Responses of host energy status, intestinal structure and gut microbiota during post-hibernation recovery in high- and low-altitude populations of the plateau frog Rana kukunoris.

BACKGROUND: Hibernation is an important seasonal strategy that enables amphibians to cope with low temperature and food scarcity. However, how high- and low-altitude amphibian populations differ in host energy status, digestive system structure and gut microbiota during post-hibernation recovery remains insufficiently understood. OBJECTIVE: This study aimed to evaluate post-hibernation changes in host energy status, intestinal structure and gut microbial composition and functional potential in high- and low-altitude populations of the plateau frog Rana kukunoris. METHODS: We compared high-altitude and low-altitude populations of R. kukunoris before and after hibernation by integrating morphological traits, whole-animal metabolic rate, digestive tract length, small-intestinal histology and shotgun metagenomic profiles of small-intestinal contents. RESULTS: After hibernation, both populations showed significant decreases in body mass, liver mass and hepatosomatic index, together with increased whole-animal metabolic rate, indicating a transition from energy reserve depletion to metabolic recovery. The hepatosomatic index showed a significant altitude × stage interaction, suggesting stronger relative liver energy depletion in the low-altitude population. Digestive system analysis showed that the low-altitude population exhibited more pronounced structural remodeling, including shortened digestive tract length, increased muscularis thickness and reduced epithelial thickness after hibernation, whereas the high-altitude population showed a relatively conservative response. Metagenomic analysis showed that alpha diversity remained relatively stable, whereas beta diversity, dominant microbial taxa and functional potential shifted among groups. Microbial functional profiles were mainly associated with metabolism, nutrient transformation and carbohydrate utilization. CONCLUSION: Post-hibernation recovery in R. kukunoris involves cross-level parallel responses in host energy status, digestive system structure and gut microbiota. High- and low-altitude populations may adopt different physiological recovery strategies after hibernation, providing new evidence for understanding seasonal adaptation in amphibians inhabiting cold environments.