tetano
Editor, Senior Moderator
Kawai Y, Kimura Y, Lezhava A, Kanamori H, Usui K, et al. (2012) One-Step Detection of the 2009 Pandemic Influenza A(H1N1) Virus by the RT-SmartAmp Assay and Its Clinical Validation. PLoS ONE 7(1): e30236. doi:10.1371/journal.pone.0030236
Yuki Kawai1, Yasumasa Kimura1, Alexander Lezhava1, Hajime Kanamori1, Kengo Usui1, Takeshi Hanami1, Takahiro Soma1, Jean-?tienne Morlighem1, Satomi Saga1, Yuri Ishizu1, Shintaro Aoki1, Ryuta Endo1, Atsuko Oguchi-Katayama1, Yasushi Kogo1, Yasumasa Mitani1, Takefumi Ishidao1, Chiharu Kawakami2, Hideshi Kurata2, Yumiko Furuya3, Takayuki Saito3, Norio Okazaki3, Masatsugu Chikahira4, Eiji Hayashi5, Sei-ichi Tsuruoka5, Tokumichi Toguchi5, Yoshitomo Saito6, Toshiaki Ban6, Shinyu Izumi7, Hideko Uryu7, Koichiro Kudo7, Yuko Sakai-Tagawa8, Yoshihiro Kawaoka8, Aizan Hirai5, Yoshihide Hayashizaki1, Toshihisa Ishikawa1*
1 Omics Science Center, RIKEN Yokohama Institute, Tsurumi-ku, Yokohama, Japan, 2 Yokohama City Institute of Health, Isogo-ku, Yokohama, Japan, 3 Kanagawa Prefectural Institute of Public Health, Chigasaki, Japan, 4 Hyogo Prefectural Institute of Public Health and Consumer Sciences, Hyogo-ku, Kobe, Japan, 5 Chiba Prefectural Togane Hospital, Togane, Japan, 6 Isumi Medical Center, Isumi, Japan, 7 National Center for Global Health and Medicine, Shinjuku-ku, Tokyo, Japan, 8 Institute of Medical Science, University of Tokyo, Shirokanedai, Minato-ku, Tokyo, Japan
Abstract Top
Background
In 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.
Methodology
To 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 Conclusions
We 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.
full article
http://www.plosone.org/article/info:doi/10.1371/journal.pone.0030236
Yuki Kawai1, Yasumasa Kimura1, Alexander Lezhava1, Hajime Kanamori1, Kengo Usui1, Takeshi Hanami1, Takahiro Soma1, Jean-?tienne Morlighem1, Satomi Saga1, Yuri Ishizu1, Shintaro Aoki1, Ryuta Endo1, Atsuko Oguchi-Katayama1, Yasushi Kogo1, Yasumasa Mitani1, Takefumi Ishidao1, Chiharu Kawakami2, Hideshi Kurata2, Yumiko Furuya3, Takayuki Saito3, Norio Okazaki3, Masatsugu Chikahira4, Eiji Hayashi5, Sei-ichi Tsuruoka5, Tokumichi Toguchi5, Yoshitomo Saito6, Toshiaki Ban6, Shinyu Izumi7, Hideko Uryu7, Koichiro Kudo7, Yuko Sakai-Tagawa8, Yoshihiro Kawaoka8, Aizan Hirai5, Yoshihide Hayashizaki1, Toshihisa Ishikawa1*
1 Omics Science Center, RIKEN Yokohama Institute, Tsurumi-ku, Yokohama, Japan, 2 Yokohama City Institute of Health, Isogo-ku, Yokohama, Japan, 3 Kanagawa Prefectural Institute of Public Health, Chigasaki, Japan, 4 Hyogo Prefectural Institute of Public Health and Consumer Sciences, Hyogo-ku, Kobe, Japan, 5 Chiba Prefectural Togane Hospital, Togane, Japan, 6 Isumi Medical Center, Isumi, Japan, 7 National Center for Global Health and Medicine, Shinjuku-ku, Tokyo, Japan, 8 Institute of Medical Science, University of Tokyo, Shirokanedai, Minato-ku, Tokyo, Japan
Abstract Top
Background
In 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.
Methodology
To 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 Conclusions
We 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.
full article
http://www.plosone.org/article/info:doi/10.1371/journal.pone.0030236