Biology subjects

Sidebottom, A. M.

Publications and source records attributed to Sidebottom, A. M..

At least 19 recordsLinked to original sources

bioRxiv2026

SpiE is a distinct gut microbial 4-cholesten-3-one 5β-reductase that acts with SpiR to convert cholesterol to coprostanol

Gut microbes convert cholesterol into coprostanol, a poorly absorbed sterol that is largely excreted in feces and is linked to lower serum cholesterol levels. However, the enzymes responsible for this process remain poorly characterized. The second step of the pathway, cholestenone reduction into coprostanone, has long been established, yet the enzyme(s) responsible were unknown. Here, we use comparative genomics to identify SpiE, a cholestenone (4-cholesten-3-one) reductase that is part of the steroid processing (spi) pathway with SpiR. We confirm that SpiE binds to and reduces both cholestenone and progesterone into 5{beta}-reduced forms. SpiR and SpiE together convert cholesterol into coprostanol, indicating that both enzymes are sufficient for coprostanol formation. SpiE belongs to the short-chain dehydrogenase (SDR) family and represents an evolutionarily distinct lineage of SDR ene-reductases. Metagenomic analyses reveal that spiE relative abundance is strongly correlated with stool coprostanol (Spearman's {rho}=0.81, p=5.99e-65). In longitudinal patient cohorts, antibiotic treatment coincided with loss of SpiE and coprostanol production. Post antibiotics, the emergence of distinct SpiE-encoding genera rescued coprostanol production in some individuals. Our findings provide the biochemical basis of the cholesterol to coprostanol pathway and lay the groundwork for future studies linking it to host physiology.

microbiology↗

bioRxiv2026

Epithelial-Intrinsic Alterations and Maladaptation to Luminal Metabolites Underlie Persistent Crohn's Disease Pathogenesis

In patients with Crohns disease, the noninflamed intestinal epithelium exhibits inflammation-like transcriptional signatures that persist even during clinical remission. However, it is unclear whether this signature is due to sustained environmental triggers or to the epitheliums intrinsic immunological adaptions. To disentangle environmental and cell-intrinsic effects on disease signatures, we used noninflamed intestinal tissue biopsies from patients with active and inactive Crohns disease, as well as control donors, to generate donor-matched organoids enriched for intestinal epithelial cells. We collected single-cell RNA-seq and single-cell ATAC-seq data from intestinal tissues, and from their matched organoids before and after stimulation with luminal metabolites of patients. This approach allowed us to distinguish between the effects of extrinsic factors and the intrinsic alterations that persist when cells are removed from their local environment. We found that organoid cells retained epithelial-autonomous disease phenotypes, even when derived from patients in remission, whereas signatures from tissue cells showed evidence of immune- and microbial-epithelial crosstalk. In contrast to healthy control organoids, organoids from Crohns disease patients exhibited selective activation of disease-associated transcriptional changes and environment-responsive inflammatory chromatin remodeling after exposure to patient-derived luminal metabolites. Overall, our study suggests that disease-associated epithelial states are shaped by both cell-intrinsic dysregulation and microenvironmental cues.

immunology↗

bioRxiv2026

Home is Where the Heterogeneity Is: Housing Facility-level Differences in the Gut Microbiome and Metabolic Phenotype Confound Arsenic Effects on Glucose Homeostasis in Male Mice

Inorganic arsenic (iAs) exposure is linked to impaired glucose homeostasis and type 2 diabetes, yet the magnitude and direction of reported effects vary substantially across studies and populations. The gut microbiome is both a target and a mediator of arsenic toxicity, suggesting that pre-exposure community composition may modulate the development of metabolic dysfunction. To test this, we conducted parallel 50 ppm iAs drinking-water exposures in male C57BL/6J mice at two animal facilities. Results were compared across facilities for metabolic phenotypes, hepatic arsenic levels, targeted and untargeted metabolomics, and shotgun metagenomics. Hepatic arsenic confirmed comparable exposure at both sites; however, the housing facility explained more variance than the iAs treatment group across every data layer. Baseline microbial communities and metabolic phenotypes at each institution differed, and this difference propagated into the iAs treatment effect. Critically, iAs exposure impaired glucose clearance at one site while trending toward improvement at the other. Facility explained 19 to 26% of variance in microbiome, bile acid, polar, and untargeted metabolite ordinations, while iAs treatment did not reach significance. A random forest classifier identified the facility with 96% cross-validated accuracy from 22 microbial species, whereas treatment classification did not exceed 67% accuracy. Functional metagenomic analyses revealed nearly 11,733 (63%) of genes were differentially abundant between facilities compared 139 with iAs treatment. Our results indicate that identical genetics and exposure may produce differential metabolic outcomes on different microbial backgrounds. Characterizing the baseline microbiome and metabolome is therefore critical both for identifying which individuals are most susceptible to the metabolic effects of arsenic exposure and for potentially reducing the risk of exposure through modulation of the gut microbiome.

