Multiple-probe-assisted DNA capture and amplification for high-throughput African swine fever virus detection

文献类型： 外文期刊

第一作者： Wang, Huicong

作者： Wang, Huicong;Pian, Hongru;Fan, Lihua;Zheng, Zhi;Wang, Huicong;Pian, Hongru;Fan, Lihua;Zheng, Zhi;Li, Jian;Yang, Jifei

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关键词： African swine fever; DNA capture; Laboratory diagnosis; High-throughput; Without DNA extraction

期刊名称：APPLIED MICROBIOLOGY AND BIOTECHNOLOGY （ 影响因子：5.0； 五年影响因子：5.2 ）

ISSN： 0175-7598

年卷期： 2023 年 107 卷 2-3 期

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收录情况： SCI

摘要： African swine fever (ASF) is one of the most devastating infectious diseases affecting domestic pigs and wild boar. The grave socio-economic impact of African swine fever infection at a global level makes large-scale rapid and robust diagnosis a critical step towards effective control. Here, we describe multiple-probe-assisted DNA capture and amplification technology (MADCAT) - a novel, sensitive, simple, and high-throughput method for detecting ASFV directly from whole blood or other complex matrices. Through a unique DNA capture approach which specifically captures the target DNA onto 96-well plate for subsequent amplification, MADCAT abandons the complicated extraction protocol and achieves ultrafast and high-throughput detection. The sample-to-result time for 96 samples is about 90 min, as compared with the 3-4 h time of the conventional real-time qPCR method. The limit of detection (LOD) of MADCAT is 0.5 copies/mu L blood and is 5 times more sensitive than an extraction-based qPCR assay when testing serially diluted whole blood samples. The assay is 100% specific against other common swine pathogens. In the clinical diagnosis of 96 field samples, all 22 positive samples were correctly identified with lower Ct values than extraction-based qPCR, confirming its high diagnostic sensitivity (100%). Owing to its high-throughput, specific high sensitivity, and direct detection features, MADCAT shows great potential for use in large-scale ASFV surveillance and monitoring for effective disease control.

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