Collaborative evaluation of a method for the detection of Norwalk virus in shellfish tissues by PCR

Author:

Atmar R L1,Neill F H1,Woodley C M1,Manger R1,Fout G S1,Burkhardt W1,Leja L1,McGovern E R1,Le Guyader F1,Metcalf T G1,Estes M K1

Affiliation:

1. Department of Medicine, Baylor College of Medicine, Houston, Texas 77030, USA.

Abstract

A multicenter, collaborative trial was performed to evaluate the reliability and reproducibility of a previously described method for the detection of Norwalk virus in shellfish tissues with the PCR (R.L. Atmar, F. H. Neill, J. L. Romalde, F. Le Guyader, C. M. Woodley, T. G. Metcalf, and M. K. Estes, Appl. Environ. Microbiol. 61:3014-3018, 1995). Virus was added to the stomachs and hepatopancreatic tissues of oysters or hard-shell clams in the control laboratory, the samples were shipped to the participating laboratories, and viral nucleic acids were extracted and then detected by reverse transcription-PCR. The sensitivity and specificity of the assay were 85 and 91%, respectively, when results were determined by visual inspection of ethidium bromide-stained agarose gels; the test sensitivity and specificity improved to 87 and 100%, respectively, after confirmation by hybridization with a digoxigenin-labeled, virus-specific probe. We have demonstrated that this method can be implemented successfully by several laboratories to detect Norwalk virus in shellfish tissues.

Publisher

American Society for Microbiology

Subject

Ecology,Applied Microbiology and Biotechnology,Food Science,Biotechnology

Reference33 articles.

1. Detection and differentiation of antigenically distinct small roundstructured viruses (Norwalk-like viruses) by reverse transcription-PCR and Southern hybridization;Ando T.;J. Clin. Microbiol.,1995

2. Atmar R. L. and J. A. Englund. Laboratory methods for the diagnosis of viral diseases. In A. S. Evans and R. Kaslow (ed.) Viral infections of humans in press. Plenum Medical Book Company New York.

3. Detection of enteric viruses in oysters by using the polymerase chain reaction;Atmar R. L.;Appl. Environ. Microbiol.,1993

4. Detection of Norwalk virus and hepatitis A virus in shellfish tissues with the PCR;Atmar R. L.;Appl. Environ. Microbiol.,1995

5. Application of the polymerase chain reaction in environmental microbiology;Bej A. K.;PCR Methods Appl.,1992

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