Active and secretory IgA-coated bacterial fractions elucidate the dysbiosis in Clostridium difficile infection

The onset of C. difficile infection (CDI) has been associated with the treatment with wide-spectrum antibiotics. Antibiotic treatment alters the activity of gut commensals and it may result in modified patterns of immune responses to pathogens. To study these mechanisms during CDI, we separated bacteria with high cellular RNA content (the active bacteria) and their inactive counterparts by fluorescence activated cell sorting (FACS) of the faecal bacterial suspension. The gut dysbiosis due to the antibiotic treatment may results in modification of immune recognition of intestinal bacteria. The immune recognition patterns were assessed by FACS of bacterial fractions either coated or not by intestinal secretory immunoglobulin A (SIgA). We described the taxonomic distributions of these four bacterial fractions (active vs. inactive and SIgA-coated vs. non-SIgA-coated) by massive 16S rDNA amplicon sequencing and quantified the proportion of C. difficile toxin genes in the samples. The overall gut microbiome composition was more robustly influenced by antibiotics than by the proper C. difficile toxins. Bayesian networks revealed that the C. difficile cluster was preferentially SIgA-coated during CDI. In contrast, in the CDI- group Fusobacterium was the characteristic genus of SIgA-opsonized fraction. Lactobacillales and Clostridium cluster IV were mostly inactive in CDI+ patients. In conclusion, despite the proportion of C. difficile in the gut is very low, it is able to initiate infection during the gut dysbiosis caused by environmental stress (antibiotic treatment) as a consequence of decreased activity of protecting bacteria.