Project description:We compared gene expression in the small intestine (ileum) of mice that were either (i) germ-free, (ii) colonized with a conventional mouse cecal microbiota, (iii) colonized with a conventional zebrafish gut microbiota, or (iv) colonized with Pseudomonas aeruginosa PAO1. Experiment Overall Design: Adult germ-free NMRI mice were colonized with either (i) a conventional mouse cecal microbiota harvested from adult Swiss-Webster mice (5 biological replicates), (ii) a conventional zebrafish intestinal microbiota harvested from adult C32 zebrafish (3 biological replicates), or (iii) a culture of Pseudomonas aeruginosa PAO1 (5 biological replicates). 14 days after colonization, total RNA was prepared from the ileum of each animal, with total RNA prepared from adult germ-free NMRI mouse ileum serving as negative controls (5 biological replicates). RNA was used as template to generate cRNA for hybridization to Affymetrix 430 v2 Mouse GeneChips.
Project description:Advanced age is associated with chronic low-grade inflammation, which is usually referred to as inflammaging. Elderly are also known to have an altered gut microbiota composition. However, whether inflammaging is a cause or consequence of an altered gut microbiota composition is not clear. In this study gut microbiota from young or old conventional mice was transferred to young germ-free mice. Four weeks after gut microbiota transfer immune cell populations in spleen, Peyer’s patches, and mesenteric lymph nodes from conventionalized germ-free mice were analyzed by flow cytometry. In addition, whole-genome gene expression in the ileum was analyzed by microarray. Gut microbiota composition of donor and recipient mice was analyzed with 16S rDNA sequencing. Here we show by transferring aged microbiota to young germ-free mice that certain bacterial species within the aged microbiota promote inflammaging. This effect was associated with lower levels of Akkermansia and higher levels of TM7 bacteria and Proteobacteria in the aged microbiota after transfer. The aged microbiota promoted inflammation in the small intestine in the germ-free mice and enhanced leakage of inflammatory bacterial components into the circulation was observed. Moreover, the aged microbiota promoted increased T cell activation in the systemic compartment. In conclusion, these data indicate that the gut microbiota from old mice contributes to inflammaging after transfer to young germ-free mice. Overall design: Gut microbiota from young or old conventional mice was transferred to young germ-free mice. Four weeks after gut microbiota transfer whole-genome gene expression in the distal ileum was analyzed by microarray.
Project description:Gut microbiome research is rapidly moving towards the functional characterization of the microbiota by means of shotgun meta-omics. Here, we selected a cohort of healthy subjects from an indigenous and monitored Sardinian population to analyze their gut microbiota using both shotgun metagenomics and shotgun metaproteomics. We found a considerable divergence between genetic potential and functional activity of the human healthy gut microbiota, in spite of a quite comparable taxonomic structure revealed by the two approaches. Investigation of inter-individual variability of taxonomic features revealed Bacteroides and Akkermansia as remarkably conserved and variable in abundance within the population, respectively. Firmicutes-driven butyrogenesis (mainly due to Faecalibacterium spp.) was shown to be the functional activity with the higher expression rate and the lower inter-individual variability in the study cohort, highlighting the key importance of the biosynthesis of this microbial by-product for the gut homeostasis. The taxon-specific contribution to functional activities and metabolic tasks was also examined, giving insights into the peculiar role of several gut microbiota members in carbohydrate metabolism (including polysaccharide degradation, glycan transport, glycolysis and short-chain fatty acid production). In conclusion, our results provide useful indications regarding the main functions actively exerted by the gut microbiota members of a healthy human cohort, and support metaproteomics as a valuable approach to investigate the functional role of the gut microbiota in health and disease.
Project description:Mardinoglu2015 - Generic mouse genome-scale
metabolic network (MMR)
This model is described in the article:
The gut microbiota modulates
host amino acid and glutathione metabolism in mice.
Mardinoglu A, Shoaie S, Bergentall
M, Ghaffari P, Zhang C, Larsson E, Bäckhed F, Nielsen
Mol. Syst. Biol. 2015; 11(10):
The gut microbiota has been proposed as an environmental
factor that promotes the progression of metabolic diseases.
