Higher Butyrate Concentrations Research Articles

Butyrate induces higher host transcriptional changes to inhibit porcine epidemic diarrhea virus strain CV777 infection in porcine intestine epithelial cells

Newborn piglets’ health is seriously threatened by the porcine epidemic diarrhea virus (PEDV), which also has a significant effect on the pig industry. The gut microbiota produces butyrate, an abundant metabolite that modulates intestinal function through many methods to improve immunological and intestinal barrier function. The objective of this investigation was to ascertain how elevated butyrate concentrations impacted the host transcriptional profile of PEDV CV777 strain infection. Our findings showed that higher concentrations of butyrate have a stronger inhibitory effect on PEDV CV777 strain infection. According to RNA-seq data, higher concentrations of butyrate induced more significant transcriptional changes in IPEC-J2 cells, and signaling pathways such as PI3K-AKT may play a role in the inhibition of PEDV CV777 strain by high concentrations of butyrate. Ultimately, we offer a theoretical and experimental framework for future research and development of novel approaches to harness butyrate’s antiviral infection properties.

Impact of Western Diet on Enterohemorrhagic Escherichia coli Colonization in the Human In Vitro Mucosal Artificial Colon as Mediated by Gut Microbiota.

Enterohemorrhagic Escherichia coli (EHEC) is a major food-borne pathogen that causes human disease ranging from diarrhea to life-threatening complications. Accumulating evidence demonstrates that the Western diet enhances the susceptibility to enteric infection in mice, but the effect of diet on EHEC colonization and the role of human gut microbiota remains unknown. Our research aimed to investigate the effects of a Standard versus a Western diet on EHEC colonization in the human in vitro Mucosal ARtificial COLon (M-ARCOL) and the associated changes in the gut microbiota composition and activities. After donor selection using simplified fecal batch experiments, two M-ARCOL bioreactors were inoculated with a human fecal sample (n = 4) and were run in parallel, one receiving a Standard diet, the other a Western diet and infected with EHEC O157:H7 strain EDL933. EHEC colonization was dependent on the donor and diet in the luminal samples, but was maintained in the mucosal compartment without elimination, suggesting a favorable niche for the pathogen, and may act as a reservoir. The Western diet also impacted the bacterial short-chain fatty acid and bile acid profiles, with a possible link between high butyrate concentrations and prolonged EHEC colonization. The work demonstrates the application of a complex in vitro model to provide insights into diet, microbiota, and pathogen interactions in the human gut.

Enhancing Biohydrogen Production: The Role of Iron-Based Nanoparticles in Continuous Lactate-Driven Dark Fermentation of Powdered Cheese Whey

Here, a comprehensive investigation was conducted under various operational strategies aimed at enhancing biohydrogen production via dark fermentation, with a specific focus on the lactate metabolic pathway, using powdered cheese whey as a substrate. Initially, a batch configuration was tested to determine both the maximum hydrogen yield (100.2 ± 4.2 NmL H2/g CODfed) and the substrate (total carbohydrates) consumption efficiency (94.4 ± 0.8%). Subsequently, a transition to continuous operation was made by testing five different operational phases: control (I), incorporation of an inert support medium for biomass fixation (II), addition of carbon-coated, zero-valent iron nanoparticles (CC-nZVI NPs) at 100 mg/L (III), and supplementation of Fe2O3 nanoparticles at concentrations of 100 mg/L (IV) and 300 mg/L (V). The results emphasized the critical role of the support medium in stabilizing the continuous system. On the other hand, a remarkable increase of 10% in hydrogen productivity was observed with the addition of Fe2O3 NPs (300 mg/L). The analysis of the organic acids’ composition unveiled a positive correlation between high butyrate concentrations and improved volumetric hydrogen production rates (25 L H2/L-d). Moreover, the presence of iron-based NPs effectively regulated the lactate concentration, maintaining it at low levels. Further exploration of the bacterial community dynamics revealed a mutually beneficial interaction between lactic acid bacteria (LAB) and hydrogen-producing bacteria (HPB) throughout the experimental process, with Prevotella, Clostridium, and Lactobacillus emerging as the predominant genera. In conclusion, this study highlighted the promising potential of nanoparticle addition as a tool for boosting biohydrogen productivity via lactate-driven dark fermentation.

