AS 3896:2017 Waters – Examination for Legionella spp. including Legionella pneumophila
8.5.2 Procedure
Selected colonies shall be subcultured onto plates of BCYE and either BCYE-Cys or BA . and incubated in a humid atmosphere at 36°+2°C for up to 3 days. Subculturing shall be carried out on the first examination if colonies are seen that are presumptive Legionella spp. Additional subculturing shall be carried out if new colony types that are presumptive legionellae appear on subsequent examinations.
8.5.3 Interpretation
Microorganisms that grow on BCYE but not on BCYE-Cys or BA, are Legionella spp. These microorganisms may be further tested in accordance with Clause 8.6. . Microorganisms that grow on BCYE-Cys or BA are not Legionella spp.
NOTE: Care should be taken to ensure that Legionella spp. are not missed on the occasions when a contaminant organism is inadvertently transferred with the presumptive legionellae onto the BCYE-Cys or BA. This may be indicated by mixed growth on the BCYE plates.
When all colonies of a given colony type fail to confirm, and the colony type is present on the other retained plates, further colonies need to be selected for confirmation. Colonies are selected from all other retained plates with that colony type according to Clause 8.4(c) and taken through to confirmation. The highest confirmed count obtained from a single plate of the retained set, shall be used for reporting purposes.
NOTE: This method uses multiple treatments and different isolation media with different performance characteristics. The different combinations of treatment and isolation media would be expected to have differing recoveries for different species and strains of Legionella in any one sample. Therefore， the highest count is considered to be the result of the optimum medium/treatment combination.
8.6 Legionella speciation and typing
Speciation and typing tests for Legionella that have been validated to be fit for purpose may be performed if required. Commercial systems shall be used in accordance with manufacturers’instructions.