Refined diet consumption increases neuroinflammatory signalling through bile acid dysmetabolism.

Over recent decades, dietary patterns have changed significantly due to the increasing availability of convenient, ultra-processed refined foods. Refined foods are commonly depleted of key bioactive compounds, which have been associated with several deleterious health conditions. As the gut microbiome can influence the brain through a bidirectional communication system known as the 'microbiota-gut-brain axis', the consumption of refined foods has the potential to affect cognitive health. In this study, multi-omics approaches were employed to assess the effect of a refined diet on the microbiota-gut-brain axis, with a particular focus on bile acid metabolism. Mice maintained on a refined low-fat diet (rLFD), consisting of high sucrose, processed carbohydrates and low fibre content, for eight weeks displayed significant gut microbial dysbiosis, as indicated by diminished alpha diversity metrics (p < 0.05) and altered beta diversity (p < 0.05) when compared to mice receiving a chow diet. Changes in gut microbiota composition paralleled modulation of the metabolome, including a significant reduction in short-chain fatty acids (acetate, propionate and n-butyrate; p < 0.001) and alterations in bile acid concentrations. Interestingly, the rLFD led to dysregulated bile acid concentrations across both the colon (p < 0.05) and the brain (p < 0.05) which coincided with altered neuroinflammatory gene expression. In particular, the concentration of TCA, TDCA and T-α-MCA was inversely correlated with the expression of NF-κB1, a key transcription factor in neuroinflammation. Overall, our results suggest a novel link between a refined low-fat diet and detrimental neuronal processes, likely in part through modulation of the microbiota-gut-brain axis and bile acid dysmetabolism.

Maternal gut microbiota Bifidobacterium promotes placental morphogenesis, nutrient transport and fetal growth in mice.

The gut microbiota plays a central role in regulating host metabolism. While substantial progress has been made in discerning how the microbiota influences host functions post birth and beyond, little is known about how key members of the maternal gut microbiota can influence feto-placental growth. Notably, in pregnant women, Bifidobacterium represents a key beneficial microbiota genus, with levels observed to increase across pregnancy. Here, using germ-free and specific-pathogen-free mice, we demonstrate that the bacterium Bifidobacterium breve UCC2003 modulates maternal body adaptations, placental structure and nutrient transporter capacity, with implications for fetal metabolism and growth. Maternal and placental metabolome were affected by maternal gut microbiota (i.e. acetate, formate and carnitine). Histological analysis of the placenta confirmed that Bifidobacterium modifies placental structure via changes in Igf2P0, Dlk1, Mapk1 and Mapk14 expression. Additionally, B. breve UCC2003, acting through Slc2a1 and Fatp1-4 transporters, was shown to restore fetal glycaemia and fetal growth in association with changes in the fetal hepatic transcriptome. Our work emphasizes the importance of the maternal gut microbiota on feto-placental development and sets a foundation for future research towards the use of probiotics during pregnancy.

Macrophage metabolism in the intestine is compartment specific and regulated by the microbiota.

Intestinal macrophages play a vital role in the maintenance of gut homeostasis through signals derived from the microbiota. We previously demonstrated that microbial-derived metabolites can shape the metabolic functions of macrophages. Here, we show that antibiotic-induced disruption of the intestinal microbiota dramatically alters both the local metabolite environment and the metabolic functions of macrophages in the colon. Broad-spectrum antibiotic administration in mice increased the expression of the large neutral amino acid transporter LAT1 and accordingly, amino acid uptake. Subsequently, antibiotic administration enhanced the metabolic functions of colonic macrophages, increasing phosphorylation of components of mammalian/mechanistic target of rapamycin signalling pathways, with increased expression of genes involved in glycolysis and oxidative phosphorylation (OXPHOS), increased mitochondrial function, increased rate of extracellular acidification (ECAR; measure of glycolysis) and increased rate of oxygen consumption (OCR; measure of OXPHOS). Small bowel macrophages were less metabolically active than their colonic counterparts, with macrophage metabolism in the small intestine being independent of the microbiota. Finally, we reveal tissue-resident Tim4+  CD4+ macrophages exhibit enhanced fatty acid uptake alongside reduced fatty acid synthesis compared to recruited macrophages. Thus, the microbiota shapes gut macrophage metabolism in a compartment-specific manner, with important implications for monocyte recruitment and macrophage differentiation.

