Human Gut Research Articles

IL-21 conditions antigen-presenting human γδ T-cells to promote IL-10 expression in naïve and memory CD4+ T-cells

Abstract Direct interaction between T-cells exerts a major influence on tissue immunity and inflammation across multiple body sites including the human gut, which is highly enriched in ‘unconventional’ lymphocytes such as γδ T-cells. We previously reported that microbial activation of human Vγ9/Vδ2+ γδ T-cells in the presence of the mucosal damage-associated cytokine IL-15 confers the ability to promote epithelial barrier defence, specifically via induction of IL-22 expression in conventional CD4+ T-cells. In the current report, we assessed whether other cytokines enriched in the gut milieu also functionally influence microbe-responsive Vγ9/Vδ2 T-cells. When cultured in the presence of IL-21, Vγ9/Vδ2 T-cells acquired the ability to induce expression of the immunoregulatory cytokine IL-10 in both naïve and memory CD4+ T-cells, at levels surpassing those induced by monocytes or monocyte-derived DCs. These findings identify an unexpected influence of IL-21 on Vγ9/Vδ2 T-cell modulation of CD4+ T-cell responses. Further analyses suggested a possible role for CD30L and/or CD40L reverse signalling in mediating IL-10 induction by IL-21 conditioned Vγ9/Vδ2 T-cells. Our findings indicate that the local microenvironment exerts a profound influence on Vγ9/Vδ2 T-cell responses to microbial challenge, leading to induction of distinct functional profiles among CD4+ T-cells that may influence inflammatory events at mucosal surfaces. Targeting these novel pathways may offer therapeutic benefit in disorders such as inflammatory bowel disease.

Rapid biotransformation of STW 5 constituents by human gut microbiome from IBS- and non-IBS donors.

STW 5 is an herbal medicinal product with proven clinical efficacy in the treatment of functional gastrointestinal disorders, like functional dyspepsia and irritable bowel syndrome (IBS). The effects of STW 5 on fecal microbial communities and metabolite production effects have been studied in an experimental model with fecal samples from individuals with and without IBS. While only minor taxonomic disparities were noted between IBS- and non-IBS samples and upon treatment with STW 5, rapid metabolic turnover of STW 5 components into specific degradation products with reported anti-inflammatory, cytoprotective, or spasmolytic activities was observed, which may be relevant for the pharmacological activity of STW 5.

Genomic Insights Into Enterococcus mundtii 203: A Promising Probiotic Candidate Isolated From Camel Feces.

Enterococcus, a common commensal organism in the human gut, exhibits a dual nature with certain strains offering probiotic benefits, while others are associated with nosocomial infections. In this study, we conducted a comprehensive examination of the genome of Enterococcus mundtii strain 203 to assess its probiotic potential and safety profile. The complete genome sequencing, assembly, and annotation were performed, followed by bioinformatics analysis. Our investigation reveals a detailed characterization of the Enterococcus mundtii 203 genome, originally isolated from camel feces, with a size of 3,053,234 bases and a GC content of 38.4%. Importantly, our analysis suggests that this strain poses no risk as a human pathogen due to the absence of antibiotic resistance determinants and virulence factors. The genome harbors a multitude of genes responsible for lactic acid production, bioactive peptide synthesis, adhesion molecule expression, resistance to harsh gut conditions, and enhancement of host metabolism. These findings underline the potential probiotic functionality of Enterococcus mundtii 203, positioning it as a promising candidate. Notably, our study did not identify any sequences related to insertion elements or CRISPR-Cas fragments.

