Haplotype-resolved chromosome-level genome assemblies of four Diamesa species reveal the genetic basis of cold tolerance and high-altitude adaptations in arctic chironomids

Arctic and alpine insects face extreme environmental stressors, yet the genomic basis of their adaptation remains poorly understood. Here, we present the first haplotype-resolved, chromosome-level genomes for four species of Diamesa (Diptera: Chironomidae), a genus of cold-adapted midges inhabiting glacial and high-altitude freshwater ecosystems. Using PacBio HiFi sequencing and Hi-C scaffolding, we assembled high-quality genomes with chromosome-level resolution and high k-mer completeness. Phylogenomic analyses support Diamesinae as sister to other Chironomidae except Podonominae, and genomic comparisons provide evidence for introgression between the evolutionary distinct D. hyperborea and D. tonsa . Comparative genomic analyses across 20 Diptera species revealed significant gene family contractions in Diamesa associated with oxygen transport and metabolism, suggesting adaptations to high-altitude, low-oxygen environments. Conversely, expansions were detected in histone-related and Toll-like receptor gene families, likely enhancing chromatin remodeling and immune regulation under cold stress. A single gene family encoding glucose dehydrogenase was significantly expanded across all cold-adapted species studied, implicating its role in cryoprotectant synthesis and oxidative stress mitigation. Notably, Diamesa species exhibit the largest gene family contraction at any node, with minimal overlap in expansions with other cold-adapted Diptera, indicating lineage-specific adaptation. Our findings support the hypothesis that genome size condensation and selective gene family changes underpin survival in cold environments. These genome assemblies represent a valuable resource for investigating adaptation, speciation, and conservation in cold-specialist insects. Future work integrating gene expression and population genomics will further illuminate the evolutionary resilience of Diamesa in a warming world.