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Svetlana N. Dedysh - One of the best experts on this subject based on the ideXlab platform.
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Genomic Determinants of Phototrophy in Methanotrophic Alphaproteobacteria
Microbiology, 2019Co-Authors: Kirill K Miroshnikov, S. E. Belova, Svetlana N. DedyshAbstract:— Aerobic methanotrophic bacteria are an ecologically important group of microorganisms, which are functionally specialized in oxidation of the greenhouse gas methane. Recent insights into the growing pool of available genome sequences from methanotrophs revealed a number of as-yet-unknown metabolic capabilities of these bacteria. Thus, the genes indicative of aerobic anoxygenic photosynthesis by means of the photosystem II characteristic of purple bacteria were revealed in the genome of an obligate methanotroph Methylocapsa palsarum NE2^T. Advanced search for genomic determinants of phototrophy in other methanotrophs confirmed their occurrence in a number of methanotrophic Alphaproteobacteria , including Methylocella silvestris BL2^T and TVC, Methylocystis rosea SV97^T and GW6, as well as Methylocystis spp. strains SB2 and MitZ-2018. Genomes of these methanotrophs contained the pufABCLM gene clusters encoding the light-harvesting complex, bch/chl genes responsible for bacteriochlorophyll biosynthesis, and the pucC gene essential for bacteriochlorophyll transport, as well as the crtFDC , crtL and crtB genes responsible for carotenoid biosynthesis. Organization of these gene clusters was conserved within each methanotroph species and was highly similar in Methylocapsa and Methylocella strains. A number of rearrangements, including inverse localization of the genes encoding bacteriochlorophyll and carotenoid biosynthesis, were observed in the genomes of Methylocystis species. The presence of pufLM genes was also revealed in a new isolate of Methylocapsa palsarum , strain NSB8, which was obtained in this study from a tundra wetland of European Northern Russia. The presence of phototrophy-related genes in all available strains of the abovementioned species indicates their functional importance for these bacteria and suggests realization of the phototrophic potential under certain environmental conditions.
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complete genome sequence of the aerobic facultative methanotroph Methylocella tundrae strain t4
Microbiology resource announcements, 2019Co-Authors: Muhammad Farhan Ul Haque, Andrew T Crombie, Svetlana N. Dedysh, Theo A Van Alen, Mike S M Jetten, Huub Op Den J M Camp, Maartje A H J Van KesselAbstract:Methylocella tundrae T4T is a facultative aerobic methanotroph which was isolated from an acidic tundra wetland and possesses only a soluble methane monooxygenase. The complete genome, which includes two megaplasmids, was sequenced using a combination of Illumina and Nanopore technologies. One of the megaplasmids carries a propane monooxygenase gene cluster.
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Methylocella: a gourmand among methanotrophs.
Trends in microbiology, 2014Co-Authors: Peter F. Dunfield, Svetlana N. DedyshAbstract:A recent article in Nature describes the ability of Methylocella silvestris to grow simultaneously on methane and longer chain alkanes, something never before observed in the microbial world. It adds to a growing list of unique metabolic traits that distinguish Methylocella from any other bacterium.
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Methyloferula stellata gen. nov., sp. nov., an acidophilic, obligately methanotrophic bacterium that possesses only a soluble methane monooxygenase.
International journal of systematic and evolutionary microbiology, 2010Co-Authors: Alexey V Vorobev, Werner Liesack, Allyson L. Brady, Peter F. Dunfield, Mohamed Baani, Nina V Doronina, Svetlana N. DedyshAbstract:Two strains of aerobic methanotrophic bacteria, AR4(T) and SOP9, were isolated from acidic (pH 3.8-4.0) Sphagnum peat bogs in Russia. Another phenotypically similar isolate, strain LAY, was obtained from an acidic (pH 4.0) forest soil in Germany. Cells of these strains were Gram-negative, non-pigmented, non-motile, thin rods that multiplied by irregular cell division and formed rosettes or amorphous cell conglomerates. Similar to Methylocella species, strains AR4(T), SOP9 and LAY possessed only a soluble form of methane monooxygenase (sMMO) and lacked intracytoplasmic membranes. Growth occurred only on methane and methanol; the latter was the preferred growth substrate. mRNA transcripts of sMMO were detectable in cells when either methane or both methane and methanol were available. Carbon was assimilated via the serine and ribulose-bisphosphate (RuBP) pathways; nitrogen was fixed via an oxygen-sensitive nitrogenase. Strains AR4(T), SOP9 and LAY were moderately acidophilic, mesophilic organisms capable of growth between pH 3.5 and 7.2 (optimum pH 4.8-5.2) and at 4-33 °C (optimum 20-23 °C). The major cellular fatty acid was 18 : 1ω7c and the quinone was Q-10. The DNA G+C content was 55.6-57.5 mol%. The isolates belonged to the family Beijerinckiaceae of the class Alphaproteobacteria and were most closely related to the sMMO-possessing methanotrophs of the genus Methylocella (96.4-97.0 % 16S rRNA gene sequence similarity), particulate MMO (pMMO)-possessing methanotrophs of the genus Methylocapsa (96.1-97.0 %), facultative methylotrophs of the genus Methylovirgula (96.1-96.3 %) and non-methanotrophic organotrophs of the genus Beijerinckia (96.5-97.0 %). Phenotypically, strains AR4(T), SOP9 and LAY were most similar to Methylocella species, but differed from members of this genus by cell morphology, greater tolerance of low pH, detectable activities of RuBP pathway enzymes and inability to grow on multicarbon compounds. Therefore, we propose a novel genus and species, Methyloferula stellata gen. nov., sp. nov., to accommodate strains AR4(T), SOP9 and LAY. Strain AR4(T) ( = DSM 22108(T) = LMG 25277(T) = VKM B-2543(T)) is the type strain of Methyloferula stellata.
