Three multiplex hemi-nested RT-PCR assays were developed to detect simultaneously 12 RNA respiratory viruses: influenza viruses A, B and C, human respiratory syncytial virus (hRSV), human metapneumovirus (hMPV), parainfluenza virus types 1–4 (PIV-1, -2, -3 and -4), human coronavirus OC43 and 229E (HCoV) and rhinovirus (hRV). An internal amplification control was included in one of the RT-PCR assays. The RT-PCR multiplex 1 and the hemi-nested multiplex 1 detected 1 and 0.1 TCID50 of RSV A, respectively, and 0.01 and 0.001 TCID50 of influenza virus A/H3N2, respectively. Two hundred and three nasal aspirates from hospitalised children were retrospectively tested in comparison with two conventional methods: direct immunofluorescence assay and viral isolation technique. Almost all samples (89/91) that were positive by immunofluorescence assay and/or viral isolation technique were detected by the multiplex assay. This method also detected an additional 85 viruses and 33 co-infections. The overall sensitivity (98%), rapidity and enhanced efficiency of these multiplex hemi-nested RT-PCR assays suggest that they would be a significant improvement over conventional methods for the detection of a broad spectrum of respiratory viruses.
【저자키워드】 Influenza virus, Human coronavirus, rhinovirus, respiratory syncytial virus, parainfluenza virus, human metapneumovirus, Multiplex RT-PCR assay, 【초록키워드】 children, nasal, Influenza virus, RT-PCR, RNA, respiratory viruses, amplification, sensitivity, Multiplex assay, rhinovirus, respiratory syncytial virus, RSV, immunofluorescence assay, Isolation, human coronavirus OC43, multiplex, parainfluenza virus, development, RT-PCR assay, 229E, Efficiency, RT-PCR assays, positive, métapneumovirus, hospitalised, TCID50, detected, tested, detect, included, virus, influenza virus, respiratory RNA virus,