Physiological processes of yeast cultures followed by changes in cell Surface potential

LAVAISSE, L.; HOLLMAN, A.; NAZARENO M. A.; DISALVO, A.

San Miguel de Tucumán

Congreso; XI Congreso Argentino de Microbiología General; 2015

Asociación Civil de Microbiología General

Traditional techniques used for monitoring the viability and the growth of cell cultures such as measurements of turbidity of cell suspensions (as absorbance) exhibit some limitation because provides information of the bulk of the cell culture reporting an averaged state considering the whole cell population, without distinguishing alive cells from dead cells. On the other hand, the colony forming units (CFU) method that reports viable cells is very sensitive but depends on plating conditions and is relatively time-consuming. Finally, methylene blue staining distinguishes between alive and dead cells, but does not offer the same level of sensitivity. In contrast, zeta potential determination by optical microscopy provides information about the state of a single cell because it is able to measure cell surface charges in individual cells of the suspension. The goal of this work is to determine changes in the surface properties of cells in suspensions following the zeta potential and relate it to the physiological states. Forthis purpose Saccharomyces cerevisiae isolated from instant dry yeast commercial powder was chosen as a biological model system. Yeasts were grown in standard medium (Sabouraud glucose broth). At different incubation times, culture samples werecollected to evaluate yeast growth, performing CFU, OD600nm and following variations in the electrophoretic mobility of cells in an electric field. Data obtained show that changes in optical density during exponential growth phase are concomitant with a change in zeta potential values. A clear correlation between optical density and the zeta potential as a bulk measure was observed. A closer inspection of the zeta potential of individual cells allowed establishing that a distribution of cells with different zeta potentials exists in each sample. This suggests that different metabolic stages converge at each time of culture when averaged values are taken by turbidity. After the exponential phase, two main jumps in zeta potential values are observed. The first one matches with the transition from exponential to stationary phase, and the second one with the transition of late stationary to death phase. These results were confirmed by viability staining in an inverted fluorescence microscope and colony counting plate. It is concluded that zeta potential is a valuable tool for following the growing of suspended cells. It denotes that cells in different metabolic states coexist in a culture, revealing a delicate balance of populations. In comparison to turbidity it allow to anticipate the beginning of the death phase. The surface changes observed are possibly related to metabolic changes taken place during its growing. It is expected that the alteration of this balance predicts the changes on the growth cell phase.