Background: Asymptomatic  C. difficile colonization is believed to predispose to subsequent C.  difficile infection (CDI). While emerging insights into the role of the  commensal microbiota in mediating colonization resistance against C.  difficile have associated CDI with specific microbial components,  corresponding prospectively collected data on colonization with C.  difficile are largely unavailable.

Methods: C.  difficile status was assessed by GDH EIA and real-time PCR targeting the  toxin A (tcdA) and B (tcdB) genes. 16S V3 and V4 gene sequencing  results from fecal samples of patients tested positive for C. difficile  were analyzed by assessing alpha and beta diversity, LefSe, and the  Piphillin functional inference approach to estimate functional capacity.

Results: 1506  patients were recruited into a prospective observational study  (DRKS00005335) upon admission into one of five academic hospitals. 936  of them provided fecal samples on admission and at discharge and were  thus available for longitudinal analysis. Upon hospital admission, 5.5%  (83/1506) and 3.7% (56/1506) of patients were colonized with toxigenic  (TCD) and non-toxigenic C. difficile (NTCD), respectively. During  hospitalization, 1.7% (16/936) acquired TCD. Risk factors for  acquisition of TCD included pre-existing lung diseases, lower GI  endoscopy and antibiotics. Species protecting against hospital-related  C. difficile acquisition included Gemmiger spp., Odoribacter  splanchnicus, Ruminococcus bromii and other Ruminococcus spp..  Metagenomic pathway analysis identified steroid biosynthesis as the most  underrepresented metabolic pathway in patients who later acquire C.  difficile colonization.

Conclusions: Gemmiger  spp., Odoribacter splanchnicus, Ruminococcus bromii and other  Ruminococci were associated with a decreased risk of C. difficile  acquisition.

Keywords: Gemmiger spp; bile acids; microbiota; toxigenic C. difficile.