A 1000-cow study across four European countries was undertaken to understand to what extent ruminant microbiomes can be controlled by the host animal and to identify characteristics of the host rumen microbiome axis that determine productivity and methane emissions. A core rumen microbiome, phylogenetically linked and with a preserved hierarchical structure, was identified. A 39-member subset of the core formed hubs in co-occurrence networks linking microbiome structure to host genetics and phenotype (methane emissions, rumen and blood metabolites, and milk production efficiency). These phenotypes can be predicted from the core microbiome using machine learning algorithms. The heritable core microbes, therefore, present primary targets for rumen manipulation toward sustainable and environmentally friendly agriculture.
- MeSH
- bachor metabolismus MeSH
- fenotyp MeSH
- fylogeneze MeSH
- kohortové studie MeSH
- krev metabolismus MeSH
- methan metabolismus MeSH
- mléko metabolismus MeSH
- skot genetika mikrobiologie MeSH
- střevní mikroflóra genetika fyziologie MeSH
- zvířata MeSH
- Check Tag
- skot genetika mikrobiologie MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Background: The rumen microbiota is one of the most complex consortia of anaerobes, involving archaea, bacteria, protozoa, fungi and phages. They are very effective at utilizing plant polysaccharides, especially cellulose and hemicelluloses. The most important hemicellulose decomposers are clustered with the genus Butyrivibrio. As the related species differ in their range of hydrolytic activities and substrate preferences, Butyrivibrio fibrisolvens was selected as one of the most effective isolates and thus suitable for proteomic studies on substrate comparisons in the extracellular fraction. The B. fibrisolvens genome is the biggest in the butyrivibria cluster and is focused on "environmental information processing" and "carbohydrate metabolism". Methods: The study of the effect of carbon source on B. fibrisolvens 3071 was based on cultures grown on four substrates: xylose, glucose, xylan, xylan with 25% glucose. The enzymatic activities were studied by spectrophotometric and zymogram methods. Proteomic study was based on genomics, 2D electrophoresis and nLC/MS (Bruker Daltonics) analysis. Results: Extracellular β-endoxylanase as well as xylan β-xylosidase activities were induced with xylan. The presence of the xylan polymer induced hemicellulolytic enzymes and increased the protein fraction in the interval from 40 to 80 kDa. 2D electrophoresis with nLC/MS analysis of extracellular B. fibrisolvens 3071 proteins found 14 diverse proteins with significantly different expression on the tested substrates. Conclusion: The comparison of four carbon sources resulted in the main significant changes in B. fibrisolvens proteome occurring outside the fibrolytic cluster of proteins. The affected proteins mainly belonged to the glycolysis and protein synthesis cluster.
- Publikační typ
- časopisecké články MeSH
The diversity of the methanogenic archaea associated with the six segments of the horse and donkey hindgut (caecum, right ventral colon, left ventral colon, left dorsal colon, right dorsal colon, and rectum) was analyzed using 16S rDNA gene clone library. A total of 641 positive clones, 321 from the horse and 320 from the donkey hindgut, were examined by the RFLP, revealing 9 different ribotypes, 8 in the horse and 5 in the donkey hindgut. In both the animals Methanobacteriales (Methanobrevibacter-like sequences) and Methanomicrobiales (Methanocorpusculum-like sequences) were detected as the dominant orders followed by the uncultured Methanomassiliicoccales. The composition of the equine archaeal community was found to be dependent on the gut region. In both the two animals no Methanobrevibacter-like clones were detected in the caeca, which were instead inhabited by the Methanocorpusculum-like archeons. The Methanosarcinales were found only in distal regions of the horse hindgut.
