Bilophila Research Articles

P924 A unique gut microbiome signature can help differentiate Crohn`s Disease and isolated Terminal Ileitis

Abstract Background With increasing number of ileal intubations, isolated terminal ileal ulcers(TI) are often detected during colonoscopies. The etiology of these lesions have not been well-defined and differentiating TI from early Crohn’s Disease (CD) is a commonly encountered diagnostic challenge. CD is associated with significant dysbiosis; however, the gut microbiome of TI is poorly studied. We aimed to compare the gut microbial profile between CD and isolated non-specific TI. Methods Fecal samples were collected from treatment naïve patients (26 CD;13 TI) and healthy volunteers (29) without the use of antibiotics, probiotics, and prebiotics for at least 6 weeks. CD was confirmed by endoscopic and histological examination. TI was diagnosed based on isolated terminal ileal erosions/ulcers, normal small bowel endoscopy, non-specific inflammation with no granulomas/ cryptitis/crypt abscess on biopsy, and no history of NSAIDs. Fecal DNA was isolated and sequenced(16S-V4). Taxonomical classification was performed by Amplicon Sequence Variant approach and analysed by STAMP (Statistical Analysis of Metagenomic Profiles) using Welch`s T-Test. PICRUSt (Phylogenetic Investigation of Communities by Reconstruction of Unobserved States) was used for predictive metagenomics. Significant metabolic pathways were corrected by Storey FDR (q<0.05). Fecal Calprotectin (FCal) was measured using QuantOnCal rapid test. Results Microbiome data analysis revealed a progressive decline in α-diversity(p=0.002) in CD and different β-diversity (Bray-Curtis dissimilarity p=0.001) compared to TI and healthy(Fig1). Overall Firmicutes were lower(p=9.81e-4) while Desulfobacterota were higher(p=8.65e-3) in CD than TI(Fig2A). Faecalibacterium prausnitzii (p=0.04) and Roseburia inulinivorans (p=0.03) were markedly reduced while Parabacteroides distasonis (p=9.91e-3) and Bilophila wadsworthia (p=7.36e-3) were elevated in CD (Fig3). A microbial signature covering 8 non-overlapping taxa was deduced to distinguish CD and TI (3 taxa reduced, 5 elevated in CD compared to TI) (Fig2B). PICRUSt analysis indicated 18 significant metabolic pathways differentiating CD and TI. Bacterial stress alarmone PPGPP-MET was significantly higher in CD (p=2.04e-3) (Fig4). FCal was significantly higher in CD with mean value (±SD) as 549.12±390.4 and 167.15±191.8µg/g in TI (p=0.002). Conclusion A distinct gut microbiome profile was observed in TI compared to CD. Exploitation of these differences can potentiate differential diagnosis of CD and TI. Further research on the spectrum of dysbiosis is warranted. This may help in differentiating patients with early-CD who require active therapy from those with non-specific TI ulcers that resolve over time.

Alterations in Gut Microbiota Composition in Patients with COVID-19: A Pilot Study of Whole Hypervariable 16S rRNA Gene Sequencing.

It is crucial to consider the importance of the microbiome and the gut-lung axis in the context of SARS-CoV-2 infection. This pilot study examined the fecal microbial composition of patients with COVID-19 following a 3-month recovery. Using for the first time metagenomic analysis based on all hypervariable regions (V1-V9) of the 16S rRNA gene, we have identified 561 microbial species; however, 17 were specific only for the COVID-19 group (n = 8). The patients' cohorts revealed significantly greater alpha diversity of the gut microbiota compared to healthy controls (n = 14). This finding has been demonstrated by operational taxonomic units (OTUs) richness (p < 0.001) and Chao1 index (p < 0.01). The abundance of the phylum Verrucomicrobia was 30 times higher in COVID-19 patients compared to healthy subjects. Accordingly, this disproportion was also noted at other taxonomic levels: in the class Verrucomicrobiae, the family Verrucomicrobiaceae, and the genus Akkermansia. Elevated pathobionts such as Escherichia coli, Bilophila wadsworthia, and Parabacteroides distasonis were found in COVID-19 patients. Considering the gut microbiota's ability to disturb the immune response, our findings suggest the importance of the enteric microbiota in the course of SARS-CoV-2 infection. This pilot study shows that the composition of the microbial community may not be fully restored in individuals with SARS-CoV-2 following a 3-month recovery.

