Discovery, Isolation and Characterization of Bacteriophages Specific for Edwardsiella ictaluri

Abstract

Two novel bacteriophages that infect Edwardsiella ictaluri, the causative agent of enteric septicemia of catfish (ESC), were isolated from aquaculture pond water and from a diseased channel catfish. Both phages (FeiDWF and FeiAU) produced clear plaques ranging from 0.5 to11 mm in diameter, and have an icosahedral head that is approximately 50 nm in diameter with a non-rigid tail structure. The phages are tentatively classified in the family Siphoviridae. Phages FeiDWF and FeiAU are double-stranded DNA viruses with approximate genome sizes of 39 and 45 kb, respectively, with many restriction fragments in common as revealed by multiple restriction digests. Phage infection is calcium-dependent, with an optimal concentration of 500 µM CaCl2 resulting in a increase in plaque forming unit per milliliters (PFU ml-1) of over three orders of magnitude compared to media without supplemental CaCl2. Phage titers were also highest under the ideal growth conditions for E. ictaluri, with an optimal temperature range of 25-30°C. The latent period for phages is approximately 40 min with an estimated burst size of 270 viral particles per bacterial cell. At a multiplicity of infection (MOI) of 0.1 (phage: host), an E. ictaluri culture was reduced in colony forming units per milliliters (CFU ml-1) by over six orders of magnitude within a 6 h incubation period. All E. ictaluri strains tested were susceptible to phage infection with variable plaquing efficiencies. These phages have not been observed to produce plaques on any other bacterial species. The specificity of these bacteriophages for E. ictaluri isolates makes them attractive candidates as diagnostic agents and for bacteriophage therapy of ESC.