Dual screen for gut metabolites suppressing enterobacterial growth and invasiveness reveals structure–activity relationships among anti-infective indoles

Abstract

The antibiotic resistance crisis has made the characterization of new anti-infective molecules a pressing matter. Molecules that suppress bacterial growth and survival, virulence, or a 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, e.g., for medium-chain fatty acids, bile acids, purine nucleotides, and indoles. 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 its concentration-dependent suppressive effect on type-III-secretion-system virulence protein expression, flagellar motility (for Salmonella), and enterobacterial growth. By contrast, N1-methylation markedly attenuated the activity of indole and its C-derivatized versions. This study, hence, offers assays for dual growth and virulence analysis of invasive enterobacteria exposed to anti-infective candidate molecules, and informs on the structure–activity relationships among indole metabolites.