Short-chain Fatty Acid Profiles Research Articles

A Multi-Omics Protocol for Swine Feces to Elucidate Longitudinal Dynamics in Microbiome Structure and Function.

Swine are regarded as promising biomedical models, but the dynamics of their gastrointestinal microbiome have been much less investigated than that of humans or mice. The aim of this study was to establish an integrated multi-omics protocol to investigate the fecal microbiome of healthy swine. To this end, a preparation and analysis protocol including integrated sample preparation for meta-omics analyses of deep-frozen feces was developed. Subsequent data integration linked microbiome composition with function, and metabolic activity with protein inventories, i.e., 16S rRNA data and expressed proteins, and identified proteins with corresponding metabolites. 16S rRNA gene amplicon and metaproteomics analyses revealed a fecal microbiome dominated by Prevotellaceae, Lactobacillaceae, Lachnospiraceae, Ruminococcaceae and Clostridiaceae. Similar microbiome compositions in feces and colon, but not ileum samples, were observed, showing that feces can serve as minimal-invasive proxy for porcine colon microbiomes. Longitudinal dynamics in composition, e.g., temporal decreased abundance of Lactobacillaceae and Streptococcaceae during the experiment, were not reflected in microbiome function. Instead, metaproteomics and metabolomics showed a rather stable functional state, as evident from short-chain fatty acids (SCFA) profiles and associated metaproteome functions, pointing towards functional redundancy among microbiome constituents. In conclusion, our pipeline generates congruent data from different omics approaches on the taxonomy and functionality of the intestinal microbiome of swine.

Plasma Short Chain Fatty Acids As a Predictor of Response to Therapy for Life-Threatening Acute Graft-Versus-Host Disease

Introduction Minnesota High Risk and steroid-refractory acute graft-versus-host disease (aGVHD) are life-threatening complications of allogeneic hematopoietic cell transplantation (HCT). Recent data suggests dysbiosis may relate to GVHD outcomes. Bacteria in the lower gastrointestinal tract (LGI) produce short chain fatty acids (SCFAs) as fuel for enterocytes and modulators of mucosal immunity. LGI damage caused by aGVHD and antibiotic administration may be detrimental to SCFAs production and thus intestinal repair. However, limited data exists on how plasma SCFA levels vary between aGVHD patients who respond to treatment and those who do not. This study examined how aGVHD treatment response relates to plasma levels of five common SCFAs: acetate, propionate, butyrate, isovalerate, and valerate. Patients and Methods Serial plasma samples (n=221) were collected from 49 patients who underwent treatment for Minnesota High Risk or steroid-dependent/refractory aGVHD on NCT02525029 at study baseline and at 7, 14, 28 and 56 days post-treatment initiation. GVHD severity was graded using Minnesota criteria. Patients were categorized by their response to therapy at day 28 (complete/partial response [CR/PR], n=35 versus no response or death [NR], n=14). Plasma SCFA levels were quantified via liquid chromatography and cytokines measured by multiplex cytokine array. Changes in SCFA over time were assessed by repeated measures ANOVA. SCFA levels responders versus non-responders at individual time points were compared using Mann-Whitney testing and principal component analysis (PCA). SCFA, cytokines, and GVHD biomarkers including ST2, REG3a, and AREG were correlated using Spearman's rho with Bonferroni correction for multiple comparisons. Results Patients with CR/PR had higher levels of propionate (p=0.02) and butyrate (p=0.008) in comparison to NR among all samples analyzed (Figure 1a). Only butyrate levels varied significantly over time (p=0.018, with a significant difference in CR/PR vs NR at day 7, Figure 1b). The overall metabolomic profile of CR/PR patients was more stable than NR patients as determined by PCA. Neither initial clinical nor histologic grade of LGI aGVHD were associated with baseline plasma SCFA. Recent Clostridium difficile infection (n=5, p=NS) and total parenteral nutrition use (n=26, p=NS) did not influence baseline SCFA concentrations. Patients with high levels of butyrate also had high propionate (rho 0.9, p<0.001) and valerate (rho 0.84, p<0.001) and low isovalerate (rho -0.34, p<0.001). Plasma levels of interleukin-6 were higher in patients with low propionate and valerate (rho -0.33, p=0.008 and rho -0.36, p=0.002 respectively). Discussion Plasma SCFA profiles in patients with life-threatening GVHD suggest that differences in intestinal commensal microbe-derived SCFA production may relate to response to treatment. Patients responding to therapy have higher levels of plasma propionate and butyrate, especially early in the course of therapy. Patients with NR at day 28 appear to have increasing butyrate over time, possibly due to survival bias and/or stabilization on 2nd line therapy. Patients with higher concentrations of propionate and valerate have lower circulating IL-6 levels, which may indicate reduced systemic inflammation. Figure. (A) Comparison of short chain fatty acid (SCFA) plasma concentrations and clinical response (complete/partial response [CR/PR] vs no response [NR]) among all samples and (B) with butyrate over time. (C) Correlation of plasma SCFA, cytokines, and graft-versus-host disease biomarkers. Disclosures MacMillan: Equillium, Inc.: Consultancy; Mesoblast: Consultancy; Talaris Therapeutics, Inc: Consultancy; Fate Therapeutics, Inc.: Consultancy; Angiocrine Biosciences, Inc.: Consultancy. Rashidi:Synthetic Biologics: Other: DSMC member (1 trial) and related honorarium. Weisdorf:Incyte: Research Funding; FATE Therapeutics: Consultancy. Blazar:Tmunity: Other: Co-founder; Fate Therapeutics Inc.: Research Funding; KidsFirst Fund: Research Funding; Childrens' Cancer Research Fund: Research Funding; BlueRock Therapeutics: Research Funding; BlueRock Therapeuetic: Consultancy; Magenta Therapeutics: Consultancy. Betts:Patent Pending: Patents & Royalties: Dr. Betts has a pending patent WO2017058950A1: Methods of treating transplant rejection. This includes the use of JAK inhibitors. Neither he nor his institution have received payment related to claims described in the patent.. Holtan:Incyte: Consultancy; Generon: Consultancy; CSL Behring: Other: Clinical trial data adjudication; BMS: Consultancy.

