Digestive systems are dynamic, contractile ecosystems that continually shape the physical and chemical niches inhabited by gut microbes. Patterns of mixing and propulsion generate flow, shear, and nutrient gradients that select for microbial traits such as adhesion, biofilm formation, and metabolic timing. Microbial metabolites, in turn, influence smooth muscle excitability and enteric neural circuits, creating bidirectional feedback that structures microbial communities and affects host physiology. We argue that gastrointestinal motility, an ancient and nearly universal feature of metazoan digestive systems, functions as an evolutionary scaffold, linking microbial spatial organization with host neuromuscular diversification. Comparative evidence from cnidarians to mammals highlights how these interactions have shaped both microbial ecology and host adaptation, with relevance for understanding modern dysmotility-microbiome disorders.

Abstract Prebiotic supplementation a well-established impact on early-life gut microbiota, yet how substrate type and baseline community structure interact to shape microbial and metabolic responses remains incompletely understood. Using donor-derived infant faecal fermentations, a defined synthetic microbial community (SynCom), and multi-layered metabolomics, we demonstrate that prebiotic responses are strongly dependent on feeding-mode and mediated by species-specific metabolic interactions. While both human milk oligosaccharides (HMOs) and galacto-oligosaccharides (GOS) robustly promoted bifidobacterial expansion, modulation of Bacteroides/Phocaeicola species varied according to feeding background and substrate identity with 2′-fucosyllactose (2′FL) selectively alleviating growth inhibition in breast milk-derived communities. Mechanistic dissection using SynCom revealed extensive cross-feeding networks and substrate-dependent interaction rewiring, identifying Phocaeicola vulgatus as a context-dependent ecological hub that disproportionately shaped succinate and propionate production and altered Escherichia coli metabolic activity without affecting its abundance. Competitive outcomes were determined not only by environmental acidification, but also by the chemical identity of fermentation products, with lactate exerting species-specific inhibitory effects independent of bulk pH changes. Using an in-house ex vivo IntestiFlow gut organ platform, we further show that metabolites produced by the defined community induced a strong but reversible suppression of colonic peristaltic activity, an effect not recapitulated by SCFAs alone. Biogenic amine metabolites, including phenethylamine, partially reproduced key aspects of this response, suggesting a role for specific microbial fermentation products in modulating intestinal motility. Together, these findings demonstrate that early-life prebiotic responses extend beyond uniform bifidogenic expansion and instead emerge from interaction-driven metabolic specialization within feeding-mode-specific community contexts, with direct functional consequences for infant intestinal physiology.

Pseudomonas fluorescens is a Gram-negative bacterium with a remarkable metabolic and physiological versatility that enables it to adapt and colonize diverse ecological niches, including the human small intestine. While serotonin is primarily found in high concentrations in gut tissue, its levels in the lumen can be elevated in conditions such as celiac disease, where P. fluorescens is also found in increased abundance. The potential effects of serotonin on P. fluorescens in such contexts remain unclear. We demonstrate that P. fluorescens metabolizes serotonin primarily into 5-hydroxyindole-3-acetic acid (5-HIAA) and, to a lesser extent, into 5-hydroxytryptophol and N-acetylserotonin. Gene expression analysis revealed significant changes in oxidative stress-related pathways over time, and proteomic analysis confirmed the shifts seen particularly in amino acid catabolic pathways. Serotonin metabolism also enhanced bacterial resistance to oxidative stress, suggesting a protective role. The findings reveal a novel mechanism by which serotonin modulates the metabolism and stress responses of P. fluorescens. This study provides insight into how P. fluorescens adapts to serotonin-rich environments, such as in celiac disease, and may inform future research on microbial interactions with host-derived metabolites in disease contexts.

Mangiferin (MAN), a natural C-glycosylxanthone, is recognized for its health-promoting effects in traditional medicinal preparations. However, its poor bioavailability and limited intestinal permeability restrict its direct biological activity in vivo. Previous studies have suggested a potential bacterial breakdown of MAN into norathyriol (NOR), an aglycone with significantly higher bioavailability and absorption. Yet, the prevalence of MAN-metabolizing microbes, the presence of MAN or NOR within the gut microbial community, and their effects on the composition and metabolic activity of the gut microbiome remain unclear. In this study, fecal samples from healthy adult volunteers treated with MAN revealed its conversion to NOR, with interindividual variation attributed to the uncultured bacterial strain CAKRHR01 sp934339005. While MAN had minimal impact on microbial composition and metabolic activity, NOR treatment significantly increased pH, reduced overall bacterial cell counts, and selectively suppressed short-chain fatty acid-producing bacteria, including Faecalibacterium prausnitzii as well as urate consumers, such as Enterocloster bolteae. These findings underscore the potential of NOR to modulate gut microbial activity and highlight the importance of understanding microbiome-mediated metabolism when assessing the health implications of phytochemicals.

