Integrated lipidomics and gut microbiome analyses to understand metabolic adaptation during hibernation in Pelodiscus sinensis

Hibernation is the most critical and vulnerable stage in the life cycle of animals, during which organisms must maintain energy homeostasis and undergo metabolic regulation under prolonged low temperatures and fasting conditions. Turtles, as ancient ectothermic vertebrates and one of the most threatened animal groups, remain poorly understood regarding how gut microbial communities participate in lipid metabolic remodeling through metabolic regulatory networks during hibernation. In this study, the Chinese soft-shelled turtle ( Pelodiscus sinensis ) was used to investigate metabolic adaptation during the summer active (SA), hibernation (H), and early arousal (EA) periods through integrated analysis of lipidomics, gut microbiome, and AMPK–mTOR pathway-related gene expression. The results showed that the hepatic lipid composition underwent significant remodeling during hibernation, with glycerophospholipid (GP) and sphingolipid (SP) metabolism identified as the major altered pathways. Gut microbiota composition also changed markedly, characterized by an increased relative abundance of Proteobacteria and decreased relative abundances of Firmicutes and Campylobacterota during the H and EA periods. Within AMPK–mTOR pathway-related genes, AMPKα1 and AMPKβ1 were downregulated, whereas mTOR signaling components ( MTOR , S6K1 , S6K2 , and 4EBP1 ) were upregulated. Correlation analysis further indicated that gut microbiota remodeling and AMPK–mTOR mediated energy regulation were associated with the maintenance of energy homeostasis and cellular stability during hibernation by influencing membrane lipid metabolism reprogramming. This study provides new insights into the coordinated metabolic adaptation mechanisms underlying turtle hibernation, enriches the physiological theory of reptilian hibernation, and offers a theoretical basis for artificial overwintering management and conservation research of threatened turtle species under changing environmental conditions.