Flow cytometry is increasingly employed by drinking water providers. Its use with appropriate fluorescent stains allows the distinction between intact and membrane-damaged bacteria, which makes it ideally suited for assessment of disinfection efficiency. In contrast to plate counting, the technology allows the visualization of the gradual loss of membrane integrity. Although this sensitivity per se is very positive, it creates the problem of how this detailed viability information compares with binary plate counts where a colony is either formed or not. Guidelines are therefore needed to facilitate interpretation of flow cytometry results and to determine a degree of membrane damage where bacteria can be considered 'dead'. In this study we subjected Escherichia coli and environmental microorganisms in real water to increasing chlorine concentrations. Resulting flow cytometric patterns after membrane integrity staining were compared with culturability and in part with redox activity. For laboratory-grown bacteria, culturability was lost at lower disinfectant concentrations than membrane integrity making the latter a conservative viability parameter. No recovery from chlorine was observed for four days. For real water, loss of membrane integrity had to be much more substantial to completely suppress colony formation, probably due to the heterogenic composition of the natural microbial community with different members having different susceptibilities to the disinfectant.

Please refer to any applicable publisher terms of use. Highlights  Different turbidities have different effects on chlorine disinfection  Humic acids interfere with chlorination at ≥ 1NTU (2 mg/L)  Chalk does not interfere with chlorination up to 5 NTU  Both culture and flow cytometry lead to same conclusions 3 AbstractChlorine is globally the most widely used chemical for water disinfection. Whereas disinfection efficiency is well known to depend on water pH and temperature, the effect of turbidity is less well studied. Although turbidity is measured online in most drinking water works and most countries where regulations exist have set limits of < 1 NTU for water leaving the works, the composition of turbidity is typically unknown. Given the heterogeneous nature of substances contributing to turbidity, the aim of this work was to study the effect of selected compounds on chlorination efficacy. The effect of humic acids and chalk on the inactivation of the indicator bacteria Escherichia coli and Enterococcus faecalis was assessed at neutral pH at different turbidity levels using both plate counting and flow cytometry in combination with membrane integrity staining. For humic acids, a turbidity of 1 NTU (corresponding to 2 mg/L) was identified as a critical threshold, which when exceeded was found to have a negative impact on chlorine disinfection. Chalk, on the other hand, had no measurable impact up to 5 NTU. The observation applied to both bacterial species with identical conclusions from the two diagnostic methods. Results corroborate that different turbidity causing substances affect chlorination efficiency to very different extents with chlorine demand by organic material probably being the most important determinant. In case of turbidities > 1 NTU, turbidity measurement benefits from the consideration of the organic content as mere NTU values do not allow predicting an impact on chlorination efficiency.