Synbiotics combine the concepts of probiotics and prebiotics to synergistically enhance the health-associated effects of both components. Previously, we have shown that the intestinal persistence of inulin-utilizing L. plantarum Lp900 is significantly increased in rats fed an inulin-supplemented, high-calcium diet. Here we employed a competitive population dynamics approach to demonstrate that inulin and GOS can selectively enrich L. plantarum strains that utilize these substrates for growth during in vitro cultivation, but that such enrichment did not occur during intestinal transit in rats fed a GOS or inulin-supplemented diet. The intestinal persistence of all L. plantarum strains increased irrespective of their prebiotic utilization phenotype, which was dependent on the calcium level of the diet. Analysis of fecal microbiota and intestinal persistence decline rates indicated that prebiotic utilization capacity did not selectively stimulate intestinal persistence in prebiotic supplemented diets. Moreover, microbiota and organic acid profile analyses indicate that the prebiotic utilizing probiotic strains are vastly outcompeted by the endogenous prebiotic-utilizing microbiota, and that the collective enhanced persistence of all L. plantarum strains is most likely explained by their well-established tolerance to organic acids.

The gut microbiota plays a pivotal role in health and disease. The use of probiotics as microbiota-targeted therapies is a promising strategy to improve host health. However, dynamic molecular mechanisms are often not elucidated, especially when targeting the small intestinal microbiota. Here, we show that supplementation of a probiotic formula (Ecologic®825) to the adult human small intestinal ileostoma microbiota counteracts the growth of Enterococcaceae and Enterobacteriaceae and reduces ethanol production, leading to major changes in nutrient utilization and resistance to perturbations. The observed alterations coincided with an initial increase in lactate production and decrease in pH by the probiotics, followed by a sharp increase in the levels of butyrate and propionate. Additionally, increased production of multiple N -acyl amino acids was detected in the stoma samples supplemented with the probiotic formula. Overall, this study shows how network theory can be used to improve the current or identify novel microbiota-targeted therapies. The outcome may help further understand the reported effects of these probiotic formula on the host.

The gut microbiota plays a pivotal role in health and disease. The use of probiotics as microbiota-targeted therapies is a promising strategy to improve host health. However, the molecular mechanisms involved in such therapies are often not well understood, particularly when targeting the small intestinal microbiota. In this study, we investigated the effects of a probiotic formula (Ecologic®825) on the adult human small intestinal ileostoma microbiota. The results showed that supplementation with the probiotic formula led to a reduction in the growth of pathobionts, such as Enterococcaceae and Enterobacteriaceae, and a decrease in ethanol production. These changes were associated with significant alterations in nutrient utilization and resistance to perturbations. These probiotic mediated alterations which coincided with an initial increase in lactate production and decrease in pH were followed by a sharp increase in the levels of butyrate and propionate. Moreover, the probiotic formula increased the production of multiple N-acyl amino acids in the stoma samples. The study demonstrates the utility of network theory in identifying novel microbiota-targeted therapies and improving existing ones. Overall, the findings provide insights into the dynamic molecular mechanisms underlying probiotic therapies, which can aid in the development of more effective treatments for a range of conditions.

ABSTRACT Intestinal microbiota and microbiota-derived metabolites play a key role in regulating the host physiology. Recently, we have identified a gut-bacterial metabolite, namely 5-hydroxyindole, as a potent stimulant of intestinal motility via its modulation of L-type voltage-gated calcium channels located on the intestinal smooth muscle cells. Dysregulation of L-type voltage-gated calcium channels is associated with various gastrointestinal motility disorders, including constipation, making L-type voltage-gated calcium channels an important target for drug development. Nonetheless, the majority of currently available drugs are associated with alteration of the gut microbiota. Using 16S rRNA sequencing this study shows that, when administered orally, 5-hydroxyindole has only marginal effects on the rat cecal microbiota. Molecular dynamics simulations propose potential-binding pockets of 5-hydroxyindole in the α1 subunit of the L-type voltage-gated calcium channels and when its stimulatory effect on the rat colonic contractility was compared to 16 different analogues, ex-vivo, 5-hydroxyindole stood as the most potent enhancer of the intestinal contractility. Overall, the present findings imply a potential role of microbiota-derived metabolites as candidate therapeutics for targeted treatment of slow intestinal motility-related disorders including constipation.

