Epigenetic Clock in Bears: A Simple Cost-Effective Blood DNA Methylation-Based Age Estimation Method Applicable to Multiple Bear Species.

Age is an essential factor to understand the life history and demographic parameters of wildlife. Previously, we established an age estimation method for brown bears based on blood DNA methylation level. In this study, we first applied the brown bear-specific age estimation model to other bear species, including Asian black, polar, sun, and Andean bears. Using blood DNA, we performed bisulfite pyrosequencing to determine the methylation levels at four cytosine-phosphate-guanine (CpG) sites adjacent to a single gene, . The best model specific to brown bears estimated their ages with satisfactory accuracy, with mean absolute error (MAE) of 1.5, 2.1, 2.2, and 0.4 years for Asian black (52 samples from 16 captive and 36 wild bears), polar (27 samples from 21 captive bears), sun bears (11 samples from 8 captive bears), and Andean bears (one captive bear), respectively. Then, we established an Asian black bear-specific age estimation model and a common age estimation model applicable for other bear species (i.e., a pan-bear model) using the methylation levels of the four CpG sites. The best model specific to Asian black bears had high accuracy with MAE of 1.1 after leave-one-out cross-validation (LOOCV). In addition, the best pan-bear model achieved accuracy with MAE of 1.3, 1.2, 2.1, and 2.2 years after LOOCV for brown, Asian black, polar, and sun bears, respectively. The results suggested that the pan-bear age estimation model using the aging marker (CpG sites adjacent to ) is a simple, highly accurate, and cost-effective tool that is applicable to Ursidae.