The early life microbiota protects neonatal mice from pathological small intestinal epithelial cell shedding.

The early life gut microbiota plays a crucial role in regulating and maintaining the intestinal barrier, with disturbances in these communities linked to dysregulated renewal and replenishment of intestinal epithelial cells. Here we sought to determine pathological cell shedding outcomes throughout the postnatal developmental period, and which host and microbial factors mediate these responses. Surprisingly, neonatal mice (Day 14 and 21) were highly refractory to induction of cell shedding after intraperitoneal administration of liposaccharide (LPS), with Day 29 mice showing strong pathological responses, more similar to those observed in adult mice. These differential responses were not linked to defects in the cellular mechanisms and pathways known to regulate cell shedding responses. When we profiled microbiota and metabolites, we observed significant alterations. Neonatal mice had high relative abundances of Streptococcus, Escherichia, and Enterococcus and increased primary bile acids. In contrast, older mice were dominated by Candidatus Arthromitus, Alistipes, and Lachnoclostridium, and had increased concentrations of SCFAs and methyamines. Antibiotic treatment of neonates restored LPS-induced small intestinal cell shedding, whereas adult fecal microbiota transplant alone had no effect. Our findings further support the importance of the early life window for microbiota-epithelial interactions in the presence of inflammatory stimuli and highlights areas for further investigation.

Effects of in vitro metabolism of a broccoli leachate, glucosinolates and S-methylcysteine sulphoxide on the human faecal microbiome.

PURPOSE: Brassica are an important food source worldwide and are characterised by the presence of compounds called glucosinolates. Studies indicate that the glucosinolate derived bioactive metabolite sulphoraphane can elicit chemoprotective benefits on human cells. Glucosinolates can be metabolised in vivo by members of the human gut microbiome, although the prevalence of this activity is unclear. Brassica and Allium plants also contain S-methylcysteine sulphoxide (SMCSO), that may provide additional health benefits but its metabolism by gut bacteria is not fully understood. METHODS: We examined the effects of a broccoli leachate (BL) on the composition and function of human faecal microbiomes of five different participants under in vitro conditions. Bacterial isolates from these communities were then tested for their ability to metabolise glucosinolates and SMCSO. RESULTS: Microbial communities cultured in vitro in BL media were observed to have enhanced growth of lactic acid bacteria, such as lactobacilli, with a corresponding increase in the levels of lactate and short-chain fatty acids. Members of Escherichia isolated from these faecal communities were found to bioconvert glucosinolates and SMCSO to their reduced analogues. CONCLUSION: This study uses a broccoli leachate to investigate the bacterial-mediated bioconversion of glucosinolates and SMCSO, which may lead to further products with additional health benefits to the host. We believe that this is the first study that shows the reduction of the dietary compound S-methylcysteine sulphoxide by bacteria isolated from human faeces.

Identification of Genes Required for Glucan Exopolysaccharide Production in Lactobacillus johnsonii Suggests a Novel Biosynthesis Mechanism.

