Fecal Profiles Research Articles

Fecal metabonomics combined with 16S rDNA sequencing to analyze the changes of gut microbiota in rats fed with different protein source diets.

When blended, animal and plant proteins can complement each other in terms of amino acid composition and release time. In this study, we investigated whether the blended protein diet has a better feeding effect than the single protein diet, and to reveal the differences in growth and intestinal microbiota composition caused by the blended protein diet. Forty Sprague Dawley (SD) rats received diets with different protein sources, including casein (C), whey protein (WP), black soybean protein (BSP), and black soybean-whey blended protein (BS-WP), for eight weeks. To investigate the effects of blended protein supplement on gut microbiota and metabolites, we performed a high throughput 16S rDNA sequencing and fecal metabolomics profiling. In addition, we determined growth and serum biochemical indices, and conducted intestinal morphology analyses. Compared to those in the BSP and WP groups, the daily body weight gain and feed conversion efficiency increased in the BS-WP group. Serum biochemical indices indicated that the protein utilization efficiency of the WP and BS-WP groups was relatively high, and the BS-WP blended protein diet improved the protein adoption rate. The BS-WP blended protein diet also improved intestinal tissue morphology and promoted intestinal villi development compared to the single protein diets. Furthermore, dietary protein altered the composition of gut microbiota, the gut microbial diversity of rats fed with the BS-WP diet was significantly (P < 0.05) higher than that of the other groups. The difference in dietary protein corresponded with an alteration of fecal amino acids and their metabolites, and tryptophan and tyrosine metabolism were the key mechanisms leading to the changes in fecal microbial composition. Dietary protein sources played an important role in the growth and development of rats by influencing intestinal metabolism and microbial composition. The BS-WP blended protein diet was more conducive to nutrient absorption than the single protein diet. Furthermore, blended protein increased the diversity of intestinal microbes and aided the establishment of intestinal barrier function.

Fecal microbiota transplantation for refractory immune-checkpoint-inhibitor colitis.

2657 Background: Immune-related colitis (irColitis) is associated with significant morbidity and mortality among patients treated with immune checkpoint inhibition (ICI). The gut microbiota has been implicated in the pathophysiology of irColitis. We hypothesized that abnormal fecal microbiome features would be present at the time of irColitis onset, and that restoring the microbiome to a healthy state would mitigate disease severity. Methods: We present fecal microbiota profiles from N = 18 patients with irColitis and describe our clinical experience of N = 5 patients treated with healthy donor fecal microbial transplantation (FMT). Recipients of ICI between May 2019 and June 2021 at Memorial Sloan Kettering Cancer Center whose stool samples were investigated for diarrhea were included in our dataset (N = 100). N = 18 patients with a diagnosis of irColitis were identified. Patients that had other causes of diarrhea were classified as part of the “no-colitis” comparator group (N = 32). All patients with infectious causes of diarrhea ( C. difficile, other GI pathogen) were excluded from the analysis. Stool samples were profiled by whole metagenomic shotgun sequencing. FMT products from individual healthy donors were procured from the OpenBiome fecal bank and delivered via colonoscopy. Results: There was no difference in alpha diversity between the irColitis and no-colitis groups. Patients in the irColitis group had significantly higher relative abundance of Proteobacteria at the time of symptom onset compared to the no-colitis group (p = 0.04). Escherichia spp. and Klebsiella spp. were associated with irColitis, also described in multivariate analyses. Five patients with irColitis refractory to steroids and biologics received healthy-donor FMT, with initial clinical improvement in irColitis symptoms observed in four of five (80%) patients. Two out of five subsequently exhibited recurrence of irColitis symptoms following a course of antibiotics for pneumonia and urinary tract infection, respectively. Both received a second “salvage” FMT that was, again, followed by clinical improvement of irColitis. In these two cases, fecal profiles demonstrated dysbiotic shifts in the gut microbiome post-antibiotics characterized by Proteobacteria expansion, which was salvaged post salvage FMT, mirroring clinical resolution of irColitis symptoms. Conclusions: We observed that Proteobacteria expansion is characteristic of irColitis at time of symptom onset. FMT was followed by clinical improvements in several cases of refractory irColitis. Strategies to restore or prevent microbiome injury—with a particular focus on salvage FMT after antibiotic injury—in the context of immunotherapy toxicities could be further explored in prospective clinical trials.

