A Model for Dental Age Verification Using Ultrastructural Imaging for Modern and Fossil Representatives of the Rhinocerotidae Family

SIMPLE SUMMARY: Determining the age of an animal based on remains, especially fossil remains, is challenging. The age of an individual can be estimated from ultrastructural analyses of teeth. Therefore, in this study, an attempt was made to establish a relationship between the age of a rhinoceros and the ultrastructure of its tooth. The subject of these examinations was a tooth that originated from a female white rhinoceros that died at Kiev Zoo. The age of the female was known. She did not give birth during her life, so pregnancy did not influence the tooth ultrastructure. For ultrastructural examination, ground sections of the tooth were obtained and slides were observed under a light microscope in white light as well as in polarized light with the use of a lambda filter. In addition, gold-coated preparations were imaged in a scanning electron microscope. Three sections were cut out of the specimen: Horizontal through the tooth crown, horizontal through the upper part of the root, and longitudinal. In subsequent stages they were investigated to see the annual growth lines of mineralized dental tissues of cement and dentine. These lines were counted from the root canal center to the cheek surface of the tooth. The most satisfactory results were obtained on the horizontal section through the upper part of the root, where distinct growth lines were observed in the dentine, and their number for both roots was consistent with the known chronological age of the animal. ABSTRACT: The analyses were performed on a right third premolar (P(3)) of a white rhinoceros female (Ceratotherium simum, Burchell 1817). The specimen was born in captivity at London Zoo (Zoological Society of London), then in the 1970s transferred to Kiev Zoo (Peremohy Avenue), Ukraine, and was kept there until it died at a documented chronological age of 48 years. The female died because of its age, which indicates it was kept in good conditions adequate to the requirements of this species. Photographs and micrographs with radiological documentation were taken on the said tooth. Its structural characteristics were determined, and on the occlusal surface areas and points of anatomical constitution of its crown were identified. The tooth was also histologically evaluated via sections taken horizontally in a mesial-distal plane through the crown, horizontally in a mesial-distal plane through the coronal portion of the root, and longitudinally in a lingual-buccal plane through the crown and the root. Preparations with ground sections were made and observed in white, polarized, and reflected light. In the subsequent stage X-ray and SEM imaging has also been used, for analysis of the distribution of annual growth layers of mineralized dental tissues of cement and dentine, counted from the root canal center to the buccal surface. An attempt was also made to confirm the annual season in which the animal died, based on cement growth lines. It was observed that the growth lines were visible in all the analyzed sections, in dentine and cement. In the cement, the lines were relatively few and did not represent the attested age of the animal. The analysis of the coloration of the cement lines indicated that the animal was regularly fed a diet that was not seasonally differentiated. From the X-ray examination comes a conclusion that the animal did not suffer from periodontal diseases. Visible growth lines were observed on the dentine. On the horizontal section through the crown growth lines in the dentine were few and unclear. On the longitudinal section, both on the caudal and rostral roots, these lines were clearly visible and much more numerous than expected considering the known age of the animal, as more than 50 were counted. On horizontal sections through the upper part of both roots, distinct growth lines were observed in the dentine, and their number—48 for both roots—corresponded precisely to the age of the animal. The results of our study indicate that this method has significant potential for application to verify the age at death for modern and fossil representatives of rhinoceros.