Eboigbodin Kevin, Filén Sanna, Ojalehto Tuomas, Brummer Mirko, Elf Sonja, Pousi Kirsi, Hoser Mark
Research and Development, Orion Diagnostica Oy, P. O. BOX 83, FI-02101, Espoo, Finland.
Molecular Biology, GeneForm Technologies, Broadstairs, UK.
Appl Microbiol Biotechnol. 2016 Jun;100(12):5559-67. doi: 10.1007/s00253-016-7491-y. Epub 2016 Apr 11.
Rapid and accurate diagnosis of influenza viruses plays an important role in infection control, as well as in preventing the misuse of antibiotics. Isothermal nucleic acid amplification methods offer significant advantages over the polymerase chain reaction (PCR), since they are more rapid and do not require the sophisticated instruments needed for thermal cycling. We previously described a novel isothermal nucleic acid amplification method, 'Strand Invasion Based Amplification' (SIBA®), with high analytical sensitivity and specificity, for the detection of DNA. In this study, we describe the development of a variant of the SIBA method, namely, reverse transcription SIBA (RT-SIBA), for the rapid detection of viral RNA targets. The RT-SIBA method includes a reverse transcriptase enzyme that allows one-step reverse transcription of RNA to complementary DNA (cDNA) and simultaneous amplification and detection of the cDNA by SIBA under isothermal reaction conditions. The RT-SIBA method was found to be more sensitive than PCR for the detection of influenza A and B and could detect 100 copies of influenza RNA within 15 min. The development of RT-SIBA will enable rapid and accurate diagnosis of viral RNA targets within point-of-care or central laboratory settings.
快速准确地诊断流感病毒在感染控制以及防止抗生素滥用方面发挥着重要作用。等温核酸扩增方法相较于聚合酶链反应（PCR）具有显著优势，因为它们速度更快，且不需要热循环所需的精密仪器。我们之前描述了一种新型等温核酸扩增方法——“基于链入侵的扩增”（SIBA®），该方法具有高分析灵敏度和特异性，用于检测DNA。在本研究中，我们描述了SIBA方法的一个变体——逆转录SIBA（RT - SIBA）的开发，用于快速检测病毒RNA靶标。RT - SIBA方法包括一种逆转录酶，它允许RNA一步逆转录为互补DNA（cDNA），并在等温反应条件下通过SIBA对cDNA进行同时扩增和检测。结果发现，RT - SIBA方法在检测甲型和乙型流感方面比PCR更灵敏，并且能够在15分钟内检测到100份流感RNA。RT - SIBA的开发将能够在即时护理点或中心实验室环境中对病毒RNA靶标进行快速准确的诊断。