microbiology↗

bioRxiv2026

Distribution of luxS and production of autoinducer-2 among gut Bacteroidales

One of the best studied quorum sensing (QS) molecules, autoinducer-2 (AI-2), regulates processes in numerous bacteria. LuxS is an enzyme of the activated methyl cycle which, along with MtnN, converts S-adenosyl-homocysteine to homocysteine, releasing DPD (4,5-dihydroxy-2,3-pentanedione) which is spontaneously converted to AI-2. Many bacteria do not encode MtnN-LuxS and instead encode SahH, which directly converts S-adenosyl-homocysteine to homocysteine without AI-2 production. The genomes of some gut Bacteroidales were shown to contain luxS, however, these reports, as well as reports of the production of AI-2 by gut Bactereoidales have been inconsistent. We performed a comprehensive analysis of the distribution of luxS and sahH in Bacteroidota with an in-depth exploration of gut Bacteroidales. The data suggest that the ancestral Bacteroidota contained sahH, with numerous independent replacements with mtnN-luxS during diversification. In Bacteroidaceae, Parabacteroides, and many Prevotellaceae, mtnN-luxS or sahH are present in the same genetic region, adjacent to yfhO. Using Bacteroides fragilis, which contains sahH, and Bacteroides uniformis and Phocaeicola vulgatus, whose genomes contain mtnN-luxS, we show that luxS-containing strains produce AI-2. Transcriptomic analyses and gnotobiotic mouse experiments using wild-type strains and mtnN-luxS and sahH genetic swaps showed that Bacteroidaceae species do not respond to AI-2 under the conditions tested. However, analyses of 15 human gut metagenomic datasets show that mtnN-luxS containing Bacteroidales species average between 5.1 - 33% of the total bacteria in these various human gut microbiome datasets, and therefore, likely produce substantial amounts of AI-2 sensed by responsive gut symbionts and pathogens. ImportanceHere, we show that gut Bacteroidales possess one of two pathways to complete the activated methyl cycle, one that produces the QS molecule AI-2. We clarify conflicting data regarding AI-2 production in gut Bacteroidales and show that luxS is present in many gut Bacteroidales species, but lacking in species such as Bacteroides thetaiotaomicron and Bacteroides fragilis. We find that prevalent and abundant human gut Bacteroidales species including Bacteroides uniformis and Phocaeicola vulgatus produce substantial amounts of AI-2. While our data do not show that Bacteroidales sense or respond to AI-2 under the conditions tested, analyses of human gut metagenomic data reveal that AI-2 producing Bacteroidales comprise a large proportion of the gut bacteria of both industrialized and non-industrialized human populations.

microbiology↗

bioRxiv2026

Microbiome histidine competition mediates dietary control of systemic imidazole propionate

The gut microbiome produces numerous metabolites that influence mammalian health. While microbiome composition and diet influence metabolite concentrations, how these factors interact remains incompletely defined. Here we find production of imidazole propionate (ImP), a microbial metabolite associated with cardiometabolic and neurodegenerative diseases, is determined by the balance of competing metabolic pathways that catabolize histidine to ImP or short-chain fatty acids (SCFAs). We show glutamate serves as a preferred substrate that selectively inhibits histidine conversion to SCFAs, redirecting flux to increased ImP production across mouse- and human-derived microbial communities. We find dietary monosodium glutamate (MSG) acting via this mechanism boosts ImP production in the mouse gut, transiently impairing glucose tolerance and increasing systemic ImP. These findings show that predictable interactions between dietary substrate and microbial competition control systemic ImP levels, providing a mechanistic framework for understanding microbiome metabolite production more broadly.

