AUTHOR=Lu Chenze , Wang Jingwen , Pan Leiming , Gu Xiuying , Lu Wenjing , Chen Di , Zhang Cen , Ye Qin , Xiao Chaogeng , Liu Pengpeng , Tang Yulong , Tang Biao , Huang Guangrong , Fang Jiehong , Jiang Han 

TITLE=Rapid detection of multiple resistance genes to last-resort antibiotics in Enterobacteriaceae pathogens by recombinase polymerase amplification combined with lateral flow dipstick

JOURNAL=Frontiers in Microbiology

VOLUME=13

YEAR=2023

URL=https://www.frontiersin.org/journals/microbiology/articles/10.3389/fmicb.2022.1062577

DOI=10.3389/fmicb.2022.1062577

ISSN=1664-302X

ABSTRACT=<p>The worrying emergence of multiple resistance genes to last-resort antibiotics in food animals and human populations throughout the food chain and relevant environments has been increasingly reported worldwide. <italic>Enterobacteriaceae</italic> pathogens are considered the most common reservoirs of such antibiotic resistance genes (ARGs). Thus, a rapid, efficient and accurate detection method to simultaneously screen and monitor such ARGs in <italic>Enterobacteriaceae</italic> pathogens has become an urgent need. Our study developed a recombinase polymerase amplification (RPA) assay combined with a lateral flow dipstick (LFD) for simultaneously detecting predominant resistance genes to last-resort antibiotics of <italic>Enterobacteriaceae</italic> pathogens, including <italic>mcr-1</italic>, <italic>bla<sub>NDM-1</sub></italic> and <italic>tet(X4)</italic>. It is allowed to complete the entire process, including crude DNA extraction, amplification as well as reading, within 40 min at 37°C, and the detection limit is 10<sup>1</sup> copies/μl for <italic>mcr-1</italic>, <italic>bla<sub>NDM-1</sub></italic> and <italic>tet(X4)</italic>. Sensitivity analysis showed obvious association of color signals with the template concentrations of <italic>mcr-1</italic>, <italic>bla<sub>NDM-1</sub></italic> and <italic>tet(X4)</italic> genes in <italic>Enterobacteriaceae</italic> pathogens using a test strip reader (<italic>R</italic><sup>2</sup> = 0.9881, <italic>R</italic><sup>2</sup> = 0.9745, and <italic>R</italic><sup>2</sup> = 0.9807, respectively), allowing for quantitative detection using multiplex RPA-LFD assays. Therefore, the RPA-LFD assay can suitably help to detect multiple resistance genes to last-resort antibiotics in foodborne pathogens and has potential applications in the field.</p>