Functional genomics reveals key gene families and molecular pathways for extreme cold survival in the khapra beetle

The khapra beetle (Trogoderma granarium Everts) is among the world’s most destructive invasive pests, threatening food security and international trade. Its larvae exhibit exceptional resilience to extreme cold, enabling expansion beyond typical habitats. However, the molecular mechanisms underlying its cold tolerance remain largely unexplored due to limited genomic resources. We generated the first chromosome-level genome assembly for T. granarium (327 Mb, contig N50 = 883 kb), identifying 19,112 protein-coding genes across 9 chromosomes (99.0% BUSCO completeness). Comparative genomics revealed significant expansions in gene families associated with stress response, detoxification, and metabolism, including P450s, HSPs, and the insulin/IGF/relaxin family. Transcriptomic analysis and RNA interference targeting INSR, PIK3CB, and ADCY9 demonstrated marked effects on mortality and development following cold exposure. Crucially, functional validation revealed distinct survival roles: while INSR and PIK3CB are essential for sustaining basal homeostasis under stress, ADCY9 specifically mediates the response to cold adaptation. Additionally, miRNAs such as tca-miR-305-3p suggest post-transcriptional regulation of cold-responsive genes within the longevity and insulin signaling pathways. This study elucidates the genetic basis of T. granarium’s resilience, uncovering an integrated survival strategy involving both robust metabolic maintenance and targeted adaptive responses. These findings provide high-quality genomic resources and identify actionable molecular targets for the development of quarantine measures and invasion prevention strategies.