Microbial communities, functional, and flavor differences among three different-colored high-temperature Daqu: A comprehensive metagenomic, physicochemical, and electronic sensory analysis.

High-temperature Daqu (HTD) is the starter for producing sauce-flavor Baijiu, with different-colored Daqu (white, yellow, and black) reflecting variations in fermentation chamber conditions, chemical reactions, and associated microbiota. Understanding the relationship between Daqu characteristics and flavor/taste is challenging yet vital for improving Baijiu fermentation. This study utilized metagenomic sequencing, physicochemical analysis, and electronic sensory evaluation to compare three different-colored HTD and their roles in fermentation. Fungi and bacteria dominated the HTD-associated microbiota, with fungi increasing as the fermentation temperature rose. The major fungal genera were Aspergillus (40.17%) and Kroppenstedtia (21.16%), with Aspergillus chevalieri (25.65%) and Kroppenstedtia eburnean (21.07%) as prevalent species. Microbial communities, functionality, and physicochemical properties, particularly taste and flavor, were color-specific in HTD. Interestingly, the microbial communities in different-colored HTDs demonstrated robust functional complementarity. White Daqu exhibited non-significantly higher α-diversity compared to the other two Daqu. It played a crucial role in breaking down substrates such as starch, proteins, hyaluronic acid, and glucan, contributing to flavor precursor synthesis. Yellow Daqu, which experienced intermediate temperature and humidity, demonstrated good esterification capacity and a milder taste profile. Black Daqu efficiently broke down raw materials, especially complex polysaccharides, but had inferior flavor and taste. Notably, large within-group variations in physicochemical quality and microbial composition were observed, highlighting limitations in color-based HTD quality assessment. Water content in HTD was associated with Daqu flavor, implicating its crucial role. This study revealed the complementary roles of the three HTD types in sauce-flavor Baijiu fermentation, providing valuable insights for product enhancement.

Bifidobacterium bifidum FJSWX19M5 alleviated 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced chronic colitis by mitigating gut barrier injury and increasing regulatory T cells.

Probiotics have been evaluated as alternative approaches for preventing the relapse of Crohn's disease (CD). Previously, we observed strain-specific anti-inflammatory properties of Bifidobacterium bifidum in 2,4,6-trinitrobenzene sulfonic acid (TNBS) acute colitis models. In this study, we further assessed the effects of several B. bifidum strains on colonic damage, fibrosis, inflammatory factors, intestinal microbial and metabolic profiles, and peripheral regulatory T cells (Tregs) in the context of TNBS chronic colitis in mice. These results indicated that B. bifidum FJSWX19M5, but not FXJWS17M4, ameliorated body weight loss, reduced colonic shortening and injury, decreased markers of gut inflammation, and rebalanced colonic metabolism in TNBS-treated mice. FJSWX19M5 supplementation also promoted Treg cell differentiation and intestinal barrier restoration compared to other strains. All living B. bifidum strains (FJSWX19M5, FXJWS17M4 and FHENJZ3M6) seemed to restore the disruption of the gut microbiota caused by TNBS. The co-culture of B. bifidum strains and mesenteric lymph node cells from TNBS-treated mice showed that those strains with anti-colitis could induce higher IL-10 levels and a lower ratio of IL-22/IL-10 and IL-17/IL-10 when compared to those strains that were not protective. Furthermore, heat-killed FJSWX19M5 exhibited a relief effect on colitis-related symptoms (including body weight loss, colonic shortening and injury). These data imply that specific B. bifidum strains or their lysates may be the current therapeutic alternatives for CD.

Species- or genus-dependent immunostimulatory effects of gut-derived potential probiotics.

