Abstract
BackgroundThe morphology of yeast cells changes during budding, depending on the growth rate and cultivation conditions. A photo-optical microscope was adapted and used to observe such morphological changes of individual cells directly in the cell suspension. In order to obtain statistically representative samples of the population without the influence of sampling, in situ microscopy (ISM) was applied in the different phases of a Saccharomyces cerevisiae batch cultivation. The real-time measurement was performed by coupling a photo-optical probe to an automated image analysis based on a neural network approach.ResultsAutomatic cell recognition and classification of budding and non-budding cells was conducted successfully. Deviations between automated and manual counting were considerably low. A differentiation of growth activity across all process stages of a batch cultivation in complex media became feasible. An increased homogeneity among the population during the growth phase was well observable. At growth retardation, the portion of smaller cells increased due to a reduced bud formation. The maturation state of the cells was monitored by determining the budding index as a ratio between the number of cells, which were detected with buds and the total number of cells. A linear correlation between the budding index as monitored with ISM and the growth rate was found.ConclusionIt is shown that ISM is a meaningful analytical tool, as the budding index can provide valuable information about the growth activity of a yeast cell, e.g. in seed breeding or during any other cultivation process. The determination of the single-cell size and shape distributions provided information on the morphological heterogeneity among the populations. The ability to track changes in cell morphology directly on line enables new perspectives for monitoring and control, both in process development and on a production scale.
Highlights
The morphology of yeast cells changes during budding, depending on the growth rate and cultivation conditions
The morphology of single cells is traditionally measured with microscopy
The present study aims to achieve a further development stage by monitoring the maturation state of the budding yeast Saccharomyces cerevisiae with in situ microscopy (ISM) on a single-cell level
Summary
The morphology of yeast cells changes during budding, depending on the growth rate and cultivation conditions. The real-time measurement was performed by coupling a photo-optical probe to an automated image analysis based on a neural network approach. Due to a certain relationship between form and function, the growth state of cells, and even the production performance can be investigated on the basis of cell size and other morphological features. As such features are determined off line or at line, they. Neither a representative sample is obtained, since only a few cells are counted at certain specific time points, nor the heterogeneity of the cell population is considered. If conventional microscopy is coupled to a sampling tube and
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