Clostridium difficile Modulates the Gut Microbiota by Inducing the Production of Indole, an Interkingdom Signaling and Antimicrobial Molecule

Abstract

Clostridium (Clostridioides) difficile infection (CDI) is associated with dysbiosis. C. difficile has a characteristic propensity to persist and recur 1 to 4 weeks after treatment, but the mechanism is unknown. We hypothesized that C. difficile may persist by manipulating the intestinal microenvironment, thereby hampering gut microbiota reconstitution following antibiotic-mediated dysbiosis. By screening stools from CDI patients for unique markers, a metabolite identified to be indole by mass spectrometry and Fourier transform infrared spectroscopy was identified. The average fecal indole concentration detected in CDI patients (n = 216; mean, 1,684.0 ± 84.4 µM) was significantly higher than in stools of patients with non-C. difficile diarrhea (n = 204; mean, 762.8 ± 53.8 µM). Certain intestinal bacteria, but not C. difficile, produce indole, a potent antimicrobial antioxidant. Remarkably, C. difficile induced other indole-producing gut microbes to produce increasing amounts of indole. Furthermore, a C. difficile accessory gene regulator 1 quorum sensing system mutant cannot induce indole, but complementation of the mutant strain with the wild-type gene restored its ability to induce indole production. Indole tolerance assays indicated that the amount of indole required to inhibit growth of most gut-protective bacteria was within the range detected in the CDI stools. We think that a high indole level limits the growth of beneficial indole-sensitive bacteria in the colon and alters colonization resistance and this might allow C. difficile to proliferate and persist. Together, these results reveal a unique mechanism of C. difficile persistence and provide insight into complex interactions and chemical warfare among the gut microbiota. IMPORTANCE Clostridium difficile infection is the leading cause of hospital-acquired and antibiotic-associated diarrhea worldwide. C. difficile flourishes in the colon after the diversity of the beneficial and protective gut microbiota have been altered by antibiotic therapy. C. difficile tends to persist, as does dysbiosis, encouraging recurrence a few days to weeks after treatment, and this further complicates treatment options. Here, we show that C. difficile might persist by manipulating the indigenous microbiota to produce indole, a bioactive molecule that inhibits the growth and reconstitution of the protective gut microbiota during infection. This discovery may explain a unique strategy C. difficile uses to control other bacteria in the colon and provide insight into the complex interactions and chemical warfare among the gut microbiota.

Dissecting individual pathogen-commensal interactions within a complex gut microbiota community

“…Small but significant increases in the numbers of B. dorei, B. ovatus, E. coli, F. prausnitzii , and R. gnavus was observed when C. difficile was cocultured with the commensal community. Although of the nine species, B. ovatus, B. thetaiotaomicron, B. adolescentis, E. coli and F. prausnitzii are indole producers 57–60 , no inhibitory effects were seen. Given that C. difficile was instead inhibited, it is possible that the bacteria were unable to reach sufficient numbers to influence indole production.…”

“…In a successful infection, C. difficile has been reported to control the microbiota by modulating bacterial metabolism, including the production of indole 56,57 . It does this through influencing the expression of tryptophanase (tnaA) in other species; C. difficile is thought to limit the recovery of the microbiota through indole-mediated inhibition of growth of protective gut bacteria 57 . Small but significant increases in the numbers of B. dorei, B. ovatus, E. coli, F. prausnitzii , and R. gnavus was observed when C. difficile was cocultured with the commensal community.…”

Flexible cobamide metabolism in Clostridioides (Clostridium) difficile 630 Δerm

“…In a biassociation, Bacteroides thetaiotaomicron can break down host mucin and produce sialic acid, which can be used by C. difficile for expansion in the gut (19). C. difficile can also induce other members of the microbiota to produce indole, which is thought to create a more favorable environment for the pathogen by inhibiting competing microbes (20).…”

Biphasic chemotaxis of Escherichia coli to the microbiota metabolite indole

“…Indole promotes homeostasis in the microbial community of the GI tract by inhibiting virulence and colonization of host cells by several pathogens, including enterohemorrhagic E. coli (24,25), Salmonella enterica (22,26), and Vibrio cholerae (27). Indole also promotes phenotypes linked to virulence and colonization in other pathogens such as Pseudomonas aeruginosa and Clostridium difficile (23,28,29). These varying effects of indole probably depend on its concentrations, which can fluctuate over a wide range (0.2 to 6.5 mM) in the GI tract (30).…”