Characterization of Bacteriophages Active Against Escherichia coli O157:H7

REBECCA OOT, RAUL RAYA, SARAH PERIGO, ANNA CASTANO, MICHAEL DYEN, RICHARD BAUMANN, TODD CALLAWAY, ELIZABETH KUTTER, ANDREW BRABBAN

Atlanta, Georgia, USA

Congreso; 2005 General Meeting American Society for Microbiology; 2005

American Society for Microbiology, USA

Escherichia coli O157:H7 is a gram negative, facultative anaerobe that lives commensally in the gut of ruminants. It is also a food-borne pathogen of great concern in the US. Virulent bacteriophages show great promise in eliminating this pathogen from one of the main reservoirs that lead to human exposure, the gastrointestinal tracts of livestock. Naturally occurring bacteriophages active against O157:H7 have been isolated from sheep fecal material, the most efficient of which, CEV1 (T4-like Myovirus), CEV2 and CEV5 (both T5-like Siphoviridae), were selected for characterization. These phages have been analyzed using PFGE, PCR, growth curve experiments, genome restriction enzyme digests, EOP (efficiency of plating), host range on both the E. coli collection of reference (ECOR) and a number of pathogenic strains, electron micrographs and DNA sequencing. All three infect a broad range of pathogenic E. coli O157:H7 strains. CEV1 has a distinct host range on ECOR from CEV2 and CEV5, emphasizing probable differences in receptors. Infection characteristics under varying conditions are being examined in both single phage and cocktail growth curve experiments using (nonpathogenic) O157:H7 strain NCTC 12900 (ATCC 700728). In cocktail infection experiments using all 3 phages, lysis is complete by 60 min, bacterial survivors fall 4-5 logs and the only survivors observed are mucoidmutants. Some mutants are still sensitive to one or more of the phages in the cocktail, while others are resistant to these and all other tested phages. These various kinds of resistant mutants are being used as selective strains to characterize new phages and also to analyze cocktail-infection phage progeny.