ABSTRACT: Intestinal phages are abundant and important components of gut microbiota, yet the isolated and characterized representatives that infect abundant gut bacteria are sparse. Here we describe the isolation of human intestinal phages infecting Bacteroidesuniformis. Bacteroides is one of the most common bacterial groups in the global human gut microbiota; however, to date not many Bacteroides specific phages are known. Phages isolated in this study belong to a novel viral genus, Bacuni, within the Siphoviridae family. Their genomes encode diversity-generating retroelements (DGR), which were shown in other bacteriophages to promote phage adaptation to rapidly changing environmental conditions and to broaden their host range. Three isolated phages showed 99.83% genome identity but one of them infected a distinct B. uniformis strain. The tropism of Bacuni phages appeared to be dependent on the interplay of DGR mediated sequence variations of gene encoding putative phage fimbrial tip proteins and mutations in host genes coding for outer-membrane proteins. We found prophages with up to 85% amino acid similarity over two-thirds of the Bacuni phage genome in the B. acidifaciens and Prevotella sp. genomes. Despite the abundance of Bacteroides within the human microbiome, we found Bacuni phages only in a limited subset of published gut metagenomes.
Project description:BACKGROUND:Diversity-generating retroelements (DGRs) are genetic cassettes that selectively mutate target genes to produce hypervariable proteins. First characterized in Bordetella bacteriophage BPP-1, the DGR creates a hypervariable phage tail fiber that enables host tropism switching. Subsequent surveys for DGRs conclude that the majority identified to date are bacterial or archaeal in origin. This work examines bacteriophage and bacterial genomes for novel phage-encoded DGRs. RESULTS:This survey discovered 92 DGRs that were only found in phages exhibiting a temperate lifestyle. The majority of phage-encoded DGRs were identified as prophages in bacterial hosts from the phyla Bacteroidetes, Proteobacteria, and Firmicutes. Sequence reads from these previously unidentified prophages were present in viral metagenomes (viromes), indicating these prophages can produce functional viruses. Five phages possessed hypervariable proteins with structural similarity to the tail fiber of BPP-1, whereas the functions of the remaining DGR target proteins were unknown. A novel temperate phage that harbors a DGR cassette targeting a protein of unknown function was induced from Bacteroides dorei. This phage, here named Bacteroides dorei Hankyphage, lysogenizes 13 different Bacteroides species and was present in 34% and 21% of whole-community metagenomes and human-associated viromes, respectively. CONCLUSIONS:Here, the number of known DGR-containing phages is increased from four to 92. All of these phages exhibit a temperate lifestyle, including a cosmopolitan human-associated phage. Targeted hypervariation by temperate phages may be a ubiquitous mechanism underlying phage-bacteria interaction in the human microbiome.
Project description:Bacteroides spp. are dominant components of the phylum Bacteroidetes in the gut microbiota and prosper in glycan enriched environments. However, knowledge of the machinery of specific species isolated from humans (like Bacteroides uniformis) contributing to the utilization of dietary and endogenous sources of glycans and their byproducts is limited. We have used the cutting-edge nanopore-based technology to sequence the genome of B. uniformis CECT 7771, a human symbiont with a proven pre-clinical efficacy on metabolic and immune dysfunctions in obesity animal models. We have also used massive sequencing approaches to distinguish the genome expression patterns in response to carbon sources of different complexity during growth. At genome-wide level, our analyses globally demonstrate that B. uniformis strains exhibit an expanded glycolytic capability when compared with other Bacteroides species. Moreover, by studying the growth and whole-genome expression of B. uniformis CECT 7771 in response to different carbon sources, we detected a differential growth fitness and expression patterns across the genome depending on the carbon source of the culture media. The dietary fibers used exerted different effects on B. uniformis CECT 7771 activating different molecular pathways and, therefore, allowing the production of different metabolite types with potential impact on gut health. The genome and transcriptome analysis of B. uniformis CECT 7771, in response to different carbon sources, shows its high versatility to utilize both dietary and endogenous glycans along with the production of potentially beneficial end products for both the bacterium and the host, pointing to a mechanistic basis of a mutualistic relationship.
Project description:This study investigated the immune mechanisms whereby administration of Bacteroides uniformis CECT 7771 reduces metabolic dysfunction in obesity. C57BL/6 adult male mice were fed a standard diet or a Western diet high in fat and fructose, supplemented or not with B. uniformis CECT 7771 for 14 weeks. B. uniformis CECT 7771 reduced body weight gain, plasma cholesterol, triglyceride, glucose, and leptin levels; and improved oral glucose tolerance in obese mice. Moreover, B. uniformis CECT 7771 modulated the gut microbiota and immune alterations associated with obesity, increasing Tregs and reducing B cells, total macrophages and the M1/M2 ratio in both the gut and epididymal adipose tissue (EAT) of obese mice. B. uniformis CECT 7771 also increased the concentration of the anti-inflammatory cytokine IL-10 in the gut, EAT and peripheral blood, and protective cytokines TSLP and IL-33, involved in Treg induction and type 2 innate lymphoid cells activation, in the EAT. It also restored the obesity-reduced TLR5 expression in the ileum and EAT. The findings indicate that the administration of a human intestinal bacterium with immunoregulatory properties on the intestinal mucosa helps reverse the immuno-metabolic dysfunction caused by a Western diet acting over the gut-adipose tissue axis.
