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the first true obligately syntrophic propionate-oxidizing bacterium, pelotomaculum schinkii sp. nov., co-cultured with methanospirillum hungatei, and emended description of the genus pelotomaculum.a gram-positive, spore-forming, syntrophic propionate-oxidizing bacterium, pelotomaculum schinkii sp. nov. strain hh(t), was isolated as a co-culture with methanospirillum hungatei jf-1(t) from anaerobic, freeze-dried granular sludge obtained from an upflow anaerobic sludge bed reactor treating sugar beet wastewater. the bacterium converted propionate to acetate in co-culture with methanospirillum hungatei jf-1(t) or methanobacterium formicicum mf(nt), but not in co-culture with methanobrevibact ...200516014504
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.200717085694
phylogenetic analysis of bacterial and archaeal species in symptomatic and asymptomatic endodontic infections.phylogenetic analysis of bacterial and archaeal 16s rrna was used to examine polymicrobial communities within infected root canals of 20 symptomatic and 14 asymptomatic patients. nucleotide sequences from approximately 750 clones amplified from each patient group with universal bacterial primers were matched to the ribosomal database project ii database. phylotypes from 37 genera representing actinobacteria, bacteroidetes, firmicutes, fusobacteria and proteobacteria were identified. results were ...200717172525
rapid clonal expansion and prolonged maintenance of memory cd8+ t cells of the effector (cd44highcd62llow) and central (cd44highcd62lhigh) phenotype by an archaeosome adjuvant independent of tlr2.vaccines capable of eliciting long-term t cell immunity are required for combating many diseases. live vectors can be unsafe whereas subunit vaccines often lack potency. we previously reported induction of cd8(+) t cells to ag entrapped in archaeal glycerolipid vesicles (archaeosomes). in this study, we evaluated the priming, phenotype, and functionality of the cd8(+) t cells induced after immunization of mice with ova-methanobrevibacter smithii archaeosomes (ms-ova). a single injection of ms-ov ...200717277146
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 c ...200516085849
characterization of a 4-methylbenzoate-degrading methanogenic consortium as determined by small-subunit rdna sequence analysis.a methanogenic consortium that degrades 4-methylbenzoate (mba) as the sole carbon and energy source was successfully enriched in an upflow anaerobic sludge bed bioreactor and studied. electron microscopic observation showed that long rods with a distinct collar feature resembling desulfomonile tiedjei rods were the predominant population, and that these rods formed a close spatial orientation with methanobrevibacter-like bacteria. in addition, thin filaments and bamboo-shaped filaments that high ...200116233021
anaerobic degradation of lactate by syntrophic associations of methanosarcina barkeri and desulfovibrio species and effect of h(2) on acetate degradation.when grown in the absence of added sulfate, cocultures of desulfovibrio desulfuricans or desulfovibrio vulgaris with methanobrevibacter smithii (methanobacterium ruminantium), which uses h(2) and co(2) for methanogenesis, degraded lactate, with the production of acetate and ch(4). when d. desulfuricans or d. vulgaris was grown in the absence of added sulfate in coculture with methanosarcina barkeri (type strain), which uses both h(2)-co(2) and acetate for methanogenesis, lactate was stoichiometr ...198116345708
fermentation of cellulose to methane and carbon dioxide by a rumen anaerobic fungus in a triculture with methanobrevibacter sp. strain ra1 and methanosarcina barkeri.the fermentation of cellulose by a rumen anaerobic fungus in the presence of methanobrevibacter sp. strain ra1 and methanosarcina barkeri strain 227 resulted in the formation of 2 mol each of methane and carbon dioxide per mol of hexose fermented. coculture of the fungus with either methanobrevibacter sp. or m. barkeri produced 0.6 and 1.3 mol of methane per mol of hexose, respectively. acetate, formate, ethanol, hydrogen, and lactate, which are major end products of cellulose fermentation by th ...198216346048
gelrite as an agar substitute for the cultivation of mesophilic methanobacterium and methanobrevibacter species.gelrite was evaluated as a gelling agent for the growth of mesophilic methanobacterium and methanobrevibacter species. gelrite was shown to be superior to agar in its gel strength and clarity. viable cell counts and colony sizes of methanobacterium species were greater on gelrite-based medium compared with agar-based medium.198516346906
direct characterization of methanogens in two high-rate anaerobic biological reactors.the methanogenic flora from two types of turbulent, high-rate reactors was studied by immunologic methods as well as by phase-contrast, fluorescence, and scanning electron microscopy. the reactors were a fluidized sand-bed biofilm anitron reactor and an ultrafiltration membrane-associated suspended growth mars reactor (both trademarks of air products and chemicals, inc., allentown, pa.). conventional microscopic methods revealed complex mixtures of microbes of a range of sizes and shapes, among ...198816347581
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 s ...199116348508
influence of metronidazole, co, co(2), and methanogens on the fermentative metabolism of the anaerobic fungus neocallimastix sp. strain l2.the effects of metronidazole, co, methanogens, and co(2) on the fermentation of glucose by the anaerobic fungus neocallimastix sp. strain l2 were investigated. both metronidazole and co caused a shift in the fermentation products from predominantly h(2), acetate, and formate to lactate as the major product and caused a lower glucose consumption rate and cell protein yield. an increased lactate dehydrogenase activity and a decreased hydrogenase activity were observed in cells grown under both cul ...199316349022
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 ho ...199416349350
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-con ...200616451188
say hello to our little friends. 200717633442
methanobrevibacter ruminantium as an indicator of domesticated-ruminant fecal pollution in surface waters.a pcr-based assay (mrnif) targeting the nifh gene of methanobrevibacter ruminantium was developed to detect fecal pollution from domesticated ruminants in environmental water samples. the assay produced the expected amplification product only when the reaction mixture contained dna extracted from m. ruminantium culture, bovine (80%), sheep (100%), and goat (75%) feces, and water samples from a bovine waste lagoon (100%) and a creek contaminated with bovine lagoon waste (100%). the assay appears ...200717827308
