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Development of a reverse transcription recombinase-aided amplification assay for detection of Getah virus.

ABSTRACT: GETV, an arbo-borne zoonotic virus of the genus Alphavirus, which causes diarrhea and reproduction disorders in swine, lead to serious economic losses to the swine industry in China. At present, the existing methods for GETV detection are time-consuming and low sensitivity, so, a rapid, accurate and sensitive GETV detection method is urgently needed. In this study, a fluorescent reverse transcription recombinase-assisted amplification method (RT-RAA) was successfully established for the rapid detection of GETV. The sensitivity of this method to GETV was 8 copies/reaction and 20 TCID₅₀/reaction. No cross-reaction with other viruses. A total of 118 samples were prepared for GETV detection using fluorescent RT-RAA and SYBR Green I RT-qPCR, the coincidence rate of the two methods was 100%. The results suggest that the RT-RAA method is rapid, sensitive and specific for GETV detection and can be applied in the clinical.

SUBMITTER: Nie M

PROVIDER: S-EPMC8501081 | biostudies-literature | 2021 Oct

REPOSITORIES: biostudies-literature

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Development of a reverse transcription recombinase-aided amplification assay for detection of Getah virus.

Nie Mincai M Deng Huidan H Zhou Yuancheng Y Sun Xiangang X Huang Yao Y Zhu Ling L Xu Zhiwen Z

Scientific reports 20211008 1

GETV, an arbo-borne zoonotic virus of the genus Alphavirus, which causes diarrhea and reproduction disorders in swine, lead to serious economic losses to the swine industry in China. At present, the existing methods for GETV detection are time-consuming and low sensitivity, so, a rapid, accurate and sensitive GETV detection method is urgently needed. In this study, a fluorescent reverse transcription recombinase-assisted amplification method (RT-RAA) was successfully established for the rapid de ...[more]

PMID: 34625631

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Project description:Newcastle disease is an acute and highly contagious disease of poultry caused by Newcastle disease virus infection, which does great harm to the poultry industry all over the world. To diagnose the disease simply and quickly, 2 detection methods were established based on reverse transcription recombinase-aided amplification (RT-RAA) technology. One is reverse transcription recombinase-aided amplification-lateral flow dipstick (RT-RAA-LFD) that is to combine RT-RAA with lateral flow dipstick; the other is real-time fluorescence-based reverse transcription recombinase-aided amplification (RF-RT-RAA) that is the combination of RT-RAA and exo probe. In this study, the reaction conditions such as reaction temperature and reaction time of the 2 methods were optimized, and their specificity and sensitivity were tested. The results showed that the RT-RAA-LFD method could be used to complete reaction within 23 min, and its lowest detectable limit was 102 copies/?L, 10 times higher than that of the conventional PCR method (103 copies/?L); the RF-RT-RAA method could be used to complete reaction within 26 min, and its lowest detectable limit was 10 copies/?L, 100 times higher than that of conventional PCR method (103 copies/?L), and it was as sensitive as real-time fluorescence-based quantitative PCR (10 copies/?L). The 2 methods had no cross reaction to the nucleic acid of other avian pathogens and showed good specificity. A total of 86 clinical samples suspected of the Newcastle disease virus were tested by conventional PCR, real-time fluorescence-based quantitative PCR, RT-RAA-LFD, and RF-RT-RAA. Based on the commonly used conventional PCR method, the other 3 detection methods had a coincidence rate of higher than 93%. In summary, RT-RAA-LFD and RF-RT-RAA had high specificity, sensitivity, and efficiency, which were suitable for clinical and laboratory diagnosis, respectively, and provided technical support for the prevention and control of Newcastle disease.

Dataset Information

Development of a reverse transcription recombinase-aided amplification assay for detection of Getah virus.

Publications

Development of a reverse transcription recombinase-aided amplification assay for detection of Getah virus.

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