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<OL> <LI> To develop physical and biological strategies to ensure inactivation of E. coli O157:H7 on the surface of composting heaps of animal manure. <LI> To delineate the conditions under which internalization and surface contamination of leafy greens by E. coli O157:H7 would occur in field conditions. <LI> To assess the quantitative transfer of E. coli O157:H7 among leafy greens during processing operations of conveyance, dewatering, and mechanical shredding. <LI> To investigate the potential for using processing water as a contamination marker for leafy greens. <LI> To assess the efficacy of intervention strategies (sanitizing and physical treatments such as long wave X-rays) for enhancing the safety and shelf-life of fresh-cut leafy greens contaminated with E. coli O157:H7. <LI> To characterize survival and growth of E. coli O157:H7 in contaminated leafy greens during post-harvest storage and distribution conditions. <LI> To develop a mathematical risk model for E. coli O157:H7 contamination associated with the production and sale of leafy greens and identify candidate risk mitigation strategies. <LI> To disseminate research outcomes and potential management strategies through annual steering committee meetings and regional stakeholder meetings.
NON-TECHNICAL SUMMARY: Contamination of leafy greens by E. coli O157:H7 in the field and during processing and storage, efficacy of low-energy X-rays intervention treatments, and detection of contamination through monitoring of process water will be addressed. Risk assessment models associated with the production and sale of leafy greens will be developed to assist with the identification of risk mitigation strategies. <P> APPROACH: GFP-labeled strains of E. coli O157:H7 will be used throughout the project. Bacteriophages and competitive exclusion microorganisms will be applied to compost piles to serve as biological control agents for E. coli O157:H7. To assist in thermal inactivation of E. coli O157:H7 contaminating the surface of fresh compost, piles will be covered with mature compost, straw, or pine shavings and turf reinforcement mesh. In growth chamber studies, leafy green plants will be subjected to varying levels of soil fertility, heat stress, water stress, and biological control agents to determine the impact on internalization of E. coli O157:H7 into the plant through the roots. In separate studies, plant leaves will also be exposed to E. coli O157:H7 dispersed either in water or a manure exudate to determine effect on the pathogen's survival. In field studies, the ability of lettuce, spinach, and parsley to internalize avirulent strains of E. coli O157:H7 as contamination level and plant maturity varies will be assessed. Using a combination of internally contaminated, surface-inoculated, and uninoculated leafy greens, the degree of cross-contamination will be monitored for E. coli O157:H7 at three key processing points: 1) conveyance on a conveyor belt; 2) dewatering step; and 3) mechanical shredding. A protocol for detecting low numbers of E. coli O157:H7 in lettuce process waters using a tangential flow filtration system, immunomagnetic separation system, and real-time PCR will be developed and evaluated in fresh-cut processing facilities and growing fields. To determine the potential for low-energy X-ray treatments to inactivate E. coli O157:H7 in leafy greens, a quantitative relationship between radiation dose and inactivation will be developed. In addition, the dose distribution within a commercial package of leafy greens will be mapped and validations of overall lethality in packages of leafy greens subjected to low-energy X-ray treatments will be conducted. The behavior of E. coli O157:H7 inoculated onto the spinach leaf surface or internalized will be compared under different packaging conditions and storage. Data collected in this project as well as other studies focusing on contamination at farm, factory, and retail/home will be used to used to develop exposure assessment estimations. Using surrogate and E. coli O157 outbreak data, hazard characterizations will also be assessed and combined with the exposure assessment models for risk characterization of E. coli O157:H7 in leafy greens. From these models and data, critical control points and possible mitigation strategies will be identified using a Food Safety Objective (FSO) approach. Dissemination of the project's research outcomes and proposed management strategies will be through scientific symposia, extension agent workshops, and a grower meeting.