Univariate Analysis of Short-Chain Fatty Acids Related to Sudden Infant Death Syndrome.

Sudden infant death syndrome (SIDS) is defined as the death of a child under one year of age, during sleep, without apparent cause, after exhaustive investigation, so it is a diagnosis of exclusion. SIDS is the principal cause of death in industrialized countries. Inborn errors of metabolism (IEM) have been related to SIDS. These errors are a group of conditions characterized by the accumulation of toxic substances usually produced by an enzyme defect and there are thousands of them and included are the disorders of the -oxidation cycle, similarly to what can affect the metabolism of different types of fatty acid chain (within these, short chain fatty acids (SCFAs)). In this work, an analysis of postmortem SCFAs profiles of children who died due to SIDS is proposed. Initially, a set of features containing SCFAs information, obtained from the NIH Common Fund’s National Metabolomics Data Repository (NMDR) is submitted to an univariate analysis, developing a model based on the relationship between each feature and the binary output (death due to SIDS or not), obtaining 11 univariate models. Then, each model is validated, calculating their receiver operating characteristic curve (ROC curve) and area under the ROC curve (AUC) value. For those features whose models presented an AUC value higher than 0.650, a new multivariate model is constructed, in order to validate its behavior in comparison to the univariate models. In addition, a comparison between this multivariate model and a model developed based on the whole set of features is finally performed. From the results, it can be observed that each SCFA which comprises of the SFCAs profile, has a relationship with SIDS and could help in risk identification.

Microbial signature in IgE-mediated food allergies

BackgroundMultiple studies suggest a key role for gut microbiota in IgE-mediated food allergy (FA) development, but to date, none has studied it in the persistent state.MethodsTo characterize the gut microbiota composition and short-chain fatty acid (SCFAs) profiles associated with major food allergy groups, we recruited 233 patients with FA including milk (N = 66), sesame (N = 38), peanut (N = 71), and tree nuts (N = 58), and non-allergic controls (N = 58). DNA was isolated from fecal samples, and 16S rRNA gene sequences were analyzed. SCFAs in stool were analyzed from patients with a single allergy (N = 84) and controls (N = 31).ResultsThe gut microbiota composition of allergic patients was significantly different compared to age-matched controls both in α-diversity and β-diversity. Distinct microbial signatures were noted for FA to different foods. Prevotella copri (P. copri) was the most overrepresented species in non-allergic controls. SCFAs levels were significantly higher in the non-allergic compared to the FA groups, whereas P. copri significantly correlated with all three SCFAs. We used these microbial differences to distinguish between FA patients and non-allergic healthy controls with an area under the curve of 0.90, and for the classification of FA patients according to their FA types using a supervised learning algorithm. Bacteroides and P. copri were identified as taxa potentially contributing to KEGG acetate-related pathways enriched in non-allergic compared to FA. In addition, overall pathway dissimilarities were found among different FAs.ConclusionsOur results demonstrate a link between IgE-mediated FA and the composition and metabolic activity of the gut microbiota.

Prebiotic Xylo-Oligosaccharides Ameliorate High-Fat-Diet-Induced Hepatic Steatosis in Rats.

