Nested multiplex PCR assay for detection of human enteric viruses in shellfish and sewage.

Environmental samples and contaminated shellfish present frequently low concentrations of more than one viral species. For this reason, a nested multiplex RT-PCR was developed for the detection of adenoviruses, enteroviruses and hepatitis A viruses in different environmental samples such as urban sewage and shellfish. This assay will save time and cost for detection of these enteric viruses with a smaller sample volume, which otherwise can be a limiting factor in routine analysis. The limit of detection was approximately 1 copy for adenovirus and 10 copies for enterovirus and hepatitis A virus per PCR reaction using titrated cell-cultured viruses as template material. In shellfish and environmental samples, this multiplex PCR was optimized to detect all three viruses simultaneously when the concentration of each virus was equal or lower than 1000 copies per PCR reaction. This is the level found predominantly in the environment and in shellfish when the numbers of fecal bacterial and phage indicators are low. The detection of human adenoviruses by PCR has been suggested as a molecular index of fecal contamination of human origin in the environment and food and the multiplex assay developed may be a tool for evaluating the presence of viral contamination in shellfish and water and to expand microbiological control to include viral markers.

Sporadic cases of acute autochthonous hepatitis E in Spain.

BACKGROUND/AIMS: In industrialized countries hepatitis E virus (HEV) infection is rare and its diagnosis is difficult because the utility of available tests is not well established. METHODS: We studied the presence of acute HEV infection markers in a cluster of 11 cases of acute hepatitis with IgG anti-HEV antibodies. RESULTS: Three cases were confirmed as acute hepatitis E and 8 as presumptive hepatitis E, two as a past HEV infection and one could not be determined. Three different HEV strains were identified in serum from 3 patients. Two strains belonged to genotype 3, the predominant genotype found in local urban sewage and the other strain belonged to genotype 1 and was considered an imported strain. CONCLUSIONS: Our findings demonstrate the presence of some autochthonous, sporadic acute hepatitis E cases as well as an imported case in our area and the transitory nature of virological and serological markers for HEV.