Amoebae in domestic water systems: resistance to disinfection treatments and implication in Legionella persistence

Abstract

Aims: Monitoring of microbial changes during and after application of various disinfection treatments in a model domestic water system. Methods and Results: A pilot‐scale domestic water system consisting of seven galvanized steel re‐circulation loops and copper dead legs was constructed. Culture techniques, confocal laser scanning microscopy after fluorescent in situ hybridization and viability staining with the BacLight® LIVE/DEAD kit were used for planktonic and biofilm flora monitoring. Before starting the treatments, the system was highly contaminated with Legionella pneumophila and biofilm populations mainly consisted of β‐proteobacteria. In the water and the biofilm of the loops, continuous application of chlorine dioxide (0·5 mg l−1), or chlorine (2·5 mg l−1) were very effective in reducing the microbial flora, including L. pneumophila. Heterotrophic bacteria, although strongly reduced, were still detectable after ozone application (0·5 mg l−1), whereas with monochloramine (0·5 mg l−1) and copper–silver ionization (0·8/0·02 mg l−1), the contamination remained significantly higher. Monochloramine and copper–silver did not remove the biofilm. During copper–silver application, Legionella re‐growth was observed. Only chlorine dioxide led to detectable effects in the dead leg. Amoebae could not be eliminated, and after interrupting the treatments, L. pneumophila quickly recovered their initial levels, in all cases. Conclusions: Chlorine dioxide, applied as a continuous treatment, was identified in this study as the most efficient for controlling L. pneumophila in a domestic water system. Chlorine dioxide showed a longer residual activity, leading to improved performance in the dead leg. Amoebae resisted to all the treatments applied and probably acted as reservoirs for L. pneumophila, allowing a quick re‐colonization of the system once the treatments were interrupted. Significance and Impact of the Study: Control of microbial contamination requires maintenance of a constant disinfectant residual throughout the water system. Treatment strategies targeting free‐living amoebae should lead to improved control of L. pneumophila. Such treatment strategies still have to be investigated.

Transcriptional profiling of Legionella pneumophila biofilm cells and the influence of iron on biofilm formation

“…Unfortunately, these treatments generally do not lead to a total eradication of the bacterium, and recolonization occurs as soon as the treatments are interrupted (Thomas et al, 2004). Resistance of L. pneumophila to disinfection is due not only to its capacity to enter amoebae, where bacteria could be protected from biocides, but also to its association with biofilms (Abu Kwaik et al, 1998;Thomas et al, 2004;Saby et al, 2005). However, only very little information regarding the physiological state and gene expression of L. pneumophila within biofilms is currently available.…”

Cellular Response of the Amoeba Acanthamoeba castellanii to Chlorine, Chlorine Dioxide, and Monochloramine Treatments

“…Intracellular L. pneumophila is more resistant to biocides (3,4,14). Consequently, free-living amoebae, such as Acanthamoeba, have been recently described as important targets in water treatment (19,30). Common biocides used in water treatment (i.e., chlorine, monochloramine, and chlorine dioxide) were assessed against pathogenic protozoa such as Giardia, Cryptosporidium, and to a lesser extent Entamoeba histolytica (20).…”

Knowledge to Predict Pathogens: Legionella pneumophila Lifecycle Critical Review Part I Uptake into Host Cells

“…As such, it is important that treatments can penetrate the extracellular polymeric substance (EPS) matrix of the biofilm and persist in large premise plumbing systems [160,161]. Chlorine dioxide was shown to be most effective in reducing L. pneumophila levels in biofilms in copper piping, compared to chlorine, monochloramine, electro-chlorination, ozone, and copper-silver ionization, due to its longer residual activity and its ability to penetrate the biofilm [162]. However, the amoeba in the biofilm were resistant to all forms of treatment and L. pneumophila was able to regrow after short periods of non-treatment [162,163].…”