microbiology↗

bioRxiv2026

Gut Microbiota-derived Acetate Safeguards the Colonic Epithelial Acetyl-CoA Reserve to Avert Colonic Senescence

Aging is a major driver of tissue senescence, but little is known about the development of age-unrelated tissue senescence. Here we show that nucleocytosolic acetyl-CoA deficiency in colon epithelial cells, caused by Acly ablation and bacterial depletion, triggers age-independent, p53-dependent colonic senescence leading to severe systemic inflammation. Acetate supplementation, acetate-producing bacterial transplantation, targeted depletion of senescent cells or treatment with lysine deacetylase inhibitors blocks colonic senescence and inflammatory injury. Spontaneous colonic senescence develops following simultaneous deletion of epithelial Acly and Acss2, confirming that microbe-derived acetate maintains the epithelial acetyl-CoA pool via ACSS2 to avert colonic senescence. Mechanistically, acetyl-CoA deficiency deprives a cohort of mitochondrial and nuclear proteins of acetylation, leading to increased oxidative stress and DNA repair stress that trigger cellular senescence. Among these proteins, ATP5F1A-K161 acetylation and H4-K5/8 acetylation are required to protect colonic epithelial cells from developing senescence. These observations unveil a previously unknown mechanism that governs colonic senescence.

physiology↗

bioRxiv2025

Intracellular acidification by bacteria-derived valeric acid is a mechanism of trans-kingdom ecology against Candida parapsilosis colonization

In hematopoietic cell transplant patients, intestinal Candida parapsilosis expansion and translocation cause life-threatening candidemia, yet how commensal intestinal bacteria prevent Candida expansion remains incompletely defined. Here, we trained a machine learning model on supernatant metabolomic profiles of commensal bacteria to identify bacteria-derived inhibitors of fungal growth, with valeric and butyric acid as top hits. Experimental validation confirmed in silico predictions in three systems. First, in patient fecal samples, valeric and butyric acid levels inversely correlated with C. parapsilosis growth. Second, in vitro, valeric acid potently inhibited C. parapsilosis growth by causing intracellular acidification. Third, administration of glycerol valerate, and free or microencapsulated valeric acid blunted C. parapsilosis growth at murine intestinal sites where valeric acid could be detected. Thus, machine learning could identify a mechanistic driver of trans-kingdom ecology limiting C. parapsilosis intestinal expansion and may inform strategies to reduce patient risk of developing candidiasis during high-risk periods.

microbiology↗

bioRxiv2025

Gut microbes mediate the synergistic effects of dietary cholesterol and saturated fat in driving fibrosing MASH

Metabolic dysfunction-associated steatotic liver disease (MASLD) affects approximately one-third of the global population and can progress to metabolic dysfunction-associated steatohepatitis (MASH) with fibrosis, increasing risk of cirrhosis, hepatocellular carcinoma, and mortality. Gut microbes driven by diets high in saturated fat, simple sugar, and cholesterol contribute to disease progression, yet underlying mechanisms remain undefined. We explored the independent and synergistic effects of dietary saturated fat and cholesterol on MASH development using specific pathogen-free (SPF) and germ-free (GF) mice. We demonstrate that 1) both dietary cholesterol and saturated fat are required to induce fibrosing MASH in SPF mice, whereas GF mice are protected, 2) saturated fat and cholesterol individually alter gut microbial membership, potentially via altered bile acid metabolism, while their combination promotes a distinct composition, including an increase in Parasutterella spp. which correlates with hepatic fibrosis, and 3) diluted cecal contents from SPF, but not GF, mice fed high-fat, high-cholesterol diets are enriched in deoxycholic acid and activate human hepatic stellate cells in vitro, suggesting a mechanistic link between dietary lipid-induced microbiota and liver fibrogenesis. These findings reveal how specific Western dietary components shape the gut microbiota and contribute to hepatic liver fibrosis via stellate activation, offering potential targets for therapeutic intervention in MASLD/MASH.

physiology↗

bioRxiv2025

Lantibiotic-producing bacteria impact microbiome resilience and colonization resistance

