In Vitro Fermentation of NUTRIOSE? FB06, a Wheat Dextrin Soluble Fibre,in a Continuous Culture Human Colonic Model System

Wheat dextrin soluble fibre may have metabolic and health benefits, potentially acting via mechanisms governed by the selective modulation of the human gut microbiota. Our aim was to examine the impact of wheat dextrin on the composition and metabolic activity of the gut microbiota. We used a validated in vitro three-stage continuous culture human colonic model (gut model) system comprised of vessels simulating anatomical regions of the human colon. To mimic human ingestion, 7 g of wheat dextrin (NUTRIOSE^® FB06) was administered to three gut models, twice daily at 10.00 and 15.00, for a total of 18 days. Samples were collected and analysed for microbial composition and organic acid concentrations by 16S rRNA-based fluorescence in situ hybridisation and gas chromatography approaches, respectively. Wheat dextrin mediated a significant increase in total bacteria in vessels simulating the transverse and distal colon, and a significant increase in key butyrate-producing bacteria Clostridium cluster XIVa and Roseburia genus in all vessels of the gut model. The production of principal short-chain fatty acids, acetate, propionate and butyrate, which have been purported to have protective, trophic and metabolic host benefits, were increased. Specifically, wheat dextrin fermentation had a significant butyrogenic effect in all vessels of the gut model and significantly increased production of acetate (vessels 2 and 3) and propionate (vessel 3), simulating the transverse and distal regions of the human colon, respectively. In conclusion, wheat dextrin NUTRIOSE^® FB06 is selectively fermented in vitro by Clostridium cluster XIVa and Roseburia genus and beneficially alters the metabolic profile of the human gut microbiota.     

Kinetic modelling of lactate utilization and butyrate production by key human colonic bacterial species

Butyrate is the preferred energy source for colonocytes and has an important role in gut health; in contrast, accumulation of high concentrations of lactate is detrimental to gut health. The major butyrate-producing bacterial species in the human colon belong to the Firmicutes. Eubacterium hallii and a new species, Anaerostipes coli SS2/1, members of clostridial cluster XIVa, are able to utilize lactate and acetate via the butyryl CoA : acetate CoA transferase route, the main metabolic pathway for butyrate synthesis in the human colon. Here we provide a mathematical model to analyse the production of butyrate by lactate-utilizing bacteria from the human colon. The model is an aggregated representation of the fermentation pathway. The parameters of the model were estimated using total least squares and maximum likelihood, based on in vitro experimental data with E. hallii L2-7 and A. coli SS2/1. The findings of the mathematical model adequately match those from the bacterial batch culture experiments. Such an in silico approach should provide insight into carbohydrate fermentation and short-chain fatty acid cross-feeding by dominant species of the human colonic microbiota.     

Effect of garlic powder on the growth of commensal bacteria from the gastrointestinal tract

Garlic (Allium sativum) is considered one of the best disease-preventive foods. We evaluated in vitro the effect of a commercial garlic powder (GP), at concentrations of 0.1% and 1% (w/v), upon the viability of representative gut bacteria. In pure culture studies, Lactobacillus casei DSMZ 20011 was essentially found to be resistant to GP whereas a rapid killing effect of between 1 and 3 log CFU/ml reduction in cell numbers was observed with Bacteroides ovatus, Bifidobacterium longum DSMZ 20090 and Clostridium nexile A2-232. After 6h incubation, bacterial numbers increased steadily and once the strains became resistant they retained their resistant phenotype upon sub-culturing. A colonic model was also used to evaluate the effect of GP on a mixed bacterial population representing the microbiota of the distal colon. Lactic acid bacteria were found to be more resistant to GP compared to the clostridial members of the gut microbiota. While for most bacteria the antimicrobial effect was transient, the lactobacilli showed a degree of resistance to garlic, indicating that its consumption may favour the growth of these beneficial bacterial species in the gut. Garlic intake has the potential to temporarily modulate the gut microbiota.     

