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

Zheng Bian; Wenbo Liu; Junhua Jin; Yanling Hao; Linshu Jiang; Yuanhong Xie; Hongxing Zhang

doi:10.1371/journal.pone.0278869

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

PLoS One. 2022 Dec 12;17(12):e0278869. doi: 10.1371/journal.pone.0278869. eCollection 2022.

Authors

Zheng Bian¹, Wenbo Liu¹, Junhua Jin¹, Yanling Hao², Linshu Jiang³, Yuanhong Xie¹, Hongxing Zhang¹

Affiliations

¹ Beijing Laboratory of Food Quality and Safety, Beijing Key Laboratory of Agricultural Product Detection and Control of Spoilage Organisms and Pesticide Residue, Beijing Engineering Technology Research Center of Food Safety Immune Rapid Detection, College of Food Science and Engineering, Beijing University of Agriculture, Beijing, China.
² Department of Nutrition and Health, Key Laboratory of Functional Dairy, Co-constructed by Ministry of Education and Beijing Government, China Agricultural University, Beijing, China.
³ Animal Science and Technology College, Beijing University of Agriculture, Beijing, China.

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.

Copyright: © 2022 Bian et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Escherichia coli / genetics
Humans
Nucleic Acid Amplification Techniques / methods
Nucleotidyltransferases
Recombinases*
Sensitivity and Specificity
Shigella* / genetics

Substances

Recombinases
Nucleotidyltransferases

Grants and funding

We thank the Research project of Beijing Municipal Commission of Education, this work was supported by the Research project of Beijing Municipal Commission of Education (KM201810020016). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.