Combinatorial effects of tryptophan derivatives serotonin and indole on virulence modulation of enteric pathogens

There is a wealth of signals present in the human gut that mediate host-microbiota communication and intersect with the gut-brain-axis. There is differential spatial localization of the tryptophan derivatives serotonin and indole in the gut, which are important cues for enteric pathogens to find their colonization niche. Both signals are sensed by enteric pathogens such as enterohemorrhagic <i>Escherichia coli</i> (EHEC) and <i>Citrobacter rodentium</i>, a murine pathogen extensively employed as a surrogate animal model for EHEC. EHEC and <i>C. rodentium</i> virulence determinants include genes necessary for the attaching and effacing (AE) lesion formation on enterocytes, which are contained within the locus of enterocyte effacement (LEE) pathogenicity island. Both signals inhibit LEE expression <i>in vitro</i> and during mammalian infection. The roles of indole and serotonin in virulence have been initially interrogated separately. Here, we show the combinatorial effect of these signals antagonizing each other's activity on bacterial virulence <i>in vitro</i> and during murine infection. The role of both signals <i>in vivo</i> was interrogated by manipulating the serotonin levels in mice through pharmacological and genetic strategies to increase the levels of serotonin in the gut by inhibiting the serotonin reuptake transporter (SERT). Simultaneously, indole levels were altered through infection with <i>C. rodentium</i> strains that either produce or don't indole or recolonizing the microbiota with WT or Δ<i>tnaA</i> (does not produce indole) strains of <i>Bacteroides thetaiotaomicron</i>. Individually increasing the levels of serotonin and indole in the gut decreased <i>C. rodentium</i> pathogenesis. However, when both signals were elevated simultaneously, they antagonized each other's activity.IMPORTANCEPathogens sense a plethora of signals within the gut to successfully establish colonization by precise regulation of virulence gene expression within the right niche. Our study shows that it is crucial to not disregard the interaction of different signaling mechanisms to understand the complexity of virulence regulation in enteric pathogens. Even though serotonin and indole are both tryptophan derivatives with similar structures that individually decrease bacterial virulence, combinatorial sensing of these two signals cancels out each other's effect. Understanding these sensing mechanisms provides a better insight into potential therapeutic approaches against enteric infections.