S1033 Objective 1. Develop or improve methods for control or elimination of pathogens in pre-and post harvest environments including meat, poultry, seafood, fruits and vegetables and nutmeats. <P>S1033 Objective 3. Investigate factors leading to the emergence, persistence and elimination of antimicrobial resistance in food processing and animal production environments
NON-TECHNICAL SUMMARY: Foodborne bacterial pathogens have been associated with food product recalls, illnesses and deaths in the United States. Foodborne bacterial pathogens undergo various stresses and possess the ability to adapt to various food processing conditions including cold, heat, acid, alkali or to sub-lethal doses of antimicrobials that they encounter during processing and in finished food products. Due to short generation times, foodborne pathogenic bacteria are very adaptable to surrounding environments and resistant surviving cells share genetic information between offspring to reestablish. Resistant forms of bacterial strains have appeared in all major target foodborne bacterial pathogens that affect raw and ready-to-eat food products of both plant and animal origin. Even though some resistant microflora are not human pathogens but they can still transfer genetic information to the target pathogenic foodborne bacteria in which they coexist. For example, human non-pathogenic fluoroquinolone resistant Campylobacter are commonly found in retail raw chicken carcass rinses along with human pathogenic species C. jejuni for which current intervention strategies are inadequate. Another pathogen, L. monocytogenes cells can survive in the absence of nutrients for very long periods and can infect when favorable conditions reappear. There is a critical need for pinpointing the survival patterns and global changes that take place in stress-adapted or antimicrobial/antibiotic-resistant forms of pathogen cells under various harsh conditions. Food industries spend as much as 300 million yearly on antimicrobials and there is a demand for diverse classes of antimicrobials that can limit or destroy the growth of pathogen cells without inducing resistance forms. To achieve high target efficacy for antimicrobials, there are critical gaps in our understanding of how various food components, packing conditions and target organisms interact. The full spectrums of stress- induced proteins that govern the pathogen cell adaptation or destruction have not been elucidated for major target foodborne bacterial pathogens in various food products. This work will lead to improved strategies for food safety by addressing the emerging threats of stress-adapted and antimicrobial-resistant or antibiotic-resistant forms of target foodborne pathogens with primary emphasis on L. monocytogenes and Campylobacter besides Vibrio spp., E. coli O157:H7 and Salmonella in target food products of animal and plant origin. <P>
APPROACH: The bactericidal effectiveness of lauric arginate (LAE) on the surfaces of meats and fresh fruits and vegetables will be determined against inactivation, survival and growth of L. monocytogenes, E. coli O157:H7 and Salmonella. The rate of death of E. coli O157:H7 and Salmonella on lettuce, spinach, tomato, and hot pepper will be determined 4C, 7C and 20C for up to 7 to 21 days after LAE treatment. Five strain mixtures of E. coli O157:H7 or Salmonella will be applied on the intact skin surfaces of above products by spot-inoculation method. These spots will be air dried for 2 h at room temperature in a BSL-2 laminar flow hood followed by 24 h incubation at 4C for bacterial attachment. Lauric arginate at FDA-approved concentration of 200 ppm (calculated based on pre-weighed samples) will be sprayed on the product surfaces as a fine-mist spray. Populations of pathogen cells surviving or growing on the products will be recovered by vigorous vortexing of samples or by stomaching for 2 min. Sub-sample surface rinses will be plated on CT-SMAC for E. coli O157:H7 or XLD agar for enumeration of Salmonella CFU counts. New methods for isolation, detection and enumeration of non-stressed, stressed, antimicrobial/antibiotic-resistant foodborne bacterial pathogen cells occurring in food products will be developed using chromogenic-agar based direct plating methods and by rapid antibody-based or PCR assays. Diversity of naturally occurring antibiotic resistant strains of Campylobacter and other foodborne pathogens will be isolated and confirmed by species-specific set of primers against target antibiotic resistance genes. Non-adapted and adapted Listeria monocytogenes cells to cold, acid, alkali, osmotic, and to sublethal concentrations of different antimicrobials will be detected by chromogenic agar assays with b-glycine and salt or with other compounds that enhance the recovery of stress adapted cells. The effect of pH, storage temperature, antimicrobial concentration, packaging environment and shelf life on the growth, survival and destruction of non-stressed, stressed and antimicrobial/antibiotic-resistant foodborne target bacterial pathogen cells will be determined in media and food matrices. Interactions of organic acids, salts, and other lethal factors on stress-adaption of pathogens will be determined. Novel antimicrobials (lauric arginate, Listex P100 bacteriophage and other) will be evaluated as a bactericidal surface treatment in various food products for achieving target quantitative reductions against pathogen cells. Monoclonal antibodies, proteomic and genomic approaches will be used to identify the regulatory networks of stress-adapted or resistant foodborne bacterial pathogen cells in food products. Difference in protein expression between cold-adapted and non-cold adapted L. monocytogenes will be identified by proteomic approaches by employing a set of mutant strains (cspA, cspAB, cspAD, and cspABD). Cells grown in rich media versus those grown in food products will be compared for stress protein expressions. The target foodborne bacterial pathogens include L. monocytogenes and C. jejuni besides E. coli O157:H7, Salmonella and Vibrio spp.