Development and evaluation of loop-mediated isothermal amplification-based kit for rapid detection of Pseudomonas aeruginosa in packaged drinking water

Abstract:

[Background] Pseudomonas aeruginosa is a major pathogen that needed to be intensively monitored in the production activities by packaged drinking water enterprises. With continuous development of the molecular detection technology, it is very important to develop simple and efficient detection products to detect P. aeruginosa in packaged drinking water. [Objective] To develop loop-mediated isothermal amplification (LAMP) based kit for rapid detecting P. aeruginosa and evaluate its detecting efficacy in packaged drinking water. [Methods] We optimized the LAMP reaction system and confirmed the components of the kit. The main reaction reagents were made into powder pattern by freeze drying technology. Then the specificity, sensitivity, repeatability, shelf life and other performance indexes of the kit were evaluated. [Results] All P. aeruginosa standard strains and isolated strains were detected positive, while all non-P. aeruginosa standard strains and isolated strains were detected negative, showing that the detection had no cross reaction. The minimum test limit of the kit was calculated to 18 CFU/mL, and detection specificity, accuracy and sensitivity of the kit were preeminent. The detection result was highly consistent with that obtained by conventional microbial detection method. The intra-batch and inter-batch testing repetition rates were all 100%. The kit could be preserved for more than 12 months at 4 °C, and its efficacy was mostly maintained for more than 72 h at 42 °C. [Conclusion] The kit has good performance with stable and reliable test results which make it applicable for rapid detection of P. aeruginosa in packaged drinking water.