Waku-Kouomou Diane, Alla Amal, Blanquier Bariza, Jeantet Damien, Caidi Hayat, Rguig Ahmed, Freymuth François, Wild Fabian T
INSERM U404-Immunity and Vaccination, IFR128, 21 Avenue Tony Garnier, 69365 Lyon Cédex 03, France.
J Clin Microbiol. 2006 Feb;44(2):487-94. doi: 10.1128/JCM.44.2.487-494.2006.
Real-time PCR has been developed to genotype measles virus (MV) isolates. MV strains circulating in epidemics in Gabon in 1984, Cameroon in 2001, Morocco in 2003, and France in 2004 were investigated. We developed a real-time amplification refractory mutation system PCR (RT-AMRS PCR) using SYBR green fluorescent dye. Six pairs of primers for RT-ARMS PCR were designed to specifically amplify genotypes A, B2, B3.1, B3.2, C2, and D7. Genotypes could be differentiated by melting curve analysis. All strains were also confirmed by direct sequencing. Using the result obtained by direct sequencing and phylogenetic analysis as the reference, the accuracy of MV by RT-ARMS PCR and melting curve analysis was 97%. However, the latter method is more rapid and sensitive than the former method. This method could be a useful tool for molecular epidemiological studies of MV, providing an efficient alternative for large-scale studies.
实时聚合酶链反应(PCR)已被用于麻疹病毒(MV)分离株的基因分型。对1984年在加蓬、2001年在喀麦隆、2003年在摩洛哥以及2004年在法国流行期间传播的MV毒株进行了调查。我们使用SYBR绿色荧光染料开发了一种实时扩增阻滞突变系统PCR(RT-AMRS PCR)。设计了6对用于RT-ARMS PCR的引物,以特异性扩增A、B2、B3.1、B3.2、C2和D7基因型。通过熔解曲线分析可以区分基因型。所有毒株也通过直接测序得到了确认。以直接测序和系统发育分析获得的结果为参考,RT-ARMS PCR和熔解曲线分析对MV的分型准确率为97%。然而,后一种方法比前一种方法更快速、灵敏。该方法可能是MV分子流行病学研究的有用工具,为大规模研究提供了一种有效的替代方法。