Bone metabolism and evolutionary origin of osteocytes: Novel application of FIB-SEM tomography.

Lacunae and canaliculi spaces of osteocytes are remarkably well preserved in fossilized bone and serve as an established proxy for bone cells. The earliest bone in the fossil record is acellular (anosteocytic), followed by cellular (osteocytic) bone in the jawless relatives of jawed vertebrates, the osteostracans, about 400 million years ago. Virtually nothing is known about the physiological pressures that would have initially favored osteocytic over anosteocytic bone. We apply focused ion beam-scanning electron microscopy tomography combined with machine learning for cell detection and segmentation to image fossil cell spaces. Novel three-dimensional high-resolution images reveal areas of low density around osteocyte lacunae and their canaliculi in osteostracan bone. This provides evidence for demineralization that would have occurred in vivo as part of osteocytic osteolysis, a mechanism of mineral homeostasis, supporting the hypothesis that a physiological demand for phosphorus was the principal driver in the initial evolution of osteocytic bone.

Rarity of congenital malformation and deformity in the fossil record of vertebrates - A non-human perspective.

OBJECTIVE: A malformed pectoral joint of the middle Devonian antiarch fish Asterolepis ornata is described, and a survey of congenital malformations in the fossil record is provided. MATERIALS: The specimen of A. ornata (MB.f.73) from Ehrman in Latvia, stored at the Museum für Naturkunde Berlin, Germany. METHODS: A. ornata was macroscopically and radiologically investigated, and the overview on congenital malformation was based on an extensive literature survey. RESULTS: In the deformed joint of A. ornata, the articular surfaces and muscle attachment sites are greatly reduced, indicating restricted mobility. Congenital malformations can be found since the middle Silurian and affect all groups of vertebrates, but they are rare. Teeth and the vertebral column are the most commonly affected anatomical regions, and the mechanisms causing these malformations probably remained the same through geological time. CONCLUSIONS: Micro-CT of the deformed joint shows no disturbance of the normal trabecular pattern and no evidence of trauma or disease, suggesting a congenital hypoplasia, although an acquired deformity cannot be ruled out completely. SIGNIFICANCE: Congenital malformations, even those that are rare, were part of the common history of vertebrates for more than 400 million years. LIMITATIONS: Epidemiologic measures like incidence and prevalence usually cannot be applied to define rare diseases in the fossil record. SUGGESTIONS FOR FURTHER RESEARCH: A broadly based analysis of species of fossil vertebrates with numerus recovered specimens (e.g. many bony fishes, amphibians, certain dinosaurs) might statistically affirm the occurrence of malformations and possible correlations with the paleoenvironment.

Do Meristic Characters Used in Phylogenetic Analysis Evolve in an Ordered Manner?

The use of ordered characters in phylogenetic analysis has been inconsistent throughout the history of phylogenetic inference. It has become more widespread in recent years, and some have advocated that all characters representing continuous or meristic traits should be ordered as a matter of course. Here, using the example of dental evolution, we examine two factors that may impact on whether meristic characters actually evolve in an ordered manner: the regulatory hierarchy governing the development of teeth that allows large sections of the entire tooth row to be suppressed in a single transition and regionalization of the tooth row where different modules have a degree of independence in their evolution. These are studied using both empirical and simulated data. Models of evolution of such characters are examined over molecular phylogenies to see if ordered or unordered models fit best. Simulations of tooth-row evolution are designed to incorporate changes in region size and multiple levels of developmental control to suppress individual regions or the entire row. The empirical analyses show that in a clade with largely homodont dentition the characters evolve in an ordered manner, but if dentition is heterodont with distinct regionalization their evolution better fits an unordered model. In the simulations, even if teeth are added and removed from the tooth row in an ordered manner, dividing the row into independently evolving modules can lead to characters covering multiple modules better fitting an unordered model of evolution. Adding the ability to suppress regions or the entire tooth row has a variable effect depending on the rates of suppression relative to the rates of addition and subtraction of individual teeth. We therefore advise not following a single policy when deciding whether to order meristic traits but to base the decision on a priori knowledge of the focal clade's evolution and developmental biology. [Discrete characters; ordered characters; phylogeny; teeth.].

Histological skeletochronology indicates developmental plasticity in the early Permian stem lissamphibian Doleserpeton annectens.

