To examine the long-term infectivity of human norovirus in water, 13 study subjects were challenged at different time points with groundwater spiked with the prototype human norovirus, Norwalk virus. Norwalk virus spiked in groundwater remained infectious after storage at room temperature in the dark for 61 days (the last time point tested). The Norwalk virus-seeded groundwater was stored for 1,266 days and analyzed, after RNase treatment, by reverse transcription-quantitative PCR (RT-qPCR) to detect Norwalk virus RNA contained within intact capsids. Norwalk virus RNA within intact capsids was detected in groundwater for 1,266 days, with no significant log 10 reduction throughout 427 days and a significant 1.10-log 10 reduction by day 1266. Purified Norwalk virus RNA (extracted from Norwalk virus virions) persisted for 14 days in groundwater, tap water, and reagent-grade water. This study demonstrates that Norwalk virus in groundwater can remain detectable for over 3 years and can remain infectious for at least 61 days. (ClinicalTrials.gov identifier NCT00313404.)

Contamination of oysters with human noroviruses (HuNoV) constitutes a human health risk and may lead to severe economic losses in the shellfish industry. There is a need to identify a technology that can inactivate HuNoV in oysters. In this study, we conducted a randomized, double-blinded clinical trial to assess the effect of high hydrostatic pressure processing (HPP) on Norwalk virus (HuNoV genogroup I.1) inactivation in virus-seeded oysters ingested by subjects. Forty-four healthy, positive-secretor adults were divided into three study phases. Subjects in each phase were randomized into control and intervention groups. Subjects received Norwalk virus (8FIIb, 1.0 ؋ 10 4 genomic equivalent copies) in artificially seeded oysters with or without HPP treatment (400 MPa at 25°C, 600 MPa at 6°C, or 400 MPa at 6°C for 5 min). HPP at 600 MPa, but not 400 MPa (at 6°or 25°C), completely inactivated HuNoV in seeded oysters and resulted in no HuNoV infection among these subjects, as determined by reverse transcription-PCR detection of HuNoV RNA in subjects' stool or vomitus samples. Interestingly, a white blood cell (granulocyte) shift was identified in 92% of the infected subjects and was significantly associated with infection (P ‫؍‬ 0.0014). In summary, these data suggest that HPP is effective at inactivating HuNoV in contaminated whole oysters and suggest a potential intervention to inactivate infectious HuNoV in oysters for the commercial shellfish industry.

Noroviruses (NoV) are the main cause of epidemic gastroenteritis worldwide. We have previously shown that both an individual's secretor status (genetic) and induction of Norwalk virus-specific salivary IgA (immune) are important predictors of infection with Norwalk virus (NV). We still do not understand the factors that mediate the immune response, and infection, to Norwalk virus. To address this need, we have extended our analysis of immunological, genetic, and demographic parameters of 109 volunteers challenged with various doses of Norwalk virus through univariate and multivariate regression analyses. We found that gender, ethnicity, levels of pre-challenge NV-specific serum IgG or levels of pre-challenge NV-specific salivary IgA were not significant predictors of infection or development of a NV-specific antibody response. The presence but not magnitude of pre-challenge serum NV-specific IgG and secretor positive status were significantly associated with infection. Dose of inoculum received and age of the individual may also be predictors of infection and development of an NV-specific antibody response. We observed significant collinearity between the age of the volunteer and the presence of pre-challenge serum NV-specific IgG. Lastly, we found a strong association between levels of salivary and serum NV-specific antibody levels, after adjusting for covariates. Taken together, these data will be used to better understand the immune response to NV infection and design effective NoV vaccines to protect at risk populations including children, food handlers, and the elderly.

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