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Methylocapsa aurea sp. nov., a facultative methanotroph possessing a particulate methane monooxygenase, and emended description of the genus Methylocapsa.
International Journal of Systematic and Evolutionary Microbiology, 2010Co-Authors: Peter F. Dunfield, Alexey Vorobev, Svetlana E. Belova, Sabrina L. Cornish, Svetlana N. DedyshAbstract:An aerobic, methanotrophic bacterium, designated KYGT, was isolated from a forest soil in Germany. Cells of strain KYGT were Gram-negative, non-motile, slightly curved rods that multiplied by binary fission and produced yellow colonies. The cells contained intracellular granules of poly-β-hydroxybutyrate at each cell pole, a particulate methane monooxygenase (pMMO) and stacks of intracytoplasmic membranes (ICMs) packed in parallel along one side of the cell envelope. Strain KYGT grew at pH 5.2–7.2 and 2–33 °C and could fix atmospheric nitrogen under reduced oxygen tension. The major cellular fatty acid was C18 : 1 ω7c (81.5 %) and the DNA G+C content was 61.4 mol%. Strain KYGT belonged to the family Beijerinckiaceae of the class Alphaproteobacteria and was most closely related to the obligate methanotroph Methylocapsa acidiphila B2T (98.1 % 16S rRNA gene sequence similarity and 84.7 % pmoA sequence similarity). Unlike Methylocapsa acidiphila B2T, which grows only on methane and methanol, strain KYGT was able to grow facultatively on acetate. Facultative acetate utilization is a characteristic of the methanotrophs of the genus Methylocella, but the genus Methylocella does not produce pMMO or ICMs. Strain KYGT differed from Methylocapsa acidiphila B2T on the basis of substrate utilization pattern, pigmentation, pH range, cell ultrastructure and efficiency of dinitrogen fixation. Therefore, we propose a novel species, Methylocapsa aurea sp. nov., to accommodate this bacterium. The type strain is KYGT (=DSM 22158T =VKM B-2544T).
Andrew T Crombie - One of the best experts on this subject based on the ideXlab platform.
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Facultative methanotrophs - diversity, genetics, molecular ecology and biotechnological potential: a mini-review.
Microbiology (Reading England), 2020Co-Authors: Muhammad Farhan Ul Haque, Andrew T CrombieAbstract:Methane-oxidizing bacteria (methanotrophs) play a vital role in reducing atmospheric methane emissions, and hence mitigating their potent global warming effects. A significant proportion of the methane released is thermogenic natural gas, containing associated short-chain alkanes as well as methane. It was one hundred years following the description of methanotrophs that facultative strains were discovered and validly described. These can use some multi-carbon compounds in addition to methane, often small organic acids, such as acetate, or ethanol, although Methylocella strains can also use short-chain alkanes, presumably deriving a competitive advantage from this metabolic versatility. Here, we review the diversity and molecular ecology of facultative methanotrophs. We discuss the genetic potential of the known strains and outline the consequent benefits they may obtain. Finally, we review the biotechnological promise of these fascinating microbes.
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Genome Scale Metabolic Model of the versatile methanotroph Methylocella silvestris.
