Molecular Adaptations and Sex-Biased Regulatory Pathways in Multi-Year Diapause of the Cabbage Beetle Colaphellus bowringi: A Duration-Resolved Transcriptomic Analysis

The cabbage beetle Colaphellus bowringi, a key pest of cruciferous crops, exhibits a multiyear dormancy strategy characterized by pronounced interindividual variation in diapause duration: individuals entering summer or winter diapause-though initiated synchronously within each seasonal cohort-may remain in diapause for periods ranging from several months to as long as 5 years. To elucidate the molecular underpinnings of this extended dormancy and its sex-specific regulatory patterns, we performed a duration-resolved, sex-stratified transcriptomic analysis. Adult males and females were sampled after 0.5, 1, and 2 years of diapause durations and subjected to high-throughput RNA sequencing. De novo assembly yielded 66,379 unigenes, of which 16,132 (24.3%) were removed as non-arthropod contamination, leaving 50,247 unigenes that were used for all downstream annotation and expression analyses. Comparative transcriptomic profiling revealed a conserved adaptive strategy during prolonged diapause: patterns consistent with coordinated suppression of energy-intensive metabolic processes coupled with increased transcript abundance of genes in cellular protection pathways-including antioxidant defense, xenobiotic detoxification, and protein homeostasis mechanisms. Sex-biased transcriptional responses were observed: females showed comparable or higher numbers of differentially expressed genes in matched duration comparisons, with sustained transcript abundance of genes associated with reproductive processes and somatic maintenance, whereas males displayed greater duration-dependent transcriptional remodeling, particularly in signaling cascades and physiological homeostasis networks. KEGG pathway enrichment analysis identified sustained activation of glutathione metabolism, autophagy, and lysosomal degradation pathways throughout diapause; notably, extended diapause (2 years) was specifically associated with significant enrichment of immune response and lipid metabolism pathways. Duration-dependent dynamics in gene expression underscore that diapause is not a static quiescent state but a highly regulated, multi-stage process involving duration-specific transcriptional reconfiguration. This study establishes the first duration-resolved, sex-resolved transcriptomic resource for C. bowringi and advances mechanistic understanding of long-term insect diapause-thereby informing the development of targeted, environmentally sustainable pest management strategies.