Periodic Reporting for period 1 - ENEVOLVE (A novel approach to characterizing prismless enamel in modern and fossil reptiles)
Reporting period: 2020-10-01 to 2022-09-30
ENEVOLVE is a multi-disciplinary study of the properties of dental enamel in living and extinct reptiles through beyond-state-of-the-art techniques. Palaeontologists have long used teeth to interpret the diets of extinct species. As the hardest parts of the vertebrate body, teeth are the most likely to be preserved in the fossil record, and their use in palaeontology relies on extrapolating tooth shape, structure, and function in living animals to similar fossil analogues. While extant-to-extinct comparisons work well for mammals, because of an already detailed understanding of the material properties of mammal dental tissues, the same cannot be said for the teeth of reptiles. Reptiles have much thinner, structurally simpler enamel with poorly understood material properties in most living species, let alone in fossil groups. Enamel is also unique among dental tissues in that it is unable to re-grow or regenerate once it is worn away. As such, enamel structure and chemistry in most vertebrates has likely been under intense selection to optimize its function. Understanding how structurally simpler forms of enamel – like those of reptiles – cope with the stresses and strains of different diets would not only improve palaeontologists’ ability to interpret tooth form and function in the fossil record, but could lead to bio-inspired solutions for artificial enamel regrowth in humans.
In order to better understand tooth form and function in living and extinct reptiles, ENEVOLVE used advanced X-ray, mass spectrometry, and electron-based imaging, as well as nanomechanical testing to reveal the material properties of key tooth morphotypes in reptiles. These included the serrated cutting teeth of tyrannosaurid dinosaurs and Komodo dragons, the piercing and crushing teeth of living and fossil crocodiles and their relatives, and the grinding teeth of reptilian herbivores. The Research Objectives (ROs) of ENEVOLVE are to (1) characterize the spatial distribution and composition of the dental hard tissues in modern and fossil teeth of specific morphotypes; (2-3) quantify enamel and dentine crystal structure and chemical composition in extant tooth morphotypes and some of their extinct analogues; (4) identify structure-function relationships in reptile enamels.
In order to better understand tooth form and function in living and extinct reptiles, ENEVOLVE used advanced X-ray, mass spectrometry, and electron-based imaging, as well as nanomechanical testing to reveal the material properties of key tooth morphotypes in reptiles. These included the serrated cutting teeth of tyrannosaurid dinosaurs and Komodo dragons, the piercing and crushing teeth of living and fossil crocodiles and their relatives, and the grinding teeth of reptilian herbivores. The Research Objectives (ROs) of ENEVOLVE are to (1) characterize the spatial distribution and composition of the dental hard tissues in modern and fossil teeth of specific morphotypes; (2-3) quantify enamel and dentine crystal structure and chemical composition in extant tooth morphotypes and some of their extinct analogues; (4) identify structure-function relationships in reptile enamels.
ENEVOLVE has achieved several important milestones:
Successful collection and tissue-level characterization of fossil and extant reptile teeth: The success of ENEVOLVE hinged on choosing appropriate pairs of fossil and extant tooth types to compare and contrast in terms of their structure, chemistry, and material properties. The PI and Dr. LeBlanc were able to establish collaborations with the UK’s only crocodile zoo “Crocodiles of the World”, Imperial College London, The Natural History Museum, the University of Alberta (Canada), and The Museum of Life Sciences (King’s College London) to acquire dental specimens. This project sampled the serrated teeth of Komodo dragons and tyrannosaurid dinosaurs, multiple types of grasping and crushing teeth of extant and extinct crocodylians, and the unusual grinding teeth of an extinct group of reptiles known as sphenodontians for the proposed downstream analyses.
Successful applications and significant results from seven synchrotron experiments: The project relied on synchrotron-based experiments (X-ray diffraction/XRD, fluorescence/XRF, and nanotomography/XNT), which require the submission of competitive project proposals for instrument time at a select number of synchrotron facilities across the globe. From January 2021 to the end of the project (October 2022), ENEVOLVE resulted in six successful synchrotron beam time proposals directly related to the objectives of the project, and an additional synchrotron experiment aimed at developing a new imaging technique (experiment #7) at three synchrotron facilities in Paris, Grenoble, and Oxford. These multi-day experiments yielded several terabytes of quantitative data on enamel and dentine crystal structure in reptiles (tyrannosaurid dinosaurs, fossil and modern crocodylians) that are being analyzed for at least three key ENEVOLVE publications.
Significant results addressing original Research Objectives: The combination of all of these techniques have yielded at least three important results that are currently being prepared for publication: the unexpected structural and chemical complexity of serrated reptile teeth, the effects of fossilization on the material properties of reptile teeth, and the first report of mechanically sensitive enamel in reptiles. The results and impact of these are discussed in the subsequent section.
So far, these results have been presented at institutional seminars and most recently at the Society of Vertebrate Paleontology annual conference held in Toronto, Canada (Nov. 2022).