16s rdna directed pcr primers and detection of methanogens in the bovine rumen.to assess the diversity of ruminal methanogens in a grazing cow, and develop pcr primers targeting the predominant methanogens.200616478508
salt tolerance of archaeal extremely halophilic lipid membranes.the membranes of extremely halophilic archaea are characterized by the abundance of a diacidic phospholipid, archaetidylglycerol methylphosphate (pgp-me), which accounts for 50-80 mol% of the polar lipids, and by the absence of phospholipids with choline, ethanolamine, inositol, and serine head groups. these membranes are stable in concentrated 3-5 m nacl solutions, whereas membranes of non-halophilic archaea, which do not contain pgp-me, are unstable and leaky under such conditions. by x-ray di ...200616484230
evaluation of methanogenic strains and their ability to endure aeration and water stress.during periods of drainage, both water stress and oxygen can cause damage to indigenous methanogens. in the present study, we evaluated the tolerance of seven methanogenic strains (methanobrevibacter arboriphilicus, methanobacterium formicicum, methanococcus vannielii, methanospirillum hungatei, methanoculleus olentangyi, methanoplanus limicola, and methanosarcina mazei) to long-term exposure to air/nitrogen and drying. we found that these methanogenic strains except for m. limicola and m. olent ...200817990030
anaerobic transformation of furfural by methanococcus deltae (delta)lh.methanococcus deltae (delta)lh was grown on h(inf2)-co(inf2) in the presence of various concentrations of furfural. furfural at higher concentrations, namely, 20 and 25 mm, inhibited growth of this organism. at concentration of 5 and 10 mm, no inhibition of growth was observed. the other methanogens in this study were not inhibited by 10 mm furfural. among the methanogens tested, m. deltae was capable of transforming furfural, whereas methanobacterium thermoautotrophicum marburg, methanosarcina ...199716535618
identification and quantification of archaea involved in primary endodontic infections.members of the domain archaea, one of the three domains of life, are a highly diverse group of prokaryotes, distinct from bacteria and eukaryotes. despite their abundance and ubiquity on earth, including their close association with humans, animals, and plants, so far no pathogenic archaea have been described. as some archaea live in close proximity to anaerobic bacteria, for instance, in the human gut system and in periodontal pockets, the aim of our study was to assess whether archaea might po ...200616597851
[diversity and function of symbiotic microbes in the gut of lower termites].the gut of lower termites harbor a complex microbial community, including eukaryotic flagellates and prokaryotic bacteria and archaea, which play important roles in the wood-cellulose digestion of these termites. the hindguts of lower termites are characterized with an enlarged paunch with steep oxygen and hydrogen gradient and not randomly distributed abundant microorganisms. the symbiotic flagellates in the gut of lower termites, which are normally associated with epibionts or endobionts, are ...200616933630
dynamics of a pig slurry microbial community during anaerobic storage and management.the microbial community of a pig slurry on a farm was monitored for 6 months using both molecular and cultural approaches. sampling was carried out at all the different stages of effluent handling, from the rearing build-up to slurry spreading. total dna of each sample was extracted and analyzed by pcr-single-strand conformation polymorphism (sscp) analysis using primers targeting the 16s rrna genes from the archaeal and bacterial domains and also the eubacterium-clostridium, bacillus-streptococ ...200616672505
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. pig ...200616782812
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 acetoge ...199716887606
effect of methanobrevibacter sp mf1 inoculation on glycoside hydrolase and polysaccharide depolymerase activities, wheat straw degradation and volatile fatty acid concentrations in the rumen of gnotobiotically-reared lambs.four naturally born lambs were placed in sterile isolators 24 h after birth before the natural establishment of cellulolytic microorganisms and archaea methanogens. at the age of 6 weeks they were inoculated with pure cultures of the strains fd1 and 007 of ruminococcus flavefaciens and at the age of 4 months with a pure culture of methanobrevibacter sp. mf1. following the establishment of mf1, the population of r. flavefaciens slightly increased in the rumen of the four lambs, there was also an ...199716887613
application of pseudomurein endoisopeptidase to fluorescence in situ hybridization of methanogens within the family methanobacteriaceae.in situ detection of methanogens within the family methanobacteriaceae is sometimes known to be unsuccessful due to the difficulty in permeability of oligonucleotide probes. pseudomurein endoisopeptidase (pei), a lytic enzyme that specifically acts on their cell walls, was applied prior to 16s rrna-targeting fluorescence in situ hybridization (fish). for this purpose, pure cultured methanogens within this family, methanobacterium bryantii, methanobrevibacter ruminantium, methanosphaera stadtmana ...200616950902
in vitro bacterial growth and in vivo ruminal microbiota populations associated with bloat in steers grazing wheat forage.the role of ruminal bacteria in the frothy bloat complex common to cattle grazing winter wheat has not been previously determined. two experiments, one in vitro and another in vivo, were designed to elucidate the effects of fresh wheat forage on bacterial growth, biofilm complexes, rumen fermentation end products, rumen bacterial diversity, and bloat potential. in exp. 1, 6 strains of ruminal bacteria (streptococcus bovis strain 26, prevotella ruminicola strain 23, eubacterium ruminantium b1c23, ...200616971591
molecular phylogenetic diversity of the microbial community associated with a high-temperature petroleum reservoir at an offshore oilfield.the microbial community and its diversity in production water from a high-temperature, water-flooded petroleum reservoir of an offshore oilfield in china were characterized by 16s rrna gene sequence analysis. the bacterial and archaeal 16s rrna gene clone libraries were constructed from the community dna and, using sequence analysis, 388 bacterial and 220 archaeal randomly selected clones were clustered with 60 and 28 phylotypes, respectively. the results showed that the 16s rrna genes of bacter ...200717286581
shifts in methanogenic subpopulations measured with antibody probes in a fixed-bed loop anaerobic bioreactor treating sulfite evaporator condensate.a fixed-bed loop, high-rate anaerobic bioreactor treating sulfite evaporator condensate was sampled when it reached steady state and afterwards following perturbations during a 14-month period. by using immunotechnology, it was observed that shifts in methanogenic subpopulations occurred in association with perturbations, such as restarting and relocating the biomass into a different tank. methanogens related to methanobacterium bryantii mohg and methanobrevibacter smithii ali were numerous thro ...198916347990
