are a potential source of novel colistin-resistance genes in European coastal environments.

Colistin is a widespread last resort antibiotic for treatment of multidrug-resistant bacteria. The recent worldwide emergence of colistin resistance (Col-R) conferred by -1 in human pathogens has raised concern, but the putative sources and reservoirs of novel genes in the marine environment remain underexplored. We observed a high prevalence of Col-R, particularly in isolated from European coastal waters by using the same cohorts of oysters as bioaccumulators in three sites across Europe. The high sequence diversity found in the A gene family was geographically structured, particularly for three novel gene variants, which were restricted to the Mediterranean (France, Spain) and occurred as a A-A operon. The RstA/RstB two component system was shown to control both the A-A operon and the Col-R phenotype. The analysis of 29 427 genomes revealed that this mechanism of intrinsic resistance is prevalent and specific to the Harveyi clade, which includes the human pathogens and . The operon conferred colistin-resistance when transferred to sensitive non- strains. In general, A gene variants are widespread and evolved with the lineage. They occur in clade-specific genomic environments, suggesting that expression responds to distinct environmental signals across the phylogeny. However, we also identified mobile A paralogues that have been recently transferred between and within clades. This highlights as a potential source of Col-R mechanisms, emphasizing the need for enhanced surveillance to prevent colistin-resistant infections in coastal areas.