Dual Screen for Gut Metabolites Suppressing Enterobacterial Growth and Invasiveness Reveals Structure – Activity Relationships among Anti-Infective Indoles

The antibiotic resistance crisis makes characterization of new anti-infective molecules a pressing matter. Molecules that suppress bacterial growth and survival, virulence, or the combination of these traits, all warrant further exploration. Naturally occurring microbe-host ecosystems, such as the human gut, provide incompletely tapped resources in this regard. We developed a flexible platform to parallelly assess how gut metabolites affect the growth and epithelial cell invasion capacity of the enteropathogens Salmonella enterica Typhimurium ( Salmonella ) and Shigella flexneri . By screening a gut metabolite library, the assays identified multiple anti-infective compound classes and extended previously reported antibacterial activities for e.g. medium chain fatty acids, bile acids, purine nucleotides, and indole. Importantly, a targeted follow-up screen combined with chemical biology iterations showed how the anti-infective activity of indole is impacted by its derivatization. Specifically, a methyl group at either of the carbons of the indole scaffold potentiated the suppressive effect on type-III-secretion-system-mediated virulence, flagellar motility (for Salmonella ), and growth, in a concentration-dependent manner. By contrast, N1-methylation markedly attenuated the activity of indole and its C-derivatized versions. The study, hence, offers assays for dual growth and virulence analysis of invasive enterobacteria exposed to anti-infective candidate molecules, and informs on structure–activity relationships among indole metabolites.