Comprehensive assessment of bacterial community viability in natural whey starter cultures by flow cytometric approach

The natural whey starter consists of a lactic microbial community perpetuated daily by fermenting the whey from the previous day’s processing. These natural cultures play a crucial role in the acidification process of the cheese mass, which must lead to the complete breakdown of lactose in the first few hours after its production. The lactic acid bacteria in the whey starter act first, through their metabolism, as living cells and then as released enzymes, participating in the complex biochemical phenomena that occur during cheese ripening. The viability of the cells plays a key role in the acidifying activity required during the first hours of curd acidification, in parallel to the microbial composition. Analytical methods based on plate counts have long been used to evaluate the number of cultured cells. The disadvantage of these methods is that they have high variability and take days to obtain results. In this work, we focused on evaluating the physiological state of the lactic bacterial cell community of natural whey starter samples. This work involved a comparison of bacterial enumeration by classical methods using whey agar medium, MRS, and M17, and flow cytometry on natural whey samples. The flow cytometry results were in good agreement with a tendency towards overestimation. Flow cytometry has also introduced other parameters to assess natural whey by measuring cell physiological status. In addition, two fluorescent dyes used in flow cytometry resulted in the assessment of cell wall damage and metabolic activity and therefore enabled the estimation of the number of cells even under suboptimal physiological states. In conclusion, we discovered that determining the physiological condition of the cells served as a potential indicator for predicting its acidifying activity.