Successful collection and tissue-level characterization of fossil and extant reptile teeth: The success of ENEVOLVE hinged on choosing appropriate pairs of fossil and extant tooth types to compare and contrast in terms of their structure, chemistry, and material properties. The PI and Dr. LeBlanc were able to establish collaborations with the UK’s only crocodile zoo “Crocodiles of the World”, Imperial College London, The Natural History Museum, the University of Alberta (Canada), and The Museum of Life Sciences (King’s College London) to acquire dental specimens. This project sampled the serrated teeth of Komodo dragons and tyrannosaurid dinosaurs, multiple types of grasping and crushing teeth of extant and extinct crocodylians, and the unusual grinding teeth of an extinct group of reptiles known as sphenodontians for the proposed downstream analyses.
Successful applications and significant results from seven synchrotron experiments: The project relied on synchrotron-based experiments (X-ray diffraction/XRD, fluorescence/XRF, and nanotomography/XNT), which require the submission of competitive project proposals for instrument time at a select number of synchrotron facilities across the globe. From January 2021 to the end of the project (October 2022), ENEVOLVE resulted in six successful synchrotron beam time proposals directly related to the objectives of the project, and an additional synchrotron experiment aimed at developing a new imaging technique (experiment #7) at three synchrotron facilities in Paris, Grenoble, and Oxford. These multi-day experiments yielded several terabytes of quantitative data on enamel and dentine crystal structure in reptiles (tyrannosaurid dinosaurs, fossil and modern crocodylians) that are being analyzed for at least three key ENEVOLVE publications.
Significant results addressing original Research Objectives: The combination of all of these techniques have yielded at least three important results that are currently being prepared for publication: the unexpected structural and chemical complexity of serrated reptile teeth, the effects of fossilization on the material properties of reptile teeth, and the first report of mechanically sensitive enamel in reptiles. The results and impact of these are discussed in the subsequent section.
So far, these results have been presented at institutional seminars and most recently at the Society of Vertebrate Paleontology annual conference held in Toronto, Canada (Nov. 2022).
Unexpected complexity of enamel in the serrated teeth of Komodo dragons and tyrannosaurid dinosaurs: The results from several of the synchrotron experiments (1, 4, 5 and preliminary experiment) have shown that despite having thin enamel coatings, the serrated teeth of tyrannosaurid dinosaurs exhibit unexpected structural complexity that can be quantified and visualized in XRD analyses. Tyrannosaurid enamel shows structural adaptations for mitigating microfracture formation along the cutting edges, which more closely resembles enamel specializations in the grinding teeth of herbivorous dinosaurs. Our comparisons with the extant analogue, the Komodo dragon, revealed for the first time that some lizards have iron-coated cutting edges- allowing them to maintain a more resilient serrated edge to the teeth through chemical alteration of enamel rather than structural modification. These adaptations were thought to be restricted to mammals.
Fossilization alters the material properties of reptilian dentine and enamel differently: Synchrotron XRF experiments have shown that fossil reptile teeth have undergone significant chemical alterations compared to their extant counterparts. Moreover, these alterations affected the enamel and dentine differently and significantly altered the mechanical properties of enamel and dentine such that they become nearly indistinguishable during nanomechanical testing. This challenges previous assumptions about the ability to extract biologically relevant mechanical properties from fossilized teeth and reinforces the need to select appropriate extant analogues for extrapolating material properties of extinct reptile dentitions.
Some reptiles evolved “sensitive” enamel: XNT analysis of fossil sphenodontian teeth from the Triassic of England has revealed that some reptiles with grinding teeth have evolved a specialized form of enamel that must have been sensitive to chemical and mechanical stimuli. Tomography scans of <1µm resolution provided the opportunity to map small tubules extending from the sensitive dentine, across the enamel-dentine boundary, through the entire enamel, and out onto the external surfaces of the teeth, The connections between the outer environment and the innermost pulp of the tooth would normally be a pathway for infection, however, in these reptiles, enamel and dentine tubules must have provided vital sensitivity for the teeth, co-opting enamel into a new function never seen before in reptiles.
Fossilization alters the material properties of reptilian dentine and enamel differently: Synchrotron XRF experiments have shown that fossil reptile teeth have undergone significant chemical alterations compared to their extant counterparts. Moreover, these alterations affected the enamel and dentine differently and significantly altered the mechanical properties of enamel and dentine such that they become nearly indistinguishable during nanomechanical testing. This challenges previous assumptions about the ability to extract biologically relevant mechanical properties from fossilized teeth and reinforces the need to select appropriate extant analogues for extrapolating material properties of extinct reptile dentitions.
Some reptiles evolved “sensitive” enamel: XNT analysis of fossil sphenodontian teeth from the Triassic of England has revealed that some reptiles with grinding teeth have evolved a specialized form of enamel that must have been sensitive to chemical and mechanical stimuli. Tomography scans of <1µm resolution provided the opportunity to map small tubules extending from the sensitive dentine, across the enamel-dentine boundary, through the entire enamel, and out onto the external surfaces of the teeth, The connections between the outer environment and the innermost pulp of the tooth would normally be a pathway for infection, however, in these reptiles, enamel and dentine tubules must have provided vital sensitivity for the teeth, co-opting enamel into a new function never seen before in reptiles.