In vitro digestion and fermentation of animal-derived fermentable substrates using canine and feline faecal inoculum

Abstract There is increasing interest in understanding the fermentative benefits of animal-derived fermentable substrates (ADFS) in pet foods. While previous research has assessed various ADFS using faecal inoculum derived from the cat, there is no published literature available for the dog. Additionally, very little is understood of the fermentation profiles of ADFS, such as skin and bone. Therefore, faeces were collected from a cohort of cats and dogs fed a complete and balanced high-protein diet and a selection of substrates were analysed in this study. Individual ADFS (tendon, bone cake, mechanically deboned meat (MDM), corium and hydrolysed collagen (a positive control)) were digested in vitro, followed by fermentation using either canine or feline faecal inoculum. Concentrations of butyrate, indole and ammonia were determined after 24 h of fermentation. Regardless of whether cat or dog faecal inoculum was used, fermentation of hydrolysed collagen produced the highest (P < 0.01) concentrations of butyrate and ammonia and the lowest concentrations of indole. For the other substrates, there were differences in the fermentation profiles between the canine and feline inocula. During the feline faecal fermentations, in comparison to the other substrates bone cake produced high (P < 0.05) butyrate concentrations, whereas in the dog faecal fermentations, MDM resulted in high (P < 0.05) butyrate concentrations. In conclusion, ADFS from different alternative co-products lead to different fermentation products, providing valuable information which may be considered in canine and feline dietary formulations.

CO2 supply is a powerful tool to control homoacetogenesis, chain elongation and solventogenesis in ethanol and carboxylate fed reactor microbiomes.

Anaerobic fermentation technology enables the production of medium chain carboxylates and alcohols through microbial chain elongation. This involves steering reactor microbiomes to yield desired products, with CO2 supply playing a crucial role in controlling ethanol-based chain elongation and facilitating various bioprocesses simultaneously. In the absence of CO2 supply (Phase I), chain elongation predominantly led to n-caproate with a high selectivity of 96 Cmol%, albeit leaving approximately 80% of ethanol unconverted. During this phase, C. kluyveri and Proteiniphilum-related species dominated the reactors. In Phase II, with low CO2 input (2.0NmL L-1min-1), formation of n-butyrate, butanol, and hexanol was stimulated. Increasing CO2 doses in Phase III (6 NmLL-1min-1) led to CO2 utilization via homoacetogenesis, coinciding with the enrichment of Clostridium luticellarii, a bacterium that can use CO2 as an electron acceptor. Lowering CO2 dose to 0.5NmL L-1min-1 led to a shift in microbiome composition, diminishing the dominance of C. luticellarii while increasing C. kluyveri abundance. Additionally, other Clostridia, Proteiniphilum, and Lactobacillus sakei-related species became prevalent. This decrease in CO2 load from 6 to 0.5NmL L-1min-1 minimized excessive ethanol oxidation from 30%-50% to 0%-3%, restoring a microbiome favoring net n-butyrate consumption and n-caproate production. The decreased ethanol oxidation coincided with the resurgence of hydrogen formation at partial pressures above 1%. High concentrations of butyrate, caproate, and ethanol in the reactor, along with low acetate concentration, promoted the formation of butanol and hexanol. It is evident that CO2 supply is indispensable for controlling chain elongation in an open culture and it can be harnessed to stimulate higher alcohol formation or induce CO2 utilization as an electron acceptor.

Contrary effects of the gut metabolites deoxycholate and butyrate on the acetylcholine-evoked calcium response in an enteroendocrine cell model

The interaction of nutrients and bacterial-derived products and their enteroendocrine effects are still enigmatic. We used the human enteroendocrine tumour cell line P-STS isolated from the terminal ileum as cellular sentinel. In this model, P-STS cells react to acetylcholine or histamine with an increase in intracellular calcium mediated by T-type voltage-gated calcium channels followed by serotonin secretion. Incubation with the secondary bile acid deoxycholate inhibited the calcium response to acetylcholine but not to histamine. No response to other nutrients or bacterial products tested (i.e. palmitate, lactate, acetate, proprionate, butyrate, the OR51E1 ligand isovalerate and the HCA2 ligand β-hydroxybutyrate) could be observed. The bile acid receptor TGR5 agonist CCDC did not inhibit the acetylcholine-induced calcium response, and the free fatty acid receptor 2 agonist 4-CMTB had no enhancing effect on the calcium response to acetylcholine. Only butyrate had an enhancing effect on the calcium response in P-STS cells when added for short-time pre-incubation before challenge with acetylcholine. From our data we propose i) an inhibitory effect of deoxycholate at the muscarinic acetylcholine receptor M3 without involvement of TGR5, and that ii) high butyrate concentrations derived from bacterial metabolism of carbohydrates might increase intestinal release of serotonin and potentially also other gut hormones and satiety-inducing peptides independently of known butyrate receptors, thereby influencing intestinal motility and ion secretion.

