Application of bacteriophages EP75 and EP335 efficiently reduces viable cell counts of Escherichia coli O157 on beef and vegetables.

Shiga toxin producing Escherichia coli (STEC) are common etiological agents of food borne illnesses and outbreaks, most often caused by consuming contaminated beef products, followed by raw vegetables and dairy products. Patients infected with E. coli O157 are more likely hospitalized than patients infected with non-O157 STEC, making E. coli O157 an important target for microbiological interventions. We show that a cocktail of bacteriophages EP75 and EP335 effectively reduces E. coli O157 on beef, romaine lettuce, spinach, and zucchini. Treatment of contaminated beef with either 2 × 107 or 1 × 108 PFU/cm2 of bacteriophage cocktail EP75/EP335 resulted in reductions of 0.8-1.1 log10 CFU/cm2 and 0.9-1.3 log10 CFU/cm2, respectively (P < 0.0001). Similarly, bacteriophage treatments of contaminated romaine lettuce, zucchini, or spinach showed significant (P < 0.05) E. coli O157 reductions of 0.7-1.9 log10 CFU/cm2 (2 × 107 PFU/cm2), and 1.4-2.4 log10 CFU/cm2 (1 × 108 PFU/cm2). An E. coli O157 reduction of 0.9 log10 and 2.0 log10 was observed already 30 min after phage application of 1 × 108 PFU/cm2 on beef and romaine lettuce, respectively. These data show that bacteriophages EP75 and EP335 can be effectively used as a processing aid on beef and vegetables, and thereby can aid industry to reduce the risk of E. coli O157 food poisoning.

Structural and functional characterization of the receptor binding proteins of Escherichia coli O157 phages EP75 and EP335.

Bacteriophages (phages) are widely used as biocontrol agents in food and as antibacterial agents for treatment of food production plant surfaces. An important feature of such phages is broad infectivity towards a given pathogenic species. Phages attach to the surfaces of bacterial cells using receptor binding proteins (RBPs), namely tail fibers or tailspikes (TSPs). The binding range of RBPs is the primary determinant of phage host range and infectivity, and therefore dictates a phage's suitability as an antibacterial agent. Phages EP75 and EP335 broadly infect strains of E. coli serotype O157. To better understand host recognition by both phages, here we focused on characterizing the structures and functions of their RBPs. We identified two distinct tail fibers in the genome of the podovirus EP335: gp12 and gp13. Using fluorescence microscopy, we reveal how gp13 recognizes strains of E. coli serotypes O157 and O26. Phage EP75 belongs to the Kuttervirus genus within the Ackermannviridae family and features a four TSP complex (TSPs 1-4) that is universal among such phages. We demonstrate enzymatic activity of TSP1 (gp167) and TSP2 (gp168) toward the O18A and O157 O-antigens of E. coli, respectively, as well as TSP3 activity (gp169.1) against O4, O7, and O9 Salmonella O-antigens. TSPs of EP75 present high similarity to TSPs from E. coli phages CBA120 (TSP2) and HK620 (TSP1) and Salmonella myovirus Det7 (TSP3), which helps explain the cross-genus infectivity observed for EP75.

Complete Genome Sequences of Escherichia coli Phages vB_EcoM-EP75 and vB_EcoP-EP335.

Phages vB_EcoM-EP75 (EP75) and vB_EcoP-EP335 (EP335) specifically infect Shiga toxin (Stx)-producing Escherichia coli (STEC) O157 strains. EP75 has a genome size of 158,143 bp and belongs to the genus Vi1virus The genome size of EP335 is 76,622 bp, and it belongs to the genus Phieco32virus.