Supplementation with soluble or insoluble rice-bran fibers increases short-chain fatty acid producing bacteria in the gut microbiota in vitro

IntroductionStudies have shown that a diet high in fiber and prebiotics has a positive impact on human health due largely to the fermentation of these compounds by the gut microbiota. One underutilized source of fiber may be rice bran, a waste product of rice processing that is used most frequently as an additive to livestock feed but may be a good source of fibers and other phenolic compounds as a human diet supplement. Previous studies focused on specific compounds extracted from rice bran showed that soluble fibers extracted from rice bran can improve glucose response and reduce weight gain in mouse models. However, less is known about changes in the human gut microbiota in response to regular rice bran consumption.MethodsIn this study, we used a Simulator of the Human Intestinal Microbial Ecology (SHIME®) to cultivate the human gut microbiota of 3 different donors in conditions containing either soluble or insoluble fiber fractions from rice bran. Using 16S rRNA amplicon sequencing and targeted metabolomics via Gas Chromatography-Mass Spectrometry, we explored how gut microbial communities developed provided different supplemental fiber sources.ResultsWe found that insoluble and soluble fiber fractions increased short-chain fatty acid production, indicating that both fractions were fermented. However, there were differences in response between donors, for example the gut microbiota from donor 1 increased acetic acid production with both fiber types compared with control; whereas for donors 2 and 3, butanoic acid production increased with ISF and SF supplementation. Both soluble and insoluble rice bran fractions increased the abundance of Bifidobacterium and Lachnospiraceae taxa.DiscussionOverall, analysis of the effect of soluble and insoluble rice bran fractions on the human in vitro gut microbiota and the metabolites produced revealed individually variant responses to these prebiotics.

Conjugated linoleic acid metabolite impact in colorectal cancer: a potential microbiome-based precision nutrition approach.

Colorectal cancer (CRC) is the second most deadly and the third most diagnosed cancer in both sexes worldwide. CRC pathogenesis is associated with risk factors such as genetics, alcohol, smoking, sedentariness, obesity, unbalanced diets, and gut microbiota dysbiosis. The gut microbiota is the microbial community living in symbiosis in the intestine, in a dynamic balance vital for health. Increasing evidence underscores the influence of specific gut microbiota bacterial species on CRC incidence and pathogenesis. In this regard, conjugated linoleic acid (CLA) metabolites produced by certain gut microbiota have demonstrated an anticarcinogenic effect in CRC, influencing pathways for inflammation, proliferation, and apoptosis. CLA production occurs naturally in the rumen, and human bioavailability is through the consumption of food derived from ruminants. In recent years, biotechnological attempts to increase CLA bioavailability in humans have been unfruitful. Therefore, the conversion of essential dietary linoleic acid to CLA metabolite by specific intestinal bacteria has become a promising process. This article reviews the evidence regarding CLA and CLA-producing bacteria as therapeutic agents against CRC and investigates the best strategy for increasing the yield and bioavailability of CLA. Given the potential and limitations of the present strategies, a new microbiome-based precision nutrition approach based on endogenous CLA production by human gut bacteria is proposed. A literature search in the PubMed and PubMed Central databases identified 794 papers on human gut bacteria associated with CLA production. Of these, 51 studies exploring association consistency were selected. After excluding 19 papers, due to health concerns or discrepancies between studies, 32 papers were selected for analysis, encompassing data for 38 CLA-producing bacteria, such as Bifidobacterium and Lactobacillus species. The information was analyzed by a bioinformatics food recommendation system patented by our research group, Phymofood (EP22382095). This paper presents a new microbiome-based precision nutrition approach targeting CLA-producing gut bacterial species to maximize the anticarcinogenic effect of CLA in CRC.

Comparison of fidaxomicin, thuricin CD, vancomycin and nisin highlights the narrow spectrum nature of thuricin CD