Beneficial metabolic transformations and prebiotic potential of hemp bran and its alcalase hydrolysate, after colonic fermentation in a gut model

Hemp seed bran (HB) is an industrial food byproduct that is generally discarded. Knowledge on the functional capabilities of HB is limited and it is not known the impact of HB on human colon microbiota, where vegetable fibers are metabolized. In this work, we investigated in depth the prebiotic potential of HB and HB protein extract hydrolyzed by alcalase (HBPA) in comparison to fructooligosaccharides (FOS) after human distal colonic fermentation using MICODE (multi-unit in vitro colon gut model). During the 24 h of fermentation, metabolomics (SPME GC/MS) and microbiomics (MiSeq and qPCR) analyses were performed. The results indicated that HBPA on a colonic fermentation had a higher prebiotic index than HB (p < 0.05), and slightly lower to that of FOS (p > 0.05). This feature was described and explained as HBPA colonic fermentation produces beneficial organic fatty acids (e.g. Pentanoic and Hexanoic acids); reduces detrimental phenol derivates (e.g. p-Cresol); produces bioactives VOCs (e.g. Acetophenone or 4-Terpineol); increases beneficial bacteria (e.g. 1.76 fold and 2.07 fold more of Bifidobacterium bifidum and Bacteroides fragilis, respectively) and limits opportunistic bacteria (e.g. 3.04 fold and 2.07 fold less of Bilophila wadsworthia and Desulfovibrio, respectively). Our study evidenced the prebiotic role of HB and HBPA, and within the principles of OneHealth it valorizes a byproduct from the queen plant of sustainable crops as a food supplement.

TRAUMATIC BRAIN INJURY-INDUCED INFLAMMATION AND GASTROINTESTINAL MOTILITY DYSFUNCTION.

Background: Traumatic brain injury (TBI) is a significant cause of morbidity and mortality in the United States, with an annual cost of 60 billion dollars. There is evidence suggesting that in the post-TBI period, the gastrointestinal tract plays a central role in driving organ and immune dysfunction and may be the source of increased circulating proinflammatory mediators. In this study, we examined systemic inflammation and bacterial dysbiosis in patients who sustained a TBI with or without polytrauma. Using a mouse model of TBI, we further show how neuroinflammation after TBI is potentially linked to disruptions in gut homeostasis such as intestinal transit and inflammation. Methods: During a study of trauma patients performed from September 1, 2018, to September 1, 2019, at a single, level 1 trauma center, TBI patients aged 21 to 95 years were enrolled. Patients were categorized as TBI based on evidence of acute abnormal findings on head computed tomographic scan, which was a combination of isolated TBI and TBI with polytrauma. Blood and stool samples were collected between 24 h and 3 days after admission. Twelve plasma samples and 10 fecal samples were used for this study. Healthy control samples were obtained from a healthy control biobank. We examined systemic inflammation and bacterial changes in patients who sustained a TBI. In addition, TBI was induced in 9- to 10-week-old male mice; we assessed neuroinflammation, and intestine transit (motility) and bacterial changes 24 h after TBI. Results: When compared with healthy controls, TBI patients had increased systemic inflammation as evidenced by increased levels of IFN-γ and MCP-1 and a trend toward an increase of IL-6 and IL-8 ( P = 0.0551 and P = 0.0549), respectively. The anti-inflammatory cytokine, IL-4, was also decreased in TBI patients. Although there was a trend of an increase in copy number of Enterobacteriaceae and a decrease in copy number of Lactobacillus in both patients and mice after TBI, these trends were not found to be significantly different. However, TBI significantly increased the copy number of another potential pathogenic bacteria Bilophila wadsworthia in TBI patients compared with healthy controls. After a moderate TBI, mice had increased expression of TNF-α, IL-6 and IL-1β, CXCL1, s100a9, and Ly6G and decreased IL-10 in the brain lesion after TBI. This accompanied decreased transit and increased TNF-α in the small intestine of mice after TBI. Conclusions: Our findings suggest that TBI increases systemic inflammation, intestinal dysfunction, and neuroinflammation. More studies are needed to confirm whether changes in intestinal motility play a role in post-TBI neuroinflammation and cognitive deficit.

Abstract 3524: Role of the gut microbiota in the development of immune-mediated diarrhea and colitis induced by immune checkpoint inhibitors

