Development of a rapid method for a human pollution source tracking marker using enterococcus surface protein (esp) in Enterococcus faecium

Enumeration of fecal indicator bacteria alone will not provide information on the sources of contamination in water, thus microbial source tracking is necessary to identify the source of pollution. Major limitations with microbial source tracking methods are time consumption and the cost of the technique. The objective of the first portion of the study was to develop a rapid method for source tracking human pollution using a host specific genetic marker "esp" gene. Methods (2 & 3) were compared to the conventional method (1) for the detection of the esp marker in E. faecium from raw sewage. The elimination of the DNA extraction process and the removal of the enrichment process from the conventional method were assessed. Ninety seven membranes from 10 different raw sewage samples were analyzed for comparing method 1 and method 2. The rapid method limit of sensitivity was 46±27 cfu/membrane compared to the 45±28 cfu/membrane of the conventional method. In method 3 (without enrichment) the sensitivity was 26±40 cfu/membrane (n=78). The second objective was to evaluate the marker in the sewage. The marker was present in all the raw sewage samples (n=23) and in 2/10 effluent samples. The marker was absent in all the non human samples (n=23) and was present in 2/25 environmental samples processed. In a blind study the marker correctly identified 6/7 USGS source samples from human and was absent in all the non human samples processed (n=7) from different geographical regions.