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Applied and Environmental Microbiology, June 2009, p. 3379-3388, Vol. 75, No. 11
0099-2240/09/$08.00+0     doi:10.1128/AEM.02302-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Quantification of Human Polyomaviruses JC Virus and BK Virus by TaqMan Quantitative PCR and Comparison to Other Water Quality Indicators in Water and Fecal Samples{triangledown}

Shannon M. McQuaig,1 Troy M. Scott,2 Jerzy O. Lukasik,2 John H. Paul,3 and Valerie J. Harwood1*

Department of Biology, University of South Florida, Tampa, Florida 33620,1 Biological Consulting Services of North Florida, Gainesville, Florida 32609,2 College of Marine Science, University of South Florida, St. Petersburg, Florida 337013

Received 7 October 2008/ Accepted 24 March 2009

In the United States, total maximum daily load standards for bodies of water that do not meet bacterial water quality standards are set by each state. The presence of human polyomaviruses (HPyVs) can be used as an indicator of human-associated sewage pollution in these waters. We have developed and optimized a TaqMan quantitative PCR (QPCR) assay based on the conserved T antigen to both quantify and simultaneously detect two HPyVs; JC virus and BK virus. The QPCR assay was able to consistently quantify ≥10 gene copies per reaction and is linear over 5 orders of magnitude. HPyVs were consistently detected in human waste samples (57 of 64) and environmental waters with known human fecal contamination (5 of 5) and were not amplified in DNA extracted from 127 animal waste samples from 14 species. HPyV concentrations in sewage decreased 81.2 and 84.2% over 28 days incubation at 25 and 35°C, respectively. HPyVs results were compared to Escherichia coli, fecal coliform, and enterococci concentrations and the presence of three other human-associated microbes: Bacteroidetes, Methanobrevibacter smithii, and adenovirus. HPyVs were the most frequently detected of these in human and contaminated environmental samples and were more human specific than the Bacteroidetes (HF183) or M. smithii. HPyVs and M. smithii more closely mimicked the persistence of adenovirus in sewage than the other microbes. The use of this rapid and quantitative assay in water quality research could help regulatory agencies to identify sources of water pollution for improved remediation of contaminated waters and ultimately protect humans from exposure to pathogens.

* Corresponding author. Mailing address: Department of Biology, SCA 110, University of South Florida, 4202 E. Fowler Ave., Tampa, FL 33620. Phone: (813) 974-1524. Fax: (813) 974-3263. E-mail: vharwood{at}cas.usf.edu

{triangledown} Published ahead of print on 3 April 2009.

Applied and Environmental Microbiology, June 2009, p. 3379-3388, Vol. 75, No. 11
0099-2240/09/$08.00+0     doi:10.1128/AEM.02302-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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