Molecular Detection of Legionella spp. and their associations with Mycobacterium spp., Pseudomonas aeruginosa and amoeba hosts in a drinking water distribution system

AIMS: This study investigated waterborne opportunistic pathogens (OPs) including potential hosts, and evaluated the use of Legionella spp. for indicating microbial water quality for OPs within a full‐scale operating drinking water distribution system (DWDS). METHODS AND RESULTS: To investigate the occurrence of specific microbial pathogens within a major city DWDS we examined large volume (90 l drinking water) ultrafiltration (UF) concentrates collected from six sites between February, 2012 and June, 2013. The detection frequency and concentration estimates by qPCR were: Legionella spp. (57%/85 cell equivalent, CE l⁻¹), Mycobacterium spp. (88%/324 CE l⁻¹), Pseudomonas aeruginosa (24%/2 CE l⁻¹), Vermamoeba vermiformis (24%/2 CE l⁻¹) and Acanthamoeba spp. (42%/5 cyst equivalent, CE l⁻¹). There was no detection of the following microorganisms: human faecal indicator Bacteroides (HF183), Salmonella enterica, Campylobacter spp., Escherichia coli O157:H7, Giardia intestinalis, Cryptosporidium spp. or Naegleria fowleri. There were significant correlations between the qPCR signals of Legionella spp. and Mycobacterium spp., and their potential hosts V. vermiformis and Acanthamoeba spp. Sequencing of Legionella spp. demonstrated limited diversity, with most sequences coming from two dominant groups, of which the larger dominant group was an unidentified species. Other known species including Legionella pneumophila were detected, but at low frequency. The densities of Legionella spp. and Mycobacterium spp. were generally higher (17 and 324 folds, respectively) for distal sites relative to the entry point to the DWDS. CONCLUSIONS: Legionella spp. occurred, had significant growth and were strongly associated with free‐living amoebae (FLA) and Mycobacterium spp., suggesting that Legionella spp. could provide a useful DWDS monitoring role to indicate potential conditions for non‐faecal OPs. SIGNIFICANCE AND IMPACT OF THE STUDY: The results provide insight into microbial pathogen detection that may aid in the monitoring of microbial water quality within DWDS prior to customer exposures.