The immune regulatory effects of probiotics have been widely recognized to be strain-specific. However it is unknown if there is a species- or genus-dependent manner. In this study, we use an in vitro mesenteric lymph node (MLN) model to systematically evaluate the immunostimulatory effects of gut-derived potential probiotics. The results exhibit an obvious species or genus consensus immune response pattern. RNA-seq shows that T cell-dependent B cell activation and antibody responses may be inherent to this model. Of the five tested genera, Akkermansia spp. and Clostridium butyrium directly activate the immune response in vitro, as indicated by the secretion of interleukin-10. Bifidobacterium spp. and Bacteroides spp. activate immune response with the help of stimuli (anti-CD3 and anti-CD28 antibodies). Lactobacillus spp. blunt the immune response with or without stimuli. Further investigations show that the cell surface protein of A. muciniphila AH39, which may serve as a T cell receptor cognate antigen, might evoke an in vitro immune activation. In vivo, oral administration of A. muciniphila AH39 influences the proportion of T regulatory cells (Tregs) in MLNs and the spleen under homeostasis in both specific pathogen-free and germ-free mice. All these findings indicate the distinct effects of different genera or species of potential gut-derived probiotics on intestinal and systemic immunity.

Characteristics of an In Vitro Mesenteric Lymph Node Cell Suspension Model and Its Possible Association with In Vivo Functional Evaluation.

In a previous study, we uncovered three immune-responsive patterns of gut microbes using an in vitro mesenteric lymph node cell suspension model, abbreviated as the MLN model hereafter. We used Akkermansia muciniphila and Clostridium butyricum as the first group directly inducing an immune response, Bifidobacterium sp. and Bacteroides sp. as the second group evoking an immune response with the help of stimuli (anti-CD3 and anti-CD28 antibodies), and Lactobacillus sp. as the third group blunting the immune response with or without stimuli. Our group previously clarified the immune-activation characteristics of A. muciniphila and linked its in vivo immune induction effect in GF and SPF mice under homeostasis. In the present study, we supplemented the characteristics of C. butyricum and B. bifidum in the in vitro MLN model and addressed the specific elements of the model. Finally, we used an in vivo TNBS-challenge model to show the functional differences between these species with different response patterns in vitro. The results showed that C. butyricum and B. bifidum evoked an immune response in vitro in a dose-dependent and strain-unique manner. Although TLR2, rather than TLR4, is indispensable for immune activation in the present in vitro model, it may not involve interaction between TLR2 and bacterial ligands. Like the PBMC model, the present in vitro MLN model is highly dependent on cell resources and should be given more attention when used to conduct a quantitative comparison. Finally, a mixture of two strong immunogenic strains, A. muciniphila and C. butyricum, significantly increased the mortality of TNBS-challenged (2,4,6-trinitrobenzene sulfonic acid, TNBS) mice, indicating a possible link between the in vitro MLN model and in vivo functional evaluation. However, more evidence is needed to clarify the associations and underlying mechanisms.

Protective effects of Bacteroides fragilis against lipopolysaccharide-induced systemic inflammation and their potential functional genes.

Bacteroides fragilis, one of the potential next-generation probiotics, has been demonstrated to alleviate inflammation-associated diseases. In this study, we compare the anti-inflammatory effects of six Bacteroides fragilis strains on systemic inflammation and link their strain-specific characteristics, both physiologically and genetically, to their function. A lipopolysaccharide (LPS)-induced systemic inflammation model in mice was used as an in vivo model to compare the effects of different B. fragilis strains. Short-chain fatty acids (SCFAs) were measured by gas chromatography-mass spectrometry (GC-MS). The in vitro immunomodulatory properties were evaluated in LPS-stimulating RAW264.7 cell lines. Orthologous gene clusters were compared using OrthoVenn2. The results indicate a strain-specific in vitro anti-inflammatory effect. Effective strains induce higher colon SCFAs in vivo and interleukin-10 (IL-10) production in vitro. Comparative genomic analysis showed that the SCFA-inducing strains possess three genes relating to carbohydrate metabolism (GH2, GH35 families) and binding and transportation (SusD), all of which are associated with niche fitness and expansion. IL-10-inducing strains share a highly similar gene, wbjE, which may result in a distinct O-antigen structure of LPS and influence their immunomodulatory properties. B. fragilis is strain-specific against LPS-induced systemic inflammation in mice. The beneficial effects of a specific strain may be attributed to its SCFA and IL-10 inducing abilities. Strain-specific potential genes can be excavated to link these differences.

