Blood DNA Methylation Responses to Seasonal Environmental Variation in the Giant Panda (Ailuropoda melanoleuca): A Whole-Genome Bisulfite Sequencing Analysis of Multiple Individuals Across Multiple Time Points

Seasonal environmental variation is a widespread stressor for wildlife and can induce plastic changes in DNA methylation. Blood DNA methylation is an integrative molecular indicator of overall physiological status, yet its seasonal dynamics in the giant panda remain unclear. This study aimed to profile genome-wide blood methylation patterns in giant pandas under natural field conditions and to reveal the epigenetic strategies underlying their seasonal adaptation. Twelve blood samples were collected from three wild-trained giant pandas in the Daxiangling Nature Reserve during winter (n = 6) and summer (n = 6) between 2023 and 2025 and were subjected to whole-genome bisulfite sequencing. The genome-wide mC/C ratio (4.18–4.98%) was dominated by the CG context. Winter–summer comparison identified 7113 differentially methylated regions and 3063 differentially methylated genes. These genes were enriched in transcriptional regulation, cell surface receptor signaling, miRNA metabolism, and MAPK, Ras, and T/B cell receptor signaling pathways. Winter-hypermethylated genes were enriched in both immune defense and energy-consuming processes, whereas winter-hypomethylated genes showed limited enrichment, indicating that hypermethylation-mediated suppression dominates seasonal adaptation. This study reveals, for the first time, a seasonal methylation fingerprint in giant panda blood characterized by a “summer immune enhancement, winter energy conservation” pattern. As sampling occurred in natural habitats, these differences reflect integrated epigenetic responses to temperature, photoperiod, food availability, and pathogen pressure rather than single-factor effects. These findings provide molecular evidence for seasonal adaptation in endangered mammals and candidate markers for captive panda management and wild-release assessment.