ABSTRACT Vancomycin and metronidazole are commonly used treatments for Clostridioides difficile infection (CDI). However, these antibiotics have been associated with high levels of relapse in patients. Fidaxomicin is a new treatment for CDI that is described as a narrow spectrum antibiotic that is minimally active on the commensal bacteria of the gut microbiome. The aim of this study was to compare the effect of fidaxomicin on the human gut microbiome with a number of narrow (thuricin CD) and broad spectrum (vancomycin and nisin) antimicrobials. The spectrum of activity of each antimicrobial was tested against 47 bacterial strains by well-diffusion assay. Minimum inhibitory concentrations (MICs) were calculated against a select number of these strains. Further, a pooled fecal slurry of 6 donors was prepared and incubated for 24 h with 100 µM of each antimicrobial in a mini-fermentation system together with a no-treatment control. Fidaxomicin, vancomycin, and nisin were active against most gram positive bacteria tested in vitro, although fidaxomicin and vancomycin produced larger zones of inhibition compared to nisin. In contrast, the antimicrobial activity of thuricin CD was specific to C. difficile and some Bacillus spp. The MICs showed similar results. Thuricin CD exhibited low MICs (<3.1 µg/mL) for C. difficile and Bacillus firmus, whereas fidaxomicin, vancomycin, and nisin demonstrated lower MICs for all other strains tested when compared to thuricin CD. The narrow spectrum of thuricin CD was also observed in the gut model system. We conclude that the spectrum of activity of fidaxomicin is comparable to that of the broad-spectrum antibiotic vancomycin in vitro and the broad spectrum bacteriocin nisin in a complex community.

Selective impact of three homogenous polysaccharides with different structural characteristics from Grifola frondosa on human gut microbial composition and the structure-activity relationship

Natural polysaccharides interact with gut microbes to enhance human well-being. Grifola frondosa is a polysaccharides-rich edible and medicinal mushroom. The prebiotic potential of G. frondosa polysaccharides has been explored in recent years, however, the relationship between their various structural features and prebiotic activities is poorly understood. In this study, three homogenous polysaccharides GFP10, GFP21 and GFP22 having different molecular weights (Mw), monosaccharide compositions and glycosidic linkages were purified from G. frondosa, and their effects on intestinal microbial composition were compared. GFP10 was a fucomannogalactan with an Mw of 23.0 kDa, and it selectively inhibited Enterobacter, while GFP21 was a fucomannogalactoglucan with an Mw of 18.6 kDa, and it stimulated Catenibacterium. GFP22 was a 4.9 kDa mannoglucan that selectively inhibited Klebsiella and boosted Bifidobacterium, Catenibacterium and Phascolarctobacterium, and prominently promoted the production of short-chain fatty acids (SCFAs). The selective modulation of gut microbiota by polysaccharides was structure-dependent. A relatively lower Mw and a high proportion of glycosidic linkages like T-Glcp, 1,3-Glcp, 1,3,6-Glcp and 1,4-Glcp might be more easily utilized to produce SCFAs and beneficial for the proliferation of Catenibacterium and Phascolarctobacterium. This research provided a valuable resource for further exploring the structure-activity relationship and prebiotic activity of G. frondosa polysaccharides.

A low-cost, easy-to-use prototype bioreactor model for the investigation of human gut microbiota: validation using a prebiotic treatment

In vitro gut models allow for the study of the impact of molecules on human gut microbiota composition and function without the implication of the host. However, current models, such as the Simulator of Human Intestinal Microbial Ecosystem (SHIME®), are expensive, time-consuming, and require specialized personnel. Homemade in vitro models that lessen these issues have limited evidence of their humanlike functionality. In this study, we present the development of a low-cost and easy-to-use bioreactor with the proven functionality of human microbiota. In our model, we evaluated the capability of replicating human gut microbiota growth and the response of the human bacterial community to a prebiotic, resistant starch, particularly resistant starch type 2 (RS2). Our bioreactor produced an environment that was stable for pH, temperature, and anaerobic conditions. The bioreactor consistently cultivated bacterial communities over a 48 h time period, replicating the composition of the gut microbiota and the associated metabolite production response to RS2, in line with prior human studies. In response to the RS2 prebiotic, we observed an increase in Bifidobacterium adolescentis and Bifidobacterium faecale and an increase in the production of the short-chain fatty acids such as acetate, propionate, and isobutyrate. Taken together, these data demonstrate that our low-cost and user-friendly prototype bioreactor model provides a favorable environment for the growth of human gut microbiota and can mimic its response to a prebiotic.

A perfusion host-microbe bioreactor (HMB) system that captures dynamic interactions of secreted metabolites between epithelial cells cocultured with a human gut anaerobe.

