Applicability of Novel Bacteriophage Treatments to Reduce Shiga Toxin-producing Escherichia coli on Leafy Greens

Introduction: Shiga toxigenic Escherichia coli (STEC) is a major group of foodborne pathogens, accounting for multiple outbreaks and has also become a threat for the fresh-produce industry. Control of these pathogens remains a challenge even with the adaptation of modern practices and technologies. It is therefore important to develop effective and novel control strategies. Bacteriophage-based treatments could offer a natural, specific, and effective strategy to control STEC.

Purpose: Evaluate the effectiveness of STEC-specific bacteriophages on leafy greens.

Methods: Bacteriophages, isolated from cattle operations, showing lytic activity towards diverse STEC strains were used to prepare antibacterial treatments in phosphate buffered saline (PBS). Leafy greens tested were spinach and romaine lettuce. Prepared leafy green samples were transferred to petri-plates containing moistened filter paper and spot-inoculated with 5 log CFU/ml of either individual strains of E. coli O45, O103, O111, O121, O26, or O145 or cocktail of E. coli O157:H7. Leaves were spray-treated with phages (8 log PFU/ml) or PBS-control, using airbrush filled with treatment solution. Treated leaves were stored for 72 h at 4°C. Surviving bacteria were enumerated at 0 h, 24 h, and 72 h and data analyzed using one way ANOVA (P<0.05). 

Results: Leafy greens treated with phage cocktail showed an immediate reduction (1.4-3.0 logs) in E. coli O157:H7 populations, compared to the positive or PBS control (P<0.05) at 0 h. Similarly, leafy greens contaminated with non-O157 STECs and treated with individual phages, also showed an immediate reduction (1.3 to 3.0 logs) in pathogen populations, compared to the positive or PBS control (P<0.05) at 0 h. Phage-treated leafy greens continued to show significant reductions (1.4 to 3.0 logs; P<0.05) in both E. coli O157:H7 and non-O157 STEC populations at 24 and 72 h, compared to the positive or PBS control.   

Significance: Bacteriophages could potentially be used as antimicrobial treatments to reduce or inactivate STEC populations on leafy greens.