effect of methanobrevibacter smithii on xylanolytic activity of anaerobic ruminal fungi.three different ruminal anaerobic fungi, neocallimastix frontalis pnk2, sphaeromonas communis b7, and piromonas communis b19, were grown axenically or in coculture with methanobrevibacter smithii on xylan. n. frontalis and s. communis in monoculture and coculture accumulated xylobiose, xylose, and arabinose in the growth medium; arabinose was not metabolized, but xylobiose and xylose were subsequently used. the transient accumulation of xylose was much less evident in cocultures. both the rate a ...199016348244
quantitative microbiological analysis of bacterial community shifts in a high-rate anaerobic bioreactor treating sulfite evaporator condensate.the bacterial population of a high-rate, anaerobic, fixed-bed loop reactor treating sulfite evaporator condensate from the pulp industry was studied over a 14-month period. this period was divided into seven cycles that included a startup at the beginning of each cycle. some 82% of the total biomass was immobilized on and between the porous glass rings filling the reactor. the range of the total number of microorganisms in these biofilms was 2 x 10 to 7 x 10 cells per ml. enumeration and charact ...199016348253
syntrophic growth on formate: a new microbial niche in anoxic environments.anaerobic syntrophic associations of fermentative bacteria and methanogenic archaea operate at the thermodynamic limits of life. the interspecies transfer of electrons from formate or hydrogen as a substrate for the methanogens is key. contrary requirements of syntrophs and methanogens for growth-sustaining product and substrate concentrations keep the formate and hydrogen concentrations low and within a narrow range. since formate is a direct substrate for methanogens, a niche for microorganism ...200818708519
molecular diversity of methanogens in feedlot cattle from ontario and prince edward island, canada.the molecular diversity of rumen methanogens in feedlot cattle and the composition of the methanogen populations in these animals from two geographic locations were investigated using 16s rrna gene libraries prepared from pooled pcr products from 10 animals in ontario (127 clones) and 10 animals from prince edward island (114 clones). a total of 241 clones were examined, with methanobrevibacter ruminantium accounting for more than one-third (85 clones) of the clones identified. from these 241 cl ...200717483285
postinoculation protozoan establishment and association patterns of methanogenic archaea in the ovine rumen.association patterns between archaea and rumen protozoa were evaluated by analyzing archaeal 16s rrna gene clone libraries from ovine rumen inoculated with different protozoa. five protozoan inoculation treatments, fauna free (negative control), holotrich and cellulolytic protozoa, isotricha and dasytricha spp., entodinium spp., and total fauna (type a) were tested. we used denaturing gradient gel electrophoresis, quantitative pcr, and phylogenetic analysis to evaluate the impact of the protozoa ...200717513586
genomic and metabolic adaptations of methanobrevibacter smithii to the human gut.the human gut is home to trillions of microbes, thousands of bacterial phylotypes, as well as hydrogen-consuming methanogenic archaea. studies in gnotobiotic mice indicate that methanobrevibacter smithii, the dominant archaeon in the human gut ecosystem, affects the specificity and efficiency of bacterial digestion of dietary polysaccharides, thereby influencing host calorie harvest and adiposity. metagenomic studies of the gut microbial communities of genetically obese mice and their lean litte ...200717563350
quantitation and diversity analysis of ruminal methanogenic populations in response to the antimethanogenic compound bromochloromethane.methyl coenzyme-m reductase a (mcra) clone libraries were generated from microbial dna extracted from the rumen of cattle fed a roughage diet with and without supplementation of the antimethanogenic compound bromochloromethane. bromochloromethane reduced total methane emissions by c. 30%, with a resultant increase in propionate and branched chain fatty acids. the mcra clone libraries revealed that methanobrevibacter spp. were the dominant species identified. a decrease in the incidence of methan ...200717949432
simultaneous methanogenesis and oxygen reduction by methanobrevibacter cuticularis at low oxygen fluxes.methanogenic archaea are often encountered in habitats that are not entirely anoxic in space or time. recent biochemical and genomic studies have revealed the capacity of methanogens to reduce molecular oxygen. o(2) reduction by methanobrevibacter species was investigated. cell suspensions incubated in agar tubes under a headspace of h(2)-co(2) and increasing concentrations of o(2) formed a distinct growth band, which coincided with the oxic-anoxic interface and indicated that the influx of o(2) ...200717961173
evidence of increased diversity of methanogenic archaea with plant extract supplementation.this study evaluated the effects of selected essential oils on archaeal communities using the ovine rumen model. forty weaned canadian arcott ewes, fed with barley-based diet, were allotted to one of three essential oil supplementation treatments or a control (10 ewes per treatment) for 13 weeks. the treatments were cinnamaldehyde, garlic oil, juniper berry oil, and a control with no additive. rumen content was sampled after slaughter and grouped by treatment by combining subsamples from each an ...200818075710
effect of tea saponin on methanogenesis, microbial community structure and expression of mcra gene, in cultures of rumen micro-organisms.to determine the in-vitro effect and mode of action of tea saponin on the rumen microbial community and methane production.200819146532
evaluations of different hypervariable regions of archaeal 16s rrna genes in profiling of methanogens by archaea-specific pcr and denaturing gradient gel electrophoresis.different hypervariable (v) regions of the archaeal 16s rrna gene (rrs) were compared systematically to establish a preferred v region(s) for use in archaea-specific pcr-denaturing gradient gel electrophoresis (dgge). the pcr products of the v3 region produced the most informative dgge profiles and permitted identification of common methanogens from rumen samples from sheep. this study also showed that different methanogens might be detected when different v regions are targeted by pcr-dgge. die ...200818083874
