to the limited amount of fossilized skin impressions that have been discovered. Thus far, the fossil record of sauropod integument fossils include titanosaur embryos from Patagonia, possible keratinous diplodocid dorsal spines, track ways with foot impressions, and other isolated skin impressions found in association with sauropod body fossils. Several prominent integument fossils have been found at the Mother's Day Quarry, located in the Bighorn Basin, Montana. These discoveries may bring new important information about diplodocids, specifically Diplodocus sp. Here we describe newly uncovered fossilized skin that gives evidence of scale diversity in the genus Diplodocus. The scales themselves represent tubercles, and exhibit various shapes including rectangular, ovoid, polygonal, and globular scales. The tubercles are small in size, the biggest of which only reach about 10mm in length. Considering how diverse the scale shapes are in such a small area of skin, it is possible that these distinct scale shapes may represent a transition on the body from one region to another: perhaps from the abdomen to dorsal side, or abdomen to shoulder. Based on analysis of extant integument and scale orientation of crocodilians, it is possible to hypothesize on the location of the integument relative to the body as well as the size and relative maturational status of the individual.

Musculoskeletal modelling of the Nile crocodile (Crocodylus niloticus) hindlimb: Effects of limb posture on leverage during terrestrial locomotion.

We developed a three-dimensional, computational biomechanical model of a juvenile Nile crocodile (Crocodylus niloticus) pelvis and hindlimb, composed of 47 pelvic limb muscles, to investigate muscle function. We tested whether crocodiles, which are known to use a variety of limb postures during movement, use limb orientations (joint angles) that optimise the moment arms (leverages) or moment-generating capacities of their muscles during different limb postures ranging from a high walk to a sprawling motion. We also describe the three-dimensional (3D) kinematics of the crocodylian hindlimb during terrestrial locomotion across an instrumented walkway and a treadmill captured via X-ray Reconstruction of Moving Morphology (biplanar fluoroscopy; 'XROMM'). We reconstructed the 3D positions and orientations of each of the hindlimb bones and used dissection data for muscle lines of action to reconstruct a focal, subject-specific 3D musculoskeletal model. Motion data for different styles of walking (a high, crouched, bended and two types of sprawling motion) were fed into the 3D model to identify whether any joints adopted near-optimal poses for leverage across each of the behaviours. We found that (1) the hip adductors and knee extensors had their largest leverages during sprawling postures and (2) more erect postures typically involved greater peak moment arms about the hip (flexion-extension), knee (flexion) and metatarsophalangeal (flexion) joints. The results did not fully support the hypothesis that optimal poses are present during different locomotory behaviours because the peak capacities were not always reached around mid-stance phase. Furthermore, we obtained few clear trends for isometric moment-generating capacities. Therefore, perhaps peak muscular leverage in Nile crocodiles is instead reached either in early/late stance or possibly during swing phase or other locomotory behaviours that were not studied here, such as non-terrestrial movement. Alternatively, our findings could reflect a trade-off between having to execute different postures, meaning that hindlimb muscle leverage is not optimised for any singular posture or behaviour. Our model, however, provides a comprehensive set of 3D estimates of muscle actions in extant crocodiles which can form a basis for investigating muscle function in extinct archosaurs.

Osteological description of casque ontogeny in the southern cassowary (Casuarius casuarius) using micro-CT imaging.

Extant cassowaries (Casuarius) are unique flightless birds found in the tropics of Indo-Australia. They have garnered substantial attention from anatomists with focus centered on the bony makeup and function of their conspicuous cranial casques, located dorsally above the orbits and neurocranium. The osteological patterning of the casque has been formally described previously; however, there are differing interpretations between authors. These variable descriptions suggest that an anatomical understanding of casque anatomy and its constituent elements may be enhanced by developmental studies aimed at further elucidating this bizarre structure. In the present study, we clarify casque osteology of the southern cassowary (C. casuarius) by detailing casque anatomy across an extensive growth series for the first time. We used micro-computed tomography (µCT) imaging to visualize embryonic development and post-hatching ontogeny through adulthood. We also sampled closely related emus (Dromaius novaehollandiae) and ostriches (Struthio camelus) to provide valuable comparative context. We found that southern cassowary casques are comprised of three paired (i.e., nasals, lacrimals, frontals) and two unpaired elements (i.e., mesethmoid, median casque element). Although lacrimals have rarely been considered as casque elements, the contribution to the casque structure was evident in µCT images. The median casque element has often been cited as a portion of the mesethmoid. However, through comparisons between immature C. casuarius and D. novaehollandiae, we document the median casque element as a distinct unit from the mesethmoid.

Ontogeny of a sexually selected structure in an extant archosaur Gavialis gangeticus (Pseudosuchia: Crocodylia) with implications for sexual dimorphism in dinosaurs.

Despite strong evidence for sexual selection in various display traits and other exaggerated structures in large extinct reptiles, such as dinosaurs, detecting sexual dimorphism in them remains difficult. Their relatively small sample sizes, long growth periods, and difficulties distinguishing the sexes of fossil specimens mean that there are little compelling data on dimorphism in these animals. The extant gharial (Gavialis gangeticus) is a large and endangered crocodylian that is sexually dimorphic in size, but males also possesses a sexually selected structure, the ghara, which has an osteological correlate in the presence of a fossa associated with the nares. This makes the species a unique model for potentially assessing dimorphism in fossil lineages, such as dinosaurs and pterosaurs, because it is a large, slow-growing, egg-laying archosaur. Here we assess the dimorphism of G. gangeticus across 106 specimens and show that the presence of a narial fossa diagnoses adult male gharials. Males are larger than females, but the level of size dimorphism, and that of other cranial features, is low and difficult to detect without a priori knowledge of the sexes, even with this large dataset. By extension, dimorphism in extinct reptiles is very difficult to detect in the absence of sex specific characters, such as the narial fossa.

