The flagellin FliC of Clostridium difficile is responsible for pleiotropic gene regulation during in vivo infection

Clostridium difficile is the main agent responsible for hospital acquired antibiotic associated diarrhoea. In recent years, epidemic strains have emerged causing more severe infections. Whilst C. difficile has two major virulence factors, toxins TcdA and TcdB, it is generally accepted that other virulence components of the bacterium contribute to disease. Previously, it has been suggested that flagella expression from pathogenic bacteria might be implicated in virulence. In a recent study, we observed an increased mortality in a gnotobiotic mouse model when animals were colonized with an isogenic fliC mutant constructed in the PCR-ribotype 027 (B1/NAP1) strain R20291, while animals survived when colonized by the parental strain or after colonization by other high-toxin-producing C. difficile strains. To understand the reasons for this increased virulence, we compared the global gene expression profiles between the fliC-R20291 mutant and its parental strain using an in vitro and in vivo transcriptomic approach. The latter made use of the gnotobiotic mouse model. Interestingly, in the fliC mutant, we observed considerable up-regulation of genes involved in mobility, membrane transport systems (PTS, ABC transporters), carbon metabolism, known virulence factors and sporulation. A smaller but significant up-regulation of genes involved in cell growth, fermentation, metabolism, stress and antibiotic resistance was also apparent. All of these genes may be associated with the increased virulence of the fliC-R20921 mutant. We confirmed that the fliC mutation is solely responsible for the observed changes in gene expression in the mutant strain since expression profiles were restored to that of the wild-type strain in the fliC-complemented strain. Thus, the absence of FliC is directly or indirectly involved in the high mortality observed in the fliC mutant infected animals. Therefore, we provide the first evidence that when the major structural component of the flagellum is neutralized, deregulation of gene expression can occur during infection

Known and putative virulence factor genes differentially expressed <i>in vivo i</i>n the <i>fliC</i> mutant compared to strain R20291.

<p>ND: non determined.</p

Flagellar genes differentially expressed <i>in vivo</i> in the <i>fliC</i> mutant.

<p>*From F1 region (late-stage flagellar genes) and **F3 region (early-stage flagellar genes) of the flagellar operon from <i>C. difficile</i> 630 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0096876#pone.0096876-Aubry1" target="_blank">[39]</a>.</p

Functional clusters of <i>in vivo</i> differentially expressed genes.

<p>Differentially expressed genes in the <i>fliC</i> mutant compared to wild-type R20291 from caeca of 14 h post-infection mice were classified in functional groups according to their involvement in the biological process categories. (<b>A</b>) The number of analyzed genes is represented in green bars and the number of significantly differentially expressed genes is shown in black bars. Percentages of differentially regulated genes are indicated at right. (<b>B</b>) The numbers of up- and down-regulated genes for each cluster are indicated in green and black bars, respectively.</p

Membrane transport genes highly differentially expressed <i>in vivo</i> in the <i>fliC</i> mutant compared to strain R20291.

<p>Membrane transport genes highly differentially expressed <i>in vivo</i> in the <i>fliC</i> mutant compared to strain R20291.</p

Ability of sporulation of the <i>fliC</i> mutant compared to wild-type R20291 <i>in vivo</i> and <i>in vitro</i>.

<p>(<b>A</b>) Groups of 6 axenic mice were infected by oral gavage route with 1×10⁸<i>C. difficile</i> CFU. The <i>C. difficile</i> faecal vegetative cells and spores were measured by determining the concentration of CFU in faeces at 14 h post-infection by homogenising and plating on appropriated agar medium after heat shock (for spores) or not (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0096876#s4" target="_blank">Materials and Methods</a>). Data represent the ratio of spores per vegetative cells. (<b>B</b>) Cultures in BHIS broth in anaerobic conditions were prepared from 2 successive subcultures as indicated in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0096876#s4" target="_blank">Materials and Methods</a>. After heat shock, spores were quantified (CFU/ml) by performing serial dilutions and spread plating on BHIS agar supplemented with 0.1% bile salt taurocholate to induce germination. Data represent the ratio of spores per total cells. The presented values are the mean of 3 different cultures. Statistically significant difference is indicated by * for p<0,05.</p

Les archives de l’invention

Les historiens ont longtemps privilégié le facteur technique dans l’approche des révolutions industrielles. Dans cette logique monocausale, le progrès technique était assimilé à une succession d’inventions apparues dans des secteurs pionniers, moteurs de la croissance, entraînant le reste de l’économie, dite traditionnelle, dans son sillage. L’un des paradoxes de cette approche consistait à valoriser l’innovation tout en évitant d’interroger les pratiques inventives. La dynamique interne du progrès technique et les traits de génie des inventeurs tenaient lieu de modèles explicatifs. La remise en cause de ces approches suscite bien des interrogations de méthode. Comment repérer les formes de l’invention ordinaire, en cerner les acteurs ? Comment assigner une origine à des nouveautés dont l’antériorité se perd dans la mémoire commune ? Comment appréhender des savoirs pratiques instables et non codifiés que ne livrent pas les corpus constitués de sources ? Comment concilier les définitions construites de l’invention et de l’inventeur, les catégories déjà forgées par les institutions et le corps social, et les mentions informelles ou indirectes de l’invention ? Ces questions débordent l’écrit. Cet ouvrage, issu d’un colloque international tenu à Paris en 2003, élargit le concept de sources : au-delà des « sources-textes », il considère les dessins, les enregistrements sonores, les instruments et outils, les installations, les échantillons, les modèles, les prototypes, etc. Il propose une réflexion originale sur le statut des archives de l’invention, sur leur mode de production et sur les méthodologies mises en œuvre dans leur exploitation

Staphylococcus capitis isolated from bloodstream infections: a nationwide 3-month survey in 38 neonatal intensive care units

International audienceTo increase the knowledge about S. capitis in the neonatal setting, we conducted a nationwide 3-month survey in 38 neonatal intensive care units (NICUs) covering 56.6% of French NICU beds. We demonstrated 14.2% of S. capitis BSI (S.capBSI) among nosocomial BSIs. S.capBSI incidence rate was 0.59 per 1000 patient-days. A total of 55.0% of the S.capBSIs were late onset catheter-related BSIs. The S. capitis strains infected preterm babies (median gestational age 26 weeks, median birth weight 855 g). They were resistant to methicillin and aminoglycosides and belonged to the NRCS-A clone. Evolution was favorable in all but one case, following vancomycin treatment