Musculoskeletal modeling of sprawling and parasagittal forelimbs provides insight into synapsid postural transition.

The sprawling-parasagittal postural shift was a major transition during synapsid evolution, underpinned by reorganization of the forelimb, and considered key to mammalian ecological diversity. Determining when and how this transition occurred in the fossil record is challenging owing to limited comparative data on extant species. Here, we built forelimb musculoskeletal models of three extant taxa that bracket sprawling-parasagittal postures-tegu lizard, echidna, and opossum-and tested the relationship between three-dimensional joint mobility, muscle action, and posture. Results demonstrate clear functional variation between postural grades, with the parasagittal opossum occupying a distinct region of pose space characterized by a highly retracted and depressed shoulder joint that emphasizes versatility and humeral elevation. Applying our data to the fossil record support trends of an increasingly retracted humerus and greater elevation muscle moment arms indicative of more parasagittal postures throughout synapsid evolution.

Validation of an Echidna Forelimb Musculoskeletal Model Using XROMM and diceCT.

In evolutionary biomechanics, musculoskeletal computer models of extant and extinct taxa are often used to estimate joint range of motion (ROM) and muscle moment arms (MMAs), two parameters which form the basis of functional inferences. However, relatively few experimental studies have been performed to validate model outputs. Previously, we built a model of the short-beaked echidna (Tachyglossus aculeatus) forelimb using a traditional modelling workflow, and in this study we evaluate its behaviour and outputs using experimental data. The echidna is an unusual animal representing an edge-case for model validation: it uses a unique form of sprawling locomotion, and possesses a suite of derived anatomical features, in addition to other features reminiscent of extinct early relatives of mammals. Here we use diffusible iodine-based contrast-enhanced computed tomography (diceCT) alongside digital and traditional dissection to evaluate muscle attachments, modelled muscle paths, and the effects of model alterations on the MMA outputs. We use X-ray Reconstruction of Moving Morphology (XROMM) to compare ex vivo joint ROM to model estimates based on osteological limits predicted via single-axis rotation, and to calculate experimental MMAs from implanted muscles using a novel geometric method. We also add additional levels of model detail, in the form of muscle architecture, to evaluate how muscle torque might alter the inferences made from MMAs alone, as is typical in evolutionary studies. Our study identifies several key findings that can be applied to future models. 1) A light-touch approach to model building can generate reasonably accurate muscle paths, and small alterations in attachment site seem to have minimal effects on model output. 2) Simultaneous movement through multiple degrees of freedom, including rotations and translation at joints, are necessary to ensure full joint ROM is captured; however, single-axis ROM can provide a reasonable approximation of mobility depending on the modelling objectives. 3) Our geometric method of calculating MMAs is consistent with model-predicted MMAs calculated via partial velocity, and is a potentially useful tool for others to create and validate musculoskeletal models. 4) Inclusion of muscle architecture data can change some functional inferences, but in many cases reinforced conclusions based on MMA alone.

Evolution of the patella and patelloid in marsupial mammals.

The musculoskeletal system of marsupial mammals has numerous unusual features beyond the pouch and epipubic bones. One example is the widespread absence or reduction (to a fibrous "patelloid") of the patella ("kneecap") sesamoid bone, but prior studies with coarse sampling indicated complex patterns of evolution of this absence or reduction. Here, we conducted an in-depth investigation into the form of the patella of extant marsupial species and used the assembled dataset to reconstruct the likely pattern of evolution of the marsupial patella. Critical assessment of the available literature was followed by examination and imaging of museum specimens, as well as CT scanning and histological examination of dissected wet specimens. Our results, from sampling about 19% of extant marsupial species-level diversity, include new images and descriptions of the fibrocartilaginous patelloid in Thylacinus cynocephalus (the thylacine or "marsupial wolf") and other marsupials as well as the ossified patella in Notoryctes 'marsupial moles', Caenolestes shrew opossums, bandicoots and bilbies. We found novel evidence of an ossified patella in one specimen of Macropus rufogriseus (Bennett's wallaby), with hints of similar variation in other species. It remains uncertain whether such ossifications are ontogenetic variation, unusual individual variation, pathological or otherwise, but future studies must continue to be conscious of variation in metatherian patellar sesamoid morphology. Our evolutionary reconstructions using our assembled data vary, too, depending on the reconstruction algorithm used. A maximum likelihood algorithm favours ancestral fibrocartilaginous "patelloid" for crown clade Marsupialia and independent origins of ossified patellae in extinct sparassodonts, peramelids, notoryctids and caenolestids. A maximum parsimony algorithm favours ancestral ossified patella for the clade [Marsupialia + sparassodonts] and subsequent reductions into fibrocartilage in didelphids, dasyuromorphs and diprotodonts; but this result changed to agree more with the maximum likelihood results if the character state reconstructions were ordered. Thus, there is substantial homoplasy in marsupial patellae regardless of the evolutionary algorithm adopted. We contend that the most plausible inference, however, is that metatherians independently ossified their patellae at least three times in their evolution. Furthermore, the variability of the patellar state we observed, even within single species (e.g. M. rufogriseus), is fascinating and warrants further investigation, especially as it hints at developmental plasticity that might have been harnessed in marsupial evolution to drive the complex patterns inferred here.

