APOBEC3G-Induced Hypermutation of Human Immunodeficiency Virus Type-1 Is Typically a Discrete “All or Nothing” Phenomenon

The rapid evolution of Human Immunodeficiency Virus (HIV-1) allows studies of ongoing host–pathogen interactions. One key selective host factor is APOBEC3G (hA3G) that can cause extensive and inactivating Guanosine-to-Adenosine (G-to-A) mutation on HIV plus-strand DNA (termed hypermutation). HIV can inhibit this innate anti-viral defense through binding of the viral protein Vif to hA3G, but binding efficiency varies and hypermutation frequencies fluctuate in patients. A pivotal question is whether hA3G-induced G-to-A mutation is always lethal to the virus or if it may occur at sub-lethal frequencies that could increase viral diversification. We show in vitro that limiting-levels of hA3G-activity (i.e. when only a single hA3G-unit is likely to act on HIV) produce hypermutation frequencies similar to those in patients and demonstrate in silico that potentially non-lethal G-to-A mutation rates are ∼10-fold lower than the lowest observed hypermutation levels in vitro and in vivo. Our results suggest that even a single incorporated hA3G-unit is likely to cause extensive and inactivating levels of HIV hypermutation and that hypermutation therefore is typically a discrete “all or nothing” phenomenon. Thus, therapeutic measures that inhibit the interaction between Vif and hA3G will likely not increase virus diversification but expand the fraction of hypermutated proviruses within the infected host

Detection of enterovirus in environmental waters: A new optimized method compared to commercial real-time RT-qPCR kits

International audienceDespite the progress in water and wastewater treatment technologies, waterborne diseases are still amajor concern of public health. In the reported water-related outbreaks, viruses constitute one of themain causal agents. Enteroviruses are one of the most viruses monitored in water and are often usedas an indicator of viral pollution. Isolation and identification of this virus are now regularly based onmolecular tools. However published or commercial protocols for detection of these viruses in waterare frequently lacking of validation processes and performance evaluation in such complex samples. Amethod for enterovirus detection in environmental water has been developed, its performance has beenevaluated and compared with several commercial kits.The sensitivity of commercial methods in clinical samples, ranged between 89% and 100%, while thesensitivity in seeded environmental matrices fell between 16% and 91%. This method showed the bestperformance in environmental samples and was subsequently applied on surface and treated wastewa-ter. The results showed the large dissemination of enteroviruses in an urbanized river. The results alsoemphasized the importance of good knowledge of the method’s limits for its utilization in environmentalsamples in order to minimize false negatives and to avoid underestimating viral concentration

Hydrophobic Organic Matter Promotes Coxsackievirus B5 Stabilization and Protection from Heat

International audienceIn urban rivers, many of which are used for drinking water production, viruses encounter a range of particulate, colloidal, and dissolved organic and inorganic compounds. To date, the impact of environmental organic matter on virus persistence in the environment has received little attention. In the present study, fresh water was fractioned to separate particulate natural organic matter from dissolved forms. Each fraction was tested for its ability to promote coxsackievirus B5 resistance to heat inactivation. Our results demonstrate that, at natural concentrations, environmental waters contain particulate or dissolved compounds that are able to protect viruses from heat. We also show that hydrophobic compounds promote an efficient protection against heat inactivation. This study suggests that local conditions encountered by viruses in the environment could greatly impact their persistence

Interaction of Human Enteric Viruses with Microbial Compounds: Implication for Virus Persistence and Disinfection Treatments

International audienceAlthough the interaction between phages and bacteria has already been well described, it only recently emerged that human viruses also interact with bacteria in the mammalian gut. We studied whether this interaction could occur in tap water and thus confer enteric viruses protection against temperature and the classical disinfection treatments used in drinking water production. We demonstrated that the addition of lipopolysaccharide or peptidoglycan of bacterial origin to enterovirus provides thermal protection through stabilization of the viral capsid. This interaction plays a role when viruses are exposed to disinfection that targets the capsid, but less so when the virus genome is directly targeted. The interaction seems to be serotype-specific, suggesting that the capsid protein sequence could be important. The protection is linked to a direct association between viral particles and bacterial compounds as observed by microscopy. These results show that bacterial compounds present in the environment can affect virus inactivation

Deciphering the diversities of astroviruses and noroviruses in wastewater treatment plant effluents by a high-throughput sequencing method

International audienceAlthough clinical epidemiology lists human enteric viruses to be among the primary causes of acute gastroenteritis in the human population, their circulation in the environment remains poorly investigated. These viruses are excreted by the human population into sewers and may be released into rivers through the effluents of wastewater treatment plants (WWTPs). In order to evaluate the viral diversity and loads in WWTP effluents of the Paris, France, urban area, which includes about 9 million inhabitants (approximately 15% of the French population), the seasonal occurrence of astroviruses and noroviruses in 100 WWTP effluent samples was investigated over 1 year. The coupling of these measurements with a high-throughput sequencing approach allowed the specific estimation of the diversity of human astroviruses (human astrovirus genotype 1 [HAstV-1], HAstV-2, HAstV-5, and HAstV-6), 7 genotypes of noroviruses (NoVs) of genogroup I (NoV GI.1 to NoV GI.6 and NoV GI.8), and 16 genotypes of NoVs of genogroup II (NoV GII.1 to NoV GII.7, NoV GII.9, NoV GII.12 to NoV GII.17, NoV GII.20, and NoV GII.21) in effluent samples. Comparison of the viral diversity in WWTP effluents to the viral diversity found by analysis of clinical data obtained throughout France underlined the consistency between the identified genotypes. However, some genotypes were locally present in effluents and were not found in the analysis of the clinical data. These findings could highlight an underestimation of the diversity of enteric viruses circulating in the human population. Consequently, analysis of WWTP effluents could allow the exploration of viral diversity not only in environmental waters but also in a human population linked to a sewerage network in order to better comprehend viral epidemiology and to forecast seasonal outbreaks. © 2015, American Society for Microbiology

Effect of concentration on virus removal for ultrafiltration membrane in drinking water production

International audienceRemoval of pathogenic microorganisms as viruses during drinking water production was evaluated by ultrafiltration. Two enteric viruses (ADV 41 and CV-B5) were compared to the MS2 bacteriophage, largely used in literature and by membrane producers as enteric virus surrogate. The effect of feed concentration of viruses on the ultrafiltration efficiency has been assessed. For the three viruses, low retentions about 1 log were observed at the lowest concentrations. At higher concentrations, an increase of removal up to 3.0 log for CV-B5 and MS2 phage and 3.5 log for ADV 41 was observed. These results highlight the potential overestimation of UF efficiency during laboratory experiments realized at high concentrations, compared to low concentrations found in environmental resources used for drinking water production. Virus removals with Evian water and real groundwater were compared and groundwater achieved similar or slightly higher removals for the three viruses. Finally, impact of membrane ageing after chlorine exposure was checked. It was observed that membrane degradations, visible by a water permeability increase with exposure dose did not affect the removal of viruses at low feed concentrations

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

International audienceAfter 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

Retention and integrity of drinking water production membranes towards nanoparticles and viruses

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Integrity of ultrafiltration membranes (UF) towards nanoparticles (NPs) and viruses for drinking water production

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Rétention des nanoparticules et virus : intégrité des membranes d’ultrafiltration dans la production d’eau potable

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