Fresh produce can serve as a vehicle for foodborne pathogens, posing serious public health risks. These pathogens, including Shiga toxin-producing Escherichia coli (STEC), Salmonella enterica, and Listeria monocytogenes, can contaminate agricultural environments through multiple pathways. The first part of the research includes the factors that affect the survival of STEC, S. enterica, and L. monocytogenes in soil extracts simulating flooded or stagnant water conditions. Chemical analysis revealed that high-nutrient extracts, characterized by elevated nitrogen, phosphorus, and carbon levels, promoted pathogen persistence, whereas native microbiomes played a critical role in reducing pathogen populations. Microbiome analysis indicated higher microbial diversity in low-nutrient extracts, suggesting complex interactions that influence pathogen survival. Additionally, a long short-term memory (LSTM) model accurately predicted pathogen survival based on soil parameters, demonstrating the potential for predictive tools in food safety risk assessments. The second phase of this research examined the impact of cold storage on the survival and transition to physiological state of STEC O157:H7 on romaine lettuce. Lettuce harvested at 9 °C and stored at 2 °C for five days exhibited increased dormancy, transitioning into persister and viable but non-culturable (VBNC) states. STEC cell populations including persister cells showed reduced virulence, acid tolerance, and chlorine tolerance over time, underscoring the need to consider storage-induced physiological changes in microbial risk assessments. Finally, we investigated how enrichment time, strain variability, and persister cell populations influenced STEC O157:H7 detection using a commercially available qPCR-based system. Detection rates improved significantly with 24-hour enrichment compared to 8-hour enrichment, and the Yuma, AZ 2018 outbreak strain demonstrated higher detection rates than other strains. Additionally, samples with persister cell populations exceeding 68% were more likely to be detected, highlighting the importance of optimizing enrichment protocols to enhance detection reliability. This study underscores the significance of soil ecology, post-harvest storage conditions, and detection methodologies in improving microbial risk assessments in the fresh produce supply chain. Fresh produce can sometimes be contaminated with pathogens. These pathogens can be deadly and can contaminate produce by various means. The first part of the research includes the factors that affect the survival of STEC, S. enterica, and L. monocytogenes in soil extracts simulating flooded or stagnant water conditions. Chemical analysis revealed higher total nitrogen, phosphorus, and carbon in high-nutrient extracts, which supported pathogen survival, whereas the presence of native microbiomes reduced pathogen levels. Microbiome analysis showed greater diversity in low-nutrient extracts, indicating distinct microbial interactions that influence pathogen persistence. A long short-term memory (LSTM) model effectively predicted pathogen survival based on soil parameters, underscoring the potential for predictive tools in food safety risk assessments. The second part of the research includes evaluation of the impact of cold storage on the survival and physiological state of Shiga toxin-producing E. coli (STEC) O157:H7 on romaine lettuce. Lettuce harvested at 9 °C followed by storage at 2 °C for five days exhibited increased transformation into dormant states, including persister and viable but non-culturable (VBNC) cells. These dormant cells showed reduced virulence, acid tolerance, and chlorine tolerance over time. These findings highlight the need to account for storage-induced physiological changes when assessing microbial risks in the food supply chain. Finally, we investigated how enrichment time, strain variability, and persister cell populations influence STEC O157:H7 detection using the commercially available qPCR-based detection system. Detection rates increased significantly with 24-hour enrichment compared to 8-hour enrichment and the Yuma, AZ 2018 outbreak strain exhibited higher detection rates than other strains. Samples with persister cell percentages exceeding 68% were more likely to be detected, emphasizing the importance of optimizing enrichment protocols to improve detection reliability. This study highlights the importance of soil ecology, storage conditions, and detection methods, for better risk assessment analysis.
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