Microbial cell factories, 2020Co-Authors: Sergio Bordel, Andrew T Crombie, Raúl MuñozAbstract:Methylocella silvestris is a facultative aerobic methanotrophic bacterium which uses not only methane, but also other alkanes such as ethane and propane, as carbon and energy sources. Its high metabolic versatility, together with the availability of tools for its genetic engineering, make it a very promising platform for metabolic engineering and industrial biotechnology using natural gas as substrate. The first Genome Scale Metabolic Model for M. silvestris is presented. The model has been used to predict the ability of M. silvestris to grow on 12 different substrates, the growth phenotype of two deletion mutants (ΔICL and ΔMS), and biomass yield on methane and ethanol. The model, together with phenotypic characterization of the deletion mutants, revealed that M. silvestris uses the glyoxylate shuttle for the assimilation of C1 and C2 substrates, which is unique in contrast to published reports of other methanotrophs. Two alternative pathways for propane metabolism have been identified and validated experimentally using enzyme activity tests and constructing a deletion mutant (Δ1641), which enabled the identification of acetol as one of the intermediates of propane assimilation via 2-propanol. The model was also used to integrate proteomic data and to identify key enzymes responsible for the adaptation of M. silvestris to different substrates. The model has been used to elucidate key metabolic features of M. silvestris, such as its use of the glyoxylate shuttle for the assimilation of one and two carbon compounds and the existence of two parallel metabolic pathways for propane assimilation. This model, together with the fact that tools for its genetic engineering already exist, paves the way for the use of M. silvestris as a platform for metabolic engineering and industrial exploitation of methanotrophs.
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Novel facultative Methylocella strains are active methane consumers at terrestrial natural gas seeps.
Microbiome, 2019Co-Authors: Muhammad Farhan Ul Haque, Andrew T CrombieAbstract:Natural gas seeps contribute to global climate change by releasing substantial amounts of the potent greenhouse gas methane and other climate-active gases including ethane and propane to the atmosphere. However, methanotrophs, bacteria capable of utilising methane as the sole source of carbon and energy, play a significant role in reducing the emissions of methane from many environments. Methylocella-like facultative methanotrophs are a unique group of bacteria that grow on other components of natural gas (i.e. ethane and propane) in addition to methane but a little is known about the distribution and activity of Methylocella in the environment. The purposes of this study were to identify bacteria involved in cycling methane emitted from natural gas seeps and, most importantly, to investigate if Methylocella-like facultative methanotrophs were active utilisers of natural gas at seep sites. The community structure of active methane-consuming bacteria in samples from natural gas seeps from Andreiasu Everlasting Fire (Romania) and Pipe Creek (NY, USA) was investigated by DNA stable isotope probing (DNA-SIP) using 13C-labelled methane. The 16S rRNA gene sequences retrieved from DNA-SIP experiments revealed that of various active methanotrophs, Methylocella was the only active methanotrophic genus common to both natural gas seep environments. We also isolated novel facultative methanotrophs, Methylocella sp. PC1 and PC4 from Pipe Creek, able to utilise methane, ethane, propane and various non-gaseous multicarbon compounds. Functional and comparative genomics of these new isolates revealed genomic and physiological divergence from already known methanotrophs, in particular, the absence of mxa genes encoding calcium-containing methanol dehydrogenase. Methylocella sp. PC1 and PC4 had only the soluble methane monooxygenase (sMMO) and lanthanide-dependent methanol dehydrogenase (XoxF). These are the first Alphaproteobacteria methanotrophs discovered with this reduced functional redundancy for C-1 metabolism (i.e. sMMO only and XoxF only). Here, we provide evidence, using culture-dependent and culture-independent methods, that Methylocella are abundant and active at terrestrial natural gas seeps, suggesting that they play a significant role in the biogeochemical cycling of these gaseous alkanes. This might also be significant for the design of biotechnological strategies for controlling natural gas emissions, which are increasing globally due to unconventional exploitation of oil and gas.
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complete genome sequence of the aerobic facultative methanotroph Methylocella tundrae strain t4
Microbiology resource announcements, 2019Co-Authors: Muhammad Farhan Ul Haque, Andrew T Crombie, Svetlana N. Dedysh, Theo A Van Alen, Mike S M Jetten, Huub Op Den J M Camp, Maartje A H J Van KesselAbstract:Methylocella tundrae T4T is a facultative aerobic methanotroph which was isolated from an acidic tundra wetland and possesses only a soluble methane monooxygenase. The complete genome, which includes two megaplasmids, was sequenced using a combination of Illumina and Nanopore technologies. One of the megaplasmids carries a propane monooxygenase gene cluster.
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Draft Genome Sequence of Methylocella silvestris TVC, a Facultative Methanotroph Isolated from Permafrost.