Lactobacillus johnsonii FI9785 makes two capsular exopolysaccharides-a heteropolysaccharide (EPS2) encoded by the eps operon and a branched glucan homopolysaccharide (EPS1). The homopolysaccharide is synthesized in the absence of sucrose, and there are no typical glucansucrase genes in the genome. Quantitative proteomics was used to compare the wild type to a mutant where EPS production was reduced to attempt to identify proteins associated with EPS1 biosynthesis. A putative bactoprenol glycosyltransferase, FI9785_242 (242), was less abundant in the Δeps_cluster mutant strain than in the wild type. Nuclear magnetic resonance (NMR) analysis of isolated EPS showed that deletion of the FI9785_242 gene (242) prevented the accumulation of EPS1, without affecting EPS2 synthesis, while plasmid complementation restored EPS1 production. The deletion of 242 also produced a slow-growth phenotype, which could be rescued by complementation. 242 shows amino acid homology to bactoprenol glycosyltransferase GtrB, involved in O-antigen glycosylation, while in silico analysis of the neighboring gene 241 suggested that it encodes a putative flippase with homology to the GtrA superfamily. Deletion of 241 also prevented production of EPS1 and again caused a slow-growth phenotype, while plasmid complementation reinstated EPS1 synthesis. Both genes are highly conserved in L. johnsonii strains isolated from different environments. These results suggest that there may be a novel mechanism for homopolysaccharide synthesis in the Gram-positive L. johnsoniiIMPORTANCE Exopolysaccharides are key components of the surfaces of their bacterial producers, contributing to protection, microbial and host interactions, and even virulence. They also have significant applications in industry, and understanding their biosynthetic mechanisms may allow improved production of novel and valuable polymers. Four categories of bacterial exopolysaccharide biosynthesis have been described in detail, but novel enzymes and glycosylation mechanisms are still being described. Our findings that a putative bactoprenol glycosyltransferase and flippase are essential to homopolysaccharide biosynthesis in Lactobacillus johnsonii FI9785 indicate that there may be an alternative mechanism of glucan biosynthesis to the glucansucrase pathway. Disturbance of this synthesis leads to a slow-growth phenotype. Further elucidation of this biosynthesis may give insight into exopolysaccharide production and its impact on the bacterial cell.

Fine-Tuning of Sirtuin 1 Expression Is Essential to Protect the Liver From Cholestatic Liver Disease.

Cholestasis comprises aetiologically heterogeneous conditions characterized by accumulation of bile acids in the liver that actively contribute to liver damage. Sirtuin 1 (SIRT1) regulates liver regeneration and bile acid metabolism by modulating farnesoid X receptor (FXR); we here investigate its role in cholestatic liver disease. We determined SIRT1 expression in livers from patients with cholestatic disease, in two experimental models of cholestasis, as well as in human and murine liver cells in response to bile acid loading. SIRT1-overexpressing (SIRToe ) and hepatocyte-specific SIRT1-KO (knockout) mice (SIRThep-/- ) were subjected to bile duct ligation (BDL) and were fed with a 0.1% DDC (3,5-diethoxycarbonyl-1,4-dihydrocollidine) diet to determine the biological relevance of SIRT1 during cholestasis. The effect of NorUDCA (24-norursodeoxycholic acid) was tested in BDL/SIRToe mice. We found that SIRT1 was highly expressed in livers from cholestatic patients, mice after BDL, and Mdr2 knockout mice (Mdr2-/- ) animals. The detrimental effects of SIRT1 during cholestasis were validated in vivo and in vitro. SIRToe mice showed exacerbated parenchymal injury whereas SIRThep-/- mice evidenced a moderate improvement after BDL and 0.1% DDC feeding. Likewise, hepatocytes isolated from SIRToe mice showed increased apoptosis in response to bile acids, whereas a significant reduction was observed in SIRThep-/- hepatocytes. Importantly, the decrease, but not complete inhibition, of SIRT1 exerted by norUDCA treatment correlated with pronounced improvement in liver parenchyma in BDL/SIRToe mice. Interestingly, both SIRT1 overexpression and hepatocyte-specific SIRT1 depletion correlated with inhibition of FXR, whereas modulation of SIRT1 by NorUDCA associated with restored FXR signaling. Conclusion: SIRT1 expression is increased during human and murine cholestasis. Fine-tuning expression of SIRT1 is essential to protect the liver from cholestatic liver damage.

APOE genotype influences the gut microbiome structure and function in humans and mice: relevance for Alzheimer's disease pathophysiology.

