<Ol> <LI> Develop or improve methods for control or elimination of pathogens in pre-and post harvest environments. <LI> Develop rapid, molecular-based detection methods for pathogens in pre-and post-harvest environments.<LI>Investigate factors leading to the emergence, persistence and elimination of antimicrobial resistance in food processing and animal production environments. </ol> Outputs: <li> Validated decontamination methods that can be used by the fruit, vegetable, seafood, meat and poultry industry to enhance the safety of their finished product <LI>Outreach/extension education and training materials for regulatory personnel, producers, processors, consumers
NON-TECHNICAL SUMMARY: Despite the availability of a variety of antimicrobial decontamination methods, food contamination and foodborne diseases continue to occur. The need for effective antimicrobials as well as food safety education and training is ever present. Furthermore, widely used cultural-based pathogen detection methods are time consuming, and often lack sensitivity and selectivity. The use of molecular based methods can overcome many of the limitations of cultural-based methods, in particular, sensitivity, selectivity and speed. A limitation of PCR-based molecular detection methods is the inability to differential between dead or live cells. Our work with the use of ethidium monoazide coupled with PCR will allow for the detection of only live pathogens. The outcomes of this project would contribute to improving food safety by way of the use of highly effective novel antimicrobial compounds and the ability to detect live pathogens in a rapid, sensitive and selctive manner.
APPROACH: Novel antimicrobial compounds will be tested to determine their effectiveness against pathogens. Natural and synthetic compounds to be tested will include organic acids, bacteriocins, zinc and magnesium oxide nanoparticles. Pathogens, including Escherichia coli, Salmonella and Listeria monocytogenes will be the target organisms. The antimicrobial compounds will be tested in various food matrices. PCR- and novel biosensor-based techniques will be improved upon as techniques for rapid and simultaneous detection of foodborne pathogens, including E. coli O157:H7, Salmonella and L. monocytogenes. DNA and RNA sequences that would make suitable primers for simultaneous detection of target pathogens will be continually searched in the literature. Other biosensor-based rapid methods, such as the use of quantum dots and other nanomaterials, will also be explored to achieve the objectives. Artificially and naturally contaminated food samples will be tested to optimize PCR-based methods, specifically by removing food inhibitors that typically limit the sensitivity of the technique. Foodborne pathogens can be induced to the viable-but-nonculturable (VBNC) state when in the presence of commonly used antimicrobial preservatives or food processes. We will investigate the VBNC phenomenon in E. coli O157:H7, Salmonella and L. monocytogenes in meat products and meat processing equipment following routine cleaning and sanitizing protocols and subsequent storage in various packaging and refrigeration conditions. The meat products will be artificially inoculated with varying concentrations of the pathogens and exposed to different intervention steps, while equipment will be artificially contaminated with the pathogens and the product processed as routinely done, followed by typical cleaning and sanitizing steps. VBNC cells will be monitored by the use of ethidium monoazide (EMA), a stain that can selectively penetrate dead cells because of their compromised membrane/cell wall systems and bind to the intracellular DNA. We will use EMA coupled with PCR to monitor for the presence and recovery of only viable and VBNC cells. The data obtained from this project will be shared with stakeholders via publications, news outlets and classroom activities.