A subset of commensal bacterial strains secrete bacteriocins, such as lantibiotics, to establish and protect their niche in the gut. Because the antimicrobial spectrum of lantibiotics includes opportunistic pathogens, such as vancomycin-resistant Enterococcus faecium (VRE), they may provide an approach to reduce antibiotic-resistant infections. The impact of lantibiotic-producing bacteria on the complex microbial populations constituting the microbiome, however, remains poorly defined. We find that genes encoding lanthipeptides, including lantibiotics, are commonly present in the microbiomes of healthy humans and in dysbiotic microbiomes of hospitalized patients. In fecal samples collected from hospitalized patients, bacterial species encoding lantibiotic genes are present in greater abundance than lantibiotic-deficient strains of the same species. We demonstrate that the lantibiotic-producing bacterium, Blautia pseudococcoides SCSK, prevents intestinal recolonization of mice by a wide range of commensal species following antibiotic-induced dysbiosis and markedly reduces fecal concentrations of microbiota-derived metabolites associated with mucosal immune defenses. Lantibiotic-mediated dysbiosis results in sustained loss of colonization resistance against Klebsiella pneumoniae and Clostrioides difficile infection. Our findings reveal the potential impact of lantibiotic-producing bacterial species on microbiome resilience and susceptibility to infection following antibiotic treatment.

microbiology↗

bioRxiv2025

DNA-utilization loci enable exogenous DNA metabolism in gut Bacteroidales

The human gut microbiome plays a central role in nutrient metabolism, yet the fate of exogenous nucleic acids within this ecosystem remains poorly understood. Here, we show that multiple Bacteroidales species efficiently metabolize exogenous DNA, with Bacteroides thetaiotaomicron converting it into the deaminated nucleobases uracil and xanthine. Using genetic and biochemical approaches, we identify ddbABCDEF, a six-gene locus encoding secreted nucleases and an outer membrane transporter, essential for exogenous DNA metabolism in B. thetaiotaomicron. Colonization of gnotobiotic mice with ddbABCDEF mutants reveals that this pathway significantly alters nucleobase pools in the gut. Comparative genomics demonstrate that ddbABCDEF is evolutionarily related to a natural transformation system present in Bacteroidota and has diversified into four distinct subtypes, each linked to unique DNA-processing activities in closely related gut Bacteroidales strains. These findings thus establish DNA as a metabolic substrate in the gut microbiome and reveal a distinctive pathway for nucleobase production with implications for host-microbe interactions. SIGNIFICANCE STATEMENTThe gut microbiome plays a crucial role in nutrient metabolism, yet the fate of extracellular DNA within this ecosystem remains poorly understood. This study identifies Bacteroidales species that actively metabolize extracellular DNA, revealing a conserved pathway that converts DNA-derived nucleotides into deaminated nucleobases. We show that Bacteroides thetaiotaomicron utilizes a specialized genetic locus, ddbABCDEF, to facilitate this process, influencing nucleobase availability in the gut. Comparative genomic analyses suggest that ddbABCDEF is evolutionarily linked to bacterial natural transformation systems but has diverged into distinct metabolic subtypes. These findings establish DNA as a metabolic substrate in the gut microbiome, with potential implications for microbial ecology, host-microbe interactions, and gut health.

microbiology↗

bioRxiv2024

Fecal Metabolite Quantitation for Rapid Assessment of the Gut Microbiome

The intestinal microbiome is composed of myriad microbial species that produce metabolites that impact host health. While loss of bacterial species and beneficial metabolites from the fecal microbiome is associated with development of a range of diseases and medical complications, there are currently no diagnostic tests that rapidly identify individuals with microbiome deficiencies. Reduced concentrations of fecal butyrate and deoxycholic acid are associated with adverse clinical outcomes and result from the loss of a subset of health-associated bacterial species. We present a rapid diagnostic test based on 3-nitrophenylhydrazine derivatization and ultrahigh-performance liquid chromatography-mass spectrometry that measures fecal butyrate and deoxycholic acid concentrations as markers of microbiome function. A matrix-matched calibration curve was developed using a simulated fecal mixture to optimize accuracy and facilitate adherence to clinical laboratory regulations. The assay showed an analytical measurement range from 4.3-3030.1 {micro}M (LLOQ = 9.75 {micro}M) for butyrate and from 0.9- 64.9 {micro}M (LLOQ = 0.9 {micro}M) for deoxycholic acid. Precision demonstrated a coefficient of variation <15% at all QC levels tested. The assay can be performed in under an hour from extraction to provision of quantitative results, enabling the rapid identification of patients with defective microbiome function.

