Flow cytometric assessment of the effects of chlorine, chloramine, and UV on bacteria by using nucleic acid stains and 5-cyano-2,3-ditolyltetrazolium chloride

Abstract

Flow cytometry based on nucleic acid stains and CTC was established and optimized.

Membrane of S. aureus is more resistant to chlorine/chloramine than E. coli.

The metabolic activity of bacteria persisted after the cytomembranewas damaged.

S. aureus showed more resistance to UV irradiation than E. coli by FCM.

MP-UV was a stronger inhibitor of metabolic activity than LP-UV.

Flow cytometry (FCM) has been widely used in multi-parametric assessment of cells in various research fields, especially in environmental sciences. This study detected the metabolic activity of Escherichia coli and Staphylococcus aureus by using an FCM method based on 5-cyano-2,3-ditolyltetrazolium chloride (CTC); the accuracy of this method was enhanced by adding SYTO 9 and 10% R2A broth. The disinfection effects of chlorine, chloramine, and UV were subsequently evaluated by FCM methods. Chlorine demonstrated stronger and faster destructive effects on cytomembrane than chloramine, and nucleic acids decomposed afterwards. The metabolic activity of the bacteria persisted after the cytomembranewas damaged as detected using CTC. Low-pressure (LP) UV or medium-pressure (MP) UV treatments exerted no significant effects on membrane permeability. The metabolic activity of the bacteria decreased with increasing UV dosage, and MP-UV was a stronger inhibitor of metabolic activity than LP-UV. Furthermore, the membrane of Gram-positive S. aureus was more resistant to chlorine/chloramine than that of Gram-negative E. coli. In addition, S. aureus showed higher resistance to UV irradiation than E. coli.

Graphical abstract

Keywords

Flow cytometry / Escherichia coli / Staphylococcusaureus / UV / CTC / SYTO 9

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