The effects of diet and gut microbiota on the regulation of intestinal mucin glycosylation.

Intestinal mucins glycosylation is regulated by host cues and environmental signals from the microbiome and diets. However, the mechanisms responsible for the dialogue between these three factors and mucin glycosylation in the digestive environment of the host are not well understood. In this review, the dynamic alterations of mucin glycosylation induced by immune responses to gut diseases are summarized. The various types of interactions between mucin glycans and gut microbes, including adhesins, glycosidases, metabolic products and surface components, are discussed. The mechanisms that determine how dietary components (fat, fiber, prebiotics, protein, and food additives) affect intestinal mucin glycosylation and maintain mucosal homeostasis are identified. A potential framework for individualized dietary recommendations is proposed for the prevention of abnormal mucin glycosylation driven by immune dysregulation, gut microbiome alterations and other factors. This review may provide a basis for future research on glycosylation-inspired therapies for gut diseases.

Dose-dependent effects of lead induced gut injuries: An in vitro and in vivo study.

Lead (Pb) toxicity has been widely studied, but its dose-dependent toxic effects on the gut remain unclear, therefore, the aim of this study was to evaluate the effects of different doses of Pb exposure on the gut microbiota and gut barrier in vitro and in vivo. The HT-29 cell model was used to determine the Pb-induced effects on cell viability, reactive oxygen species (ROS), and tight junction proteins (TJPs) in vitro, and C57BL/6 mice models exposed to 0, 20, 100, 500, and 1000 mg/kg Pb were used to investigate the Pb-induced dose-dependent effects on the gut microbiota, TJP expression, and colon histopathology. Our results showed that the exposure of HT-29 cells to 8 mM Pb decreased cell viability by 50%, elevated ROS levels by 200%, and suppressed the expression of the TJPs, zonula occludens-1 (ZO-1) and occludin by 23% and 35%, respectively. Consistently, Pb-exposed mice showed significant increases in colon tissue damage and inflammation and reductions in ZO-1 mRNA levels in a dose-dependent manner. The occludin mRNA levels decreased in the 500 and 1000 mg/kg groups. At the genus level, the relative abundance of Coprococcus and Oscillospira decreased and that of Lactobacillus increased in linear manner with the Pb exposure dose. PICRUSt analysis based on 16S rRNA sequencing revealed Pb dose-dependent alterations in metabolism through the gut microbiota. These findings suggest that Pb exposure can not only disrupt the barrier by generating oxidative stress, but can also induce gut dysbiosis, colon tissue damage, and gut inflammation in a dose-dependent manner.

Effects of acute oral lead exposure on the levels of essential elements of mice: a metallomics and dose-dependent study.

BACKGROUND AND OBJECTIVES: Lead (Pb) has been reported to disturb the metabolism of essential elements, such as calcium (Ca), magnesium (Mg), iron (Fe) and zinc (Zn) in vivo. This study focused on the relationship between various dose of Pb and the essential elements. METHODS: 50 healthy male C57BL/6 mice underwent oral administration of 0.2 mL lead acetate trihydrate solution (0, 20, 100, 500, and 1000 mg Pb/day/kg body weight) for 3 days. The concentrations of Pb and four essential elements (Ca, Zn, Fe and Mg) in the blood, kidney, liver, bone and brain were quantified with inductively coupled plasma mass spectrometry. RESULTS: Various doses of Pb led to significant increases in the contents of Ca, Fe and Zn in the liver, and decreased contents of Mg and Fe in the blood in a dose-dependent pattern. The Pb dose of 20 mg/kg reduced the concentration of bone Ca, which did not continue to show an obvious decline with continued increases in the oral Pb dose. Pb also caused alterations in the Mg distribution pattern, and decreased the correlation of Mg, Ca and Zn in the brain, both findings were dose-dependent. In addition to the changes in metallomics, the related oxidative stress was exacerbated, but no significant changes were detected in hepatic and renal histopathological lesions after a short period of Pb exposure. CONCLUSIONS: This study contributes to a thorough analysis of the Pb-poisoning mechanism, and indicates that the concentrations of essential elements could be used as sensitive toxicological indicators of Pb exposure.

