Rapid Concentration of Viruses from Water

In the U.S., several high profile outbreaks of waterborne illness during the past 15 years have highlighted the need for solutions to drinking water contamination. Several recent studies suggest that approximately 20% of the surface and ground source waters in the U.S. are contaminated with viruses. However, there is very little data on virus occurrence in drinking water, which is mostly due to the absence of efficient methodologies that enable concentration and detection of viruses from water samples. Most current methods are labor or time intensive, expensive, or impractical. Therefore, there is an acute need for the development of concentration and detection technologies that enable efficient assay of viruses in water samples and these methods should be designed to be conducted directly in the field. We have designed a concentration device, that can capture viruses from large amounts of water, and the device completely eliminates the need for a secondary concentration step, and also increases the speed of concentration. The objective of this project is the development of an integrated detection method that includes capture of viruses from large volumes of water followed by detection of viruses by multiples real-time, reverse-transcriptase polymerase chain reaction. This project directly targets the following 2008 SBIR Phase I Research Topic: D. Drinking Water and Water Monitoring; Improved detection and measurement techniques for microbial pathogens. During Phase 1, two Specific Aims will be completed including: (1) Optimization of the sampling and concentration device, and demonstration of the ability to simultaneously capture two viruses from moderate (40-60 liters) volumes of tap water, river water, and irrigation water within 1-3 hours; and (2) Demonstration that both viral RNA and DNA can be simultaneously extracted directly without elution, and detected and quantified by a multiples real-time RT-PCR assay. During the Phase II period we plan to develop a multiplex isothermal RT-PCR assay, so that both the viral capture and detection steps can be completed directly in the field. Also, we will conduct in-field sampling of ground and surface water sources, and demonstrate the ability of the integrated capture and detection method to assay for the presence of viruses in these waters. It is expected that water testing companies, research laboratories, municipalities, and government agencies will make use of the capture and concentration device, either in combination with their own downstream detection assays, or with the isothermal-based RT-PCR assay that will be created during Phase II.