Development and structural characteristics of pseudoosteodentine in the Pacific cutlassfish, Trichiurus lepturus.

Depending on the mineralization pattern of dentine, teeth can be divided into three histological types (orthodont, osteodont, and pseudoosteodont type). However, the development and structural characteristics of pseudoosteodentine has not been systematically investigated yet. Here, the teeth of Trichiurus lepturus were selected for revealing a maturation process during pseudoosteodentine formation and describing ultrastructural details of pseudoosteodentine architecture. Micro-computed tomography, scanning electron microscopy, hematoxylin-eosin, Masson staining and immunohistochemistry using a dentine sialophosphoprotein (DSPP) antibody were used to analyze the microstructure and the development of the dentine. Compared with Muraenesox cinereus orthodentine, the ultrastructure of pseudoosteodentine, dentine development, the localization and migration of odontoblasts during odontogenesis in T. lepturus were observed in detail. In pseudoosteodentine, orthodentine and osteodentine all contain similar tubule-like structures and tubule openings. Labeled by DSPP immunohistochemistry for secretory odontoblasts, the organic matrix of pseudoosteodentine forms in two stages: secreting matrix by orthodentine odontoblasts with inverted nuclear polarity and the formation of osteodentine by stellate odontoblasts throughout the dental papilla. Our findings increase the understanding of the odontogenesis and structure of pseudoosteodentine and might provide a new model for the study of biomineralization and tooth development.

Complex enameloid microstructure of †Ischyrhiza mira rostral denticles.

Serving in a foraging or self-defense capacity, pristiophorids, pristids, and the extinct sclerorhynchoids independently evolved an elongated rostrum lined with modified dermal denticles called rostral denticles. Isolated rostral denticles of the sclerorhynchoid Ischyrhiza mira are commonly recovered from Late Cretaceous North American marine deposits. Although the external morphology has been thoroughly presented in the literature, very little is known about the histological composition and organization of these curious structures. Using acid-etching techniques and scanning electron microscopy, we show that the microstructure of I. mira rostral denticles are considerably more complex than that of previously described dermal denticles situated elsewhere on the body. The apical cap consists of outer single crystallite enameloid (SCE) and inner bundled crystallite enameloid (BCE) overlying a region of orthodentine. The BCE has distinct parallel bundled enameloid (PBE), tangled bundled enameloid (TBE), and radial bundled enameloid (RBE) components. Additionally, the cutting edge of the rostral denticle is produced by a superficial layer of SCE and a deeper ridges/cutting edge layer (RCEL) of the BCE. The highly organized enameloid observed in the rostral denticles of this batomorph resembles that of the multifaceted tissue architecture observed in the oral teeth of selachimorphs and demonstrates that dermal scales have the capacity to evolve histologically similar complex tooth-like structures both inside and outside the oropharyngeal cavity.

Evaluación de la mineralización en mandíbula y diente de trucha arcoíris (Oncorhynchus mykiss) mediante microscopía electrónica de barrido de presión variable acoplada a detector de espectroscopía de rayos X de energía dispersiva / Evaluation of mineralization in jaw and teeth of rainbow trout (Oncorhynchus mykiss) using variable pressure scanning electron microscopy coupled to energy-dispersive X-ray spectroscopy detector