Abstract The accelerating use of combined immune checkpoint inhibitors (ICI; anti-PD-1 plus anti-CTLA-4 antibodies) enhanced therapeutic responses in various cancers while significantly increasing immune-related adverse events (irAE). The mechanism involved in the occurrence of immune-mediated diarrhea and colitis (IMDC) and its potential implication on ICI efficacy remain unclear. The gut microbiome is progressively recognized as a key biomarker of ICI efficacy and preliminary results suggest its potential association with irAE. However, the link between microbiota and IMDC still needs to be elucidated. In this study, we investigated IMDC-associated intestinal bacteria.We performed metagenomics sequencing of baseline pre-ICI fecal samples from 26 advanced melanoma patients and recorded IMDC. Culturomics using MALDI-TOF was used to isolate bacteria from 19 patient samples: 11 IMDC-free (fIMDC), 5 baseline pre-IMDC (pIMDC) and 8 during IMDC (dIMDC). Serum multiplex cytokine panel was measured for 36 patients. Then, we induced colitis with Dextran Sulfate Sodium (DSS) in MCA-205 murine tumor model treated with oral supplementation of human bacteria and ICI.Metagenomics profiling demonstrated lower α-diversity at baseline in patients that developed irAE. Moreover, pIMDC samples present a lower Simpson α-diversity than fIMDC samples (p=0.08). ß-diversity was also different between both groups (p=0.10) suggesting a different baseline microbiome composition. Lefse analysis revealed an overrepresentation of Enterococcus faecium and Bilophila wadsworthia in pIMDC samples compared to fIMDC samples. Culturomics showed that dIMDC samples exhibited a shift in gut microbiota with a significant decrease in bacterial diversity mostly anaerobic species and an enrichment of Clostridium paraputricum, Clostridium perfringens, Paraclostridium bifermentans and Paeniclostridium sordelii compared to fIMDC and pIMDC samples. Immunological analysis of serum revealed that IMDC patients had a higher concentration of pro-inflammatory cytokines including GMCSF (p=0.04), IL12-23p40 (p=0.04) and IL-16 (p=0.03) compared to fIMDC patients. In our DSS-induced colitis mouse model, we showed that P. bifermentans oral supplementation related with diarrhea and a shorter colon length, indicating more severe colitis. Moreover, we showed that P. bifermentans increased anti-CLTA-4 and anti-PD-1 anti-tumor activity.Results showed that gut microbiota composition of dIMDC patients differed from that of pIMDC and fIMDC patients with a decrease of diversity and colonization of P.bifermentans and P.sordelii. In IMDC patients, we isolated bacteria that aggravated colitis in murine models and are known to be associated with inflammatory bowel disease. These results provide more insight on potential microbiome-modifying strategies to damper IMDC and increase anti-cancer efficacy. Citation Format: Khoudia Diop, Benlaïfaoui Myriam, Wiam Belkaïd, Alexis Nolin-Lapalme, Julie Malo, Marion Tonneau, Ponce Mayra, Afnan Al-Saleh, Catalin Mihalcioiu, Ian Watson, Arielle Elkrief, Karla Lee, Meriem Messaoudene, Bertrand Routy, Corentin Richard. Role of the gut microbiota in the development of immune-mediated diarrhea and colitis induced by immune checkpoint inhibitors [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3524.

Shotgun metagenomic sequencing revealed the prebiotic potential of a grain-based diet in mice

In the present study, we elucidated the effect of grain-based (GB) diet containing both soluble and insoluble fibers and purified ingredients-based (PIB) diet containing only insoluble fiber, namely cellulose on mice gut microbiome using whole shotgun based metagenomic sequencing. Although the fiber content in both diet types is the same (5%) the presence of soluble fiber only in the GB diet differentiates it from the PIB diet. The taxonomic analysis of sequenced reads reveals a significantly higher enrichment of probiotic Lactobacilli in the GB group as compared to the PIB group. Further, the enhancement of energy expensive cellular processes namely, cell cycle control, cell division, chromosome partitioning, and transcription is observed in the GB group which could be due to the metabolization of the soluble fiber for faster energy production. In contrast, a higher abundance of cellulolytic bacterial community namely, the members of family Lachnospiraceae and Ruminococcaceae and the metabolism functions are found in the PIB group. The PIB group shows a significant increase in host-derived oligosaccharide metabolism functions indicating that they might first target the host-derived oligosaccharides and self-stored glycogen in addition to utilising the available cellulose. In addition to the beneficial microbial community variations, both the groups also exhibited an increased abundance of opportunistic pathobionts which could be due to an overall low amount of fiber in the diet. Furthermore, backtracing analysis identified probiotic members of Lactobacillus, viz., L. crispatus ST1, L. fermentum CECT 5716, L. gasseri ATCC 33323, L. johnsonii NCC 533 and L. reuteri 100-23 in the GB group, while Bilophila wadsworthia 3_1_6, Desulfovibrio piger ATCC 29098, Clostridium symbiosum WAL-14163, and Ruminococcaceae bacterium D16 in the PIB group. These data suggest that Lactobacilli, a probiotic community of microorganisms, are the predominant functional contributors in the gut of GB diet-fed mice, whereas pathobionts too coexisted with commensals in the gut microbiome of the PIB group. Thus at 5% fiber, GB modifies the gut microbial ecology more effectively than PIB and the inclusion of soluble fiber in the GB diet may be one of the primary factors responsible for this impact.

