Quantitative Analysis of Biofilm Susceptibility against a Model Disinfectant

In water distribution systems, biofilm survival has been reported despite the regulatory presence of residual disinfectants. Natural organic matter (NOM), which is ubiquitous in drinking water systems, contributes to biofilm growth as a carbon source as well as increasing disinfectant demand. In this study, strains from an opportunistic pathogen, Pseudomonas aeruginosa (both wild type and mutant strains) with different extracellular polymeric substance (EPS) secretion capabilities were used to cultivate single species biofilms. Biofilms were grown in a continuous flow system under low nutrient condition simulating drinking water distribution system. After chlorine disinfection, biofilm was visualized using a confocal laser scanning microscope (CLSM) followed by image analysis to quantify biofilm EPS content and viability spatial distribution. The survival rate of detached cells from PAO1 biofilm was analyzed by flow cytometry to differentiate live, dead and membrane compromised cells considering the presence or absence of disinfectant consumption substrate (NOM). Both biofilm and detachment viability were confirmed by plate count method.The results show that the amount of EPS is positively related to biofilm viability. Resistance was significantly enhanced in EPS overproduction biofilm compared to EPS deficient strains. Due to chemical reaction between NOM and residual disinfectants, the presence of NOM improved detached biofilm resistance to chlorine residuals.