Lifestyle-dependent evolution and CtrA-mediated lifestyle transitions shape phage resistance in marine Roseobacter

Marine bacteria alternate between planktonic and surface-attached lifestyles, facing continuous phage predation. However, how these lifestyles shape resistance evolution remains poorly understood. Using a Roseobacter model strain, we demonstrate that surface-attached populations exhibit 26-fold higher survivability than planktonic counterparts during lytic phage infection. This advantage emerges through the evolution of heterogeneous subpopulations exhibiting diverse resistance phenotypes, a pattern absent in planktonic populations. Whole-genome sequencing of 139 heritable phage-resistant mutants revealed fundamentally divergent mutational patterns, with planktonic populations predominantly harboring tandem repeat mutations, whereas surface-attached populations favor non-coding mutations. Despite this, both lifestyles independently converged on mutations in the CtrA phosphorelay system, identifying CtrA as a previously unrecognized evolutionary target of phage-driven selection and triggering planktonic-to-surface-attached switch. Further analyses revealed systematic downregulation of motility genes and enhancement of biofilm formation, mechanistically linking phage resistance to lifestyle transitions. The identified CtrA mutations occur in regions highly conserved across ecologically important marine Alphaproteobacteria (Rhodobacterales) that are known to switch between planktonic and surface-attached states, suggesting lifestyle-dependent evolutionary trajectories may broadly shape phage resistance in marine ecosystems.