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PubMed1991

Diversity and population dynamics of methanogenic bacteria in a granular consortium.

Upflow anaerobic sludge blanket bioreactor granules were used as an experimental model microbial consortium to study the dynamics and distribution of methanogens. Immunologic methods revealed a considerable diversity of methanogens that was greater in mesophilic granules than in the same granules 4 months after a temperature shift from 38 to 55 degrees C. During this period, the sizes of the methanogenic subpopulations changed with distinctive profiles after the initial reduction caused by the shift. Methanogens antigenically related to Methanobrevibacter smithii PS and ALI, Methanobacterium hungatei JF1, and Methanosarcina thermophila TM1 increased rapidly, reached a short plateau, and then fell to lower concentrations that persisted for the duration of the experiment. A methanogen related to Methanogenium cariaci JR1 followed a similar profile at the beginning, but it soon diminished below detection levels. Methanothrix rods weakly related to the strain Opfikon increased rapidly, reaching a high-level, long-lasting plateau. Two methanogens related to Methanobrevibacter arboriphilus AZ and Methanobacterium thermoautotrophicum DeltaH emerged from very low levels before the temperature shift and multiplied to attain their highest numbers 4 months after the shift. Histochemistry and immunohistochemistry revealed thick layers, globular clusters, and lawns of variable density which were distinctive of the methanogens related to M. thermoautotrophicum DeltaH, M. thermophila TM1, and M. arboriphilus AZ and M. soehngenii Opfikon, respectively, in thin sections of granules grown at 55 degrees C for 4 months. Mesophilic granules showed a different pattern of methanogenic subpopulations.

Antibodies, Archaeal↗

PubMed1994

Anaerobic degradation of propionate by a mesophilic acetogenic bacterium in coculture and triculture with different methanogens.

A mesophilic acetogenic bacterium (MPOB) oxidized propionate to acetate and CO(2) in cocultures with the formate- and hydrogen-utilizing methanogens Methanospirillum hungatei and Methanobacterium formicicum. Propionate oxidation did not occur in cocultures with two Methanobrevibacter strains, which grew only with hydrogen. Tricultures consisting of MPOB, one of the Methanobrevibacter strains, and organisms which are able to convert formate into H(2) plus CO(2) (Desulfovibrio strain G11 or the homoacetogenic bacterium EE121) also degraded propionate. The MPOB, in the absence of methanogens, was able to couple propionate conversion to fumarate reduction. This propionate conversion was inhibited by hydrogen and by formate. Formate and hydrogen blocked the energetically unfavorable succinate oxidation to fumarate involved in propionate catabolism. Low formate and hydrogen concentrations are required for the syntrophic degradation of propionate by MPOB. In triculture with Methanospirillum hungatei and the aceticlastic Methanothrix soehngenii, propionate was degraded faster than in biculture with Methanospirillum hungatei, indicating that low acetate concentrations are favorable for propionate oxidation as well.

Journal Article↗

PubMed1999

Phylogenetic diversity of symbiotic methanogens living in the hindgut of the lower termite Reticulitermes speratus analyzed by PCR and in situ hybridization.

A phylogenetic analysis of the sequences of 60 clones of archaeal small-subunit rRNA genes amplified from the termite Reticulitermes speratus revealed that most of them (56 clones) clustered in the genus Methanobrevibacter. Three clones were classified in the order Thermoplasmales. The Methanobrevibacter-related symbionts were detected by in situ hybridization analysis.

Animals↗

PubMed2026

Comparative analysis of rumen metagenomes with dietary supplementation of 3-nitrooxypropanol revealed divergent modes of action in hydrogen metabolism and reductant pathways between beef and dairy cattle.

