Reuterin disrupts Clostridioides difficile metabolism and pathogenicity through reactive oxygen species generation

Gut Microbes. 2020 Nov 9;12(1):1788898. doi: 10.1080/19490976.2020.1795388.


Antibiotic resistance is one of the world's greatest public health challenges and adjunct probiotic therapies are strategies that could lessen this burden. Clostridioides difficile infection (CDI) is a prime example where adjunct probiotic therapies could decrease disease incidence through prevention. Human-derived Lactobacillus reuteri is a probiotic that produces the antimicrobial compound reuterin known to prevent C. difficile colonization of antibiotic-treated fecal microbial communities. However, the mechanism of inhibition is unclear. We show that reuterin inhibits C. difficile outgrowth from spores and vegetative cell growth, however, no effect on C. difficile germination or sporulation was observed. Consistent with published studies, we found that exposure to reuterin stimulated reactive oxygen species (ROS) in C. difficile, resulting in a concentration-dependent reduction in cell viability that was rescued by the antioxidant glutathione. Sublethal concentrations of reuterin enhanced the susceptibility of vegetative C. difficile to vancomycin and metronidazole treatment and reduced toxin synthesis by C. difficile. We also demonstrate that reuterin is protective against C. difficile toxin-mediated cellular damage in the human intestinal enteroid model. Overall, our results indicate that ROS are essential mediators of reuterin activity and show that reuterin production by L. reuteri is compatible as a therapeutic in a clinically relevant model.

Keywords: Clostridioides difficile; Lactobacillus reuteri; Reuterin; enteroids; metabolism; organoids; oxidative stress; probiotics; reactive oxygen species.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Anti-Bacterial Agents / pharmacology
  • Clostridioides difficile / drug effects*
  • Clostridioides difficile / growth & development
  • Clostridioides difficile / metabolism
  • Clostridioides difficile / pathogenicity
  • Drug Synergism
  • Epithelial Cells / drug effects
  • Epithelial Cells / microbiology
  • Glyceraldehyde / analogs & derivatives*
  • Glyceraldehyde / metabolism
  • Glyceraldehyde / pharmacology
  • Humans
  • Limosilactobacillus reuteri / metabolism
  • Organoids / drug effects
  • Organoids / microbiology
  • Oxidative Stress / drug effects
  • Probiotics / metabolism
  • Propane / metabolism
  • Propane / pharmacology*
  • Reactive Oxygen Species / metabolism*
  • Spores, Bacterial / drug effects
  • Spores, Bacterial / growth & development


  • Anti-Bacterial Agents
  • Reactive Oxygen Species
  • 3-hydroxypropionaldehyde
  • Glyceraldehyde
  • Propane