The human gut microbiome expands the host’s metabolic functionality approximately 10-fold and subsequently contributes to host health by providing otherwise inaccessible nutrients. Many metabolic products derived from the activity of this diverse microbiome, along with host metabolism, are bioactive and contribute to functions beyond basic cellular maintenance. These bioactive compounds can regulate enteric pathogens through two-component system signaling and contribute to host neurological disorders like depression and schizophrenia through the gut-brain axis. The control of bioactive metabolites, whether through dietary alterations, microbiome manipulation, or some combination thereof, presents an intriguing and promising approach to not only controlling enteric infections but also as possible biomarkers or treatments for a range of human neurological disorders. Though promising, exogenous fine control of metabolism is difficult and the mechanistic interactions of metabolites with their microbial and human targets remains understudied due to the inherent complexity of metabolism and the notable inter- and intra-individual variation of the gut microbiome.

However challenging, disentangling the potential widespread effects of bioactive metabolites on host neurological health is imperative for the development of future therapeutics and biomarkers. In Chapter 2 we attempted to sort out some of this relationship between shared microbe-host metabolites and human neurological disorders through a deep dive into existing literature. Many metabolic products from tryptophan, an essential amino acids for humans and a central substrate for protein catabolism and energy production, have gained notoriety in this field for their potential role as markers or agents of common neurological disorders. Anthranilate, one intermediate product of the tryptophan-reliant kynurenine pathway, is particularly interesting given current literature supports that this compound may be neuroprotective or neurotoxic depending on context. Further investigation of this compound revealed that common neurological disorders like depression, Schizophrenia, and Alzheimer’s may be linked together by the activity of the anthranilic metabolic pathway. This search additionally exposed the significant role gut microorganisms play in altering the precarious balance of circulating tryptophan-derived metabolites, the variation of which can have serious implications for host health.

Given the clear importance of microbial metabolites to host neurological health, we hypothesized host-derived metabolites could likewise control the activity of microorganisms in the gut. Indeed, a thorough review on the effect of host-derived metabolic products on enteric microorganisms supported the concept of metabolite-driven bi-directional communication and interkingdom signaling. Two-component signaling systems, ubiquitous in prokaryotes, are one mechanism by which bacteria translate the host-produced extracellular small molecules into intracellular responses, such as the induction of virulence factors or repression of extraneous metabolic functions. A follow-up search of the Joint Genome Institute’s Integrated Microbial Genomes and Microbiomes (IMG/M) platform revealed genes encoding two-component systems are omnipresent in the metagenomes of human gut microbes, lending support to the importance of better understanding how host-derived substrates influence activity in the gut microbiome for better or worse.

The strong body of literature supporting the integral role of metabolic substrates as contributors to host health and regulators of enteric microorganisms led us to propose the addition of exogenous dietary substrates in vitro would alter the metabolic crosstalk between enteric pathogen Salmonella enterica sv. Typhimurium and human colonic epithelial cells. In vitro work using colonic epithelial cells and S. Typhimurium treated with either human milk oligosaccharides (HMO) or mannanoligosaccharides (BioMos®), evaluated using dual RNAseq and untargeted metabolomics, exposed clear and substrate-dependent metabolic shifts in both host and pathogen. Intriguingly, both substrates reduced overall host-pathogen association, but BioMos® treatment shifted S. Typhimurium towards more energetically favorable aerobic respiration while HMO did not induce this same shift, although both treatments did increase the expression of common virulence factors. Also notable was the induction of amino acid metabolism in BioMos® treated S. Typhimurium, a likely consequence of the inclusion of compounds beyond mannanoligosaccharides in this commercial prebiotic product. Although reduction in pathogenic association was decreased by both treatments, the distinct metabolic and gene expression profiles of host and pathogen in each condition suggests prebiotic supplements alter gut activity irrespective of the presence of commensal bacteria, their intended targets, and may actually induce virulent activity in dysbiotic gut environments, contrary to the goal of their application.

This work demonstrates the hypotheses that 1) tryptophan-derived metabolites from shared host-microbe metabolism can indicate and/or alter host neurology through circulation of small metabolites; 2) host metabolic products can likewise alter microbial function in the gastrointestinal tract, including but not limited to, regulating virulence factors via interactions with prokaryotic two-component systems; and 3) exogenous addition of dietary prebiotics shifts the shared metabolome of host epithelial cells and pathogenic S. Typhimurium, alongside simultaneous regulation of redox related metabolism in both cell types in a prebiotic-dependent manner. Taken together this work validates not only the importance of small metabolites in host health but also the potential for enteric pathogens to manipulate the tightly regulated metabolic relationship between host and gut microbes. Findings here additionally suggest that diet-driven manipulation of gut microbiota (for example the consumption of prebiotics) holds great promise as a next generation therapeutic approach, but widespread and off-target impacts remain a concern for widespread application at this stage in prebiotic research.

The growing demand for high quality food resulting from the increase in world population has incentivized the agriculture industry to increase their food production. However, large amounts of biomass and byproducts from food production are sent to landfills. Many of these wastes, despite no longer being suitable for human consumption, are often still rich in nutrients and represent valuable alternatives to complement animal feed. Livestock, especially ruminants, are an integral part in agriculture as they can convert plant biomass that cannot be digested by humans into high quality animal products such meat, milk and wool. The ability of ruminant animals to degrade recalcitrant plant fibers is due to the symbiotic relationship with the microorganisms that reside in the rumen and that can digest and convert plant biomass into nutrients and metabolic intermediates that can be further utilized by the host animals. Some of these microorganisms produce greenhouse gases, such as methane (CH4), which is released into the atmosphere where it has detrimental effects on the environment. Many byproducts from agriculture may contain bioactive compounds such as phenolics, organosulfur, terpenoids, and fatty acids that can alter rumen fermentation and even inhibit methanogenesis. California is the largest producer of agricultural products in the United States (U.S.), with a large arsenal of byproducts that could hold the key to improve ruminant nutrition and also have the potential to reduce enteric methane production. As of today, there is still only a limited number of studies that have investigated the potential of California byproducts to reduce enteric fermentation. In the work presented here, a panel of byproducts from California agriculture was subjected to chemical profiling and subsequent in vitro rumen fermentation. In addition to providing first insights into the potential effect of the tested local byproducts this work also provides a framework and the standard operating procedures that should be employed for future analyses to identify potential methane inhibitor and to enable comparisons of results that will be generated over time and from different researchers and institutes. A standardization of the analytical methods to evaluate rumen response will be key to not only identify novel strategies but also provide the framework to study the molecular drivers of methane inhibition and to optimize future methane mitigation strategies.