Mise au point d'une méthode de dénombrement de la totalité des cellules de levures de la surface d'un fromage à pâte molle

Abstract

A method was developed allowing Coulter Counter numeration of yeasts in soft cheeses without interference of casein micelles and fat globules. Casein and fat of a soft cheese suspension were completely solubilized by treating samples with 19 volumes of Driehuis and Teernstra (1992) transparisation reagent buffered at pH 12. The best conditions of transparisation were 30 min at 37 °C. The yeast dry weight, turbidimetric absorbance, microscopic count and Petri dish count methods were chosen as reference methods. After transparisation, the dry weight method could not be used because an irreversible adsorption of transparisation reagent on yeast cells took place and this reaction was not reproducible. Treatment of pure cultures of Kluyveromyces lactis and Geotrichum candidum with transparisation reagent reduced the cellular turbidity respectively by 30 ± 3% and 23 ± 3%. The turbidity reduction by transparisation depends on the type of strains and general form of cells. The turbidity method could be used for a strain in pure culture but not for the yeast mixture that exists on soft cheeses. It seemed that these yeast strains were relatively more resistant to the transparisation reagent than lactic acid bacteria. The transparisation killed the majority of initial viable cells of yeasts but it did not change their general morphology. For this reason, the Coulter Counter method could be used. Transparisation treatment dissociated aggregated cells and/or myceliums. Before and after transparisation treatment, the Coulter Counter numeration values could be plotted against microscopical counts. After transparisation of mixtures of yeast strains in the presence of soft cheese components, the Coulter Counter method allows a rapid determination of total yeast cell concentration and of a cell size distribution. It also allows an estimation of total cell concentration for each strain and its distribution, if the difference between the average diameters of yeast strains was higher than 1 μm. © 1995 Elsevier/INRA.