Apolipoprotein E (APOE) genotype is the strongest prevalent genetic risk factor for Alzheimer's disease (AD). Numerous studies have provided insights into the pathologic mechanisms. However, a comprehensive understanding of the impact of APOE genotype on microflora speciation and metabolism is completely lacking. In this study, we investigated the association between APOE genotype and the gut microbiome composition in human and APOE-targeted replacement (TR) transgenic mice. Fecal microbiota amplicon sequencing from matched individuals with different APOE genotypes revealed no significant differences in overall microbiota diversity in group-aggregated human APOE genotypes. However, several bacterial taxa showed significantly different relative abundance between APOE genotypes. Notably, we detected an association of Prevotellaceae and Ruminococcaceae and several butyrate-producing genera abundances with APOE genotypes. These findings were confirmed by comparing the gut microbiota of APOE-TR mice. Furthermore, metabolomic analysis of murine fecal water detected significant differences in microbe-associated amino acids and short-chain fatty acids between APOE genotypes. Together, these findings indicate that APOE genotype is associated with specific gut microbiome profiles in both humans and APOE-TR mice. This suggests that the gut microbiome is worth further investigation as a potential target to mitigate the deleterious impact of the APOE4 allele on cognitive decline and the prevention of AD.-Tran, T. T. T., Corsini, S., Kellingray, L., Hegarty, C., Le Gall, G., Narbad, A., Müller, M., Tejera, N., O'Toole, P. W., Minihane, A.-M., Vauzour, D. APOE genotype influences the gut microbiome structure and function in humans and mice: relevance for Alzheimer's disease pathophysiology.

Serine-rich repeat protein adhesins from Lactobacillus reuteri display strain specific glycosylation profiles.

Lactobacillus reuteri is a gut symbiont inhabiting the gastrointestinal tract of numerous vertebrates. The surface-exposed serine-rich repeat protein (SRRP) is a major adhesin in Gram-positive bacteria. Using lectin and sugar nucleotide profiling of wild-type or L. reuteri isogenic mutants, MALDI-ToF-MS, LC-MS and GC-MS analyses of SRRPs, we showed that L. reuteri strains 100-23C (from rodent) and ATCC 53608 (from pig) can perform protein O-glycosylation and modify SRRP100-23 and SRRP53608 with Hex-Glc-GlcNAc and di-GlcNAc moieties, respectively. Furthermore, in vivo glycoengineering in E. coli led to glycosylation of SRRP53608 variants with α-GlcNAc and GlcNAcß(1→6)GlcNAcα moieties. The glycosyltransferases involved in the modification of these adhesins were identified within the SecA2/Y2 accessory secretion system and their sugar nucleotide preference determined by saturation transfer difference NMR spectroscopy and differential scanning fluorimetry. Together, these findings provide novel insights into the cellular O-protein glycosylation pathways of gut commensal bacteria and potential routes for glycoengineering applications.

Acute Consumption of Flavan-3-ol-Enriched Dark Chocolate Affects Human Endogenous Metabolism.

Flavan-3-ols and methylxanthines have potential beneficial effects on human health including reducing cardiovascular risk. We performed a randomized controlled crossover intervention trial to assess the acute effects of consumption of flavan-3-ol-enriched dark chocolate, compared with standard dark chocolate and white chocolate, on the human metabolome. We assessed the metabolome in urine and blood plasma samples collected before and at 2 and 6 h after consumption of chocolates in 42 healthy volunteers using a nontargeted metabolomics approach. Plasma samples were assessed and showed differentiation between time points with no further separation among the three chocolate treatments. Multivariate statistics applied to urine samples could readily separate the postprandial time points and distinguish between the treatments. Most of the markers responsible for the multivariate discrimination between the chocolates were of dietary origin. Interestingly, small but significant level changes were also observed for a subset of endogenous metabolites. 1H NMR revealed that flavan-3-ol-enriched dark chocolate and standard dark chocolate reduced urinary levels of creatinine, lactate, some amino acids, and related degradation products and increased the levels of pyruvate and 4-hydroxyphenylacetate, a phenolic compound of bacterial origin. This study demonstrates that an acute chocolate intervention can significantly affect human metabolism.

Structure and biosynthesis of two exopolysaccharides produced by Lactobacillus johnsonii FI9785.

