Phage Resistance in Vibrio harveyi: Evolutionary strategies and physiological trade-offs

Bacteriophages impose strong selective pressure on bacterial populations, shaping host evolution and driving the emergence of phage-resistant variants. While receptor loss is a well-characterized resistance mechanism, the broader physiological consequences and transcriptional reprogramming associated with resistance remain underexplored, particularly in marine pathogens. Here, we investigate the molecular basis of phage resistance in Vibrio harveyi , an opportunistic aquaculture pathogen, using a combination of time-resolved transcriptomics, whole-genome variant analysis, and phenotypic assays. Exposure of wild-type V. harveyi to the lytic phage Virtus revealed a triphasic transcriptional response: early activation of SOS and stress pathways, mid-phase biosynthetic and translational upregulation, and late-stage repression of metabolism and envelope remodeling consistent with virion maturation. In parallel, we characterized 11 naturally evolved phage-resistant mutants that arose spontaneously under phage selection, identifying three mechanistic categories based on their mutational profiles: PilT -associated mutants, MSHA pilus-associated mutants, and hypermutators with defects in DNA mismatch repair. PilT and MSHA mutants harbored single-point mutations that triggered targeted envelope remodeling and transcriptional shifts. Hypermutators displayed extensive genomic variation and global transcriptional insulation, including persistent SOS activation, suppression of secretion systems, and metabolic downregulation. Despite achieving resistance, several mutants exhibited fitness trade-offs, including impaired growth, attenuated virulence in fish larvae, and altered antibiotic susceptibility. Comparative transcriptomics revealed a shared resistance signature across distinct lineages, marked by consistent upregulation of arginine catabolism and repression of Type VI secretion system components. These findings suggest that structural mutations can act as molecular toggles, globally reprogramming the host’s transcriptional state to resist infection. Our results demonstrate that phage resistance in V. harveyi is a multilayered adaptation involving mutational switching, envelope restructuring, and transcriptional rewiring. This work provides a system-level framework for understanding natural phage resistance and has implications for the development of sustainable phage therapy strategies.