Diet Quality and Fecal Bile Acid Composition

Background: Bile acids (BAs) are steroid metabolites generated by the liver from cholesterol. Their function is to support digestion of dietary fat, cholesterol, and fat-soluble vitamins. The gut microbiota metabolizes BAs that escape intestinal reabsorption to secondary BAs and other metabolites that are linked to increased risk of diseases including colorectal cancer (CRC). Diet plays a major role in BA metabolism and may be an important target for optimizing host-microbial BA metabolism for cancer prevention. Purpose: To examine fecal BA metabolites in relation to diet quality (Healthy Eating Index-2015, HEI-2015) among urban middle to older age adults. Methods: 135 participants were recruited from two academic medical centers. Stool, two 24-hour diet recalls, anthropometrics, and survey data were collected. Fecal BA profiling was completed using LC/MS. Diet quality was calculated using USDA's HEI-2015. Descriptive and inferential statistics were conducted. Results: Participants were 57% female, 51% African/American Black, mean age of 59.6 (±6.2) years and mean BMI of 31.4 (±6.9) kg/m2. Using USDA's cut-point for poor diet quality (HEI-2015 &amp;lt; 51 points), participants with poor diet quality were significantly more likely to be classified with obesity, self-report as African American/Black, report lower educational attainment, have increased secondary BAs, deoxycholic acid (DCA) and lithocholic acid (LCA), and lower 3-oxo-lithocholic acid and iso-allo-deoxycholic acid derivatives. In linear models, HEI-2015 (continuous) was a significant inverse predictor of DCA and LCA when controlling for BMI, race, and education. BMI was a significant positive predictor of DCA and LCA when controlling for HEI-2015, race, education. African American/Black was a significant inverse predictor of 3-oxo-lithocholic acid and iso-allo-deoxycholic acid when controlling for HEI-2015, BMI, and education. Conclusion. Poor diet quality and elevated BMI is associated with increased secondary BAs that have been linked to increased risk of CRC. Improving diet quality and reducing BMI may optimize host-microbial BA metabolism to prevent disease. African American/Black was an independent predicator of lower anti-inflammatory BA derivatives which warrants further investigation.

High-resolution analyses of associations between medications, microbiome, and mortality in cancer patients

Discerning the effect of pharmacological exposures on intestinal bacterial communities in cancer patients is challenging. Here, we deconvoluted the relationship between drug exposures and changes in microbial composition by developing and applying a new computational method, PARADIGM (parameters associated with dynamics of gut microbiota), to a large set of longitudinal fecal microbiome profiles with detailed medication-administration records from patients undergoing allogeneic hematopoietic cell transplantation. We observed that several non-antibiotic drugs, including laxatives, antiemetics, and opioids, are associated with increased Enterococcus relative abundance and decreased alpha diversity. Shotgun metagenomic sequencing further demonstrated subspecies competition, leading to increased dominant-strain genetic convergence during allo-HCT that is significantly associated with antibiotic exposures. We integrated drug-microbiome associations to predict clinical outcomes in two validation cohorts on the basis of drug exposures alone, suggesting that this approach can generate biologically and clinically relevant insights into how pharmacological exposures can perturb or preserve microbiota composition. The application of a computational method called PARADIGM to a large dataset of cancer patients' longitudinal fecal specimens and detailed daily medication records reveals associations between drug exposures and the intestinal microbiota that recapitulate invitro findings and are also predictive of clinical outcomes.

Vitamin A deficiency from maternal gestation may contribute to autistic-like behaviors and gastrointestinal dysfunction in rats through the disrupted purine and tryptophan metabolism

Vitamin A deficiency (VAD) has been linked to autism spectrum disorder (ASD) in multiple studies, and autistic children with gastrointestinal (GI) symptoms have been found to have lower VA levels than those without GI symptoms. However, the exact mechanism by which VAD causes both core symptoms and GI symptoms in ASD is ill defined. We constructed VAD and vitamin A normal (VAN) rat models from maternal gestation onwards. Autism-related behaviors were tested using the open-field test and the three-chamber test, and GI function was assessed with the GI transit time, the colonic transit time and fecal water content. Untargeted metabolomic analysis on the prefrontal cortex (PFC) and fecal samples was performed. VAD rats displayed autistic-like behaviors and impaired GI function compared to VAN rats. Metabolic profiles of both PFC and feces from VAD and VAN rats were significantly different. The differential metabolites in both PFC and feces between the VAN and VAD rats were mostly enriched in the purine metabolic pathway. Moreover, the most significantly affected metabolic pathway in PFC of VAD rats was the phenylalanine, tyrosine and tryptophan biosynthesis pathway, and the most remarkably altered metabolic pathway in the feces of VAD rats was the tryptophan metabolism pathway. These results indicate that VAD starting from maternal gestation might be linked to core symptoms of ASD and its GI co-occurring disorders through the purine and tryptophan-related metabolism disorders.