Lactobacillus plantarum CCFM8661 modulates bile acid enterohepatic circulation and increases lead excretion in mice.

The management of lead (Pb) exposure and toxicity remains a major public health priority worldwide. In our previous study, the probiotic strain Lactobacillus plantarum CCFM8661 prevented Pb absorption in mice via intestinal sequestration. This follow-up study aimed to evaluate the additional protective mechanism of L. plantarum CCFM8661 with a focus on its regulation of enterohepatic circulation. We first confirmed the relationship between the enterohepatic circulations of Pb and bile acid (BA) by administering a BA sequestrant, cholestyramine, to mice with a high Pb burden. Our data further showed that L. plantarum CCFM8661 significantly induced hepatic BA synthesis, enhanced bile flow and biliary glutathione output, and increased fecal BA excretion in the mice, which in turn increased biliary Pb output and enhanced fecal Pb excretion. This regulation was associated with the alterations in the expression of target genes in the enterohepatic farnesoid X receptor-fibroblast growth factor (FXR-FGF15) axis and could be reversed using an FXR agonist, GW4064. Pre-treatment with antibiotics also abolished the L. plantarum CCFM8661-induced effects on BA and Pb enterohepatic circulation. These results suggest that L. plantarum CCFM8661 induces fecal Pb excretion by regulating BA enterohepatic circulation. This regulation is associated with the down-regulation of the FXR-FGF15 axis and is partly dependent on the gut microbiota of mice.

Oral Supplementation of Lead-Intolerant Intestinal Microbes Protects Against Lead (Pb) Toxicity in Mice.

Oral exposure to the heavy metal lead (Pb) causes various dysfunctions in animals. However, the influence of gut bacteria on Pb absorption, bioaccumulation, and excretion is largely unknown. In this study, we use a mouse model to investigate the relationship between gut microbiota, Pb-intolerant intestinal microbes and Pb toxicity. First, mice were treated with a broad-spectrum antibiotic cocktail to deplete their gut microbiota, and were then acutely and orally exposed to Pb at 1304 mg/kg for 3 days. Compared to the control mice, antibiotic-treated mice had increased Pb concentrations in the blood and primary organs and decreased Pb fecal concentrations, suggesting that gut microbiota limited the Pb burden that developed from acute oral Pb exposure. Next, three Pb-intolerant gut microbes, Akkermansia muciniphila, Faecalibacterium prausnitzii, and Oscillibacter ruminantium, were orally administered to mice, and their effects against Pb toxicity were evaluated. F. prausnitzii treatment significantly promoted the fecal Pb excretion and reduced Pb concentrations in blood (from 152.70 ± 25.62 µg/dL to 92.20 ± 24.33 µg/dL) and primary tissues. Supplementation with O. ruminantium significantly decreased Pb concentrations in blood (from 152.70 ± 25.62 µg/dL to 104.60 ± 29.85 µg/dL) and kidney (from 7.30 ± 1.08 µg/g to 5.64 ± 0.79 µg/g). Treatment with F. prausnitzii and O. ruminantium also upregulated tight junction (TJ) protein expression and the production of short-chain fatty acids by colonic microbiota, and showed protective effects against liver and kidney toxicity. These results indicate the potential for reducing Pb toxicity by the modulation of gut microbiota.