Enhanced biological phosphorus removal systems adapting to seawater intrusion: Impacts of varying carbon source on metabolic adaptation in Candidatus Accumulibacter phosphatis

In enhanced biological phosphorus removal (EBPR) systems, seawater intrusion may introduce uncertainty into the metabolism of Polyphosphate Accumulating Organisms (PAOs), especially when volatile fatty acid (VFA) profiles derived from fermentation liquids serves as the carbon source. This study investigated the impacts of varying sea salt salinity levels on the metabolism of Accumulibacter with different VFA as carbon sources. As sea salt salinity increased from 0.5 % to 3.5 %, catabolism (phosphorus release, glycogen and PHA biodegradation) was inhibited to a much lower degree compared to anabolism (phosphorus uptake, glycogen and PHA synthesis). Under seawater conditions, the intracellular free Mg2+ was partially excreted, leading to a subsequent reduction in glycogen metabolism and PHA synthesis. The decrease in PHA synthesis resulted in the diminished aerobic metabolic activity and polyphosphatase (PPX) enzyme activity, collectively causing decreased phosphorus uptake. Moreover, additional energy consumption was used to alleviate osmotic stress, which limited the energy used for esterase activity, translation activity, and phosphorus and glycogen metabolism, resulting in a decrease in growth activity. When using butyrate as a carbon source, Accumulibacter exhibited the highest metabolic activity (49.8 %), making it more adaptable to seawater conditions. The clade IIF was the dominant Accumulibacter in seawater conditions and could synthesize more PHA with butyrate, isobutyrate and valerate as a carbon source, respectively, stabilizing aerobic metabolism. Therefore, fermentation liquid containing high concentrations of butyrate and the activated sludge dominated by clade IIF have the potential to enhance the stabilization of EBPR systems in wastewater treatment plants infiltrated by seawater.

Microbiome analysis of thai traditional fermented soybeans reveals short-chain fatty acid-associated bacterial taxa

Thua Nao is a Thai traditional fermented soybean food and low-cost protein supplement. This study aimed to evaluate the bacterial community in Thua Nao from northern Thailand and assess potentially active short-chain fatty acids (SCFAs)-related bacteria. Sixty-five Thua Nao consisting of 30 wet and 35 dried samples were collected from six provinces: Chiang Rai, Chiang Mai, Mae Hong Son, Lampang, Lamphun, and Phayao. Bacterial diversity was significantly higher in the wet samples than in the dried samples. The dominant phyla were Firmicutes (92.7%), Proteobacteria (6.7%), Actinobacteriota (0.42%), and Bacteroidota (0.26%). The genus Bacillus (67%) was the most represented in all samples. Lactobacillus, Enterococcus, and Globicatella were enriched in the wet samples. Assessment of the SCFA-microbiota relationships revealed that high butyrate and propionate concentrations were associated with an increased Clostridiales abundance, and high acetate concentrations were associated with an increased Weissella abundance. Wet products contained more SCFAs, including acetate (P = 2.8e−08), propionate (P = 0.0044), butyrate (P = 0.0021), and isovalerate (P = 0.017), than the dried products. These results provide insight into SCFA-microbiota associations in Thua Nao, which may enable the development of starter cultures for SCFA-enriched Thua Nao production.

Syntrophic Butyrate-Oxidizing Consortium Mitigates Acetate Inhibition through a Shift from Acetoclastic to Hydrogenotrophic Methanogenesis and Alleviates VFA Stress in Thermophilic Anaerobic Digestion