Exopolysaccharides were isolated and purified from Lactobacillus johnsonii FI9785, which has previously been shown to act as a competitive exclusion agent to control Clostridium perfringens in poultry. Structural analysis by NMR spectroscopy revealed that L. johnsonii FI9785 can produce two types of exopolysaccharide: EPS-1 is a branched dextran with the unusual feature that every backbone residue is substituted with a 2-linked glucose unit, and EPS-2 was shown to have a repeating unit with the following structure: -6)-α-Glcp-(1-3)-ß-Glcp-(1-5)-ß-Galf-(1-6)-α-Glcp-(1-4)-ß-Galp-(1-4)-ß-Glcp-(1-. Sites on both polysaccharides were partially occupied by substituent groups: 1-phosphoglycerol and O-acetyl groups in EPS-1 and a single O-acetyl group in EPS-2. Analysis of a deletion mutant (ΔepsE) lacking the putative priming glycosyltransferase gene located within a predicted eps gene cluster revealed that the mutant could produce EPS-1 but not EPS-2, indicating that epsE is essential for the biosynthesis of EPS-2. Atomic force microscopy confirmed the localization of galactose residues on the exterior of wild type cells and their absence in the ΔepsE mutant. EPS2 was found to adopt a random coil structural conformation. Deletion of the entire 14-kb eps cluster resulted in an acapsular mutant phenotype that was not able to produce either EPS-2 or EPS-1. Alterations in the cell surface properties of the EPS-specific mutants were demonstrated by differences in binding of an anti-wild type L. johnsonii antibody. These findings provide insights into the biosynthesis and structures of novel exopolysaccharides produced by L. johnsonii FI9785, which are likely to play an important role in biofilm formation, protection against harsh environment of the gut, and colonization of the host.

Early adaptation to oxygen is key to the industrially important traits of Lactococcus lactis ssp. cremoris during milk fermentation.

BACKGROUND: Lactococcus lactis is the most used species in the dairy industry. Its ability to adapt to technological stresses, such as oxidative stress encountered during stirring in the first stages of the cheese-making process, is a key factor to measure its technological performance. This study aimed to understand the response to oxidative stress of Lactococcus lactis subsp. cremoris MG1363 at the transcriptional and metabolic levels in relation to acidification kinetics and growth conditions, especially at an early stage of growth. For those purposes, conditions of hyper-oxygenation were initially fixed for the fermentation. RESULTS: Kinetics of growth and acidification were not affected by the presence of oxygen, indicating a high resistance to oxygen of the L. lactis MG1363 strain. Its resistance was explained by an efficient consumption of oxygen within the first 4 hours of culture, leading to a drop of the redox potential. The efficient consumption of oxygen by the L. lactis MG1363 strain was supported by a coherent and early adaptation to oxygen after 1 hour of culture at both gene expression and metabolic levels. In oxygen metabolism, the over-expression of all the genes of the nrd (ribonucleotide reductases) operon or fhu (ferrichrome ABC transports) genes was particularly significant. In carbon metabolism, the presence of oxygen led to an early shift at the gene level in the pyruvate pathway towards the acetate/2,3-butanediol pathway confirmed by the kinetics of metabolite production. Finally, the MG1363 strain was no longer able to consume oxygen in the stationary growth phase, leading to a drastic loss of culturability as a consequence of cumulative stresses and the absence of gene adaptation at this stage. CONCLUSIONS: Combining metabolic and transcriptomic profiling, together with oxygen consumption kinetics, yielded new insights into the whole genome adaptation of L. lactis to initial oxidative stress. An early and transitional adaptation to oxidative stress was revealed for L. lactis subsp. cremoris MG1363 in the presence of initially high levels of oxygen. This enables the cells to maintain key traits that are of great importance for industry, such as rapid acidification and reduction of the redox potential of the growth media.

A novel Mediterranean diet-inspired supplement ameliorates cognitive, microbial, and metabolic deficits in a mouse model of low-grade inflammation.

