Yihong Hu1, Zhenzhou Wan2, Yonglin Mu3, Yi Zhou4, Jia Liu5, Ke Lan6, Chiyu Zhang7. 1. Pathogen Discovery and Big Data Center, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, China. yhhu@ips.ac.cn. 2. Medical Laboratory of Taizhou Fourth People's Hospital, Taizhou, China. wanlv@126.com. 3. Key Laboratory of Computational Biology, Chinese Academy of Sciences-Max Planck Partner Institute for Computational Biology, Chinese Academy of Sciences, Shanghai, China. muyonglin@picb.ac.cn. 4. Pathogen Discovery and Big Data Center, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, China. yizhou@ips.ac.cn. 5. Pathogen Discovery and Big Data Center, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, China. liuj@yiruibio.com.cn. 6. State Key Laboratory of Virology, College of Life Sciences, Medical Research Institute, Wuhan University, Wuhan, China. klan@whu.edu.cn. 7. Pathogen Discovery and Big Data Center, Institut Pasteur of Shanghai, Chinese Academy of Sciences, Shanghai, China. zhangcy1999@ips.ac.cn.
Abstract
INTRODUCTION: Human respiratory syncytial virus (hRSV) is a common respiratory virus closely related to respiratory tract infection (RTI). Rapid and accurate detection of hRSV is urgently needed to reduce the high morbidity and mortality due to hRSV infection. METHODOLOGY: Here, we established a highly sensitive and specific reverse-transcription loop-mediated isothermal amplification (RT-LAMP) assay for the rapid detection of A and B group hRSV simultaneously. The specific primer sets for hRSV A and B groups were designed in the M and M2-2 gene, respectively. SYTO 9 was used as the fluorescent dye for real-time monitoring of the amplification of hRSV RNA without cross reaction between hRSV A and B. RESULTS: The limit of detection (LOD) of our new method was 281.17 50% tissue culture infective doses (TCID50)/mL for hRSV A and 1.58 TCID50/mL for hRSV B. Using 90 clinical samples, a comparison to traditional RT-PCR was performed to validate this assay. The positivity rate of RT-LAMP and RT-PCR were 67.8% and 55.6%, respectively, and the positivity rate of RT-LAMP was significantly higher than RT-PCR (χ2 test, P < 0.01). CONCLUSIONS: Compared with traditional RT-PCR method, the newly developed fluorescent RT-LAMP combined with well-designed primers and SYTO 9 is quite sensitive, specific, rapid and well applicable to hRSV clinical diagnosis. Copyright (c) 2019 Yihong Hu, Zhenzhou Wan, Yonglin Mu, Yi Zhou, Jia Liu, Ke Lan, Chiyu Zhang.
INTRODUCTION:Human respiratory syncytial virus (hRSV) is a common respiratory virus closely related to respiratory tract infection (RTI). Rapid and accurate detection of hRSV is urgently needed to reduce the high morbidity and mortality due to hRSV infection. METHODOLOGY: Here, we established a highly sensitive and specific reverse-transcription loop-mediated isothermal amplification (RT-LAMP) assay for the rapid detection of A and B group hRSV simultaneously. The specific primer sets for hRSV A and B groups were designed in the M and M2-2 gene, respectively. SYTO 9 was used as the fluorescent dye for real-time monitoring of the amplification of hRSV RNA without cross reaction between hRSV A and B. RESULTS: The limit of detection (LOD) of our new method was 281.17 50% tissue culture infective doses (TCID50)/mL for hRSV A and 1.58 TCID50/mL for hRSV B. Using 90 clinical samples, a comparison to traditional RT-PCR was performed to validate this assay. The positivity rate of RT-LAMP and RT-PCR were 67.8% and 55.6%, respectively, and the positivity rate of RT-LAMP was significantly higher than RT-PCR (χ2 test, P < 0.01). CONCLUSIONS: Compared with traditional RT-PCR method, the newly developed fluorescent RT-LAMP combined with well-designed primers and SYTO 9 is quite sensitive, specific, rapid and well applicable to hRSVclinical diagnosis. Copyright (c) 2019 Yihong Hu, Zhenzhou Wan, Yonglin Mu, Yi Zhou, Jia Liu, Ke Lan, Chiyu Zhang.
Authors: Abhijeet A Bakre; Les P Jones; Hailey K Bennett; Davis E Bobbitt; Ralph A Tripp Journal: J Virol Methods Date: 2020-11-30 Impact factor: 2.014
Authors: E Alperay Tarim; Betul Karakuzu; Cemre Oksuz; Oyku Sarigil; Melike Kizilkaya; Mahmoud Khatib A A Al-Ruweidi; Huseyin Cagatay Yalcin; Engin Ozcivici; H Cumhur Tekin Journal: Emergent Mater Date: 2021-03-25