Legionella spp. survival after different disinfection procedures: Comparison between conventional culture, qPCR and EMA–-qPCR

The development of rapid and sensitive methods for the detection and quantification of Legionella viable cells is essential for monitoring water quality and preventing legionellosis. The aim of this study was to verify the applicability of a quantitative PCR (qPCR) method used in combination with ethidium monoazide (EMA) to the quantification of Legionella spp. in samples collected from swimming pools, water recirculation systems and hot water systems in two fitness clubs. This molecular technique (EMA–-qPCR) allows the amplification of target DNA from culturable and viable cells, but prevents the amplification of DNA from non-viable cells. The effectiveness of this new method able to detect alive legionellae was also compared with conventional qPCR and culture method. Our results confirm that EMA–-qPCR allows to discriminate the non-viable cells from those viable and that it is particularly indicated for monitoring the effectiveness of thermal treatments for the Legionella contamination control in water environments, also providing information about the presence of Viable But Non-Culturable (VBNC) cells. Other Gram-negative bacteria typically associated with biofilm were identified in samples taken from swimming pools and balance tanks, suggesting that also the presence of biofilm should be monitored for a more general view of water contamination.