Plant cell wall composition modulates the gut microbiota and metabolites in in-vitro fermentation

This research investigated the effect of different types of plant cell wall fibres, including cereal (i.e., barley, sorghum, and rice), legume (i.e., pea, faba bean, and mung bean), and tuber (potato, sweet potato, and yam) cell wall fibres on in vitro faecal fermentation profiles and gut microbiota composition. The cell wall composition, specifically the content of lignin and pectin, was found to have a significant influence on the gut microbiota and fermentation outcomes. Compared with type I cell walls (legume and tuber) which have high pectin content, the type II cell walls (cereal) which are high in lignin but low in pectin had a lower fermentation rates and less short-chain fatty acid production. The redundancy analysis showed samples with similar fibre composition and fermentation profiles clustered together, and the principal coordinate analysis revealed separation among different types of cell walls and closer proximity among the same cell wall types. These findings emphasize the importance of cell wall composition in shaping the microbial community during fermentation and contribute to a better understanding of the relationship between plant cell walls and gut health. This research has practical implications for the development of functional foods and dietary interventions.

A Fecal MicroRNA Signature by Small RNA Sequencing Accurately Distinguishes Colorectal Cancers: Results From a Multicenter Study

Fecal tests currently used for colorectal cancer (CRC) screening show limited accuracy in detecting early tumors or precancerous lesions. In this respect, we comprehensively evaluated stool microRNA (miRNA) profiles as biomarkers for noninvasive CRC diagnosis. A total of 1273 small RNA sequencing experiments were performed in multiple biospecimens. In a cross-sectional study, miRNA profiles were investigated in fecal samples from an Italian and a Czech cohort (155 CRCs, 87 adenomas, 96 other intestinal diseases, 141 colonoscopy-negative controls). A predictive miRNA signature for cancer detection was defined by a machine learning strategy and tested in additional fecal samples from 141 CRC patients and 80 healthy volunteers. miRNA profiles were compared with those of 132 tumors/adenomas paired with adjacent mucosa, 210 plasma extracellular vesicle samples, and 185 fecal immunochemical test leftover samples. Twenty-five miRNAs showed altered levels in the stool of CRC patients in both cohorts (adjusted P < .05). A 5-miRNA signature, including miR-149-3p, miR-607-5p, miR-1246, miR-4488, and miR-6777-5p, distinguished patients from control individuals (area under the curve [AUC], 0.86; 95% confidence interval [CI], 0.79-0.94) and was validated in an independent cohort (AUC, 0.96; 95% CI, 0.92-1.00). The signature classified control individuals from patients with low-/high-stage tumors and advanced adenomas (AUC, 0.82; 95% CI, 0.71-0.97). Tissue miRNA profiles mirrored those of stool samples, and fecal profiles of different gastrointestinal diseases highlighted miRNAs specifically dysregulated in CRC. miRNA profiles in fecal immunochemical test leftover samples showed good correlation with those of stool collected in preservative buffer, and their alterations could be detected in adenoma or CRC patients. Our comprehensive fecal miRNome analysis identified a signature accurately discriminating cancer aimed at improving noninvasive diagnosis and screening strategies.

Fecal Microbial Structure and Metabolic Profile in Post-Weaning Diarrheic Piglets.

(1) Background: Piglet diarrhea is one of the most serious diseases in pigs and has brought great economic losses to the pig industry. Alteration of the gut microbiota is an important factor in the etiology of piglet diarrhea. Therefore, this study aimed to analyze the differences in the gut microbial structures and fecal metabolic profile between post-weaning diarrhea and healthy Chinese Wannan Black pigs. (2) Methods: An integrated approach of 16S rRNA gene sequencing combined with LC/MS-based metabolomics was employed in this study. (3) Results: We found an increase in the relative abundance of the bacterial genus Campylobacter and a decrease in phylum Bacteroidetes and the species Streptococcus gallolyticus subsp. macedonicus. (S. macedonicus) in piglet diarrhea. Meanwhile, obvious changes in the fecal metabolic profile of diarrheic piglets were also detected, particularly higher levels of polyamines (spermine and spermidine). Moreover, there were substantial associations between the disturbed gut microbiota and the altered fecal metabolites, especially a strong positive relationship between spermidine and Campylobacter. (4) Conclusions: These observations may provide novel insights into potential etiologies related to post-weaning diarrhea and further enhance our understanding of the role of gut microbiota in host homeostasis and in modulating gut microbial structure.

