Detection and serotyping of rotaviruses in stool specimens by using reverse transcription and polymerase chain reaction amplification

✍ Scribed by Hiroshi Ushijima; Hinako Koike; Atsushi Mukoyama; Ayako Hasegawa; Shuichi Nishimura; Jon Gentsch

• Publisher: John Wiley and Sons
• Year: 1992
• Tongue: English
• Weight: 662 KB
• Volume: 38
• Category: Article
• ISSN: 0146-6615
• DOI: 10.1002/jmv.1890380412

✦ Synopsis

Abstract

Direct rotavirus serotyping (VP7, G type) in stool specimens was carried out by reverse transcription and polymerase chain reaction amplification (RT‐PCR) and compared to serotyping by enzyme immunoassay with monoclonal antibodies (EIA‐MAb). The methods used for double‐stranded (ds) RNA extraction, RT‐PCR amplification, and the primers used were modified from previous reports [Gouvea et al.: Journal of Clinical Microbiology 29:519–523, 1990; Gentsch et al.: Journal of Clinical Microbiology, 1992]. For samples that were positive by both methods, the serotypes obtained were identical, however RT‐PCR typing was found to be considerably more sensitive (70.4% samples serotyped) than EIA‐MAb (35.6% of samples serotyped). The overall sensitivities for detection of rotavirus in stool samples by latex agglutination, enzyme immunoassay, electron microscopy, polyacrylamide gel electrophoresis, and RT‐PCR were essentially the same. The results confirm that RT‐PCR typing (genotyping) is extremely valuable for G typing of samples which cannot be typed by EIA‐MAb. We also developed a PCR confirmation technique for serotypes 1, 2, and 4. © 1992 Wiley‐Liss, Inc.