A Strategy for Detection of Viruses in Groundwater by PCR

Abstract

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.

Chapter 7 Global Supply of Virus-Safe Drinking Water

“…Therefore, in addition to wastewater overflows treated effluents will have to be taken into account as a significant source of viral pollution for wastewater used in agriculture and for surface waters used for aquaculture and fishery, recreational purposes or as a source for drinking water production. Moreover, both raw and treated sewage may contaminate groundwater through leakage (Abbaszadegan et al, 1999;Borchardt, 2003;Borchardt et al, 2004).…”

Waterborne Transmission of Enteric Viruses and Their Impact on Public Health

“…The water contamination by these agents was for domestic wastewater vector, which then can contaminate ground water and surfaces. Human enteric viruses can contaminate humans through the ingestion of contaminated water or other types of contact, such as recreational activities in contaminated marine waters (Abbaszadegan et al, 1999).…”

Assessment of the stability of human viruses and coliphage in groundwater by PCR and infectivity methods

“…The Polymerase Chain Reaction (PCR) is a well established and rapid method, which can be used to identify the presence of a specific virus based on the detection of its genetic material. It is a commonly used and valuable tool for assessing the microbial quality of water resources (Abbaszadegan et al 1999;Powell et al 2003). Virus detection by PCR may provide a more sensitive method of detection than cell culture assay because of the amplification process used, with low copy numbers of nucleic acid able to be detected.…”

“…It has traditionally been assumed that the use of PCR as a water quality monitoring tool will overestimate the level of microbial hazard. PCR should result in more virus being detected than cell culture assays as viral nucleic acid can be present when there are no infectious viruses present (Abbaszadegan et al 1999). However, recent studies have reported correlations between detection of virus genomes and infectivity under some specific conditions.…”

“…spring or well) . In recent years in the USA, viruses have been detected by PCR in groundwater, including in drinking water production wells and in confined and unconfined aquifers (Abbaszadegan et al 1999;Borchardt et al 2003Borchardt et al , 2007. In the UK, norovirus and enterovirus were detected by PCR at depths of up to 47 m in unconfined Permo-Triassic sandstone aquifers (Powell et al 2003).…”