RESUMEN: En salmonicultura se utilizan imágenes de rayos X, para evaluar la columna vertebral y las aletas, pero estas no permiten cuantificar los minerales que constituyen el hueso, para esto se utilizan otras técnicas que son destructivas. La Microscopía Electrónica de Barrido de presión variable (VP SEM) acoplada al detector espectroscopia de rayos X de energía dispersiva (EDX) acoplado, nos permite analizar la microestructura y a la vez determinar elementos químicos, porcentaje y distribución presentes en puntos específicos en una muestra. Se utilizaron 5 truchas control y 5 con deformidad mandibular, de agua dulce en etapa de smolt, se analizó la estructura y mineralización del hueso, se eligieron en promedio 6 puntos de cuantificación por región de interés y se analizaron mediante Microscopía Electrónica de Barrido (VP SEM-EDX). Los datos obtenidos muestran porcentaje en masa de Ca promedio en hueso articular de truchas fueron de 8.07 % y de 14.48 % en truchas con deformidad mandibular y control respectivamente y el porcentaje en masa promedio de P es de 4.07 % y 7.60 %, en truchas con deformidad mandibular y control respectivamente. Se identificó además otros elementos presentes en la muestra como Mg, Na, C, O, N, S, F, Zn, Al y Fe, con especial interés en el aumento de carbono en las muestras analizadas con deformidad mandibular y la presencia de aluminio en todas las muestras. La Técnica de VP SEM-EDX, permite evaluar de forma directa, sin destrucción de la muestra y con una preparación mínima de la muestra. En el hueso, la aplicación más frecuente de SEM-EDX es la medición del contenido de Ca y P y la relación que existe entre estos elementos Ca/P, en la muestra. Paralelamente la técnica nos permite la detección de otros microelementos provenientes del agua o de la alimentación y que eventualmente pueden provocar alteraciones en los peces, confirmando la hipótesis que el microanálisis elemental tiene utilidad para la salmonicultura.

SUMMARY: In salmon farming systems, X-ray images are used to evaluate the spine and fins, but these do not allow quantifying the minerals that make up the bone, for different techniques that are destructive are used. Variable Pressure Scanning Electron Microscopy (VP SEM) coupled to an Energy Dispersive X-ray spectroscopy detector (EDX), allows us to analyze the microstructure and at the same time determine chemical elements, percentages, and distribution present at specific points in a sample. Five control and five jaw deformity trout, from freshwater and in the smolt stage were used. The structure and mineralization of the bone were analyzed, an average six quantification points were chosen per region of interest (ROI) and then they were analyzed by Scanning Electron Microscopy (VP SEM-EDX). The data obtained have shown the average mass percentage of calcium in trout joint bone was 8.07 % and 14.48 % in jaw deformity and control trout, respectively; and the average mass percentage of phosphorus is 4.07 % and 7.60 %, in jaw deformity and control trout, respectively. Other elements present in the sample were also identified, such as magnesium, sodium, carbon, oxygen, nitrogen, sulphur, fluorine, zinc, aluminium, and iron, with special interest the increase of carbon in the analyzed samples with mandibular deformity and the presence of aluminum in all samples. The VP SEM-EDX Technique allows direct evaluation, without destruction of the sample and with minimal sample preparation. In bone, the most frequent application of SEM-EDX is the measurement of the content of calcium (Ca) and phosphorus (P) and the relationship that exists between these elements, calcium/phosphorus (Ca/P), in the sample. At the same time, the technique allows us to detect other microelements from water or food that can eventually cause alterations in fish, confirming the hypothesis that elemental microanalysis is useful for salmon farming.

Toxicity Assessment of Long-Term Exposure to Non-Thermal Plasma Activated Water in Mice.

Non-thermal plasma activated water (PAW) has recently emerged as a powerful antimicrobial agent. Despite numerous potential bio-medical applications, studies concerning toxicity in live animals, especially after long-term exposure, are scarce. Our study aimed to assess the effects of long-term watering with PAW on the health of CD1 mice. PAW was prepared from distilled water with a GlidArc reactor according to a previously published protocol. The pH was 2.78. The mice received PAW (experimental group) or tap water (control group) daily for 90 days as the sole water source. After 90 days, the following investigations were performed on the euthanatized animals: gross necropsy, teeth mineral composition, histopathology, immunohistochemistry, hematology, blood biochemistry, methemoglobin level and cytokine profile. Mice tolerated PAW very well and no adverse effects were observed during the entire period of the experiment. Histopathological examination of the organs and tissues did not reveal any structural changes. Moreover, the expression of proliferation markers PCNA and Ki67 has not been identified in the epithelium of the upper digestive tract, indicating the absence of any pre- or neoplastic transformations. The results of our study demonstrated that long-term exposure to PAW caused no toxic effects and could be used as oral antiseptic solution in dental medicine.

