Bacterial Pathogen Amplification & Real-Time Detection

DESCRIPTION (provided by applicant): As we all know, bio-terrorism in America is a reality. However in addition to the Category A agents like anthrax, Yersinia pestis and smallpox, which are difficult to safely grow and disseminate, exist the Category B agents that could be used to infect our food or water supply. These organisms include bacterial pathogens, protozoa, and viruses. In addition to these natural pathogenic organisms they could also be genetically engineered to increase their virulence or to resist standard antibiotic treatments. Therefore new methods for rapid sensitive food and waterborne pathogen detection are greatly needed, especially if they can also be used to identify drug sensitivity within these organisms. Bio-terrorism using a food pathogen is not just a hypothetical threat to America. A religious cult in Dalles Oregon sickened at least 751 people by contaminating food in grocery stores and restaurants in the fall of 1984. The group simply grew cultures of the food pathogen Salmonella typhimurium that they obtained from their local scientific supply house and sprinkled the cultures on produce in the grocery stores and the restaurant salad bars. If the group had used a more deadly pathogen like Salmonella typhi that causes typhoid fever many people would certainly have died. The overall goal of this program is to develop an integrated isothermal DNA amplification and a probe array detection slide capable of rapidly identifying a variety of food and waterborne pathogens. All of the NIAID Biodefense Category B food and waterborne bacterial pathogens E. coli, Vibrio cholera, Shigella dysentery, Salmonella species, Listeria monocytogenes, Camphylobacter jejuni, and Yersinia enterocolitica will be detected in this program. A single integrated slide capable of isothermal amplifying and detecting all of these organisms in real-time in a closed sealed device is proposed. The program can also distinguish live organisms from dead organisms killed by the food or water sanitation process. Also, the test can be used to identify the antibiotic sensitivity of the pathogen to identify genetically altered organisms.