Abundant coating of Clostridium difficile by unspecific intestinal IgA in infected patients gut. active-IgAcoated-CDI

C. difficile infection (CDI) is particularly associated with antibiotic treatment, which causes microbial dysbiosis in the gut. However, when microbial composition of CDI+ and CDI- fecal samples are compared in metagenomic studies, C. difficile is not differentiating the two groups. Antibiotics act on bacterial activity (by blocking or killing) and influence the way how the immune system recognize pathogens. In this work we aimed to clarify the onset of CDI, by studying active and IgA-opsonized bacterial fractions of total faecal microbiota. The cohort included patients under multiple antibiotic treatment or no antibiotics at all; 12 of them were CDI positive and 12 CDI negative. Fluorescence activated cell sorting was used to separate bacteria into active and inactive fractions (active-F and inactive-F) and into IgA-opsonized or not. Then 16S rRNA gene amplicons from the four fractions of each of the 24 patients were characterized by Illumina sequencing. The bacterial composition of the four fractions significantly differed (p-value < 0.05). The taxonomical distributions of active was influenced more by the type of antibiotics than by the load of C. difficile toxins. For example, Enterococcus sp. was one of the determinant species of the active bacteria (p-value < 0.01) of patients who took anti-CDI antibiotics. The composition of IgA-pos-F was shaped by antibiotics and also by recurrence of CDI and ulcerative colitis. The proportion of the C. difficile cluster opsonized by IgA was significantly increased (p-value < 0.01) in CDI positive patients. In constrast, in CDI negative patients C. difficile cells were significatively represented in the fraction without IgA opsonization. In conclusion, in CDI positive patients, the infection was mainly driven by the action of immune system, independently on the overall microbial composition which is, on the other side, shaped by the proper antibiotics treatment.