An Ultrastructural Study on the Effect of High Temperatures on Teeth and Restorative Materials That Aids in the Identification of Human Remains.

INTRODUCTION: In most disasters, teeth are the only means of positive identification of an otherwise unrecognizable body, as there has been tremendous increase in the use of dental restorations that have different resistance to prolonged high temperature which is an important aid in identifying burned victims. Application of SEM/EDS in forensics was found useful in areas where there is a need for good imaging with high magnification combined with elemental analysis. The objective of this study was to analyze incineration effects on teeth and restorative materials using SEM/EDS. Materials and Method. 128 extracted teeth were collected, 96 were restored with silver amalgam, composite, and GIC of 32 each, and crown preparation was done in 32 teeth for which metal ceramic crowns were prepared. These teeth were subjected to 4 different temperatures (500°C, 700°C, 900°C, and 1100°C) for 20 minutes, and they were analyzed macroscopically and by using SEM for the changes subsequent to their exposure to such high temperatures. RESULTS: All the restorations which were very difficult to identify by naked eye were identified with the help of SEM/EDS. CONCLUSION: Elemental analysis of the specific restorative material proves to be an essential tool for the forensic odontologist.

Novel Approach to Tooth Chemistry. Quantification of the Dental-Enamel Junction.

The dentin-enamel junction (DEJ) is known for its special role in teeth. Several techniques were applied for the investigation of the DEJ in human sound molar teeth. The electron (EPMA) and proton (PIXE) microprobes gave consistent indications about the variability of elemental concentrations on this boundary. The locally increased and oscillating concentrations of Mg and Na were observed in the junction, in the layer adhering to the enamel and covering roughly half of the DEJ width. The chemical results were compared with the optical profiles of the junction. Our chemical and optical results were next compared with the micromechanical results (hardness, elastic modulus, friction coefficient) available in the world literature. A strong correlation of both result sets was proven, which testifies to the self-affinity of the junction structures for different locations and even for different kinds of teeth and techniques applied for studies. Energetic changes in tooth strictly connected with crystallographic transformations were calculated, and the minimum energetic status was discovered for DEJ zone. Modeling of both walls of the DEJ from optical data was demonstrated. Comparing the DEJ in human teeth with the same structure found in dinosaur, shark, and alligator teeth evidences the universality of dentin enamel junction in animal world. The paper makes a contribution to better understanding the joining of the different hard tissues.

Ultrastructural studies of developing egg tooth in grass snake Natrix natrix (Squamata, Serpentes) embryos, supported by X-ray microtomography analysis.

The egg tooth development is similar to the development of all the other vertebrate teeth except earliest developmental stages because the egg tooth develops directly from the oral epithelium instead of the dental lamina similarly to null generation teeth. The developing egg tooth of Natrix natrix changes its curvature differently than the egg tooth of the other investigated unidentates due to the presence of the rostral groove. The developing grass snake egg tooth comprises dental pulp and the enamel organ. The fully differentiated enamel organ consists of outer enamel epithelium, stellate reticulum, and ameloblasts in its inner layer. The enamel organ directly in contact with the oral cavity is covered with periderm instead of outer enamel epithelium. Stellate reticulum cells in the grass snake egg tooth share intercellular spaces with the basal part of ameloblasts and are responsible for their nutrition. Ameloblasts during egg tooth differentiation pass through the following stages: presecretory, secretory, and mature. The ameloblasts from the grass snake egg tooth show the same cellular changes as reported during mammalian amelogenesis but are devoid of Tomes' processes. Odontoblasts of the developing grass snake egg tooth pass through the following classes: pre-odontoblasts, secretory odontoblasts, and ageing odontoblasts. They have highly differentiated secretory apparatus and in the course of their activity accumulate lipofuscin. Grass snake odontoblasts possess processes which are poor in organelles. In developing egg tooth cilia have been identified in odontoblasts, ameloblasts and cells of the stellate reticulum. Dental pulp cells remodel collagen matrix during growth of the grass snake egg tooth. They degenerate in a way previously not described in other teeth.

