f In this study we tested 39 Lactococcus lactis strains isolated from diverse habitats for their robustness under heat and oxidative stress, demonstrating high diversity in survival (up to 4 log units). Strains with an L. lactis subsp. lactis phenotype generally displayed more-robust phenotypes than strains with an L. lactis subsp. cremoris phenotype, whereas the habitat from which the strains had been isolated did not appear to influence stress survival. Comparison of the stress survival phenotypes with already available comparative genomic data sets revealed that the absence or presence of specific genes, including genes encoding a GntR family transcriptional regulator, a manganese ABC transporter permease, a cellobiose phosphotransferase system (PTS) component, the FtsY protein, and hypothetical proteins, was associated with heat or oxidative stress survival. Finally, 14 selected strains also displayed diversity in survival after spray drying, ranging from 20% survival for the most robust strains, which appears acceptable for industrial application, to 0.1% survival for the least-tolerant strains. The high and low levels of survival upon spray drying correlated clearly with the combined robustness under heat and oxidative stress. These results demonstrate the relevance of screening culture collections for robustness under heat and oxidative stress on top of the typical screening for acidifying and flavor-forming properties. Based on their spoilage-preventing and flavor-enhancing characteristics, lactic acid bacteria (LAB) have been employed since ancient times in the fermentation of foods, e.g., fruits, vegetables, cereal grains, meat, and milk (1, 2). Nowadays, many of these processes have been industrialized, and fermentation is typically initiated by the addition of starter cultures, which contain high concentrations of one or multiple LAB strains (1, 2). As starter cultures require metabolic activity to contribute to the taste and texture of the fermentation end products, there has been an increasing industrial interest in studying robustness phenotypes during industrial production and processing (1), which involves preservation by either freezing or drying techniques (3-5). The major disadvantages of frozen starter cultures are the inconvenience and costs of transport and storage at low temperature, and, therefore, drying techniques are preferred (3-5). During spray drying, cultures are exposed to severe heat and oxidative stress (6, 7), typically resulting in lower survival rates of starter cultures than freeze-drying (3, 4). Therefore, freeze-drying is currently the most often applied industrial drying method (3-5). However, spray drying appears a more cost-effective and energy-efficient drying alternative for the preservation of starter cultures (3-5), providing strains that display high robustness under the stresses encountered in this process can be identified. This appears feasible, as studies on stress phenotypes typically result in highly strainspecific robustness phenotypes, e.g., for the gastro...

BackgroundLactococcus lactis is industrially employed to manufacture various fermented dairy products. The most cost-effective method for the preservation of L. lactis starter cultures is spray drying, but during this process cultures encounter heat and oxidative stress, typically resulting in low survival rates. However, viability of starter cultures is essential for their adequate contribution to milk fermentation, supporting the ambition to better understand and improve their robustness phenotypes.ResultsThis study describes a transcriptome-phenotype matching approach in which the starter L. lactis MG1363 was fermented under a variety of conditions that differed in the levels of oxygen and/or salt, as well as the fermentation pH and temperature. Samples derived from these fermentations in the exponential phase of bacterial growth were analyzed by full-genome transcriptomics and the assessment of heat and oxidative stress phenotypes. Variations in the fermentation conditions resulted in up to 1000-fold differences in survival during heat and oxidative stress. More specifically, aeration during fermentation induced protection against heat stress, whereas a relatively high fermentation temperature resulted in enhanced robustness towards oxidative stress. Concomitantly, oxygen levels and fermentation temperature induced differential expression of markedly more genes when compared with the other fermentation parameters. Correlation analysis of robustness phenotypes and gene expression levels revealed transcriptome signatures for oxidative and/or heat stress survival, including the metC-cysK operon involved in methionine and cysteine metabolism. To validate this transcriptome-phenotype association we grew L. lactis MG1363 in the absence of cysteine which led to enhanced robustness towards oxidative stress.ConclusionsOverall, we demonstrated the importance of careful selection of fermentation parameters prior to industrial processing of starter cultures. Furthermore, established stress genes as well as novel genes were associated with robustness towards heat and/or oxidative stress. Assessment of the expression levels of this group of genes could function as an indicator for enhanced selection of fermentation parameters resulting in improved robustness during spray drying. The increased robustness after growth without cysteine appeared to confirm the role of expression of the metC-cysK operon as an indicator of robustness and suggests that sulfur amino acid metabolism plays a pivotal role in oxidative stress survival.Electronic supplementary materialThe online version of this article (doi:10.1186/s12934-014-0148-6) contains supplementary material, which is available to authorized users.