A Rapid Detection Method for H3 Avian Influenza Viruses Based on RT–RAA
Author:
Li Jiaqi1, Cui Huan12, Zhang Yuxin1, Wang Xuejing3, Liu Huage3, Mu Yingli1, Wang Hongwei1, Chen Xiaolong1, Dong Tongchao1, Zhang Cheng12, Chen Ligong1
Affiliation:
1. College of Veterinary Medicine, Hebei Agricultural University, Baoding 071000, China 2. Hebei Key Laboratory of Analysis and Control of Zoonotic Pathogenic Microorganism, College of Veterinary Medicine, Hebei Agricultural University, Baoding 071000, China 3. The Animal Husbandry and Veterinary Institute of Hebei, Baoding, 071001, China
Abstract
The continued evolution of H3 subtype avian influenza virus (AIV)—which crosses the interspecific barrier to infect humans—and the potential risk of genetic recombination with other subtypes pose serious threats to the poultry industry and human health. Therefore, rapid and accurate detection of H3 virus is highly important for preventing its spread. In this study, a method based on real-time reverse transcription recombinase-aided isothermal amplification (RT–RAA) was successfully developed for the rapid detection of H3 AIV. Specific primers and probes were designed to target the hemagglutinin (HA) gene of H3 AIV, ensuring highly specific detection of H3 AIV without cross-reactivity with other important avian respiratory viruses. The results showed that the detection limit of the RT–RAA fluorescence reading method was 224 copies/response within the 95% confidence interval, while the detection limit of the RT–RAA visualization method was 1527 copies/response within the same confidence interval. In addition, 68 clinical samples were examined and the results were compared with those of real-time quantitative PCR (RT–qPCR). The results showed that the real-time fluorescence RT–RAA and RT–qPCR results were completely consistent, and the kappa value reached 1, indicating excellent correlation. For visual detection, the sensitivity was 91.43%, the specificity was 100%, and the kappa value was 0.91, which also indicated good correlation. In addition, the amplified products of RT–RAA can be visualized with a portable blue light instrument, which enables rapid detection of H3 AIV even in resource-constrained environments. The H3 AIV RT-RAA rapid detection method established in this study can meet the requirements of basic laboratories and provide a valuable reference for the early diagnosis of H3 AIV.
Funder
Hebei Agricultural University Hebei Province Central Guide Local Science and Technology special project Key Research and Development Program of Hebei Province Hebei Agriculture Research System Shijiazhuang Science and Technology Programme
Reference36 articles.
1. Nanomaterial-based biosensors for avian influenza virus: A new way forward;Fadhilah;Talanta,2023 2. At the centre: Influenza A virus ribonucleoproteins;Eisfeld;Nat. Rev. Microbiol.,2015 3. Lycett, S.J., Duchatel, F., and Digard, P. (2019). A brief history of bird flu. Philos. Trans. R. Soc. Lond. Ser. B Biol. Sci., 374. 4. Wang, Y., Song, T., Li, K., Jin, Y., Yue, J., Ren, H., and Liang, L. (2019). Different Subtypes of Influenza Viruses Target Different Human Proteins and Pathways Leading to Different Pathogenic Phenotypes. BioMed Res. Int., 2019. 5. Liu, J.H., Chang, C.C., Chen, C.W., Wong, L.T., and Chu, Y.W. (2019). Conservation region finding for influenza A viruses by machine learning methods of N-linked glycosylation sites and B-cell epitopes. Math. Biosci., 315.
|
|