PCR/DGGE and 16S rRNA gene library analysis of the colonic microbiota of HLA-B27/beta2-microglobulin transgenic rats

AIMS: To determine the phylogenetic composition of the colonic microbiota of transgenic (TG) HLA-B27 rats using 16S ribosomal RNA (rRNA) gene sequences obtained from denaturing gradient gel electrophoresis (DGGE) gels and sequences from a 16S rRNA gene library. METHODS AND RESULTS: Colonic microbiota of TG and nontransgenic (NT) rats harboured by 10-week-old and 6-month-old animals was screened using PCR/DGGE. Six months old TG rats had marked inflammation of the colon compared with 10-week-old TG and NT rats. The DGGE profiles of rats with inflamed colon were similar from rat to rat (Dice's Similarity Coefficient proximal colon 73%, distal colon 83%) whereas profiles from animals without inflammation were dissimilar (52-64%). Identifications of bacterial origins of 16S rRNA gene sequences obtained from DGGE gels (200 bp) and from 16S rRNA clones (450 bp) of the colonic microbiota of diseased rats gave sequences most closely phylogenetically affiliated with uncultured or unknown bacteria. CONCLUSIONS: PCR/DGGE was shown to be an effective method to compare the colonic microbiota composition of TG and NT rats relative to the progression of inflammatory disease. Sequencing of 16S rRNA gene fragments from DGGE gels or 16S rRNA gene clones from a random library showed that uncultured or unknown bacteria were most commonly detected by both methods. It can be concluded that it would be better in future studies to search for the antigens produced by the gut microbiota against which the dysfunctional immune system reacts rather than seek phylogenetic associations. SIGNIFICANCE AND IMPACT OF THE STUDY: PCR/DGGE can be used as a rapid initial screening method to compare the composition of bacterial communities of initially unknown composition that are associated with the development of intestinal disease.     

Beneficial Effects of Celastrol on Immune Balance by Modulating Gut Microbiota in Experimental Ulcerative Colitis Mice

Ulcerative colitis (UC) is a chronic inflammatory bowel disease caused by many factors including colonic inflammation and microbiota dysbiosis. Previous studies have indicated that celastrol (CSR) has strong anti-inflammatory and immune-inhibitory effects. Here, we investigated the effects of CSR on colonic inflammation and mucosal immunity in an experimental colitis model, and addressed the mechanism by which CSR exerts the protective effects. We characterized the therapeutic effects and the potential mechanism of CSR on treating UC using histological staining, intestinal permeability assay, cytokine assay, flow cytometry, fecal microbiota transplantation (FMT), 16S rRNA sequencing, untargeted metabolomics, and cell differentiation. CSR administration significantly ameliorated the dextran sodium sulfate (DSS)-induced colitis in mice, which was evidenced by the recovered body weight and colon length as well as the decreased disease activity index (DAI) score and intestinal permeability. Meanwhile, CSR down-regulated the production of pro-inflammatory cytokines and up-regulated the amount of anti-inflammatory mediators at both mRNA and protein levels, and improved the balances of Treg/Th1 and Treg/Th17 to maintain the colonic immune homeostasis. Notably, all the therapeutic effects were exerted in a gut microbiota-dependent manner. Furthermore, CSR treatment increased the gut microbiota diversity and changed the compositions of the gut microbiota and metabolites, which is probably associated with the gut microbiota-mediated protective effects. In conclusion, this study provides the strong evidence that CSR may be a promising therapeutic drug for UC.     

Saccharomyces boulardii interferes with Shigella pathogenesis by postinvasion signaling events

