Strain-Dependent Transcriptome Signatures for Robustness in Lactococcus lactis

Abstract

Recently, we demonstrated that fermentation conditions have a strong impact on subsequent survival of Lactococcus lactis strain MG1363 during heat and oxidative stress, two important parameters during spray drying. Moreover, employment of a transcriptome-phenotype matching approach revealed groups of genes associated with robustness towards heat and/or oxidative stress. To investigate if other strains have similar or distinct transcriptome signatures for robustness, we applied an identical transcriptome-robustness phenotype matching approach on the L. lactis strains IL1403, KF147 and SK11, which have previously been demonstrated to display highly diverse robustness phenotypes. These strains were subjected to an identical fermentation regime as was performed earlier for strain MG1363 and consisted of twelve conditions, varying in the level of salt and/or oxygen, as well as fermentation temperature and pH. In the exponential phase of growth, cells were harvested for transcriptome analysis and assessment of heat and oxidative stress survival phenotypes. The variation in fermentation conditions resulted in differences in heat and oxidative stress survival of up to five 10-log units. Effects of the fermentation conditions on stress survival of the L. lactis strains were typically strain-dependent, although the fermentation conditions had mainly similar effects on the growth characteristics of the different strains. By association of the transcriptomes and robustness phenotypes highly strain-specific transcriptome signatures for robustness towards heat and oxidative stress were identified, indicating that multiple mechanisms exist to increase robustness and, as a consequence, robustness of each strain requires individual optimization. However, a relatively small overlap in the transcriptome responses of the strains was also identified and this generic transcriptome signature included genes previously associated with stress (ctsR and lplL) and novel genes, including nanE and genes encoding transport proteins. The transcript levels of these genes can function as indicators of robustness and could aid in selection of fermentation parameters, potentially resulting in more optimal robustness during spray drying.

Characterization of a Prophage-Free Derivative Strain of Lactococcus lactis ssp. lactis IL1403 Reveals the Importance of Prophages for Phenotypic Plasticity of the Host

“…This suggests that the effects observed here were caused by as-yet-unidentified bIL311- and bIL312-encoded resistance function(s). It has been shown recently that two late bIL311 genes, orf17 and orf18 , and one bIL312 gene, orf21 , are expressed in the host IL1403 strain ( Dijkstra et al, 2016 ). The predicted Orf17 protein shows homology to proteins from the GNAT family of acetyltransferases.…”

“…The genome of IL1403 contains six prophage-like elements, which represent in total 6.7% of the IL1403 chromosome ( Chopin et al, 2001 ). Transcriptional analyses of IL1403 and other lactococcal strains have revealed that, in addition to the genes encoding repressor proteins, other phage-specific genes are expressed in the lysogenic state under different growth conditions ( Boyce et al, 1995 ; Blatny et al, 2003 ; Seegers et al, 2004 ; Xie et al, 2004 ; Cretenet et al, 2011 ; Dijkstra et al, 2016 ; Visweswaran et al, 2017 ). As yet, though, it is unknown if or how these prophage-encoded proteins affect the physiology of the lactococcal cell.…”

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

“…The fact that these could still be enriched despite the apparent counter selection in the intermediate culturing step showcases the dramatically increased robustness phenotypes that are present in small subpopulations of cells in a culture. Moreover, no significant correlation was observed between growth rate and stress survival of the isolates after two cycles of heat stress, demonstrating that an increase in robustness can be achieved without necessarily reducing the growth rate, as has been previously demonstrated in multiple L. lactis strains as well (Dijkstra et al, 2014a , 2016 ).…”

“…For example, previous studies demonstrated that a relatively high oxygen level during fermentation resulted in an increased robustness toward heat stress of L. lactis strain MG1363 and exposure to mild acid stress increased survival of strain NCDO712 during subsequent lethal doses of acid, heat, osmotic, oxidative, and ethanol stress (O'Sullivan and Condon, 1997 ; Dijkstra et al, 2014a ). However, these adaptation methods can complicate the fermentation process, effects of adaptation conditions on robustness vary among strains (Dijkstra et al, 2016 ) and robustness is considered to be only improved for a short period in contrast to mutants obtained from evolution and selection experiments.…”

“…cremoris strain SK11, which has been isolated from a cheese starter culture, has been previously demonstrated to be relatively sensitive toward heat stress and spray drying (Dijkstra et al, 2014b ). Furthermore, heat stress survival of SK11 was shown to be highly influenced by fermentation conditions (Dijkstra et al, 2016 ). In this study, we repeatedly exposed strain SK11 to heat stress to select derivatives with improved robustness, which is relevant for survival during spray drying (Santivarangkna et al, 2008 ; Dijkstra et al, 2014b ).…”