Molecular anatomy of the alligator dorsal telencephalon.

The evolutionary relationships of the mammalian neocortex and avian dorsal telencephalon (DT) nuclei have been debated for more than a century. Despite their central importance to this debate, nonavian reptiles remain underexplored with modern molecular techniques. Reptile studies harbor great potential for understanding the changes in DT organization that occurred in the early evolution of amniotes. They may also help clarify the specializations in the avian DT, which comprises a massive, cell-dense dorsal ventricular ridge (DVR) and a nuclear dorsal-most structure, the Wulst. Crocodilians are phylogenetically and anatomically attractive for DT comparative studies: they are the closest living relatives of birds and have a strikingly bird-like DVR, but they also possess a highly differentiated reptile cerebral cortex. We studied the DT of the American alligator, Alligator mississippiensis, at late embryonic stages with a panel of molecular marker genes. Gene expression and cytoarchitectonic analyses identified clear homologs of all major avian DVR subdivisions including a mesopallium, an extensive nidopallium with primary sensory input territories, and an arcopallium. The alligator medial cortex is divided into three components that resemble the mammalian dentate gyrus, CA fields, and subiculum in gene expression and topography. The alligator dorsal cortex contains putative homologs of neocortical input, output, and intratelencephalic projection neurons and, most notably, these are organized into sublayers similar to mammalian neocortical layers. Our findings on the molecular anatomy of the crocodilian DT are summarized in an atlas of the alligator telencephalon.

Neocortical Association Cell Types in the Forebrain of Birds and Alligators.

The avian dorsal telencephalon has two vast territories, the nidopallium and the mesopallium, both of which have been shown to contribute substantially to higher cognitive functions. From their connections, these territories have been proposed as equivalent to mammalian neocortical layers 2 and 3, various neocortical association areas, or the amygdala, but whether these are analogies or homologies by descent is unknown. We investigated the molecular profiles of the mesopallium and the nidopallium with RNA-seq. Gene expression experiments established that the mesopallium, but not the nidopallium, shares a transcription factor network with the intratelencephalic class of neocortical neurons, which are found in neocortical layers 2, 3, 5, and 6. Experiments in alligators demonstrated that these neurons are also abundant in the crocodilian cortex and form a large mesopallium-like structure in the dorsal ventricular ridge. Together with previous work, these molecular findings indicate a homology by descent for neuronal cell types of the avian dorsal telencephalon with the major excitatory cell types of mammalian neocortical circuits: the layer 4 input neurons, the deep layer output neurons, and the multi-layer intratelencephalic association neurons. These data raise the interesting possibility that avian and primate lineages evolved higher cognitive abilities independently through parallel expansions of homologous cell populations.

The evolution of the manus of early theropod dinosaurs is characterized by high inter- and intraspecific variation.

The origin of the avian hand, with its reduced and fused carpals and digits, from the five-fingered hands and complex wrists of early dinosaurs represents one of the major transformations of manus morphology among tetrapods. Much attention has been directed to the later part of this transition, from four- to three-fingered taxa. However, earlier anatomical changes may have influenced these later modifications, possibly paving the way for a later frameshift in digit identities. We investigate the five- to four-fingered transition among early dinosaurs, along with changes in carpus morphology. New three-dimensional reconstructions from computed tomography data of the manus of the Triassic and Early Jurassic theropod dinosaurs Coelophysis bauri and Megapnosaurus rhodesiensis are described and compared intra- and interspecifically. Several novel findings emerge from these reconstructions and comparisons, including the first evidence of an ossified centrale and a free intermedium in some C. bauri specimens, as well as confirmation of the presence of a vestigial fifth metacarpal in this taxon. Additionally, a specimen of C. bauri and an unnamed coelophysoid from the Upper Triassic Hayden Quarry, New Mexico, are to our knowledge the only theropods (other than alvarezsaurs and birds) in which all of the distal carpals are completely fused together into a single unit. Several differences between the manus of C. bauri and M. rhodesiensis are also identified. We review the evolution of the archosauromorph manus more broadly in light of these new data, and caution against incorporating carpal characters in phylogenetic analyses of fine-scale relationships of Archosauromorpha, in light of the high degree of observed polymorphism in taxa for which large sample sizes are available, such as the theropod Coelophysis and the sauropodomorph Plateosaurus. We also find that the reduction of the carpus and ultimate loss of the fourth and fifth digits among early dinosaurs did not proceed in a neat, stepwise fashion, but was characterized by multiple losses and possible gains of carpals, metacarpals and phalanges. Taken together, the high degree of intra- and interspecific variability in the number and identities of carpals, and the state of reduction of the fourth and fifth digits suggest the presence of a 'zone of developmental variability' in early dinosaur manus evolution, from which novel avian-like morphologies eventually emerged and became channelized among later theropod clades.