Caractérisation des propriétés d'adhésion de Streptococcus thermophilus LMD-9 aux cellules épithéliales intestinales : 1. Rôle des protéines de surface dans la résistance aux sels biliaires et dans l’adhésion 2. Impact de l’adhésion sur l’expression des gènes eucaryotes et bactériens

Abstract : Lactic acid bacteria are of great economic interest because of their use in the food industry. Among them, Streptococcus thermophilus (ST) is of major interest since it is the most used after Lactococcus lactis, for the manufacture of fermented dairy products and cheese. In addition, this bacterium is the only streptococcus to benefit from GRAS status (Generally Recognized As Safe). Beside its interest in the dairy industry, ST has beneficial effects on human intestinal health. Although these effects are widely documented, the probiotic status of ST remains to be consolidated. Therefore, studies are currently being conducted in order to select strains of ST with a high probiotic potential. Among criteria that are important to select ST strains include their ability to survive stress the drastic conditions of the digestive tract (DT) and their ability to adhere to intestinal cells. In this context, the aim of this thesis was to investigate firstly the in vitro adhesion capacity of the ST LMD-9 strain to different human intestinal cell lines and to evaluate the survival of this strain to bile salt stress. In order to highlight the potential role of some surface proteins in these two processes, three mutants derived from this strain and inactivated in the genes prtS (parietal protease), srtA (sortase A) and mucBP (Mucin Binding-protein) were also included in this study. Secondly, the impact of LMD-9 adhesion was analyzed, on one hand on the expression of some mucin-encoding genes in eukaryotic cells, and on the other hand on the expression of genes that would be specifically induced during adhesion process using the R-IVET (Recombinase-based In Vivo Expression Technology) approach. The results obtained demonstrated the ability of LMD-9 to survive up to the concentration of 3 mM of bile salts and that the PrtS, SrtA and MucBP surface proteins would be involved in the resistance to this stress. Our results also showed that the LMD-9 strain was capable of adhering to three cell lines used suggesting that this strain could interact with different mucins that may encounter in the DT. Moreover, the involvement of some surface proteins in the adhesion of LMD-9 has been found to be dependent on the surface characteristics of these cell lines, whether they are enterocytic (Caco-2) or mucus-secreting cells (HT29-MTX and HT29-CL.16E). Regarding the impact of LMD-9 adhesion on MUC2 and MUC5AC gene expression, no effect has been observed on the transcript level under our experimental conditions. Furthermore, for the first time, our results allowed us to identify genes specifically induced in the LMD-9 strain during adhesion process to epithelial cells. We have thus shown that the LMD-9 adhesion does not depend solely on surface proteins, but other functions and metabolic pathways are also involved. This thesis work contributes thus to new knowledge related to (i) the choice of the cellular model in in vitro bacterial adhesion studies, (ii) the ability of LMD-9 to survive bile salt stress by involving some surface proteins and (iii) understanding the molecular mechanisms of LMD-9 adhesion to epithelial cells
Document type :
Theses
File URL :
http://docnum.univ-lorraine.fr/ulprive/DDOC_T_2018_0123_BENEUX.pdf
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https://hal.univ-lorraine.fr/tel-01923777
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Submitted on : Thursday, November 15, 2018 - 2:24:00 PM
Last modification on : Monday, April 8, 2019 - 10:11:14 AM

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  • HAL Id : tel-01923777, version 1

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Mounira Kebouchi. Caractérisation des propriétés d'adhésion de Streptococcus thermophilus LMD-9 aux cellules épithéliales intestinales : 1. Rôle des protéines de surface dans la résistance aux sels biliaires et dans l’adhésion 2. Impact de l’adhésion sur l’expression des gènes eucaryotes et bactériens. Ingénierie des aliments. Université de Lorraine, 2017. Français. ⟨NNT : 2017LORR0209⟩. ⟨tel-01923777⟩

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