methanobrevibacter phylotypes are the dominant methanogens in sheep from venezuela.rumen methanogens in sheep from venezuela were examined using 16s rrna gene libraries and denaturing gradient gel electrophoresis (dgge) profiles prepared from pooled and individual pcr products from the rumen contents from 10 animals. a total of 104 clones were examined, revealing 14 different 16s rrna gene sequences or phylotypes. of the 14 phylotypes, 13 (99 of 104 clones) belonged to the genus methanobrevibacter, indicating that the genus methanobrevibacter is the most dominant component of ...200818165875
[molecular diversity of rumen archaea from jinnan cattle].molecular diversities of rumen archaea of jinnan (south shanxi province, china) cattle was analyzed and compared by 16s rrna gene sequencing from three clone libraries generated using three different archaea-specific primer sets, respectively. dna from rumen of 4 jinnan cattle was extracted, and methanogen 16s rrna gene was amplified using archaea-specific primer sets. three clone libraries were generated by using vector pgem-t and cloning into e.coli jm109. one hundred clones were randomly pick ...200818338569
molecular diversity analysis of rumen methanogenic archaea from goat in eastern china by dgge methods using different primer pairs.to screen a pair of primers suitable for denaturing gradient gel electrophoretic (dgge) analysis of ruminal methanogenic archaea and to detect the archaeal communities in the rumen of goat.200919416460
molecular analysis of persistent periradicular lesions and root ends reveals a diverse microbial profile.the microbial etiology of periradicular lesions is recognized; however, the bacterial profile of these lesions is not well elucidated. the purpose of this study was to determine the frequency of bacterial colonization in these lesions and to characterize the bacterial community present in root ends and periradicular lesions.200919567313
prevalence and molecular diversity of archaea in subgingival pockets of periodontitis patients.the aim of this study was to investigate the prevalence and molecular diversity of archaea in the subgingival crevices of patients with chronic periodontitis.200919572899
archaeosome adjuvants: immunological capabilities and mechanism(s) of action.archaeosomes (liposomes comprised of glycerolipids of archaea) constitute potent adjuvants for the induction of th1, th2 and cd8(+) t cell responses to the entrapped soluble antigen. archaeal lipids are uniquely constituted of ether-linked isoprenoid phytanyl cores conferring stability to the membranes. additionally, varied head groups displayed on the glycerol-lipid cores facilitate unique immunostimulating interactions with mammalian antigen-presenting cells (apcs). the polar lipid from the ar ...200818343538
volatilisation of metals and metalloids: an inherent feature of methanoarchaea?as shown by recent studies, anaerobic members of archaea and bacteria are involved in processes that transform ionic species of metals and metalloids (arsenic, antimony, bismuth, selenium, tellurium and mercury) into volatile and mostly toxic derivatives (mainly methyl derivatives or hydrides). since the fact that these transformations proceed in both environmental settings and in parts of the human body, we have to consider that these processes also interfere directly with human health. the div ...200818396004
t-rflp-based mcra gene analysis of methanogenic archaea in association with oral infections and evidence of a novel methanobrevibacter phylotype.increasing evidence suggests a role for methanogenic archaea (methanogens) in human health and disease via syntrophic interactions with bacteria. here we assessed the prevalence and distribution of methanogens and possible associations with bacteria in oral biofilms.200919702957
human methanogen diversity and incidence in healthy and diseased colonic groups using mcra gene analysis.the incidence and diversity of human methanogens are insufficiently characterised in the gastrointestinal tract of both health and disease. a pcr and clone library methodology targeting the mcra gene was adopted to facilitate the two-fold aim of surveying the relative incidence of methanogens in health and disease groups and also to provide an overview of methanogen diversity in the human gastrointestinal tract.200818492229
structural characterization of archaeal lipid mucosal vaccine adjuvant and delivery (amvad) formulations prepared by different protocols and their efficacy upon intranasal immunization of mice.intranasal administration of ovalbumin (ova) formulated in an archaeal lipid mucosal vaccine adjuvant and delivery (amvad) system prepared by the addition of cacl2 to small unilamellar archaeosomes (liposomes made from archaeal polar lipids) containing encapsulated ova, was recently shown to elicit strong and sustained ova-specific mucosal and systemic immune responses. in this study, we show that the centrifugation/washing and antigen quantization steps required in the standard protocol for obt ...200818569448
granular sludge formation in upflow anaerobic sludge blanket (uasb) reactors.the state of the art for upflow anaerobic sludge blanket (uasb) reactors is discussed, focusing on the microbiology of immobilized anaerobic bacteria and the mechanism of granule formation. the development of granular sludge is the key factor for successful operation of the uasb reactors. criteria for determining if granular sludge has developed in a uasb reactor is given based on the densities and diameters of the granular sludge. the shape and composition of granular sludge can vary significan ...199618623574
[detection and diversity analysis of rumen methanogens in the co-cultures with anaerobic fungi].rumen methanogen diversity in the co-cultures with anaerobic fungi from goat rumen was analyzed. mix-cultures of anaerobic fungi and methanogens were obtained from goat rumen using anaerobic fungal medium and the addition of penicillin and streptomycin and then subcultured 62 times by transferring cultures every 3 - 4d. total dna from the original rumen fluid and subcultured fungal cultures was used for pcr/dgge and rflp analysis. 16s rdna of clones corresponding to representative otus were sequ ...200617302147
methanobrevibacter millerae sp. nov. and methanobrevibacter olleyae sp. nov., methanogens from the ovine and bovine rumen that can utilize formate for growth.four formate-utilizing methanogens were isolated from ovine (strain km1h5-1p(t)) and bovine (strains ak-87, ocp and za-10(t)) rumen contents. based on 16s rrna gene sequence analysis, the methanogen strains were found to belong to the order methanobacteriales in the genus methanobrevibacter. strains za-10(t) and km1h5-1p(t) gained energy for growth by the reduction of co(2) to ch(4) using h(2) or formate exclusively as electron donors. increasing formate concentrations to 220 mm in batch culture ...200717329767
