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Applied and Environmental Microbiology, November 2008, p. 6839-6847, Vol. 74, No. 22
0099-2240/08/$08.00+0     doi:10.1128/AEM.00601-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Temporal Assessment of the Impact of Exposure to Cow Feces in Two Watersheds by Multiple Host-Specific PCR Assays {triangledown}

Yong-Jin Lee,1 Marirosa Molina,2* Jorge W. Santo Domingo,3 Jonathan D. Willis,2 Michael Cyterski,2 Dinku M. Endale,4 and Orin C. Shanks3

National Research Council Research Associateship Programs, U.S. Environmental Protection Agency, Athens, Georgia 30605,1 U.S. Environmental Protection Agency, Office of Research and Development, National Exposure Research Laboratory, Ecosystems Research Division, Ecosystems Assessment Branch, Athens, Georgia 30605,2 U.S. Environmental Protection Agency, Office of Research and Development, National Risk Management Research Laboratory, Cincinnati, Ohio 45268,3 U.S. Department of Agriculture Agricultural Research Service, Watkinsville, Georgia 306774

Received 12 March 2008/ Accepted 7 September 2008

Exposure to feces in two watersheds with different management histories was assessed by tracking cattle feces bacterial populations using multiple host-specific PCR assays. In addition, environmental factors affecting the occurrence of these markers were identified. Each assay was performed using DNA extracts from water and sediment samples collected from a watershed directly impacted by cattle fecal pollution (WS1) and from a watershed impacted only through runoff (WS2). In WS1, the ruminant-specific Bacteroidales 16S rRNA gene marker CF128F was detected in 65% of the water samples, while the non-16S rRNA gene markers Bac1, Bac2, and Bac5 were found in 32 to 37% of the water samples. In contrast, all source-specific markers were detected in less than 6% of the water samples from WS2. Binary logistic regressions (BLRs) revealed that the occurrence of Bac32F and CF128F was significantly correlated with season as a temporal factor and watershed as a site factor. BLRs also indicated that the dynamics of fecal-source-tracking markers correlated with the density of a traditional fecal indicator (P < 0.001). Overall, our results suggest that a combination of 16S rRNA gene and non-16S rRNA gene markers provides a higher level of confidence for tracking unknown sources of fecal pollution in environmental samples. This study also provided practical insights for implementation of microbial source-tracking practices to determine sources of fecal pollution and the influence of environmental variables on the occurrence of source-specific markers.

* Corresponding author. Mailing address: U.S. EPA, 960 College Station Road, Athens, GA 30605. Phone: (706) 355-8113. Fax: (706) 355-8104. E-mail: molina.marirosa{at}epa.gov

{triangledown} Published ahead of print on 19 September 2008.

Applied and Environmental Microbiology, November 2008, p. 6839-6847, Vol. 74, No. 22
0099-2240/08/$08.00+0     doi:10.1128/AEM.00601-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.





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