Understanding the importance of the gut microbiota (GM) in non-alcoholic fatty liver disease (NAFLD) has raised the hope for therapeutic microbes. We have shown that high hepatic fat content associated with low abundance of Faecalibacterium prausnitzii in humans and, further, the administration of F. prausnitzii prevented NAFLD in mice. Here, we aimed at targeting F. prausnitzii by prebiotic xylo-oligosaccharides (XOS) to treat NAFLD. First, the effect of XOS on F. prausnitzii growth was assessed in vitro. Then, XOS was supplemented or not with high (HFD, 60% of energy from fat) or low (LFD) fat diet for 12 weeks in Wistar rats (n = 10/group). XOS increased F. prausnitzii growth, having only a minor impact on the GM composition. When supplemented with HFD, XOS ameliorated hepatic steatosis. The underlying mechanisms involved enhanced hepatic β-oxidation and mitochondrial respiration. Nuclear magnetic resonance (1H-NMR) analysis of cecal metabolites showed that, compared to the HFD, the LFD group had a healthier cecal short-chain fatty acid profile and on the HFD, XOS reduced cecal isovalerate and tyrosine, metabolites previously linked to NAFLD. Cecal branched-chain fatty acids associated positively and butyrate negatively with hepatic triglycerides. In conclusion, XOS supplementation can ameliorate NAFLD by improving hepatic oxidative metabolism and affecting GM.

Association of Moderate Beer Consumption with the Gut Microbiota and SCFA of Healthy Adults.

Fermented alcoholic drinks’ contribution to the gut microbiota composition is mostly unknown. However, intestinal microorganisms can use compounds present in beer. This work explored the associations between moderate consumption of beer, microbiota composition, and short chain fatty acid (SCFA) profile. Seventy eight subjects were selected from a 261 healthy adult cohort on the basis of their alcohol consumption pattern. Two groups were compared: (1) abstainers or occasional consumption (ABS) (n = 44; <1.5 alcohol g/day), and (2) beer consumption ≥70% of total alcohol (BEER) (n = 34; 200 to 600 mL 5% vol. beer/day; <15 mL 13% vol. wine/day; <15 mL 40% vol. spirits/day). Gut microbiota composition (16S rRNA gene sequencing) and SCFA concentration were analyzed in fecal samples. No differences were found in α and β diversity between groups. The relative abundance of gut bacteria showed that Clostridiaceae was lower (p = 0.009), while Blautia and Pseudobutyrivibrio were higher (p = 0.044 and p = 0.037, respectively) in BEER versus ABS. In addition, Alkaliphilus, in men, showed lower abundance in BEER than in ABS (p = 0.025). Butyric acid was higher in BEER than in ABS (p = 0.032), and correlated with Pseudobutyrivibrio abundance. In conclusion, the changes observed in a few taxa, and the higher butyric acid concentration in consumers versus non-consumers of beer, suggest a potentially beneficial effect of moderate beer consumption on intestinal health.

Animal, feed and rumen fermentation attributes associated with methane emissions from sheep fed brassica crops.

Methane emissions from ruminants enhance global warming and lead to a loss of feed energy. The emissions are low when fed brassica crops, but the factors contributing to low emissions are unknown. A meta-analysis was conducted with individual animal data collected from seven experiments. In these experiments, methane emissions were measured using respiration chambers. Animal characteristics, feed chemical composition and rumen fermentation parameters were included for the analysis using multiple regression models. Feed intake level, animal live weight and age were animal factors that were weakly and negatively related to methane yield (g/dry matter intake). The duration in which sheep were fed brassica crops was a significant contributor in the model, suggesting that the effect on emissions diminishes with time. Among a range of feed chemical composition characters, acid detergent fibre and hot-water-soluble carbohydrate contributed significantly to the model, suggesting that both structural and soluble carbohydrates affect methane formation in the rumen. There was no significant correlation between the concentration of sulphate in brassicas and emissions, but nitrate was moderately and negatively correlated with methane yield (r=-.53). Short-chain fatty acid profiles in the rumen of animals fed brassicas were different from those fed pasture, but these parameters only moderately correlated to methane emissions (r=.42). Feeding forage rape resulted in low rumen pH. The pH before morning feeding was strongly correlated to methane yield (r=.90). Rumen pH, together with microbial communities mediated by pH, might lead to low emissions. Bacteria known to produce hydrogen were relatively less abundant in the rumen contents of brassica-fed animals than pasture-fed animals. In conclusion, animal and feed factors, rumen fermentation and microbial communities all affect methane emissions to some extent. The interactions of these factors with each other thus contribute to methane emissions from brassica-fed sheep.