The Mediterranean diet (MD) and its bioactive constituents have been advocated for their neuroprotective properties along with their capacity to affect gut microbiota speciation and metabolism. Mediated through the gut brain axis, this modulation of the microbiota may partly contribute to the neuroprotective properties of the MD. To explore this potential interaction, we evaluated the neuroprotective properties of a novel bioactive blend (Neurosyn240) resembling the Mediterranean diet in a rodent model of chronic low-grade inflammation. Behavioral tests of cognition, brain proteomic analysis, 16S rRNA sequencing, and 1H NMR metabolomic analyses were employed to develop an understanding of the gut-brain axis interactions involved. Recognition memory, as assessed by the novel object recognition task (NOR), decreased in response to LPS insult and was restored with Neurosyn240 supplementation. Although the open field task performance did not reach significance, it correlated with NOR performance indicating an element of anxiety related to this cognitive change. Behavioral changes associated with Neurosyn240 were accompanied by a shift in the microbiota composition which included the restoration of the Firmicutes: Bacteroidota ratio and an increase in Muribaculum, Rikenellaceae Alloprevotella, and most notably Akkermansia which significantly correlated with NOR performance. Akkermansia also correlated with the metabolites 5-aminovalerate, threonine, valine, uridine monophosphate, and adenosine monophosphate, which in turn significantly correlated with NOR performance. The proteomic profile within the brain was dramatically influenced by both interventions, with KEGG analysis highlighting oxidative phosphorylation and neurodegenerative disease-related pathways to be modulated. Intriguingly, a subset of these proteomic changes simultaneously correlated with Akkermansia abundance and predominantly related to oxidative phosphorylation, perhaps alluding to a protective gut-brain axis interaction. Collectively, our results suggest that the bioactive blend Neurosyn240 conferred cognitive and microbiota resilience in response to the deleterious effects of low-grade inflammation.

Exploring the sequence-function space of microbial fucosidases.

Microbial α-L-fucosidases catalyse the hydrolysis of terminal α-L-fucosidic linkages and can perform transglycosylation reactions. Based on sequence identity, α-L-fucosidases are classified in glycoside hydrolases (GHs) families of the carbohydrate-active enzyme database. Here we explored the sequence-function space of GH29 fucosidases. Based on sequence similarity network (SSN) analyses, 15 GH29 α-L-fucosidases were selected for functional characterisation. HPAEC-PAD and LC-FD-MS/MS analyses revealed substrate and linkage specificities for α1,2, α1,3, α1,4 and α1,6 linked fucosylated oligosaccharides and glycoconjugates, consistent with their SSN clustering. The structural basis for the substrate specificity of GH29 fucosidase from Bifidobacterium asteroides towards α1,6 linkages and FA2G2 N-glycan was determined by X-ray crystallography and STD NMR. The capacity of GH29 fucosidases to carry out transfucosylation reactions with GlcNAc and 3FN as acceptors was evaluated by TLC combined with ESI-MS and NMR. These experimental data supported the use of SSN to further explore the GH29 sequence-function space through machine-learning models. Our lightweight protein language models could accurately allocate test sequences in their respective SSN clusters and assign 34,258 non-redundant GH29 sequences into SSN clusters. It is expected that the combination of these computational approaches will be used in the future for the identification of novel GHs with desired specificities.

Maternal and cord blood LC-HRMS metabolomics reveal alterations in energy and polyamine metabolism, and oxidative stress in very-low birth weight infants.

To assess the global effect of preterm birth on fetal metabolism and maternal-fetal nutrient transfer, we used a mass spectrometric-based chemical phenotyping approach on cord blood obtained at the time of birth. We sampled umbilical venous, umbilical arterial, and maternal blood from mothers delivering very-low birth weight (VLBW, with a median gestational age and weight of 29 weeks, and 1210 g, respectively) premature or full-term (FT) neonates. In VLBW group, we observed a significant elevation in the levels and maternal-fetal gradients of butyryl-, isovaleryl-, hexanoyl- and octanoyl-carnitines, suggesting enhanced short- and medium chain fatty acid ß-oxidation in human preterm feto-placental unit. The significant decrease in glutamine-glutamate in preterm arterial cord blood beside lower levels of amino acid precursors of Krebs cycle suggest increased glutamine utilization in the fast growing tissues of preterm fetus with a deregulation in placental glutamate-glutamine shuttling. Enhanced glutathione utilization is likely to account for the decrease in precursor amino acids (serine, betaine, glutamate and methionine) in arterial cord blood. An increase in both the circulating levels and maternal-fetal gradients of several polyamines in their acetylated form (diacetylspermine and acetylputrescine) suggests an enhanced polyamine metabolic cycling in extreme prematurity. Our metabolomics study allowed the identification of alterations in fetal energy, antioxidant defense, and polyamines and purines flux as a signature of premature birth.

