Abstract
Mustelinae are among the most diverse and taxonomically complex subfamilies within the Mustelidae, yet their evolutionary history and genetic diversity remain largely unexplored at the whole-genome level. Here, we present the first comprehensive comparative and phylogenomic study of this lineage, integrating nuclear and mitochondrial genomes from 10 species across the Holarctic and Indomalayan realms. Our dataset includes two novel genome assemblies (Mustela strigidorsa, M. sibirica) and an improved genome for M. nivalis, enabling robust cross-species analyses of genome size, chromosomal evolution, genetic diversity, and demographic history. We uncover striking inter- and intraspecific variation in genome-wide heterozygosity and genome size, with evidence of marked homozygosity in some Asian lineages (M. eversmanii, M. sibirica, M. strigidorsa) and remarkable genetic diversity in widespread species such as M. nivalis and M. erminea. Phylogenomic results support the previously suggested split of M. richardsonii from M. erminea, but we found no evidence for speciation within M. nivalis. Ancestral reconstruction of chromosomal rearrangements revealed key chromosomal fissions that shaped the Mustelinae radiation, including early events predating the divergence of modern Mustela species. The results confirmed the suggested ancestral karyotypes of Mustela (2n = 44) and Mustelinae (2n = 42). Finally, demographic reconstructions exposed species-specific responses to Quaternary climatic cycles, ranging from long-term resilience in M. nivalis to repeated population bottlenecks in M. putorius and M. sibirica. Collectively, our findings establish a genomic foundation for future evolutionary and conservation genomic research on this emblematic Mustelidae lineage.