Highlights d Lactobacillus species sense intestinal iron levels and attenuate host iron absorption d Microbial metabolites DAP and reuterin are novel HIF-2a inhibitors d Gut microbial metabolites regulate intestinal iron storage via ferritin regulation d Gut microbiota can be therapeutically targeted for ironrelated disorders

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Although the microbiota in the proximal gastrointestinal (GI) tract have been implicated in health and disease, much about these microbes remains understudied compared to those in the distal GI tract. This study characterized the microbiota across multiple proximal GI sites over time in healthy individuals. As part of a study of the pharmacokinetics of oral mesalamine administration, healthy, fasted volunteers (n = 8; 10 observation periods total) were orally intubated with a four-lumen catheter with multiple aspiration ports. Samples were taken from stomach, duodenal, and multiple jejunal sites, sampling hourly (≤7 h) to measure mesalamine (administered att = 0), pH, and 16S rRNA gene-based composition. We observed a predominance ofFirmicutesacross proximal GI sites, with significant variation compared to stool. The microbiota was more similar within individuals over time than between subjects, with the fecal microbiota being unique from that of the small intestine. The stomach and duodenal microbiota displayed highest intraindividual variability compared to jejunal sites, which were more stable across time. We observed significant correlations in the duodenal microbial composition with changes in pH; linear mixed models identified positive correlations with multipleStreptococcusoperational taxonomic units (OTUs) and negative correlations with multiplePrevotellaandPasteurellaceaeOTUs. Few OTUs correlated with mesalamine concentration. The stomach and duodenal microbiota exhibited greater compositional dynamics than the jejunum. Short-term fluctuations in the duodenal microbiota were correlated with pH. Given the unique characteristics and dynamics of the proximal GI tract microbiota, it is important to consider these local environments in health and disease states.IMPORTANCEThe gut microbiota are linked to a variety of gastrointestinal diseases, including inflammatory bowel disease. Despite this importance, microbiota dynamics in the upper gastrointestinal tract are understudied. Our article seeks to understand what factors impact microbiota dynamics in the healthy human upper gut. We found that the upper gastrointestinal tract contains consistently prevalent bacterial OTUs that dominate the overall community. Microbiota variability is highest in the stomach and duodenum and correlates with pH.

The diets of industrialized countries reflect the increasing use of processed foods, often with the introduction of novel food additives. Xanthan gum is a complex polysaccharide with unique rheological properties that have established its use as a widespread stabilizer and thickening agent 1 . However, little is known about its direct interaction with the gut microbiota, which plays a central role in digestion of other, chemically-distinct dietary fiber polysaccharides. Here, we show that the ability to digest xanthan gum is surprisingly common in industrialized human gut microbiomes and appears to be contingent on the activity of a single bacterium that is a member of an uncultured bacterial genus in the family Ruminococcaceae. We used a combination of enrichment culture, multi-omics, and recombinant enzyme studies to identify and characterize a complete pathway in this uncultured bacterium for the degradation of xanthan gum. Our data reveal that this keystone degrader cleaves the xanthan gum backbone with a novel glycoside hydrolase family 5 (GH5) enzyme before processing the released oligosaccharides using additional enzymes. Surprisingly, some individuals harbor a Bacteroides species that is capable of consuming oligosaccharide products generated by the keystone Ruminococcaceae or a purified form of the GH5 enzyme. This Bacteroides symbiont is equipped with its own distinct enzymatic pathway to cross-feed on xanthan gum breakdown products, which still harbor the native linkage complexity in xanthan gum, but it cannot directly degrade the high molecular weight polymer. Thus, the introduction of a common food additive into the human diet in the past 50 years has promoted the establishment of a food chain involving at least two members of different phyla of gut bacteria.

Dietary fiber provides a variety of microbiota-mediated benefits ranging from anti-inflammatory metabolites to pathogen colonization resistance. A healthy gut microbiota protects against Clostridioides difficile colonization. Manipulation of these microbes through diet may increase colonization resistance to improve clinical outcomes. The primary objective of this study was to identify how the dietary fiber xanthan gum affects the microbiota and C. difficile colonization. We added 5% xanthan gum to the diet of C57BL/6 mice and examined its effect on the microbiota through 16S rRNA gene amplicon sequencing and short-chain fatty acid analysis. Following either cefoperazone or an antibiotic cocktail administration, we challenged mice with C. difficile and measured colonization by monitoring the CFU. Xanthan gum administration is associated with increases in fiber-degrading taxa and shortchain fatty acid concentrations. However, by maintaining both the diversity and absolute abundance of the microbiota during antibiotic treatment, the protective effects of xanthan gum administration on the microbiota were more prominent than the enrichment of these fiber-degrading taxa. As a result, mice that were on the xanthan gum diet experienced limited to no C. difficile colonization. Xanthan gum administration alters mouse susceptibility to C. difficile colonization by maintaining the microbiota during antibiotic treatment. While antibiotic-xanthan gum interactions are not well understood, xanthan gum has previously been used to bind drugs and alter their pharmacokinetics. Thus, xanthan gum may alter the activity of the oral antibiotics used to make the microbiota susceptible. Future research should further characterize how this and other common dietary fibers interact with drugs. IMPORTANCE A healthy gut bacterial community benefits the host by breaking down dietary nutrients and protecting against pathogens. Clostridioides difficile capitalizes on the absence of this community to cause diarrhea and inflammation. Thus, a major clinical goal is to find ways to increase resistance to C. difficile colonization by either supplementing with bacteria that promote resistance or a diet to enrich for those already present in the gut. In this study, we describe an interaction between xanthan gum, a human dietary additive, and the microbiota resulting in an altered gut environment that is protective against C. difficile colonization.