This Article Full Text (PDF) Other Versions of this Article: AEM.02474-06v1 73/8/2416    most recent Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Shanks, O. C. Articles by Graham, J. E. Search for Related Content PubMed PubMed Citation Articles by Shanks, O. C. Articles by Graham, J. E. Agricola Articles by Shanks, O. C. Articles by Graham, J. E.

 Previous Article  |  Next Article 

Appl. Environ. Microbiol. doi:10.1128/AEM.02474-06
Copyright (c) 2006, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Identification of Bacterial DNA Markers for the Detection of Human Fecal Pollution in Water

U.S. Environmental Protection Agency, Office of Research and Development, National Risk Management Research Laboratory, 26 West Martin Luther King Drive, MS-387, Cincinnati, OH; Department of Microbiology and Immunology, and Department of Biology, University of Louisville, Louisville, KY

^* To whom correspondence should be addressed. Email: santodomingo.jorge{at}epa.gov.

	Abstract

We used genome fragment enrichment (GFE) and bioinformatics to identify several microbial DNA sequences with high potential for use as markers in PCR assays for detection of human fecal contamination in water. Following competitive solution phase hybridization of total DNA from human and pig fecal samples, 351 plasmid clones were sequenced, and determined to define 289 different genomic DNA regions. These putative human-specific fecal bacterial DNA sequences were then analyzed by dot blot hybridization, which confirmed that 98% were present in the source human fecal microbial community, and absent from the original pig fecal DNA extract. Comparative sequence analyses of these sequences suggested that a large number (43.5%) were predicted to encode bacterial secreted or surface-associated proteins. Deoxyoligonucleotide primers capable of annealing to a subset of 26 of the candidate sequences predicted to encode factors involved in interactions with host cells were then used in the polymerase chain reaction (PCR), and did not amplify markers in DNA from any additional pig fecal specimens. These 26 PCR assays exhibited a range of specificity in tests with 11 other animal sources, with more than half amplifying markers only in specimens from dogs or cats. Four assays were more specific, detecting markers only in human specimens, including those from 18 different human populations examined. We then demonstrated the potential utility of these assays by using them to detect human fecal contamination in several impacted watersheds.

This article has been cited by other articles:

• Lu, J., Santo Domingo, J. W., Lamendella, R., Edge, T., Hill, S. (2008). Phylogenetic Diversity and Molecular Detection of Bacteria in Gull Feces. Appl. Environ. Microbiol. 74: 3969-3976 [Abstract] [Full Text]  
• Shanks, O. C., Atikovic, E., Blackwood, A. D., Lu, J., Noble, R. T., Domingo, J. S., Seifring, S., Sivaganesan, M., Haugland, R. A. (2008). Quantitative PCR for Detection and Enumeration of Genetic Markers of Bovine Fecal Pollution. Appl. Environ. Microbiol. 74: 745-752 [Abstract] [Full Text]