Selenium-dependent biogenesis of formate dehydrogenase in Campylobacter jejuni is controlled by the fdhTU accessory genes.

The food-borne bacterial pathogen Campylobacter jejuni efficiently utilizes organic acids such as lactate and formate for energy production. Formate is rapidly metabolized via the activity of the multisubunit formate dehydrogenase (FDH) enzyme, of which the FdhA subunit is predicted to contain a selenocysteine (SeC) amino acid. In this study we investigated the function of the cj1500 and cj1501 genes of C. jejuni, demonstrate that they are involved in selenium-controlled production of FDH, and propose the names fdhT and fdhU, respectively. Insertional inactivation of fdhT or fdhU in C. jejuni resulted in the absence of FdhA and FdhB protein expression, reduced fdhABC RNA levels, the absence of FDH enzyme activity, and the lack of formate utilization, as assessed by (1)H nuclear magnetic resonance. The fdhABC genes are transcribed from a single promoter located two genes upstream of fdhA, and the decrease in fdhABC RNA levels in the fdhU mutant is mediated at the posttranscriptional level. FDH activity and the ability to utilize formate were restored by genetic complementation with fdhU and by supplementation of the growth media with selenium dioxide. Disruption of SeC synthesis by inactivation of the selA and selB genes also resulted in the absence of FDH activity, which could not be restored by selenium supplementation. Comparative genomic analysis suggests a link between the presence of selA and fdhTU orthologs and the predicted presence of SeC in FdhA. The fdhTU genes encode accessory proteins required for FDH expression and activity in C. jejuni, possibly by contributing to acquisition or utilization of selenium.

Using a multi-omic approach to investigate the mechanism of 12-bis-THA activity against Burkholderia thailandensis.

Burkholderia pseudomallei is the causative agent of the tropical disease, melioidosis. It is intrinsically resistant to many antimicrobials and treatment requires an onerous regimen of intravenous and orally administered drugs. Relapse of disease and high rates of mortality following treatment are common, demonstrating the need for new anti-Burkholderia agents. The cationic bola-amphiphile, 12,12'-(dodecane-1,12-diyl) bis (9-amino-1,2,3,4-tetrahydroacridinium), referred to as 12-bis-THA, is a molecule with the potential to treat Burkholderia infections. 12-bis-THA spontaneously forms cationic nanoparticles that bind anionic phospholipids in the prokaryotic membrane and are readily internalized. In this study, we examine the antimicrobial activity of 12-bis-THA against strains of Burkholderia thailandensis. As B. pseudomallei produces a polysaccharide capsule we first examined if this extra barrier influenced the activity of 12-bis-THA which is known to act on the bacterial envelope. Therefore two strains of B. thailandensis were selected for further testing, strain E264 which does not produce a capsule and strain E555 which does produce a capsule that is chemically similar to that found in B. pseudomallei. In this study no difference in the minimum inhibitory concentration (MIC) was observed when capsulated (E555) and unencapsulated (E264) strains of B. thailandensis were compared, however time-kill analysis showed that the unencapsulated strain was more susceptible to 12-bis-THA. The presence of the capsule did not affect the membrane permeation of 12-bis-THA at MIC concentrations. Proteomic and metabolomic analyses showed that 12-bis-THA causes a shift in central metabolism away from glycolysis and glyoxylate cycle, and suppressed the production of the F1 domain of ATP synthase. In summary, we provide insight into the molecular mechanisms underpinning the activity of 12-bis-THA against B. thailandensis and discuss its potential for further development.

Microbial-derived metabolites as a risk factor of age-related cognitive decline and dementia.