Saccharomyces boulardii is gaining in popularity as a treatment for a variety of diarrheal diseases as well as inflammatory bowel disease. This study was designed to examine the effect of this yeast on infection by Shigella flexneri, a highly infectious and human host-adapted enteric pathogen. We investigated key interactions between the bacteria and host cells in the presence of the yeast in addition to a number of host responses including proinflammatory events and markers. Although the presence of the yeast during infection did not alter the number of bacteria that was able to attach or invade human colon cancer-derived T-84 cells, it did positively impact the tight junction protein zonula occluden-2 and significantly increase the barrier integrity of model epithelia. The yeast also decreased ERK, JNK, and NF-kappaB activation in response to S. flexneri, events likely responsible for the observed reductions in IL-8 secretion and the transepithelial migration of polymorphonuclear leukocytes across T-84 monolayers. These results, suggesting that the yeast allowed for a dampened inflammatory response, were confirmed in vivo utilizing a highly relevant model of human fetal colonic tissue transplanted into scid mice. Furthermore, a cell-free S. boulardii culture supernatant was also capable of reducing IL-8 secretion by infected T-84 cells. These data suggest that although the use of S. boulardii during infection with S. flexneri may alleviate symptoms associated with the inflammatory response of the host, it would not prevent infection.     

Potential probiotic Kluyveromyces marxianus B0399 modulates the immune response in Caco-2 cells and peripheral blood mononuclear cells and impacts the human gut microbiota in an in vitro colonic model system

Considering the increase in the consumption of yeasts as human probiotics, the aim of this study was to broadly investigate the beneficial properties of the lactic yeast Kluyveromyces marxianus (formerly Kluyveromyces fragilis) B0399. Several potential probiotic traits of K. marxianus B0399 were investigated by using in vitro assays, including adhesion and immune modulation, and the effect of the administration of 10(7) CFU/day of K. marxianus B0399 on the composition and metabolic activity of the human intestinal microbiota was investigated in a 3-stage continuous-culture system simulating the human colon. We demonstrated that this strain was highly adhesive to human enterocyte-like Caco-2 cells and modulated the immune response, inducing proinflammatory cytokines in peripheral blood mononuclear cells (PBMCs). In the presence of inflammatory stimulation with lipopolysaccharide (LPS), K. marxianus B0399 provoked decreases in the levels of production of proinflammatory cytokines in PBMCs and Caco-2 cells, thus ameliorating the inflammatory response. Furthermore, K. marxianus B0399 impacted the colonic microbiota, increasing the bifidobacterial concentration in the stages of the colonic model system simulating the proximal and transverse colon. The amounts of the short-chain fatty acids acetate and propionate also increased following yeast supplementation. Finally, K. marxianus B0399 was found to induce a decrease of the cytotoxic potential of the culture supernatant from the first stage of the colonic model system. The effects of K. marxianus B0399 on adhesion, immune function, and colonic microbiota demonstrate that this strain possesses a number of beneficial and strain-specific properties desirable for a microorganism considered for application as a probiotic.     

Like Will to Like: Abundances of Closely Related Species Can Predict Susceptibility to Intestinal Colonization by Pathogenic and Commensal Bacteria

The intestinal ecosystem is formed by a complex, yet highly characteristic microbial community. The parameters defining whether this community permits invasion of a new bacterial species are unclear. In particular, inhibition of enteropathogen infection by the gut microbiota ( = colonization resistance) is poorly understood. To analyze the mechanisms of microbiota-mediated protection from Salmonella enterica induced enterocolitis, we used a mouse infection model and large scale high-throughput pyrosequencing. In contrast to conventional mice (CON), mice with a gut microbiota of low complexity (LCM) were highly susceptible to S. enterica induced colonization and enterocolitis. Colonization resistance was partially restored in LCM-animals by co-housing with conventional mice for 21 days (LCM^con21). 16S rRNA sequence analysis comparing LCM, LCM^con21 and CON gut microbiota revealed that gut microbiota complexity increased upon conventionalization and correlated with increased resistance to S. enterica infection. Comparative microbiota analysis of mice with varying degrees of colonization resistance allowed us to identify intestinal ecosystem characteristics associated with susceptibility to S. enterica infection. Moreover, this system enabled us to gain further insights into the general principles of gut ecosystem invasion by non-pathogenic, commensal bacteria. Mice harboring high commensal E. coli densities were more susceptible to S. enterica induced gut inflammation. Similarly, mice with high titers of Lactobacilli were more efficiently colonized by a commensal Lactobacillus reuteri ^RR strain after oral inoculation. Upon examination of 16S rRNA sequence data from 9 CON mice we found that closely related phylotypes generally display significantly correlated abundances (co-occurrence), more so than distantly related phylotypes. Thus, in essence, the presence of closely related species can increase the chance of invasion of newly incoming species into the gut ecosystem. We provide evidence that this principle might be of general validity for invasion of bacteria in preformed gut ecosystems. This might be of relevance for human enteropathogen infections as well as therapeutic use of probiotic commensal bacteria.     