BACKGROUND: The compound 3-nitrooxypropanol (3-NOP), an inhibitor of methyl-coenzyme M reductase (MCR), reduces enteric methane production in both beef and dairy cattle. Although the proposed mechanisms of 3-NOP involve on inhibiting the activity of MCR in vivo, it is unknown how this process could affect rumen microbiome as a whole and if it differs between beef and dairy cattle. This study conducted a comparative analysis of the rumen microbiome and its functional shifts in four different cattle studies (two beef and two dairy cattle studies) that evaluated 3-NOP supplementation using metataxonomics and metagenomics. RESULTS: Comparative analysis of 281 rumen metataxonomic datasets (143 beef and 138 dairy cattle) revealed that dietary supplementation with 3-NOP affected rumen bacteria and methanogens. Further, comparative analysis of 54 metagenomic datasets (24 beef and 30 dairy cattle) revealed that 3-NOP inhibited mcrA, decreased the abundances of Methanobrevibacter gottschalkii and the protozoal species Isotricha prostoma, while increased the abundances of Methanobrevibacter ruminantium and Methanosphaera sp., Prevotella sp. was a significant bacterial taxon in both beef and dairy cattle, contributing to various pathways such as propionate and butyrate production. Its increased abundance after 3-NOP supplementation may also be linked to the decrease in Isotricha prostoma. Hydrogenotrophic methanogenesis decreased after 3-NOP supplementation with the abundance of genes involved in methylenetetrahydromethanopterin dehydrogenase decreased in beef cattle, while that of 4Fe-4S ferredoxin gene decreased in dairy cattle. The abundance of protozoal Polyplastron multivesiculatum increased after long-term 3-NOP supplementation in beef cattle, potentially due to changes in hydrogen (H2) partial pressure. During 3-NOP-mediated methanogenesis reduction, abundance of genes encoding methanogenic hydrogenase and H2 producing hydrogenase were decreased, while those encoding H2 sensory hydrogenase increased. Acyl-CoA dehydrogenase gene involved in propionate and butyrate production pathways increased in both beef and dairy cattle, while nitrite reductase increased specifically in beef cattle, indicating a rise in alternative H2 sinks. Video Abstract CONCLUSION: Our findings revealed broad effects of 3-NOP on rumen microbiome and functions in vivo, with varied effects in beef and dairy cattle, which provide mechanistic insights into the supplementation of 3-NOP in both beef and dairy cattle, supporting its more sustainable and effective use in the future.

Metagenome↗

PubMed2001

Inhibition of growth of methane-producing bacteria of the ruminant forestomach by hydroxymethylglutaryl-SCoA reductase inhibitors.

Two hydroxymethylglutaryl-SCoA (HMG-CoA) reductase inhibitors, mevastatin and lovastatin, inhibited the in vitro growth and production of CH4 of strains of Methanobrevibacter isolated from the rumen. Mevastatin or lovastatin did not inhibit growth of species of rumen bacteria that are essential for fermenting cellulose, starch and other plant polysaccharides to acetate, propionate, and butyrate. Approximately 4 nmol of lovastatin per milliliter resulted in 50% growth inhibition of Methanobrevibacter strain ZA10 and concentrations > or =10 nmol per milliliter completely inhibited growth and CH4 formation. Results of in vitro growth studies suggest that supplementation of ruminant feeds with HMG-CoA inhibitors could decrease ruminant methane production and increase the efficiency of feed utilization by domestic ruminants.

Animals↗

PubMed1991

[Methanogens from feces of patients with intestine disorder].

Fecal specimens from 10 colorectal cancer, 1 duodenum diverticulosis and 2 healthy adults were examined. Nine fecal enrichments contained Methanobrevibacter. The percentage of methanogen positive individuals was 69% Methanobrevibacter were isolated from fecal enrichments of 4 colorectal cancer and 1 duodenum diverticulosis. The percentage of breath methane positive individuals and methane production had significantly increased in colorectal cancer patients. It will be hopeful that methane production in breath samples at the end of exhalation will became a rapid, simple and non-penetrative method for monitoring precancerous colonic or rectal carcinogenesis and monitoring the relapse of individuals with carcinosectomy.

Breath Tests↗

PubMed2006

A humanized gnotobiotic mouse model of host-archaeal-bacterial mutualism.

Our colons harbor trillions of microbes including a prominent archaeon, Methanobrevibacter smithii. To examine the contributions of Archaea to digestive health, we colonized germ-free mice with Bacteroides thetaiotaomicron, an adaptive bacterial forager of the polysaccharides that we consume, with or without M. smithii or the sulfate-reducing bacterium Desulfovibrio piger. Whole-genome transcriptional profiling of B. thetaiotaomicron, combined with mass spectrometry, revealed that, unlike D. piger, M. smithii directs B. thetaiotaomicron to focus on fermentation of dietary fructans to acetate, whereas B. thetaiotaomicron-derived formate is used by M. smithii for methanogenesis. B. thetaiotaomicron-M. smithii cocolonization produces a significant increase in host adiposity compared with monoassociated, or B. thetaiotaomicron-D. piger biassociated, animals. These findings demonstrate a link between this archaeon, prioritized bacterial utilization of polysaccharides commonly encountered in our modern diets, and host energy balance.

Animals↗

PubMed2004

Archaeosomes as adjuvants for combination vaccines.