The human microbiota impacts a variety of diseases and responses to therapeutics. Due to a lack of robust in vitro models, detailed mechanistic explanations of host-microbiota interactions cannot often be recapitulated. We describe the design and development of a novel, versatile and modular in vitro system that enables indirect coculture of human epithelial cells with anaerobic bacteria for the characterization of host-microbe secreted metabolite interactions. This system was designed to compartmentalize anaerobes and human cells in separate chambers conducive to each organism's requisite cell growth conditions. Using perfusion, fluidic mixing, and automated sample collection, the cells continuously received fresh media, while in contact with their corresponding compartments conditioned supernatant. Supernatants from each chamber were collected in a cell-free time-resolved fashion. The system sustained low oxygen conditions in the anaerobic chamber, while also supporting the growth of a representative anaerobe (Bacteroides thetaiotaomicron) and a human colonic epithelial cell line (Caco-2) in the aerobic chamber. Caco-2 global gene expression changes in response to coculture with B. thetaiotaomicron was characterized using RNA sequencing. Extensive, targeted metabolomics analysis of over 150 central carbon metabolites was performed on the serially collected supernatants. We observed broad metabolite changes in host-microbe coculture, compared to respective mono-culture controls. These effects were dependent both on sampling time and the compartment probed (apical vs. basolateral). Coculturing resulted in the depletion of several important metabolites, including guanine, uridine 5'-monophosphate, asparagine, and thiamine. Additionally, while Caco-2 cells cultured alone predominantly affected the basolateral metabolite milieu, increased abundance of 2,3-dihydroxyisovalerate and thymine on the basolateral side, occurred when the cells were cocultured with B. thetaiotaomicron. Thus, our system can capture the dynamic, competitive and cooperative processes between host cells and gut microbes.

The gut microbiome and HLA-B27-associated anterior uveitis: a case-control study

BackgroundThe human gut microbiome (GM) is involved in inflammation and immune response regulation. Dysbiosis, an imbalance in this ecosystem, facilitates pathogenic invasion, disrupts immune equilibrium, and potentially triggers diseases including various human leucocyte antigen (HLA)-B27-associated autoinflammatory and autoimmune diseases such as inflammatory bowel disease (IBD) and spondyloarthropathy (SpA). This study assesses compositional and functional alterations of the GM in patients with HLA-B27-associated non-infectious anterior uveitis (AU) compared to healthy controls.MethodsThe gut metagenomes of 20 patients with HLA-B27-associated non-infectious AU, 21 age- and sex-matched HLA-B27-negative controls, and 6 HLA-B27-positive healthy controls without a history of AU were sequenced using the Illumina NovaSeq 6000 platform for whole metagenome shotgun sequencing. To identify taxonomic and functional features with significantly different relative abundances between groups and to identify associations with clinical metadata, the multivariate association by linear models (MaAsLin) R package was applied.ResultsSignificantly higher levels of the Eubacterium ramulus species were found in HLA-B27-negative controls (p = 0.0085, Mann-Whitney U-test). No significant differences in microbial composition were observed at all other taxonomic levels. Functionally, the lipid IVA biosynthesis pathway was upregulated in patients (p < 0.0001, Mann-Whitney U-test). A subgroup analysis comparing patients with an active non-infectious AU to their age- and sex-matched HLA-B27-negative controls, showed an increase of the species Phocaeicola vulgatus in active AU (p = 0.0530, Mann-Whitney U-test). An additional analysis comparing AU patients to age- and sex-matched HLA-B27-positive controls, showed an increase of the species Bacteroides caccae in controls (p = 0.0022, Mann-Whitney U-test).ConclusionIn our cohort, non-infectious AU development is associated with compositional and functional alterations of the GM. Further research is needed to assess the causality of these associations, offering potentially novel therapeutic strategies.