Dietary glycated protein modulates the colonic microbiota towards a more detrimental composition in ulcerative colitis patients and non-ulcerative colitis subjects

Aim:  To investigate the effect of native, heated and glycated bovine serum albumin (BSA) on the ulcerative colitis (UC) and non-UC colonic microbiota in vitro .
Methods and Results:  Continuous flow culture (CFC) models of the human colonic microbiota inoculated with faeces from UC and non-UC volunteers were maintained on BSA as growth substrate. Changes in bacterial populations and short-chain fatty acids were determined. UC and non-UC microbiota differed significantly in microbial populations, with elevated numbers of sulfate-reducing bacteria (SRB) and clostridia in the microbiota from UC patients. Compared with native BSA, glycated BSA modulated the gut microbiota of UC patients in vitro towards a more detrimental community structure with significant increases in putatively harmful bacteria (clostridia, bacteroides and SRB; P  < 0·009) and decreases in dominant and putatively beneficial bacterial groups (eubacteria and bifidobacteria; P  < 0·0004). The levels of beneficial short-chain fatty acids were significantly decreased by heated or glycated BSA, but were increased significantly by native BSA.
Conclusion:  The UC colonic microbiota maintained in CFC was significantly modified by glycated BSA.
Significance and Impact of the Study:  Results suggest that dietary glycated protein may impact upon the composition and activity of the colonic microbiota, an important environmental variable in UC.     

The bacterial community in the gut of the Cockroach Shelfordella lateralis reflects the close evolutionary relatedness of cockroaches and termites

Termites and cockroaches are closely related, with molecular phylogenetic analyses even placing termites within the radiation of cockroaches. The intestinal tract of wood-feeding termites harbors a remarkably diverse microbial community that is essential for the digestion of lignocellulose. However, surprisingly little is known about the gut microbiota of their closest relatives, the omnivorous cockroaches. Here, we present a combined characterization of physiological parameters, metabolic activities, and bacterial microbiota in the gut of Shelfordella lateralis, a representative of the cockroach family Blattidae, the sister group of termites. We compared the bacterial communities within each gut compartment using terminal-restriction fragment length polymorphism (T-RFLP) analysis and made a 16S rRNA gene clone library of the microbiota in the colon-the dilated part of the hindgut with the highest density and diversity of bacteria. The colonic community was dominated by members of the Bacteroidetes, Firmicutes (mainly Clostridia), and some Deltaproteobacteria. Spirochaetes and Fibrobacteres, which are abundant members of termite gut communities, were conspicuously absent. Nevertheless, detailed phylogenetic analysis revealed that many of the clones from the cockroach colon clustered with sequences previously obtained from the termite gut, which indicated that the composition of the bacterial community reflects at least in part the phylogeny of the host.     

The Effect of Selected Synbiotics on Microbial Composition and Short-Chain Fatty Acid Production in a Model System of the Human Colon

Background

Prebiotics, probiotics and synbiotics can be used to modulate both the composition and activity of the gut microbiota and thereby potentially affecting host health beneficially. The aim of this study was to investigate the effects of eight synbiotic combinations on the composition and activity of human fecal microbiota using a four-stage semicontinuous model system of the human colon.