The present study evaluated the potential of archaesomes, prepared from the total polar lipids extracted from Methanobrevibacter smithii, as adjuvants for combination (multivalent) vaccines. Groups of Balb/c mice were immunized subcutaneously at day 0 and 21 with one of the following vaccines: trivalent vaccine formulated by the simultaneous co-encapsulation of bovine serum albumine (BSA), ovalbumin (OVA) and hen egg lysozyme (HEL) into archaeosomes (CEC vaccine); an univalent archaeosome vaccine (UVE vaccine) containing either BSA, OVA or HEL; or an admixture vaccine (AMC vaccine) consisting of the three UVE vaccines. Serum specific antibody (IgG + M) responses were determined at day 32, 112 and 203, and specific IgG1 and IgG2a responses were determined at day 112. Mice immunized with the CEC of AMC vaccine developed strong and sustained specific antibody responses to all three antigens at a magnitude similar to those seen in control mice immunized with UVE vaccines. Moreover, the serum BSA-, OVA-, and HEL-specific IgG1 and IgG2a levels in the CEC and AMC immunized mice were overall comparable to those of the UVE immunized control mice. Boosting CEC and AMC vaccinated mice with antigens alone at day 203 elicited strong antibody memory responses, comparable to those in the UVE vaccinated groups. These results show that archaeosomes could be used as adjuvants in developing combination vaccines.

Adjuvants, Immunologic↗

PubMed2006

H2 and acetate transfers during xylan fermentation between a butyrate-producing xylanolytic species and hydrogenotrophic microorganisms from the human gut.

The aim of this work was to investigate in vitro interrelationships during xylan fermentation between an H2 and butyrate-producing xylanolytic species recently isolated in our laboratory from human faeces and identified as Roseburia intestinalis and the H2-utilizing acetogen Ruminococcus hydrogenotrophicus or the methanogen Methanobrevibacter smithii. H2 transfer between M. smithii or Ru. hydrogenotrophicus and the xylanolytic species was evidenced, confirming the great potential of these H2-consuming microorganisms to reutilize fermentative H2 during fibre fermentation in the gut. In addition, acetate transfer was demonstrated between the xylanolytic Roseburia sp. and the acetogenic species, both metabolites transfers leading to butyric fermentation of oat xylan without production of H2.

Acetates↗

PubMed2006

Identification and quantification of methanogenic Archaea in adult chicken ceca.

By using molecular methods for the identification and quantification of methanogenic archaea in adult chicken ceca, 16S rRNA genes of 11 different phylotypes, 10 of which were 99% similar to Methanobrevibacter woesei, were found. Methanogen populations, as assessed by cultivation, and the 16S rRNA copy number were between 6.38 and 8.23 cells/g (wet weight) and 5.50 and 7.19 log(10)/g (wet weight), respectively.

Animals↗

PubMed1988

Quantitative immunologic analysis of the methanogenic flora of digestors reveals a considerable diversity.

To determine which methanogens occur in digestors, we performed a quantitative immunologic analysis of a variety of samples. A comprehensive panel of calibrated polyclonal antibody probes of predefined specificity spectra was used. This allowed precise identification of bacteria by antigenic fingerprinting. A considerable diversity of methanogens was uncovered, much larger than previously reported, encompassing at least 14 strains of 11 species. Strategies were developed to measure the load of any given methanogen in a sample and to compare samples quantitatively. Two methanogens were found to predominate which were antigenically closely related with either Methanobacterium formicicum MF or Methanobrevibacter arboriphilus AZ. Fundamental data, probes, and methods are now available to monitor methanogenic subpopulations during digestor operation and thus learn about their respective roles and predictive significance.

Antibodies, Archaeal↗

PubMed2005

Structure and topology of microbial communities in the major gut compartments of Melolontha melolontha larvae (Coleoptera: Scarabaeidae).