Abstract Small intestinal bacteria harboring tyrosine decarboxylase (TDC) activity was shown to reduce levels of the main treatment for Parkinson’s disease (PD), levodopa. However, it remains obscure whether the relative abundance of bacterial-TDC in the oral cavity and/or feces contributes to the decreased levodopa bioavailability and the frequency of dosage regimen of levodopa treatment. Shotgun metagenomic sequencing of saliva and fecal samples collected from 19 PD patients requiring various levodopa dosage regimens (consumed between 2 and 7 times per day; dosage range between 300-1400 mg) and 16 age-matched household spousal healthy control subjects was employed. The composition of TDC- harboring bacteria differed between oral and fecal samples, as well as between fecal samples from PD patients compared to their controls. TDC-bacterial activity showed significant negative correlation with plasma levodopa levels measured over five hours in all PD patients, while TDC-bacterial relative abundance correlated positively with the daily levodopa dosage in PD patients on high frequency levodopa dosage regimen. PD fecal samples had markedly lower relative abundance of short-chain fatty acids-producing bacteria. Functional analyses revealed differences in microbiota metabolism in PD involving lower vitamin B12 biosynthesis. Our data revealed a significant contribution of TDC to the decreased levodopa bioavailability and differences in fecal, but not the overall oral microbiota composition and metabolism between PD patients and their controls. These findings suggest that PD-associated alterations of the gut, but not oral microbiome, may render considerable functional gene profiling differences affecting levodopa bioavailability, host metabolism and motor-fluctuations.

The gut microbiota is in continuous interaction with the intestinal mucosa via metabolic, neuro-immunological, and neuroendocrine pathways. Disruption in levels of antimicrobial peptides produced by the enteroendocrine cells, such as catestatin, has been associated with changes in the gut microbiota and imbalance in intestinal homeostasis. However, whether the changes in the gut microbiota have a causational role in intestinal dyshomeostasis has remained elusive. To this end, we performed reciprocal fecal microbial transplantation in wild-type mice and mice with a knockout in the catestatin coding region of the chromogranin-A gene (CST-KO mice). Combined microbiota phylogenetic profiling, RNA sequencing, and transmission electron microscopy were employed. Fecal microbiota transplantation from mice deficient in catestatin (CST-KO) to microbiota-depleted wild-type mice induced transcriptional and physiological features characteristic of a distorted colon in the recipient animals, including impairment in tight junctions, as well as an increased collagen area fraction indicating colonic fibrosis. In contrast, fecal microbiota transplantation from wild-type mice to microbiota-depleted CST-KO mice reduced collagen fibrotic area, restored disrupted tight junction morphology, and altered fatty acid metabolism in recipient CST-KO mice. This study provides a comprehensive overview of the murine metabolic- and immune-related cellular pathways and processes that are co-mediated by the fecal microbiota transplantation and supports a prominent role for the gut microbiota in the colonic distortion associated with the lack of catestatin in mice. Overall, the data show that the gut microbiota may play a causal role in the development of features of intestinal inflammation and metabolic disorders, known to be associated with altered levels of catestatin and may, thus, provide a tractable target in the treatment and prevention of these disorders.

