Resistance Without Clones: Accessory Genome Dynamics Shape Antimicrobial Resistance in Corynebacterium striatum

Corynebacterium striatum has emerged as a multidrug-resistant opportunistic pathogen; however, the population-scale genomic architecture underlying resistance dissemination remains incompletely resolved. Existing studies are limited in sampling depth and thus have not comprehensively quantified the structural coupling between pangenome compartmentalization, resistance localization, and mobilome association. Here, we assembled and analyzed the largest C. striatum dataset to date, comprising 839 high-quality genomes. The pangenome consisted of 6,361 gene families and exhibited a weakly open architecture (γ = 0.102; R² = 0.989), with a conserved core backbone (31.3%) embedded within an expansive accessory reservoir. Gene frequency spectra displayed pronounced bimodality, and functional analyses revealed clear stratification: core genes were enriched for essential information-processing and metabolic functions, whereas accessory compartments - particularly the cloud - contained disproportionately high fractions of poorly characterized genes. A total of 35 antimicrobial resistance (AMR) gene families were identified and were strictly confined to shell and cloud compartments (χ² p = 2.9 × 10⁻⁸), with no resistance determinants detected in the core genome. AMR genes were strongly enriched on plasmid-associated contigs (odds ratio = 417.38, p < 1 × 10⁻¹⁶) and exhibited significant proximity to mobile genetic elements (odds ratio = 24.41, p < 1 × 10⁻¹⁶). Phylogenetic mapping demonstrated that resistance burden and multidrug resistance phenotypes are broadly distributed across diverse lineages rather than restricted to a dominant clonal expansion. Together, these findings define a constrained yet mobility-enriched genomic architecture that facilitates repeated, lineage-independent acquisition of antimicrobial resistance, highlighting the central role of accessory genome dynamics in genomic surveillance of emerging opportunistic pathogens.