Development of Genomics and Molecular Biology-Based Methods for Tracking and Controlling Foodborne Microoganisms in Foods and the Environment

NON-TECHNICAL SUMMARY: Food safety concerns touch everyone on a daily basis. Despite recent advances that protect the United State's food supply, reports of illness and death caused by bacterial contamination of foods remain common. According to the United States Centers for Disease Control, pathogens typically transmitted through food, including Escherichia coli O157:H7, Listeria, non-Typhi Salmonella, and Shigella, caused 2621, 896, 45322, and 16168 cases of disease in 2005, respectively. These numbers represent a small fraction of the actual cases of disease, as only 3% of all foodborne illnesses are likely reported to health agencies Many foodborne illnesses are caused by zoonotic (ie. have an animal reservoir) organisms that contaminate food when they are shed into the environment. For example, E. coli O157:H7 asymptomatically colonizes the gastrointestinal system of cattle, are shed into the environment in feces, and may subsequently contaminate animals or plants present on the farm that are intended for human consumption. Efforts to reduce the burden of foodborne illness can therefore target one of three areas: 1) Prevention of the colonization of animals with the pathogen (via vaccines, etc); 2) Development of vaccines and therapeutics for reducing disease in humans; 3) Reducing the burden of the organism in the environment; 4) Development of control measures for reducing the burden of organisms throughout the food chain. This project proposes research into the mechanisms that zoonotic foodborne pathogens use to survive in the environment, and the development of DNA-based methods for tracking the transmission of these organisms from farm-to-fork. This data will lead to science-based approaches of reducing the burden of these organisms on farms, and for identifying control points that are necessary to reduce the spread of these organisms into the food supply.

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APPROACH: 1) Various genetic methods, such as transposon mutagenesis, signature tagged mutagenesis, and microarray analysis, will be evaluated for their usefulness in identifying genes and biochemical pathways that may be important for the long-term survival of pathogens in laboratory models of soil and manure environments. Specific regulons, or large number of genes under the control of a single genetic element, will also be inactivated and mutants tested for survival in these laboratory models. Where applicable, mutants that show decreased survival will be further analyzed to identify the mechanism behind the reduced fitness. 2) Other laboratory model systems of environmental survival (such as survival on plant surfaces) will be developed in collaboration with Dr. Timothy McNellis (Department of Plant Pathology, Penn State), and the strains identified in objective 1 will be screened for defects in survival in these models. Genetic methods developed in objective 1 will also be used to identify additional genes and biochemical pathways important for fitness in these models of environmental survival. 3) DNA sequencing of genes conserved in various strains of different foodborne microorganisms will be used to identify small sequence changes, or Single Nucleotide Polymorphisms (SNPs) that may be useful for differentiating strains of the same bacterial species. Libraries of different strains will be screened for various SNPs, to determine which of these molecular markers may be useful in tracking the movement of microorganisms within the food system. Alternative methods of identifying microorganisms within food systems, such as quantitative PCR, will also be investigated.