A PCR assay To discriminate human and ruminant feces on the basis of host differences in Bacteroides-Prevotella genes encoding 16S rRNA

Abstract

Our purpose was to develop a rapid, inexpensive method of diagnosing the source of fecal pollution in water. In previous research, we identified Bacteroides-Prevotella ribosomal DNA (rDNA) PCR markers based on analysis. These markers length heterogeneity PCR and terminal restriction fragment length polymorphism distinguish cow from human feces. Here, we recovered 16S rDNA clones from natural waters that were close phylogenetic relatives of the markers. From the sequence data, we designed specific PCR primers that discriminate human and ruminant sources of fecal contamination.

FIG. 1

Phylogenetic relationships among partial 16S rDNA sequences (558 positions) of clones recovered from Tillamook Bay water samples (TB). HF and CF are host-specific genetic markers identified from human and cow fecal clone libraries, respectively. The tree was inferred by neighbor joining. Numbers above the internal branches are percentages of bootstrap replicates that support the branching order. Bootstrap values below 50% are not shown. Bootstrap values for branches a and b dropped from 68 to 47 and 76 to 40, respectively, when TB147 was added to the analysis. The sequence from Cytophaga fermentans was used to root the tree.