Genomics & Informatics

Korea Genome Organization (한국유전체학회)
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Development of reverse transcription loop-mediated isothermal amplification assays for point-of-care testing of avian influenza virus subtype H5 and H9

  • Zhang, Songzi (Department of Biomedicine & Health Sciences, Graduate School, The Catholic University of Korea) ;
  • Shin, Juyoun (ConnectaGen) ;
  • Shin, Sun (Department of Microbiology, College of Medicine, The Catholic University of Korea) ;
  • Chung, Yeun-Jun (Department of Biomedicine & Health Sciences, Graduate School, The Catholic University of Korea)
  • Received : 2020.10.06
  • Accepted : 2020.10.16
  • Published : 2020.12.31
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Abstract

Avian influenza (AIV) outbreaks can induce fatal human pulmonary infections in addition to economic losses to the poultry industry. In this study, we aimed to develop a rapid and sensitive point-of-care AIV test using loop-mediated isothermal amplification (LAMP) technology. We designed three sets of reverse transcription LAMP (RT-LAMP) primers targeting the matrix (M) and hemagglutinin (HA) genes of the H5 and H9 subtypes. RT-LAMP targeting the universal M gene was designed to screen for the presence of AIV and RT-LAMP assays targeting H5-HA and H9-HA were designed to discriminate between the H5 and H9 subtypes. All three RT-LAMP assays showed specific amplification results without nonspecific reactions. In terms of sensitivity, the detection limits of our RT-LAMP assays were 100 to 1,000 RNA copies per reaction, which were 10 times more sensitive than the detection limits of the reference reverse-transcription polymerase chain reaction (RT-PCR) (1,000 to 10,000 RNA copies per reaction). The reaction time of our RT-LAMP assays was less than 30 min, which was approximately four times quicker than that of conventional RT-PCR. Altogether, these assays successfully detected the existence of AIV and discriminated between the H5 or H9 subtypes with higher sensitivity and less time than the conventional RT-PCR assay.

Keywords

Acknowledgement

This work was supported by a grant from the National Research Foundation of Korea (2017R1E1A1A01074913 and 2017M3C9A6047615). We thank KREONET (Korea Research Environment Open NETwork) and KISTI (Korea Institute of Science and Technology Information) for allowing us to use their network infrastructure.

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