Gut Microbial Signatures of Distinct Trimethylamine N-Oxide Response to Raspberry Consumption.

The aim of this exploratory study was to evaluate the gut microbial signatures of distinct trimethylamine N-oxide (TMAO) responses following raspberry consumption. Investigations were carried out in 24 subjects at risk of developing metabolic syndrome who received 280 g/day of frozen raspberries for 8 weeks. Blood and stool samples were collected at weeks 0 and 8. Inter-individual variability in plasma TMAO levels was analyzed, 7 subjects were excluded due to noninformative signals and 17 subjects were kept for analysis and further stratified according to their TMAO response. Whole-metagenome shotgun sequencing analysis was used to determine the impact of raspberry consumption on gut microbial composition. Before the intervention, the relative abundance of Actinobacteriota was significantly higher in participants whose TMAO levels increased after the intervention (p = 0.03). The delta TMAO (absolute differences of baseline and week 8 levels) was positively associated with the abundance of gut bacteria such as Bilophila wadsworthia (p = 0.02; r2 = 0.37), from the genus Granulicatella (p = 0.03; r2 = 0.48) or the Erysipelotrichia class (p = 0.03; r2 = 0.45). Changes in the gut microbial ecology induced by raspberry consumption over an 8-week period presumably impacted quaternary amines-utilizing activity and thus plasma TMAO levels.

Compositional Alterations of Gut Microbiota in Patients with Diabetic Kidney Disease and Type 2 Diabetes Mellitus

PurposeEmerging evidence has revealed that gut microbiota plays a pivotal role in the pathogenesis of type 2 diabetes mellitus (T2DM) and diabetic kidney disease (DKD). However, few studies have used metagenomic sequencing to analyze the alterations of gut microbiota community in patients with early-stage DKD.MethodsWe carried out metagenomic sequencing in fecal samples of 10 DKD patients (DKD group) and 10 T2DM patients who appeared to be less prone to DKD (non-DKD group), aiming to compare the composition and function of gut microbiota between the DKD and non-DKD groups.ResultsThe gut microbial community of the DKD group was significantly different from that of the non-DKD group, characterized by a marked increase in phylum Proteobacteria, genus Selenomonadales, Neosynechococcus, Shigella, Bilophila, Acidaminococcus, species, Escherichia coli, Bacteroides plebeius, Megasphaera elsdenii, Acidaminococcus unclassified, and Bilophila wadsworthia. The amounts of species Citrobacter farmeri and Syntrophaceticus schinkii were significantly and positively correlated with the urinary albumin creatinine ratio in the DKD group. Furthermore, functional analysis based on dbCAN and KEGG databases showed aberrant lipopolysaccharide (LPS) biosynthesis and carbohydrate metabolism in the gut microbiome of the DKD group.ConclusionOur findings provided evidence for alterations in the composition and function of gut microbiota in patients with DKD versus the non-DKD group. These data may contribute to a more comprehensive understanding of the pathological mechanisms of DKD.

Alteration of Gut Microbiome in Patients With Schizophrenia Indicates Links Between Bacterial Tyrosine Biosynthesis and Cognitive Dysfunction

BackgroundSchizophrenia (SCZ) is a heterogeneous neuropsychiatric disorder for which current treatment has insufficient efficacy and severe adverse effects. The modifiable gut microbiome might be a potential target for intervention to improve neurobiological functions through the gut-microbiome-brain axis. MethodsIn this case-control study, gut microbiota of 132 patients with SCZ and increased waist circumference were compared with gut microbiota of two age- and sex-matched control groups, composed of 132 healthy individuals and 132 individuals with metabolic syndrome. Shotgun sequencing was used to characterize fecal samples at the taxonomic and functional levels. Cognition of the patients with SCZ was evaluated using the Brief Assessment of Cognition instrument. ResultsSCZ gut microbiota differed significantly from those of healthy control subjects and individuals with metabolic syndrome in terms of richness and global composition. SCZ gut microbiota were notably enriched in Flavonifractor plautii, Collinsella aerofaciens, Bilophila wadsworthia, and Sellimonas intestinalis, while depleted in Faecalibacterium prausnitzii, Ruminococcus lactaris, Ruminococcus bicirculans, and Veillonella rogosae. Functional potential of the gut microbiota accounted for 11% of cognition variability. In particular, the bacterial functional module for synthesizing tyrosine, a precursor for dopamine, was in SCZ cases positively associated with cognitive score (ρ = 0.34, q ≤ .1). ConclusionsOverall, this study shows that the gut microbiome of patients with SCZ differs greatly from that of healthy control subjects or individuals with metabolic syndrome. Cognitive function of patients with SCZ is associated with the potential for gut bacterial biosynthesis of tyrosine, a precursor for dopamine, suggesting that gut microbiota might be an intervention target for alleviation of cognitive dysfunction in SCZ.

