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Applied and Environmental Microbiology, February 1999, p. 444-449, Vol. 65, No. 2
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

A Strategy for Detection of Viruses in Groundwater by PCR

Morteza Abbaszadegan,1,* Peter Stewart,1 and Mark LeChevallier2

American Water Works Service Company, Inc., Belleville, Illinois 62220,1 and American Water Works Service Company, Inc., Voorhees, New Jersey 080432

Received 16 July 1998/Accepted 2 October 1998

We evaluated the use of the PCR for detection of enteric viruses in groundwater. To do this, we used an improved sample-processing technique and a large-volume amplification protocol. The objective of this study was to use advanced molecular techniques to develop a rapid and simple method which can be used by the water industry for detection of viral contamination in a variety of water samples. The strategy described here fulfills the water industry's need for a rapid, reliable, easily performed method for analyzing groundwater for virus contamination. Viruses were detected after concentration from at least 400 gallons (1,512 liters) of water by a filter adsorption and elution method, which resulted in a concentrate containing viruses. A total of 150 samples were analyzed by performing cell culture assays for enteroviruses and by performing reverse transcription PCR (RT-PCR) analyses for enteroviruses, hepatitis A virus, and rotavirus. Thirteen samples (8.7%) produced cellular cytopathic effects when the Buffalo green monkey cell line was used. When primers specific for enteroviruses were used in RT-PCR, 40 of 133 samples (30.1%) tested positive for the presence of enterovirus RNA. When hepatitis A virus-specific primers were used, 12 of 139 samples (8.6%) were considered positive for the presence of hepatitis A viral RNA. The RT-PCR analysis performed with rotavirus-specific primers identified 18 of 130 samples (13.8%) that were positive for rotavirus RNA sequences. Our sample-processing technique and large-volume PCR protocol (reaction volume, 300 µl) resulted in sufficient removal or dilution of inhibitors so that more than 95% of the samples could be assayed by PCR. Because of its sensitivity for detecting viral nucleic acid sequences, PCR analysis should produce more positive results than cell culture analysis. Since either cell culture analysis or PCR can reveal only a "snapshot" of the quality of the groundwater being sampled, PCR seems to be a desirable rapid initial screening tool.

* Corresponding author. Mailing address: American Water Works Service Company, Inc., Quality Control & Research Laboratory, 1115 South Illinois Street, Belleville, IL 62220. Phone: (618) 235-9771. Fax: (618) 235-6349. E-mail: mabbasza{at}bellevillelab.com.

Applied and Environmental Microbiology, February 1999, p. 444-449, Vol. 65, No. 2
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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