Evaluation of an extraction-precipitation method for recovering hepatitis A virus and poliovirus from hardshell clams (mercenaria mercenaria)

Abstract

Epidemiological evidence of shellfish mediated transmission of enteric viral diseases and the inability of bacterial indicators to reliably predict viral contamination of shellfish highlight the risks associated with shellfish consumption. This problem suggests the need for direct virological examination of shellfish, and a number of methods have been developed to recover enteroviruses from clams and other bivalve molluscs. These methods have been evaluated primarily for recovery of poliovirus, and there has been little or no testing of their ability to recover hepatitis A virus (HAV). The present study was done to evaluate a new extraction-precipitation method for recovery of both poliovirus and HAV from experimentally contaminated hard clams. In the method 50-gram clam samples are homogenized after supplementing with saline and beef extract and then extracted with an equal volume of fluorocarbon. After centrifugation, viruses in the supernatant are precipitated with 12% polyethylene glycol, resuspended in buffered saline and assayed in cell cultures. From a series of replicate trials using clams seeded with test viruses, this method gave poliovirus and HAV recoveries of about 53 and 22%, respectively. Although the recovery of HAV was lower than that of poliovirus, it was considered adequate for further application. In subsequent studies using clams experimentally contaminated by virus uptake from water, the recovery method detected naturally accumulated HAV and poliovirus in clams. The method evaluated in this study is one of the few documented as capable of detecting HAV as well as enteroviruses in hard clams.