Gut Microbiota Composition Is Related to AD Pathology.

IntroductionSeveral studies have reported alterations in gut microbiota composition of Alzheimer’s disease (AD) patients. However, the observed differences are not consistent across studies. We aimed to investigate associations between gut microbiota composition and AD biomarkers using machine learning models in patients with AD dementia, mild cognitive impairment (MCI) and subjective cognitive decline (SCD).Materials and MethodsWe included 170 patients from the Amsterdam Dementia Cohort, comprising 33 with AD dementia (66 ± 8 years, 46%F, mini-mental state examination (MMSE) 21[19-24]), 21 with MCI (64 ± 8 years, 43%F, MMSE 27[25-29]) and 116 with SCD (62 ± 8 years, 44%F, MMSE 29[28-30]). Fecal samples were collected and gut microbiome composition was determined using 16S rRNA sequencing. Biomarkers of AD included cerebrospinal fluid (CSF) amyloid-beta 1-42 (amyloid) and phosphorylated tau (p-tau), and MRI visual scores (medial temporal atrophy, global cortical atrophy, white matter hyperintensities). Associations between gut microbiota composition and dichotomized AD biomarkers were assessed with machine learning classification models. The two models with the highest area under the curve (AUC) were selected for logistic regression, to assess associations between the 20 best predicting microbes and the outcome measures from these machine learning models while adjusting for age, sex, BMI, diabetes, medication use, and MMSE.ResultsThe machine learning prediction for amyloid and p-tau from microbiota composition performed best with AUCs of 0.64 and 0.63. Highest ranked microbes included several short chain fatty acid (SCFA)-producing species. Higher abundance of [Clostridium] leptum and lower abundance of [Eubacterium] ventriosum group spp., Lachnospiraceae spp., Marvinbryantia spp., Monoglobus spp., [Ruminococcus] torques group spp., Roseburia hominis, and Christensenellaceae R-7 spp., was associated with higher odds of amyloid positivity. We found associations between lower abundance of Lachnospiraceae spp., Lachnoclostridium spp., Roseburia hominis and Bilophila wadsworthia and higher odds of positive p-tau status.ConclusionsGut microbiota composition was associated with amyloid and p-tau status. We extend on recent studies that observed associations between SCFA levels and AD CSF biomarkers by showing that lower abundances of SCFA-producing microbes were associated with higher odds of positive amyloid and p-tau status.

The antioxidant tempol transforms gut microbiome to resist obesity in female C3H mice fed a high fat diet

The nitroxide, Tempol, prevents obesity related changes in mice fed a high fat diet (HFD). The purpose of this study was to gain insight into the mechanisms that result in such changes by Tempol in female C3H mice. Microarray methodology, Western blotting, bile acid analyses, and gut microbiome sequencing were used to identify multiple genes, proteins, bile acids, and bacteria that are regulated by Tempol in female C3H mice on HFD. The effects of antibiotics in combination with Tempol on the gut microflora were also studied. Adipose tissue, from Tempol treated mice, was analyzed using targeted gene microarrays revealing up-regulation of fatty acid metabolism genes (Acadm and Acadl > 4-fold, and Acsm3 and Acsm5 > 10-fold). Gene microarray studies of liver tissue from mice switched from HFD to Tempol HFD showed down-regulation of fatty acid synthesis genes and up-regulation of fatty acid oxidation genes. Analyses of proteins involved in obesity revealed that the expression of aldehyde dehydrogenase 1A1 (ALDH1A1) and fasting induced adipose factor/angiopoietin-like protein 4 (FIAF/ANGPTL4) was altered by Tempol HFD. Bile acid studies revealed increases in cholic acid (CA) and deoxycholic acid (DCA) in both the liver and serum of Tempol treated mice. Tempol HFD effect on the gut microbiome composition showed an increase in the population of Akkermansia muciniphila, a bacterial species known to be associated with a lean, anti-inflammatory phenotype. Antibiotic treatment significantly reduced the total level of bacterial numbers, however, Tempol was still effective in reducing the HFD weight gain. Even after antibiotic treatment Tempol still positively influenced several bacterial species such as as Akkermansia muciniphila and Bilophila wadsworthia. The positive effects of Tempol moderating weight gain in female mice fed a HFD involves changes to the gut microbiome, bile acids composition, and finally to changes in genes and proteins involved in fatty acid metabolism and storage.

Preparation and Characterization of a New Monoclonal Antibody Specific Against Lawsonia intracellularis and Its Application in Indirect Immunofluorescence and Immunocytochemistry Assay.