Effects of Ganoderma lucidum polysaccharide peptide ameliorating cyclophosphamide-induced immune dysfunctions based on metabolomics analysis.

Ganoderma lucidum polysaccharide peptide (GLPP) is one of the most abundant constituents of Ganoderma lucidum (G. lucidum), with a wide range of functional activities. The present study investigated the immunomodulatory effects of GLPP in cyclophosphamide (CTX)-induced immunosuppressive mice. The results showed that 100 mg/kg/day of GLPP administration significantly alleviated CTX-induced immune damage by improving immune organ indexes, earlap swelling rate, the index of carbon phagocytosis and clearance value, secretion of cytokines (TNF-α, IFN-γ, and IL-2), and immunoglobulin A(IgA) in the mice. Furthermore, ultra-performance liquid chromatography with mass/mass spectrometry (UPLC-MS/MS) was conducted to identify the metabolites, followed by biomarker and pathway analysis. The results showed that GLPP treatment alleviated CTX-induced alterations in the fecal metabolome profile, including arachidonic acid (AA), leukotriene D4 (LTD4), indole-3-ethanol, and formyltetrahydrofolate (CF), by reversing citric acid, malic acid, cortisol, and oleic acid. These results support the concept that GLPP exhibits immunomodulatory activity via the folate cycle, methionine cycle, TCA cycle, fatty acid biosynthesis and metabolism, glycerophospholipid metabolism, AA metabolism, and cAMP pathways. In conclusion, the results could be helpful to understand the use of GLPP to clarify the immunomodulatory mechanism and be used as immunostimulants to prevent CTX-induced side effects in the immune system.

Alterations in the Gut Microbiome and Metabolisms in Pregnancies with Fetal Growth Restriction.

Fetuses diagnosed with fetal growth restriction (FGR) are at an elevated risk of stillbirth and adulthood morbidity. Gut dysbiosis has emerged as one of the impacts of placental insufficiency, which is the main cause of FGR. This study aimed to characterize the relationships among the intestinal microbiome, metabolites, and FGR. Characterization was conducted on the gut microbiome, fecal metabolome, and human phenotypes in a cohort of 35 patients with FGR and 35 normal pregnancies (NP). The serum metabolome was analyzed in 19 patients with FGR and 31 normal pregnant women. Multidimensional data was integrated to reveal the links between data sets. A fecal microbiota transplantation mouse model was used to determine the effects of the intestinal microbiome on fetal growth and placental phenotypes. The diversity and composition of the gut microbiota were altered in patients with FGR. A group of microbial species altered in FGR closely correlated with fetal measurements and maternal clinical variables. Fecal and serum metabolism profiles were distinct in FGR patients compared to those in the NP group. Altered metabolites were identified and associated with clinical phenotypes. Integrated multi-omics analysis revealed the interactions among gut microbiota, metabolites, and clinical measurements. Microbiota from FGR gravida transplanted to mice progestationally induced FGR and placental dysfunction, including impaired spiral artery remodeling and insufficient trophoblast cell invasion. Taken together, the integration of microbiome and metabolite profiles from the human cohort indicates that patients with FGR endure gut dysbiosis and metabolic disorders, which contribute to disease pathogenesis. IMPORTANCE Downstream of the primary cause of fetal growth restriction are placental insufficiency and fetal malnutrition. Gut microbiota and metabolites appear to play an important role in the progression of gestation, while dysbiosis induces maternal and fetal complications. Our study elaborates the significant differences in microbiota profiles and metabolome characteristics between women with FGR and normal pregnancies. This is the first attempt so far that reveals the mechanistic links in multi-omics in FGR, providing a novel insight into host-microbe interaction in placenta-derived diseases.