Miniaturization: How many cells are needed to build a tooth?

BACKGROUND: Organs that develop early in life, and are replaced by a larger version as the animal grows, often represent a miniature version of the adult organ. Teeth constituting the first functional dentition in small-sized teleost fish, such as medaka (Oryzias latipes), are examples of such miniature organs. With a dentin cone as small as the size of one human cell, or even smaller, these teeth raise the question how many dentin-producing cells (odontoblasts) are required to build such a tooth, and whether this number can be as little as one. RESULTS: Based on detailed observations with transmission electron microscopy (TEM) and TEM-based 3D-reconstructions, we show that only one mesenchymal cell qualifies as a true odontoblast. A second mesenchymal cell potentially participates in dentin formation, but only at a late stage of tooth development. Moreover, the fate of these cells appears to be specified very early during tooth development. CONCLUSIONS: Our observations indicate that in this system, one single odontoblast fulfills roles normally exerted by a large and communicating cell population. First-generation teeth in medaka thus provide an exciting model to study integration of multiple functions into a single cell.

Spectroscopic and microscopic examination of teeth exposed to green tea at different temperatures.

Tea is a popular beverage consumed at different temperatures. The effect of tea on teeth at different temperatures has not been studied previously. The present study used an in vitro green tea immersed tooth model at different tea temperatures (hot and cold) compared to an in vivo tea administration model allowing rats to drink tea over the course of a week. The elements present in tea leaves were identified by Inductively Coupled Plasma Mass Spectrometry (ICP-MS) and compared to the elements in teeth (enamel surface) using Laser-Induced Breakdown Spectroscopy (LIBS). Here, LIBS demonstrated in vivo and in vitro green tea treatments resulted in a significant increase in the mineral elements found in enamel. For the in vitro assessment, elements in enamel varied based on cold-tea and hot-tea treatment; however, hot water reduced the elements in enamel. Atomic force microscopy found the in vivo tea group had a higher roughness average (RA) compared with the in vivo water group. Cold tea and hot tea in vitro groups demonstrated lower RA than in vitro water controls. Scanning electron microscopy found hot water induced cracks more than 1.3µm in enamel while cold tea and hot tea promoted the adhering of extrinsic matter to teeth. Overall, teeth treated to high temperature lost the mineral phase leading to demineralization. Our results indicate that green tea protects enamel, but its protective action in dental structures is enhanced at cold temperature.

Fossil microbial shark tooth decay documents in situ metabolism of enameloid proteins as nutrition source in deep water environments.

Alteration of organic remains during the transition from the bio- to lithosphere is affected strongly by biotic processes of microbes influencing the potential of dead matter to become fossilized or vanish ultimately. If fossilized, bones, cartilage, and tooth dentine often display traces of bioerosion caused by destructive microbes. The causal agents, however, usually remain ambiguous. Here we present a new type of tissue alteration in fossil deep-sea shark teeth with in situ preservation of the responsible organisms embedded in a delicate filmy substance identified as extrapolymeric matter. The invading microorganisms are arranged in nest- or chain-like patterns between fluorapatite bundles of the superficial enameloid. Chemical analysis of the bacteriomorph structures indicates replacement by a phyllosilicate, which enabled in situ preservation. Our results imply that bacteria invaded the hypermineralized tissue for harvesting intra-crystalline bound organic matter, which provided nutrient supply in a nutrient depleted deep-marine environment they inhabited. We document here for the first time in situ bacteria preservation in tooth enameloid, one of the hardest mineralized tissues developed by animals. This unambiguously verifies that microbes also colonize highly mineralized dental capping tissues with only minor organic content when nutrients are scarce as in deep-marine environments.

Using tooth enamel microstructure to identify mammalian fossils at an Eocene Arctic forest.

