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ARS Home » Midwest Area » Madison, Wisconsin » U.S. Dairy Forage Research Center » Environmentally Integrated Dairy Management Research » Research » Publications at this Location » Publication #263553
Title: Method 1615 measurment of Enterovirus and Norovirus occurrence in water by culture and qRT-PCR

Author
item FOUT, G - Environmental Protection Agency (EPA)
item BRINKMAN, N - Environmental Protection Agency (EPA)
item CASHDOLLAR, J - Environmental Protection Agency (EPA)
item GRIFFIN, S - Environmental Protection Agency (EPA)
item MCMINN, B - Environmental Protection Agency (EPA)
item RHODES, E - Environmental Protection Agency (EPA)
item VARUGHESE, E - Environmental Protection Agency (EPA)
item GRIMM, A - Environmental Protection Agency (EPA)
item Spencer, Susan
item Borchardt, Mark

Submitted to: Government Publication/Report
Publication Type: Government Publication
Publication Acceptance Date: 6/14/2011
Publication Date: 12/1/2011
Citation: Fout, G.S., Brinkman, N.E., Cashdollar, J.L., Griffin, S.M., Mcminn, B.R., Rhodes, E.R., Varughese, E.A., Grimm, A.G., Spencer, S.K., Borchardt, M.A. 2011. Method 1615 measurment of Enterovirus and Norovirus occurrence in water by culture and qRT-PCR. Government Publication/Report. EPA/600/R-10/181.

Interpretive Summary:

Technical Abstract: Viruses that may be present in environmental or finished drinking waters are concentrated from passage through electropositive filters. Viruses are eluted from the filters with a beef extract reagent and concentrated using organic flocculation. A portion of the concentrated eluate is then inoculated onto replicate flasks of BGM cells to measure infectious viruses. Cultures are examined for the development of cytopathic effects for two weeks and then re-passaged onto fresh cultures for confirmation. Virus concentration in each sample is calculated in terms of the most probable number (MPN) of infectious units per liter using EPA’s MPN calculator. For molecular assays, the concentrated eluate is concentrated again by centrifugal ultrafiltration. The RNA is then extracted from the concentrate and tested for enterovirus and norovirus RNA using real time quantitative reverse transcription-polymerase chain reaction (qRT-PCR). Virus concentrations for the molecular assay are calculated in terms of genomic copies of viral RNA per liter based upon a standard curve.