Gut microbiome combined with metabolomics reveals biomarkers and pathways in central precocious puberty

BackgroundCentral precocious puberty (CPP) is a common disease in prepubertal children and results mainly from disorders in the endocrine system. Emerging evidence has highlighted the involvement of gut microbes in hormone secretion, but their roles and downstream metabolic pathways in CPP remain unknown.MethodsTo explore the gut microbes and metabolism alterations in CPP, we performed the 16S rRNA sequencing and untargeted metabolomics profiling for 91 CPP patients and 59 healthy controls. Bioinformatics and statistical analyses, including the comparisons of alpha and beta diversity, abundances of microbes, were undertaken on the 16S rRNA gene sequences and metabolism profiling. Classifiers were constructed based on the microorganisms and metabolites. Functional and pathway enrichment analyses were performed for identification of the altered microorganisms and metabolites in CPP.ResultsWe integrated a multi-omics approach to investigate the alterations and functional characteristics of gut microbes and metabolites in CPP patients. The fecal microbiome profiles and fecal and blood metabolite profiles for 91 CPP patients and 59 healthy controls were generated and compared. We identified the altered microorganisms and metabolites during the development of CPP and constructed a machine learning-based classifier for distinguishing CPP. The Area Under Curves (AUCs) of the classifies were ranged from 0.832 to 1.00. In addition, functional analysis of the gut microbiota revealed that the nitric oxide synthesis was closely associated with the progression of CPP. Finally, we investigated the metabolic potential of gut microbes and discovered the genus Streptococcus could be a candidate molecular marker for CPP treatment.ConclusionsOverall, we utilized multi-omics data from microorganisms and metabolites to build a classifier for discriminating CPP patients from the common populations and recognized potential therapeutic molecular markers for CPP through comprehensive analyses.

Effect of short-term consumption of yellow peas as noodles on the intestinal environment: A single-armed pre-post comparative pilot study.

Legumes contain dietary fiber and resistant starch, which are beneficial to the intestinal environment. Here, we investigated the effects of yellow pea noodle consumption on the gut microbiota and fecal metabolome of healthy individuals. This single-armed pre-post comparative pilot study evaluated eight healthy female participants who consumed yellow pea noodles for 4 weeks. The gut microbiota composition and fecal metabolomic profile of each participant were evaluated before (2 weeks), during (4 weeks), and after (4 weeks) daily yellow pea noodle consumption. 16S rRNA gene sequencing was performed on stool samples, followed by clustering of operational taxonomic units using the Cluster Database at High Identity with Tolerance and integrated QIIME pipeline to elucidate the gut microbiota composition. The fecal metabolites were analyzed using capillary electrophoresis time-of-flight mass spectrometry and liquid chromatography time-of-flight mass spectrometry. Compared to day 0, the relative abundances of five bacterial genera (Bacteroides, Bilophila, Hungatella, Parabacteroides, and Streptococcus) in the intestinal microbiota significantly decreased, wherein those of Bifidobacterium longum and Ruminococcus bromii were increased on day 29 and decreased to the basal level (day 0) on day 57. Fecal metabolomic analysis identified 11 compounds showing significant fluctuations in participants on day 29 compared to day 0. Although the average levels of short-chain fatty acids in participants did not differ significantly on day 29 compared to those on day 0, the levels tended to increase in individual participants with >8% relative abundance of R. bromii in their gut microbiota. In conclusion, incorporating yellow peas as a daily staple may confer human health benefits by favorably altering the intestinal environment.

18 Fecal microbial metabolite profiles are unaltered by laminitis history

Microbial metabolites of the equine gastrointestinal tract may confer information about health and susceptibility to disease. Laminitis is often associated with metabolic disturbances, but little is known of its impact on functional microbial metabolism. A pilot study compared metabolomic profiles of feces collected from 55 native ponies, comprising n = 15 pairs of previously laminitic (PL) and non-laminitic (NL) ponies matched for location, diet, and management, and n = 25 ponies suspected of having an active episode of laminitis (SL) but without clinical diagnosis. Targeted liquid chromatography tandem mass spectrometry (LC-MS/MS) analysis was used. Univariate analyses (ANOVA with Fisher's least significant difference comparisons of PL, NL and SL groups) was performed with significance set at P < 0.05 following Benjamini-Hochberg correction. Principal component analysis (PCA) wasapplied to visualize clustering of individuals. Analysis was performed using the MetaboAnalyst 5.0 web-based interface. Of a panel of 167 metabolites, 104 were identified in the feces, however, none were significantly different between the laminitis status groups. Assessment of the PCA scores plot revealed no clustering of individuals by metabolic profile based on laminitis status. The similar fecal metabolite profile of previously laminitic and non-laminitic ponies indicates that laminitis may not result in significant long-term, alteration of microbial metabolism. Future work to combine these data with that of microbial abundance and species diversity in the feces may reveal further insights into the functional microbial metabolism in laminitic and healthy animals.