the genome sequence of the rumen methanogen methanobrevibacter ruminantium reveals new possibilities for controlling ruminant methane emissions.methane (ch(4)) is a potent greenhouse gas (ghg), having a global warming potential 21 times that of carbon dioxide (co(2)). methane emissions from agriculture represent around 40% of the emissions produced by human-related activities, the single largest source being enteric fermentation, mainly in ruminant livestock. technologies to reduce these emissions are lacking. ruminant methane is formed by the action of methanogenic archaea typified by methanobrevibacter ruminantium, which is present in ...201020126622
relationships between anaerobic consortia and removal efficiencies in an uasb reactor degrading 2,4 dichlorophenol (dcp).to gain more insight into the interactions between anaerobic bacteria and reactor performances (chemical oxygen demand-cod, 2,4 dichlorophenol-2,4 dcp removals, volatile fatty acid-vfa, and methane gas productions) and how they depended on operational conditions the microbial variations in the anaerobic granular sludge from an upflow anaerobic sludge blanket (uasb) reactor treating 2,4 dcp was studied. the study was composed of two parts. in the first part, the numbers of methanogens and acedoge ...200817379394
identification of pseudomurein cell wall binding domains.methanothermobacter thermautotrophicus is a methanogenic gram-positive microorganism with a cell wall consisting of pseudomurein. currently, no information is available on extracellular pseudomurein biology and so far only two prophage pseudomurein autolysins, peiw and peip, have been reported. in this paper we show that peiw and peip contain two different n-terminal pseudomurein cell wall binding domains. this finding was used to identify a novel domain, pb007923, on the m. thermautotrophicus g ...200617427286
analysis of methanogen diversity in the rumen using temporal temperature gradient gel electrophoresis: identification of uncultured methanogens.a temporal temperature gradient gel electrophoresis (ttge) method was developed to determine the diversity of methanogen populations in the rumen. tests with amplicons from genomic dna from 12 cultured methanogens showed single bands for all strains, with only two showing apparently comigrating bands. fingerprints of methanogen populations were analyzed from dna extracted from rumen contents from two cattle and four sheep grazing pasture. for one sheep, dilution cultures selective for methanogen ...200717431710
[phylogenetic analysis of symbiotic archaea in the gut of reticulitermes chinensis snyder].to analyze the phylotypes of symbiotic archaea in the gut of reticulitermes chinensis snyder by using non-cultivating method.200920222453
phylogenetic analysis and fluorescence in situ hybridization detection of archaeal and bacterial endosymbionts in the anaerobic ciliate trimyema compressum.the anaerobic free-living ciliate, trimyema compressum, is known to harbor both methanogenic archaeal and bacterial symbionts in the cytoplasm. to clarify their phylogenetic belongings, a full-cycle rrna approach was applied to this symbiosis. phylogenetic analysis showed that the methanogenic symbiont was related to methanobrevibacter arboriphilicus, which was distantly related to symbionts found in other trimyema species. this result suggested that trimyema species do not require very specific ...200717468963
diversity of the resident microbiota in a thermophilic municipal biogas plant.biogas plants continuously convert biological wastes mainly into a mixture of methane, co2 and h2o-a conversion that is carried out by a consortium of bacteria and archaea. especially in the municipal plants dedicated towards waste treatment, the reactor feed may vary considerably, exposing the resident microbiota to a changing variety of substrates. to evaluate how and if such changes influence the microbiology, an established biogas plant (6,600 m3, up to 600 m3 biogas per h) was followed over ...200818820906
safety of intranasally administered archaeal lipid mucosal vaccine adjuvant and delivery (amvad) vaccine in mice.the safety profile of a recently described novel archaeal lipid mucosal vaccine adjuvant and delivery (amvad) system capable of eliciting robust antigen-specific mucosal and systemic immune responses was evaluated in female balb/c mice (10/group) using ovalbumin (ova) antigen. mice were intranasally immunized (0, 7, and 21 days) with a vaccine comprising 1 microg ova (0.05 mg/kg body weight) formulated in 0.04 mg total polar lipids extract (2.17 mg/kg body weight) of methanobrevibacter smithii c ...200818821397
effects of the gut microbiota on host adiposity are modulated by the short-chain fatty-acid binding g protein-coupled receptor, gpr41.the distal human intestine harbors trillions of microbes that allow us to extract calories from otherwise indigestible dietary polysaccharides. the products of polysaccharide fermentation include short-chain fatty acids that are ligands for gpr41, a g protein-coupled receptor expressed by a subset of enteroendocrine cells in the gut epithelium. to examine the contribution of gpr41 to energy balance, we compared gpr41-/- and gpr41+/+ mice that were either conventionally-raised with a complete gut ...200818931303
thermophilic degradation of phenolic compounds in lab scale hybrid up flow anaerobic sludge blanket reactors.this study describes the feasibility of anaerobic degradation of united states environmental protection agency (usepa) listed 4-chloro-2-nitrophenol (4c-2-np), 2-chloro-4-nitrophenol (2c-4-np), 2-chloro-5-methylphenol (2c-5-mp) from a simulated wastewater using four identical 7l bench scale hybrid up flow anaerobic sludge blankets (huasbs) at five different hydraulic retention times (hrts) under thermophilic condition (55+/-3 degrees c). the substrate to co-substrate ratios were maintained betwe ...200918986764
long-term monensin supplementation does not significantly affect the quantity or diversity of methanogens in the rumen of the lactating dairy cow.a long-term monensin supplementation trial involving lactating dairy cattle was conducted to determine the effect of monensin on the quantity and diversity of rumen methanogens in vivo. fourteen cows were paired on the basis of days in milk and parity and allocated to one of two treatment groups, receiving (i) a control total mixed ration (tmr) or (ii) a tmr with 24 mg of monensin premix/kg of diet dry matter. rumen fluid was obtained using an ororuminal probe on day -15 (baseline) and days 20, ...200919028912
quantitative detection of culturable methanogenic archaea abundance in anaerobic treatment systems using the sequence-specific rrna cleavage method.a method based on sequence-specific cleavage of rrna with ribonuclease h was used to detect almost all known cultivable methanogens in anaerobic biological treatment systems. to do so, a total of 40 scissor probes in different phylogeny specificities were designed or modified from previous studies, optimized for their specificities under digestion conditions with 32 methanogenic reference strains, and then applied to detect methanogens in sludge samples taken from 6 different anaerobic treatment ...200919212429
