Clostridium difficile infection (CDI) is typically associated with disturbed gut microbiota and changes related to decreased colonization resistance against C. difficile are well described.
However, nothing is known about possible effects of C. difficile on gut microbiota restoration during or after CDI.
In this study, we have mimicked such a situation by using C. difficile conditioned medium of six different C. difficile strains belonging to PCR ribotypes 027 and 014/020 for cultivation of fecal microbiota.
A marked decrease of microbial diversity was observed in conditioned medium of both tested ribotypes. The majority of differences occurred within the phylum Firmicutes, with a general decrease of gut commensals with putative protective functions (i.e. Lactobacillus, Clostridium_XIVa) and an increase in opportunistic pathogens (i.e. Enterococcus). Bacterial populations in conditioned medium differed between the two C. difficile ribotypes, 027 and 014/020 and are likely associated with nutrient availability. Fecal microbiota cultivated in medium conditioned by E. coli, Salmonella Enteritidis or Staphylococcus epidermidis grouped together and was clearly different from microbiota cultivated in C. difficile conditioned medium suggesting that C. difficile effects are specific.
Our results show that the changes observed in microbiota of CDI patients are partially directly influenced by C. difficile.
Clostridium difficile is a nosocomial pathogen prevalent in hospitals worldwide and increasingly common in the community.
Sequence differences have been shown to be present in the Surface Layer Proteins (SLPs) from different C. difficileribotypes (RT) however whether these differences influence severity of infection is still not clear.
We used a molecular evolutionary approach to analyse SLPs from twenty-sixC. difficile RTs representing different slpA sequences. We demonstrate that SLPs from RT 027 and 078 exhibit evidence of positive selection (PS).
We compared the effect of these SLPs to those purified from RT 001 and 014, which did not exhibit PS, and demonstrate that the presence of sites under positive selection correlates with ability to activate macrophages.
SLPs from RTs 027 and 078 induced a more potent response in macrophages, with increased levels of IL-6, IL-12p40, IL-10, MIP-1α, MIP-2 production relative to RT 001 and 014. Furthermore, RTs 027 and 078 induced higher expression of CD40, CD80 and MHC II on macrophages with decreased ability to phagocytose relative to LPS.
These results tightly link sequence differences in C. difficile SLPs to disease susceptibility and severity, and suggest that positively selected sites in the SLPs may play a role in driving the emergence of hyper-virulent strains.