Compatible solutes in lactic acid bacteria subjected to water stress

The goal of the research project described in this thesis was to investigate the protective effect of compatible solutes on tactic acid bacteria subjected to drying. Dried preparations of lactic acid bacteria are applied as starter cultures in feed and food industries. Dried starter cultures compare favourable to frozen cultures because of lower transport and storage costs. During desiccation by drying a considerable inactivation occurs resulting in low survival and subsequently, reduced residual activity. Physiological adaptation of tactic acid bacteria using compatible solutes prior to the drying process increases survival and activity after drying. Possibly, this results in a commercially attractive dried starter culture.Compatible solutes accumulate in cells during decreased water activity (Chapter 1). Reduced water activity can be accomplished either by addition of high concentrations of osmolytes, such as salts and sugars to a system (osmotic stress) or by removing the water of the system by drying (desiccation stress). Research on compatible solutes in bacteria has mainly been focused on the former system. In this thesis both systems have been investigated.Prior to desiccation by drying cells of Lactobacillus plantarum were subjected to osmotic stress by the addition of salt to the growth medium. Additionally, the compatible solute betaine was included in this stressed medium! As a result, cells cultured in the presence of salt and betaine showed enhanced survival after drying (Chapter 2).To investigate if L. plantarum accumulated compatible solutes like betaine 13C-nuclear magnetic resonance ( 13C-NMR) analysis were performed of cellular extracts of stressed cells. Betaine was accumulated together with a novel compatible solute identified as carnitine (Chapter 3).In addition to the results described in chapter 2 and 3, other tactic acid bacteria were tested for their ability to accumulate betaine and carnitine. Subsequently, the effect of betaine on survival after drying was tested (Chapter 4). Lactobacillus bulgaricus, a salintracellulart-sensitive bacterial strain, accumulated neither betaine nor carnitine and hence survival after drying remained low compared to strains that were more salt-tolerant. Only L. plantarum raised both levels of intracellular betaine and carnitine when the salt concentration of the medium was raised, whereas Lactobacillus halotolerans and Enterococcus faecium only raised betaistructurallyne levels .Molecules, structurally related to betaine or carnitine, improved growth of L. plantarum subjected to osmotic stress (Chapter 3). The betaine related compounds choline and acetylchotine were accumulated during osmotic stress conditions (Chapter 5). In other bacteria, the positive charged choline is converted into the zwitterionic betaine. Presumably, L. plantarum balanced the charge of choline (or acetylcholine) by the accumulation of the anionic amino acid glutamate. L. plantarum, also accumulated carnitine related compounds such as acetylcholine and propionylcholine (Chapter 6). These accumulated molecules are unchanged consequently, glutamate levels in cells now decreased.Accumulation of compatible solutes increased survival of several tactic acid bacteria strains subjected to drying. It would now be interesting to continue research by identifying the protective role of these solutes in desiccated cells and compare the results with cells which have not accumulated compatible solutes.