Carbohydrate Metabolism Is Essential for the Colonization of Streptococcus thermophilus in the Digestive Tract of Gnotobiotic Rats

Streptococcus thermophilus is the archetype of lactose-adapted bacterium and so far, its sugar metabolism has been mainly investigated in vitro. The objective of this work was to study the impact of lactose and lactose permease on S. thermophilus physiology in the gastrointestinal tract (GIT) of gnotobiotic rats. We used rats mono-associated with LMD-9 strain and receiving 4.5% lactose. This model allowed the analysis of colonization curves of LMD-9, its metabolic profile, its production of lactate and its interaction with the colon epithelium. Lactose induced a rapid and high level of S. thermophilus in the GIT, where its activity led to 49 mM of intra-luminal L-lactate that was related to the induction of mono-carboxylic transporter mRNAs (SLC16A1 and SLC5A8) and p27Kip1 cell cycle arrest protein in epithelial cells. In the presence of a continuous lactose supply, S. thermophilus recruited proteins involved in glycolysis and induced the metabolism of alternative sugars as sucrose, galactose, and glycogen. Moreover, inactivation of the lactose transporter, LacS, delayed S. thermophilus colonization. Our results show i/that lactose constitutes a limiting factor for colonization of S. thermophilus, ii/that activation of enzymes involved in carbohydrate metabolism constitutes the metabolic signature of S. thermophilus in the GIT, iii/that the production of lactate settles the dialogue with colon epithelium. We propose a metabolic model of management of carbohydrate resources by S. thermophilus in the GIT. Our results are in accord with the rationale that nutritional allegation via consumption of yogurt alleviates the symptoms of lactose intolerance

Lactobacillaceae and Cell Adhesion: Genomic and Functional Screening

The analysis of collections of lactic acid bacteria (LAB) from traditional fermented plant foods in tropical countries may enable the detection of LAB with interesting properties. Binding capacity is often the main criterion used to investigate the probiotic characteristics of bacteria. In this study, we focused on a collection of 163 Lactobacillaceace comprising 156 bacteria isolated from traditional amylaceous fermented foods and seven strains taken from a collection and used as controls. The collection had a series of analyses to assess binding potential for the selection of new probiotic candidates. The presence/absence of 14 genes involved in binding to the gastrointestinal tract was assessed. This enabled the detection of all the housekeeping genes (ef-Tu, eno, gap, groEl and srtA) in the entire collection, of some of the other genes (apf, cnb, fpbA, mapA, mub) in 86% to 100% of LAB, and of the other genes (cbsA, gtf, msa, slpA) in 0% to 8% of LAB. Most of the bacteria isolated from traditional fermented foods exhibited a genetic profile favorable for their binding to the gastrointestinal tract. We selected 30 strains with different genetic profiles to test their binding ability to non-mucus (HT29) and mucus secreting (HT29-MTX) cell lines as well as their ability to degrade mucus. Assays on both lines revealed high variability in binding properties among the LAB, depending on the cell model used. Finally, we investigated if their binding ability was linked to tighter cross-talk between bacteria and eukaryotic cells by measuring the expression of bacterial genes and of the eukaryotic MUC2 gene. Results showed that wild LAB from tropical amylaceous fermented food had a much higher binding capacity than the two LAB currently known to be probiotics. However their adhesion was not linked to any particular genetic equipment

Les animaux gnotobiotiques : pour mieux comprendre le microbiote intestinal

Gnotobiotic animals to improve our understanding of the intestinal microbiota A large number of data on microbiota has emerged recently, but functional interactions between the microbiota and intestinal cells remain poorly understood, even though they are absolutely essential for health. In this context, we chose two examples of gnotobiotic models used to analyse the dialogue between intestinal cells and microbiota. Animals mono-associated with Bacteroides thetaiotaomicron show the mutual effects between a commensal bacterium and the host. The study of gnotobiotic animals carrying yogurt bacteria helped suggest new pathways for the regulation of sugar metabolism in lactic acid bacteria, and new molecular pathways by which these bacteria may influence the physiology of the host.De nombreuses données récentes concernent le microbiote intestinal, mais les interactions fonctionnelles qui s’établissent entre ce microbiote et les entérocytes restent encore peu connues, alors qu’elles sont tout à fait essentielles pour la santé. Nous avons choisi deux exemples illustrant l’utilisation d’animaux gnotobiotiques pour analyser le dialogue entre les cellules intestinales et le microbiote. Les animaux mono-associés avec la bactérie commensale Bacteroides thetaiotaomicron révèlent les effets mutuels qui existent entre une bactérie commensale et l’hôte. L’étude d’animaux gnotobiotiques, porteurs de bactéries du yaourt, permet de proposer des nouvelles voies de régulation du métabolisme des sucres des bactéries lactiques et de nouvelles voies moléculaires par lesquelles ces bactéries pourraient influencer la physiologie de l’hôte.Thomas Muriel, Wrzosek Laura, Rul Françoise, Langella Philippe. Les animaux gnotobiotiques : pour mieux comprendre le microbiote intestinal. In: Bulletin de l'Académie Vétérinaire de France tome 166 n°1, 2013. pp. 19-23

The role of aminopeptidase PepS in the growth of <em>Streptococcus thermophilus</em> is not restricted to nitrogen nutrition

International audienceAims: To investigate the effect of an absence of aminopeptidase PepS on the growth of Streptococcus thermophilus on different media and at different temperatures. Methods and Results: Using gene interruption, a negative mutant of the Strep. thermophilus CNRZ385 strain was constructed for the aminopeptidase PepS (strain D pepS). Checks were first of all made using biochemical assays that the D pepS strain lacks the peptide hydrolase activity of aminopeptidase PepS. It was demonstrated that the absence of the aminopeptidase PepS exerted a negative effect on growth whatever the culture medium (M17, chemically defined medium, milk). The role of aminopeptidase PepS in growth was enhanced at a high temperature (45°C vs 37°C). The D pepS strain was more resistant to lysozyme than the wild-type strain. Conclusions: We were able to demonstrate that aminopeptidase PepS probably plays a pleiotropic role through its involvement in growth via nitrogen nutrition, as well as via other cellular functions⁄metabolisms (such as peptidoglycane metabolism). Significance and Impact of the Study: This study constitutes the first report on the role of a member of the M29 MEROPS family of metallopeptidases (http:// merops.sanger.ac.uk/)

Activation of ERK, Controlled by Rac1 and Cdc42 via Akt, Is Required for Anoikis

International audienceWe have recently reported that two Rho family GTPases, Rac1 and Cdc42, are intimately involved in the control of cell survival of murine fibroblasts linked to adherence to the extracellular matrix. Inhibition of either Rac1 or Cdc42 signaling in adherent cells mimics the loss of anchorage and efficiently induces apoptosis in both immortalized and primary cells. In both cases cell death is dependent on the wild-type p53 tumor suppressor and is accompanied by activation of endogenous p53. Here, we describe that the inhibition of Rac1 or Cdc42 signaling leads to MAPK ERK activation via a pathway involving PI(3)K, Akt, Raf, and MEK, but not Ras. The moderate level of ERK activation that accompanies anoikis is an essential component of proapoptotic signaling; whereas sustained, high-intensity ERK signaling promotes survival in the same experimental system