Effectiveness of CSE to counterstain particles and dead bacterial cells with permeabilised membranes: application to viability assessment in waters

• Published in: FEMS microbiology letters 1999-09, Vol.178 (2), p.219-226
• Main Authors: Catala, Philippe, Parthuisot, Nathalie, Bernard, Laetitia, Baudart, Julia, Lemarchand, Karine, Lebaron, Philippe
• Format: Article
• Language: English
• Subjects: Bacteria; Bacteria - growth & development; Bacteria - isolation & purification; Bacteriological methods and techniques used in bacteriology; Bacteriological Techniques; Bacteriology; Biological and medical sciences; Cell Membrane Permeability; Colony Count, Microbial; Cytometry; Dyes; Environmental Sciences; Esterase; Flow Cytometry; Fluorescent Dyes; Fluorescent indicators; Fundamental and applied biological sciences. Psychology; Heat treatment; Membranes; Microbiology; Natural environment; Natural waters; Scanning cytometry; Seawater; Solid phases; Staining and Labeling; Strains (organisms); Toxicity; Viability; Viability dye; Water Microbiology; Water Supply
• ISSN: 0378-1097; 1574-6968
• DOI: 10.1111/j.1574-6968.1999.tb08680.x

Abstract The CSE dye (Chemunex, Maisons-Alfort, France) was combined with an activity marker to improve bacterial activity assessment in natural waters. Its effectiveness to counterstain dead cells with permeabilised membranes was investigated on live and dead cells of a variety of strains from collections or isolated from the natural environment. Cells were killed by heat treatment. For all strains tested, the fluorescent dye showed an intense staining of killed cells having permeabilised membranes while no significant signal was detected when applied to live cells. Furthermore, the CSE dye had no toxicity on viable cells. Then, CSE was combined with the ChemChrome V6 dye (Chemunex) to assess the activity of bacterial cells in different waters. Both fluorescences were analysed simultaneously by solid-phase cytometry. The active cell counts were sometimes lower when both dyes were combined suggesting that CSE was able to counterstain cells having a residual esterase activity and compromised membranes. These cells were subtracted from the active cell counts determined with ChemChrome V6. In most samples, active cell counts were congruent with those determined by the direct viable count method.