| This work explores the molecular, cellular and ecological intricacies of the bacterial response to the drinking water disinfectant monochloramine using Escherichia coli and Mycobacterium avium as model bacteria. It was found that exposure of M. avium to a sub-lethal dose of monochloramine resulted in rapid cell wall permeabilization and intracellular thiol oxidation. The oxidative stress (OxyR) response was induced very strongly and rapidly and many virulence-associated genes also were upregulated, though whether this response increases M. avium virulence to humans must be further studied. The role of environmental conditions in inducing monochloramine resistance of E. coli also was explored. Growth of E. coli in either biofilm mode or at a suboptimal temperature of 20°C increased its resistance to monochloramine. Comparative transcriptional profiling of cells grown in biofilm mode, at 20°C, or after monochloramine exposure was performed in order to define a "drinking water stressome", which was characterized by widespread metabolic inhibition, regulation of redox-active genes, and induction of osmotic and cell envelope stress responses. Overall, there appears to be extensive overlap between response to monochloramine and to other stresses, such as general oxidative stress and osmotic stress. Finally, the relevance of the interaction of M. avium and Acanthamoeba was explored as a possible survival mechanism in drinking water treatment and distribution. M. avium formed stable infections within a range of Acanthamoeba strains, maintaining its viability for at least 28 days. Acanthamoeba-associated M. avium was much more resistant to monochloramine than M. avium alone and inactivation kinetics of intracellular M. avium exposed to monochloramine closely matched the inactivation kinetics of Acanthamoeba castellanii Neff, suggesting that acanthamoebal inactivation may be a useful surrogate for intracellular M. avium inactivation. Overall, this research underscores the importance of biological processes in drinking water treatment and distribution, characterizing the biological complexity of bacterial response to monochloramine, the complexities emerging from response to conditions typically found in drinking water distribution, and the interactions of bacterial pathogens with free-living amoebae. |
