Relevance of F-Specific RNA Bacteriophages in Assessing Human Norovirus Risk in Shellfish and Environmental Waters

Hartard, Cédric; Banas, Sandrine; Loutreul, Julie; Rincé, Alain; Benoit, Fabienne; Boudaud, Nicolas; Gantzer, Christophe

doi:10.1128/aem.01528-16

Cited by 25 publications

(12 citation statements)

References 61 publications

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“…The use of a qualitative approach (37) underlined that more than 90% of the specimens were positive for FRNAPH infectious particles (i.e., 84.7% were positive for infectious FRNAPH-I and 64.9% were positive for infectious FRNAPH-II). We confirmed, in accordance with other works, the lack of a relation between the regulatory indicator E. coli and infectious viral indicators, such as FRNAPH (Fisher's exact test, P ϭ 0.8374) (24,41,44). Compared to the results obtained during a 1-year period of surveillance of marketed oysters, for which only 34% and 7% of the samples were positive for infectious FRNAPH-I and -II, respectively, the oysters sampled in the present study seemed to be more impacted by fecal pollution when the same qualitative method was used (37).…”

Section: Discussionsupporting

confidence: 92%

“…In the light of this, a strict comparison should be made only by investigating the genomes of both NoV and FRNAPH. In doing so, a good correlation between the presence of NoV and that of FRNAPH, especially when human FRNAPH-II is considered, has been observed (24,26). Nevertheless, this observation was made with a limited number of data.…”

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confidence: 96%

“…Indeed, even though it has never been demonstrated in oysters, it is very well recognized that viral genomes have a better persistence than infectious viruses in the environment, and thus, only a negligible part of the detected genomes in fact corresponds to infectious particles (19)(20)(21)(22), especially after inactivation by UV light (23). More alarmingly, while the absence of genomes is necessarily a sign of the absence of the corresponding infectious viruses, the strict application of the ISO 15216-1 standard expresses a theoretical limit of detection (LOD) of greater than the 18 particles, usually defined to be the minimal NoV infective dose (24,25). It can be 10 to 100 times higher because of the small volume analyzed by reverse transcription (RT)-PCR, the low recovery rate during genome extraction, and the presence of (RT-)PCR inhibitors.…”

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confidence: 99%

“…A recent review of the literature shows the advantages and limitations of FRNAPH as viral indicators in shellfish management (42). FRNAPH have been discredited because, even though a number of studies have pointed out a positive correlation between the presence of FRNAPH and that of enteric viruses in shellfish (24,26,(43)(44)(45)(46)(47), others have expressed opposite views (48)(49)(50). A closer look reveals that most studies compare the detection of infectious FRNAPH with that of the human NoV genome.…”

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confidence: 99%

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F-Specific RNA Bacteriophages, Especially Members of Subgroup II, Should Be Reconsidered as Good Indicators of Viral Pollution of Oysters

Hartard

Leclerc

Rivet

et al. 2018

Appl Environ Microbiol

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Norovirus (NoV) is the leading cause of gastroenteritis outbreaks linked to oyster consumption. In this study, we investigated the potential of F-specific RNA bacteriophages (FRNAPH) as indicators of viral contamination in oysters by focusing especially on FRNAPH subgroup II (FRNAPH-II). These viral indicators have been neglected because their behavior is sometimes different from that of NoV in shellfish, especially during the depuration processes usually performed before marketing. However, a significant bias needs to be taken into account. This bias is that, in the absence of routine culture methods, NoV is targeted by genome detection, while the presence of FRNAPH is usually investigated by isolation of infectious particles. In this study, by targeting both viruses using genome detection, a significant correlation between the presence of FRNAPH-II and that of NoV in shellfish collected from various European harvesting areas impacted by fecal pollution was observed. Moreover, during their depuration, while the long period of persistence of NoV was confirmed, a similar or even longer period of persistence of the FRNAPH-II genome, which was over 30 days, was observed. Such a striking genome persistence calls into question the relevance of molecular methods for assessing viral hazards. Targeting the same virus (i.e., FRNAPH-II) by culture and genome detection in specimens from harvesting areas as well as during depuration, we concluded that the presence of genomes in shellfish does not provide any information on the presence of the corresponding infectious particles. In view of these results, infectious FRNAPH detection should be reconsidered as a valuable indicator in oysters, and its potential for use in assessing viral hazard needs to be investigated. This work brings new data about the behavior of viruses in shellfish, as well as about the relevance of molecular methods for their detection and evaluation of the viral hazard. First, a strong correlation between the presence of F-specific RNA bacteriophages of subgroup II (FRNAPH-II) and that of norovirus (NoV) in shellfish impacted by fecal contamination has been observed when both viruses are detected using molecular approaches. Second, when reverse transcription-PCR and culture are used to detect FRNAPH-II in shellfish, it appears that the genomes of the viruses present a longer period of persistence than infectious virus, and thus, virus genome detection fails to give information about the concomitant presence of infectious viruses. Finally, this study shows that FRNAPH persist at least as long as NoV does. These data are major arguments to reconsider the potential of FRNAPH as indicators of shellfish viral quality.

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Section: Discussionsupporting

confidence: 92%

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confidence: 96%

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confidence: 99%

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confidence: 99%

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F-Specific RNA Bacteriophages, Especially Members of Subgroup II, Should Be Reconsidered as Good Indicators of Viral Pollution of Oysters

Hartard

Leclerc

Rivet

et al. 2018

Appl Environ Microbiol

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“…Four genogroups of FRNAPH have been described and commonly circulate in the environment (23). Among them, genogroups II (FRNAPH-II) and III (FRNAPH-III) are mainly associated with human fecal pollution (24), while genogroups I (FRNAPH-I) and IV (FRNAPH-IV) are mainly related to animal fecal pollution. A positive correlation has recently been demonstrated by Hartard et al between the genomes of FRNAPH-II and NoV in oysters during purification processes (22), showing that FRNAPH-II could be proposed as a potential indicator to evaluate NoV risk in shellfish (25).…”

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confidence: 99%

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

Leduc

Leclerc

Challant

et al. 2020

Appl Environ Microbiol

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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.

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Novel Approaches for Detecting Water-Associated Pathogens

Maurya

Kumar

Saxena

2019

Water-Associated Infectious Diseases

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Relevance of F-Specific RNA Bacteriophages in Assessing Human Norovirus Risk in Shellfish and Environmental Waters

Cited by 25 publications

References 61 publications

F-Specific RNA Bacteriophages, Especially Members of Subgroup II, Should Be Reconsidered as Good Indicators of Viral Pollution of Oysters

F-Specific RNA Bacteriophages, Especially Members of Subgroup II, Should Be Reconsidered as Good Indicators of Viral Pollution of Oysters

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

Novel Approaches for Detecting Water-Associated Pathogens

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