Evaluation of the Levels of Metabolites in Feces of Patients with Inflammatory Bowel Diseases

Inflammatory bowel diseases (IBD), which include ulcerative colitis (UC) and Crohn’s disease (CD), are chronic intestinal inflammatory disorders with only partially known etiology. They are characterized by chronic recurrent inflammation of the intestinal mucosa and lead to a significant decrease in the quality of life and death of patients. IBD is associated with suppression of normal intestinal microflora, particularly, a decrease in the number of bacteria—producers of short chain fatty acids (SCFAs) that exhibit anti-inflammatory and protective properties. Among various methods used for correction of intestinal microflora, fecal microbiota transplantation (FMT) attracts particular attention. FMT consists of the infusion of a fecal suspension from a healthy donor to the intestinal tract of a recipient patient. Such treatment is usually accompanied by a positive therapeutic effect with the restoration of the normal intestinal microflora of the recipient patient. A significant drawback of this method is the lack of standardization. Evaluation of metabolites produced by intestinal microflora, particularly, SCFAs, may be used for objective assessment of the functional state of the intestinal microbiota and, consequently, the success of the FMT procedure. Using gas chromatography and nuclear magnetic resonance spectroscopy, we have analyzed the concentrations and molar ratios of SCFAs from feces of 60 healthy donors. The results obtained by these two methods were in good accord with each other and with the published data. The analysis of SCFAs in the feces of patients with UC (19 patients) and CD (17 patients) revealed a general decrease in the concentration of SCFAs in the studied groups as compared with controls and significant fluctuations in the values of the studied group. Using limited sets of IBD patients (6 patients with UC and 5 patients with CD) we determined concentrations of SCFAs before and within 30 days of observation after FMT. In the selected time period, FMT exerts an effect on the SCFAs levels. Despite the fact that within 30 days of observation the SCFAs profiles, specifically attributed to healthy donors, have not been fully restored, the tendency to their achievement was clearly observed in both sets of UC and CD patients.

Daily Fermented Whey Consumption Alters the Fecal Short-Chain Fatty Acid Profile in Healthy Adults.

Gut microbiota influences many aspects of host health including immune, metabolic, and gut health. We examined the effect of a fermented whey concentrate (FWC) drink rich in L-(+)-Lactic acid, consumed daily, in 18 healthy men (n = 5) and women (n = 13) in free-living conditions.Objective: The aims of this 6-weeks pilot trial were to (i) identify changes in the gut microbiota composition and fecal short chain fatty acid (SCFA) profile, and (ii) to monitor changes in glucose homeostasis.Results: Total fecal SCFA (mM) concentration remained constant throughout the intervention. Proportionally, there was a significant change in the composition of different SCFAs compared to baseline. Acetate levels were significantly reduced (−6.5%; p < 0.01), coupled to a significant increase in the relative amounts of propionate (+2.2%; p < 0.01) and butyrate (+4.2%; p < 0.01), respectively. No changes in the relative abundance of any specific bacteria were detected. No significant changes were observed in glucose homeostasis in response to an oral glucose tolerance test.Conclusion: Daily consumption of a fermented whey product led to significant changes in fecal SCFA metabolite profile, indicating some potential prebiotic activity. These changes did not result in any detectable differences in microbiota composition. Post-hoc analysis indicated that baseline microbiota composition might be indicative of participants likely to see changes in SCFA levels. However, due to the lack of a control group these findings would need to be verified in a rigorously controlled trial. Future work is also required to identify the biological mechanisms underlying the observed changes in microbiota activity and to explore if these processes can be harnessed to favorably impact host health.Clinical Trial Registration: www.clinicaltrials.gov, identifier NCT03615339; retrospectively registered on 03/08/2018.

In search for interplay between stool microRNAs, microbiota and short chain fatty acids in Crohn\u2019s disease - a preliminary study

BackgroundInflammatory bowel diseases are classic polygenic disorders, with genetic loads that reflect immunopathological processes in response to the intestinal microbiota. Herein we performed the multiomics analysis by combining the large scale surveys of gut bacterial community, stool microRNA (miRNA) and short chain fatty acid (SCFA) signatures to correlate their association with the activity of Crohn’s disease (CD).MethodsDNA, miRNA, and metabolites were extracted from stool samples of 15 CD patients, eight with active disease and seven in remission, and nine healthy individuals. Microbial, miRNA and SCFA profiles were assessed using datasets from 16S rRNA sequencing, Nanostring miRNA and GC-MS targeted analysis, respectively.ResultsPairwise comparisons showed that 9 and 23 taxa differed between controls and CD patients with active and inactive disease, respectively. Six taxa were common to both comparisons, whereas four taxa differed in CD patients. α-Diversity was lower in both CD groups than in controls. The levels of 13 miRNAs differed (p-value < 0.05; FC > 1.5) in CD patients and controls before FDR correction and 4 after. Of six SCFAs, the levels of two differed significantly (p-value < 0.05, FC > 1.5) in CD patients and controls, and the levels of four differed in patients with active and inactive CD. PLS-DA revealed models with smallest error rate for controls in bacterial component and inactive disease in metabolites.ConclusionA complex interrelationship may exist between gut dysbiosis, miRNA profiling and SCFA level in response to intestinal inflammation.

