Table_2_Transcriptome Analysis of a Spray Drying-Resistant Subpopulation Reveals a Zinc-Dependent Mechanism for Robustness in L. lactis SK11.xlsx

The viability of starter cultures is essential for an adequate contribution to the fermentation process and end-product. Therefore, robustness during processing and storage is an important characteristic of starter culture strains. For instance, during spray drying cells are exposed to heat and oxidative stress, generally resulting in loss of viability. In this study, we exposed the industrially relevant but stress-sensitive Lactococcus lactis strain SK11 to two cycles of heat stress, with intermediate recovery and cultivation at moderate temperatures. After these two cycles of heat exposure, the abundance of robust derivatives was increased as compared with the original culture, which enabled isolation of heat-resistant subpopulations displaying up to 1,000-fold enhanced heat stress survival. Moreover, this heat-resistant subpopulation demonstrated an increased survival during spray drying. Derivatives from two independent lineages displayed different transcriptome changes as compared with the wild type strain, indicating that the increased robustness within these lineages was established by different adaptive strategies. Nevertheless, an overlap in differential gene expression in all five derivatives tested in both lineages included three genes in an operon involved in zinc transport. The link between zinc homeostasis and heat stress survival in L. lactis was experimentally established by culturing of the wild type strain SK11 in medium with various levels of zinc ions, which resulted in alterations in heat stress survival phenotypes. This study demonstrates that robust derivatives of a relatively sensitive L. lactis strain can be isolated by repeated exposure to heat stress. Moreover, this work demonstrates that transcriptome analysis of these robust derivatives can provide clues for improvement of the robustness of the original strain. This could boost the industrial application of strains with specific desirable traits but inadequate robustness characteristics.