Fecal metabolomics analysis for deciphering the lipid-lowering effect of Qizhi capsule on high-fat feed induced hyperlipidemia.

Qizhi capsule (QZC), a Chinese patent drug, has been utilized to treat hyperlipidemia. The present study aims to investigate the lipid-lowering effect of QZC, as well as the mechanism of action for treating hyperlipidemia. High-fat diet (HFD) induced hyperlipidemia rats were administrated with different doses of QZC for 28 days, and atorvastatin calcium tablets was used as the positive control. Serum total cholesterol (TC), triglycerides (TG), low-density lipoprotein cholesterol (LDL-C), and high-density lipoprotein cholesterol (HDL-C) levels were used to evaluate the effectiveness of QZC treatment. The metabolic profiles of feces were analyzed by UPLC-MS-based metabolomics approach coupled with multivariate data analysis. The levels of serum TC, TG, LDL-C, and HDL-C were significantly reversed in QZC treatment groups, showing a similar or even better treatment effect compared with the atorvastatin calcium group. Thirty-two potential fecal biomarkers related to hyperlipidemia were identified. QZC could partially recover the disturbed metabolic pathways of alpha-linolenic acid metabolism, sphingolipid metabolism, glycerophospholipid metabolism, and glycosylphosphatidylinositol (GPI)-anchor biosynthesis. Meanwhile, the signal pathways of regulation of lipid metabolism by peroxisome proliferator-activated receptor α (PPARα), PPARα activates gene expression, and transcriptional regulation of white adipocyte differentiation can be also regulated by QZC. The lipid-lowering effect of QZC was confirmed by both serum biochemistry and metabolomics analysis. The beneficial effects of QZC were mainly attributed to the correction of metabolic disorders and the maintenance of the dynamic balance of metabolites.

Characterizing the metabolomic signature of attention-deficit hyperactivity disorder in twins

Emerging evidence implicate the gut microbiota as a potential susceptibility factor in attention-deficit hyperactivity disorder (ADHD), a common multifactorial neurodevelopmental condition. However, little is known about the biochemical signature of ADHD, including the metabolic contribution of the microbiota via the gut-brain axis, and the relative contribution of genetics and environmental factors. Here, we perform unbiased metabolomic profiling of urine and fecal samples collected from a well-characterized Swedish twin cohort enriched for ADHD (33 ADHD, 79 non-ADHD), using 1H nuclear magnetic resonance spectroscopy and liquid chromatography-mass spectrometry. Our results highlight sex-specific patterns in the metabolic phenotype of individuals with ADHD. Specifically, the urine profile of males, but not females, with ADHD was characterized by greater excretion of hippurate, a product of microbial-host co-metabolism that can cross the blood-brain-barrier with bioactivity of potential relevance to ADHD. This trans-genomic metabolite was also negatively correlated with IQ in males and was significantly correlated with fecal metabolites associated with gut microbial metabolism. The fecal profile of ADHD individuals was characterized by increased excretion of stearoyl-linoleoyl-glycerol, 3,7-dimethylurate, and FAD and lower amounts of glycerol 3-phosphate, thymine, 2(1H)-quinolinone, aspartate, xanthine, hypoxanthine, and orotate. These changes were independent of ADHD medication, age, and BMI. Furthermore, our specific twins’ models revealed that many of these gut metabolites had a stronger genetic influence than environmental. These findings suggest that metabolic disturbances in ADHD, involving combined gut microbial and host metabolic processes, may largely derive from gene variants previously linked to behavioral symptoms in this disorder.This article is part of the Special Issue on “Microbiome & the Brain: Mechanisms & Maladies”.

From adenoma to CRC stages: the oral-gut microbiome axis as a source of potential microbial and metabolic biomarkers of malignancy