Methods and Findings

Carbohydrates were selected by their ability to enhance growth of the probiotic bacteria Lactobacillus acidophilus NCFM (NCFM) and Bifidobacterium animalis subsp. lactis Bl-04 (Bl-04) under laboratory conditions. The most effective carbohydrates for each probiotic were further investigated, using the colonic model, for the ability to support growth of the probiotic bacteria, influence the composition of the microbiota and stimulate formation of short-chain fatty acids (SCFA).The following combinations were studied: NCFM with isomaltulose, cellobiose, raffinose and an oat β-glucan hydrolysate (OBGH) and Bl-04 with melibiose, xylobiose, raffinose and maltotriose. All carbohydrates showed capable of increasing levels of NCFM and Bl-04 during fermentations in the colonic model by 10³–10⁴ fold and 10–10² fold, respectively. Also the synbiotic combinations decreased the modified ratio of Bacteroidetes/Firmicutes (calculated using qPCR results for Bacteroides-Prevotella-Porphyromonas group, Clostridium perfringens cluster I, Clostridium coccoides - Eubacterium rectale group and Clostridial cluster XIV) as well as significantly increasing SCFA levels, especially acetic and butyric acid, by three to eight fold, as compared to the controls. The decreases in the modified ratio of Bacteroidetes/Firmicutes were found to be correlated to increases in acetic and butyric acid (p = 0.04 and p = 0.03, respectively).

Conclusions

The results of this study show that all synbiotic combinations investigated are able to shift the predominant bacteria and the production of SCFA of fecal microbiota in a model system of the human colon, thereby potentially being able to manipulate the microbiota in a way connected to human health.     

Alteration of the gastrointestinal microbiota of mice by edible blue-green algae

Aims:  To characterize the effect of edible blue-green algae (cyanobacteria) on the gastrointestinal microbiota of mice.
Methods and Results:  C57BL/6J mice were fed a diet supplemented with 0% or 5% dried Nostoc commune , Spirulina platensis or Afanizominon flos-aquae (w/w) for 4 weeks. Molecular fingerprinting of the colonic microbiota using denaturing gradient gel electrophoresis revealed that administration of N. commune induced major alterations in colonic microbiota composition, while administration of S. platensis or A. flos-aquae had a more subtle impact. Community profile analysis revealed that administration of N. commune did not reduce microbial diversity indices of the colonic microbiota. Despite its pronounced effects on the bacterial composition in the colon, total bacterial numbers in the gut of mice fed N. commune were not reduced as assessed by quantitative real-time PCR and bacteriological culture.
Conclusions:  The results presented here show that administration of blue-green algae, and especially N. commune , alters colonic microbiota composition in mice with limited effects on total bacterial numbers or microbial diversity.
Significance and Impact of the Study:  Blue-green algae are consumed in many countries as a source of nutrients and to promote health, and they are intensively studied for their pharmaceutical value. Given the importance of the gut microbiota for many host functions, the effects of blue-green algae on gut microbial ecology revealed during this study should be considered when using them as food supplements or when studying their pharmaceutical properties.     

Human milk oligosaccharides shorten rotavirus-induced diarrhea and modulate piglet mucosal immunity and colonic microbiota

The impact of human milk oligosaccharides (HMO) on mucosal immunity, gut microbiota and response to rotavirus (RV) infection was investigated in the piglet model. Newborn piglets were fed with formula alone (FF) or formula supplemented with 4 g l⁻¹ HMO (HMO) or a prebiotic mixture of 9:1 short-chain galactooligosaccharides (3.6 g l⁻¹) and long-chain fructooligosaccharides (0.4 g l⁻¹) (PRE) (n=19–21 per group) for 15 days. Piglets (n=7–8) in each dietary group were orally infected with porcine rotavirus (RV) OSU strain on d10, and stool consistency was assessed daily. Blood, small intestine and colonic contents were collected at day 15. Serum RV-specific antibody concentrations, intestinal histomorphology, RV non-structural protein-4 (NSP4) and cytokine mRNA expression were assessed. Colonic content pH, dry matter (DM) and short-chain fatty acid concentrations were measured. Ascending colonic microbiota was analyzed by 16S rRNA gene v1-3 region pyrosequencing. HMO- and PRE-fed groups had shorter duration of diarrhea than FF piglets. Infection changed intestinal histomorphology, increased serum RV-specific antibody response and intestinal RV NSP4 expression, and modulated ileal cytokine expression. HMO enhanced T helper type 1 (interferon-gamma) and anti-inflammatory (interleukin-10) cytokines in the ileum, while prebiotics promoted RV-specific immunoglobulin M response to the infection. RV infection and HMO supplementation altered intraluminal environment and gut microbiota. HMO increased pH and lowered DM of colonic contents and enhanced the abundance of unclassified Lachnospiraceae, which contains numerous butyrate-producing bacteria. In conclusion, HMO and prebiotics did not prevent the onset of RV infection but reduced the duration of RV-induced diarrhea in piglets, in part, by modulating colonic microbiota and immune response to RV infection.     