Pectoral girdle and forelimb musculoskeletal function in the echidna (Tachyglossus aculeatus): insights into mammalian locomotor evolution.

Although evolutionary transformation of the pectoral girdle and forelimb appears to have had a profound impact on mammalian locomotor and ecological diversity, both the sequence of anatomical changes and the functional implications remain unclear. Monotremes can provide insight into an important stage of this evolutionary transformation, due to their phylogenetic position as the sister-group to therian mammals and their mosaic of plesiomorphic and derived features. Here we build a musculoskeletal computer model of the echidna pectoral girdle and forelimb to estimate joint ranges of motion (ROM) and muscle moment arms (MMA)-two fundamental descriptors of biomechanical function. We find that the echidna's skeletal morphology restricts scapulocoracoid mobility and glenohumeral flexion-extension compared with therians. Estimated shoulder ROMs and MMAs for muscles crossing the shoulder indicate that morphology of the echidna pectoral girdle and forelimb is optimized for humeral adduction and internal rotation, consistent with limited in vivo data. Further, more muscles act to produce humeral long-axis rotation in the echidna compared to therians, as a consequence of differences in muscle geometry. Our musculoskeletal model allows correlation of anatomy and function, and can guide hypotheses regarding function in extinct taxa and the morphological and locomotor transformation leading to therian mammals.

Evolution of the patellar sesamoid bone in mammals.

The patella is a sesamoid bone located in the major extensor tendon of the knee joint, in the hindlimb of many tetrapods. Although numerous aspects of knee morphology are ancient and conserved among most tetrapods, the evolutionary occurrence of an ossified patella is highly variable. Among extant (crown clade) groups it is found in most birds, most lizards, the monotreme mammals and almost all placental mammals, but it is absent in most marsupial mammals as well as many reptiles. Here, we integrate data from the literature and first-hand studies of fossil and recent skeletal remains to reconstruct the evolution of the mammalian patella. We infer that bony patellae most likely evolved between four and six times in crown group Mammalia: in monotremes, in the extinct multituberculates, in one or more stem-mammal genera outside of therian or eutherian mammals and up to three times in therian mammals. Furthermore, an ossified patella was lost several times in mammals, not including those with absent hindlimbs: once or more in marsupials (with some re-acquisition) and at least once in bats. Our inferences about patellar evolution in mammals are reciprocally informed by the existence of several human genetic conditions in which the patella is either absent or severely reduced. Clearly, development of the patella is under close genomic control, although its responsiveness to its mechanical environment is also important (and perhaps variable among taxa). Where a bony patella is present it plays an important role in hindlimb function, especially in resisting gravity by providing an enhanced lever system for the knee joint. Yet the evolutionary origins, persistence and modifications of a patella in diverse groups with widely varying habits and habitats-from digging to running to aquatic, small or large body sizes, bipeds or quadrupeds-remain complex and perplexing, impeding a conclusive synthesis of form, function, development and genetics across mammalian evolution. This meta-analysis takes an initial step toward such a synthesis by collating available data and elucidating areas of promising future inquiry.

Cellular preservation of musculoskeletal specializations in the Cretaceous bird Confuciusornis.

