Background. Clostridium difficile infection (CDI) is a leading cause of infectious diarrhea 1, and the Centers for Disease Control and Prevention has listed C. difficile as an urgent threat to antimicrobial resistance 2. Currently vancomycin is used as first line therapy, although approximately 25% of patients relapse after treatment and often require fecal microbial transplantation (FMT) 3. C. difficile pathogenicity requires the activities of its toxins, TcdA and TcdB 4, but the T cell-mediated response to these toxins remains uncharacterized.
Methods. We enrolled a cohort of patients with newly acquired CDI, a cohort with relapsing CDI, and healthy volunteers with no history of CDI. Toxin-specific CD4+ T cell responses were measured using a whole blood flow cytometry assay that measures induced co-expression of CD25 and OX40 following 44h incubation with antigen.
Results. In patients with recurring CDI, CD4+ T cell responses to TcdB, but not TcdA, were significantly higher than for healthy controls and newly acquired CDI. Notably, levels of anti-TcdA/TcdB IgG or IgA antibodies were not different between patients and controls. In both patient cohorts, TcdB-specific CD4+ T cells were functionally heterogeneous, although strongly biased towards a Th17 cell phenotype. Interestingly, TcdB-specific Th17 cells were significantly reduced in recurring, compared to newly acquired, CDI and did not change post-FMT. We observed the TCR repertoire of TcdB-specific CD4+ T cells was polyclonal and we have identified several HLA-DR7 and DR15 restricted TcdB epitopes recognised by these cells.
Conclusion. This is the first investigation of T cell immunity to C. difficile toxins, and identifies anti-TcdB CD4+ T cells, but not anti-TcdB antibody levels, as a marker of patients with active disease. Analysing toxin-specific CD4+ T cell responses has the potential to predict relapse and provides insight into how CD4+ T cell memory develops in response to a prevalent bacterial pathogen.