Metabolite profiling reveals the interaction of chitin-glucan with the gut microbiota

ABSTRACT Dietary fibers are considered beneficial nutrients for health. Current data suggest that their interaction with the gut microbiota largely contributes to their physiological effects. In this context, chitin-glucan (CG) improves metabolic disorders associated with obesity in mice, but its effect on gut microbiota has never been evaluated in humans. This study explores the effect of a 3-week intervention with CG supplementation in healthy individuals on gut microbiota composition and bacterial metabolites. CG was given to healthy volunteers (n = 15) for three weeks as a supplement (4.5 g/day). Food diary, visual analog and Bristol stool form scales and a “quality of life” survey were analyzed. Among gut microbiota-derived metabolites, bile acids (BA), long- and short-chain fatty acids (LCFA, SCFA) profiling were assessed in stool samples. The gut microbiota (primary outcome) was analyzed by Illumina sequencing. A 3-week supplementation with CG is well tolerated in healthy humans. CG induces specific changes in the gut microbiota composition, with Eubacterium, Dorea and Roseburia genera showing the strongest regulation. In addition, CG increased bacterial metabolites in feces including butyric, iso-valeric, caproic and vaccenic acids. No major changes were observed for the fecal BA profile following CG intervention. In summary, our work reveals new potential bacterial genera and gut microbiota-derived metabolites characterizing the interaction between an insoluble dietary fiber -CG- and the gut microbiota.

Effect of Alliaceae Extract Supplementation on Performance and Intestinal Microbiota of Growing-Finishing Pig

Simple SummaryThe increasing interest in phytogenics for use with livestock, especially swine and poultry, is mainly due to their antimicrobial, antioxidant, growth-promoting, and gut microbiome modulation properties, which makes them ideal candidates to mitigate the negative effects of the ban on antibiotic growth promoters in the European Union. We tested the ability of Allium spp. extract (containing garlic and onion), one of the best-known phytogenics, used in pig feed, to improve growth performance through modulation of the microbiome and changes in the metabolism of short-chain fatty acids in the gut tract. The promising results obtained in the present study suggested that Allium spp. extracts had the potential to be used in feeding pigs to improve growth performances by modulating the microbiota and metabolism of short-chain fatty acids.The aim of the present study was to ascertain whether an Allium spp. extract rich in organosulfur compounds, such as propyl thiosulfonate (PTSO), added to the feed of growing-finishing pigs at 5 g/kg enhances growth performance or affects the fecal microbiome, the levels of short-chain fatty acids, or the antioxidant capacity of the animals. Fifty male growing pigs (large white) of 23.07 ± 2.87 kg average body weight were randomly allotted to two treatments in a 103-day trial. The trial was divided into two periods, an initial growing phase (56-days) and a finishing phase (47-days). Two dietary treatments for each phase (growing and finishing) were used: a control diet (CON) and an experimental diet consisting of the control diet to which 5 g/kg of Allium spp. extract was added to substitute sepiolite (GAR). Throughout the study, body weight, average daily gain (kg/day, ADG), feed intake (kg/day), and feed conversion ratio (kg/kg) were measured, while the backfat thickness and muscle depth were determined at the end of the study. Besides, feces samples were taken for bacterial counts by means of real-time PCR and short-chain fatty acid (SCFA) profile determination, and the antioxidant capacity was assessed in serum and saliva. In the animals receiving Allium spp. extract (5 g/kg) in the feed, ADG increased (p < 0.05) throughout the trial, Salmonella spp. and Clostridium spp. counts in feces had decreased (p < 0.05) when measured on day 56, and, by day 103, Salmonella spp., Clostridium spp., and Enterobacteriaceae counts had decreased (p < 0.05) and Lactobacillus spp. counts had increased (p < 0.01) in feces. Regarding the SCFA profile in feces and antioxidant capacity measured in serum and saliva, supplementation with Allium spp. extract significantly increased the levels of propionic, isobutyric, and isovaleric acids and the percentage of total branched fatty acids, while the c2/c3 and (c2 + c4)/c3 ratios were lower (p < 0.05) in feces; the Trolox equivalent antioxidant capacity and the cupric reducing antioxidant capacity levels in serum were significantly higher in the same pigs on day 103 than on day 0. Consequently, based on the current results, Allium spp. extract rich in organosulfur compounds, added to the diet at 5 g/kg, had a beneficial effect on the microbiota and would seem to be a possible alternative for increasing the growth performance of growing-finishing pigs. However, further studies on the effects of Allium spp. supplementation on carcass quality are necessary.

Rumen Microbiome and Metabolome of Tibetan Sheep (Ovis aries) Reflect Animal Age and Nutritional Requirement.

