Free Chlorine and Peroxynitrite Alter the Capsid Structure of Human Norovirus GII.4 and Its Capacity to Bind Histo-Blood Group Antigens.

Human noroviruses (HuNoVs) are one of the leading causes of acute gastroenteritis worldwide. HuNoVs are frequently detected in water and foodstuffs. Free chlorine and peroxynitrite (ONOO-) are two oxidants commonly encountered by HuNoVs in humans or in the environment during their natural life cycle. In this study, we defined the effects of these two oxidants on GII.4 HuNoVs and GII.4 virus-like particles (VLPs). The impact on the capsid structure, the major capsid protein VP1 and the ability of the viral capsid to bind to histo-blood group antigens (HBGAs) following oxidative treatments were analyzed. HBGAs are attachment factors that promote HuNoV infection in human hosts. Overall, our results indicate that free chlorine acts on regions involved in the stabilization of VP1 dimers in VLPs and affects their ability to bind to HBGAs. These effects were confirmed in purified HuNoVs. Some VP1 cross-links also take place after free chlorine treatment, albeit to a lesser extent. Not only ONOO- mainly produced VP1 cross-links but can also dissociate VLPs depending on the concentration applied. Nevertheless, ONOO- has less effect on HuNoV particles.

The effect of proteolytic enzymes and pH on GII.4 norovirus, during both interactions and non-interaction with Histo-Blood Group Antigens.

Human noroviruses (HuNoVs) are the leading cause of acute gastroenteritis worldwide. Histo-Blood Groups Antigens (HBGAs) have been described as attachment factors, promoting HuNoV infection. However, their role has not yet been elucidated. This study aims to evaluate the ability of HBGAs to protect HuNoVs against various factors naturally found in the human digestive system. The effects of acid pH and proteolytic enzymes (pepsin, trypsin, and chymotrypsin) on GII.4 virus-like particles (VLPs) and GII.4 HuNoVs were studied, both during interactions and non-interaction with HBGAs. The results showed that GII.4 VLPs and GII.4 HuNoVs behaved differently following the treatments. GII.4 VLPs were disrupted at a pH of less than 2.0 and in the presence of proteolytic enzymes (1,500 units/mL pepsin, 100 mg/mL trypsin, and 100 mg/mL chymotrypsin). VLPs were also partially damaged by lower concentrations of trypsin and chymotrypsin (0.1 mg/mL). Conversely, the capsids of GII.4 HuNoVs were not compromised by such treatments, since their genomes were not accessible to RNase. HBGAs were found to offer GII.4 VLPs no protection against an acid pH or proteolytic enzymes.

Effect of natural ageing and heat treatments on GII.4 norovirus binding to Histo-Blood Group Antigens.

Human noroviruses (HuNoVs) are the leading cause of viral foodborne outbreaks worldwide. To date, no available methods can be routinely used to detect infectious HuNoVs in foodstuffs. HuNoVs recognize Histo-Blood Group Antigens (HBGAs) through the binding pocket (BP) of capsid protein VP1, which promotes infection in the host cell. In this context, the suitability of human HBGA-binding assays to evaluate the BP integrity of HuNoVs was studied on GII.4 virus-like particles (VLPs) and GII.4 HuNoVs during natural ageing at 20 °C and heat treatments. Our results demonstrate that this approach may reduce the over-estimation of potential infectious HuNoVs resulting from solely using the genome detection, even though some limitations have been identified. The specificity of HBGA-binding to the BP is clearly dependent on the HGBA type (as previously evidenced) and the ionic strength of the media without disturbing such interactions. This study also provides new arguments regarding the ability of VLPs to mimic HuNoV behavior during inactivation treatments. The BP stability of VLPs was at least 4.3 fold lower than that of HuNoVs at 20 °C, whereas capsids of both particles were disrupted at 72 °C. Thus, VLPs are relevant surrogates of HuNoVs for inactivation treatments inducing significant changes in the capsid structure.