Genome announcements, 2018Co-Authors: Jing Wang, Andrew T Crombie, Muhammad Farhan Ul Haque, Kan Geng, Lorna E. Street, Philip A. Wookey, Jennifer PratscherAbstract:Permafrost environments play a crucial role in global carbon and methane cycling. We report here the draft genome sequence of Methylocella silvestris TVC, a new facultative methanotroph strain, isolated from the Siksik Creek catchment in the continuous permafrost zone of Inuvik (Northwest Territories, Canada).
Peter F. Dunfield - One of the best experts on this subject based on the ideXlab platform.
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differential transcriptional activation of genes encoding soluble methane monooxygenase in a facultative versus an obligate methanotroph
Microorganisms, 2018Co-Authors: Angela V Smirnova, Peter F. DunfieldAbstract:Methanotrophs are a specialized group of bacteria that can utilize methane (CH4) as a sole energy source. A key enzyme responsible for methane oxidation is methane monooxygenase (MMO), of either a soluble, cytoplasmic type (sMMO), or a particulate, membrane-bound type (pMMO). Methylocella silvestris BL2 and Methyloferula stellata AR4 are closely related methanotroph species that oxidize methane via sMMO only. However, Methyloferula stellata is an obligate methanotroph, while Methylocella silvestris is a facultative methanotroph able to grow on several multicarbon substrates in addition to methane. We constructed transcriptional fusions of the mmo promoters of Methyloferula stellata and Methylocella silvestris to a promoterless gfp in order to compare their transcriptional regulation in response to different growth substrates, in the genetic background of both organisms. The following patterns were observed: (1) The mmo promoter of the facultative methanotroph Methylocella silvestris was either transcriptionally downregulated or repressed by any growth substrate other than methane in the genetic background of Methylocella silvetris; (2) Growth on methane alone upregulated the mmo promoter of Methylocella silvetris in its native background but not in the obligate methanotroph Methyloferula stellata; (3) The mmo promoter of Methyloferula stellata was constitutive in both organisms regardless of the growth substrate, but with much lower promoter activity than the mmo promoter of Methylocella silvetris. These results support a conclusion that a different mode of transcriptional regulation of sMMO contributes to the facultative lifestyle of Methylocella silvetris compared to the obligate methanotroph Methyloferula stellata.
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Methylocella: a gourmand among methanotrophs.
Trends in microbiology, 2014Co-Authors: Peter F. Dunfield, Svetlana N. DedyshAbstract:A recent article in Nature describes the ability of Methylocella silvestris to grow simultaneously on methane and longer chain alkanes, something never before observed in the microbial world. It adds to a growing list of unique metabolic traits that distinguish Methylocella from any other bacterium.
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The (d)evolution of methanotrophy in the Beijerinckiaceae--a comparative genomics analysis.
The ISME journal, 2013Co-Authors: Ivica Tamas, Angela V Smirnova, Peter F. DunfieldAbstract:The alphaproteobacterial family Beijerinckiaceae contains generalists that grow on a wide range of substrates, and specialists that grow only on methane and methanol. We investigated the evolution of this family by comparing the genomes of the generalist organotroph Beijerinckia indica, the facultative methanotroph Methylocella silvestris and the obligate methanotroph Methylocapsa acidiphila. Highly resolved phylogenetic construction based on universally conserved genes demonstrated that the Beijerinckiaceae forms a monophyletic cluster with the Methylocystaceae, the only other family of alphaproteobacterial methanotrophs. Phylogenetic analyses also demonstrated a vertical inheritance pattern of methanotrophy and methylotrophy genes within these families. Conversely, many lateral gene transfer (LGT) events were detected for genes encoding carbohydrate transport and metabolism, energy production and conversion, and transcriptional regulation in the genome of B. indica, suggesting that it has recently acquired these genes. A key difference between the generalist B. indica and its specialist methanotrophic relatives was an abundance of transporter elements, particularly periplasmic-binding proteins and major facilitator transporters. The most parsimonious scenario for the evolution of methanotrophy in the Alphaproteobacteria is that it occurred only once, when a methylotroph acquired methane monooxygenases (MMOs) via LGT. This was supported by a compositional analysis suggesting that all MMOs in Alphaproteobacteria methanotrophs are foreign in origin. Some members of the Beijerinckiaceae subsequently lost methanotrophic functions and regained the ability to grow on multicarbon energy substrates. We conclude that B. indica is a recidivist multitroph, the only known example of a bacterium having completely abandoned an evolved lifestyle of specialized methanotrophy.
