Establishment of LAMP Rapid Detection Method for Pseudomonas aeruginosa and Verification of Droplet Digital PCR

Objective:A rapid method for detecting Pseudomonas aeruginosa was establish by loop-mediated isothermal amplification,and verified by microdroplet digital PCR assay. Methods:Using the rpod gene of Pseudomonas aeruginosa as the target gene,a rapid detection method for Pseudomonas aeruginosa in juice drink was established by design of primers. For the reliability,specificity and sensitivity of the method was verified by the droplet digital PCR. Results:The established LAMP rapid detection method had obviously color change. In the detection of droplets digital PCR,the concentration of Pseudomonas aeruginosa suspension was used as abscissa,and the number of copy amplified by digital PCR was used as ordinate. The standard curve was y=0.07497x-9.3516.The linear relationship was good,R²=0.9999. The detection range of Pseudomonas aeruginosa was 1.5×10²~1.5×10⁵ CFU/mL The method had good specificity,high sensitivity and accuracy. The detection limit of standard Pseudomonas aeruginosa was 1 ng/μL. The standard deviation were 0.57,0.71,4.24,14.8,and the RSD value between 0.66% to 22.8%. 128 samples of juice drinks were detected,126 samples were not colored,and 2 samples were colored. The values were determined by ddPCR method,and the concentrations were 1.64×10⁴ and 2.29×10² CFU/mL,respectively.