The rumen microbiota plays an important role in animal functional attributes. These microbes are indispensable for the normal physiological development of the rumen, and may also convert the plant polysaccharides from grass into available milk and meat, making it highly valuable to humans. Exploring the microbial composition and metabolites of rumen across developmental stages is important for understanding ruminant nutrition and metabolism. However, relatively few reports have investigated the microbiome and metabolites across developmental stages in ruminants. Using 16S rRNA gene sequnecing, metabolomics and high-performance liquid chromatography techniques, we compared the rumen microbiota, metabolites and short chain fatty acids (SCFAs) between lambs and sub-adult Tibetan sheep (Ovis aries) from Qinghai-Tibetan Plateau. Bacteroidetes and Spirochaetae were enriched in sub-adult sheep, while Firmicutes and Tenericutes were more abundant in young individuals. The sub-adult individuals had higher alpha diversity values than those in young sheep. Metabolomics analysis showed that the content of essential amino acids and related gene functional pathways in rumen were different between the lambs and sub-adult population. L-Leucine that participates in valine, leucine and isoleucine biosynthesis was more abundant in the lambs, while phenylethylamine that takes part in phenylalanine metabolism was more enriched in the sub-adults. Both rumen microbial community structures and metabolite profiles were impacted by age, but rumen SCFA concentration was relatively stable between different age stages. Some specific microbes (e.g., Clostridium and Ruminococcaceae) were positively associated with L-Leucine but negatively correlated with phenylethylamine, implying that rumen microbes may play different roles for metabolite production at different ages. Mantel test analysis showed that rumen microbiota was significantly correlated with metabolomics and SCFA profiles. Our results indicates the close relationship between microbial composition and metabolites, and also reveal different nutritional requirement for different ages in ruminants, thus having important significance for regulating animal nutrition and metabolism by microbiome intervention.

A Marine-Derived, Multi-mineral Supplement Influences Bacterial Fermentation and Short Chain Fatty Acid Profile In Vitro.

Aquamin is a calcium-rich multi-mineral supplement derived from the red marine algae, Lithothamnion species. Calcium supplementation has been shown to exert a prebiotic-like effect on the gut microbiota and has been associated with distinct changes in lactate and short chain fatty acid (SCFA) production. Irritable bowel syndrome (IBS) subtype is associated with changes in SCFA levels compared with healthy controls. Using an ex vivo simulation model, and a fecal inoculum from a patient diagnosed with IBS, we evaluated the effects of Aquamin (at 6 and 30 mg/mL) on SCFAs and lactate production, pH and gas production, and human microbiota composition. Our results demonstrate that Aquamin increased SCFA production (acetate and propionate by 8% and 24%, respectively, at 30 mg/mL dose), significantly decreased lactate production (30 mg/mL), and increased colonic fluid pH without inducing changes in colonic gas production or gastrointestinal (GI) microbiota composition. These results indicate that Aquamin may play a role in optimizing GI microbial function in an ex vivo setting.

Evaluation of fecal fermentation profile and bacterial community in organically fed dairy cows consuming forage-rich diets with different particle sizes

Organic cattle farming encourages the use of forage-rich diets, and the reduction of particle size has been suggested as an approach to improve forage utilization and enhance nutrient intake of cows. However, reducing forage particle size increases passage rate, as well as the flow of potentially fermentable nutrients out of the rumen, and the consequences for hindgut fermentation have not been evaluated yet. This study evaluated the effects of decreasing dietary forage particle size on the fecal short-chain fatty acid (SCFA) profile and the bacterial community structure of dairy cows fed forage-based rations. Twenty-one organically fed lactating Holstein cows (4 primiparous and 17 multiparous; mean and standard deviation 703 ± 65 kg body weight, 135 ± 104 days in milk) were divided into 2 groups and fed 1 of 2 diets for 34 d. Diets contained 20% concentrate and 80% forage (dry matter basis), and were fed either as a control with a forage geometric mean particle size of 52 mm (CON; 11 cows) or as a diet with the forage particle size reduced to a geometric mean size of 7 mm (RED; 10 cows). Fecal samples were collected at the end of the experiment, and samples were immediately frozen at -20°C. Samples were analyzed for SCFA, and the fecal bacterial community was evaluated using 16S rRNA sequencing. Data showed that the concentration of total SCFA was not affected by treatment, but the proportion of propionate, a key glucogenic precursor in cattle, tended to be greater for RED (13.3 and 13.8 ± 0.1%, respectively). The predominant bacterial phyla, including Firmicutes (58.0 ± 0.7%), Bacteroidetes (26.9 ± 0.4%), and Verrucomicrobia (4.0 ± 0.4%), were not affected by forage particle size. Family Lachnospiraceae increased in relative abundance when the RED diet was fed (12.1 and 13.9 ± 0.5% for CON and RED, respectively), and genera Acetitomaculum (1.1 and 1.8 ± 0.2%), Turicibacter (0.7 and 0.9 ± 0.1%), and Ruminobacter (0.1 and 0.4 ± 0.1%) increased in relative abundance when RED was fed. In addition, relative abundance of some fecal bacterial taxa was correlated with major fecal SCFA and pH. Reducing the particle size of forages, from 52 to 7 mm geometric mean particle size, maintained fecal concentration of total SCFA and tended to enhance propionate concentration, without risk of dysbiosis. Thus, results suggest that reduction of forage particle size represents an effective approach to optimizing forage utilization while maintaining hindgut fermentation and fecal bacterial diversity in dairy cows fed forage-rich diets.