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Methyloferula stellata gen. nov., sp. nov., an acidophilic, obligately methanotrophic bacterium that possesses only a soluble methane monooxygenase.
International journal of systematic and evolutionary microbiology, 2010Co-Authors: Alexey V Vorobev, Werner Liesack, Allyson L. Brady, Peter F. Dunfield, Mohamed Baani, Nina V Doronina, Svetlana N. DedyshAbstract:Two strains of aerobic methanotrophic bacteria, AR4(T) and SOP9, were isolated from acidic (pH 3.8-4.0) Sphagnum peat bogs in Russia. Another phenotypically similar isolate, strain LAY, was obtained from an acidic (pH 4.0) forest soil in Germany. Cells of these strains were Gram-negative, non-pigmented, non-motile, thin rods that multiplied by irregular cell division and formed rosettes or amorphous cell conglomerates. Similar to Methylocella species, strains AR4(T), SOP9 and LAY possessed only a soluble form of methane monooxygenase (sMMO) and lacked intracytoplasmic membranes. Growth occurred only on methane and methanol; the latter was the preferred growth substrate. mRNA transcripts of sMMO were detectable in cells when either methane or both methane and methanol were available. Carbon was assimilated via the serine and ribulose-bisphosphate (RuBP) pathways; nitrogen was fixed via an oxygen-sensitive nitrogenase. Strains AR4(T), SOP9 and LAY were moderately acidophilic, mesophilic organisms capable of growth between pH 3.5 and 7.2 (optimum pH 4.8-5.2) and at 4-33 °C (optimum 20-23 °C). The major cellular fatty acid was 18 : 1ω7c and the quinone was Q-10. The DNA G+C content was 55.6-57.5 mol%. The isolates belonged to the family Beijerinckiaceae of the class Alphaproteobacteria and were most closely related to the sMMO-possessing methanotrophs of the genus Methylocella (96.4-97.0 % 16S rRNA gene sequence similarity), particulate MMO (pMMO)-possessing methanotrophs of the genus Methylocapsa (96.1-97.0 %), facultative methylotrophs of the genus Methylovirgula (96.1-96.3 %) and non-methanotrophic organotrophs of the genus Beijerinckia (96.5-97.0 %). Phenotypically, strains AR4(T), SOP9 and LAY were most similar to Methylocella species, but differed from members of this genus by cell morphology, greater tolerance of low pH, detectable activities of RuBP pathway enzymes and inability to grow on multicarbon compounds. Therefore, we propose a novel genus and species, Methyloferula stellata gen. nov., sp. nov., to accommodate strains AR4(T), SOP9 and LAY. Strain AR4(T) ( = DSM 22108(T) = LMG 25277(T) = VKM B-2543(T)) is the type strain of Methyloferula stellata.
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Methylocapsa aurea sp. nov., a facultative methanotroph possessing a particulate methane monooxygenase, and emended description of the genus Methylocapsa.
International Journal of Systematic and Evolutionary Microbiology, 2010Co-Authors: Peter F. Dunfield, Alexey Vorobev, Svetlana E. Belova, Sabrina L. Cornish, Svetlana N. DedyshAbstract:An aerobic, methanotrophic bacterium, designated KYGT, was isolated from a forest soil in Germany. Cells of strain KYGT were Gram-negative, non-motile, slightly curved rods that multiplied by binary fission and produced yellow colonies. The cells contained intracellular granules of poly-β-hydroxybutyrate at each cell pole, a particulate methane monooxygenase (pMMO) and stacks of intracytoplasmic membranes (ICMs) packed in parallel along one side of the cell envelope. Strain KYGT grew at pH 5.2–7.2 and 2–33 °C and could fix atmospheric nitrogen under reduced oxygen tension. The major cellular fatty acid was C18 : 1 ω7c (81.5 %) and the DNA G+C content was 61.4 mol%. Strain KYGT belonged to the family Beijerinckiaceae of the class Alphaproteobacteria and was most closely related to the obligate methanotroph Methylocapsa acidiphila B2T (98.1 % 16S rRNA gene sequence similarity and 84.7 % pmoA sequence similarity). Unlike Methylocapsa acidiphila B2T, which grows only on methane and methanol, strain KYGT was able to grow facultatively on acetate. Facultative acetate utilization is a characteristic of the methanotrophs of the genus Methylocella, but the genus Methylocella does not produce pMMO or ICMs. Strain KYGT differed from Methylocapsa acidiphila B2T on the basis of substrate utilization pattern, pigmentation, pH range, cell ultrastructure and efficiency of dinitrogen fixation. Therefore, we propose a novel species, Methylocapsa aurea sp. nov., to accommodate this bacterium. The type strain is KYGT (=DSM 22158T =VKM B-2544T).