Lower Eocene (Wasatchian-aged) sediments of the Margaret Formation on Ellesmere Island in Canada's High Arctic preserve evidence of a rainforest inhabited by alligators, turtles, and a diverse mammalian fauna. The mammalian fossils are fragmentary and often poorly preserved. Here, we offer an alternative method for their identification. Among the best preserved and extensive of the Eocene Arctic forests is the Strathcona Fiord Fossil Forest, which contains permineralized in situ tree stumps protruding from a prominent coal seam, but a paucity of vertebrate fossils. In 2010 and 2018, we recovered mammalian tooth fragments at the fossil forest, but they are so incomplete as to be undiagnostic by using their external morphology. We used a combination of light microscopy and SEM analysis to study the enamel microstructure of two tooth fragments from the fossil forest-NUFV2092B and 2092E. The results of our analysis indicate that NUFV2092B and 2092E have Coryphodon-enamel, which is characterized by vertical bodies that manifest as bands of nested chevrons or treelike structures visible in the tangential section under light microscopy. This enamel type is not found in other mammals known from the Arctic. Additionally, when studied under SEM, the enamel of NUFV2092B and 2092E has rounded prisms that open to one side and are surrounded by interprismatic matrix that is nearly parallel to the prisms, which also occurs in Coryphodon enamel, based on prior studies. The tooth fragments reported here, along with some poorly preserved bone fragments, thus far are the only documented vertebrate fossils from the Strathcona Fiord Fossil Forest. However, fossils of Coryphodon occur elsewhere in the Margaret Formation, so its presence at the fossil forest is not surprising. What is novel in our study is the way in which we identified the fossils using their enamel microstructure.

Radula formation in two species of Conoidea (Gastropoda).

The radula is the basic feeding structure in gastropod molluscs and exhibits great morphological diversity that reflects the exceptional anatomical and ecological diversity occurring in these animals. This uniquely molluscan structure is formed in the blind end of the radular sac by specialized cells (membranoblasts and odontoblasts). Secretion type, and the number and shape of the odontoblasts that form each tooth characterize the mode of radula formation. These characteristics vary in different groups of gastropods. Elucidation of this diversity is key to identifying the main patterns of radula formation in Gastropoda. Of particular interest would be a phylogenetically closely related group that is characterized by high variability of the radula. One such group is the large monophyletic superfamily Conoidea, the radula of which is highly variable and may consist of the radular membrane with five teeth per row, or the radular membrane with only two or three teeth per row, or even just two harpoon-like teeth per row without a radular membrane. We studied the radulae of two species of Conoidea (Clavus maestratii Kilburn, Fedosov & Kantor, 2014 [Drilliidae] and, Lophiotoma acuta (Perry, 1811) [Turridae]) using light and electron microscopy. Based on these data and previous studies, we identify the general patterns of the radula formation for all Conoidea: the dorsolateral position of two groups of odontoblasts, uniform size, and shape of odontoblasts, folding of the radula in the radular sac regardless of the radula configuration. The morphology of the subradular epithelium is most likely adaptive to the radula type.

Seeing is believing? When scanning electron microscopy (SEM) meets clinical dentistry: The replica technique.

In restorative dentistry, the in situ replication of intra-oral situations, is based on a non-invasive and non-destructive scanning electron microscopy (SEM) evaluation method. The technique is suitable for investigation restorative materials and dental hard- and soft-tissues, and its interfaces. Surface characteristics, integrity of interfaces (margins), or fracture analysis (chipping, cracks, etc.) with reliable resolution and under high magnification (from ×50 to ×5,000). Overall the current study aims to share detailed and reproducible information about the replica technique. Specific goals are: (a) to describe detailed each step involved in producing a replica of an intra-oral situation, (b) to validate an integrated workflow based on a rational sequence from visual examination, to macrophotography and SEM analysis using the replica technique; (c) to present three clinical cases documented using the technique. A compilation of three clinical situations/cases were analyzed here by means the replica technique showing a wide range of possibilities that can be reached and explored with the described technique. This guidance document will contribute to a more accurate use of the replica technique and help researchers and clinicians to understand and identify issues related to restorative procedures under high magnification.

A structural biorhythm related to human sexual dimorphism.