In anaerobic digestion (AD), butyrate is degraded by syntrophic consortium, but can accumulate in highly loaded AD systems. The effect of butyrate on the AD process attracts much less attention than propionate or acetate. In this work, an enrichment culture of the thermophilic butyrate-oxidizing syntrophic consortium was obtained by gradually increasing the initial butyrate concentration from 20 to 170 mM. Surprisingly, even the highest butyrate concentration did not significantly inhibit the methanogenic community, and the stage of acetate degradation was the limiting overall rate of the process. At 170 mM butyrate, the bacterial community changed towards the dominance of syntrophic acetate-oxidizing (SAO) bacteria related to Syntrophaceticus (42.9%), Syntrophomonas (26.2%) and Firmicutes (26.2%), while the archaeal community experienced a sharp decrease in the abundance of Methanosarcina thermophila (from 86.0 to 25.0%) and increase in Methanothermobacter thermautotrophicus (from 3.2 to 53.1%) and Methanomassiliicoccus (from 3.2 to 21.9%). Thus, the shift from acetoclastic methanogenesis to SAO coupled to hydrogenotrophic methanogenesis occurred as an adaptive strategy to overcome high acetate (~200 mM) build-up. Bioaugmentation with the obtained enrichment culture was effective in mitigating the butyrate-dominated VFA build-up during the AD of readily biodegradable waste, increasing the methane production rate, methane yield and volatile solids removal by more than 3.5, 6.2 and 2.9 times, respectively. Our study revealed that the thermophilic butyrate-oxidizing consortia as bioaugmented culture could be the potential strategy to alleviate the high organic load and VFA stress of AD.

Intensification of Hydrogen Production by a Co-culture ofSyntrophomonas wolfei and Rhodopseudomonas palustrisEmploying High Concentrations of Butyrate as a Substrate.

The purpose of this study is to present an effective form of developing a sequential dark (DF) and photo (PF) fermentation using volatile fatty acids (VFAs) and nitrogen compounds as bonding components between both metabolic networks of microbial growing in each fermentation. A simultaneous (co-)culture of Syntrophomonas wolfei (with its ability to consume butyrate and produce acetate) and Rhodopseudomonas palustris (that can use the produced acetate as a carbon source) performed a syntrophic metabolism. The former bacteria consumed the acetate/butyrate mixture reducing the butyrate concentration below 2.0g/L, permitting Rhodopseudomonas palustris to produce hydrogen. Considering that the inoculum composition (Syntrophomonas wolfei/Rhodopseudomonas palustris) and the nitrogen source (yeast extract) define the microbial biomass specific productivity and the butyrate consumption, a response surface methodology defined the best inoculum design and yeast extract (YE) yielding to the highest biomass concentration of 1.1g/L after 380.00h. A second culture process (without a nitrogen source) showed the biomass produced in the previous culture process yields to produce a total cumulated hydrogen concentration of 3.4 mmol. This value was not obtained previously with the pure strain Rhodopseudomonas palustris if the culture medium contained butyrate concentration above 2.0g/L, representing a contribution to the sequential fermentation scheme based on DF and PF.

Developing a synbiotic biotherapy to prevent allergic responses to food

Abstract Over the past generation, the prevalence of food allergies has risen dramatically, but the development of treatments has lagged. The commensal gut microbiome is critically involved in regulating allergic responses to food. Our laboratory has described a humanized model of cow’s milk allergy (CMA) in which germ-free mice are colonized with feces from healthy or CMA infants and sensitized to the cow’s milk allergen b-lactoglobulin (BLG). The healthy infant microbiota and specifically a single Clostridial species, Anaerostipes caccae, protect against the development of anaphylaxis, while the CMA microbiota does not induce this protection. We isolated a novel sub-strain of A. caccae (A. caccae_lahuc) from the feces of a healthy infant, sequenced its genome, and characterized its growth on various substrates in vitro. A. caccae_lahuc produces high concentrations of butyrate, a key immunoregulatory molecule in the gut which we hypothesize is critical to its protective impact. A. caccae_lahuc engrafts readily into the dysbiotic CMA microbiota, but co-delivery with lactulose is required for measurable butyrate production in vivo. Lactulose undergoes primary degradation by bacteria in the CMA feces which release small metabolites that A. caccae_lahuc can ferment into butyrate. Preliminary data show that this novel synbiotic biotherapy (A. caccae_lahuc + lactulose) prevents allergic responses to BLG in CMA-colonized mice as measured by serum mMCPT-1 and core body temperature upon challenge, and type 2 cytokine production in BLG-stimulated splenocytes. Further exploration into the mechanisms behind this therapeutic effect will contribute toward the larger goal of developing bacterial biotherapeutics against food allergies.

Effects of Total Dietary Fiber on Cecal Microbial Community and Intestinal Morphology of Growing White Pekin Duck.