Small Intestinal Bacterial Overgrowth (SIBO) and Twelve Groups of Related Diseases—Current State of Knowledge

The human gut microbiota creates a complex microbial ecosystem, characterized by its high population density, wide diversity, and complex interactions. Any imbalance of the intestinal microbiome, whether qualitative or quantitative, may have serious consequences for human health, including small intestinal bacterial overgrowth (SIBO). SIBO is defined as an increase in the number of bacteria (103–105 CFU/mL), an alteration in the bacterial composition, or both in the small intestine. The PubMed, Science Direct, Web of Science, EMBASE, and Medline databases were searched for studies on SIBO and related diseases. These diseases were divided into 12 groups: (1) gastrointestinal disorders; (2) autoimmune disease; (3) cardiovascular system disease; (4) metabolic disease; (5) endocrine disorders; (6) nephrological disorders; (7) dermatological diseases; (8) neurological diseases (9); developmental disorders; (10) mental disorders; (11) genetic diseases; and (12) gastrointestinal cancer. The purpose of this comprehensive review is to present the current state of knowledge on the relationships between SIBO and these 12 disease groups, taking into account risk factors and the causal context. This review fills the evidence gap on SIBO and presents a biological–medical approach to the problem, clearly showing the groups and diseases having a proven relationship with SIBO, as well as indicating groups within which research should continue to be expanded.

Structural analysis and in vitro fermentation characteristics of an Avicennia marina fruit RG-I pectin as a potential prebiotic

Avicennia marina (Forssk.) Vierh. is a highly salt-tolerant mangrove, and its fruit has been traditionally used for treating constipation and dysentery. In this study, a pectin (AMFPs-0-1) was extracted and isolated from this fruit for the first time, its structure was analyzed, and the effects on the human gut microbiota were investigated. The results indicated that AMFPs-0-1 with a molecular weight of 798 kDa had a backbone consisting of alternating →2)-α-L-Rhap-(1→ and →4)-α-D-GalpA-(1→ residues and side chains composed of →3-α-L-Araf-(1→-linked arabinan with a terminal β-L-Araf, →5-α-L-Araf-(1→-linked arabinan, and →4)-β-D-Galp-(1→-linked galactan that linked to the C-4 positions of all α-L-Rhap residues in the backbone. It belongs to a type I rhamnogalacturonan (RG-I) pectin but has no arabinogalactosyl chains. AMFPs-0-1 could be consumed by human gut microbiota and increase the abundance of some beneficial bacteria, such as Bifidobacterium, Mitsuokella, and Megasphaera, which could help fight digestive disorders. These findings provide a structural basis for the potential application of A. marina fruit RG-I pectic polysaccharides in improving human intestinal health.

2 Another renaissance for bile acid gastrointestinal microbiology

Abstract The field of bile acid microbiology in the gastrointestinal tract is going through a current rebirth after a peak of activity in the late 1970s and early 1980s. This renewed activity is a result of many factors including the discovery near the turn of the century that bile acids are potent signaling molecules, technological advances in next generation sequencing, computation, culturomics, gnotobiology, and metabolomics. The current state of the field with particular emphasis on questions that have remained unanswered for many decades in both bile acid synthesis by the host, and metabolism by the gut microbiota will be addressed. Current knowledge of established enzymatic pathways including bile salt hydrolase, hydroxysteroid dehydrogenases involved in the oxidation and epimerization of bile acid hydroxy groups, the Hylemon-Bjӧrkhem Pathway (HBP) of bile acid C7-dehydroxylation, and the formation of secondary allo-bile acids are described. Aspects of bile acid conjugation and esterification, and evidence for bile acid C3- and C12-dehydroxylation that are less well understood, but potentially critical for our understanding of bile acid metabolism in the human gut will be covered. The physiological consequences of bile acid metabolism for human health, important caveats and cautionary notes on experimental design and interpretation of bile acid metabolism are also explored.