Proliferative enteropathy (PE) is an infectious enteric disease caused by Lawsonia intracellularis (L. intracellularis) and is endemic in pig herds worldwide. However, a L. intracellularis-specific monoclonal antibody plays an important role in the evaluation of L. intracellularis infection in vitro. Therefore, the objective of this study was to produce and identify the characteristics of a new monoclonal antibody against the outer membrane protein (Omp2) of L. intracellularis and apply it in an indirect immunofluorescence assay (IFA) and immunocytochemistry (IHC). The results indicated that three highly specific monoclonal antibodies against the Omp2 protein (4D9, 3G2, and 7G5) of L. intracellularis were obtained by using purified Omp2 as an immunogen, the titers of ascitic fluids of 4D9, 3G2, and 7G5 cells were 1:2,048,000, 1:512,000, and 1:256,000, respectively. IFA analysis showed that the 4D9, 3G2, and 7G5 have no cross-reactivity with other enteric bacteria commonly found in the ilea of pigs or closely related to L. intracellularis, such as Desulfovibrio, Bilophila wadsworthia (B. wadsworthia), Salmonella choleraesuis (S. choleraesuis), Salmonella typhimurium (S. typhimurium), Escherichia coli (E. coli), and Brachyspira hyodysenteriae (B. hyodysenteriae). IFA and IHC results indicated that the monoclonal antibodies can be successfully used as primary antibodies to detect L. intracellularis in infected cells and in the crypt of the ileum from infected tissues of PE. Our findings suggested that the new monoclonal antibody specific against L. intracellularis will be useful for the evaluation of L. intracellularis infection in vivo and in vitro.

20.5 Preliminary Evidence for Associations of the Infant Gut Colonization With Parent-Infant Relationship, Maternal Depression, Stress, and Parenting Styles

The objectives of this presentation are to: 1) examine associations among infant gut colonization based on microbiome measurements in the infant population and maternal depressive symptoms and distress; 2) examine associations among infant microbiome and parent-infant interactions during a vaccination procedure; and 3) examine how infant cries correlate with their microbiome at 4 months of age. For this pilot study, we recruited a prospective cohort of infants (n = 31) and parents (n = 31). We examined the association between the infant gut microbiome at 4 months during well-child visits (WCVs), maternal depression (by Patient Health Questionnaire-9 [PHQ-9]), parent and child characteristics and life stress (by Parenting Stress Index [PSI-IV]), and parent-infant interactions (by Measure of Adult and Infant Soothing and Distress [MAISD]). For each of the variables, we looked at 3 types of associations to identify the component of the microbiome revealed by whole-genome shotgun sequencing: β-diversity (via Bray-Curtis [BC] and Jaccard distances), α-diversity (via Shannon and Inverse-Simpson indexes), and differential abundance (via linear models for differential abundance [LinDA]). Infant cries in response to vaccination were assessed for spectral characteristics, with higher frequencies being associated with greater stress response (mid frequency: 1000-5000 Hz, high frequency: 5000 Hz+). Taxonomic classification identified 19 phyla, 417 genera, and 1246 species. The PHQ-9 scores obtained at 2 months were associated with 4 bacterial species (Ruminococcus flavefaciens, Actinomyces johnsonii, [Clostridium] dakarense, and Bilophila wadsworthia; false discovery rate [FDR] < 0.1) and β-diversity (p = 0.006-BC; p = 0.005-Jaccard). Infant cries with greater high-frequency band power (p < 0.03) and a greater high-to-mid frequency ratio metrics (p < 0.05) associated with higher infant stress were related to lower α-diversity. There were no correlations between PHQ-9 at 4 months, PSI-IV, and microbiome diversity. Data from our study point to a connection between higher perception of stress in infants reflected in their cries and lower microbiome diversity (higher diversity more desirable). We found that parental depressive symptoms influenced their infants’ microbiome. More studies are needed to explore ways in which the quality of the infant-caregiver relationship influences intestinal microbiota.

Alterations in the gut microbiota contribute to cognitive impairment induced by the ketogenic diet and hypoxia

Many genetic and environmental factors increase susceptibility to cognitive impairment (CI), and the gut microbiome is increasingly implicated. However, the identity of gut microbes associated with CI risk, their effects on CI, and their mechanisms remain unclear. Here, we show that a carbohydrate-restricted (ketogenic) diet potentiates CI induced by intermittent hypoxia in mice and alters the gut microbiota. Depleting the microbiome reduces CI, whereas transplantation of the risk-associated microbiome or monocolonization with Bilophila wadsworthia confers CI in mice fed a standard diet. B.wadsworthia and the risk-associated microbiome disrupt hippocampal synaptic plasticity, neurogenesis, and gene expression. The CI is associated with microbiome-dependent increases in intestinal interferon-gamma (IFNg)-producing Th1 cells. Inhibiting Th1 cell development abrogates the adverse effects of both B.wadsworthia and environmental risk factors on CI. Together, these findings identify select gut bacteria that contribute to environmental risk for CI in mice by promoting inflammation and hippocampal dysfunction.