The gut microbiota is in continuous interaction with the innermost layer of the gut, namely the epithelium. One of the various functions of the gut epithelium, is to keep the microbes at bay to avoid overstimulation of the underlying mucosa immune cells. To do so, the gut epithelia secrete a variety of antimicrobial peptides, such as chromogranin A (CgA) peptide catestatin (CST: hCgA 352-372 ). As a defense mechanism, gut microbes have evolved antimicrobial resistance mechanisms to counteract the killing effect of the secreted peptides. To this end, we treated wild-type mice and CST knockout (CST-KO) mice (where only the 63 nucleotides encoding CST have been deleted) with CST for 15 consecutive days. CST treatment was associated with a shift in the diversity and composition of the microbiota in the CST-KO mice. This effect was less prominent in WT mice. Levels of the microbiota-produced short-chain fatty acids, in particular, butyrate and acetate were significantly increased in CST-treated CST-KO mice but not the WT group. Both CST-treated CST-KO and WT mice showed a significant increase in microbiota-harboring phosphoethanolamine transferase-encoding genes, which facilitate their antimicrobial resistance. Finally, we show that CST was degraded by Escherichia coli via an omptin-protease and that the abundance of this gene was significantly higher in metagenomic datasets collected from patients with Crohn’s disease but not with ulcerative colitis. Overall, this study illustrates how the endogenous antimicrobial peptide, CST, shapes the microbiota composition in the gut and primes further research to uncover the role of bacterial resistance to CST in disease states such as inflammatory bowel disease.

Abstract Background: Intestinal microbiota and microbiota-derived metabolites play a key role in regulating the host physiology, including gut motility. We have previously shown that a wide variety of gut bacteria can produce 5-hydroxyindole from its precursor, dietary supplement and antidepressant, 5-hydroxytryptophan. When 5-hydroxyindole was administered orally in rats, it significantly accelerated the total gut transit time. Results: Here we show, using 16S rRNA sequencing, that 5-hydroxyindole has no effect on the bacterial richness. No separation between the treated and untreated groups was detected based on their microbial composition as shown by principal component analysis. A significant increase in the abundance of only two genera, Shuttleworthia and Prevotellaceae_UCG-001, was detected in the 5-hydroxyindole treated group compared to the vehicle group. No significant association was found between the reduced total gut transit time as a result of 5-hydroxyindole administration and the cecal bacterial composition. Conclusions: Together, our study infers a marginal effect of 5-hydroxyindole or its subsequent accelerated gut motility on the cecal microbiota composition of rats. The results urge further investigation of the impact of 5-hydroxyindole oral administration in humans, which will support its potential application as a treatment for slow intestinal motility disorders.

Fructo-oligosaccharides (FOS), inulin, and galacto-oligosaccharides (GOS) are widely recognized prebiotics that profoundly affect the intestinal microbiota, including stimulation of bifidobacteria and lactobacilli, and are reported to elicit several health benefits. The combination of dietary FOS and inulin with calcium phosphate was reported to stimulate commensal Lactobacillus populations and protect the host against pathogenic Enterobacteriaceae, but little is known about the effects of GOS in diets with a different level of calcium phosphate. We investigated the microbiome changes elicited by dietary supplementation with GOS or inulin using diets with high (100 mmol/kg) and low (30 mmol/kg) calcium phosphate levels in adult Wistar rats. Rats were acclimatized to the respective experimental diets for 14 days, after which fecal material was collected, DNA was extracted from fecal material, and the V3‑V4 region of the bacterial 16S rRNA gene was amplified with PCR, followed by microbial composition analysis. In tandem, the organic acid profiles of the fecal material were analyzed. Feeding rats non-supplemented (no prebiotic-added) diets revealed that diets rich in calcium phosphate favored members of the Firmicutes and increased fecal lactic, succinic, acetic, propionic, and butyric acid levels. In contrast, relatively low dietary calcium phosphate levels promoted the abundance of mucin degrading genera like Akkermansia and Bacteroides, and resulted in increased fecal propionic acid levels and modest increases in lactic and butyric acid levels. Irrespective of the calcium phosphate levels, supplementation with GOS or inulin strongly stimulated Bifidobacterium, while only high calcium phosphate diets increased the endogenous Faecalibaculum populations. Despite the prebiotic’s substantial difference in chemical structure, sugar composition, oligomer size, and the microbial degradation pathway involved in their utilization, inulin and GOS modulated the gut microbiota very similarly, in a manner that strongly depended on the dietary calcium phosphate level. Therefore, our study implies that the collection of detailed diet information including micronutrient balance is necessary to correctly assess diet-driven microbiota analysis.