Life on earth is regulated by biological rhythms, some of which oscillate with a circadian, monthly or lunar cycle. Recent research suggests that there is a near weekly biorhythm that may exert an influence on human skeletal growth. Evidence for the timing of this biorhythm is retained in tooth enamel as the periodicity of Retzius lines. Studies report that Retzius periodicity (RP) relates to adult human stature and enamel thickness. Adult human stature is sexually dimorphic, and so is enamel thickness of maxillary third molars (M3) but not mandibular M3. Yet, previous studies report sex differences in RP are apparent in some populations but not others, and it is unknown if dimorphism in enamel thickness relates to RP. To further our understanding of this biorhythm we analysed sex-related variation in RP and its relationship with enamel thickness in a sample of M3's (n = 94) from adults in Northern Britain. Results reveal RP was significantly higher in our sample of female molars compared to those of males, which is consistent with the previously reported correlation between the biorhythm and adult stature. The RP of maxillary M3 related to sex differences in enamel thickness, but this relationship was not present in mandibular M3. Our results support previous findings suggesting that this biorhythm is sexually dimorphic and provide the first evidence that RP may be one factor influencing sex differences in enamel thickness. Our study also shows that correlations between RP and enamel thickness appear to be most readily detected for tooth types with sufficiently wide ranges of enamel thickness variation, as is the case for maxillary but not mandibular M3. Achieving a sufficient sample size was critical for detecting a sex difference in periodicity.

Unique three-dimensional structure of a fish pharyngeal jaw subjected to unusually high mechanical loads.

We examine the structure of the bone of the pharyngeal jaws of a large fish, the black drum (Pogonias cromis), that uses its tooth-jaw complex to crush hard-shelled bivalve mollusks. During mastication huge compressive forces are concentrated in a tiny zone at the tooth-bone interface. We report on the structure of this bone, with emphasis on its contact with the teeth, at different hierarchical levels and in 3D. Micro-CT shows that the molariform teeth do not have roots and are supported by a circular narrow bony rim that surrounds the periphery of the tooth base. The lower pharyngeal jaw is highly porous, as seen by reflected light microscopy and secondary electron microscopy (SE-SEM). Porosity decreases close to the bone-tooth interface and back-scattered electron (BSE-SEM) microscopy shows a slight elevation in mineral density. Focused ion beam - scanning electron microscopy (FIB-SEM) in the serial surface view (SSV) mode reveals a most surprising organization at the nanoscale level: parallel arrays of mineralized collagen fibrils surrounding channels of ~100 nm diameter, both with their long axes oriented along the load direction. The channels are filled with organic matter. These fibril-channel arrays are surrounded by a highly disordered mineralized material. This unusual structure clearly functions efficiently under compression, but the precise way by which this unique arrangement achieves this function is unknown.

Ultramicrostructural reductions in teeth: implications for dietary transition from non-avian dinosaurs to birds.

BACKGROUND: Tooth morphology within theropod dinosaurs has been extensively investigated and shows high disparity throughout the Cretaceous. Changes or diversification in feeding ecology, i.e., adoption of an herbivorous diet (e.g., granivorous), is proposed as a major driver of tooth evolution in Paraves (e.g., Microraptor, troodontids and avialans). Here, we studied the microscopic features of paravian non-avian theropod and avialan teeth using high-spatial-resolution synchrotron transmission X-ray microscopy and scanning electron microscopy. RESULTS: We show that avialan teeth are characterized by the presence of simple enamel structures and a lack of porous mantle dentin between the enamel and orthodentin. Reduced internal structures of teeth took place independently in Early Cretaceous birds and a Microraptor specimen, implying that shifts in diet in avialans from that of closely related dinosaurs may correlate with a shift in feeding ecology during the transition from non-avian dinosaurs to birds. CONCLUSION: Different lines of evidence all suggest a large reduction in biting force affecting the evolution of teeth in the dinosaur-bird transition. Changes in teeth microstructure and associated dietary shift may have contributed to the early evolutionary success of stemward birds in the shadow of other non-avian theropods.

