McCullough, H. C. | Search bioRxiv

bioRxiv2024

Diversity in Chemical Subunits and Linkages: A Key Molecular Determinant of Microbial Richness, Community Structure, Interactions, and Substrate Utilization

Dietary fibers play a significant role in shaping the composition and function of microbial communities in the human colon. Our understanding of the specific chemical traits of dietary fibers that influence microbial diversity, interactions, and function remains limited. Towards filling this knowledge gap, we developed a novel measure, termed Chemical Subunits and Linkages (CheSL) Shannon diversity, to characterize the effects of carbohydrate complexity on human fecal bacteria cultured in vitro under controlled, continuous flow conditions using media that systematically varied in carbohydrate composition. Our analysis revealed that CheSL Shannon diversity demonstrated a strong Pearson correlation with microbial richness across multiple fecal samples and study designs. Additionally, we observed that microbial communities in media with higher CheSL Shannon diversity scores exhibited greater peptide utilization, and more connected, reproducible structures in computationally inferred microbial interaction networks. Together, these findings demonstrate that CheSL Shannon diversity can be a useful tool to quantify the effects of carbohydrate complexity on microbial diversity, metabolic potential, and interactions. Furthermore, our work highlights how robust and stable community data can be generated by engineering media composition and structure. These studies provide a valuable framework for future research on microbial community interactions and their potential impacts on host health. IMPORTANCEFor the human adult gut microbiota, higher microbial diversity strongly correlates with positive health outcomes. This correlation is likely due to increased community resilience that results from functional redundancy that can occur within diverse communities. While previous studies have shown that dietary fibers influence microbiota composition and function, we lack a complete mechanistic understanding of how differences in the composition of fibers are likely to functionally impact microbiota diversity. To address this need, we developed Chemical Subunits and Linkages (CheSL) Shannon diversity, a novel measure that describes carbohydrate complexity. Using this measure, we were able to correlate changes in carbohydrate complexity with alterations in microbial diversity and interspecies interactions. Overall, these analyses provide new perspectives on dietary optimization strategies to improve human health.

microbiology↗

bioRxiv2024

Clostridioides difficile colonization is not mediated by bile salts and requires Stickland fermentation of proline in an in vitro model of infection

Treatment with antibiotics is a major risk factor for Clostridioides difficile infection, likely due to depletion of the gastrointestinal microbiota. Two microbiota-mediated mechanisms thought to limit C. difficile colonization include conversion of conjugated primary bile salts into secondary bile salts toxic to C. difficile growth, and competition between the microbiota and C. difficile for limiting nutrients. Using a continuous flow model that simulates the nutrient conditions of the distal colon, we investigated how treatment with six clinically-used antibiotics influenced susceptibility to C. difficile infection in 12 different microbial communities cultivated from healthy individuals. Antibiotic treatment reduced microbial richness; disruption varied by antibiotic class and microbiota composition, but did not correlate with C. difficile susceptibility. Antibiotic treatment also disrupted microbial bile salt metabolism, increasing levels of the primary bile salt, cholate. However, changes in bile salt did not correlate with increased C. difficile susceptibility. Further, bile salts were not required to inhibit C. difficile colonization. We tested whether amino acid fermentation contributed to persistence of C. difficile in antibiotic- treated communities. C. difficile mutants unable to use proline as an electron acceptor in Stickland fermentation due to disruption of proline reductase (prdB-) had significantly lower levels of colonization than wild-type strains in four of six antibiotic-treated communities tested. Inability to ferment glycine or leucine as electron acceptors, however, was not sufficient to limit colonization in any communities. This data provides further support for the importance of bile salt-independent mechanisms in regulating colonization of C. difficile. IMPORTANCEC. difficile is one of the leading causes of hospital-acquired infections and antibiotic-associated diarrhea. Several potential mechanisms through which the microbiota can limit C. difficile infection have been identified and are potential targets for new therapeutics. However, it is unclear which mechanisms of C. difficile inhibition represent the best targets for development of new therapeutics. These studies demonstrate that in a complex in vitro model of C. difficile infection, colonization resistance is independent of microbial bile salt metabolism. Instead, the ability of C. difficile to colonize is dependent upon its ability to metabolize proline, although proline-dependent colonization is context-dependent and is not observed in all disrupted communities. Altogether, these studies support the need for further work to understand how bile- independent mechanisms regulate C. difficile colonization.

microbiology↗