Viral persistence in surface and drinking water: Suitability of PCR pre-treatment with intercalating dyes

- Enteric viruses are able to remain infectious for a long time in the environment.
- Free viral nucleic acids are still detectable after chlorine treatment or UV irradiation.
- Intercalating dye pre-treatment is suitable for viral analysis of drinking water.

After many outbreaks of enteric virus associated with consumption of drinking water, the study of enteric viruses in water has increased significantly in recent years. In order to better understand the dynamics of enteric viruses in environmental water and the associated viral risk, it is necessary to estimate viral persistence in different conditions. In this study, two representative models of human enteric viruses, adenovirus 41 (AdV 41) and coxsackievirus B2 (CV-B2), were used to evaluate the persistence of enteric viruses in environmental water. The persistence of infectious particles, encapsidated genomes and free nucleic acids of AdV 41 and CV-B2 was evaluated in drinking water and surface water at different temperatures (4 °C, 20 °C and 37 °C). The infectivity of AdV 41 and CV-B2 persisted for at least 25 days, whatever the water temperature, and for more than 70 days at 4 °C and 20 °C, in both drinking and surface water. Encapsidated genomes persisted beyond 70 days, whatever the water temperature. Free nucleic acids (i.e. without capsid) also were able to persist for at least 16 days in drinking and surface water. The usefulness of a detection method based on an intercalating dye pre-treatment, which specifically targets preserved particles, was investigated for the discrimination of free and encapsidated genomes and it was compared to virus infectivity. Further, the resistance of AdV 41 and CV-B2 against two major disinfection treatments applied in drinking water plants (UV and chlorination) was evaluated. Even after the application of UV rays and chlorine at high doses (400 mJ/cm2 and 10 mg.min/L, respectively), viral genomes were still detected with molecular biology methods. Although the intercalating dye pre-treatment had little use for the detection of the effects of UV treatment, it was useful in the case of treatment by chlorination and less than 1 log10 difference in the results was found as compared to the infectivity measurements. Finally, for the first time, the suitability of intercalating dye pre-treatment for the estimation of the quality of the water produced by treatment plants was demonstrated using samples from four drinking-water plants and two rivers. Although 55% (27/49) of drinking water samples were positive for enteric viruses using molecular detection, none of the samples were positive when the intercalating dye pre-treatment method was used. This could indicate that the viruses that were detected are not infectious.

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