The current study was to investigate the effects of total dietary fiber (TDF) on growth performance, cecal structure, cecal microbial community, and short-chain fatty acids (SCFAs) profiles in the cecum of growing White Pekin ducks. A total of 108 male Pekin ducks of 14-days-old were randomly allocated and fed diets containing 12.4, 14.7, and 16.2% TDF for 35 days. Each dietary treatment consisted of six replicates with six birds each. The results showed that 14.7 and 16.2% TDF treatments promoted growth performance relative to 12.4% TDF treatments (P < 0.05). A total of 14.7 and 16.2% TDF treatments significantly elevated villus height, the ratio of villus height to crypt depth and muscle layer thickness of cecum, and lowered crypt depth compared with 12.4% TDF treatment (P < 0.05). Simultaneously, 14.7 and 16.2% TDF treatments up-regulated Claudin-1 mRNA expression of barrier genes in the cecum compared with 12.4% TDF (P < 0.05). Butyrate-producing bacteria like Oscillopiraceae affiliating to the phyla Firmicutes were observed as a biomarker in the 16.2% TDF. Higher concentration of butyrate in the cecum was obtained in the 14.7% TDF compared with 12.4 and 16.2% TDF (P < 0.05). The concentrations of isobutyrate, valerate, and isovalerate in the cecum were significantly increased in the 16.2% TDF compared with 12.4 and 14.7% TDF (P < 0.05). Meanwhile, the abundance of genus UCG-005 and Enterococcus was positive correlations with isobutyrate and valerate (P < 0.05). However, the concentration of propionate in the cecum significantly decreased in 14.7 and 16.2% TDF treatments relative to 12.4% TDF treatments (P < 0.05). In summary, increasing TDF levels improved growth performance, cecal histomorphology, and barrier function of meat ducks and it might be mediated by the changes of microbiota communities, especially bloom of SCFAs-producing bacteria, which facilitated the interaction between intestinal mucosa and microbiota.

Dynamics of sulfate reduction in the thermophilic dark fermentation of sugarcane vinasse: A biohydrogen-independent approach targeting enhanced bioenergy production

Dark fermentation (DF) systems usually exploit bioH2 production aiming to improve the energetic potential (EP) of anaerobic digestion (AD) schemes. In the case of sulfate-rich substrates, the versatility of fermentative consortia may favor the establishment of sulfidogenic conditions, minimizing/eliminating the competition between sulfate-reducing bacteria (SRB) and methanogens in the sequential stage and providing sulfide-free biogas. This study details the establishment of sulfidogenesis on the thermophilic DF of sugarcane vinasse in a continuous polyurethane-filled fixed-film reactor, also estimating the EP for vinasses according to different AD-related approaches. High sodium levels naturally stimulated SRB by maintaining high pH values (> 6.0), whilst NaHCO3 dosing was required when natural buffering conditions were not maintained. High acetate (~ 3 g-COD L−1) and COD/sulfate ratio (> 25.0) levels indicated a high-quality substrate for further methanogenesis. High butyrate concentrations (~11 g-COD L−1) coupled to caproate detection indicated the occurrence of chain elongation processes, leading to low bioH2 production levels (<400 NmL-H2 L−1 d−1). However, estimates indicated 12.2%- and 51.7%-increases in the methane-based EP (MJ m−3) of sulfate-free fermented vinasse (216.72) relative to sulfate-rich fermented (193.10) and fresh (142.88) vinasses. Hence, the ultimate approach for maximizing bioenergy production from sugarcane vinasse is detailed in this study.

Oral administration of Moringa oleifera leaf powder relieves oxidative stress, modulates mucosal immune response and cecal microbiota after exposure to heat stress in New Zealand White rabbits