CD4CD8αα Lymphocytes,A Novel Human Regulatory T Cell Subset Induced by Colonic Bacteria and Deficient in Patients with Inflammatory Bowel Disease

How the microbiota affects health and disease is a crucial question. In mice, gut Clostridium bacteria are potent inducers of colonic interleukin (IL)-10-producing Foxp3 regulatory T cells (Treg), which play key roles in the prevention of colitis and in systemic immunity. In humans, although gut microbiota dysbiosis is associated with immune disorders, the underlying mechanism remains unknown. In contrast with mice, the contribution of Foxp3 Treg in colitis prevention has been questioned, suggesting that other compensatory regulatory cells or mechanisms may exist. Here we addressed the regulatory role of the CD4CD8 T cells whose presence had been reported in the intestinal mucosa and blood. Using colonic lamina propria lymphocytes (LPL) and peripheral blood lymphocytes (PBL) from healthy individuals, and those with colon cancer and irritable bowel disease (IBD), we demonstrated that CD4CD8αα (DP8α) T lymphocytes expressed most of the regulatory markers and functions of Foxp3 Treg and secreted IL-10. Strikingly, DP8α LPL and PBL exhibited a highly skewed repertoire toward the recognition of Faecalibacterium prausnitzii, a major Clostridium species of the human gut microbiota, which is decreased in patients with IBD. Furthermore, the frequencies of DP8α PBL and colonic LPL were lower in patients with IBD than in healthy donors and in the healthy mucosa of patients with colon cancer, respectively. Moreover, PBL and LPL from most patients with active IBD failed to respond to F. prausnitzii in contrast to PBL and LPL from patients in remission and/or healthy donors. These data (i) uncover a Clostridium-specific IL-10-secreting Treg subset present in the human colonic LP and blood, (ii) identify F. prausnitzii as a major inducer of these Treg, (iii) argue that these cells contribute to the control or prevention of colitis, opening new diagnostic and therapeutic strategies for IBD, and (iv) provide new tools to address the systemic impact of both these Treg and the intestinal microbiota on the human immune homeostasis.     

Paeoniflorin modulates gut microbial production of indole-3-lactate and epithelial autophagy to alleviate colitis in mice

BackgroundTotal glucosides of peony (TGP), extracted from the root and rhizome of Paeonia lactiflora Pall, has well-confirmed immunomodulatory efficacy in the clinic. However, the mechanism and active ingredients remain largely unclear.Hypothesis/PurposeOur previous study revealed a low systemic exposure but predominant gut distribution of TGP components. The aim of this study was to investigate involvement of the gut microbiota in the immunoregulatory effects and identify the active component.MethodsMice received 3% DSS to establish a model of colitis. The treatment group received TGP or single paeoniflorin (PF) or albiflorin (AF). Body weight, colon length, inflammatory and histological changes were assessed. Gut microbiota structure was profiled by 16s rRNA sequencing. Antibiotic treatment and fecal transplantation were used to explore the involvement of gut microbiota. Metabolomic assay of host and microbial metabolites in colon was performed.ResultsTGP improved colonic injury and gut microbial dysbiosis in colitis mice, and PF was responsible for the protective effects. Fecal microbiota transfer from TGP-treated mice conferred resilience to colitis, while antibiotic treatment abrogated the protective effects. Both TGP and PF decreased colonic indole-3-lactate (ILA), a microbial tryptophan metabolite. ILA was further identified as an inhibitor of epithelial autophagy and ILA supplementation compromised the benefits of TGP.ConclusionOur findings suggest that TGP acts in part through a gut microbiota-ILA-epithelial autophagy axis to alleviate colitis.     