pathology↗

bioRxiv2024

Gut Bacteria Metabolize Natural and Synthetic Steroid Hormones via the Reductive OsrABC Pathway

Steroid hormone metabolism by the gut microbiome affects host physiology, but the underlying microbial pathways remain incompletely understood. Here, we isolate a novel human gut bacterium, Clostridium steroidoreducensT strain HCS.1 that reduces cortisol and related steroid hormones to 3{beta},5{beta}-tetrahydrosteroid products. Through transcriptomics and enzymatic discovery, we establish the C. steroidoreducens OsrABC steroid hormone pathway. OsrA is a 3-oxo-{Delta}1-steroid hormone reductase that targets synthetic glucocorticoids, including prednisolone-- a frontline Crohns disease therapy. OsrB is a 3-oxo-{Delta}4-steroid reductase that converts steroid hormones to 5{beta}-dihydrosteroid intermediates, which OsrC subsequently reduces to 3{beta},5{beta}-tetrahydro products. Homologs of osrA and osrB predict steroid-reducing activity across gut bacteria and are enriched in Crohns disease patient metagenomes. Consistent with a role in modulating drug efficacy, C. steroidoreducens colonization decreases prednisolone bioavailability in gnotobiotic mice. These findings thus define a previously unrecognized pathway for microbial steroid hormone inactivation and establish a mechanistic basis for bacterial interference with anti-inflammatory therapies.

microbiology↗

bioRxiv2024

MICROBIOME-MODIFIED METABOLITES PREDICT ACUTE BRAIN INJURY OUTCOME

BACKGROUNDAcute brain injury (ABI)-mediated disruption of the microbiome may potentiate inflammation and secondary brain injury (SBI). However, microbial-specific mediators and mechanisms remain unclear. METHODSThirty-five consecutive patients with ABI admitted to the neuroscience critical care unit at the University of Chicago were prospectively studied. Injury severity at hospital admission was assessed using the Injury Severity Score (ISS) and the Glasgow Coma Scale (GCS). Final neurologic function was assessed via the Glasgow Outcome Score extended (GOSe). Serum, plasma, and stool targeted metabolomics, as well as stool shotgun metagenomics, were performed on longitudinal samples collected during hospitalization. RESULTSMultivariate analysis identified microbiome-modified metabolites that were positively and negatively associated with functional outcomes after ABI. Novel identification of conjugated bile acid (BA) species and vitamin B12 precursors indicative of outcome were detected in the first collected samples (within 48 hours). Network analysis revealed greater integration of negatively associated metabolites across tissues and identified tauro-/-muricholic acid (TMCA) as central to the cross-tissue metabolomes. CONCLUSIONSMicrobiome metabolites may be useful in assessing brain injury outcomes to inform treatment. Bile acid species transformed by the gut microbiome are predictive of ABI outcome.

neuroscience↗

bioRxiv2024

Eubacterium rectale detoxification mechanism increases resilience of the gut environment

Lachnospiraceae members were highly detected in dysbiotic IL-10 KO mice that displayed similar physiological outcomes as control mice. Lachnospiraceae is a highly diverse family of microbes that have been shown to display both commensal and pathogenic characteristics in the colon environment. We investigated the impact of genetic variation in five Lachnospiraceae strains on lowering cellular inflammation and ROS levels. Cell free spent media (CFSM) from Eubacterium rectale resulted in lowered ROS, and nitric oxide levels in stressed colon cells. We demonstrated through an array of multi omics and molecular techniques that glutathione (GSH) biosynthesized by E. rectale was able to alleviate host ROS damage. We further showed downregulation of cell stress and immune response genes by host RNA sequencing, which is evidence that E. rectale microbial products promote recovery and alleviate ROS stress. HighlightsO_LILachnospiraceae detection correlated with lower host inflammation in vertically transmitted dysbiotic mice C_LIO_LICell free spent media from E. rectale lowered ROS and nitric oxide levels when introduced to stressed Caco-2 cells more effectively than 4 other strains of Lachnospiraceae C_LIO_LIEubacterium rectale spent media products include high levels of glutathione (GSH), carbohydrates, and amino acids C_LIO_LIEubacterium rectale potential gene function for glutathione biosynthesis and metabolism found in the genome C_LIO_LILower expression of repair and stress genes of intestinal cells treated with hydrogen peroxide was observed with E. rectale CFSM as well as treatment with 15 mM GSH C_LI

microbiology↗

bioRxiv2024

Protection against Clostridioides difficile disease by a naturally avirulent C. difficile strain

