Age Analysis in Odontocetes Cetaceans PDF

Age analysis in odontocete cetaceans Age assessment in cetaceans provides relevant information on demographic characteristics of populations (e.g. longevity, growth rates, age at reproductive maturity). Likewise, it allows us to assess how age may influence the variation in biological and ecologica...

Age analysis in odontocete cetaceans Age assessment in cetaceans provides relevant information on demographic characteristics of populations (e.g. longevity, growth rates, age at reproductive maturity). Likewise, it allows us to assess how age may influence the variation in biological and ecological characteristics (e.g. diet) or even the effect of anthropogenic factors (e.g. accumulation of pollutants). In odontocetes, assessment of the age of individuals is traditionally done by counting groups of growth layers in teeth, which result from the deposition of minerals in dentin. Each growth group, composed of two layers, usually corresponds to one year. The study of age in odontocetes is traditionally done through the analysis of histological sections of teeth. Teeth histology protocol 1 – Cleaning Remove excess tissue from the teeth and rinse in tap water. Fig. 1 – Cleaned harbour porpoise (Phocoena phocoena) teeth. 2 – Decalcification Place the teeth in histological cassettes for decalcification until they are rubbery in texture, and rinse in running tap water for several hours. 3 – Sectioning In porpoises two teeth from each individual are necessary for sectioning in two orientations: one parallel to the mandible (dolphin cut) and one perpendicular to the mandible (porpoise cut). In delphinid species it is only necessary to make one longitudinal section cut, perpendicular to the mandible (dolphin cut). Fig. 2 – Sectioned harbour porpoise teeth. 4 – Dehydration Use a tissue processor for the dehydration and embedding in paraffin wax. A sequential ethanol series is used followed by xylene and molten wax. 5 – Paraffin blocks Put the tooth face down in the metal mould, then the cassette on top, and fill it with liquid paraffin. 6 – Microtome Sectioning Section longitudinally in a standard paraffin microtome. Select the most central and complete sections from each tooth and place them in histological slides. 7 – Remove paraffin wax Oven-dry the sections for 15min at 100ºC and rehydrate (xylene+ethanol series),to allow staining. 8 – Staining and Mounting Place slides in a Mayer’s hematoxylin solution to stain the sections, then rinse in tap water. Follow by placing in a weak ammonium solution and rinse again in tap water. Once slides are fully dried, cover sections with a glass coverslip using DPX-mounting medium. 9 – Reading Count growth layer groups (GLGs) in the dentine under a binocular microscope. A- Cement B- Prenatal dentin C- Neonatal line D- Most recent zone in the process of formation E- Cementum (gum area) 1- Dentin deposited during the first year 2- Dentin deposited during the second year 3- Dentin deposited during the third year 4- Dentin deposited during the fourth year 5- Dentin deposited during the fifth year 6- Formation of the sixth layer of dentin Section of 5-year-old porpoise tooth (adapted from Gaskin and Blair, 1976).