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

The CSE dye (Chemunex, Maisons-Alfort, France) was combined with an activit y marker to improve bacterial activity assessment in natural waters. Its ef fectiveness to counterstain dead cells with permeabilised membranes was inv estigated on live and dead cells of a variety of strains from collections o r isolated from the natural environment. Cells were killed by hear treatmen t. For all strains tested, the fluorescent dye showed an intense staining o f killed cells having permeabilised membranes while no significant signal w as detected when applied to live cells. Furthermore, the CSE dye had no tox icity on viable cells. Then, CSE was combined with the ChemChrome V6 dye (C hemunex) to assess the activity of bacterial cells in different waters. Bot h fluorescences were analysed simultaneously by solid-phase cytometry. The active cell counts were sometimes lower when both dyes were combined sugges ting that CSE was able to counterstain cells having a residual esterase act ivity and compromised membranes. These cells were subtracted from the activ e cell counts determined with ChemChrome V6. In most samples, active cell c ounts were congruent with those determined by the direct viable count metho d. (C) 1999 Federation of European Microbiological Societies. Published by Elsevier Science B.V. All rights reserved.