ISO 16266-2:2018

This document specifies a method for the enumeration of Pseudomonas aeruginosa in water. The method is based on the growth of target organisms in a liquid medium and calculation of the most probable number (MPN) of organisms by reference to MPN tables.
This document is applicable to a range of types of water. For example, hospital waters, drinking water and non-carbonated bottled waters intended for human consumption, groundwater, swimming pool and spa pool waters including those containing high background counts of heterotrophic bacteria.
This document does not apply to carbonated bottled waters, flavoured bottle waters, cooling tower waters or marine waters, for which the method has not been validated. These waters are, therefore, outside the scope of this document. Laboratories can employ the method presented in this document for these matrices by undertaking appropriate validation of performance of this method prior to use.
The test is based on a bacterial enzyme detection technology that signals the presence of P. aeruginosa through the hydrolysis of a 7-amino-4-methylcoumarin aminopeptidase substrate present in a special reagent. P. aeruginosa cells rapidly grow and reproduce using the rich supply of amino acids, vitamins and other nutrients present in the reagent. Actively growing strains of P. aeruginosa have an enzyme that cleaves the 7-amido-coumarin aminopeptidase substrate releasing a product which fluoresces under ultraviolet (UV) light. The test described in this document provides a confirmed result within 24 h with no requirement for further confirmation of positive wells.