Safety and functional enrichment of gut microbiome in healthy subjects consuming a multi-strain fermented milk product: a randomised controlled trial.

Many clinical studies have evaluated the effect of probiotics, but only a few have assessed their dose effects on gut microbiota and host. We conducted a randomized, double-blind, controlled intervention clinical trial to assess the safety (primary endpoint) of and gut microbiota response (secondary endpoint) to the daily ingestion for 4 weeks of two doses (1 or 3 bottles/day) of a fermented milk product (Test) in 96 healthy adults. The Test product is a multi-strain fermented milk product, combining yogurt strains and probiotic candidate strains Lactobacillus paracasei subsp. paracasei CNCM I-1518 and CNCM I-3689 and Lactobacillus rhamnosus CNCM I-3690. We assessed the safety of the Test product on the following parameters: adverse events, vital signs, hematological and metabolic profile, hepatic, kidney or thyroid function, inflammatory markers, bowel habits and digestive symptoms. We explored the longitudinal gut microbiota response to product consumption and dose, by 16S rRNA gene sequencing and functional contribution by shotgun metagenomics. Safety results did not show any significant difference between the Test and Control products whatever the parameters assessed, at the two doses ingested daily over a 4-week-period. Probiotic candidate strains were detected only during consumption period, and at a significantly higher level for the three strains in subjects who consumed 3 products bottles/day. The global structure of the gut microbiota as assessed by alpha and beta-diversity, was not altered by consumption of the product for four weeks. A zero-inflated beta regression model with random effects (ZIBR) identified a few bacterial genera with differential responses to test product consumption dose compared to control. Shotgun metagenomics analysis revealed a functional contribution to the gut microbiome of probiotic candidates.

Faecal microbiota transplantation in recurrent Clostridium difficile infection: Recommendations from the French Group of Faecal microbiota Transplantation.

Faecal microbiota transplantation is effective for treating recurrent forms of Clostridium difficile infection and its use in this indication is recommended in the most recent European and North American guidelines. In this context, faecal microbiota transplantation is beginning to be performed in France in clinical practice, while the rules governing this procedure have been defined in France only for clinical trials. To unify, secure, and evaluate practice in this field in France, the French Group of Faecal microbiota Transplantation (FGFT) was created in October 2014 with the support of the French National Society of Gastroenterology, the French Infectious Disease Society, and the National Academy of Pharmacy. We present here the deliberations of this group regarding the use of faecal microbiota transplantation for recurrent Clostridium difficile infection. The issues addressed are the indications, therapeutic sequence, delivery procedures, donor selection, methods and conditions of specimen preparation, and traceability.

The intestine and its microflora are partners for the protection of the host: report on the Danone Symposium "The Intelligent Intestine," held in Paris, June 14, 2002.

The intestine is an extremely complex living system that participates in the protection of the host through a strong defense against aggressions from the external environment. This defensive task is based on 3 constituents that are in permanent contact and dialog with each other: the microflora, mucosal barrier, and local immune system. We review herein current knowledge about these important functions. The gut microflora play a major role against exogenous bacteria through colonization resistance, but the mechanism of action is not yet established, although it is linked to the bacteria colonizing the gut. This colonization involves bacteria-bacteria dialog, bacteria-mucins interactions, and bacteria-colonocytes cross-talk associated with environmental factors. The intestinal mucosa is a cellular barrier and the main site of interaction with foreign substances and exogenous microorganisms. It is a complex physicochemical structure consisting of a mucous layer linked to cellular and stromal components that participate in the defense of the host through mucosal blood flow, mucosal secretions, epithelial cell functionals, surface hydrophobicity, and defensin production. The intestine is the primary immune organ of the body represented by the gut-associated lymphoid tissue through innate and acquired immunity. This immune system can tolerate dietary antigens and the gut-colonizing bacteria and recognizes and rejects enteropathogenic microorganisms that may challenge the body's defenses. In cooperation with these endogenous barriers, some in-transit bacteria, such as probiotics, can act as partners of the defense system of the intestine.

Inhibition of in vitro cell adherence of Clostridium difficile by Saccharomyces boulardii.

The influence on the adherence of Clostridium difficile to Vero cells of the yeast Saccharomyces boulardii, the yeast fractions (cytoplasm and cell wall) and the culture supernatant was investigated in vitro. C. difficile adherence was significantly inhibited when bacteria were pre-incubated with the whole yeast and the cell wall fraction; this adherence inhibition was dose-dependent. The cell wall fraction also acts upon the target cultured cells inasmuch as the level of adherence was significantly decreased when Vero cells were preincubated with it. The same experiments carried out in the presence of an inhibitor of serine proteases resulted in no inhibition of bacterial adherence. These results suggest that the yeast could inhibit adherence of C. difficile to cells thanks to its proteolytic activity but also through steric hindrance.

A Clostridium difficile gene encoding flagellin.

Six strains of Clostridium difficile examined by electron microscopy were found to carry flagella. The flagella of these strains were extracted and the N-terminal sequences of the flagellin proteins were determined. Four of the strains carried the N-terminal sequence MRVNTNVSAL exhibiting up to 90% identity to numerous flagellins. Using degenerate primers based on the N-terminal sequence and the conserved C-terminal sequence of several flagellins, the gene encoding the flagellum subunit (fliC) was isolated and sequenced from two virulent strains. The two gene sequences exhibited 91% inter-strain identity. The gene consists of 870 nt encoding a protein of 290 amino acids with an estimated molecular mass of 31 kDa, while the extracted flagellin has an apparent molecular mass of 39 kDa on SDS-PAGE. The FliC protein displays a high degree of identity in the N- and C-terminal amino acids whereas the central region is variable. A second ORF is present downstream of fliC displaying homology to glycosyltransferases. The fliC gene was expressed in fusion with glutathione S-transferase, purified and a polyclonal monospecific antiserum was obtained. Flagella of C. difficile do not play a role in adherence, since the antiserum raised against the purified protein did not inhibit adherence to cultured cells. PCR-RFLP analysis of amplified flagellin gene products and Southern analysis revealed inter-strain heterogeneity; this could be useful for epidemiological and phylogenetic studies of this organism.