Physicochemical gut conditions and the composition and topology of the intestinal microbiota in the major gut compartments of the root-feeding larva of the European cockchafer (Melolontha melolontha) were studied. Axial and radial profiles of pH, O2, H2, and redox potential were measured with microsensors. Terminal restriction fragment length polymorphism (T-RFLP) analysis of bacterial 16S rRNA genes in midgut samples of individual larvae revealed a simple but variable and probably nonspecific community structure. In contrast, the T-RFLP profiles of the hindgut samples were more diverse but highly similar, especially in the wall fraction, indicating the presence of a gut-specific community involved in digestion. While high acetate concentrations in the midgut and hindgut (34 and 15 mM) corroborated the presence of microbial fermentation in both compartments, methanogenesis was confined to the hindgut. Methanobrevibacter spp. were the only methanogens detected and were restricted to this compartment. Bacterial 16S rRNA gene clone libraries of the hindgut were dominated by clones related to the Clostridiales. Clones related to the Actinobacteria, Bacillales, Lactobacillales, and gamma-Proteobacteria were restricted to the lumen, whereas clones related to the beta- and delta-Proteobacteria were found only on the hindgut wall. Results of PCR-based analyses and fluorescence in situ hybridization of whole cells with group-specific oligonucleotide probes documented that Desulfovibrio-related bacteria comprise 10 to 15% of the bacterial community at the hindgut wall. The restriction of the sulfate-reducer-specific adenosine-5'-phosphosulfate reductase gene apsA to DNA extracts of the hindgut wall in larvae from four other populations in Europe suggested that sulfate reducers generally colonize this habitat.

Animals↗

PubMed2026

Multi-Omic Insights Into Mediterranean Diet-Associated Microbiota.

This study aimed to evaluate the gut microbiota and mycobiota composition, depending on the Mediterranean diet (MD) adherence, using metataxonomics. Combining metagenomics and metatranscriptomics, we also investigate the gene expression level in the bacterial community. Two groups of healthy subjects greatly differing in adherence were selected. Significant differences in microbiota composition were observed between individuals with high adherence (HAMD; mean 10.5 +/- 0.9 points) and low adherence (LAMD; 5.23 +/- 83 points). Notably, the olive oil, vegetable, and fruit consumption presented an important discriminant power between groups. Saccharomyces, Penicillium, and Candida were the most abundant genera. Mycobiota richness was higher in LAMD than in HAMD. Aspergillus was identified as a biomarker for LAMD, whereas Yarrowia, a potential probiotic, was a biomarker for HAMD. Metatranscriptomics indicated that Bacillota was the most metabolically active phylum in the gut microbiota. The low-abundant genus, Methanobrevibacter, showed high transcriptional activity, contributing to the crucial methanogenesis process. Gene expression analyses further highlighted functional differences. Overall, HAMD microbiota presented increased metabolic activity, protein synthesis, and cellular mobility. Overexpression of flagellin and urease genes may enhance immune response in HAMD. Further metatranscriptomic studies are necessary to deepen our understanding of intestinal microbiota transcriptional programs and their interactions with the diet and human health.

Humans↗

PubMed1997

H2/CO2 metabolism in acetogenic bacteria isolated from the human colon.

The present work reports on autotrophic metabolism in four H2/CO2-utilizing acetogenic bacteria isolated from the human colon (two Clostridium species, one Streptococcus species, and Ruminococcus hydrogenotrophicus). H2/CO2-utilization by these human acetogenic strains occurred during both exponential and stationary phases of growth. Acetate was the major metabolite produced by all isolates following the stoichiometric equation of reductive acetogenesis. Furthermore, the ability of these acetogenic bacteria to incorporate 13CO2 into acetate in the presence of H2 in the gas phase demonstrated the utilization of the reductive pathway of acetate formation from a one-carbon compound. Energy conservation during the autotrophic metabolism in colonic acetogens might involve sodium- or proton-chemiosmotic mechanisms. A sodium-dependent ATP generation was only demonstrated in one Clostridium species, whereas sodium could be replaced by potassium in other strains. The minimal thresholds of hydrogen uptake were determined and varied from 1100 to 3680 ppm depending on the acetogenic strain. These values appeared higher than those measured for the colonic methanogen,Methanobrevibacter smithii.

Journal Article↗

PubMed1992

Effect of coculture of anaerobic fungi isolated from ruminants and non-ruminants with methanogenic bacteria on cellulolytic and xylanolytic enzyme activities.

Neocallimastix strain N1, an isolate from a ruminant (sheep), was cocultured with three Methanobacterium formicicum strains, Methanosarcina barkeri, and Methanobrevibacter smithii. The coculture with Methanobacterium formicicum strains resulted in the highest production of cellulolytic and xylanolytic enzymes. Subsequently four anaerobic fungi, two Neocallimastix strains (N1 and N2) from a ruminant and two Piromyces species from non-ruminants (E2 and R1), were grown in coculture with Methanobacterium formicicum DSM 3637 on filter paper cellulose and monitored over a 7-day period for substrate utilisation, fermentation products, and secretion of cellulolytic and xylanolytic enzymes. Methanogens caused a shift in fermentation products to more acetate and less ethanol, lactate and succinate. Furthermore the cellulose digestion rate increased by coculture. For cocultures of Neocallimastix strains with Methanobacterium formicicum strains the cellulolytic and xylanolytic enzyme production increased. Avicelase, CMCase and xylanase were almost completely secreted into the medium, while 40-60% of the beta-glucosidase was found to be cell bound. Coculture had no significant effect on the location of cellulolytic and xylanolytic enzymes.