Monitoring horizontal antibiotic resistance gene transfer in a colonic fermentation model

The human microbiota is suggested to be a reservoir of antibiotic resistance (ABR) genes, which are exchangeable between transient colonizers and residing bacteria. In this study, the transfer of ABR genes from Enterococcus faecalis to Listeria monocytogenes and to commensal bacteria of the human gut microbiota was demonstrated in a colonic fermentation model. In the first fermentation, an E. faecalis donor harboring the marked 50-kb conjugative plasmid pRE25(*) and a chromosomal marker was co-immobilized with L. monocytogenes and infant feces. In this complex environment, the transfer of pRE25(*) to L. monocytogenes was observed. In a second fermentation, only the E. faecalis donor and feces were co-immobilized. Enumeration of pRE25(*) and the donor strain by quantitative PCR revealed an increasing ratio of pRE25(*) to the donor throughout the 16-day fermentation, indicating the transfer of pRE25(*) . An Enterococcus avium transconjugant was isolated, demonstrating that ABR gene transfer to gut commensals occurred. Moreover, pRE25(*) was still functional in both the E. avium and the L. monocytogenes transconjugant and transmittable to other genera in filter mating experiments. Our study reveals that the transfer of a multiresistance plasmid to commensal bacteria in the presence of competing fecal microbiota occurs in a colonic model, suggesting that commensal bacteria contribute to the increasing prevalence of antibiotic-resistant bacteria.     

长链菊粉对抗生素处理后小鼠肠道菌群重建的影响

【目的】评价长链菊粉对抗生素致小鼠肠道菌群失调后肠道菌群的恢复情况。【方法】选择50只健康的10周龄BALB/c小鼠,随机分为2组,其中15只为正常对照组,余下35只饮用水中含4种抗生素连续喂养7 d,诱导小鼠肠道菌群严重失调后,再随机分为长链菊粉恢复组(饮用水中添加5%(W/W)长链菊粉)和自发恢复组(饮用水为无菌水),连续处理21 d。受试小鼠在抗生素治疗后的第7天以及恢复喂养的第7、14和21天,取结肠组织进行切片然后进行HE染色分析,无菌取粪便进行16Sr RNA测序分析,观察小鼠肠道组织及菌群恢复情况。【结果】抗生素处理7 d后,小鼠结肠组织有轻微炎症,但肠道菌群严重失调。组织学分析表明,在补充长链菊粉或自发恢复21d后,结肠炎症逐渐减轻;但相比于自发恢复,长链菊粉干预延迟了结肠组织的恢复。16S r RNA基因V3–V4区扩增子测序分析显示无论是长链菊粉补充还是自发恢复都无法在属水平上恢复肠道菌群组成。尤其是长链菊粉的补充,反而导致了某些机会致病菌的选择性扩增,并提高了与肠道菌群相关的疾病途径。【结论】抗生素诱导肠道菌群严重失调后补充长链菊粉会延迟肠道菌群的重建,可能会导致潜在的不良影响。     

Empagliflozin ameliorates type 2 diabetes mellitus-related diabetic nephropathy via altering the gut microbiota

