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

• Published in: PloS one Vol. 17; no. 12; p. e0278869
• Main Authors: Bian, Zheng, Liu, Wenbo, Jin, Junhua, Hao, Yanling, Jiang, Linshu, Xie, Yuanhong, Zhang, Hongxing
• Format: Journal Article
• Language: English
• Published: United States Public Library of Science 12.12.2022; Public Library of Science (PLoS)
• Subjects: Amplification; Bacteria; Biology and Life Sciences; Biotechnology; Cross-reactivity; Design; Dysentery; E coli; Escherichia coli; Escherichia coli - genetics; Food; Food contamination; Foodborne diseases; Health aspects; Humans; Laboratories; Medicine and Health Sciences; Methods; Nucleic Acid Amplification Techniques - methods; Nucleotidyltransferases; Optimization; Probes; Reaction time; Recombinase; Recombinases; Research and Analysis Methods; Sensitivity and Specificity; Shigella; Shigella - genetics; Thermal cycling; Virulence; Waterborne diseases; China; United Kingdom > UK; Beijing China
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0278869

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 1.29×10 2 copies/μL. The detection of food sample showed that the RPA-LFD method was also verified for the detection of actual samples.