DETECTION AND TYPING OF FOOD-BORNE PATHOGENS

Objective 1: Develop rapid and effective means to separate and concentrate targeted pathogens from food matrices that can be coupled to very rapid detection methods such as real-time PCR. 1A. Develop filtration/centrifugation methods for separating and concentrating pathogenic Escherichia, Salmonella, Listeria, and Campylobacter spp. from a variety of food matrices. Optimize reagents, apparatus and conditions to achieve maximum speed and recovery with minimum detection limits. 1B. Develop DNA extraction methods providing rapid, efficient, unbiased recovery of inhibitor-free DNA from a variety of pathogens. Objective 2: Examine environmental factors and microbiological culture conditions affecting genotypes or phenotypes that are important for virulence, isolation, or detection of foodborne pathogens. 2A. Detection of foodborne threat agents (model system- pathogenic Yersinia spp.). 2B. Isolation and detection of foodborne pathogens maintaining mobile genetic elements. 2C. Enrichment of pathogens while maintaining mobile genetic elements. Objective 3: Develop protein- and nucleic acid-based methods for the multiplexed detection and characterization of food-borne pathogens. 3A. Protein-based microarray and other multiplexed methods for the analysis of foodborne pathogenic bacteria. 3B. Oligonucleotide-based microarray for multiple pathogen detection and characterization. 3C. Multiplex real-time PCR for multiple pathogen identification and quantification. Objective 4: Develop typing methods for pathogens of concern to associated food regulatory agencies. 4A. Develop Restriction Fragment Sequence Polymorphism method for typing. 4B. Fractionation of a naïve library of biorecognition elements for bacterial typing-An alternative to ¿molecular typing.¿