BackgroundHeat stress (HS) disrupts the gut barrier allowing the uptake of lipopolysaccharide (LPS) and leads to an inflammatory response and changes in gut microbiota composition. Moringa oleifera leaf powder (MOLP) has been proposed to combat HS, yet its alleviate role is currently under investigation. The current study investigated the effects of chronic HS and MOLP supplementation on changes in redox status and immune response of cecal mucosa along with alteration in cecal microbiota.MethodsA total of 21 young New Zealand White (NZW) rabbits (male) about 32 weeks old (mean body weight of 3318 ± 171 g) reared on a commercial pelleted diet were employed; divided into three groups (n = 7): control (CON, 25 °C), heat stress (HS, 35 °C for 7 h daily), and HS supplemented orally with MOLP (HSM, 35 °C) at 200 mg/kg body weight per day for 4 weeks.ResultsThe results demonstrated that MOLP supplementation increased organ index of cecal tissue compared with the HS group (P > 0.05). Levels of malonaldehyde (MDA) and activity of superoxide dismutase (SOD) as well as lactate dehydrogenase (LDH) were reduced in the cecal mucosa of the HSM group compared with the HS group. MOLP downregulated the contents of cecal mucosa LPS, several inflammatory markers (TNF-α/IL-1α/IL-1β), and myeloperoxidase (MPO) in the HSM group (P < 0.05). Secretory immunoglobulin A (SIgA) was increased in the HSM group compared with the HS group (P < 0.05). The transcriptome of cecal mucosa showed that MOLP reduced gene expression relative to several immune factors, including IL-10, IFNG, and RLA, whereas both HS and MOLP increased the gene expression of fat digestion and absorption pathway, including APOA1, FABP1, FABP2, MTTP, and LOC100344166, compared to the CON group (P < 0.001). At the phylum level, the relative abundance of Proteobacteria was increased by HS, while Actinobacteria was significantly increased by HSM compared to other groups (P < 0.05). At genus level, Papillibacter was higher in abundance in HSM groups compared to CON and HS groups (P < 0.05). Higher butyrate concentrations were observed in the HSM group than HS and CON groups (P < 0.05).ConclusionIn conclusion, HS in growing rabbits resulted in alteration of cecal microbiota at phyla level as well as increased oxidative stress and expression of mucosal inflammatory genes. Whereas, oral MOLP supplementation elevated the relative weight of cecum, affected their immunological and cecal micro-ecosystem function by improving antioxidant status and down-regulating mucosal tissue inflammatory response.

Butyrate alters lipopolysaccharide-induced inflammatory response by porcine monocytes

Abstract Continuous, low-level inflammation can have a negative impact on intestinal health. Butyrate is a short-chain fatty acid produced in the lower intestinal tract by microbial fermentation of dietary fibers and modulates immune cell responses through various mechanisms. Blood monocytes migrate into the intestine and respond to signals, including potential modulation by butyrate or in response to microbial components, such as lipopolysaccharide (LPS). To investigate the impact of butyrate on inflammatory response to LPS, porcine monocytes were either cultured with a combination of LPS and varying butyrate concentrations or pre-treated with butyrate before LPS exposure. Independent of butyrate, LPS stimulation of monocytes induced IL-1β and TNF production. Monocytes cultured just three hours (h) prior to LPS stimulation had reduced overall cytokine production, suggesting rapid phenotypic changes in monocytes. Butyrate had varying effects on the amount of cytokine produced in response to LPS. Specifically, low butyrate concentrations (0.25 mM) limited IL-1β production, but did not impact TNF production. Higher butyrate concentrations (&amp;gt; 4 mM) limited TNF production to LPS stimulation. Pre-treatment of monocytes with butyrate for 3 h prior to LPS stimulation altered IL-1β production. Pre-treatment with 0.25 mM butyrate did not limit IL-1β production, but concentrations &amp;gt; 4 mM did. In conclusion, reduction of LPS-induced cytokine production by porcine monocytes was dependent on butyrate concentration and exposure time relative to LPS exposure. Further work is ongoing to understand the mechanism of butyrate modulation of pig monocytes and inflammatory response to LPS for potential application in vivo.

