Vegetative Clostridium difficile survives in room air on moist surfaces and in gastric contents with reduced acidity: a potential mechanism to explain the association between proton pump inhibitors and C. difficile-associated diarrhea?

Abstract

Proton pump inhibitors (PPIs) have been identified as a risk factor for Clostridium difficile-associated diarrhea (CDAD), though the mechanism is unclear because gastric acid does not kill C. difficile spores. We hypothesized that the vegetative form of C. difficile, which is killed by acid, could contribute to disease pathogenesis if it survives in room air and in gastric contents with elevated pH. We compared the numbers of C. difficile spores and vegetative cells in stools of patients prior to and during the treatment of CDAD. We assessed the survival of vegetative cells on moist or dry surfaces in room air versus anaerobic conditions and in human gastric contents, in pH-adjusted gastric contents, and in gastric contents from individuals receiving PPI therapy. Stool samples obtained from patients prior to the initiation of antibiotic treatment for C. difficile contained approximately 10-fold more vegetative cells than spores. On dry surfaces, vegetative C. difficile cells died rapidly, whereas they remained viable for up to 6 h on moist surfaces in room air. Vegetative C. difficile cells had only marginal survival in gastric contents at low pH; adjustment to a pH of >5 resulted in survival similar to that in the phosphate-buffered saline control. The survival of vegetative C. difficile in gastric contents obtained from patients receiving PPIs was also increased at a pH of >5. The ability of the vegetative form of C. difficile to survive on moist surfaces and in gastric contents with an elevated pH suggests a potential mechanism by which PPI therapy could increase the risk of acquiring C. difficile.

FIG. 1.

Quantification of vegetative cells and spores (PBS) versus spores alone (ethanol) in cytotoxin-positive feces obtained from patients untreated (n = 26) or while on treatment (n = 16) for CDAD. Error bars indicate SDs. *, P < 0.001.

FIG. 2.

Survival rates of vegetative C. difficile after 1 h of exposure to unadjusted gastric contents (GC), to gastric contents adjusted to pH 4 to 7, or to PBS. Pooled data from four strains of C. difficile are shown. Gastric contents that were grossly bilious were excluded. Error bars indicate SDs. *, P < 0.0001 versus survival rate in gastric contents at pH 4; †, P < 0.0001 versus survival rate in gastric contents at pH 5; ‡, P < 0.0001 versus survival rates in gastric contents at pH 6 and 7 and in PBS.

FIG. 3.

Survival rates of vegetative C. difficile after 1 h of exposure to gastric contents (GC) from individuals taking PPIs. Results for individual donors are arranged by increasing pH of gastric contents. Pooled data from four strains of C. difficile are shown. Error bars indicate SDs. An asterisk denotes a grossly bilious sample.