Aims The purpose of this study was to evaluate the ISO 16266 membrane filtration culture-based method for the detection of Pseudomonas aeruginosa from cooling tower and related water samples and to compare the performance of the Pseudalert MPN method (Pseudalert (TM)/Quanti-Tray (R)) for the enumeration of P. aeruginosa with the ISO method. Methods and Results Samples were analysed by both methods and the generated data were analysed according to ISO 17994 and showed that Pseudalert resulted in significantly higher counts and a better recovery of P. aeruginosa than the ISO method for 10 and 100 ml sample volumes. Conclusions Pseudalert represents a significant improvement in the enumeration of P. aeruginosa from cooling tower water and related samples. The advantages of Pseudalert became apparent in a better performance, a more than 10-fold higher upper quantification limit when using Quanti-Tray/2000 and a shorter incubation time with no requirement for further confirmation, resulting in a faster reporting of results. Significance and Impact of the Study The (Pseudalert/Quanti-Tray) MPN method offers a more efficient and sensitive test for enumerating P. aeruginosa from cooling tower waters, thus significantly improving management of occupational safety during cleaning and maintenance activities of cooling towers.