Butyrate Reduces TNFα‐Induced Cell Toxicity in Rat Colonic Epithelium

Hypertension (HTN) remains one of the most concerning preventable conditions for the global human population. The colonic lumen is the site of production and transport of many vasoactive metabolites and neurotransmitters that can modulate the immune system, affect cellular metabolism, and subsequently regulate blood pressure. As a critical interface between the gut bacteria and the host, dysfunctional and inflammed colonic epithelium has been associated with HTN. Here, we investigate the role of microbial metabolites in the regulation of colonic cell homeostasis, with a special emphasis on the interaction between tumor necrosis factor α (TNFα) and a gut bacterial short chain fatty acid (SCFA) metabolite butyrate, both of which have been implicated in HTN. We hypothesized that TNFα and butyrate exert opposing effects on colonic homeostasis through a mitochondrial mechanism, which is intrinsically linked to the pathology of HTN. To investigate this, Rat Primary Colon Epithelial Cells (CECs) (Cell Biologics, Cat# RN-6047) were seeded at 10,000 cells per well in a 96 well plate at passage #5-10 and exposed to increasing concentrations of TNFα (50pg/mL to 10ng/mL) and butyrate (50 µM to 25mM) in vitro for 24hrs. Using the multiplexed Alamar Blue (Cell Viability) (Thermofisher Scientific, Cat# DAL1025) and Cytotox Green (Cytotoxicity) (Promega, Cat# G8741) assays, we show that increasing concentrations of TNFα (150pg/mL-10ng/mL) led to a concentration-dependent reduction in CEC viability (by 20-90%) and elevation in CEC cytotoxicity (by 10-40%). Interestingly, treatment with butyrate alone (500µM-25mM) also led to a concentration-dependent reduction in cell viability (by 10-70%), while an increase in CEC cytotoxicity (by 10-20%) was observed only with high non-physiologic concentrations of butyrate (2.5-25mM). Next, CECs were exposed to TNFα (1 ng/mL) in the presence and absence of the physiologic colonic concentration of butyrate (500 µM). We found that co-treatment with butyrate led to a significant reduction in TNFα-induced CEC cytotoxicity (by 15%) and improved TNFα-reduced CEC viability (by 15%). Interestingly, 24hr in vitro exposure to TNFα (50pg/mL-10ng/mL) and butyrate (50 µM to 25mM) alone reduced relative ATP levels in a concentration-dependent fashion (by 15%-60% and 15%-90% respectively). Thus, we assessed oxidative phosphorylation using Seahorse and found that both butyrate (500 µM) and TNFα (1 ng/mL) alone reduced ATP-linked respiration (by 50% each), whereas co-treatment of CECs with butyrate and TNFα produced an 85% reduction in ATP-linked mitochondrial respiration. Taken together, these experiments suggest that colonic butyrate may attenuate the epithelial cell toxicity of TNFα by reducing mitochondrial activity. Moreover, while not affecting cell cytotoxicity at physiologic concentrations, butyrate appears to be toxic at higher concentrations in vitro. This may have important implications on gut homeostasis in the context of HTN where TNFα and butyrate levels are reportedly deregulated. Further research is necessary to elucidate the specific role of TNFα on colonic cell homeostasis and how butyrate and other SCFAs may affect this in order to better understand the host-microbiota interactions in HTN.

A mixture of quebracho and chestnut tannins drives butyrate-producing bacteria populations shift in the gut microbiota of weaned piglets.

Weaning is a critical period for piglets, in which unbalanced gut microbiota and/or pathogen colonisation can contribute to diseases that interfere with animal performance. Tannins are natural compounds that could be used as functional ingredients to improve gut health in pig farming thanks to their antibacterial, antioxidant, and antidiarrhoeal properties. In this study, a mixture of quebracho and chestnut tannins (1.25%) was evaluated for its efficacy in reducing the negative weaning effects on piglet growth. Microbiota composition was assessed by Illumina MiSeq 16S rRNA gene sequencing of DNA extracted from stools at the end of the trial. Sequence analysis revealed an increase in the genera Shuttleworthia, Pseudobutyrivibrio, Peptococcus, Anaerostipes, and Solobacterium in the tannin-supplemented group. Conversely, this dietary intervention reduced the abundance of the genera Syntrophococcus, Atopobium, Mitsuokella, Sharpea, and Prevotella. The populations of butyrate-producing bacteria were modulated by tannin, and higher butyrate concentrations in stools were detected in the tannin-fed pigs. Co-occurrence analysis revealed that the operational taxonomic units (OTUs) belonging to the families Veillonellaceae, Lachnospiraceae, and Coriobacteriaceae occupied the central part of the network in both the control and the tannin-fed animals. Instead, in the tannin group, the OTUs belonging to the families Acidaminococcaceae, Alcaligenaceae, and Spirochaetaceae characterised its network, whereas Family XIII Incertae Sedis occupied a more central position than in the control group. Conversely, the presence of Desulfovibrionaceae characterised the network of the control group, and this family was not present in the network of the tannin group. Moreover, the prediction of metabolic pathways revealed that the gut microbiome of the tannin group possessed an enhanced potential for carbohydrate transport and metabolism, as well as a lower abundance of pathways related to cell wall/membrane/envelope biogenesis and inorganic ion transport. In conclusion, the tested tannins seem to modulate the gut microbiota, favouring groups of butyrate-producing bacteria.