Project description:Recent studies underscore important roles of intestinal microbiota and the bacterial lipopolysaccharides (LPS) production in the pathogenesis of liver disease. However, how gut microbiota alters in response to the development of steatosis and subsequent progression to nonalcoholic steatohepatitis (NASH) and hepatocellular carcinoma (HCC) remains unclear. We aimed to study the gut microbial changes over liver disease progression using a streptozotocin-high fat diet (STZ-HFD) induced NASH-HCC C57BL/6J mouse model that is highly relevant to human liver disease. The fecal microbiota at various liver pathological stages was analyzed by 16S rDNA gene pyrosequencing. Both UniFrac analysis and partial least squares-discriminant analysis showed significant structural alterations in gut microbiota during the development of liver disease. Co-abundance network analysis highlighted relationships between genera. Spearman correlation analysis revealed that the bacterial species, Atopobium spp., Bacteroides spp., Bacteroides vulgatus, Bacteroides acidifaciens, Bacteroides uniformis, Clostridium cocleatum, Clostridium xylanolyticum and Desulfovibrio spp., markedly increased in model mice, were positively correlated with LPS levels and pathophysiological features. Taken together, the results showed that the gut microbiota was altered significantly in the progression of liver disease. The connection between the gut microbial ecology and the liver pathology may represent potential targets for the prevention and treatment of chronic liver disease and HCC.
Project description:Dietary habits and gut microbiota play an essential role in non-alcoholic fatty liver disease (NAFLD) and related factors such as insulin resistance and de novo lipogenesis. In this study, we investigated the protective effects of <i>Bacteroides uniformis</i> CBA7346, isolated from the gut of healthy Koreans, on mice with high-fat diet (HFD)-induced NAFLD. Administration of <i>B. uniformis</i> CBA7346 reduced body and liver weight gain, serum alanine aminotransferase and aspartate aminotransferase levels, liver steatosis, and liver triglyceride levels in mice on an HFD; the strain also decreased homeostatic model assessment for insulin resistance values, as well as serum cholesterol, triglyceride, lipopolysaccharide, leptin, and adiponectin levels in mice on an HFD. Moreover, <i>B. uniformis</i> CBA7346 controlled fatty liver disease by attenuating steatosis and inflammation and regulating de novo lipogenesis-related proteins in mice on an HFD. Taken together, these findings suggest that <i>B. uniformis</i> CBA7346 ameliorates HFD-induced NAFLD by reducing insulin resistance and regulating de novo lipogenesis in obese mice.
Project description:Gut-associated phages are hypothesized to alter the abundance and activity of their bacterial hosts, contributing to human health and disease. Although temperate phages constitute a significant fraction of the gut virome, the effects of lysogenic infection are underexplored. We report that the temperate phage, Bacteroides phage BV01, broadly alters its host's transcriptome, the prominent human gut symbiont Bacteroides vulgatus. This alteration occurs through phage-induced repression of a tryptophan-rich sensory protein (TspO) and represses bile acid deconjugation. Because microbially modified bile acids are important signals for the mammalian host, this is a mechanism by which a phage may influence mammalian phenotypes. Furthermore, BV01 and its relatives in the proposed phage family Salyersviridae are ubiquitous in human gut metagenomes, infecting a broad range of Bacteroides hosts. These results demonstrate the complexity of phage-bacteria-mammal relationships and emphasize a need to better understand the role of temperate phages in the gut microbiome.
Project description:Immunoglobulin A (IgA) is essential for defense of the intestinal mucosa against harmful pathogens. Previous studies have shown that Bacteroidetes, the major phylum of gut microbiota together with Firmicutes, impact IgA production. However, the relative abundances of species of Bacteroidetes responsible for IgA production were not well understood. In the present study, we identified some specific Bacteroidetes species that were associated with gut IgA induction by hsp60-based profiling of species distribution among Bacteroidetes The levels of IgA and the expression of the gene encoding activation-induced cytidine deaminase (AID) in the large intestine lamina propria, which is crucial for class switch recombination from IgM to IgA, were increased in soluble high-fiber diet (sHFD)-fed mice. We found that Bacteroides acidifaciens was the most abundant Bacteroidetes species in both sHFD- and normal diet-fed mice. In addition, the gut IgA levels were associated with the relative abundance of Bacteroides fragilis group species such as Bacteroides faecis, Bacteroides caccae, and Bacteroides acidifaciens Conversely, the ratio of B. acidifaciens to other Bacteroidetes species was reduced in insoluble high-fiber diet fed- and no-fiber diet-fed mice. To investigate whether B. acidifaciens increases IgA production, we generated B. acidifaciens monoassociated mice and found increased gut IgA production and AID expression. Collectively, soluble dietary fiber increases the ratio of gut Bacteroides fragilis group, such as B. acidifaciens, and IgA production. This might improve gut immune function, thereby protecting against bowel pathogens and reducing the incidence of inflammatory bowel diseases.IMPORTANCE Immunoglobulin A (IgA) is essential for defense of the intestinal mucosa against harmful pathogens. Gut microbiota impact IgA production, but the specific species responsible for IgA production remain largely elusive. Previous studies have shown that IgA and Bacteroidetes, the major phyla of gut microbiota, were increased in soluble high-fiber diet-fed mice. We show here that the levels of IgA in the gut and the expression of activation-induced cytidine deaminase (AID) in the large intestine lamina propria, which is crucial for class switch recombination from IgM to IgA, were correlated with the abundance of Bacteroides fragilis group species such as Bacteroides faecis, Bacteroides caccae, and Bacteroides acidifaciens B. acidifaciens monoassociated mice increased gut IgA production and AID expression. Soluble dietary fiber may improve gut immune function, thereby protecting against bowel pathogens and reducing inflammatory bowel diseases.