Clostridioides difficile (C. difficile) strains belonging to the epidemic BI/NAP1/027 (RT027) group have been associated with increased transmissibility and disease severity. In addition to the major toxin A and toxin B virulence factors, RT027 strains also encode the CDT binary toxin. Our lab previously identified a toxigenic RT027 isolate, ST1-75, that is avirulent in mice despite densely colonizing the colon. Here, we show that co-infecting mice with the avirulent ST1-75 and virulent R20291 strains protects mice from colitis due to rapid clearance of the virulent strain and persistence of the avirulent strain. Although avirulence of ST1-75 is due to a mutation in the cdtR gene, which encodes a response regulator that modulates the production of all three C. difficile toxins, the ability of ST1-75 to protect against acute colitis is not directly attributable to the cdtR mutation. Metabolomic analyses indicate that the ST1-75 strain depletes amino acids more rapidly than the R20291 strain and supplementation with amino acids ablates ST1-75s competitive advantage, suggesting that the ST1-75 strain limits the growth of virulent R20291 bacteria by amino acid depletion. Since the germination kinetics and sensitivity to the co-germinant glycine are similar for the ST1-75 and R20291 strains, our results identify the rapidity of in vivo nutrient depletion as a mechanism providing strain-specific, virulence-independent competitive advantages to different BI/NAP1/027 strains. They also suggest that the ST1-75 strain may, as a biotherapeutic agent, enhance resistance to CDI in high-risk patients. ImportanceClostridioides difficile infections (CDI) are prevalent in healthcare settings and are associated with high recurrence rates. Therapies to prevent CDI, including recent FDA-approved live biotherapeutic products, are costly and have not been used to prevent primary infections. While a nontoxigenic C. difficile strain (NTCD-M3) protects against virulent CDI in animals and reduced CDI recurrence in a phase 2 clinical trial, protection against CDI recurrence in humans was variable and required high doses of the nontoxigenic strain. Here we show that an avirulent C. difficile isolate, ST1-75, efficiently outcompetes virulent C. difficile strains in mice when co-infected at a 1:1 ratio. Our data suggest that inter-strain competition results from ST1-75s more rapid depletion of amino acids than the virulent R20291 strain. Our study identifies inter-strain nutrient depletion as a potentially exploitable mechanism to reduce the incidence of CDI.