Microbial conversion of dietary or drug substrates into small bioactive molecules represents a regulatory mechanism by which the gut microbiota alters intestinal physiology. Here, we show that a wide variety of gut bacteria can metabolize the dietary supplement and antidepressant 5-hydroxytryptophan (5-HTP) to 5-hydroxyindole (5-HI) via the tryptophanase (TnaA) enzyme. Oral administration of 5-HTP results in detection of 5-HI in fecal samples of healthy volunteers with interindividual variation. The production of 5-HI is inhibited upon pH reduction in in vitro studies. When administered orally in rats, 5-HI significantly accelerates the total gut transit time (TGTT). Deciphering the underlying mechanisms of action reveals that 5-HI accelerates gut contractility via activation of L-type calcium channels located on the colonic smooth muscle cells. Moreover, 5-HI stimulation of a cell line model of intestinal enterochromaffin cells results in significant increase in serotonin production. Together, our findings support a role for bacterial metabolism in altering gut motility and lay the foundation for microbiota-targeted interventions.

Synbiotics combine probiotics with prebiotics to synergistically elicit a health benefit in the consumer. Previous studies have shown that prebiotics can selectively stimulate the growth in the intestine of specific bacterial strains.

Methylphenidate is one of the most widely used oral treatments for attention-deficit/hyperactivity disorder (ADHD). The drug is mainly absorbed in the small intestine and has low bioavailability. Accordingly, a high interindividual variability in terms of response to the treatment is known among ADHD patients treated with methylphenidate. Nonetheless, very little is known about the factors that influence the drug’s absorption and bioavailability. Gut microbiota has been shown to reduce the bioavailability of a wide variety of orally administered drugs. Here, we tested the ability of small intestinal bacteria to metabolize methylphenidate. In silico analysis identified several small intestinal bacteria to harbor homologues of the human carboxylesterase 1 enzyme responsible for the hydrolysis of methylphenidate in the liver into the inactive form, ritalinic acid. Despite our initial results hinting towards possible bacterial hydrolysis of the drug, up to 60% of methylphenidate is spontaneously hydrolyzed in the absence of bacteria and this hydrolysis is pH-dependent. Overall, our results indicate that the stability of methylphenidate is compromised under certain pH conditions in the presence or absence of gut microbiota.

Several Lactobacillus plantarum strains are marketed as probiotics for their potential health benefits. Prebiotics, e.g., galacto-oligosaccharides (GOS), have the potential to selectively stimulate the growth of L. plantarum probiotic strains based on their phenotypic diversity in carbohydrate utilization, and thereby enhance their health promoting effects in the host in a strain-specific manner. Previously, we have shown that GOS variably promotes the strain-specific growth of L. plantarum . In this study we investigated this variation by molecular analysis of GOS utilization by L. plantarum . HPAEC-PAD analysis revealed two distinct GOS utilization phenotypes in L. plantarum. Linking these phenotypes to the strain-specific genotypes led to the identification of a lac operon encoding a β-galactosidase ( lacA ), a permease ( lacS ), and a divergently oriented regulator ( lacR ), that are predicted to be involved in the utilization of higher degree of polymerization (DP) constituents present in GOS (specifically DP of 3–4). Mutation of lacA and lacS in L. plantarum NC8 resulted in reduced growth on GOS, and HPAEC analysis confirmed the role of these genes in the import and utilization of higher-DP GOS constituents. Overall, the results enable the design of highly-selective synbiotic combinations of L. plantarum strain-specific probiotics and specific GOS-prebiotic fractions.

SummaryMany plant genomes display high levels of repetitive sequences. The assembly of these complex genomes using short high‐throughput sequence reads is still a challenging task. Underestimation or disregard of repeat complexity in these datasets can easily misguide downstream analysis. Detection of repetitive regions by k‐mer counting methods has proved to be reliable. Easy‐to‐use applications utilizing k‐mer counting are in high demand, especially in the domain of plants. We present Kmasker plants, a tool that uses k‐mer count information as an assistant throughout the analytical workflow of genome data that is provided as a command‐line and web‐based solution. Beside its core competence to screen and mask repetitive sequences, we have integrated features that enable comparative studies between different cultivars or closely related species and methods that estimate target specificity of guide RNAs for application of site‐directed mutagenesis using Cas9 endonuclease. In addition, we have set up a web service for Kmasker plants that maintains pre‐computed indices for 10 of the economically most important cultivated plants. Source code for Kmasker plants has been made publically available at https://github.com/tschmutzer/kmasker. The web service is accessible at https://kmasker.ipk-gatersleben.de.