Examining Regional Differences in the Gut Microbiota and Their Effects on Clostridioides difficile Colonization Resistance

The mammalian gut is home to a vibrant community of microbes. The ecological interactions that shape this environment are distinct across gut locations. As host and microbial community co-evolved, they formed a complex yet stable relationship that prevents invading microorganisms, such as the spore-forming bacterium, Clostridioides difficile, from establishing within the gut. Great strides have been made over the past several years in characterizing C. difficile infection physiology, particularly in how gut microbes and their host work together to provide colonization resistance. I designed the work in this thesis to characterize how the mammalian small and large intestines shape C. difficile germination and outgrowth. Using observational and experimental approaches, I used human, murine and bioreactor models of the gut microbiota to study varying levels of ecological complexity. I show that the small intestinal microbiota is predominated by Firmicutes and that fluctuations in the microbiota are associated with changes in pH and bile acids. The bile acid population from the duodenum to the mid jejunum consists mainly of conjugated primary and secondary bile acids, with little microbial metabolism of these compounds occurring across the proximal small intestine. Since conjugated bile acids tend to promote C. difficile germination, this environment supports C. difficile’s transition from spore to vegetative cell at high efficiencies, suggesting that colonization resistance is tied to preventing the establishment of vegetative cells later in the gut. I also present work characterizing the effect of dietary xanthan gum on C. difficile colonization in a murine model of infection. Xanthan gum administration modified the microbiota and led to increased production of short chain fatty acids. However, it also interfered with the activity of the orally administered antibiotics used to render mice susceptible to C. difficile colonization. As a result, C. difficile colonization resistance was maintained in mice fed xanthan gum. Finally, I use a bioreactor model to characterize how founder effects shape microbial community establishment and influence C. difficile colonization resistance. Dilution increases the variability of the microbiota and abrogates its ability to resist invasion by a non-indigenous microbe. Additionally, we provided some reactor communities with additional concentrations of the dietary polysaccharide inulin. While some communities responded to inulin by producing additional butyrate, more dilute communities concurrently lost their ability to produce additional butyrate and resist C. difficile colonization. These data demonstrate that a particular level of microbiota cohesiveness is required to produce both functions and suggests that metabolic activity of butyrate-producing microbes is tied to colonization resistance. Together, my work demonstrates the importance of understanding the environmental interactions that shape microbial physiology, particularly that of C. difficile, in both the small and large intestines. In the small intestine, a variable microbiota with low biomass is shaped by host-driven processes, such as bile acid secretion. However, in the large intestine, a high biomass microbiota shapes the environment by metabolizing complex nutrients and preventing invasive taxa, such as C. difficile, from becoming established. Future work can leverage these findings to develop treatment methods that incorporate the dynamics of the intestinal environment to improve efficacy.