Determination of norovirus contamination in oysters from two commercial harvesting areas over an extended period, using semiquantitative real-time reverse transcription PCR

The human health risk associated with the consumption of molluscan shellfish grown in sewage-contaminated waters is well established. Noroviruses, which cause gastroenteritis, are the principal agents of shellfish-related illness. Fecal-indicator quality standards based on Escherichia coli are well established in Europe and elsewhere. However, norovirus outbreaks after consumption of shellfish meeting these standards still occur, and the need to improve consumer health protection is well recognized. Alternative approaches proposed include direct monitoring of viral pathogens and the use of alternative indicator organisms capable of providing a better indication of virus risk. This study applies a recently developed TaqMan PCR assay to assess norovirus contamination in shellfish. Comparison was made with E. coli as the existing sanitary standard and a male-specific RNA bacteriophage as a possible alternative. Two commercial pacific oyster (Crassostrea gigas) harvesting areas were monitored over a 31-month period. The results show peaks of norovirus contamination in both areas during winter months, with average levels approximately 17 times higher in oysters sampled October to March than during the remainder of the year, consistent with epidemiological data for the United Kingdom showing oyster-associated illness is confined to winter months. While there was no apparent association with E. coli, an association between levels of norovirus contamination and the malespecific RNA bacteriophage was noted, with average norovirus levels over 40 times higher in samples with male-specific RNA bacteriophage counts of > 1,000 PFU/100 g than in samples with < 100 PFU/100 g. Overall, these results suggest that norovirus monitoring in shellfish production areas could be an effective strategy for reduction of virus risk.