Anaerobiosis↗

PubMed1995

Role of formate and hydrogen in the degradation of propionate and butyrate by defined suspended cocultures of acetogenic and methanogenic bacteria.

The butyrate-degrading Syntrophospora bryantii degrades butyrate and a propionate-degrading strain (MPOB) degrades propionate in coculture with the hydrogen- and formate-utilizing Methanospirillum hungatii or Methanobacterium formicicum. However, the substrates are not degraded in constructed cocultures with two Methanobrevibacter arboriphilus strains which are only able to consume hydrogen. Pure cultures of the acetogenic bacteria form both hydrogen and formate during butyrate oxidation with pentenoate as electron acceptor and during propionate oxidation with fumarate as electron acceptor. Using the highest hydrogen and formate levels which can be reached by the acetogens and the lowest hydrogen and formate levels which can be maintained by the methanogens it appeared that the calculated formate diffusion rates are about 100 times higher than the calculated hydrogen diffusion rates.

Butyrates↗

PubMed1985

Sequence divergence of an archaebacterial gene cloned from a mesophilic and a thermophilic methanogen.

A 1.6-kb fragment of DNA from the thermophilic, methane-producing, anaerobic archaebacterium Methanobacterium thermoautotrophicum delta H has been cloned and sequenced. This DNA complements mutations in both the purE1 and purE2 loci of Escherichia coli. The sequence of the M. thermoautotrophicum DNA predicts that complementation in E. coli results from the synthesis of a polypeptide with a molecular weight of 36,249. A polypeptide apparently of this molecular weight is synthesized in E. coli minicells containing recombinant plasmids that carry the cloned fragment of methanogen DNA. We have previously cloned and sequenced a purE-complementing gene from the mesophilic methanogen Methanobrevibacter smithii. The two methanogen-derived purE-complementing genes are 53% homologous and encode polypeptides that are 45% homologous in their amino acid sequences but would be 74% homologous if conservative amino acid substitutions were considered as maintaining sequence homology. The genome of M. thermoautotrophicum has a molar G + C content of 49.7%, whereas the genome of M. smithii is 30.6% G + C. Conservation of encoded amino acids while accommodating the very different G + C contents is accomplished by use of different codons that encode the same amino acid. The majority of base changes occur at the third codon position. The intergenic regions of the cloned M. thermoautotrophicum DNA contain sequences previously identified as ribosome binding sites and as putative methanogen promoters. Although the two purE-complementing genes are apparently derived from a common ancestor, only the gene from M. smithii maintains a codon usage that conforms to the RNY rule.

Amino Acid Sequence↗

PubMed2002

Degradation of fresh ryegrass by methanogenic co-cultures of ruminal fungi grown in the presence or absence of Fibrobacter succinogenes.

The ability of five ruminal fungi in syntrophic co-culture with the methanogen Methanobrevibacter smithii to degrade perennial ryegrass ( Lolium perenne) stem fragments and leaf blades was studied to determine the susceptibilities of non-autoclaved fresh tissues to fungal degradation. Autoclaving did not significantly increase fungal degradation of stem fragments but strongly increased degradation of leaf blades by a species of Caecomyces. In methanogenic co-cultures, non-autoclaved stem fragments were degraded more extensively by Neocallimastix frontalis and Piromyces isolates than by Caecomyces isolates. The N. frontalis and Piromyces isolates showed the greatest rates of stem degradation. When interactions between Fibrobacter succinogenes and methanogenic co-cultures of fungi growing on ryegrass stem were investigated, N. frontalis inhibited F. succinogenes. This has not been observed previously. In contrast, a Caecomyces species interacted positively with F. succinogenes to increase stem degradation, suggesting that F. succinogenes and Caecomyces spp. may have complementary fibrolytic activities. All five fungi tested failed to grow on fresh non-autoclaved leaf blades. In a repeat experiment with leaves from a separate harvest, leaf blades were degraded by N. frontalis but not by a Caecomyces species. We suggest that ryegrass leaf blades may contain natural anti-fungal compounds. Our results confirm the superiority of fungi in the degradation of intact stem and indicate that in vitro studies with non-autoclaved forage tissues may yield new information on forage factors affecting rumen microbes.

Anaerobiosis↗