F-Specific RNA Bacteriophages Model the Behavior of Human Noroviruses during Purification of Oysters: the Main Mechanism Is Probably Inactivation Rather than Release

Abstract

Noroviruses (NoV) are responsible for many shellfish outbreaks. Purification processes may be applied to oysters before marketing to decrease potential fecal pollution. This step is rapidly highly effective in reducing Escherichia coli; nevertheless, the elimination of virus genomes has been described to be much slower. It is therefore important to identify (i) the purification conditions that optimize virus removal and (ii) the mechanism involved. To this end, the effects of oyster stress, nutrients, and the presence of a potential competitor to NoV adhesion during purification were investigated using naturally contaminated oysters. Concentrations of NoV (genomes) and of the viral indicator F-specific RNA bacteriophage (FRNAPH; genomes and infectious particles) were regularly monitored. No significant differences were observed under the test conditions. The decrease kinetics of both virus genomes were similar, again showing the potential of FRNAPH as an indicator of NoV behavior during purification. The T90 (time to reduce 90% of the initial titer) values were 47.8 days for the genogroup I NoV genome, 26.7 days for the genogroup II NoV genome, and 43.9 days for the FRNAPH-II genome. Conversely, monitoring of the viral genomes could not be used to determine the behavior of infectious viruses because the T90 values were more than two times lower for infectious FRNAPH (20.6 days) compared to their genomes (43.9 days). Finally, this study highlighted that viruses are primarily inactivated in oysters rather than released in the water during purification processes. IMPORTANCE This study provides new data about the behavior of viruses in oysters under purification processes and about their elimination mechanism. First, a high correlation has been observed between F-specific RNA bacteriophages of subgroup II (FRNAPH-II) and norovirus (NoV) in oysters impacted by fecal contamination when both are detected using molecular approaches. Second, when using reverse transcription-quantitative PCR and culture to detect FRNAPH-II genomes and infectious FRNAPH in oysters, respectively, it appears that genome detection provides limited information about the presence of infectious particles. The comparison of both genomes and infectious particles highlights that the main mechanism of virus elimination in oysters is inactivation. Finally, this study shows that none of the conditions tested modify virus removal.

The Utility of Dreissena polymorpha for Assessing the Viral Contamination of Rivers by Measuring the Accumulation of F-Specific RNA Bacteriophages

“…Infectious FRNAPHs appeared to be inactivated faster in water than in the mussels, which highlighted the protective effect of the mussel tissue. T90 values of~20 days are currently described in the literature for the reduction of infectious FRNAPHs in oysters at 10 • C [25,36], which is in the same range as our study.…”

“…These specific interactions favor HuNoV accumulation in digestive tissues [38,40]. The accumulation of FRNAPHs in the digestive tissue of oysters is also well documented [25,36,41]. Further experiments are required to better understand the specific and/or non-specific interactions between infectious FRNAPHs and the soft tissue of zebra mussels.…”

“…These results suggested that specific and/or non-specific interactions between the FRNAPHs and the mussel soft tissue led to low rates of depuration in D. polymorpha. As Leduc et al [36] suggested, the main mechanism involved in the depuration carried out by the mussels is not the release of FRNAPHs into the water but their inactivation in soft tissue. In this context, it is relevant to understand why and how sentinel species such as zebra mussels process high levels of viral particles in surface water.…”

Evaluation of Norovirus Reduction in Environmentally Contaminated Pacific Oysters During Laboratory Controlled and Commercial Depuration

“…In contrast, all oysters in the tested population were found to be contaminated when the mean concentration tested above 300 gcg −1 (Hunt 2019 ). Additionally, two recent studies investigated the FRNA bacteriophage type II (norovirus surrogate) removal from oysters during depuration using genomic and infectivity methods (Leduc et al 2020 ; Younger et al 2020 ). These studies indicated that viruses may either be inactivated in oysters during depuration (Leduc et al 2020 ) or, as suggested by Younger et al ( 2020 ), destroyed and removed.…”

“…Additionally, two recent studies investigated the FRNA bacteriophage type II (norovirus surrogate) removal from oysters during depuration using genomic and infectivity methods (Leduc et al 2020 ; Younger et al 2020 ). These studies indicated that viruses may either be inactivated in oysters during depuration (Leduc et al 2020 ) or, as suggested by Younger et al ( 2020 ), destroyed and removed. Despite the differences in the concluded viral reduction mechanisms these findings could indicate that the virus genome copies detected using RT-PCR methods are, at least in part, only footprints left by the infectious virus particles and they no longer have the capability of causing infection especially at low concentrations (i.e.…”