Development of a rapid isothermal amplification method for clinical detection of Pseudomonas aeruginosa

Abstract

ABSTRACT Background Pseudomonas aeruginosa (P. aeruginosa) is widely distributed in air, soil, and water, human respiratory tract, intestinal tract, and skin. It can induce bloodstream infection, urinary tract infection, gastrointestinal tract infection, respiratory tract infection, etc. Conventional bacterial isolation, culture, and identification are time-consuming, and many false negative results, which cannot meet the needs of precise clinical diagnosis，and proper treatment. This study aims to develop a rapid isothermal amplification assay of Pseudomonas aeruginosa. Methods Specific primers were designed according to the National Center for Biotechnology Information (NCBI) database based on the highly conserved sequence of Pseudomonas aeruginosa virulence factor gene lecA, and a recombinase polymerase amplification (RPA) detection method was established. The sensitivity and specificity were calculated, as well as the collection and processing of clinical samples. Results The thermostatic amplification technique for Pseudomonas aeruginosa established in this paper allows nucleic acid detection within 10 minutes without cross-amplification with other bacterial strains. 27 P. aeruginosa infections were accurately detected in 300 clinical samples. Conclusion The rapid detection system based on thermostatic amplification had shown high sensitivity and specificity in this study, indicated that this method can effectively assist clinical bacterial detection.