Quantification of Hepatitis A Virus in Shellfish by Competitive Reverse Transcription-PCR with Coextraction of Standard RNA

Abstract

ABSTRACT To quantify hepatitis A virus (HAV) in experimentally contaminated mussels, we developed an internal standard RNA with a 7-nucleotide deletion for competitive reverse transcription (RT)-PCR. Deposited directly into the sample, this standard was used both as extraction control and as quantification tool. After coextraction and competitive RT-PCR, standard and wild-type products were detected by differential hybridization with specific probes and a DNA enzyme immunoassay. The quantifiable range with this reproducible method was 10 4 to 10 7 copies of HAV/gram or 400 to 10 6 50% tissue culture infective doses/ml.