quantification of human polyomaviruses jc virus and bk virus by taqman quantitative pcr and comparison to other water quality indicators in water and fecal samples.in the united states, total maximum daily load standards for bodies of water that do not meet bacterial water quality standards are set by each state. the presence of human polyomaviruses (hpyvs) can be used as an indicator of human-associated sewage pollution in these waters. we have developed and optimized a taqman quantitative pcr (qpcr) assay based on the conserved t antigen to both quantify and simultaneously detect two hpyvs; jc virus and bk virus. the qpcr assay was able to consistently q ...200919346361
methanogen community structure in the rumens of farmed sheep, cattle and red deer fed different diets.development of inhibitors and vaccines that mitigate rumen-derived methane by targeting methanogens relies on knowledge of the methanogens present. we investigated the composition of archaeal communities in the rumens of farmed sheep (ovis aries), cattle (bos taurus) and red deer (cervus elaphus) using denaturing gradient gel electrophoresis (dgge) to generate fingerprints of archaeal 16s rrna genes. the total archaeal communities were relatively constant across species and diets, and were less ...201121255054
rumen-like methanogens identified from the crop of the folivorous south american bird, the hoatzin (opisthocomus hoazin).the hoatzin is the only known avian species with foregut fermentation. it is a primarily folivorous feeder and has a distended crop and lower/distal esophagus, which has evolved for the microbial fermentation of ingested feed. crop samples collected from 10 individual animals from the apure river area, apure state, venezuela were examined for the presence and density of methanogens using 16s rrna gene clone libraries and real-time pcr prepared from pooled and individual pcr products. a total of ...200919387486
validation and field testing of library-independent microbial source tracking methods in the gulf of mexico.water quality is frequently impacted by microbial pollution from human and animal feces. microbial source tracking (mst) can identify dominant pollution sources and improve assessment of health risk compared to indicator bacteria alone. this study aims to standardize and validate mst methods across laboratories in coastal gulf of mexico states. three laboratories evaluated library-independent mst methods for human sewage detection via conventional pcr: (1) human-associated bacteroidales, (2) hum ...200919595426
innovations in host and microbial sialic acid biosynthesis revealed by phylogenomic prediction of nonulosonic acid structure.sialic acids (sias) are nonulosonic acid (nulo) sugars prominently displayed on vertebrate cells and occasionally mimicked by bacterial pathogens using homologous biosynthetic pathways. it has been suggested that sias were an animal innovation and later emerged in pathogens by convergent evolution or horizontal gene transfer. to better illuminate the evolutionary processes underlying the phenomenon of sia molecular mimicry, we performed phylogenomic analyses of biosynthetic pathways for sias and ...200919666579
assessment of the microbial ecology of ruminal methanogens in cattle with different feed efficiencies.cattle with high feed efficiencies (designated "efficient") produce less methane gas than those with low feed efficiencies (designated "inefficient"); however, the role of the methane producers in such difference is unknown. this study investigated whether the structures and populations of methanogens in the rumen were associated with differences in cattle feed efficiencies by using culture-independent methods. two 16s rrna libraries were constructed using approximately 800-bp amplicons generate ...200919717632
high prevalence of methanobrevibacter smithii and methanosphaera stadtmanae detected in the human gut using an improved dna detection protocol.the low and variable prevalence of methanobrevibacter smithii and methanosphaera stadtmanae dna in human stool contrasts with the paramount role of these methanogenic archaea in digestion processes. we hypothesized that this contrast is a consequence of the inefficiencies of current protocols for archaeon dna extraction. we developed a new protocol for the extraction and pcr-based detection of m. smithii and m. stadtmanae dna in human stool.200919759898
monitoring bacterial community of human gut microbiota reveals an increase in lactobacillus in obese patients and methanogens in anorexic patients.studies of the bacterial communities of the gut microbiota have revealed a shift in the ratio of firmicutes and bacteroidetes in obese patients. determining the variations of microbial communities in feces may be beneficial for the identification of specific profiles in patients with abnormal weights. the roles of the archaeon methanobrevibacter smithii and lactobacillus species have not been described in these studies.200919774074
energetic analysis of the growth of methanobrevibacter arboriphilus a2 in hydrogen-limited continuous cultures.hydrogen-limited chemostat cultures of methanobrevibacter arboriphilus a2 were carried out. the available electron balance and carbon balance in m. arboriphilus a2 and other methanogenic strains grown on various substrates were well satisfied. this indicates that no extracellular organic products were formed during methanogenic growth. the molar growth yields for methane (y(x/ch(4) )) were calculated as 1.06-1.42 g cell/mol ch(4) at dilution rate (0.21-0.43 day(-1)). the smaller y(x/ch(4) ) of m ...198718576419
molecular diversity of the rumen microbiome of norwegian reindeer on natural summer pasture.the molecular diversity of the rumen microbiome was investigated in five semi-domesticated adult female norwegian reindeer (rangifer tarandus tarandus) grazing on natural summer pastures on the coast of northern norway (71.00 degrees n, 25.30 degrees e). mean population densities (numbers per gram wet weight) of methanogenic archaea, rumen bacteria and ciliate protozoa, estimated using quantitative real-time polymerase chain reaction (pcr), were 3.17x10(9), 5.17x10(11) and 4.02x10(7), respective ...200918604648
design and development of the anaerochip microarray for investigation of methanogenic communities.the aim of this study was to design a microarray targeting methanogens found in anaerobic digesters, and to apply this chip together with a cloning approach to investigate the methanogenic community present in an anaerobic digester. oligonucleotide probes were designed based on sequence differences in the 16s rrna genes in order to target microorganisms in situ. for microarray hybridisations, dna was subjected to pcr amplification of the 16s rrna gene and cy5-labeled. the microarray was tested w ...200919799937