Werner Liesack - One of the best experts on this subject based on the ideXlab platform.
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Methyloferula stellata gen. nov., sp. nov., an acidophilic, obligately methanotrophic bacterium that possesses only a soluble methane monooxygenase.
International journal of systematic and evolutionary microbiology, 2010Co-Authors: Alexey V Vorobev, Werner Liesack, Allyson L. Brady, Peter F. Dunfield, Mohamed Baani, Nina V Doronina, Svetlana N. DedyshAbstract:Two strains of aerobic methanotrophic bacteria, AR4(T) and SOP9, were isolated from acidic (pH 3.8-4.0) Sphagnum peat bogs in Russia. Another phenotypically similar isolate, strain LAY, was obtained from an acidic (pH 4.0) forest soil in Germany. Cells of these strains were Gram-negative, non-pigmented, non-motile, thin rods that multiplied by irregular cell division and formed rosettes or amorphous cell conglomerates. Similar to Methylocella species, strains AR4(T), SOP9 and LAY possessed only a soluble form of methane monooxygenase (sMMO) and lacked intracytoplasmic membranes. Growth occurred only on methane and methanol; the latter was the preferred growth substrate. mRNA transcripts of sMMO were detectable in cells when either methane or both methane and methanol were available. Carbon was assimilated via the serine and ribulose-bisphosphate (RuBP) pathways; nitrogen was fixed via an oxygen-sensitive nitrogenase. Strains AR4(T), SOP9 and LAY were moderately acidophilic, mesophilic organisms capable of growth between pH 3.5 and 7.2 (optimum pH 4.8-5.2) and at 4-33 °C (optimum 20-23 °C). The major cellular fatty acid was 18 : 1ω7c and the quinone was Q-10. The DNA G+C content was 55.6-57.5 mol%. The isolates belonged to the family Beijerinckiaceae of the class Alphaproteobacteria and were most closely related to the sMMO-possessing methanotrophs of the genus Methylocella (96.4-97.0 % 16S rRNA gene sequence similarity), particulate MMO (pMMO)-possessing methanotrophs of the genus Methylocapsa (96.1-97.0 %), facultative methylotrophs of the genus Methylovirgula (96.1-96.3 %) and non-methanotrophic organotrophs of the genus Beijerinckia (96.5-97.0 %). Phenotypically, strains AR4(T), SOP9 and LAY were most similar to Methylocella species, but differed from members of this genus by cell morphology, greater tolerance of low pH, detectable activities of RuBP pathway enzymes and inability to grow on multicarbon compounds. Therefore, we propose a novel genus and species, Methyloferula stellata gen. nov., sp. nov., to accommodate strains AR4(T), SOP9 and LAY. Strain AR4(T) ( = DSM 22108(T) = LMG 25277(T) = VKM B-2543(T)) is the type strain of Methyloferula stellata.
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Complete Genome Sequence of Beijerinckia indica subsp. indica
Journal of bacteriology, 2010Co-Authors: Ivica Tamas, Werner Liesack, Svetlana N. Dedysh, Matthew B. Stott, Maqsudul Alam, J. C. Murrell, Peter F. DunfieldAbstract:Beijerinckia indica subsp. indica is an aerobic, acidophilic, exopolysaccharide-producing, N2-fixing soil bacterium. It is a generalist chemoorganotroph that is phylogenetically closely related to facultative and obligate methanotrophs of the genera Methylocella and Methylocapsa. Here we report the full genome sequence of this bacterium.
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Complete Genome Sequence of the Aerobic Facultative Methanotroph Methylocella silvestris BL2
Journal of bacteriology, 2010Co-Authors: Andrew T Crombie, Andreas R. Theisen, Werner Liesack, Svetlana N. Dedysh, M.t. Rahman, Matthew B. Stott, Maqsudul Alam, Peter F. DunfieldAbstract:Methylocella silvestris BL2 is an aerobic methanotroph originally isolated from an acidic forest soil in Germany. It is the first fully authenticated facultative methanotroph. It grows not only on methane and other one-carbon (C1) substrates, but also on some compounds containing carbon-carbon bonds, such as acetate, pyruvate, propane, and succinate. Here we report the full genome sequence of this bacterium.
