Multiplex real-time RT-qPCR for the detection of Norovirus in bottled and tap water using murine norovirus as a process control

Authors

  • C. Hennechart-Collette,

    1. Food and Water Virology Unit, ANSES, Maisons-Alfort Laboratory for Food Safety, Maisons-Alfort Cedex, France
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  • S. Martin-Latil,

    1. Food and Water Virology Unit, ANSES, Maisons-Alfort Laboratory for Food Safety, Maisons-Alfort Cedex, France
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  • L. Guillier,

    1. Modelling of bacterial behaviour Unit, ANSES, Maisons-Alfort Laboratory for Food Safety, Maisons-Alfort Cedex, France
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  • S. Perelle

    Corresponding author
    1. Food and Water Virology Unit, ANSES, Maisons-Alfort Laboratory for Food Safety, Maisons-Alfort Cedex, France
    • Correspondence

      Sylvie Perelle, Maisons-Alfort Laboratory for Food Safety, Modelling of bacterial behaviour Unit, ANSES, 23, Avenue du Général de Gaulle, 94706 Maisons-Alfort Cedex, France. E-mail: sylvie.perelle@anses.fr

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Abstract

Aims

To provide a rapid and sensitive method for detecting NoV GI and NoV GII in water and to evaluate the use of the murine norovirus (MNV-1) as a process control.

Methods and Results

The method is based on viral concentration by filtration on electropositive filters and direct lysis of adsorbed viruses from filters before RNA extraction and RT-qPCR amplification. An one-step multiplex RT-qPCR assay was developed for the simultaneous detection of NoV GI, NoV GII and MNV-1. Then, water samples were artificially contaminated to determine mean virus recoveries and method sensitivity. The method showed a higher sensitivity for detecting NoV GII (103 genome copies per 0·5 l) than for NoV GI (104 genome copies per 0·5 l) in the presence of MNV-1 regardless of the type of water. The data also showed that MNV-1 is a robust option as process control.

Conclusions

The method described provides a valuable tool for the monitoring of potential public health risks associated with NoV contamination in drinkable water.

Significance and Impact of Study

Given the increasing evidence for NoV involvement in food outbreaks, the one-step multiplex RT-qPCR assay we used in this study would be a very useful tool to investigate NoV contamination in other food products.

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