- MeSH
- Archaea klasifikace genetika izolace a purifikace metabolismus MeSH
- biodiverzita MeSH
- DNA archebakterií genetika MeSH
- Equidae mikrobiologie MeSH
- fylogeneze MeSH
- koně mikrobiologie MeSH
- methan metabolismus MeSH
- ribozomální DNA genetika MeSH
- tlusté střevo mikrobiologie MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- srovnávací studie MeSH
High-grain feeding used in the animal production is known to affect the host rumen bacterial community, but our understanding of consequent changes in goats is limited. This study was therefore aimed to evaluate bacterial population dynamics during 20 days adaptation of 4 ruminally cannulated goats to the high-grain diet (grain: hay - ratio of 40:60). The dietary transition of goats from the forage to the high-grain-diet resulted in the significant decrease of rumen fluid pH, which was however still higher than value established for acute or subacute ruminal acidosis was not diagnosed in studied animals. DGGE analysis demonstrated distinct ruminal microbial populations in hay-fed and grain-fed animals, but the substantial animal-to-animal variation were detected. Quantitative PCR showed for grain-fed animals significantly higher number of bacteria belonging to Clostridium leptum group at 10 days after the incorporation of corn into the diet and significantly lower concentration of bacteria belonging to Actinobacteria phylum at the day 20 after dietary change. Taxonomic distribution analysed by NGS at day 20 revealed the similar prevalence of the phyla Firmicutes and Bacteroidetes in all goats, significantly higher presence of the unclassified genus of groups of Bacteroidales and Ruminococcaceae in grain-fed animals and significantly higher presence the genus Prevotella and Butyrivibrio in the forage-fed animals. The three different culture-independent methods used in this study show that high proportion of concentrate in goat diet does not induce any serious disturbance of their rumen ecosystem and indicate the good adaptive response of caprine ruminal bacteria to incorporation of corn into the diet.
- MeSH
- Actinobacteria klasifikace genetika metabolismus MeSH
- bachor mikrobiologie MeSH
- Bacteroidetes klasifikace genetika metabolismus MeSH
- Butyrivibrio klasifikace genetika metabolismus MeSH
- Clostridium klasifikace genetika metabolismus MeSH
- fermentace MeSH
- Firmicutes klasifikace genetika metabolismus MeSH
- fylogeneze MeSH
- fyziologie výživy zvířat * MeSH
- koncentrace vodíkových iontů MeSH
- kozy MeSH
- krmivo pro zvířata analýza MeSH
- kukuřice setá chemie metabolismus MeSH
- lipnicovité chemie metabolismus MeSH
- píštěl žaludku MeSH
- Prevotella klasifikace genetika metabolismus MeSH
- Ruminococcus klasifikace genetika metabolismus MeSH
- sekvenční analýza DNA MeSH
- střevní mikroflóra fyziologie MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
Genes encoding glycosyl hydrolase family 11 (GH11) xylanases and xylanases have been identified from Pseudobutyrivibrio xylanivorans. In contrast, little is known about the diversity and distribution of the GH10 xylanase in strains of P. xylanivorans. Xylanase and associated activities of P. xylanivorans have been characterized in detail in the type strain, Mz5. The aim of the present study was to identify GH10 xylanase genes in strains 2 and Mz5 of P. xylanivorans. In addition, we evaluated degradation and utilization of xylan by P. xylanivorans 2 isolated from rumen of Creole goats. After a 12-h culture, P. xylanivorans 2 was able to utilize up to 53% of the total pentose content present in birchwood xylan (BWX) and to utilize up to 62% of a ethanol-acetic acid-soluble fraction prepared from BWX. This is the first report describing the presence of GH10 xylanase-encoding genes in P. xylanivorans. Strain 2 and Mz5 contained xylanases which were related to GH10 xylanase of Butyrivibrio sp. Identifying xylanase-encoding genes and activity of these enzymes are a step toward understanding possible functional role of P. xylanivorans in the rumen ecosystem and contribute to providing an improved choice of enzymes for improving fiber digestion in ruminant animals, agricultural biomass utilization for biofuel production, and other industries.
