FLUOROMETRIC ASSESSMENT OF PSEUDOMONAS-FLUORESCENS VIABILITY AFTER FREEZE-THAWING USING ETHIDIUM-BROMIDE

The relationship between impairment of the Pseudomonas fluorescens eel envelope's permeability barrier for ethidium cation, the fluorescent moiety of ethidium bromide, and viability after freeze-thawing was inv estigated. Ethidium fluorescence in the suspension of intact bacteria did not change. Disruption of the bacterial permeability barrier by ce tyltrimethylammonium bromide (CTAB) led to ethidium fluorescence incre ase due to interaction of the fluorochrome with intracellular nucleic acids. In the suspension of freeze-thawed cells, ethidium fluorescence increased and the subsequent treatment by CTAB resulted in further fl uorescence increase up to the final level corresponding to that in CTA B-treated intact bacteria. For bacteria exposed to different freeze-th awing regimes, the relative ethidium fluorescence increase closely cor related with the relative number of fluorescing cells revealed microsc opically. In the suspension of freeze-thawed cells, the relative addit ional ethidium fluorescence increase after CTAB treatment closely corr elated with viability evaluated by plate counts. It is concluded that the fluorimetric approach may be used as a means of rapidly evaluating bacterial viability after freeze-thawing.