Factors important for persistence of Lactobacillus reuteri in the gastrointestinal tract

Lactobacillus reuteri ATCC 55730 is a commercially available probiotic i.e. health promoting bacterium. The interest for lactic acid bacteria (LAB) as probiotics might be due to the tradition of use in food preservation, their non-pathogenic behavior and, a natural occurrence in the human gastrointestinal (GI) tract. Although examples of clinically proven probiotic effects are increasing, the understandings of bacterial-host interactions behind these effects are only beginning to be unravelled. To put this thesis work into context, the field of probiotic research is described and health promoting mechanisms and perspectives of strain selection are discussed. The experimental work has focused on properties of L. reuteri ATCC 55730 that are tentatively important for persistence of this and other probiotic strains in the GI tract. To get an idea of the proteins on the surface of L. reuteri ATCC 55730, a draft genome sequence was scrutinized using bioinformatic tools. This resulted in the identification of a diverse set of 126 putative proteins with different possible means of surface attachment. The bacterium was also exposed to heat and acid stress and the response monitored by DNA microarrays. Acid stress is a well-documented way of investigating probiotic strains, since survival in the acidic GI tract is considered a prerequisite for a probiotic effect. Heat stress could be considered an artificially constructed environment for a probiotic bacterium, but is probably still relevant in elucidating mechanisms of persistence in other stressful conditions such as the passage of the GI tract. To further investigate genes revealed in the bioinformatical and DNA microarray analysis as well as a set of genes from the seven earlier identified two-component systems of L. reuteri ATCC 55730, 14 genes were mutated. These where then evaluated for ecological performance using a three-stage continuous culture system simulating a human colon using a real time PCR approach to monitor the relative abundance of the individual mutants. Two caseinolytic protease (Clp) genes, clpE and clpL that were induced in heat and acid stress respectively, both showed importance for persistence in the colon model.