Abstract
BackgroundIn 2009, a pandemic (pdm) influenza A(H1N1) virus infection quickly circulated globally resulting in about 18,000 deaths around the world. In Japan, infected patients accounted for 16% of the total population. The possibility of human-to-human transmission of highly pathogenic novel influenza viruses is becoming a fear for human health and society.MethodologyTo address the clinical need for rapid diagnosis, we have developed a new method, the “RT-SmartAmp assay”, to rapidly detect the 2009 pandemic influenza A(H1N1) virus from patient swab samples. The RT-SmartAmp assay comprises both reverse transcriptase (RT) and isothermal DNA amplification reactions in one step, where RNA extraction and PCR reaction are not required. We used an exciton-controlled hybridization-sensitive fluorescent primer to specifically detect the HA segment of the 2009 pdm influenza A(H1N1) virus within 40 minutes without cross-reacting with the seasonal A(H1N1), A(H3N2), or B-type (Victoria) viruses.Results and ConclusionsWe evaluated the RT-SmartAmp method in clinical research carried out in Japan during a pandemic period of October 2009 to January 2010. A total of 255 swab samples were collected from outpatients with influenza-like illness at three hospitals and eleven clinics located in the Tokyo and Chiba areas in Japan. The 2009 pdm influenza A(H1N1) virus was detected by the RT-SmartAmp assay, and the detection results were subsequently compared with data of current influenza diagnostic tests (lateral flow immuno-chromatographic tests) and viral genome sequence analysis. In conclusion, by the RT-SmartAmp assay we could detect the 2009 pdm influenza A(H1N1) virus in patients' swab samples even in early stages after the initial onset of influenza symptoms. Thus, the RT-SmartAmp assay is considered to provide a simple and practical tool to rapidly detect the 2009 pdm influenza A(H1N1) virus.
Highlights
The 2009 pandemic influenza A(H1N1) virus, a new strain of virus identified in Mexico in April 2009, caused outbreaks on both local and global scales with severe consequences for human health and the global economy [1,2,3,4]
A total of 255 swab samples were collected from outpatients with influenza-like illness at three hospitals and eleven clinics located in the Tokyo and Chiba areas in Japan
The 2009 pdm influenza A(H1N1) virus was detected by the reverse transcriptase (RT)-SmartAmp assay, and the detection results were subsequently compared with data of current influenza diagnostic tests and viral genome sequence analysis
Summary
The 2009 pandemic (pdm) influenza A(H1N1) virus, a new strain of virus identified in Mexico in April 2009, caused outbreaks on both local and global scales with severe consequences for human health and the global economy [1,2,3,4]. Phylogenic analyses have revealed that the 2009 pdm influenza A(H1N1) viruses in Japan differed between the very early phase and the peak phase of the pandemic [6,7]. The 2009 pdm influenza A(H1N1) virus possesses PB2 and PA genes of North American avian virus origin, a PB1 gene of human H3N2 virus origin, HA (H1), NP, and NS genes of classical swine virus origin, and NA (N1) and M genes of Eurasian avianlike swine virus origin [9,10]. In 2009, a pandemic (pdm) influenza A(H1N1) virus infection quickly circulated globally resulting in about 18,000 deaths around the world. The possibility of human-to-human transmission of highly pathogenic novel influenza viruses is becoming a fear for human health and society
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