Journal of South China University of Technology(Natural Science) >

2023 , Vol. 51 >Issue 12: 140 - 151

DOI: https://doi.org/10.12141/j.issn.1000-565X.220529

Food Science & Technology

Two Visual and Rapid Enzymatic Recombinase Amplification Methods for GI and GII Noroviruses Detection

  • YANG Yange ,
  • WU Zhanwen ,
  • LI Tao ,
  • WANG Shuai ,
  • LI Hongna ,
  • SUN Dongmei ,
  • YUAN Fei
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  • 1.Key Laboratory of Food Quality and Safety for State Market Regulation，Chinese Academy of Inspection and Quarantine，Beijing 100176，China
    2.College of Life Science and Technology，Heilongjiang Bayi Agricultural University，Daqing 163000，Heilongjiang，China
杨艳歌（1986-），女，副研究员，主要从事分子生物学研究。E-mail： yange8602@126.com

Received date: 2022-08-17

  Online published: 2023-07-12

Supported by

the Chinese Academy of Inspection and Quarantine Project(2022JK36);the National Key Research and Development Program of China(2022YFF0607900)

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Abstract

Norovirus (NoV) is one of the common foodborne viruses, and even a small amount of NoV can lead to infection and illness. However, there are currently no specific therapeutic drugs and vaccines available, making it crucial to establish a rapid detection method for early screening of NoV in food products. This study first wrapped the target genes of GⅠ and GⅡ NoV specified in GB 4 789.42 into the capsid protein of bacteriophage MS2 with armored RNA technology, creating recombinant plasmid reference samples containing both GⅠ and GⅡ NoV targets. Next, based on the Enzymatic Recombinase Amplification (ERA) technology, the primers and probes of basic and fluorescence ERA detection were designed for both GⅠ and GⅡ NoV, and the optimal primers and probes were screened through experiments. Then two visualization methods for GI and GII NoV detection were established, namely ERA chromogenic and fluorescence method, and the results can be observed with the naked eye. Furthermore, the reaction program was optimized, reducing the amplification time to 5 minutes and 8 minutes for both ERA chromogenic and fluorescence methods, respectively. By optimizing the reaction system, the volume was halved, thereby reducing the cost of detection. Under these conditions, the lowest sensitivity was 10-2、10-3 ng/μL for the recombinant plasmid reference sample, respectively. Finally, the established visual rapid detection method was applied to the detection of GⅠ and GⅡ NoV authentic specimens. And the performance parameters of the method were analyzed. The results show that the established GⅠ and GⅡ NoV ERA visual rapid detection method has good specificity, with no cross-amplification with other foodborne viruses, and can detect as low as 10 copies/μL of NoV. It meets the requirements for visual rapid screening of GⅠ and GⅡ NoV. The establishment of this method provides good technical support for the rapid screening and risk monitoring of NoV, and it is of great significance for controlling NoV outbreaks and safeguarding public health.

Key words： norovirus; recombinant plasmid; enzymatic recombinase amplification; visualization; rapid detection

Cite this article

YANG Yange , WU Zhanwen , LI Tao , WANG Shuai , LI Hongna , SUN Dongmei , YUAN Fei . Two Visual and Rapid Enzymatic Recombinase Amplification Methods for GI and GII Noroviruses Detection[J]. Journal of South China University of Technology(Natural Science), 2023 , 51(12) : 140 -151 . DOI: 10.12141/j.issn.1000-565X.220529

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