Here, we investigated how the gut microbiota modulates the
global metabolic differences in duodenum, jejunum, ileum,
colon, liver, and two white adipose tissue depots obtained from
conventionally raised (CONV-R) and germ-free (GF) mice using
gene expression data and tissue-specific genome-scale metabolic
models (GEMs). We created a generic mouse metabolic reaction
(MMR) GEM, reconstructed 28 tissue-specific GEMs based on
proteomics data, and manually curated GEMs for small intestine,
colon, liver, and adipose tissues. We used these functional
models to determine the global metabolic differences between
CONV-R and GF mice. Based on gene expression data, we found
that the gut microbiota affects the host amino acid (AA)
metabolism, which leads to modifications in glutathione
metabolism. To validate our predictions, we measured the level
of AAs and N-acetylated AAs in the hepatic portal vein of
CONV-R and GF mice. Finally, we simulated the metabolic
differences between the small intestine of the CONV-R and GF
mice accounting for the content of the diet and relative gene
expression differences. Our analyses revealed that the gut
microbiota influences host amino acid and glutathione
metabolism in mice.
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Project description:Age-dependent changes of the gut-associated microbiome have been linked to increased frailty and systemic inflammation. This study found that age-associated changes of the gut microbiome of BALB/c and C57BL/6 mice could be reverted by co-housing of aged (22 months old) and adult (3 months old) mice for 30-40 days or faecal microbiota transplantation (FMT) from adult into aged mice. This was demonstrated using high-throughput sequencing of the V3-V4 hypervariable region of bacterial 16S rRNA gene isolated from faecal pellets collected from 3-4 months old adult and 22-23 months old aged mice before and after co-housing or FMT.
Project description:The mouse stool samples were collected from different diets fed mice and bacterial cells were harvest for metaproteomic analysis for understanding the role ofdiet on gut microbiota.
Project description:Microbial functions in the host physiology are a result of co-evolution between microbial communities and their hosts. Here we show that cold exposure leads to marked shift of the microbiota composition, referred to as cold microbiota. Transplantation of the cold microbiota to germ-free mice is sufficient to increase the insulin sensitivity of the host, and enable complete tolerance to cold partly by promoting the white fat browning, leading to increased energy expenditure and fat loss. During prolonged cold however, the body weight loss is attenuated, caused by adaptive mechanisms maximising caloric uptake and increasing intestinal, villi and microvilli lengths. This increased absorptive surface is promoted by the cold microbiota - effect that can be diminished by co-transplanting the most downregulated bacterial strain from the Verrucomicrobia phylum, Akkermansia muciniphila, during the cold microbiota transfer. Our results demonstrate the microbiota as a key factor orchestrating the overall energy homeostasis during increased demand. Overall design: C57BL/6J mice were put for 31 day on cold (6C) or room temperature. Fresh feces and cecum samples were collected, immediately frozen and stored. Bacterial DNA content was extracted using QIAamp Fast DNA stool Mini Kit (Qiagen). Bacterial DNA was PCR amplified with barcoded universal bacterial primers targeting variable regionV4 of 16SrRNA gene. Samples were pooled and sequenced with Ilumina MiSeq platform. Using QIIME and custom scripts, sequences were quality filtered and demultiplexed using exact matches to the supplied DNA barcodes. Resulting sequences were then searched against the Greengenes reference database of 16S rRNA gene sequences, clustered at 97% by uclust. The longest sequence from each Operation Taxonomic Unit (OTU) thus formed was then considered as the OTU representative sequence, and assigned taxonomic classification via Mothur's Bayesian classifier, trained against the Greengenes database clustered at 98%. Each listed sample is indivudual biological replicate.
Project description:We analyzed the effects of antibiotics using a popular model of gut microbiota depletion in mice by a cocktail of antibiotics. We combined intestinal transcriptome together with metagenomic analysis of the gut microbiota to develop a new bioinformatics approach that probes the links between microbial components and host functions. We found that most antibiotic-induced alterations can be explained by three factors: depletion of the microbiota; direct effects of antibiotics on host tissues; and the effects of remaining antibiotic-resistant microbes. While microbe depletion led to down-regulation of immunity, the two other factors primarily inhibited mitochondrial gene expression and amounts of active mitochondria, and induced cell death. By reconstructing and analyzing a transkingdom network, we discovered that these toxic effects were mediated by virulence/quorum sensing in antibiotic-resistant bacteria. This series includes gene expression in the ileum of control, antibiotics (ABx)-treated, germfree, germfree-ABx-treated and mice colonized with normal or Abx-resistant microbiota. common reference design with a pool of small intestine RNA labeled with Cy3