Evaluation of a novel cocktail of six lytic bacteriophages against Shiga toxin-producing Escherichia coli in broth, milk and meat

Abstract

Phages are potentially useful as antimicrobial agents in food, especially cocktails of different phages which may prevent the development of bacterial resistance. Biocontrol assays with a six-phage cocktail, which is lytic against DH5α, an enteropathogenic (EPEC) and two Shiga-toxigenic (STEC) Escherichia coli strains, were performed in Hershey-Mg broth, milk and meat at refrigerated (4 °C), room (24 °C) and abusive (37 °C) temperatures. At 4 °C, cell counts were significantly lower (2.2-2.8 log₁₀ CFU/mL) when E. coli strains (∼10⁹ CFU/mL) were challenged against the phage cocktail (∼10⁹ PFU/mL) in Hershey-Mg broth after 24 h. However, reductions were higher (3.2-3.4 log₁₀ CFU/mL) after a 48 h exposure for all the strains tested. In addition, reduction values reached up to 3.4 log₁₀ CFU/mL (24 °C) and 3.6 log₁₀ CFU/mL (37 °C) in challenge tests after 24 h, though the reductions achieved were slightly lower after 48 h for the four E. coli strains tested. In milk, the cocktail was highly effective since bacterial counts were below the detection limit (<10¹ CFU/mL) at 4 °C, while the reductions ranged from 2 to 4 log₁₀ CFU/mL at 24 °C after a 24 h exposure. At 37 °C, DH5α was eliminated within 2 h, and an average cell decrease of 4 log₁₀ CFU/mL was observed for the three pathogenic strains tested. When the assays were performed in meat, biocontrol values ranged from 0.5 to 1.0 log₁₀ CFU/mL after 48 h at 4 °C, while a higher cell inactivation was achieved at 24 °C (2.6-4.0 log₁₀ CFU/mL) and 37 °C (3.0-3.8 log₁₀ CFU/mL). Furthermore, higher inactivation values for O157:H7 STEC (1.55 ± 0.35 log₁₀ CFU/mL) at 4 °C were obtained in meat when incubation was extended up to 6 days. As a conclusion, our six-phage cocktail was highly effective at 24 °C and 37 °C, though less effective at 4 °C in both food matrices evaluated. Thus, it might be applied against pathogenic EPEC and STEC strains to prevent foodborne diseases especially when the cold chain is lost.

Keywords: Escherichia coli; Food safety; Phage cocktail.