The hindlimb of theropod dinosaurs changed appreciably in the lineage leading to extant birds, becoming more 'crouched' in association with changes to body shape and gait dynamics. This postural evolution included anatomical changes of the foot and ankle, altering the moment arms and control of the muscles that manipulated the tarsometatarsus and digits, but the timing of these changes is unknown. Here, we report cellular-level preservation of tendon- and cartilage-like tissues from the lower hindlimb of Early Cretaceous Confuciusornis. The digital flexor tendons passed through cartilages, cartilaginous cristae and ridges on the plantar side of the distal tibiotarsus and proximal tarsometatarsus, as in extant birds. In particular, fibrocartilaginous and cartilaginous structures on the plantar surface of the ankle joint of Confuciusornis may indicate a more crouched hindlimb posture. Recognition of these specialized soft tissues in Confuciusornis is enabled by our combination of imaging and chemical analyses applied to an exceptionally preserved fossil.

Skeletal pathology and variable anatomy in elephant feet assessed using computed tomography.

Foot problems are a major cause of morbidity and mortality in elephants, but are underreported due to difficulties in diagnosis, particularly of conditions affecting the bones and internal structures. Here we evaluate post-mortem computer tomographic (CT) scans of 52 feet from 21 elephants (seven African Loxodonta africana and 14 Asian Elephas maximus), describing both pathology and variant anatomy (including the appearance of phalangeal and sesamoid bones) that could be mistaken for disease. We found all the elephants in our study to have pathology of some type in at least one foot. The most common pathological changes observed were bone remodelling, enthesopathy, osseous cyst-like lesions, and osteoarthritis, with soft tissue mineralisation, osteitis, infectious osteoarthriti, subluxation, fracture and enostoses observed less frequently. Most feet had multiple categories of pathological change (81% with two or more diagnoses, versus 10% with a single diagnosis, and 9% without significant pathology). Much of the pathological change was focused over the middle/lateral digits, which bear most weight and experience high peak pressures during walking. We found remodelling and osteoarthritis to be correlated with increasing age, more enthesopathy in Asian elephants, and more cyst-like lesions in females. We also observed multipartite, missing and misshapen phalanges as common and apparently incidental findings. The proximal (paired) sesamoids can appear fused or absent, and the predigits (radial/tibial sesamoids) can be variably ossified, though are significantly more ossified in Asian elephants. Our study reinforces the need for regular examination and radiography of elephant feet to monitor for pathology and as a tool for improving welfare.

Analysis of the moment arms and kinematics of ostrich (Struthio camelus) double patellar sesamoids.

The patella ("kneecap") is a biomechanically important feature of the tendinous insertion of the knee extensor muscles, able to alter the moment arm lengths between its input and output tendons, and so modify the mechanical advantage of the knee extensor muscle. However, patellar gearing function is little-explored outside of humans, and the patella is often simplified or ignored in biomechanical models. Here, we investigate patellar gearing and kinematics in the ostrich-frequently used as an animal analogue to human bipedal locomotion and unusual in its possession of two patellae at the knee joint. We use x-ray reconstruction of moving morphology (XROMM) techniques to capture the kinematics of the patellae in an adult ostrich cadaver, passively manipulated in flexion-extension. Moment arm ratios between the input and output tendons of each patella are calculated from kinematically determined centers of patellofemoral joint rotation. Both patellae are found to decrease the mechanical advantage of the extensor muscle-tendon complex, decreasing the tendon output force for a given muscle input force, but potentially increasing the relative speed of knee extension. Mechanically and kinematically, the proximal patella behaves similarly to the single patella of most other species, whereas the distal patella has properties of both a fixed retroarticular process and a moving sesamoid. It is still not clear why ostriches possess two patellae, but we suggest that the configuration in ostriches benefits their rapid locomotion and provides tendon protection.

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.

Three-dimensional anatomy of the ostrich (Struthio camelus) knee joint.

The three-dimensional anatomy of the ostrich (Struthio camelus) knee (femorotibial, femorofibular, and femoropatellar) joint has scarcely been studied, and could elucidate certain mechanobiological properties of sesamoid bones. The adult ostrich is unique in that it has double patellae, while another similar ratite bird, the emu, has none. Understanding why these patellae form and what purpose they may serve is dually important for future studies on ratites as well as for understanding the mechanobiological characteristics of sesamoid bone development. For this purpose, we present a three-dimensional anatomical study of the ostrich knee joint, detailing osteology, ligaments and menisci, and myology. We have identified seven muscles which connect to the two patellae and compare our findings to past descriptions. These descriptions can be used to further study the biomechanical loading and implications of the double patella in the ostrich.

