The Role of Flagella in Clostridium difficile Pathogenesis: Comparison between a Non-Epidemic and an Epidemic Strain

Abstract

Clostridium difficile is a major cause of healthcare-associated infection and inflicts a considerable financial burden on healthcare systems worldwide. Disease symptoms range from self-limiting diarrhoea to fatal pseudomembranous colitis. Whilst C. difficile has two major virulence factors, toxin A and B, it is generally accepted that other virulence components of the bacterium contribute to disease. C. difficile colonises the gut of humans and animals and hence the processes of adherence and colonisation are essential for disease onset. Previously it has been suggested that flagella might be implicated in colonisation. Here we tested this hypothesis by comparing flagellated parental strains to strains in which flagella genes were inactivated using ClosTron technology. Our focus was on a UK-outbreak, PCR-ribotype 027 (B1/NAP1) strain, R20291. We compared the flagellated wild-type to a mutant with a paralyzed flagellum and also to mutants (fliC, fliD and flgE) that no longer produce flagella in vitro and in vivo. Our results with R20291 provide the first strong evidence that by disabling the motor of the flagellum, the structural components of the flagellum rather than active motility, is needed for adherence and colonisation of the intestinal epithelium during infection. Comparison to published data on 630Δerm and our own data on that strain revealed major differences between the strains: the R20291 flagellar mutants adhered less than the parental strain in vitro, whereas we saw the opposite in 630Δerm. We also showed that flagella and motility are not needed for successful colonisation in vivo using strain 630Δerm. Finally we demonstrated that in strain R20291, flagella do play a role in colonisation and adherence and that there are striking differences between C. difficile strains. The latter emphasises the overriding need to characterize more than just one strain before drawing general conclusions concerning specific mechanisms of pathogenesis.

The Phosphotransfer Protein CD1492 Represses Sporulation Initiation in Clostridium difficile

“…7C). The timing and expression of sigD and flagellar genes during infection affect colonization by and the virulence of C. difficile and other motile pathogens and thus probably contribute to the decreased virulence of the CD1492 mutant in the animal model (43)(44)(45)(46)(47).…”

The HtrA-Like Protease CD3284 Modulates Virulence of Clostridium difficile

“…The spo0A gene, encoding the master regulator of sporulation (59, 60), was 2.4-fold and 2.3-fold downregulated in the logarithmic and stationary growth phases, respectively, and many other sporulation genes showed a similar effect (see Table S2 in the supplemental material). Furthermore, several genes encoding surface-associated proteins (such as slpA and 5 other paralogs of slpA) (16), flagellum-associated genes (18,61), and cd1581 (a gene with a potential effect on adhesion and intestinal colonization) (19) were significantly downregulated.…”

“…Similarly, there is a clear link between motility and toxin production, as the alternative sigma factor SigD is required for both (81). Interestingly, flagellar operon mutants showed changes in both toxin levels (82) and their capacity to adhere to Caco-2 cells (61). Finally, more general effects of stress (nutrient depletion or temperature) on toxin levels were noted (83,84).…”

The role of flagella in Clostridium difficile pathogenicity

“…This diversity is believed to account for the differences in motility between C. difficile 630 and R20291 [15]; Stabler et al [15] observed that C. difficile 630 was less motile than R20291. It is pertinent to note that differences in motility and number of flagella have been observed between clonal isolates of C. difficile R20291 [12,15,22]. The study of Stabler et al [15] revealed R20291 to be more motile than 630; however, research by Baban et al [12] has shown R20291 to be less motile than 630 and to have one polar flagellum, as opposed to Martin et al [22], who observed multiple flagella in the R20291 strain.…”

“…However, the contribution of flagella to the pathogenesis of C. difficile is complex and not yet fully understood. There is increasing scientific evidence to suggest that flagella play a more direct role in virulence, via modulation of toxin expression [4,12] rather than simply providing force-driven motility towards nutrients available in the gut. In this review the importance of flagella to the pathogenicity of C. difficile is discussed.…”