Phage Biology

M. Dyen1, R. Raya1,2, P. Varey1, G. Dutta1, T. Edrington3, T. Callaway3, A. D. Brabban1, E. Kutter1; 1Evergreen State College, Olympia, WA, 2Cerela, Tucuman, ARGENTINA, 3USDA Agricultural Station, College Station, TX.

Enterohemorrhagic Escherichia coli serotype O157:H7 (EHEC) is one of today’s most troubling food-borne pathogens. More than 75% of U.S. EHEC outbreaks have been linked to products derived from ruminants, the natural reservoir of this pathogen. We have characterized a new virulent bacteriophage, CEV1, isolated from the feces of sheep resistant to gastrointestinal colonization by E. coli O157:H7 infusions that generally colonize sheep very effectively. Electron micrographs showed that phage CEV1 has a prolate icosahedral head, with the contractile tail and baseplate structures characteristic of the T-4 family. Restriction enzyme analysis suggested that its dsDNA contains modified nucleotides. The main capsid protein gene of CEV1 can be amplified with standard T4 family primers and displays 94% homology with the corresponding T4 head protein (gp23), confirming that phage CEV1 belongs to the T-even family. The host range of phage CEV1 was determined on about 100 E. coli strains. CEV1 infected 16/18 of the E. coli O157:H7 strains tested and all standard lab strains. In contrast, it only could infect 8/72 of the ECOR set of coli strains; thus, it is likely to leave the normal gut flora relatively undisturbed. Single-step growth curves were carried out on a nonpathogenic E. coli 0157:H7 variant (ATCC 12900) during aerobic growth and during anaerobic respiration or fermentation in media designed to reflect the gut or rumen environment. They showed that CEV1 efficiently infects strain 12900 under all conditions, with burst sizes of about 50 Pfu but varying latent periods and delays before lysis.

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