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first genome data from uncultured upland soil cluster alpha methanotrophs provide further evidence for a close phylogenetic relationship to methylocapsa acidiphila b2 and for high affinity methanotrophy involving particulate methane monooxygenase
Applied and Environmental Microbiology, 2005Co-Authors: Peter Ricke, Michael Kube, Satoshi Nakagawa, Christoph Erkel, Richard Reinhardt, Werner LiesackAbstract:The only biological sink for atmospheric CH4 is its consumption in oxic soils (7, 51). Recently, the global terrestrial sink was estimated at 29 Tg year−1, with a wide range of uncertainty (7 to >100 Tg CH4 year−1) (51). Atmospheric CH4 is consumed in forest, agricultural, and other upland soils by aerobic methanotrophic bacteria (MB). The CH4-consuming activity of these MB is significant because the magnitude of atmospheric CH4 uptake in oxic soils is similar to the estimated excess of emissions over sinks in recent years of 37 Tg year−1 (28). Moreover, methanotrophic activity is highly susceptible to disturbance by human activities (32, 51). Cultured MB are divided into two groups, namely, type I (further divided into types I and X) and type II. They differ in their phylogenetic affiliations (Gammaproteobacteria versus Alphaproteobacteria) and in diverse biochemical and ultrastructural characteristics (23). However, this traditional concept of methanotroph classification has become much more complex by recent descriptions of the genera Methylocella (9, 12, 17) and Methylocapsa (11). These acidophilic MB, although considered members of the type II MB, possess several unique morphological and physiological characteristics, and based on 16S rRNA phylogeny, Methylocella and Methylocapsa are more closely related to acidophilic heterotrophic bacteria of the genus Beijerinckia than to alphaproteobacterial type II MB of the Methylosinus/Methylocystis group. Methane monooxygenase (MMO) catalyzes the first step in the metabolic pathway of MB, the conversion of CH4 to methanol. All cultivated MB possess a particulate, membrane-associated form (pMMO) of this enzyme, except for members of the genus Methylocella (9, 12, 17). pMMO is encoded by three consecutive open reading frames (pmoC, pmoA, and pmoB) in both type I and type II MB. The identity of the microorganisms consuming atmospheric CH4 remained unclear for many years (7, 45, 49). The apparent half-saturation constant (Km) for the oxidation of atmospheric CH4 in upland soil ranges from 0.8 to 280 nM. However, the Km of cultured type I and type II MB (0.8 to 66 μM) are 1 to 3 orders of magnitude higher, and these MB are not able to survive prolonged periods using only atmospheric CH4 (7, 44, 49). It was later shown that the apparent affinity for CH4 varies with growth conditions and that type II MB of the Methylosinus/Methylocystis group might contribute to the oxidation of atmospheric CH4 in soils (15, 16). Nonetheless, the use of pmoA as a functional gene marker for the molecular characterization of methanotrophic communities revealed that, besides members of known methanotrophic genera, a novel group of MB occurs in upland soils that consume atmospheric CH4 (4, 24, 27, 33). The most recent study suggested that these novel MB (originally named forest soil cluster but now referred to as upland soil cluster alpha [USCα]) are present mainly in acidic soils with pH values of <6 (33). Activity profiles of 13C- and 14C-labeled phospholipid fatty acids provided evidence that members of USCα are highly adapted to the consumption of atmospheric CH4 and are presumably of alphaproteobacterial origin (5, 27, 33, 46). Phylogenetic trees constructed for the pmoA gene suggested that among the cultured MB, Methylocapsa acidiphila B2 is most closely related to USCα (10). In general, pmoA and 16S rRNA phylogenies show good correlation (33, 35). Despite this, the evolutionary identity of USCα is still uncertain because (i) the pmoA gene enables the inference of putative relationships only among MB and (ii) the recent detection of multiple, diverse pmoA-like genes in single genospecies of MB implies that pmoA phylogenies have to be interpreted with caution (55). Moreover, the apparent affinity for methane exhibited by M. acidiphila B2 was 1 to 2 μM, which is similar to values measured in other cultured type I and type II MB (10). Except for phospholipid fatty acid profiles and partial pmoA sequences, no additional pheno- or genotypic information is available for USCα. The aim of our study was to gain the first insight into the genome of a USCα representative by using a metagenomic approach (13, 22). This included the extraction of high-molecular-weight (HMW) DNA from acidic forest soil, the construction of a large insert clone library in a fosmid vector, and screening of the clones by pmoA-targeted PCR. For comparison, a genomic library of M. acidiphila B2 was constructed in a bacterial artificial chromosome (BAC) vector, and genomic fragments carrying pmoA were identified. The genomic information obtained for USCα and M. acidiphila B2 should be used to clarify the phylogenetic status of USCα and to perform a detailed comparative analysis of the pmo operon.
