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

Prévost, Benoit; Goulet, Marie-Line; Lucas, Françoise; Joyeux, Marc; Moulin, Laurent; Wurtzer, Sébastien

doi:10.1016/j.watres.2015.12.049

Cited by 92 publications

(64 citation statements)

References 32 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…Members of the family of enteroviruses are known to be relatively stable in the water phase at low to medium temperatures as shown by Prevost, Goulet [32]. Coxsackievirus was used to study the persistence of enteric RNA viruses in the environment and was like HAdV 41 able to remain infectious after more than 70 days at 4°C and 20°.…”

Section: Discussionmentioning

confidence: 99%

“…The treatment of the samples has been performed according to our previous study [25] with adjustments based on the finding of Prevost, Goulet [32]. Stock solutions of EMA and PMA (both Biotium Inc, Hayward, CA) have been reconstituted with 20% dimethyl sulfoxide (DMSO; Sigma-Aldrich Co., St. Louis, MO) up to a concentration of 10 mM and stored at -20°C.…”

Section: Methodsmentioning

confidence: 99%

“…The application of the method for the distinction between infectious and non-infectious viruses has been investigated thoroughly in lab scale [25–29]. Its application also has been proposed in food safety [30, 31] and for the detection of enteric viruses in the environment [32–35]. …”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

From Lab to Lake – Evaluation of Current Molecular Methods for the Detection of Infectious Enteric Viruses in Complex Water Matrices in an Urban Area

et al. 2016

View full text Add to dashboard Cite

Quantitative PCR methods are commonly used to monitor enteric viruses in the aquatic environment because of their high sensitivity, short reaction times and relatively low operational cost. However, conclusions for public health drawn from results of such molecular techniques are limited due to their inability to determine viral infectivity. Ethidium monoazide (EMA) and propidium monoazide (PMA) are capable to penetrate the damaged or compromised capsid of the inactivated viruses and bind to the viral nucleic acids. We assessed whether dye treatment is a suitable approach to improve the ability of qPCR to distinguish between infectious and non-infectious human adenovirus, enterovirus and rotavirus A in surface water of an urban river and sewage before and after UV disinfection. Like the gold standard of cell culture assays, pretreatment EMA-/PMA-qPCR succeeded in removing false positive results which would lead to an overestimation of the viral load if only qPCR of the environmental samples was considered. A dye pretreatment could therefore provide a rapid and relatively inexpensive tool to improve the efficacy of molecular quantification methods in regards to viral infectivity.

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

From Lab to Lake – Evaluation of Current Molecular Methods for the Detection of Infectious Enteric Viruses in Complex Water Matrices in an Urban Area

et al. 2016

View full text Add to dashboard Cite

show abstract

“…Photoactivatable dyes such as propidium monoazide (PMA) and ethidium monoazide (EMA) (Biotium, Inc., Fremont, CA, USA) have been used with RT-qPCR with some success for the selective detection of infectious norovirus using RT-qPCR [8][9][10][11][12][13]. Theoretically, these dyes penetrate a damaged capsid and, in the presence of light, a covalent bond forms with the nucleic acid, resulting in a reduction in nucleic acid extraction efficiency and in the RT-qPCR signal [14].…”

Section: Introductionmentioning

confidence: 99%

Detection of Infectious Noroviruses from Wastewater and Seawater Using PEMAXTM Treatment Combined with RT-qPCR

Gyawali

Hewitt

2018

Water

View full text Add to dashboard Cite

Rapid detection of infectious noroviruses from environmental samples is essential to minimize the risk of norovirus outbreaks associated with environmental transmission. Reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) methods are rapid and sensitive, but cannot differentiate between infectious and non-infectious noroviruses. In this study, a PEMAX TM treatment followed by RT-qPCR (PEMAX TM -RT-qPCR) method was developed for murine norovirus and norovirus GI/GII, and evaluated for the selective detection of infectious viruses following heat inactivation. The norovirus PEMAX TM -RT-qPCR method was then evaluated for the selective detection of infectious viruses from environmental samples. Following heat-treatment (90 • C for 3 min), the murine norovirus PEMAX TM -RT-qPCR showed at least a 2.04 log 10 reduction in detectable virus, compared to a 0.43 log 10 reduction for RT-qPCR alone. Under the same conditions, the norovirus PEMAX TM -RT-qPCR showed a 0.34 to 0.98 log 10 (GI.3) and 0.63 to 2.06 log 10 (GII.4) reduction in detectable viruses, compared to 0.05 to 0.18 log 10 (GI.3) and 0.06 to 0.25 log 10 (GII.4) for RT-qPCR alone. Evaluation of the norovirus PEMAX TM -RT-qPCR on norovirus-contaminated influent and effluent wastewater, and seawater indicated a high proportion of non-infectious norovirus GI and GII (i.e., 56 to 100% in seawater, 32 to 76% in effluent, and 11 to 79% in influent) was present in samples. While potentially overestimating the amount of infectious noroviruses, this approach has potential to provide better information on viral infectivity than RT-qPCR alone.

show abstract

“…The latter two were at first neglected because since RNA was considered as not stable; the detected genomes were assumed to be linked to infectious viruses protected by a capsid. Today, such an assumption is no longer acceptable because it has been largely demonstrated that the persistence of the genome is much greater than that of the corresponding infectious virus in most cases (Gassilloud et al 2003;Ogorzaly et al 2010;Prevost et al 2016;Seitz et al 2011;Simonet and Gantzer 2006). Gaining knowledge about the characteristics of non-infectious viral particles, or more broadly about the molecular mechanisms of viral inactivation, has become a major objective for food and environmental virologists.…”

Section: Introductionmentioning

confidence: 99%

The Effect of Heat on the Physicochemical Properties of Bacteriophage MS2

et al. 2016

View full text Add to dashboard Cite

The differences in physicochemical characteristics between infectious and non-infectious viral particles are poorly known. Even for heat, which is known as one of the most efficient treatments to inactivate enteric viruses, the global inactivation mechanisms have not been described yet. Such knowledge would help distinguish between both types of particles and therefore clarify the interpretation of the presence of viral genomes in food after heat treatment. In this study, we examined in particular the differences in electrostatic charge and hydrophobicity between the two particle types. MS2 phage, a common surrogate for enteric viruses, was used as a model virus. The heat-induced inactivation process of the infectious phages caused hydrophobic domains to be transiently exposed and their charge to become less negative. The particles also became progressively permeable to small molecules such as SYPRO Orange dye. The presence of non-infectious phage particles in which the genome was not accessible to RNases has been clearly demonstrated. These observations were done for MS2 phages exposed to a temperature of 60 °C. When exposed to a temperature higher than their critical temperature (72 °C), the particles were disrupted and the genome became available for RNases. At lower temperatures, 60 °C in this study, the transient expression of hydrophobic domains of remaining infectious phages appeared as an interesting parameter for improving their specific detection.

show abstract

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

Cited by 92 publications

References 32 publications

From Lab to Lake – Evaluation of Current Molecular Methods for the Detection of Infectious Enteric Viruses in Complex Water Matrices in an Urban Area

From Lab to Lake – Evaluation of Current Molecular Methods for the Detection of Infectious Enteric Viruses in Complex Water Matrices in an Urban Area

Detection of Infectious Noroviruses from Wastewater and Seawater Using PEMAXTM Treatment Combined with RT-qPCR

The Effect of Heat on the Physicochemical Properties of Bacteriophage MS2

Contact Info

Product

Resources

About