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Detection method for reverse transcription recombinase-aided amplification of avian influenza virus subtypes H5, H7, and H9

文献类型： 外文期刊

作者： Zhang, Zongshu 1 ; Zhang, Zichuang 2 ; Wang, Chunguang 1 ; Zhai, Xianghe 1 ; Wang, Wenjing 1 ; Chen, Xi 1 ; Zhang, Tie 1 ;

作者机构： 1.Hebei Agr Univ, Coll Vet Med, Baoding, Peoples R China

2.Chinese Acad Agr Sci, Inst Special Anim & Plant Sci, Changchun, Peoples R China

关键词： Avian influenza virus; Reverse transcription recombinase-aided amplification; Lateral flow dipstick; Real-time fluorescence

期刊名称：BMC VETERINARY RESEARCH （ 影响因子：2.6； 五年影响因子：2.9 ）

ISSN：

年卷期： 2024 年 20 卷 1 期

页码：

收录情况： SCI

摘要： Background Avian influenza virus (AIV) not only causes huge economic losses to the poultry industry, but also threatens human health. Reverse transcription recombinase-aided amplification (RT-RAA) is a novel isothermal nucleic acid amplification technology. This study aimed to improve the detection efficiency of H5, H7, and H9 subtypes of AIV and detect the disease in time. This study established RT-RAA-LFD and real-time fluorescence RT-RAA (RF-RT-RAA) detection methods, which combined RT-RAA with lateral flow dipstick (LFD) and exo probe respectively, while primers and probes were designed based on the reaction principle of RT-RAA.Results The results showed that RT-RAA-LFD could specifically amplify H5, H7, and H9 subtypes of AIV at 37 degrees C, 18 min, 39 degrees C, 20 min, and 38 degrees C, 18 min, respectively. The sensitivity of all three subtypes for RT-RAA-LFD was 102 copies/mu L, which was 10 similar to 100 times higher than that of reverse transcription polymerase chain reaction (RT-PCR) agarose electrophoresis method. RF-RT-RAA could specifically amplify H5, H7, and H9 subtypes of AIV at 40 degrees C, 20 min, 38 degrees C, 16 min, and 39 degrees C, 17 min, respectively. The sensitivity of all three subtypes for RF-RT-RAA was 101 copies/mu L, which was consistent with the results of real-time fluorescence quantification RT-PCR, and 100 similar to 1000 times higher than that of RT-PCR-agarose electrophoresis method. The total coincidence rate of the two methods and RT-PCR-agarose electrophoresis in the detection of clinical samples was higher than 95%.Conclusions RT-RAA-LFD and RF-RT-RAA were successfully established in this experiment, with quick response, simple operation, strong specificity, high sensitivity, good repeatability, and stability. They are suitable for the early and rapid diagnosis of Avian influenza and they have positive significance for the prevention, control of the disease, and public health safety.

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