Doleserpeton annectens is a small-bodied early Permian amphibamiform, a clade of temnospondyl amphibians regarded by many workers to be on the lissamphibian stem. Most studies of this taxon have focused solely on its anatomy, but further exploration of other aspects of its paleobiology, such as developmental patterns, is critical for a better understanding of the early evolutionary history of lissamphibians. Here, we present a histological analysis of growth patterns in D. annectens that utilizes 60 femora, the largest sample size for any Paleozoic tetrapod. We identified pervasive pairs of closely spaced lines of arrested growth (LAGs), a pattern that indicates a marked degree of climatic harshness and that would result in two cessations of growth within a presumed single year. We documented a wide degree of variation compared to previous temnospondyl skeletochronological studies, reflected in the poor correlation between size and inferred age, but this observation aligns closely with patterns observed in extant lissamphibians. Furthermore, sensitivity analyses conducted by subsampling our dataset at more typical sample sizes for paleontological studies produced a wide range of results. This includes biologically improbable results and exceptionally well-fit curves that demonstrate that low sample size can produce potentially misleading artifacts. We propose that the weak correlation between age and size represents developmental plasticity in D. annectens that typifies extant lissamphibians. Detection of these patterns is likely only possible with large sample sizes in extinct taxa, and low sample sizes can produce false, misleading results that warrant caution in drawing paleobiological interpretations from such samples.

Retention of fish-like odontode overgrowth in Permian tetrapod dentition supports outside-in theory of tooth origins.

Teeth are often thought of as structures that line the margins of the mouth; however, tooth-like structures called odontodes are commonly found on the dermal bones of many Palaeozoic vertebrates including early jawless fishes. 'Odontode' is a generalized term for all tooth-like dentine structures that have homologous tissues and development. This definition includes true teeth and the odontodes of early 'fishes', which have been recently examined to gain new insights into the still unresolved origin of teeth. Two leading hypotheses are frequently referenced in this debate: the 'outside-in' hypothesis, which posits that dermal odontodes evolutionarily migrate into the oral cavity, and the 'inside-out' hypothesis, which posits that teeth originated in the oropharyngeal cavity and then moved outwards into the oral cavity. Here, we show that, unlike the well-known one-to-one replacement patterns of marginal dentition, the palatal dentition of the early Permian tetrapods, including the dissorophoid amphibian Cacops and the early reptile Captorhinus, is overgrown by a new layer of bone to which the newest teeth are then attached. This same overgrowth pattern has been well documented in dermal and oral odontodes (i.e. teeth) of early fishes. We propose that this pattern represents the primitive condition for vertebrates and may even predate the origin of jaws. Therefore, this pattern crosses the fish-tetrapod transition, and the retention of this ancestral pattern in the palatal dentition of early terrestrial tetrapods provides strong support for the 'outside-in' hypothesis of tooth origins.

Permian metabolic bone disease revealed by microCT: Paget's disease-like pathology in vertebrae of an early amniote.

Bone remodeling is an essential physiological process in growth and healing. In modern systems deviations from normal bone physiology in the form of pathologies aid in the understanding of normal bone metabolism. Here we use external morphology and X-ray microtomography to diagnose and describe a metabolic bone disease in an amniote from the early Permian. The specimen consists of two fused tail vertebrae of a small varanopid from early Permian (289 million years old) cave deposits near Richards Spur, Oklahoma, USA. Inspection of the outer morphology reveals that the fusion encompasses the vertebral centra, zygopophyses and haemal arches, with the fusion zones distinctly swollen on the left side of the specimen. With visualization of its internal structure by microCT, this specimen is diagnosed as a complex metabolic bone disease. The radiological imaging suggests a pathologically high bone turnover rate, as shown by abnormal bone formation in some areas and increased bone resorption in others. This supports that the varanopid suffered from a metabolic bone disease similar to Paget's disease of bone as seen in humans today, which is linked to both genetic and viral factors. This finding extends the occurrence of Paget-like disease to the early Permian, and-provided a viral component was present-would also be by far the oldest evidence of viral infection in the fossil record.

Histological analysis of post-eruption tooth wear adaptations, and ontogenetic changes in tooth implantation in the acrodontan squamate Pogona vitticeps.

Teeth have been a focus of research in both extinct and extant taxa alike; a significant portion of dental literature is concerned with dental patterning and replacement. Most non-mammalian vertebrates continuously replace their dentition but an anomalous group of squamates has forgone this process in only having one tooth generation; these squamates all have apically implanted teeth, a condition known as acrodonty. Acrodont dentition and various characteristics attributed to it, including a lack of replacement, have often been defined ambiguously. This study explores this type of implantation through histology in the ontogeny of the acrodont agamid Pogona vitticeps. The non-replacing teeth of this squamate provides an opportunity to study wear adaptations, maintenance of occlusion in a non-mammalian system, and most importantly post-eruption changes in the tooth bone interface. In this study the post-eruption changes combined with dental wear likely gives the appearance of acrodont implantation.

