Comparison of biofilm cell quantification methods for drinking water distribution systems

Area of Science:

• Environmental Science
• Microbiology
• Water Quality Engineering

Background:

• Drinking water quality can decline in distribution systems due to biofilm formation.
• Biofilms contribute to pathogen retention, reduced disinfectant efficacy, and undesirable taste/odor.
• Standardized methods for direct biofilm measurement in drinking water systems are lacking.

Purpose of the Study:

• To compare three methods for quantifying biofilm cells in drinking water distribution systems.
• To evaluate the performance of heterotrophic plate count (HPC), confocal laser scanning microscopy (CLSM), and flow cytometry (FCM).
• To identify suitable methods for in-situ biofilm assessment in water distribution networks.

Main Methods:

• Biofilm samples were grown in pipe loop samplers.
• Quantification methods included HPC, CLSM (biovolume), and FCM (total cell count with Syto 9).
• CLSM and FCM were assessed for their ability to quantify total biofilm, including non-viable and viable but not culturable cells.

Main Results:

• CLSM and FCM performance varied with pipe material; FCM showed a low signal-to-background ratio across all materials.
• CLSM demonstrated high signal-to-background ratios for cement and cast iron pipes.
• CLSM and Syto 9 staining allowed in-situ quantification but faced challenges with non-specific staining and auto-fluorescence on certain materials.

Conclusions:

• No single method is sufficient; multiple approaches are recommended for comprehensive biofilm assessment.
• HPC and CLSM are promising for further development and application in full-scale drinking water systems.
• Combined use of HPC and CLSM can enhance understanding and control of biofilms impacting water quality and public health.