APPLICATION OF THE POLYMERASE CHAIN-REACTION TECHNIQUE TO THE DETECTION OF PATHOGENS IN WATER

Enteric pathogens may be present in fecally contaminated waters at ext remely low concentrations. In addition, these pathogens may be injured when exposed to the environment and may not be able to grow in labora tory culture media or such media may simply not exist for their progag ation in the laboratory. It is paramount thus to use techniques which do not depend on culture techniques for the detection of these pathoge ns and that allow for the detection of single-cell concentrations. The polymerase chain reaction (PCR) technique has been shown to be an exc ellent and sensitive means of detecting pathogens in waters. Membrane filtration has been combined with PCR and DNA hybridization techniques to be able to detect the DNA equivalent of one single cell in large v olumes of water. In addition, this combination of methods allows for t he amplification of different target genes that may be present in the sample, since the membrane can be subjected to repeated amplification reactions under different conditions. A Most Probable Number PCR was d eveloped which allows for the quantification of gene copy number and t hus permits extrapolation to estimate the number of bacterial cells in the original sample.