Critical complex network structures in animal gastrointestinal tract microbiomes

BackgroundLiving things from microbes to their hosts (plants, animals and humans) interact with each other, and their relationships may be described with complex network models. The present study focuses on the critical network structures, specifically the core/periphery nodes and backbones (paths of high-salience skeletons) in animal gastrointestinal microbiomes (AGMs) networks. The core/periphery network (CPN) mirrors nearly ubiquitous nestedness in ecological communities, particularly dividing the network as densely interconnected core-species and periphery-species that only sparsely linked to the core. Complementarily, the high-salience skeleton network (HSN) mirrors the pervasive asymmetrical species interactions (strictly microbial species correlations), particularly forming heterogenous pathways in AGM networks with both “backbones” and “rural roads” (regular or weak links). While the cores and backbones can act as critical functional structures, the periphery nodes and weak links may stabilize network functionalities through redundancy.ResultsHere, we build and analyze 36 pairs of CPN/HSN for the AGMs based on 4903 gastrointestinal-microbiome samples containing 473,359 microbial species collected from 318 animal species covering all vertebrate and four major invertebrate classes. The network analyses were performed at host species, order, class, phylum, kingdom scales and diet types with selected and comparative taxon pairs. Besides diet types, the influence of host phylogeny, measured with phylogenetic (evolutionary) timeline or “age”, were integrated into the analyses. For example, it was found that the evolutionary trends of three primary microbial phyla (Bacteroidetes/Firmicutes/Proteobacteria) and their pairwise abundance-ratios in animals do not mirror the patterns in modern humans phylogenetically, although they are consistent in terms of diet types.ConclusionsOverall, the critical network structures of AGMs are qualitatively and structurally similar to those of the human gut microbiomes. Nevertheless, it appears that the critical composition (the three phyla of Bacteroidetes, Firmicutes, and Proteobacteria) in human gut microbiomes has broken the evolutionary trend from animals to humans, possibly attributable to the Anthropocene epoch and reflecting the far-reaching influences of agriculture and industrial revolution on the human gut microbiomes. The influences may have led to the deviations between modern humans and our hunter-gather ancestors and animals.

Complete genome sequence of Lacticaseibacillus paracasei A02.

Lacticaseibacillus paracasei, a facultative anaerobic gram-positive bacterium, is commonly found in the gut of humans and animals, as well as in dairy products and plant ferments. Here, we report the complete genome sequence of L. paracasei A02, whose total genome length is 3,038,472 bp, with a GC content of 46.41%.

Human gut microbiota and their production of endocannabinoid-like mediators are directly affected by a dietary oil

ABSTRACT Dietary omega-3 polyunsaturated fatty acids (n-3 PUFAs) and the gut microbiome affect each other. We investigated the impact of supplementation with Buglossoides arvensis oil (BO), rich in stearidonic acid (SDA), on the human gut microbiome. Employing the Mucosal Simulator of the Human Intestinal Microbial Ecosystem (M-SHIME), we simulated the ileal and ascending colon microbiomes of four donors. Our results reveal two distinct microbiota clusters influenced by BO, exhibiting shared and contrasting shifts. Notably, Bacteroides and Clostridia abundance underwent similar changes in both clusters, accompanied by increased propionate production in the colon. However, in the ileum, cluster 2 displayed a higher metabolic activity in terms of BO-induced propionate levels. Accordingly, a triad of bacterial members involved in propionate production through the succinate pathway, namely Bacteroides, Parabacteroides, and Phascolarctobacterium, was identified particularly in this cluster, which also showed a surge of second-generation probiotics, such as Akkermansia, in the colon. Finally, we describe for the first time the capability of gut bacteria to produce N-acyl-ethanolamines, and particularly the SDA-derived N-stearidonoyl-ethanolamine, following BO supplementation, which also stimulated the production of another bioactive endocannabinoid-like molecule, commendamide, in both cases with variations across individuals. Spearman correlations enabled the identification of bacterial genera potentially involved in endocannabinoid-like molecule production, such as, in agreement with previous reports, Bacteroides in the case of commendamide. This study suggests that the potential health benefits on the human microbiome of certain dietary oils may be amenable to stratified nutrition strategies and extend beyond n-3 PUFAs to include microbiota-derived endocannabinoid-like mediators.

