Lytic Escherichia phage OSYSP acts additively and synergistically with gaseous ozone against Escherichia coli O157:H7 on spinach leaves

Abstract

Bacteriophage and gaseous ozone are evolving as meritorious alternatives to conventional sanitizers in food postharvest applications. Here, we investigated the efficacy of sequential treatments of a lytic bacteriophage and gaseous ozone, during vacuum cooling of fresh produce, against Escherichia coli O157:H7. Spinach leaves were spot-inoculated with 10⁵-10⁷ CFU g^-1 E. coli O157:H7 B6-914 and treated with Escherichia phage OSYSP spray (10⁹ PFU g^-1), gaseous ozone, or their combination. Vacuum cooling, which preceded or followed phage application but ran concomitantly with ozone treatment, was performed in a custom-made vessel at the following process sequence: vacuum to 28.5 in. Hg, vessel pressurization to 10 psig with gas containing 1.5 g ozone/kg gas-mix, holding for 30 min, and vessel depressurization to ambient pressure. Bacteriophage or gaseous ozone inactivated E. coli O157:H7, applied at different initial populations on spinach leaves, by 1.7-2.0 or 1.8-3.5 log CFU g^-1, respectively. At the high inoculum levels tested (7.1 log CFU g^-1), sequential treatments of phage and ozone reduced E. coli O157:H7 population by 4.0 log CFU g^-1, but when treatment order was reversed (i.e., ozone followed by bacteriophage), the combination synergistically decreased pathogen's population on spinach leaves by 5.2 log CFU g^-1. Regardless the antibacterial application order, E. coli O157:H7 populations, applied initially at ~ 10⁵ CFU g^-1, were reduced below the enumeration method's detection level (i.e., < 10¹ CFU g^-1). The study proved that bacteriophage-ozone combination, applied in conjunction with vacuum cooling, is a potent pathogen intervention strategy in fresh produce post-harvest applications.