BackgroundThe dysregulation of gut microbiota can be found in patients with type 2 diabetes mellitus (T2DM)-related diabetic nephropathy (DN). Inhibitors of sodium-glucose co-transporter 2 (SGLT2) were reported to affect gut microbiota. This study aimed to identify whether empagliflozin (EMPA) attenuated DN via regulating gut microbiota.Materials and methodsThe high-fat diet (HFD) combining streptozocin (STZ) injection was performed to induce DN in mice. The therapeutic effects of EMPA were observed by staining of renal tissues and urine albumin/creatinine ratio (UACR). Mouse feces were collected for 16S rRNA sequencing. Fecal short-chain fatty acids (SCFAs) and fecal and serum lipopolysaccharide (LPS) were determined. An antibiotic-ablated model was established to confirm the role of the gut microbiota in the actions of EMPA.ResultsEMPA reduced the elevation of blood glucose and UACR caused by HFD/STZ. It inhibited the thickening of the colonic crypt and restored goblet cells and the expressions of ZO-1 and Occludin. The 16S rRNA sequencing showed that the diversity of gut microbiota was reduced after HFD/STZ treatment, while it was restored after EMPA treatment. The LPS-producing bacteria, Oscillibacter, and the SCFA-producing bacteria, Bateroid and Odoribacter, were changed after EMPA administration. The therapeutic effects of EMPA on ABX-treated mice were reduced. Meanwhile, the level of fecal SCFAs was decreased, while the levels of fecal and serum LPS were elevated, in T2DM mice, and they were negated by the administration of EMPA.ConclusionEMPA ameliorates T2DM-related DN via altering the gut microbiota, especially reducing LPS-producing bacteria and increasing SCFA-producing bacteria.     

Increased Expression of Colonic Mucosal Melatonin in Patients with Irritable Bowel Syndrome Correlated with Gut Dysbiosis

Dysregulation of the gut microbiota/gut hormone axis contributes to the pathogenesis of irritable bowel syndrome (IBS). Melatonin plays a beneficial role in gut motility and immunity. However, altered expression of local mucosal melatonin in IBS and its relationship with the gut microbiota remain unclear. Therefore, we aimed to detect the colonic melatonin levels and microbiota profiles in patients with diarrhea-predominant IBS (IBS-D) and explore their relationship in germ-free (GF) rats and BON-1 cells. Thirty-two IBS-D patients and twenty-eight healthy controls (HCs) were recruited. Fecal specimens from IBS-D patients and HCs were separately transplanted into GF rats by gavage. The levels of colon mucosal melatonin were assessed by immunohistochemical methods, and fecal microbiota communities were analyzed using 16S rDNA sequencing. The effect of butyrate on melatonin synthesis in BON-1 cells was evaluated by ELISA. Melatonin levels were significantly increased and negatively correlated with visceral hypersensitivity in IBS-D patients. GF rats inoculated with fecal microbiota from IBS-D patients had high colonic melatonin levels. Butyrate-producing Clostridium cluster XIVa species, such as Roseburia species and Lachnospira species, were positively related to colonic mucosal melatonin expression. Butyrate significantly increased melatonin secretion in BON-1 cells. Increased melatonin expression may be an adaptive protective mechanism in the development of IBS-D. Moreover, some Clostridium cluster XIVa species could increase melatonin expression via butyrate production. Modulation of the gut hormone/gut microbiota axis offers a promising target of interest for IBS in the future.     

(S)-雌马酚对肠道菌群的体外调控作用及其可代谢性研究

[目的]研究(S)-雌马酚对人体肠道菌群的体外调控作用和人体肠道菌群对(S)-雌马酚的代谢衍生作用。[方法]采用人体肠道菌群体外批量发酵、细菌16S rRNA基因高通量测序、气相色谱、液相色谱和质谱等检测(S)-雌马酚与人体肠道菌群体外相互作用。[结果]体外添加(S)-雌马酚对总体人肠道菌群结构和短链脂肪酸产量影响不明显。添加0.45 mmol/L (S)-雌马酚组与对照组相比,未检测到相对丰度发生显著变化的细菌;添加0.90 mmol/L (S)-雌马酚组与对照组相比,显著增加了肠杆菌科(Enterobacteriaceae)等条件致病菌的相对丰度,减少了潜在益生菌粪球菌属(Coprococcus)的比例。代谢分析发现,发酵培养液中(S)-雌马酚的浓度降低了约15%−30%,推测可能被微生物进一步降解或衍生修饰。[结论]从体外调控肠道菌群的角度判断,0.45 mmol/L (S)-雌马酚相对较安全,而0.90 mmol/L (S)-雌马酚可能会破坏肠道菌群平衡。(S)-雌马酚可以被人体肠道菌群进一步代谢,其特定代谢产物的结构与功能及其体内生物安全性有待进一步研究。