Multiplex Detection of Microbial Pathogens

DESCRIPTION (provided by applicant): The food and water supply in the United States is claimed to be one of the safest in the world; however, CDC estimates that >76 million people get sick, with ~3-5 billion worldwide cases of associated diarrhea. Our goal is to develop a rapid one-tube multi-organism test method to screen and identify >22 infectious organisms associated with food- and water-borne disease identified on the NIAID Priority Pathogen list. This method is based on the ability of padlock oligonucleotide probe sets to selectively identify three to four predefined target recognition sequences for each pathogenic genome of interest in parallel without loss of discrimination or sensitivity. Primer design problems that typically arise with multiplex PCR applications are eliminated with padlock probes. Phase 1 will determine the appropriate padlock probes to specifically identify the strains of interest; selectivity and sensitivity of the padlock reaction; demonstrate feasibility of the technology for universal detection and array identification with biological samples; and demonstrate practicality of magnetic based sample isolation and purification based technology. In Phase II, we will assemble the appropriate probe sets to identify >22 infectious organisms, optimize sample preparation, and perform a study to assess sensitivity and specificity on various food and liquid samples.