Application of RT-PCR for the Detection of Enteric Viruses in Oysters

Oyster samples processed by adsorption-elution-precipitation were seeded with poliovirus 1 and HAV, and cleaned and concentrated by Freon extraction (2X), PEG precipitation and chloroform extraction. Freon extraction resulted in recoveries of 63-76% for polio and 42-52% for HAV. PEG precipitation/chloroform extraction gave recoveries of 47-50% for polio and 15-19% for HAV. Treated extracts inhibited RT-PCR at 10⁻² dilutions. Inhibitors were removed by treatment with the cationic detergent CTAB or Pro-Cipitate/UF adsorption-elution-concentration. Both treatments resulted in samples on which direct RT-PCR was possible. The CTAB procedure was able to detect 78 pfu of polio and 295 pfu of HAV. The Pro-Cipitate procedure was able to detect 70 pfu polio and 2.1×10³ pfu HAV.