Molecular toolbox for the monitoring and control of water distribution biofilms

The water distribution system represents the final barrier to the degradation of treated water prior to consumption. Maintaining the integrity of these systems therefore is essential for public health. Current strategies for monitoring bacteriological water quality within distribution systems are dependent upon culture-based methods, which are affected by many factors, including conventional disinfection strategies (chlorine) that have been shown to impact the culturability of bacteria on media. Improved methods to assess the bacteriological loads and responses to treatment are needed. This thesis investigated the development and application of alternate means of analysis of water quality analytes, enabling a toolbox approach for bacteriological water quality monitoring. Tools used in this study include biomass concentration using DNA quantification, bacterial community characterisation using 16S rDNA polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE), targeted population assessment using quantitative PCR (QPCR) and in situ phylogenetic abundance using fluorescence in situ hybridisation (FISH). Conventional methods such as heterotrophic plate counts were also included. Results show that chlorine does not eradicate bacteria by the proportions generally reported using culture-reliant methods. Within both drinking and recycled water distribution systems, bacteria belonging to Enterobacteriaceae and species of Pseudomonas were detected in water containing 2 ppm free chlorine concentrations. The persistence of such bacteria in conditions prevalent within distribution systems necessitates the revision of techniques of analysis as well as improved treatment and disinfection management strategies.Furthermore, current strategies for water quality monitoring are reliant on the assessment of planktonic bacterial populations, and do not consider attached (biofilm) populations. Molecular methods of analysis showed up to 5-log more bacteria are present in biofilms compared to planktonic sources. Thus, improved methods for water quality maintenance in distribution systems need to consider the contributions of biofilms. This thesis also studied the effects of a nitric oxide (NO) donor, as NO has been shown to encourage detachment of bacterial cells from the biofilm which then enter the planktonic phase of the life cycle. The molecular toolbox analysis approach showed that effects are variable on the diverse bacterial populations present in distribution systems, and some bacteria show heightened resistance compared to other populations. Ongoing work targeting biofilms in distribution systems is required to improve conventional practices of water quality assurance in distribution systems