Project description:<h4>Background</h4>Associations have been made between obesity and reduced intestinal numbers of members of the phylum Bacteroidetes, but there is no direct evidence of the role these bacteria play in obesity. Herein, the effects of Bacteroides uniformis CECT 7771 on obesity-related metabolic and immune alterations have been evaluated.<h4>Methods and findings</h4>Adult (6-8 week) male wild-type C57BL-6 mice were fed a standard diet or a high-fat-diet HFD to induce obesity, supplemented or not with B. uniformis CECT 7771 for seven weeks. Animal weight was monitored and histologic, biochemical, immunocompetent cell functions, and features of the faecal microbiota were analysed after intervention. The oral administration of B. uniformis CECT 7771 reduced body weight gain, liver steatosis and liver cholesterol and triglyceride concentrations and increased small adipocyte numbers in HFD-fed mice. The strain also reduced serum cholesterol, triglyceride, glucose, insulin and leptin levels, and improved oral tolerance to glucose in HFD fed mice. The bacterial strain also reduced dietary fat absorption, as indicated by the reduced number of fat micelles detected in enterocytes. Moreover, B. uniformis CECT 7771 improved immune defence mechanisms, impaired in obesity. HFD-induced obesity led to a decrease in TNF-? production by peritoneal macrophages stimulated with LPS, conversely, the administration of B. uniformis CECT 7771 increased TNF-? production and phagocytosis. Administering this strain also increased TNF-? production by dendritic cells (DCs) in response to LPS stimulation, which was significantly reduced by HFD. B. uniformis CECT 7771 also restored the capacity of DCs to induce a T-cell proliferation response, which was impaired in obese mice. HFD induced marked changes in gut microbiota composition, which were partially restored by the intervention.<h4>Conclusions</h4>Altogether, the findings indicate that administration of B. uniformis CECT 7771 ameliorates HFD-induced metabolic and immune dysfunction associated with intestinal dysbiosis in obese mice.
Project description:The formulation of next-generation probiotics requires competent preclinical studies to show their efficacy and safety status. This study aims to confirm the safety of the prolonged oral use of <i>Bacteroides uniformis</i> CECT 7771, a strain that protected against metabolic disorders and obesity in preclinical trials, in a sub-chronic 90 day trial in animals. The safety assessment was conducted in male and female Wistar rats (<i>n</i> = 50) administered increasing doses (10<sup>8</sup> CFU/day, 10<sup>9</sup> CFU/day, or 10<sup>10</sup> CFU/day) of <i>B. uniformis</i> CECT 7771, 10<sup>10</sup> CFU/day of <i>B. longum</i> ATCC 15707<sup>T</sup>, which complies with the qualifying presumption of safety (QPS) status of the EU, or vehicle (placebo), as the control. Pancreatic, liver, and kidney functions and cytokine concentrations were analyzed. Bacterial translocation to peripheral tissues was evaluated, and colon integrity was investigated histologically. No adverse metabolic or tissue integrity alterations were associated with treatments; however, alanine aminotransferase levels and the ratio of anti-inflammatory to pro-inflammatory cytokines in serum indicated a potentially beneficial role of <i>B. uniformis</i> CECT 7771 at specific doses. Additionally, the microbial community structure was modified by the interventions, and potentially beneficial gut bacteria were increased. The results indicated that the oral consumption of <i>B. uniformis</i> CECT 7771 during a sub-chronic 90 day study in rats did not raise safety concerns.
Project description:Our emerging view of the gut microbiome largely focuses on bacteria, while less is known about other microbial components, such as bacteriophages (phages). Though phages are abundant in the gut, very few phages have been isolated from this ecosystem. Here, we report the genomes of 27 phages from the United States and Bangladesh that infect the prevalent human gut bacterium Bacteroides thetaiotaomicron. These phages are mostly distinct from previously sequenced phages with the exception of two, which are crAss-like phages. We compare these isolates to existing human gut metagenomes, revealing similarities to previously inferred phages and additional unexplored phage diversity. Finally, we use host tropisms of these phages to identify alleles of phage structural genes associated with infectivity. This work provides a detailed view of the gut's "viral dark matter" and a framework for future efforts to further integrate isolation- and sequencing-focused efforts to understand gut-resident phages.