The Gut Microbiome in Heart Transplantation: A Prospective Pilot Study

<h3>Purpose</h3> The contribution of the intestinal microbiome to alloimmunity in heart transplantation (HT) is unknown. We examined fecal microbiota of HT recipients at time of transplantation and characterized microbiota composition, diversity, and metabolite production. <h3>Methods</h3> In a prospective, observational, pilot study, pre and post-HT stool samples were collected and microbiota composition determined by 16S ribosomal RNA gene sequencing. Fecal short chain fatty acid (SCFA) concentrations were determined by targeted metabolomics. HT samples were compared to those of healthy controls (HC) and liver transplant (LT) subjects. <h3>Results</h3> 52 fecal samples were analyzed from 9 HT recipients (1 heart-kidney, 1 heart-liver, 1 heart-liver-kidney) and compared to 15 HC and 8 LT subjects. A measure of within-sample microbial diversity, the inverse Simpson index was lower among HT recipients before and after transplantation compared to HC, but significantly higher than LT recipients (p<0.001, Fig. 1). HT patients had higher frequencies of Lactobacillus, Enterococcus, and Faecalicoccus species than HC, while LT patients had marked losses of normal bacterial taxa (Fig. 1). Compared to LT, HT microbiota had significantly higher abundances of the butyrate-producing anaerobic bacterial species Lachnospiraceae (21% vs 4%, p<0.001) and Ruminococcaceae (12% vs 1%, p<0.001), and higher butyrate concentrations (7.1mM vs 1.1mM, p<0.001). Marked changes to the relative abundance of these species and SCFA production were observed over time for individual subjects. Analysis of beta diversity with Bray-Curtis and Unifrac PCoA demonstrated clustering by cohort. <h3>Conclusion</h3> The microbiota of HT recipients is distinct from HC or LT subjects. These intriguing and readily measurable changes to bacterial composition and SCFA production are observed across the transplantation timeline. Further recruitment, collection, and correlation to clinical outcomes is in progress.

Clove (Syzygium aromaticum) and its phytochemicals in ruminant feed: an updated review

Livestock has a significant contribution to methane emission as a portion of their ingested energy, which is wasted in the form of methane (2–15%), and ammonia (75–95%). Microbial fermentation in ruminants results in the loss of energy in methanogenesis and protein via ammonia nitrogen excretion, causing a decline in feed efficiency and acting as environmental pollutants. Antibiotics were used to reduce these losses in the rumen, but this approach was restricted in animal products. Some plants or their bioactive extracts/metabolites such as organo-sulphur compounds, saponins, essential oils, flavonoids, and tannins exhibited the potential to limit the methanogenesis by altering the rumen microflora. Plant extracts, including clove (Syzygium aromaticum) bud oil and clove extracts, were used as feed additives to manipulate rumen fermentation. Clove oil interacts with bacterial cells and inhibits the growth multiplication of methanogenic and deaminating bacteria. This results in a reduction in ammonia, nitrogen, methane, acetate concentration, and high propionate and butyrate concentrations. Eugenol is one of the bioactive constituents in clove that manipulate rumen fermentation by increasing propionate production, decrease acetate and methane production, and alter the proteolysis pattern, peptidolysis, and amino acid deamination in the rumen. The current review reports on using of clove phytochemicals and extracts in manipulating rumen fermentation to inhibit methanogenesis and energy loss as well as ammonia–nitrogen waste.

Polyvinyl Butyrate Nanoparticles as Butyrate Donors for Colitis Treatment.

Butyrate has been attracting attention for the suppression of inflammatory bowel disease (IBD). However, clinical trials of butyrate for IBD treatment have resulted in controversial outcomes, likely owing to the adverse effect of butyrate on the intestinal epithelium that was observed at high butyrate concentrations. Herein, we propose polyvinyl butyrate (PVBu) nanoparticles (NPs) as butyrate donors for delivery to the lower part of the intestine for the treatment of colitis. The PVBu NPs suppressed the inflammatory activation of macrophages in vitro, although sodium butyrate inversely further activated macrophages. Oral administration of NPs did not change the luminal concentration of free butyrate; however, NPs showed a therapeutic effect on a colitis mouse model. In addition, incorporation of vitamin D3 into the NPs enhanced the therapeutic effect on colitis. Hence, PVBu NPs are a promising therapeutic for IBD treatment, not only as a butyrate donor but also as a carrier for hydrophobic drugs like vitamin D3.