Detection of hepatitis A virus in bivalve molluscs in Southern Vietnam using RT-PCR

Vietnam has a large area for raising bivalve molluscs, especially in Southern Vietnam. However, water quality in the aquaculture areas is seriously polluted and there is a high potential of Hepatitis A virus (HAV) contamination. Bivalve molluscs are filter-feeder species; therefore the cumulative risk of pathogens in their body is high. Vietnam currently lacks surveillance data for HAV. This study was conducted to evaluate the level of microbial contamination and the prevalence of hepatitis A virus in bivalve mollusc and seawater samples in the Mekong Delta, Vietnam and to find the correlation between HAV and bacterial indicators in the samples. This study also assessed the applicability of the currently used Europe Union’s sanitary controls for consumers (regulation EC no.854/2004) to bivalve mollusc production in Vietnam.
One-hundred and twenty samples, comprising 60 seawater samples and 60 bivalve mollusc samples (clam) were collected from various regions in Ho Chi Minh City and Ben Tre province, Vietnam from August of 2008 to May of 2009. The presence of Salmonella spp., Escherichia coli and faecal coliforms were analyzed. One-hundred percent of these 120 samples were found to be contaminated with faecal coliforms and E. coli. However, Salmonella spp. contamination in the samples was low (3.33%). The level of faecal coliforms and E. coli in bivalve mollusc samples was higher than that in seawater samples. There were a significant difference in the level of faecal coliform and E. coli contamination between the wet season and the dry season in both seawater samples and bivalve mollusc samples. However, there were no significant difference in them between high tide and low tide.

The HAV from 120 samples was concentrated by using a developed virus concentration method and the viral RNA was detected by RT-hn-PCR. The RT-hn-PCR protocol was set up to detect the presence of HAV in the samples. The results showed high sensitivity with this method. Detection limit of HAV determined by the RT-hn-PCR was 1 TCID50 per 25g of shellfish homogenate or 2 liter of seawater sample. The HAV was found in 21/120 (17.5%) of the total samples, in which 4/60 (6.67%) of seawater samples and 17/60 (28.33%) of bivalve mollusc samples were contaminated with HAV. The Chi-Square test showed that there was no correlation either between HAV and E. coli or between HAV and faecal coliform in total samples and in each sampling sites.

The results of this study clearly demonstrate that E. coli and faecal coliforms are not effective indicators for microbiological safety of bivalve mollusc products in Southern Vietnam since they did not reflect the presence of HAV in the bivalve mollusc products. In Vietnam, the currently used Europe Union’s sanitary controls (EC No. 854/2004) should not be used for the sole assessment of the quality of bivalve mollusc products. Improved shellfish depuration methods are needed to obtain virus-safe shellfish and reduce the risk for public health in Vietnam.