Effect of Bacteriophages on the Growth of Flavobacterium psychrophilum and Development of Phage-Resistant Strains.

Abstract

The controlling effect of single and multiple phages on the density of Flavobacterium psychrophilum at different initial multiplicity of infection (MOI) was assessed in batch cultures to explore the potential for phage-based treatment of this important fish pathogen. A high initial phage concentration (MOI = 0.3-4) was crucial for efficient viral lysis, resulting in a 10(4)-10(5)-fold reduction of phage-sensitive cells (both single phages and phage cocktails), which was maintained throughout the incubation (>10 days). Following cell lysis, regrowth of phage-resistant strains was examined and resistant strains were isolated for further characterization. The application of a mathematical model allowed simulation of phage-host interactions and resistance development, confirming indications from strain isolations that phage-sensitive strains dominated the regrowing population (>99.8%) at low MOI and phage-resistant strains (>87.8%) dominated at high MOI. A cross-infectivity test covering 68 isolated strains and 22 phages resulted in 23 different host susceptibility patterns, with 20 of the isolates being resistant to all the applied phages. Eleven isolated strains with different susceptibility patterns had lower growth rates (0.093 to 0.31 h(-1)) than the host strain (0.33 h(-1)), while 10 of 14 examined strains had lost the ability to take up specific substrates as shown by BIOLOG profiles. Despite increased selection for phage resistance at high MOI, the results emphasize that high initial MOI is essential for fast and effective control of F. psychrophilum infection and suggest that the small populations of resistant clones had reduced competitive abilities relative to the sensitive ancestral strain.