a putative new order of methanogenic archaea inhabiting the human gut, as revealed by molecular analyses of the mcra gene.the diversity of methanogenic archaea from the gut of 6 humans was investigated by targeting mcra, a molecular metabolic marker of methanogenesis. three operational taxonomic units (otus) were recovered from about 400 clones analyzed, two of which were attributed to the expected methanobacteriales methanobrevibacter smithii (4 volunteers) and methanosphaera stadtmanae (1 volunteer). the third otu (1 volunteer) corresponded to a distant phylotype that does not cluster with any of the five methano ...200818644435
distribution of archaea in japanese patients with periodontitis and humoral immune response to the components.there is controversy regarding the existence of archaeal pathogens. periodontitis is one of the human diseases in which archaea have been suggested to have roles as pathogens. this study was performed to investigate the distribution of archaea in japanese patients with periodontitis and to examine the serum igg responses to archaeal components. subgingival plaque samples were collected from 111 periodontal pockets of 49 patients (17 with aggressive periodontitis and 32 with chronic periodontitis ...200818707623
functional and comparative metagenomic analysis of bile salt hydrolase activity in the human gut microbiome.bile salt hydrolases (bshs) catalyze the "gateway" reaction in a wider pathway of bile acid modification by the gut microbiota. because bile acids function as signaling molecules regulating their own biosynthesis, lipid absorption, cholesterol homeostasis, and local mucosal defenses in the intestine, microbial bsh activity has the potential to greatly influence host physiology. however, the function, distribution, and abundance of bsh enzymes in the gut community are unknown. here, we show that ...200818757757
characterization of variation in rumen methanogenic communities under different dietary and host feed efficiency conditions, as determined by pcr-denaturing gradient gel electrophoresis analysis.understanding ruminal methanogens is essential for greenhouse gas mitigation, as well as for improving animal performance in the livestock industry. it has been speculated that ruminal methanogenic diversity affects host feed efficiency and results in differences in methane production. this study examined methanogenic profiles in the rumen using culture-independent pcr-denaturing gradient gel electrophoresis (pcr-dgge) analysis for 56 beef cattle which differed in feed efficiency, as well as die ...201020418436
abundance and diversity of mucosa-associated hydrogenotrophic microbes in the healthy human colon.hydrogenotrophic microbiota have a significant impact on colonic health; however, little is known about their diversity and ecology in situ. here, molecular-based methods and multivariate analyses were used to examine the abundance and diversity of mucosa-associated hydrogenotrophic microbes in 90 biopsies collected from right colon, left colon and rectum of 25 healthy subjects. functional genes of all three hydrogenotrophic groups were detected in at least one colonic region of all subjects. me ...201121753800
formate-driven growth coupled with h(2) production.although a common reaction in anaerobic environments, the conversion of formate and water to bicarbonate and h(2) (with a change in gibbs free energy of δg° = +1.3 kj mol(-1)) has not been considered energetic enough to support growth of microorganisms. recently, experimental evidence for growth on formate was reported for syntrophic communities of moorella sp. strain amp and a hydrogen-consuming methanothermobacter species and of desulfovibrio sp. strain g11 and methanobrevibacter arboriphilus ...201020844539
molecular profiling and identification of methanogenic archaeal species from rabbit caecum.during a comparison of 16s rdna pcr-denaturant gradient gel electrophoresis (dgge) profiles of methanogenic archaea from rumen fluid, rabbit caecum and pig feces, a unique band common to all rabbit caecum samples was observed. dgge profiling also showed that the methanogen community from the new zealand white adult rabbits is different and less complex than the methanogen communities from the rumen and pig feces. small subunit ribosomal gene sequences of methanogenic archaea were subsequently re ...201020950344
lack of correlation between enterococcal counts and the presence of human specific fecal markers in mississippi creek and coastal waters.the objective of this study was to determine whether statistically valid correlations could be shown between enterococcal counts of samples from creek and coastal sites and the presence of two molecular, library-independent markers that specify human and/or sewage pollution. four hundred ninety samples were collected between august 2007 and april 2009 to determine enterococcal counts and the presence of genetic markers for the sewage indicator organisms methanobrevibacter smithii and bacteroidal ...201020950837
a real-time qpcr assay for the detection of the nifh gene of methanobrevibacter smithii, a potential indicator of sewage pollution.to develop a quantitative, real-time pcr assay to detect the nifh gene of methanobrevibacter smithii. methanobrevibacter smithii is a methanogenic archaea found in the intestinal tract of humans that may be a useful indicator of sewage pollution in water.201021070516
the discussion goes on: what is the role of euryarchaeota in humans?the human body (primarily the intestinal tract, the oral cavity, and the skin) harbours approximately 1,000 different bacterial species. however, the number of archaeal species known to colonize man seems to be confined to a handful of organisms within the class euryarchaeota (including methanobrevibacter smithii, m. oralis, and methanosphaera stadtmanae). in contrast to this conspicuously low diversity of archaea in humans their unique physiology in conjunction with the growing number of report ...201021253553
phylogenetic analysis of methanogens in the pig feces.in order to assess methanogen diversity in feces of pigs, archaeal 16s rrna gene clone libraries were constructed from feces of the pig. after the amplification by pcr using primers met86f and met1340r, equal quantities of pcr products from each of the five pigs were mixed together and used to construct the library. sequence analysis showed that the 74 clones were divided into ten phylotypes as defined by rflp analysis. phylogenetic analysis showed that three phylotypes were most closely affilia ...201121259005
