RESUMO
A prominent oral commensal and opportunistic pathogen, Fusobacterium nucleatum can traverse to extra-oral sites such as placenta and colon, promoting adverse pregnancy outcomes and colorectal cancer, respectively. How this anaerobe sustains many metabolically changing environments enabling its virulence potential remains unclear. Informed by our genome-wide transposon mutagenesis, we report here that the highly conserved Rnf complex, encoded by the rnfCDGEAB gene cluster, is key to fusobacterial metabolic adaptation and virulence. Genetic disruption of the Rnf complex via non-polar, in-frame deletion of rnfC (Δ rnfC ) abrogates polymicrobial interaction (or coaggregation) associated with adhesin RadD and biofilm formation. The defect in coaggregation is not due to reduced cell surface of RadD, but rather an increased level of extracellular lysine, which binds RadD and inhibits coaggregation. Indeed, removal of extracellular lysine via washing Δ rnfC cells restores coaggregation, while addition of lysine inhibits this process. These phenotypes mirror that of a mutant (Δ kamAΔ ) that fails to metabolize extracellular lysine. Strikingly, the Δ rnfC mutant is defective in ATP production, cell growth, cell morphology, and expression of the enzyme MegL that produces hydrogen sulfide from cysteine. Targeted metabolic profiling demonstrated that catabolism of many amino acids, including histidine and lysine, is altered in Δ rnfC cells, thereby reducing production of ATP and metabolites including H2S and butyrate. Most importantly, we show that the Δ rnfC mutant is severely attenuated in a mouse model of preterm birth. The indispensable function of Rnf complex in fusobacterial pathogenesis via modulation of bacterial metabolism makes it an attractive target for developing therapeutic intervention.
RESUMO
Campylobacter jejuni is a leading cause of bacterially derived gastroenteritis worldwide. Campylobacter is most commonly acquired through the consumption of undercooked poultry meat or through drinking contaminated water. Following ingestion, Campylobacter adheres to the intestinal epithelium and mucus layer, causing toxin-mediated inflammation and inhibition of fluid reabsorption. Currently, the human response to infection is relatively unknown, and animal hosts that model these responses are rare. As such, we examined patient fecal samples for the accumulation of the neutrophil protein calgranulin C during infection with Campylobacter jejuni In response to infection, calgranulin C was significantly increased in the feces of humans. To determine whether calgranulin C accumulation occurs in an animal model, we examined disease in ferrets. Ferrets were effectively infected by C. jejuni, with peak fecal loads observed at day 3 postinfection and full resolution by day 12. Serum levels of interleukin-10 (IL-10) and tumor necrosis factor alpha (TNF-α) significantly increased in response to infection, which resulted in leukocyte trafficking to the colon. As a result, calgranulin C increased in the feces of ferrets at the time when C. jejuni loads decreased. Further, the addition of purified calgranulin C to C. jejuni cultures was found to inhibit growth in a zinc-dependent manner. These results suggest that upon infection with C. jejuni, leukocytes trafficked to the intestine release calgranulin C as a mechanism for inhibiting C. jejuni growth.