- MeSH
- bachor mikrobiologie MeSH
- Bacteria klasifikace enzymologie genetika izolace a purifikace MeSH
- bakteriální proteiny chemie genetika metabolismus MeSH
- endo-1,4-beta-xylanasy chemie genetika metabolismus MeSH
- fylogeneze MeSH
- kinetika MeSH
- kozy MeSH
- xylany metabolismus MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
The comparison of the bacterial profile of intracellular (iDNA) and extracellular DNA (eDNA) isolated from cow rumen content stored under different conditions was conducted. The influence of rumen fluid treatment (cheesecloth squeezed, centrifuged, filtered), storage temperature (RT, -80 °C) and cryoprotectants (PBS-glycerol, ethanol) on quality and quantity parameters of extracted DNA was evaluated by bacterial DGGE analysis, real-time PCR quantification and metabarcoding approach using high-throughput sequencing. Samples clustered according to the type of extracted DNA due to considerable differences between iDNA and eDNA bacterial profiles, while storage temperature and cryoprotectants additives had little effect on sample clustering. The numbers of Firmicutes and Bacteroidetes were lower (P < 0.01) in eDNA samples. The qPCR indicated significantly higher amount of Firmicutes in iDNA sample frozen with glycerol (P < 0.01). Deep sequencing analysis of iDNA samples revealed the prevalence of Bacteroidetes and similarity of samples frozen with and without cryoprotectants, which differed from sample stored with ethanol at room temperature. Centrifugation and consequent filtration of rumen fluid subjected to the eDNA isolation procedure considerably changed the ratio of molecular operational taxonomic units (MOTUs) of Bacteroidetes and Firmicutes. Intracellular DNA extraction using bead-beating method from cheesecloth sieved rumen content mixed with PBS-glycerol and stored at -80 °C was found as the optimal method to study ruminal bacterial profile.
- MeSH
- bachor mikrobiologie MeSH
- Bacteroidetes klasifikace genetika izolace a purifikace MeSH
- denaturační gradientová gelová elektroforéza MeSH
- DNA bakterií izolace a purifikace MeSH
- fylogeneze MeSH
- grampozitivní bakterie klasifikace genetika izolace a purifikace MeSH
- kryoprezervace * MeSH
- kvantitativní polymerázová řetězová reakce MeSH
- odběr biologického vzorku metody MeSH
- skot MeSH
- taxonomické DNA čárové kódování * MeSH
- vysoce účinné nukleotidové sekvenování MeSH
- zvířata MeSH
- Check Tag
- skot MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
BACKGROUND: Microbial sensing by Toll-like receptors (TLR) and its negative regulation have an important role in the pathogenesis of inflammation-related cancer. In this study, we investigated the role of negative regulation of Toll-like receptors signaling and gut microbiota in the development of colitis-associated cancer in mouse model. METHODS: Colitis-associated cancer was induced by azoxymethane and dextran sodium sulfate in wild-type and in interleukin-1 receptor-associated kinase M (IRAK-M)-deficient mice with or without antibiotic (ATB) treatment. Local cytokine production was analyzed by multiplex cytokine assay or enzyme-linked immunosorbent assay, and regulatory T cells were analyzed by flow cytometry. Changes in microbiota composition during tumorigenesis were analyzed by pyrosequencing, and β-glucuronidase activity was measured in intestinal content by fluorescence assay. RESULTS: ATB treatment of wild-type mice reduced the incidence and severity of tumors. Compared with nontreated mice, ATB-treated mice had significantly lower numbers of regulatory T cells in colon, altered gut microbiota composition, and decreased β-glucuronidase activity. However, the β-glucuronidase activity was not as low as in germ-free mice. IRAK-M-deficient mice not only developed invasive tumors, but ATB-induced decrease in β-glucuronidase activity did not rescue them from severe carcinogenesis phenotype. Furthermore, IRAK-M-deficient mice had significantly increased levels of proinflammatory cytokines in the tumor tissue. CONCLUSIONS: We conclude that gut microbiota promotes tumorigenesis by increasing the exposure of gut epithelium to carcinogens and that IRAK-M-negative regulation is essential for colon cancer resistance even in conditions of altered microbiota. Therefore, gut microbiota and its metabolic activity could be potential targets for colitis-associated cancer therapy.