microbiology↗

bioRxiv2023

The Host-specific Microbiota is Required for Diet-Specific Metabolic Homeostasis

In complex mammals, the importance and host-specificity of microbial communities have been demonstrated through their positive effects on host immune fitness or performance. However, whether host metabolic physiology homeostasis depends on a specific bacterial community exclusive to the host remains unclear. Here, we show that the coevolved host-specific microbiota is required to maintain diet-specific flexible and sufficient metabolic homeostasis through a high colonization rate, modulating gut metabolites, and related targets. Using germ-free (GF) mice, we tested whether the fitness benefiting the host metabolic phenotype of microbiota was host-specific. We demonstrated that GF mice associated with exogenous microbiota (human microbiota (HM)), which exhibited different and reduced gut microbial species diversity, significantly elevated metabolic rate, and exhibited metabolic insufficiency, all characteristics of GF mice. Strikingly, the absence of the host-specific microbiome attenuated high-fat diet-specific metabolism features. Different diets caused different metabolic changes in only host-specific microbiota-associated mice, not the host-microbiota mismatched mice. While RNA sequencing revealed subtle changes in the expression of genes in the liver, GF mice and HM mice showed considerably altered expression of genes associated with metabolic physiology compared to GF mice associated with host-specific microbiota. The effect of diet outweighed microbiota in the liver transcriptome. These changes occurred in the setting of decreased luminal short-chain fatty acids (SCFAs) and the secondary bile acid (BAs) pool and downstream gut signaling targets in HM and GF mice, which affects whole-body metabolism. These data indicate that a foreign microbial community provides little metabolic benefit to the host when compared to a host-specific microbiome, due to the colonization selection pressure and microbiota-derived metabolites dysfunction. Overall, microbiome fitness effects on the host metabolic phenotype were host-specific. Understanding the impact of the host-specificity of the microbiome on metabolic homeostasis may provide important insights for building a better probiotic. HighlightsO_LIMicrobiome fitness effects on the host metabolic phenotype were host-specific in mammals. C_LIO_LIHuman microbiota-associated mice exhibited lower host metabolic fitness or performance, and similar functional costs in GF mice. C_LIO_LIDifferent diets cause different metabolic changes only in host-specific microbiota-associated mice, not the host-microbiota mismatched mice. C_LIO_LIThe defective gut microbiota in host-specific microbiota, microbial metabolites and related targets likely drive the metabolic homeostasis. C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=132 SRC="FIGDIR/small/565654v1_ufig1.gif" ALT="Figure 1"> View larger version (41K): org.highwire.dtl.DTLVardef@16a864org.highwire.dtl.DTLVardef@1894996org.highwire.dtl.DTLVardef@3e5429org.highwire.dtl.DTLVardef@77bfb_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2022

Exceptionally versatile respiratory metabolisms drive metabolite production by diverse gut bacteria

Respiratory reductases enable microbes to utilize molecules present in anaerobic ecosystems as energy-generating respiratory electron acceptors. Here we identify three taxonomically distinct families of human gut bacteria (Burkholderiaceae, Eggerthellaceae, Erysipelotrichaceae) that encode large arsenals of tens-to-hundreds of respiratory-like reductases per genome. Screening species from each family (Sutterella wadsworthensis, Eggerthella lenta, and Holdemania filiformis), we discover 22 metabolites used as respiratory electron acceptors in a species-specific manner. Identified reactions transform multiple classes of dietary- and host-derived metabolites, including bioactive molecules resveratrol and itaconate. Products of identified respiratory metabolisms highlight poorly characterized compounds, such as the itaconate-derived 2-methylsuccinate. Reductase substrate-profiling defines enzyme-substrate pairs and reveals a complex picture of reductase evolution, providing evidence that reductases with specificities for related cinnamate substrates independently emerged at least four times. These studies thus establish an exceptionally versatile form of anaerobic respiration that directly links microbial energy metabolism to the gut metabolome.

microbiology↗

bioRxiv2022

Distinct energy-coupling factor transporter subunits enable flavin acquisition and extracytosolic trafficking for extracellular electron transfer in Listeria monocytogenes

A variety of electron transfer mechanisms link bacterial cytosolic electron pools with functionally diverse redox activities in the cell envelope and extracellular space. In Listeria monocytogenes, the ApbE-like enzyme FmnB catalyzes extracytosolic protein flavinylation, covalently linking a flavin cofactor to proteins that transfer electrons to extracellular acceptors. L. monocytogenes uses an energy-coupling factor (ECF) transporter complex that contains distinct substrate-binding, transmembrane, ATPase A, and ATPase A subunits (RibU, EcfT, EcfA, and EcfA) to import environmental flavins, but the basis of extracytosolic flavin trafficking for FmnB flavinylation remains poorly defined. In this study, we show that the proteins EetB and FmnA are related to ECF transporter substrate-binding and transmembrane subunits, respectively, and are essential for exporting flavins from the cytosol for flavinylation. Comparisons of the flavin import versus export capabilities of L. monocytogenes strains lacking different ECF transporter subunits demonstrates a strict directionality of substrate-binding subunit transport but partial functional redundancy of transmembrane and ATPase subunits. Based on these results, we propose that ECF transporter complexes with different subunit compositions execute directional flavin import/export through a broadly conserved mechanism. Finally, we present genomic context analyses which show that related ECF exporter genes are distributed across the Firmicutes phylum and frequently co-localize with genes encoding flavinylated extracytosolic proteins. These findings clarify the basis of ECF transporter export and extracytosolic flavin cofactor trafficking in Firmicutes.

microbiology↗