pan-genome of the dominant human gut-associated archaeon, methanobrevibacter smithii, studied in twins.the human gut microbiota harbors three main groups of h(2)-consuming microbes: methanogens including the dominant archaeon, methanobrevibacter smithii, a polyphyletic group of acetogens, and sulfate-reducing bacteria. defining their roles in the gut is important for understanding how hydrogen metabolism affects the efficiency of fermentation of dietary components. we quantified methanogens in fecal samples from 40 healthy adult female monozygotic (mz) and 28 dizygotic (dz) twin pairs, analyzed b ...201121317366
archaeosome adjuvant overcomes tolerance to tumor-associated melanoma antigens inducing protective cd8 t cell responses.vesicles comprised of the ether glycerolipids of the archaeon methanobrevibacter smithii (archaeosomes) are potent adjuvants for evoking cd8(+) t cell responses. we therefore explored the ability of archaeosomes to overcome immunologic tolerance to self-antigens. priming and boosting of mice with archaeosome-antigen evoked comparable cd8(+) t cell response and tumor protection to an alternate boosting strategy utilizing live bacterial vectors for antigen delivery. vaccination with melanoma antig ...201021318177
prevalence and microbiological diversity of archaea in peri-implantitis subjects by 16s ribosomal rna clonal analysis.this study evaluated the prevalence and the molecular diversity of archaea in the subgingival biofilm samples of subjects with peri-implantitis.201121338359
performance of a three-stage membrane bioprocess treating the organic fraction of municipal solid waste and evolution of its archaeal and bacterial ecology.a novel three-stage bioprocess achieved 75% volatile solids (vs) removal at an organic loading rate (olr) of 4g vsl(-1)day, a solids retention time (srt) of 66days, a hydraulic retention time (hrt) of 20days, at a temperature of 35 degrees c. the bioprocess consisted of an anaerobic hydrolytic reactor (hr) where the solids and liquid fractions of the organic fraction of the municipal solid waste (ofmsw) were separated with a mesh. the leachate was pumped to a submerged anaerobic membrane bioreac ...201019857960
culture-based indicators of fecal contamination and molecular microbial indicators rarely correlate with campylobacter spp. in recreational waters.campylobacter spp. are the leading cause of gastroenteritis worldwide. most human infections result from contaminated food; however, infections are also caused by recreational waterway contamination. campylobacter culture is technically challenging and enumeration by culture-based methods is onerous. thus, we employed qpcr to quantify campylobacter spp. in fresh- and marine-water samples, raw sewage and animal feces. multiplex pcr determined whether campylobacter jejuni or c. coli, most commonly ...201122048429
microbial population dynamics during startup of a full-scale anaerobic digester treating industrial food waste in kyoto eco-energy project.the microbial community in a full-scale anaerobic digester (2300m3) treating industrial food waste in the kyoto eco-energy project was analyzed using terminal restriction fragment length polymorphism for eubacterial and archaeal 16s rrna genes. both thermophilic and mesophilic sludge of treated swine waste were seeded to the digestion tank. during the 150-day startup period, coffee grounds as a main food waste, along with potato, kelp and boiled beans, tofu, bean curd lees, and deep-fried bean c ...201020129780
development of a vaccine to mitigate greenhouse gas emissions in agriculture: vaccination of sheep with methanogen fractions induces antibodies that block methane production in vitro.to develop an understanding of the immune responses of ruminants to methanogens, and to provide proof of a concept that harnessing the immune system of ruminants is a potentially viable approach to mitigate greenhouse gas emissions from agriculture.201020200573
the role of methane in intestinal diseases.the volume of human intestinal gas is about 200 ml, and it is derived from complex physiological processes including swallowed air, diffusion from bloodstream into the lumen, and particularly intraluminal production by chemical reactions and bacterial fermentation. gas is continuously removed by eructation, anal evacuation, absorption through the intestinal mucosa, and bacterial consumption. more than 99% of it is composed of hydrogen, oxygen, carbon dioxide, nitrogen, and other odoriferous gase ...201020216536
microbial equol production attenuates colonic methanogenesis and sulphidogenesis in vitro.hydrogen gas produced during colonic fermentation is excreted in breath and flatus, or removed by hydrogen-consuming bacteria such as methanogens and sulphate-reducing bacteria. however, recent research has shown that h2 is also consumed by equol-producing bacteria during the reduction of daidzein into equol. in this study, the interactions between methanogens, sulphate-reducing, and equol-producing bacteria were investigated under in vitro simulated intestinal conditions. in the presence of dai ...201020298796
antigenic group ii chaperonin in methanobrevibacter oralis may cross-react with human chaperonin cct.methanobrevibacter oralis is an archaeal species frequently isolated from sites of severe periodontitis. however, its pathogenic roles remain unclear. here, we aimed to isolate group ii chaperonin from m. oralis and examine its antigenicity. the genes encoding two chaperonin subunits (cpn-1 and cpn-2) were cloned from m. oralis using polymerase chain reaction and genome walking procedures. recombinant proteins cpn-1 and cpn-2 were generated, and the reactivities of sera from patients with period ...201020331799
differences in the rumen methanogen populations of lactating jersey and holstein dairy cows under the same diet regimen.in the dairy cattle industry, holstein and jersey are the breeds most commonly used for production. they differ in performance by various traits, such as body size, milk production, and milk composition. with increased concerns about the impact of agriculture on climate change, potential differences in other traits, such as methane emission, also need to be characterized further. since methane is produced in the rumen by methanogenic archaea, we investigated whether the population structure of m ...201121705541
the antimicrobial resistance pattern of cultured human methanogens reflects the unique phylogenetic position of archaea.objectives methanogenic archaea are constant members of the human oral and digestive microbiota retrieved, in particular, from periodontitis lesions. the objective of the study was to determine their susceptibility to antimicrobials. methods using the macrodilution method in hungate tubes with optical microscope observation combined with monitoring methane production, we determined the antibiotic resistance characteristics of eight methanogenic archaea. results methanobrevibacter smithii strains ...201121680581
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