Structure, ontogeny and evolution of the patellar tendon in emus (Dromaius novaehollandiae) and other palaeognath birds.

The patella (kneecap) exhibits multiple evolutionary origins in birds, mammals, and lizards, and is thought to increase the mechanical advantage of the knee extensor muscles. Despite appreciable interest in the specialized anatomy and locomotion of palaeognathous birds (ratites and relatives), the structure, ontogeny and evolution of the patella in these species remains poorly characterized. Within Palaeognathae, the patella has been reported to be either present, absent, or fused with other bones, but it is unclear how much of this variation is real, erroneous or ontogenetic. Clarification of the patella's form in palaeognaths would provide insight into the early evolution of the patella in birds, in addition to the specialized locomotion of these species. Findings would also provide new character data of use in resolving the controversial evolutionary relationships of palaeognaths. In this study, we examined the gross and histological anatomy of the emu patellar tendon across several age groups from five weeks to 18 months. We combined these results with our observations and those of others regarding the patella in palaeognaths and their outgroups (both extant and extinct), to reconstruct the evolution of the patella in birds. We found no evidence of an ossified patella in emus, but noted its tendon to have a highly unusual morphology comprising large volumes of adipose tissue contained within a collagenous meshwork. The emu patellar tendon also included increasing amounts of a cartilage-like tissue throughout ontogeny. We speculate that the unusual morphology of the patellar tendon in emus results from assimilation of a peri-articular fat pad, and metaplastic formation of cartilage, both potentially as adaptations to increasing tendon load. We corroborate previous observations of a 'double patella' in ostriches, but in contrast to some assertions, we find independent (i.e., unfused) ossified patellae in kiwis and tinamous. Our reconstructions suggest a single evolutionary origin of the patella in birds and that the ancestral patella is likely to have been a composite structure comprising a small ossified portion, lost by some species (e.g., emus, moa) but expanded in others (e.g., ostriches).

Osteopathology in the feet of rhinoceroses: lesion type and distribution.

An estimated 1,170 white (Ceratotherium simum), black (Diceros bicornis), greater one-horned (Rhinoceros unicornis), and Sumatran (Dicerorhinus sumatrensis) rhinoceroses are kept in captivity worldwide, where they are a popular public attraction and serve important roles in education and conservation. Rhinoceroses in captivity are reportedly affected by a variety of foot conditions, including abscesses, nail cracking, and pododermatitis, but there are few studies reporting associated bony pathology in these species. This study aimed to describe osteopathology in rhinoceros feet and identify normal and abnormal osteologic features of rhinoceros feet. The metacarpal-tarsal and phalangeal bones from 81 feet (67 skeletal specimens and 14 cadaveric feet), derived from 27 rhinoceroses of various species, were evaluated in the study (1 black, 11 white, 2 greater one-horned, 3 Javan, 9 Sumatran, and 1 unknown). Bones were examined visually (skeletal specimens) or by computed tomography (cadaver specimens) for evidence of bony lesions. Of the 27 rhinoceroses examined, 22 showed some degree of bone pathology in at least one limb. Six broad categories of pathologic change were identified, with numbers in parentheses representing numbers of rhinoceroses with lesions in at least one limb/number of rhinoceroses examined: enthesopathy (20/27), osteoarthritis (15/27), pathologic bone remodeling (12/27), osteitis-osteomyelitis (3/27), fracture (3/8), and subluxation (3/8). The frequency of pathologic changes in fore- and hind limbs was not significantly different. Most (91%) enthesopathies were observed on the proximal phalanges of the digits, and osteoarthritis was most common in the distal interphalangeal joints of the medial and lateral digits (32 and 26%, respectively). In addition to the pathology described, all examined rhinoceroses also had multiple small surface lucencies in the distal limb bones as an apparently normal anatomic feature. This study is an important first step in identifying both normal and pathologic features of rhinoceros feet and hopefully will thereby contribute to the improved knowledge and care of these species.