The Permian reptile Opisthodontosaurus carrolli: a model for acrodont tooth replacement and dental ontogeny.

Continuous tooth replacement is common for tetrapods, but some groups of acrodont lepidosaurs have lost the ability to replace their dentition (monophyodonty). Acrodonty, where the tooth attaches to the apex of the jawbone, is an unusual form of tooth attachment that has been associated with the highly autapomorphic condition of monophyodonty. Beyond Lepidosauria, very little is known about the relationship between acrodonty and monophyodonty in other amniotes. We test for this association with a detailed study of the dentition of Opisthodontosaurus, an unusual Early Permian captorhinid eureptile with acrodont dentition. We provide clear evidence, both histological and morphological, that there were regular tooth replacement events in the lower jaw of Opisthodontosaurus, similar to its captorhinid relatives. Thus, our study of the oldest known amniote with an acrodont dentition shows that acrodonty does not inhibit tooth replacement, and that many of the characteristics assigned to lepidosaurian acrodonty are actually highly derived features of lepidosaurs that have resulted secondarily from a lack of tooth replacement. In the context of reptilian dental evolution, we propose the retention of the simple definition of acrodonty, which only pertains to the relative position of the tooth at the apex of the jaw, where the jaw possesses equal lingual and labial walls. This definition of implantation therefore focuses solely on the spatial relationship between the tooth and the jawbone, and separates this relationship from tooth development and replacement.

Histological characterization of denticulate palatal plates in an Early Permian dissorophoid.

Denticles are small, tooth-like protrusions that are commonly found on the palate of early tetrapods. Despite their widespread taxonomic occurrence and similar external morphology to marginal teeth, it has not been rigorously tested whether denticles are structurally homologous to true teeth with features such as a pulp cavity, dentine, and enamel, or if they are bony, tooth-like protrusions. Additionally, the denticles are known to occur not only on the palatal bones but also on a mosaic of small palatal plates that is thought to have covered the interpterygoid vacuities of temnospondyls through implantation in a soft tissue covering; however, these plates have never been examined beyond a simple description of their position and external morphology. Accordingly, we performed a histological analysis of these denticulate palatal plates in a dissorophoid temnospondyl in order to characterize their microanatomy and histology. The dentition on these palatal plates has been found to be homologous with true teeth on the basis of both external morphology and histological data through the identification of features such as enamel and a pulp cavity surrounded by dentine. In addition, patterns of tooth replacement and ankylosis support the hypothesis of structural homology between these tiny teeth on the palatal plates and the much larger marginal dentition. We also provide the first histological characterization of the palatal plates, including documentation of abundant Sharpey's fibres that provide a direct line of evidence to support the hypothesis of soft tissue implantation. Finally, we conducted a survey of the literature to determine the taxonomic distribution of these plates within Temnospondyli, providing a broader context for the presence of palatal plates and illustrating the importance of maintaining consistency in nomenclature.

Ontogenetic Change in the Temporal Region of the Early Permian Parareptile Delorhynchus cifellii and the Implications for Closure of the Temporal Fenestra in Amniotes.

A juvenile specimen of Delorhynchus cifellii, collected from the Early Permian fissure-fill deposits of Richards Spur, Oklahoma, permits the first detailed study of cranial ontogeny in this parareptile. The specimen, consisting of a partially articulated skull and mandible, exhibits several features that identify it as juvenile. The dermal tuberosities that ornament the dorsal side and lateral edges of the largest skull of D. cifellii specimens, are less prominent in the intermediate sized holotype, and are absent in the new specimen. This indicates that the new specimen represents an earlier ontogenetic stage than all previously described members of this species. In addition, the incomplete interdigitation of the sutures, most notably along the fronto-nasal contact, plus the proportionally larger sizes of the orbit and temporal fenestrae further support an early ontogenetic stage for this specimen. Comparisons between this juvenile and previously described specimens reveal that the size and shape of the temporal fenestra in Delorhynchus appear to vary through ontogeny, due to changes in the shape and size of the bordering cranial elements. The jugal of the juvenile specimen is tri-radiate and similar in outline with those found in other amniotes with temporal fenestrae. The available growth series of D. cifellii shows that the jugal gradually becomes a more robust, tetra-radiate element, as the proportionate size of the temporal fenestra is reduced. Ontogenetic changes of other elements that form the border of the fenestra also contribute to its reduction. This growth series provides valuable new information regarding the ontogenetic trajectory of the temporal fenestra in a Palaeozoic reptile, which may be applicable to the evolutionary event of loss of temporal fenestration in other amniotes.