Principal Component Regression for Forensic Age Determination Using the Raman Spectra of Teeth.

Raman spectra of mineralized tooth tissues were used to build a principal component regression (PCR) age determination model for forensic application. A sample of 71 teeth was obtained from donors aging from 11 to 76 years. No particular selection criteria were applied; teeth affected with various pathological processes were deliberately included to simulate a realistic forensic scenario. In order to comply with the nondestructive specimen handling, Raman spectra were collected from tooth surfaces without any previous preparation. Different tooth tissues were evaluated by collecting the spectra from three distinct sites: tooth crown, tooth neck, and root apex. Whole recorded spectra (3500-200 cm-1) were used for principal component analysis and building of the age determination model using PCR. The predictive capabilities of the obtained age determination models varied according to the spectra collection site. Optimal age determination was attained by using Raman spectra collected from cementum at root apex (R2 values of 0.84 and 0.71 for male and female donors, respectively). For optimal performance of that model, male and female donors had to be analyzed separately, as merging both genders into a single model considerably diminished its predictive capability (R2 = 0.29).

In situ 3D visualization of biomineralization matrix proteins.

Calcium biominerals occur in all major animal phyla, and through biomolecular control, exhibit such diverse structures as exoskeletons, shells, bones, teeth and earstones (otoliths). Determining the three-dimensional expression of key biomineral proteins, however, has proven challenging as typical protein identification methods either lose spatial resolution during dissolution of the mineral phase or are costly and limited to two-dimensional expression of high abundance proteins. Here we present a modification of the CLARITY and ACT-PRESTO protocols to visualize and confirm, for the first time, the timing of expression and function of two key regulators of biomineralization.

Growing up Tyrannosaurus rex: Osteohistology refutes the pygmy "Nanotyrannus" and supports ontogenetic niche partitioning in juvenile Tyrannosaurus.

Despite its iconic status as the king of dinosaurs, Tyrannosaurus rex biology is incompletely understood. Here, we examine femur and tibia bone microstructure from two half-grown T. rex specimens, permitting the assessments of age, growth rate, and maturity necessary for investigating the early life history of this giant theropod. Osteohistology reveals these were immature individuals 13 to 15 years of age, exhibiting growth rates similar to extant birds and mammals, and that annual growth was dependent on resource abundance. Together, our results support the synonomization of "Nanotyrannus" into Tyrannosaurus and fail to support the hypothesized presence of a sympatric tyrannosaurid species of markedly smaller adult body size. Our independent data contribute to mounting evidence for a rapid shift in body size associated with ontogenetic niche partitioning late in T. rex ontogeny and suggest that this species singularly exploited mid- to large-sized theropod niches at the end of the Cretaceous.

A histological and ultrastructural study of equine peripheral caries.

BACKGROUND: Peripheral caries (PC) has recently been shown to affect over half of UK and Australian horses. OBJECTIVES: To examine affected cheek teeth histologically and ultrastructurally, to assess the possible route(s) of carious infection into the dental tissues and to further describe the pathological changes in affected teeth. STUDY DESIGN: Descriptive pathological study. METHODS: Four control and 16 cheek teeth with different grades of PC were examined grossly, histologically and by electron microscopy. RESULTS: Cariogenic bacteria from plaque entered peripheral cementum perpendicular to the sides of teeth alongside Sharpey's fibres or vascular channels or more horizontally alongside exposed intrinsic fibres and cemental growth lines. Subsequent intra-cemental bacterial spread created different patterns of caries including vertical and horizontal flake-like patterns; deep, large flask-shaped or smaller superficial ellipsoid-shaped lytic lesions. Cemental caries progressed to affect enamel, dentine and even pulp. Gross examination underestimated the degree of caries observed on histological evaluation. MAIN LIMITATIONS: No dental histories were available; all teeth were examined on a single occasion. CONCLUSION: Equine PC causes different patterns of cemental lesions that appear dependent on the route of cariogenic bacterial invasion. Progression of caries can lead to pulpitis and tooth loss. Gross examination can underestimate the true extent of caries.