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An improved method to detect Norwalk-like caliciviruses in environmental samples |
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Auteurs : LOISY F. (1), Le CANN P. (1), POMMEPUY M. (1), LE GUYADER F. (1) Journal : Letters in Applied Microbiology, 2000, vol. 31, 411-414. Type : Publication Affiliation : (1) Laboratoire de Microbiologie, IFREMER, Nantes, FRANCE Résumé : An original magnetic beads RNA capture was developed for the detection of Norwalk-like virus by RT-PCR. The same oligonucleotide was used both for capture and reverse transcription of the viral RNA. The optimization studies showed that the most important parameter for sensitivity is the biotin-binding capacity of the beads. This method was found to be efficient for eliminating inhibitors in sewage samples compared with classic RT-PCR. Moreover, the sensitivity was greatly enhanced, allowing the detection of 42% positive sample after gel electrophoresis, which is fourfold greater than classic RT-PCR (11%). Beads-RT-PCR sensitivity is the same as classic RT-PCR and hybridization. Thus, this method, which is easy to perform, should be of particular interest for developing quantitative RT-PCR and sequencing. Mots Clés : Norwalk-like virus, sewage samples, Calicivirus, Caliciviridae, Réaction en Chaîne par Polymérase, RT-PCR |
