The etiologic agents of many infectious diseases are present in complex biological materials, such as blood, feces, sputum, urine, and the like. The development of an effective therapeutic regimen for each disease requires the rapid and accurate detection and identification of the associated pathogen(s). The objectives of this project are 1) to develop a method for the rapid extraction and stabilization of microbial pathogen nucleic acids (total DNA and RNA) from feces, and 2) to detect the diarrheal pathogens Campylobacter jejuni and C. coli using specific and sensitive synthetic DNA probes. A procedure using chemical detergents will be developed for the rapid extraction and stabilization of the total nucleic acids. The two Campylobacter spp. will be used in a model system to demonstrate that total nucleic acids are extracted by this procedure. The efficiency of extraction and the integrity of the total nucleic acids will be evaluated. Specific pathogen nucleic acid sequences will be detected, and the level of sensitivity will be determined using artificial samples, as well as using patient samples which have been shown to be positive for the two pathogens by traditional culture techniques. The long-term objective of this project is to develop a rapid method for nucleic acid extraction from feces or other complex biological materials for use with specific DNA probes in commercial diagnostic products.
