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Applied and Environmental Microbiology, January 1999, p. 322-326, Vol. 65, No. 1
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

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

Charlotte Arnal,1,* Virginie Ferre-Aubineau,2 Berangere Mignotte,3 Berthe Marie Imbert-Marcille,2 and Sylviane Billaudel2

Laboratoire de Virologie, Institut de Biologie, Centre Hospitalier Universitaire, 44093 Nantes Cedex,1 Université de Nantes EA1156, 44035 Nantes,2 and Laboratoire de Virologie, UMR 7564 CNRS, Faculté de Pharmacie, Université Poincaré, 54001 Nancy Cedex,3 France

Received 3 August 1998/Accepted 28 October 1998

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 104 to 107 copies of HAV/gram or 400 to 106 50% tissue culture infective doses/ml.

* Corresponding author. Mailing address: Laboratoire de Virologie, Institut de Biologie, Centre Hospitalier Universitaire, 9 Quai Moncousu, 44093 Nantes Cedex, France. Phone: (33) 2 40 08 41 05. Fax: (33) 2 44 20 41 14. E-mail: sbi{at}sante.univ-nantes.fr.

Applied and Environmental Microbiology, January 1999, p. 322-326, Vol. 65, No. 1
0099-2240/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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Copyright © 1999 by the American Society for Microbiology. All rights reserved.