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Comparison of nucleic acid extraction and reverse transcription-qPCR approaches for detection of GI and GII noroviruses in drinking water
Griffin, S., N. Brinkman, E. Hedrick, E. Rhodes, AND Shay Fout. Comparison of nucleic acid extraction and reverse transcription-qPCR approaches for detection of GI and GII noroviruses in drinking water. JOURNAL OF VIROLOGICAL METHODS. Elsevier Science Ltd, New York, NY, 199:76-85, (2014).
Noroviruses (NoVs) are responsible for a number of waterborne and foodborne gastroenteritis cases each year. They are frequently associated with human sewage, and thus a potential link between wastewater discharge and contamination of source waters exists. Subsequently, contamination of drinking water is a concern for public health and necessitates rapid and reliable methods for detection of NoVs in various water matrices. Although assays using reverse transcription-quantitative polymerase chain reaction (RT-qPCR) have been developed for NoV detection, acquisition and operation costs can be prohibitive for water and environmental laboratories if there are not easily implemented molecular assays available. This study aimed to compare three RT-qPCR assays for NoV detection in drinking water with respect to performance, costs, and analysis time. The three molecular detection approaches evaluated were: A) a newly developed and previously unpublished approach; B) an RT-qPCR procedure which includes components of a proposed European Union standard method for NoV detection; and C) a commercialized RT-qPCR assay. Each approach was evaluated by most probable number (MPN) analysis for detection of GI.1 and GII.4 NoVs from human stool. Furthermore, recoveries of spiked primary effluent in tap water concentrates were compared for each approach. Few significant differences were observed between approaches with regard to performance. However, Approach C was the most time consuming and expensive in terms of both start-up and ongoing costs. This research presents a case study of how RT-qPCR assays can be compared and how various factors may play a role in which approach laboratories choose to employ.
To submit a manuscript entitled, "Comparison of reverse transcription-qPCR approaches for detection of GI and GII noroviruses in drinking water" for publication in the Journal of Food and Environmental Virology
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Organization:U.S. ENVIRONMENTAL PROTECTION AGENCY
OFFICE OF RESEARCH AND DEVELOPMENT
NATIONAL EXPOSURE RESEARCH LAB
MICROBIOLOGICAL AND CHEMICAL EXPOSURE ASSESSMENT DIVISION
BIOHAZARD ASSESSMENT RESEARCH BRANCH