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Biological Sciences


Tracking the source of fecal pollution in surface waters has traditionally focused on the origin of enteric indicators including coliforms, enterococci, or E. coli. Recently, questions of genetic variability and environmental persistence have encouraged researchers to investigate additional animal specific indicators of fecal pollution. To date only eubacteria have been utilized as markers of human and animal-specific pollution. Here we report domestic ruminant-specific markers of fecal pollution utilizing methanogens found in the rumen. PCR primers for the mcrA gene of Methanomicrobium mobile (MMmcrA) and the nifH gene of Methanobrevibacter ruminantium (Mrnif-1) were designed, tested, and used to detect ruminant-specific pollution in fecal and environmental samples. The MMmcrA amplicons were detected in expected fecal and environmental samples (71% cow, 92% sheep, 50% goat, 100% cow lagoon samples, and 100% creek contaminated with cow lagoon waste), and were observed in only 5% of human and deer fecal samples. Mrnif-1 amplification was seen in 80% cow, 100% sheep, and 54% goat fecal samples; only 2% individual human samples were positive. No PCR amplification was observed when the MMmcrA and Mrnif-1 primers were tested against 47 bacterial stock cultures and fecal samples from 134 non-ruminant animals. More importantly, no amplification was observed in sewer samples using either primer pair. Sensitivity assays using MMmcrA primers demonstrated a detection limit of 0.01ng total DNA in bovine feces, 10ng in fecally contaminated surface water, and 5ng in cow lagoon samples.