AUTHOR=Wei Lijian , Luo Shihua , Zhou Weijie , Ren Baoyan , Li Miao , Liang Lina , Li Xuebin , Wei Guijiang 

TITLE=Rapid detection of Pseudomonas aeruginosa by glycerol one-pot RAA/CRISPR-Cas12a method

JOURNAL=Frontiers in Chemistry

VOLUME=Volume 13 - 2025

YEAR=2025

URL=https://www.frontiersin.org/journals/chemistry/articles/10.3389/fchem.2025.1654270

DOI=10.3389/fchem.2025.1654270

ISSN=2296-2646

ABSTRACT=Pseudomonas aeruginosa (PA), an opportunistic pathogen commonly responsible for hospital-acquired infections, poses significant threats to human health. To enable rapid and reliable PA detection while effectively mitigating aerosol contamination risks inherent in conventional methods. We developed a glycerol one-pot Recombinase-aided Amplification (RAA)/CRISPR-Cas12a method. Four result reading methods were established: Fluorescence Detection (FD), Blue Light Irradiation Detection (BLD), and Ultraviolet Irradiation Detection (UID), as well as Lateral Flow Chromatography Strip (LFS). The glycerol one-pot RAA-CRISPR/Cas12a method demonstrated high specificity and sensitivity in detecting the PA-specific lasB gene. The detection limit reached 1.20 × 10-4 ng/μL (fluorescence-based) and 1.20 × 10−3 ng/μL (LFS-based). In validation against 64 clinical isolates, compared to conventional PCR, the assay achieved 100% sensitivity, specificity, and accuracy in lasB detection. In conclusion, the glycerol one-pot RAA/CRISPR-Cas12a method provides a rapid, sensitive, and straightforward platform, providing a promising approach for clinical diagnosis of PA and environmental surveillance applications.