Development of a recombinase polymerase amplification assay with lateral flow dipstick (RPA-LFD) for rapid detection of Shigella spp. and enteroinvasive Escherichia coli

开发一种利用侧流试纸 (RPA-LFD) 的重组酶聚合酶扩增检测法,用于快速检测志贺氏菌和肠侵袭性大肠杆菌

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作者:Zheng Bian, Wenbo Liu, Junhua Jin, Yanling Hao, Linshu Jiang, Yuanhong Xie, Hongxing Zhang

Abstract

Shigella spp. and enteroinvasive Escherichia coli (EIEC) are widely distributed and can cause serious food-borne diseases for humans such as dysentery. Therefore, an efficient detection platform is needed to detect Shigella and EIEC quickly and sensitively. In this study, a method called recombinase polymerase amplification combined with lateral flow dipstick (RPA-LFD) was developed for rapid detection of Shigella and EIEC. RPA primers and LFD detection probes were designed for their shared virulence gene ipaH. Primers and probes were screened, and the primer concentration, and reaction time and temperature were optimized. According to the optimization results, the RPA reaction should be performed at 39°C, and when combined with LFD, it takes less than 25 min for detection with the naked eye. The developed RPA-LFD method specifically targets gene ipaH and has no cross-reactivity with other common food-borne pathogens. In addition, the minimum detection limit of RPA-LFD is 1.29×102 copies/μL. The detection of food sample showed that the RPA-LFD method was also verified for the detection of actual samples.

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