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Age estimation based on blood DNA methylation levels in brown bears
Authors:Shiori Nakamura  Jumpei Yamazaki  Naoya Matsumoto  Miho Inoue-Murayama  Huiyuan Qi  Masami Yamanaka  Masanao Nakanishi  Yojiro Yanagawa  Mariko Sashika  Toshio Tsubota  Hideyuki Ito  Michito Shimozuru
Institution:1. Laboratory of Wildlife Biology and Medicine, Faculty of Veterinary Medicine, Hokkaido University, Sapporo, Japan;2. Translational Research Unit, Veterinary Teaching Hospital, Faculty of Veterinary Medicine, Hokkaido University, Sapporo, Japan

One Health Research Center, Hokkaido University, Sapporo, Japan;3. Noboribetsu Bear Park, Noboribetsu, Japan;4. Wildlife Research Center, Kyoto University, Kyoto, Japan;5. Shiretoko Nature Foundation, Shari, Japan;6. Laboratory of Theriogenology, Faculty of Veterinary Medicine, Hokkaido University, Sapporo, Japan

Abstract:Age is an essential trait for understanding the ecology and management of wildlife. A conventional method of estimating age in wild animals is counting annuli formed in the cementum of teeth. This method has been used in bears despite some disadvantages, such as high invasiveness and the requirement for experienced observers. In this study, we established a novel age estimation method based on DNA methylation levels using blood collected from 49 brown bears of known ages living in both captivity and the wild. We performed bisulfite pyrosequencing and obtained methylation levels at 39 cytosine-phosphate-guanine (CpG) sites adjacent to 12 genes. The methylation levels of CpGs adjacent to four genes showed a significant correlation with age. The best model was based on DNA methylation levels at just four CpG sites adjacent to a single gene, SLC12A5, and it had high accuracy with a mean absolute error of 1.3 years and median absolute error of 1.0 year after leave-one-out cross-validation. This model represents the first epigenetic method of age estimation in brown bears, which provides benefits over tooth-based methods, including high accuracy, less invasiveness, and a simple procedure. Our model has the potential for application to other bear species, which will greatly improve ecological research, conservation, and management.
Keywords:age estimation  aging  brown bear  DNA methylation  epigenetic clock  wildlife management
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