Highly chlorinated Escherichia coli cannot be stained by propidium iodide

Area of Science:

• Microbiology
• Environmental Science
• Biochemistry

Background:

• Bacterial viability assays often use fluorochromes like propidium iodide (PI).
• Chlorination is known to affect the staining of bacteria by various fluorochromes.
• The impact of chlorine on PI staining specifically requires further investigation.

Purpose of the Study:

• To evaluate the effect of chlorine on propidium iodide (PI) staining of bacteria and biomolecules.
• To determine the influence of chlorine concentration on PI fluorescence intensity.
• To assess the implications of chlorine-induced damage for bacterial viability assessment.

Main Methods:

• Studied Escherichia coli in suspension and biomolecules (DNA, RNA, BSA, palmitic acid, dextran) in solution.
• Subjected samples to chlorine (bleach solution) followed by neutralization with sodium thiosulphate.
• Stained samples with propidium iodide (PI) and measured fluorescence intensity.

Main Results:

• Chlorine significantly decreased PI fluorescence intensity of DNA and RNA in solution.
• At chlorine concentrations >3 micromol/L Cl2, E. coli cells were too damaged for proper PI staining.
• Chlorine-induced nucleic acid damage resulted in PI false-negative cells, masking cell death.

Conclusions:

• Chlorine exposure compromises PI staining accuracy by damaging bacterial nucleic acids.
• PI staining may underestimate bacterial death in chlorinated environments due to false-negative results.
• Caution is advised when interpreting PI staining results from chlorinated samples.