Eggshell and environmental bacteria contribute to the intestinal microbiota of growing chickens

BackgroundThe initial intestinal microbiota acquired from different sources has profound impacts on animal health and productivity. In modern poultry production practices, the source(s) of the establishing microbes and their overall contribution during development of gastrointestinal tract communities are still unclear. Using fertilized eggs from two independent sources, we assessed the impact of eggshell- and environmental-associated microbial communities on the successional processes and bacterial community structure throughout the intestinal tract of chickens for up to 6 weeks post-hatch.ResultsCulturing and sequencing techniques identified a viable, highly diverse population of anaerobic bacteria on the eggshell. The jejunal, ileal, and cecal microbial communities for the egg-only, environment-only, and conventionally raised birds generally displayed similar successional patterns characterized by increasing community richness and evenness over time, with strains of Enterococcus, Romboutsia, and unclassified Lachnospiraceae abundant for all three input groups in both trials. Bacterial community structures differed significantly based on trial and microbiota input with the exception of the egg-exposed and conventional birds in the jejunum at week 1 and the ileum at week 6. Cecal community structures were different based on trial and microbiota input source, and cecal short-chain fatty acid profiles at week 6 highlighted functional differences as well.ConclusionWe identified distinct intestinal microbial communities and differing cecal short-chain fatty acid profiles between birds exposed to the microbiota associated with either the eggshell or environment, and those of conventionally hatched birds. Our data suggest the eggshell plays an appreciable role in the development of the chicken intestinal microbiota, especially in the jejunum and ileum where the community structure of the eggshell-only birds was similar to the structure of conventionally hatched birds. Our data identify a complex interplay between the eggshell and environmental microbiota during establishment and succession within the chicken gut. Further studies should explore the ability of eggshell- and environment-derived microbes to shape the dynamics of succession and how these communities can be targeted through interventions to promote gut health and mitigate food-borne pathogen colonization in poultry.

Promising discovery of beneficial Escherichia coli in the human gut.

Ingested dietary fibres are hydrolysed by colon microbiota to produce energy-providing short-chain fatty acids (SCFA) that stimulate anti-inflammatory effects. SCFA-producing bacteria were screened from bacteria isolated from human faeces using bromothymol blue as an acid indicator and gas chromatography for SCFA profiling. The beneficial functions (antagonistic activity, haemolytic activities, antibiotic susceptibility, mucus adherent percentage and toxin gene detection) were evaluated for the top five SCFA-producing bacteria isolated from three healthy volunteers that identified as Escherichia coli strains. They produced acetic, propionic, isobutyric, butyric, isovaleric, valeric and caproic acids at average concentrations of 15.9, 1.8, 1.1, 1.9, 1.8, 2.7 and 3.4mM, respectively. The SCFA production byE. colistrains was rapidly increased during the first 8h of incubation and gradually decreased after 16h of incubation. All E. coli strains showed acid and bile tolerance, resulting in a survival rate greater than 70% with no haemolytic activity, mucus adherence greater than 40% and susceptibility to conventional antibiotics. Hence, the selected E. coli strains exhibited promising probiotic properties with neither enterotoxin nor LPS producibility was detected. The present results confirm the existence of friendly and harmlessE. colistrains in human microbiota as potential probiotics.

Short-chain fructo-oligosaccharides supplementation to suckling piglets: Assessment of pre- and post-weaning performance and gut health.

Farmers face difficulties in redeeming their investment in larger litter sizes since this comes with larger litter heterogenicity, lower litter resilience and risk of higher mortality. Dietary oligosaccharides, given to the sow, proved beneficial for the offspring’s performance. However, giving oligosaccharides to the suckling piglet is poorly explored. Therefore, this field trial studied the effect of dietary short-chain fructo-oligosaccharides (scFOS; 1g/day; drenched) supplementation to low (LBW, lower quartile), normal (NBW, two intermediate quartiles) and high (HBW, upper quartile) birth weight piglets from birth until 7 or 21 days of age. Performance parameters, gut microbiome and short-chain fatty acids profile of feces and digesta were assessed at birth (d 0), d 7, weaning (d 21.5) and 2 weeks post-weaning (d 36.5). Additional parameters reflecting gut health (intestinal integrity and morphology, mucosal immune system) were analysed at d 36.5. Most parameters changed with age or differed with the piglet’s birth weight. Drenching with scFOS increased body weight by 1 kg in NBW suckling piglets and reduced the post-weaning mortality rate by a 100%. No clear difference in the IgG level, the microbiota composition and fermentative activity between the treatment groups was observed. Additionnally, intestinal integrity, determined by measuring intestinal permeability and regenerative capacity, was similar between the treatment groups. Also, intestinal architecture (villus lenght, crypt depth) was not affected by scFOS supplementation. The density of intra-epithelial lymphocytes and the expression profiles (real-time qPCR) for immune system-related genes (IL-10, IL-1ß, IL-6, TNFα and IFNγ) were used to assess mucosal immunity. Only IFNγ expression, was upregulated in piglets that received scFOS for 7 days. The improved body weight and the reduced post-weaning mortality seen in piglets supplemented with scFOS support the view that scFOS positively impact piglet’s health and resilience. However, the modes of action for these effects are not yet fully elucidated and its potential to improve other performance parameters needs further investigation.