BackgroundApproximately 95% of Colorectal cancers (CRC) consist of adenocarcinomas originating from colonic Adenomatous polyps (AP). Increasing importance in CRC occurrence and progression has been attributed to the gut microbiota; however, a huge proportion of microorganisms inhabit the human digestive system. So, to comprehensively study the microbial spatial variations and their role in CRC progression, from AP to the different CRC phases, a holistic vision is imperative, including the simultaneous evaluation of multiple niches from the gastrointestinal system. Through an integrated approach, we identified potential microbial and metabolic biomarkers, able to discriminate human CRC from AP and/or also the different Tumor node metastasis (TNM) staging. In addition, as the microbiota contributes to the production of essential metabolic products detectable in fecal samples, we analysed and compared metabolites obtained from CRC and AP patients by using a Nuclear magnetic resonance (NMR) approach. MethodsIn this observational study, saliva, tissue and stool samples from 61 patients, have been collected, including 46 CRC and 15 AP patients, age and sex-matched, undergoing surgery in 2018 at the Careggi University Hospital (Florence, Italy). First, the microbiota in the three-district between CRC and AP patients has been characterized, as well as in different CRC TNM stages. Subsequently, proton NMR spectroscopy has been used in combination with multivariate and univariate statistical approaches, to define the fecal metabolic profile of a restricted group of CRC and AP patients. ResultsCRC patients display a different profile of tissue and fecal microbiota with respect to AP patients. Significant differences have been observed in CRC tissue microbial clades, with a rise of the Fusobacterium genus. In addition, significant taxa increase at the genus level has been observed in stool samples of CRC patients. Furthermore, Fusobacterium found in intestinal tissue has been positively correlated with fecal Parvimonas, for the first time. Moreover, as predicted by metagenomics pathway analysis, a significant increase of lactate (p=0.037) has been observed in the CRC fecal metabolic profiles, and positively correlated with Bifidobacterium (p=0.036). Finally, minor bacterial differences in CRC patients at stage T2 (TNM classification) have been detected, with a raise of the Spirochaetota phylum in CRC samples, with a slight increase of the Alphaproteobacteria class in fecal samples. ConclusionOur results suggest the importance of microbiota communities and oncometabolites in CRC development. Further studies on CRC/AP management with a focus on CRC assessment are needed to investigate novel microbial-related diagnostic tools aimed to improve therapeutic interventions.

Lycium barbarum Oligosaccharides Alleviate Hepatic Steatosis by Modulating Gut Microbiota in C57BL/6J Mice Fed a High-Fat Diet.

High-fat diets (HFD) can promote the development of hepatic steatosis by altering the structure and composition of gut flora. In this study, the potential therapeutic mechanism of Lycium barbarum oligosaccharide (LBO) against hepatic steatosis was investigated by analyzing the changes in the intestinal flora and metabolites in mice. Mice on an HFD were administered LBO by gavage once daily for a continuous period of eight weeks. Compared with the HFD group, the levels of triglyceride (TG), alanine aminotransferase (ALT) in the serum, and hepatic TG were significantly reduced in the LBO group, and liver lipid accumulation was obviously improved. In addition, LBO could regulate the HFD-induced alteration of intestinal flora. The HFD increased the proportion of Barnesiellaceae, Barnesiella, and CHKCI001. LBO increased the proportion of Dubosiella, Eubacterium, and Lactobacillus. LBO also altered the fecal metabolic profile. Significantly different metabolites between LBO and the HFD, such as taurochenodeoxycholate, taurocholate, fluvastatin, and kynurenic acid, were related to the cholesterol metabolism, bile acid metabolism, and tryptophan metabolic pathways. In light of the above, LBO can alleviate HFD-induced NAFLD by modulating the components of the intestinal flora and fecal metabolites.

An Overview on Fecal Profiles of Amino Acids and Related Amino-Derived Compounds in Children with Autism Spectrum Disorder in Tunisia

Autism spectrum disorder (ASD) is a neurodevelopmental pathology characterized by the impairment of social interaction, difficulties in communication, and repetitive behaviors. Alterations in the metabolism of amino acids have been reported. We performed a chromatographic analysis of fecal amino acids, ammonium, biogenic amines, and gamma aminobutyric acid (GABA) in Tunisian autistic children from 4 to 10 years, and results were compared with their siblings (SIB) and children from the general population (GP). ASD presented significantly higher levels of fecal amino acids than SIB and GP; differences being more pronounced in younger (4-7 years) than in older (8-10 years) individuals whereas no changes were found for the remaining compounds. Lower levels of histidine were the only difference related with severe symptoms of autism (CARS scale). A linear discriminant analysis (LDA) based on fecal amino acid profiles clearly separated ASD, SIB, and GP at 4 to 7 years but not at more advanced age (8-10 years), evidencing more pronounced alterations in younger children. The relationship of fecal amino acids with autism needs deeper research integrating blood analytical parameters, brain metabolism, and intestinal microbiota. Fecal amino acids could be targeted for designing personalized diets to prevent or minimize cognitive impairments associated with ASD.

