Microbiome-Targeted Interventions for Colitis-Associated Bacteria

Abstract

Complex interactions between mammalian hosts and their gut microbes have evolved over many millennia and have established a sophisticated communication system that is essential for symbiosis and mutualism. Perturbations to host-microbiota homeostasis in the context of a genetically susceptible host are central to the development of inflammatory bowel disease (IBD). In-depth understanding of the underlying mechanisms that control homeostasis and dysbiosis are essential for determining how to reliably modulate the host-microbiota continuum to prevent and treat disease. However, deciphering whether alterations in the microbiota are a cause or consequence of IBD remains a considerable challenge, as is defining the role of specific microbes in the pathogenesis of disease. This thesis explores the gut microbiome in mouse models of experimental colitis and evaluates the contribution of specific microbial clades and pathways in potentiating mucosal inflammation with the goal of identifying novel microbiome-targeted interventions for disease management.

To improve our understanding of microbial dysbiosis and dysfunction in IBD, we use the TRUC (T-bet-/-RAG2-/- ulcerative colitis) mouse model to profile the gut microbiome in active disease versus treatment-induced remission. 16S ribosomal RNA gene surveys of stool from mice treated with antibiotics, immunodulatory therapies, or a fermented-milk dietary intervention reveal microbial features modified among health and disease states. Discriminatory biomarkers of active disease included increased Enterobacteriaceae and shifts from carbohydrate and energy metabolism to pathways favoring bacterial pathogenesis, specifically cell motility and two-component systems. An unexpected observation is a significant enrichment in genes for microbial benzoate degradation in active colitis. Intermediates of benzoate metabolism – catechols – share the same backbone as host catecholamines, which can signal through two-component systems to promote virulence in pathogenic Enterobacteriaceae. Based on expansions in Enterobacteriaceae and increased gene abundances for benzoate degradation, two-component systems, and bacterial motility proteins, we identify a potential signaling axis linking host adrenergic stress with enhanced bacterial virulence in a preclinical model of colitis.

Enterobacteriaceae sense and respond to microbiota-generated signals and host-derived catecholamines through the QseBC two-component quorum sensing system. QseC is a membrane-bound sensor kinase that surveys the external milieu and, upon signal detection, activates its cognate response regulator, QseB, to induce expression of virulence genes. To investigate whether blocking QseC signaling could reduce disease severity, we test the effects of a QseC inhibitor (LED209) in the TRUC, Il-10-/-, and dextran sodium-sulfate models of experimental colitis. LED209 attenuates disease across all three models, with the most striking protective effect in TRUC and dextran sodium sulfate-treated mice. LED209 also prevents the expansion of Enterobacteriaceae in Il-10-/-and dextran sodium-sulfate-exposed mice, but not in TRUC mice, indicating a potential difference in microbiota responses based on genetic context. Moreover, measuring catecholamines in cecal content and stool show that LED209 does not significantly affect the luminal catecholamine pool and thus, may not disrupt host or microbial catecholamine metabolism. Collectively, these data show that QseC inhibition can ameliorate disease in distinct models of experimental colitis and suggest a role for QseC-mediated bacterial virulence in the pathogenesis of IBD.

Although a single pathogen has not been identified as a causative agent, several bacteria continue to be implicated in the initiation and progression of IBD, including adherent-invasive Escherichia coli (AIEC). As a proof-of-principle, we genetically inactivate qseC in the Crohn’s disease-associated AIEC strain LF82. We show that absence of qseC leads to downregulated virulence gene expression and defects in flagellar assembly and motility in vitro and reduced colonization efficiency in vivo. Overall, these studies provide evidence that QseC may be an upstream virulence node utilized by colitogenic bacteria to survey their host and potentiate disease and may be a useful target for microbiota-directed therapies in IBD.