Nutrient load acts as a driver of gut microbiota load, community composition and metabolic functionality in the simulator of the human intestinal microbial ecosystem.

A recently introduced quantitative framework for gut microbiota analysis indicated that microbial load alterations can be linked to various diseases, making it essential to pinpoint its determinants. We identified nutrient load as a main driver of the quantitative microbial community composition and functionality in vitro by stepwise decreasing standardized feed concentrations from 100% to 33%, 20% and 10% in 5-day intervals. While the proportional composition and metabolic profile were mainly determined by the inter-individual variability (35% and 41%), nutrient load accounted for 58%, 23% and 65% of the observed variation in the microbial load, quantitative composition and net daily metabolite production, respectively. After the 10-fold nutrient reduction, the microbial load decreased by 79.72± 9% and 82.96± 1.66% in the proximal and distal colon, respectively, while the net total short-chain fatty acid production dropped by 79.42± 4.42% and 84.58± 2.42%, respectively. The majority of microbial taxa quantitatively decreased, whereas a select group of nutritional specialists, such as Akkermansia muciniphila and Bilophila wadsworthia, and a number of opportunistic pathogens remained unaffected. This shows that nutrient load is an important driver of the human gut microbiome and should be considered in future in vitro and in vivo dietary research.

Sulfoquinovose is a select nutrient of prominent bacteria and a source of hydrogen sulfide in the human gut

Responses of the microbiota to diet are highly personalized but mechanistically not well understood because many metabolic capabilities and interactions of human gut microorganisms are unknown. Here we show that sulfoquinovose (SQ), a sulfonated monosaccharide omnipresent in green vegetables, is a selective yet relevant substrate for few but ubiquitous bacteria in the human gut. In human feces and in defined co-culture, Eubacterium rectale and Bilophila wadsworthia used recently identified pathways to cooperatively catabolize SQ with 2,3-dihydroxypropane-1-sulfonate as a transient intermediate to hydrogen sulfide (H2S), a key intestinal metabolite with disparate effects on host health. SQ-degradation capability is encoded in almost half of E. rectale genomes but otherwise sparsely distributed among microbial species in the human intestine. However, re-analysis of fecal metatranscriptome datasets of four human cohorts showed that SQ degradation (mostly from E. rectale and Faecalibacterium prausnitzii) and H2S production (mostly from B. wadsworthia) pathways were expressed abundantly across various health states, demonstrating that these microbial functions are core attributes of the human gut. The discovery of green-diet-derived SQ as an exclusive microbial nutrient and an additional source of H2S in the human gut highlights the role of individual dietary compounds and organosulfur metabolism on microbial activity and has implications for precision editing of the gut microbiota by dietary and prebiotic interventions.

Molecular basis of C-S bond cleavage in the glycyl radical enzyme isethionate sulfite-lyase

SummaryDesulfonation of isethionate by the bacterial glycyl radical enzyme (GRE) isethionate sulfite-lyase (IslA) generates sulfite, a substrate for respiration that in turn produces the disease-associated metabolite hydrogen sulfide. Here, we present a 2.7 Å resolution X-ray structure of wild-type IslA from Bilophila wadsworthia with isethionate bound. In comparison with other GREs, alternate positioning of the active site β strands allows for distinct residue positions to contribute to substrate binding. These structural differences, combined with sequence variations, create a highly tailored active site for the binding of the negatively charged isethionate substrate. Through the kinetic analysis of 14 IslA variants and computational analyses, we probe the mechanism by which radical chemistry is used for C-S bond cleavage. This work further elucidates the structural basis of chemistry within the GRE superfamily and will inform structure-based inhibitor design of IsIA and thus of microbial hydrogen sulfide production.

Impact of dietary sulfolipid-derived sulfoquinovose on gut microbiota composition and inflammatory status of colitis-prone interleukin-10-deficient mice

