In Vitro Analysis of Probiotic Properties Related to the Adaptation of Levilactobacillus brevis to Intestinal Microenvironment and Involvement of S-Layer Proteins

Although rare, the ability to produce surface S-layer proteins is beneficially associated with particular <i>Lactobacillus</i> strains being investigated as probiotics. Therefore, this work aimed to study specific probiotic functionalities of selected <i>Levilactobacillus brevis</i> strains MB1, MB2, MB13 and MB20, isolated from human milk microbiota, and to assess the contribution of S-proteins. Firstly, Rapid Annotation using Subsystem Technology revealed that cell wall-related genes were abundant in analysed <i>L. brevis</i> genomes. Furthermore, the results demonstrated that S-proteins mediate aggregation capacity and competitive exclusion of selected pathogens by <i>L. brevis</i> strains. The improvement of Caco-2 epithelial monolayer barrier function was demonstrated by the increase in JAM-A and occludin expressions when <i>L. brevis</i> strains or S-proteins were added, with the effect being most pronounced after treatment with MB2 and S-proteins of MB1. <i>L. brevis</i> strains, especially MB20, exerted the potential to adhere to recombinant human ZG16. Strain MB2 and MB20-S-proteins improved the barrier function of HT29 epithelial monolayer, as evidenced by increased ZG16 expression. Analysed <i>L. brevis</i> strains and S-proteins differentially affected the protein expression of IL-1β, IL-6 and IL-8, and IL-10 cytokines. The most prominent effect was observed by S-proteins of MB20, since IL-1β production was decreased while IL-10 production was significantly increased.