2276-PUB: Fecal Profile of Short-Chain Fatty Acids: Associations with Environmental Factors in the First Year of Life of Women Participating in the Nutritionist’s Health Study

Introduction: Many studies report that environmental factors occurring in childhood may modulate the composition of the gut microbiota in adulthood, which in turn, produces short chain fatty acids (SCFA) with metabolic effects on inflammatory cascades associated with glycemic homeostasis. Objective: This study aims to analyze associations between type of delivery (vaginal/cesarean section), type of breast-feeding (maternal/artificial) and solid food introduction (start and quality) with fecal SCFA concentrations in women. Method: A cross-sectional study including 106 women, 28±5 years old and BMI 25±5 kg/m2, a questionnaire regarding from birth until the first year of life. For quantification of acetate, butyrate and propionate it was used gas chromatography-mass spectrometry in fecal samples. Results: Of the women surveyed 63% were cesarean delivered and 93% received maternal breast milk at some point. There was no correlation between SCFA concentration and type of delivery (p&amp;gt;0.05). Positive correlation was found between breastfeeding time and butyrate (r=0.24; p=0.02), there was a tendency with propionate (r=0.20; p=0.06). There was no correlation between time received infant formula and SCFA. There was a tendency of correlation between butyrate and beginning of cow’s milk intake (r=0.194; p=0.052). Age at solid food introduction correlates with propionate (r=0.22; p=0.026) and butyrate (r=0.25; p=0.012), and report a tendency with the acetate (r=0.18; p=0.07). In the first year of life, individuals who prefer salty crackers and pasta/potatoes had less concentration of propionate and acetate, respectively (p=0.026). Conclusion: The type of delivery was not associated with SCFA, but there was a favorable association with prolonged breastfeeding, late beginning of cow’s milk intake, introduction of solids starting after 6 months of age and lower intake of refined cereals. Disclosure M.G. Barbosa: None. I. Solar: None. R.G. Freitas: None. F.B. Ribeiro: None. C.O. Ramos: None. M.R. Sant’Ana: None. A.S. Hanada: None. N.S.K. Crouchan: None. C.M. Xavier: None. B.G. Neto: None. L.G.D. Folchetti: None. I.T. Silva: None. B. Almeida-Pititto: None. S.G. Ferreira-Vivolo: None. A.J. Vasques: None. Funding São Paulo Research Foundation (2017/10185-9, 2019/05450-0)

Functional and phylogenetic alterations in gut microbiome are linked to graft-versus-host disease severity

AbstractAcute graft-versus-host disease (aGVHD) is the main complication of hematopoietic stem cell transplantation (HSCT). Changes in gut microbiota composition have been associated with subsequent aGVHD, and reconstitution of healthy microbiota is currently being explored as a therapeutic approach. However, the specific actors in the intestinal ecosystem involved in the pathologic process at the time of aGVHD onset are not yet fully known. We prospectively collected stool samples from patients who underwent allogeneic HSCT. Patients sampled at aGVHD onset were compared with non-GVHD patients. To identify phylogenetic and functional signatures of the disease process, we determined fecal short-chain fatty acid (SFCA) profiles and used high-throughput DNA sequencing and real-time quantitative polymerase chain reaction to assess the microbiota composition. Microbiota alterations were highly specific of gastrointestinal (GI) aGVHD severity. Bacterial biomass and α-diversity were lower in severe aGVHD. We identified several bacterial signatures associated with severe aGVHD at disease onset; a negative correlation was observed with anaerobic bacteria of the Lachnospiraceae, especially the Blautia genus, and Ruminococcaceae families. In parallel, in severe aGVHD patients, we showed a dramatic decrease in the levels of the main SFCAs: acetate (75.8%), propionate (95.8%), and butyrate (94.6%). Mild aGVHD patients were characterized by conserved levels of propionate and Blautia propionate producers. Butyrate was significantly decreased in all GI aGVHD stages, representing a potential diagnostic marker of the disease. Specific microbiota and metabolic alterations were thus associated with aGVHD severity and may be useful for diagnostic and pathophysiologic purposes.