The interplay between diet, intestinal microbiota and host is a major factor impacting health. A diet rich in unsaturated fatty acids has been reported to stimulate the growth of Bilophila wadsworthia by increasing the proportion of the sulfonated bile acid taurocholate (TC). The taurine-induced overgrowth of B. wadsworthia promoted the development of colitis in interleukin-10-deficient (IL-10-/-) mice. This study aimed to investigate whether intake of the sulfonates sulfoquinovosyl diacylglycerols (SQDG) with a dietary supplement or their degradation product sulfoquinovose (SQ), stimulate the growth of B. wadsworthia in a similar manner and, thereby, cause intestinal inflammation. Conventional IL-10-/- mice were fed a diet supplemented with the SQDG-rich cyanobacterium Arthrospira platensis (Spirulina). SQ or TC were orally applied to conventional IL-10-/- mice and gnotobiotic IL-10-/- mice harboring a simplified human intestinal microbiota with or without B. wadsworthia. Analyses of inflammatory parameters revealed that none of the sulfonates induced severe colitis, but both, Spirulina and TC, induced expression of pro-inflammatory cytokines in cecal mucosa. Cell numbers of B. wadsworthia decreased almost two orders of magnitude by Spirulina feeding but slightly increased in gnotobiotic SQ and conventional TC mice. Changes in microbiota composition were observed in feces as a result of Spirulina or TC feeding in conventional mice. In conclusion, the dietary sulfonates SQDG and their metabolite SQ did not elicit bacteria-induced intestinal inflammation in IL-10-/- mice and, thus, do not promote colitis.

Berberine alters gut microbial function through modulation of bile acids

BackgroundBerberine (BBR) is a plant-based nutraceutical that has been used for millennia to treat diarrheal infections and in contemporary medicine to improve patient lipid profiles. Reduction in lipids, particularly cholesterol, is achieved partly through up-regulation of bile acid synthesis and excretion into the gastrointestinal tract (GI). The efficacy of BBR is also thought to be dependent on structural and functional alterations of the gut microbiome. However, knowledge of the effects of BBR on gut microbiome communities is currently lacking. Distinguishing indirect effects of BBR on bacteria through altered bile acid profiles is particularly important in understanding how dietary nutraceuticals alter the microbiome.ResultsGermfree mice were colonized with a defined minimal gut bacterial consortium capable of functional bile acid metabolism (Bacteroides vulgatus, Bacteroides uniformis, Parabacteroides distasonis, Bilophila wadsworthia, Clostridium hylemonae, Clostridium hiranonis, Blautia producta; B4PC2). Multi-omics (bile acid metabolomics, 16S rDNA sequencing, cecal metatranscriptomics) were performed in order to provide a simple in vivo model from which to identify network-based correlations between bile acids and bacterial transcripts in the presence and absence of dietary BBR. Significant alterations in network topology and connectivity in function were observed, despite similarity in gut microbial alpha diversity (P = 0.30) and beta-diversity (P = 0.123) between control and BBR treatment. BBR increased cecal bile acid concentrations, (P < 0.05), most notably deoxycholic acid (DCA) (P < 0.001). Overall, analysis of transcriptomes and correlation networks indicates both bacterial species-specific responses to BBR, as well as functional commonalities among species, such as up-regulation of Na+/H+ antiporter, cell wall synthesis/repair, carbohydrate metabolism and amino acid metabolism. Bile acid concentrations in the GI tract increased significantly during BBR treatment and developed extensive correlation networks with expressed genes in the B4PC2 community.ConclusionsThis work has important implications for interpreting the effects of BBR on structure and function of the complex gut microbiome, which may lead to targeted pharmaceutical interventions aimed to achieve the positive physiological effects previously observed with BBR supplementation.

Multiplexed Quantitative Assessment of the Fate of Taurine and Sulfoquinovose in the Intestinal Microbiome.

(1) Introduction: Sulfonates, which can be diet- or host-derived, are a class of compounds detected in the gut, are involved in host–microbiome interactions and have several health effects. Our aim was to develop a method to quantify five of the sulfonates in the intestine and apply it in a simplified human microbiome model. These were taurine, its metabolic precursor cysteate and one of its degradation products isethionate, as well as sulfoquinovose and one of its most relevant degradation products 2,3-dihydroxy-1-propanesulfonate. (2) Methods: An extraction and sample preparation method was developed, without the need for derivatization. To detect and quantify the extracted sulfonates, a multiplexed LC-MS/MS-MRM method was established. (3) Results: The accuracy and precision of the method were within GLP-accepted parameters. To apply this method in a pilot study, we spiked either taurine or sulfoquinovose into an in vitro simplified human microbiota model with and without Bilophila wadsworthia, a known sulfonate utilizer. The results revealed that only the culture with B. wadsworthia was able to degrade taurine, with isethionate as an intermediate. After spiking the communities with sulfoquinovose, the results revealed that the simplified human microbiome model was able to degrade sulfoquinovose to 2,3-dihydroxypropane-1-sulfonate, which was probably catalyzed by Escherichia coli. In the community with B. wadsworthia, the 2,3-dihydroxypropane-1-sulfonate produced was further degraded by B. wadsworthia to sulfide. (4) Conclusions: We successfully developed a method for sulfonate quantification and applied it in a first pilot study.