Bacteriophages drive strain diversification in a marine Flavobacterium: implications for phage resistance and physiological properties

Abstract

Genetic, structural and physiological differences between strains of the marine bacterium Cellulophaga baltica MM#3 (Flavobacteriaceae) developing in response to the activity of two virulent bacteriophages, ΦS_M and ΦS_T, was investigated during 3 weeks incubation in chemostat cultures. A distinct strain succession towards increased phage resistance and a diversification of the metabolic properties was observed. During the incubation the bacterial population diversified from a single strain, which was sensitive to 24 tested Cellulophaga phages, into a multistrain and multiresistant population, where the dominant strains had lost susceptibility to up to 22 of the tested phages. By the end of the experiment the cultures reached a quasi steady state dominated by ΦS_T-resistant and ΦS_M + ΦS_T-resistant strains coexisting with small populations of phage-sensitive strains sustaining both phages at densities of > 10⁶ plaque forming units (pfu) ml⁻¹. Loss of susceptibility to phage infection was associated with a reduction in the strains' ability to metabolize various carbon sources as demonstrated by BIOLOG assays. This suggested a cost of resistance in terms of reduced physiological capacity. However, there was no direct correlation between the degree of resistance and the loss of metabolic properties, suggesting either the occurrence of compensatory mutations in successful strains or that the cost of resistance in some strains was associated with properties not resolved by the BIOLOG assay. The study represents the first direct demonstration of phage-driven generation of functional diversity within a marine bacterial host population with significant implications for both phage susceptibility and physiological properties. We propose, therefore, that phage-mediated selection for resistant strains contributes significantly to the extensive microdiversity observed within specific bacterial species in marine environments.