Abstract 3797: Fecal miRNA profiles and gut metagenome composition in Lynch syndrome: results from a mouse model study and human subjects

Abstract Bsacground: Lynch Syndrome (LS) is an inherited cancer syndrome associated with an increased lifetime risk of colorectal cancer (CRC) and characterised by germline mutations in DNA mismatch repair (MMR) genes. Abnormalities in the function of MMR genes lead to errors during DNA replication, including microsatellite instability. Despite regular colonoscopies, up to 15% of LS patients still develop CRC. Gut microbiome, bowel inflammation, the environment and host genetic and epigenetic factors are involved in CRC development. In this sense, the concomitant analysis of stool host microRNA (miRNA) profiles and gut microbiome composition may arise to the identification of specific fecal markers in LS explaining the molecular basis for the development of cancer in these patients. PURPOSE: In the present study, the fecal miRNome and microbiome were characterized in a time progression in an in vivo model using a mismatch repair deficient mice model that recapitulates human LS. The main aim was to evaluate the changes and crosstalk of miRNAs released in the stool by the host and the microbial population residing in the gut during the carcinogenesis process induced in animals. Findings in mice were compared and correlated to data obtained from a cohort of human subjects affected by LS. Methods: We used an in vivo model for LS based on a conditional knockout mouse with the tissue-specific inactivation of Msh2 (Msh2LoxP) in the intestinal mucosa, combining Msh2LoxP allele with the Villin-Cre transgene (VCMsh2LoxP). Stool from mice were collected at different time points (6, 9 and 12 months of age). Small RNA-sequencing (sRNA-seq) and shotgun metagenomics analyses were performed in stool from all samples. In concomitance, we collected 78 stool samples of clinically diagnosed LS subjects, at different follow up time. At the sampling, 36 resulted negative for colorectal adenomas/lesions, 12 were negative but with a previous history of lesions, 23 presented a neoplasia and 6 developed a lesion in the follow-up. Twenty-two stool samples from healthy subjects were used as controls. Results: In mice, preliminary results from sRNA-seq and metagenomics analyses showed several dysregulated miRNAs and differential microbial relative abundances, either between VCMsh2loxP and controls or at different time points. In humans, we identified 41 and 17 miRNAs respectively up- and downregulated in LS subjects who presented a lesion compared to those not having any lesion at follow-up. In the same comparison the metagenomic data showed significantly different microbial abundances, including those of Eubacterium ramulus. Conclusions: This is the first study characterizing the concomitant alterations in microbial composition and host miRNome overtime in relation to the onset of cancerous/precancerous lesions due to LS. Citation Format: Giulia Francescato, Giulio Ferrero, Marc Beltrà, Sonia Tarallo, Giulia Piaggeschi, Carla Di Battista, Antonio Francavilla, Carlijn Bruggeling, Fabio Penna, Barbara Pardini, Paola Costelli, Annemarie Boleij, Alessio Naccarati. Fecal miRNA profiles and gut metagenome composition in Lynch syndrome: results from a mouse model study and human subjects. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 3797.

Longitudinal Fecal Calprotectin Profiles Characterize Disease Course Heterogeneity in Crohn’s Disease

The progressive nature of Crohn's disease is highly variable and hard to predict. In addition, symptoms correlate poorly with mucosal inflammation. There is therefore an urgent need to better characterize the heterogeneity of disease trajectories in Crohn's disease by utilizing objective markers of inflammation. We aimed to better understand this heterogeneity by clustering Crohn's disease patients with similar longitudinal fecal calprotectin profiles. We performed a retrospective cohort study at the Edinburgh IBD Unit, a tertiary referral center, and used latent class mixed models to cluster Crohn's disease subjects using fecal calprotectin observed within 5 years of diagnosis. Information criteria, alluvial plots, and cluster trajectories were used to decide the optimal number of clusters. Chi-square test, Fisher's exact test, and analysis of variance were used to test for associations with variables commonly assessed at diagnosis. Our study cohort comprised 356 patients with newly diagnosed Crohn's disease and 2856 fecal calprotectin measurements taken within 5 years of diagnosis (median 7 per subject). Four distinct clusters were identified by characteristic calprotectin profiles: a cluster with consistently high fecal calprotectin and 3 clusters characterized by different downward longitudinal trends. Cluster membership was significantly associated with smoking (P= .015), upper gastrointestinal involvement (P < .001), and early biologic therapy (P < .001). Our analysis demonstrates a novel approach to characterizing the heterogeneity of Crohn's disease by using fecal calprotectin. The group profiles do not simply reflect different treatment regimens and do not mirror classical disease progression endpoints.