The construction of a rapid and sensitive assay for Salmonella contamination of foods is proposed. Salmonella causes over 40,000 U.S. cases of food poisoning per year. The Bacterial Ice Nucleation Diagnostic (BIND-TM) assay system is based upon transduction of an ice nucleation gene by a bacteriophage specific to the target organism, followed by detection of the ice nuclei produced. During Phase I of the research several short, selectable transposons, based on Tn5 and optimized for mutagenesis of bacteriophages, were developed and characterized. These transposons were used to rapidly identify regions of phage DNA which can tolerate interruption, and are therefore suitable targets for introduction of an ice nucleation reporter gene. During Phase II, this and several other methods will be used to construct a set of phages which together identify all of the clinically important serogroups of Salmonella. The phage set used will contain at least one broad host-range phage and several more specific phage, and will be validated by assaying for primary Salmonella isolates in real food samples. Successful completion of Phase II will set the stage for rapid commercialization of the BIND-TM assay for Salmonella, and lay the foundation for development of BIND-TM assays for Listeria monocytogenes, coliform bacteria, and other food contaminants.
