Comparison of biofilm cell quantification methods for drinking water distribution systems.
Drinking water quality typically degrades after treatment during conveyance through the distribution system. Potential causes include biofilm growth in distribution pipes which may result in pathogen retention, inhibited disinfectant diffusion, and proliferation of bad tastes and odors. However, there is no standard method for direct measurement of biofilms or quantification of biofilm cells in drinking water distribution systems. Three methods are compared here for quantification of biofilm cells grown in pipe loops samplers: biofilm heterotrophic plate count (HPC), biofilm biovolume by confocal laser scanning microscopy (CLSM) and biofilm total cell count by flow cytometry (FCM) paired with Syto 9. Both biofilm biovolume by CLSM and biofilm total cell count by FCM were evaluated for quantification of the whole biofilms (including non-viable cells and viable but not culturable cells). Signal-to-background ratios and overall performance of biofilm biovolume by CLSM and biofilm total cell count by FCM were found to vary with the pipe material. Biofilm total cell count by FCM had a low signal-to-background ratio on all materials, indicating that further development is recommended before application in drinking water environments. Biofilm biovolume by CLSM showed the highest signal-to-background ratio for cement and cast iron, which suggests promise for wider application in full-scale systems. Biofilm biovolume by CLSM and Syto 9 staining allowed in-situ biofilm cell quantification thus elimination variable associated with cell detachment for quantification but had limitations associated with non-specific staining of cement and, to a lesser degree, auto-fluorescence of both cement and polyvinyl chloride materials. Due to variability in results obtained from each method, multiple methods are recommended to assess biofilm growth in drinking water distribution systems. Of the methods investigated here, HPC and CLSM and recommended for further development towards application in full-scale systems. HPC is a sample and widely applied method that quantifies viable culturable cells. CLSM analysis allows the elimination of experimental variables associated with cell detachment and affords the opportunity to evaluate biofilm components such as extracellular polymeric substances through the addition of specific probes. These two methods can be applied together to assess biofilms known to degrade treated water quality during conveyance in full-scale drinking water treatment systems. The significance of improved biofilm assessment methods for drinking water distribution systems lies in advancing understanding of biofilm growth and control mechanisms that may lead to improved water quality during conveyance and at the tap for greater public health protection.
['Waller|Sharon A|SA|', 'Packman|Aaron I|AI|', 'Hausner|Martina|M|']
D001431:Bacteriological Techniques D018441:Biofilms D015169:Colony Count, Microbial D060766:Drinking Water D005434:Flow Cytometry D008422:Materials Testing D050296:Microbial Viability D018613:Microscopy, Confocal D014871:Water Microbiology D060753:Water Quality D014881:Water Supply
Biofilm drinking water distribution confocal laser scanning microscope flow cytometry