- MeSH
- azoxymethan toxicita MeSH
- cytokiny genetika metabolismus MeSH
- gastrointestinální trakt mikrobiologie MeSH
- karcinogeny toxicita MeSH
- kinázy asociované s receptory interleukinu-1 fyziologie MeSH
- kolitida chemicky indukované komplikace MeSH
- kvantitativní polymerázová řetězová reakce MeSH
- messenger RNA genetika MeSH
- metagenom * MeSH
- myši inbrední C57BL MeSH
- myši knockoutované MeSH
- myši MeSH
- nádory tračníku etiologie metabolismus patologie MeSH
- polymerázová řetězová reakce s reverzní transkripcí MeSH
- průtoková cytometrie MeSH
- receptory interleukinu-1 metabolismus MeSH
- regulační T-lymfocyty imunologie metabolismus patologie MeSH
- signální transdukce MeSH
- síran dextranu toxicita MeSH
- toll-like receptory genetika metabolismus MeSH
- western blotting MeSH
- zvířata MeSH
- Check Tag
- mužské pohlaví MeSH
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
In our previous experiment, the ten calves originated bifidobacterial strains were administered to calves and re-isolated. Fingerprinting techniques used in this study enabled us to distinguish the surviving and non-surviving strains. Only the species Bifidobacterium animalis ssp. animalis and Bifidobacterium longum ssp. suis were found to survive in the intestine.
- MeSH
- bakteriální proteiny genetika MeSH
- Bifidobacterium klasifikace genetika izolace a purifikace MeSH
- chaperon hsp60 genetika MeSH
- feces mikrobiologie MeSH
- fylogeneze MeSH
- probiotika MeSH
- RNA ribozomální 16S MeSH
- skot mikrobiologie MeSH
- střeva mikrobiologie MeSH
- zvířata MeSH
- Check Tag
- skot mikrobiologie MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Freezing is considered to be the best method for long-term storage of bacterial DNA from feces; however this method cannot be usually applied for samples of wild primates collected in the challenging conditions of the tropical forest. In order to find an alternative conservation method of fecal samples from wild great apes, we compared freezing with other fixation methods. Fecal samples from 11 captive gorillas (Gorilla gorilla gorilla) from three Czech Zoos were stored using freezing, RNA Stabilization Reagent (RNAlater), and 96% ethanol. Subsequently, the samples were examined using culture-independent methods (PCR-DGGE, and Real-time PCR) to qualitatively and quantitatively assess fecal microbiota composition and to compare differences among the storage methods. Noticeably, freezing samples resulted in the highest recoveries of DNA. No significant differences in DNA recovery were found between freezing and using RNAlater; however, significantly lower DNA concentrations were recovered from samples stored in 96% ethanol. Using PCR-DGGE we found that either 96% ethanol, RNAlater or freezing were suitable for preserving bacterial DNA; however fingerprints obtained from RNAlater storage were more similar to those obtained from the frozen method; in comparison to the patterns resulting from storing samples in ethanol. Using qPCR, frozen samples yielded the highest values of bacterial counts, with the exception of Enterobacteriaceae, which showed the highest numbers using samples stored in ethanol. Sequences of amplicons obtained from PCR-DGGE belonged to the families Clostridiaceae, Lactobacillaceae, Staphylococcaceae, and Lachnospiraceae, phylum Firmicutes; however most amplicons showed sequence similarity to previously uncultured microorganisms. Bacteria belonging to the phylum Firmicutes were the most frequently identified species in the fecal bacterial communities of captive western gorillas. The study showed that RNAlater is an optimal storage method when freezing is not possible.
