Establishment and application of recombinase polymerase amplification combined with a lateral flow dipstick for the detection of mcr-1 in uncultured clinical samples

• Published in: International journal of antimicrobial agents 2024-05, Vol.63 (5), p.107140, Article 107140
• Main Authors: Feng, Jun, Xu, Zhen, Zhuang, Yuan, Luo, Jiayuan, Chen, Yong, Wu, Yitong, Fei, Jiayi, Liu, Mingxiang, Xia, Jiahui, Zhang, Jing, Liu, Meihua, Xie, Xiaohong, Yuan, Zhengan, Chen, Min
• Format: Article
• Language: English
• Subjects: Colistin; Lateral flow dipstick; mcr-1; Recombinase polymerase amplification; Uncultured
• ISSN: 0924-8579; 1872-7913; 1872-7913
• DOI: 10.1016/j.ijantimicag.2024.107140

•Clinical samples carrying mcr-1 gene can be detected without the need for bacterial culture.•The mcr-1 gene process, from DNA extraction to detection, completed within 20 min.•The mcr-1 gene can be detected via RPA plus LFD technology as low as 10 copies/μL.•User-friendly and suitable for field screening. The rapid dissemination of the mcr-1 gene via plasmid-mediated transfer has raised concerns regarding the efficacy of colistin as a last-resort treatment for multidrug-resistant Gram-negative bacterial infections. Current mcr-1 gene detection methods mainly focus on cultured bacteria, which is a complex and time-consuming process requiring skilled personnel, making it unsuitable for field analysis. A rapid detection technique combining recombinase polymerase amplification with a lateral flow dipstick targeting uncultured clinical samples was developed. This new method targeting the mcr-1 gene region (23 232–23 642 bp, no. KP347127.1) achieved a low detection limit of 10 copies/μL. The whole process was carried out with high specificity and was completed within 20 min. The evaluation assay was conducted using 45 human faecal samples; 16 strains yielded a 98% accuracy, closely matching antimicrobial susceptibility outcomes. The novel method integrates nucleic acid extraction, isothermal amplification, and a test assay, suggesting the potential for timely colistin resistance surveillance in frontline disease control and healthcare settings, supporting future prevention and clinical standardization efforts. [Display omitted]