A consequence of our progressively ageing global population is the increasing prevalence of worldwide age-related cognitive decline and dementia. In the absence of effective therapeutic interventions, identifying risk factors associated with cognitive decline becomes increasingly vital. Novel perspectives suggest that a dynamic bidirectional communication system between the gut, its microbiome, and the central nervous system, commonly referred to as the microbiota-gut-brain axis, may be a contributing factor for cognitive health and disease. However, the exact mechanisms remain undefined. Microbial-derived metabolites produced in the gut can cross the intestinal epithelial barrier, enter systemic circulation and trigger physiological responses both directly and indirectly affecting the central nervous system and its functions. Dysregulation of this system (i.e., dysbiosis) can modulate cytotoxic metabolite production, promote neuroinflammation and negatively impact cognition. In this review, we explore critical connections between microbial-derived metabolites (secondary bile acids, trimethylamine-N-oxide (TMAO), tryptophan derivatives and others) and their influence upon cognitive function and neurodegenerative disorders, with a particular interest in their less-explored role as risk factors of cognitive decline.

(-)-Epicatechin mitigates anxiety-related behavior in a mouse model of high fat diet-induced obesity.

Mounting evidence demonstrates that consumption of high fat diet (HFD) and subsequent development of obesity leads to alterations in cognition and mood. While obesity can affect brain function, consumption of select dietary bioactives may help prevent obesity-related cognitive decline. This study investigated the capacity of the dietary flavonoid (-)-epicatechin (EC) to mitigate HFD-induced obesity-associated alterations in memory and mood. Healthy 8-week old male C57BL/6J mice were maintained on either a control diet (10 kCal% from fat) or a HFD (45 kCal% from fat) and were supplemented with EC at 2 or 20 mg/kg body weight (B.W.) for a 24 week period. Between week 20 and 22, anxiety-related behavior, recognition memory, and spatial memory were measured. Underlying mechanisms were assessed by measuring the expression of selected genes in the hippocampus and by 16S rRNA sequencing and metabolomic analysis of the gut microbiota. 24 weeks of HFD feeding resulted in obesity, which was not affected by EC supplementation. HFD-associated increase in anxiety-related behavior was mitigated by EC in a dose-response manner and was accompanied by increased hippocampal brain-derived neurotrophic factor (BDNF), as well as partial or full restoration of glucocorticoid receptor, mineralocorticoid receptor and 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) expression. Higher EC dosage (20 mg/kg B.W.) also restored aberrant Lactobacillus and Enterobacter abundance altered by HFD and/or the associated obesity. Together, these results demonstrate how EC mitigates anxiety-related behaviors, revealing a connection between BDNF- and glucocorticoids-mediated signaling. Our findings link changes in the hippocampus and the gut microbiota in a context of HFD-induced obesity and anxiety.

Fecal microbiota transfer between young and aged mice reverses hallmarks of the aging gut, eye, and brain.

BACKGROUND: Altered intestinal microbiota composition in later life is associated with inflammaging, declining tissue function, and increased susceptibility to age-associated chronic diseases, including neurodegenerative dementias. Here, we tested the hypothesis that manipulating the intestinal microbiota influences the development of major comorbidities associated with aging and, in particular, inflammation affecting the brain and retina. METHODS: Using fecal microbiota transplantation, we exchanged the intestinal microbiota of young (3 months), old (18 months), and aged (24 months) mice. Whole metagenomic shotgun sequencing and metabolomics were used to develop a custom analysis workflow, to analyze the changes in gut microbiota composition and metabolic potential. Effects of age and microbiota transfer on the gut barrier, retina, and brain were assessed using protein assays, immunohistology, and behavioral testing. RESULTS: We show that microbiota composition profiles and key species enriched in young or aged mice are successfully transferred by FMT between young and aged mice and that FMT modulates resulting metabolic pathway profiles. The transfer of aged donor microbiota into young mice accelerates age-associated central nervous system (CNS) inflammation, retinal inflammation, and cytokine signaling and promotes loss of key functional protein in the eye, effects which are coincident with increased intestinal barrier permeability. Conversely, these detrimental effects can be reversed by the transfer of young donor microbiota. CONCLUSIONS: These findings demonstrate that the aging gut microbiota drives detrimental changes in the gut-brain and gut-retina axes suggesting that microbial modulation may be of therapeutic benefit in preventing inflammation-related tissue decline in later life. Video abstract.