Genome plasticity shapes the ecology and evolution of Phocaeicola dorei and Phocaeicola vulgatus

Phocaeicola dorei and Phocaeicola vulgatus are very common and abundant members of the human gut microbiome and play an important role in the infant gut microbiome. These species are closely related and often confused for one another; yet, their genome comparison, interspecific diversity, and evolutionary relationships have not been studied in detail so far. Here, we perform phylogenetic analysis and comparative genomic analyses of these two Phocaeicola species. We report that P. dorei has a larger genome yet a smaller pan-genome than P. vulgatus. We found that this is likely because P. vulgatus is more plastic than P. dorei, with a larger repertoire of genetic mobile elements and fewer anti-phage defense systems. We also found that P. dorei directly descends from a clade of P. vulgatus¸ and experienced genome expansion through genetic drift and horizontal gene transfer. Overall, P. dorei and P. vulgatus have very different functional and carbohydrate utilisation profiles, hinting at different ecological strategies, yet they present similar antimicrobial resistance profiles.

The mystery of rich human gut antibiotic resistome in the Yellow River with hyper-concentrated sediment-laden flow

Human gut antibiotic resistome widely occur in anoxic environments characterized by high density of bacterial cells and frequent transmission of antibiotic resistance genes (ARGs). Such resistome is greatly diluted, degraded, and restrained in the aerobic habitats within most natural rivers (regarded as “terrestrial guts”) connecting continents and the oceans. Here we implemented a large-scale monitoring campaign extending 5,200 km along the Yellow River, and provide the first integral biogeographic pattern for both ARGs and their hosts. We identified plentiful ARGs (24 types and 809 subtypes) and their hosts (24 phyla and 757 MAGs) in three media (water, suspended particulate matter (SPM), and sediment). Unexpectedly, we found diverse human gut bacteria (HGB) acting as supercarriers of ARGs in this oxygen-rich river. We further discovered that numerous microhabitats were created within stratified biofilms that surround SPMs, particularly regarding the aggregation of anaerobic HGB. These microhabitats provide numerous ideal sinks for anaerobic bacteria and facilitate horizontal transfer of ARGs within the stratified biofilms, Furthermore, the stratification of biofilms surrounding SPMs has facilitated synergy between human gut flora and denitrifiers for propagation of ARGs in the anoxic atmospheres, leading to high occurrence of human gut antibiotic resistome. SPMs play active roles in the dynamic interactions of river water and sediment, thus accelerating the evolution of riverine resistome and transmission of human gut antibiotic resistome. This study revealed the special contribution of SPMs to the propagation of ARGs, and highlighted the necessity of making alternative strategies for sustainable management of large rivers with hyper-concentrated sediment-laden flows.

Sequence similarity network analysis of drug- and dye-modifying azoreductase enzymes found in the human gut microbiome

Drug metabolism by human gut microbes is often exemplified by azo bond reduction in the anticolitic prodrug sulfasalazine. Azoreductase activity is often found in incubations with cell cultures or ex vivo gut microbiome samples and contributes to the xenobiotic metabolism of drugs and food additives. Applying metagenomic studies to personalized medicine requires knowledge of the genes responsible for sulfasalazine and other drug metabolism, and candidate genes and proteins for drug modifications are understudied. A representative gut-abundant azoreductase from Anaerotignum lactatifermentan DSM 14214 efficiently reduces sulfasalazine and another drug, phenazopyridine, but could not reduce all azo-bonded drugs in this class. We used enzyme kinetics to characterize this enzyme for its NADH-dependent reduction of these drugs and food additives and performed computational docking to provide the groundwork for understanding substrate specificity in this family. We performed an analysis of the Flavodoxin-like fold InterPro family (IPR003680) by computing a sequence similarity network to classify distinct subgroups of the family and then performed chemically-guided functional profiling to identify proteins that are abundant in the NIH Human Microbiome Project dataset. This strategy aims to reduce the number of unique azoreductases needed to characterize one protein family in the diverse set of potential drug- and dye-modifying activities found in the human gut microbiome.

Pairing metagenomics and Metaproteomics to characterize ecological niches and metabolic essentiality of gut microbiomes

Pairing metagenomics and Metaproteomics to characterize ecological niches and metabolic essentiality of gut microbiomes