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Methylocella tundrae sp nov a novel methanotrophic bacterium from acidic tundra peatlands
International Journal of Systematic and Evolutionary Microbiology, 2004Co-Authors: Svetlana N. Dedysh, Werner Liesack, Yu Yu Berestovskaya, Svetlana E. Belova, Valentina N Khmelenina, Yuri A Trotsenko, Lina V Vasylieva, Natalia E Suzina, G A ZavarzinAbstract:A novel species, Methylocella tundrae, is proposed for three methanotrophic strains (T4T, TCh1 and TY1) isolated from acidic Sphagnum tundra peatlands. These strains are aerobic, Gram-negative, non-motile, dinitrogen-fixing rods that possess a soluble methane monooxygenase and utilize the serine pathway for carbon assimilation. Strains T4T, TCh1 and TY1 are moderately acidophilic organisms capable of growth between pH 4·2 and 7·5 (optimum 5·5–6·0) and between 5 and 30 °C (optimum 15 °C). The major phospholipid fatty acid is 18 : 1ω7c. The DNA G+C content of strain T4T is 63·3 mol%. The three strains possess almost identical 16S rRNA gene sequences and are most closely related to two previously identified species of Methylocella, Methylocella palustris (97 % similarity) and Methylocella silvestris (97·5 % similarity). DNA–DNA hybridization values of strain T4T with Methylocella palustris KT and Methylocella silvestris BL2T were respectively 27 and 36 %. Thus, the tundra strains represent a novel species, for which the name Methylocella tundrae sp. nov. is proposed. Strain T4T (=DSM 15673T=NCIMB 13949T) is the type strain.
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Comparison of One- and Two-dimensional Liquid Chromatography Approaches in the Label-free Quantitative Analysis of Methylocella silvestris
2016Co-Authors: Nisha A. Patel, Andrew Crombie, Konstantinos Thalassinos, Susan E. Slade, Joanne B. Connolly, Chris Hughes, James Langridge, James H. ScrivensAbstract:The proteome of the bacterium Methylocella silvestris has been characterized using reversed phase ultra high pressure liquid chromatography (UPLC) and two-dimensional reversed phase (high pH)–reversed phase (low pH) UPLC prior to mass spectrometric analysis. Variations in protein expression levels were identified with the aid of label-free quantification in a study of soluble protein extracts from the organism grown using methane, succinate, or propane as a substrate. The number of first dimensional fractionation steps has been varied for 2D analyses, and the impact on data throughput and quality has been demonstrated. Comparisons have been made regarding required experimental considerations including total loading of biological samples required, instrument time, and resulting data file sizes. The data obtained have been evaluated with respect to number of protein identifications, confidence of assignments, sequence coverage, relative levels of proteins, and dynamic range. Good qualitative and quantitative agreement was observed between the different approaches, and the potential benefits and limitations of the reversed phase–reversed phase UPLC technique in label-free analysis are discussed. A preliminary screen of the protein regulation data has also been performed, providing evidence for a possible propane assimilation route
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Comparison of one- and two-dimensional liquid chromatography approaches in the label-free quantitative analysis of Methylocella silvestris.
Journal of proteome research, 2012Co-Authors: Nisha A. Patel, Andrew T Crombie, Konstantinos Thalassinos, Susan E. Slade, Joanne B. Connolly, Chris Hughes, James I. Langridge, James H. ScrivensAbstract:The proteome of the bacterium Methylocella silvestris has been characterized using reversed phase ultra high pressure liquid chromatography (UPLC) and two-dimensional reversed phase (high pH)–reversed phase (low pH) UPLC prior to mass spectrometric analysis. Variations in protein expression levels were identified with the aid of label-free quantification in a study of soluble protein extracts from the organism grown using methane, succinate, or propane as a substrate. The number of first dimensional fractionation steps has been varied for 2D analyses, and the impact on data throughput and quality has been demonstrated. Comparisons have been made regarding required experimental considerations including total loading of biological samples required, instrument time, and resulting data file sizes. The data obtained have been evaluated with respect to number of protein identifications, confidence of assignments, sequence coverage, relative levels of proteins, and dynamic range. Good qualitative and quantitativ...
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Bacterial flavin-containing monooxygenase is trimethylamine monooxygenase.
Proceedings of the National Academy of Sciences of the United States of America, 2011Co-Authors: Nisha A. Patel, Andrew T Crombie, James H. ScrivensAbstract:Flavin-containing monooxygenases (FMOs) are one of the most important monooxygenase systems in Eukaryotes and have many important physiological functions. FMOs have also been found in bacteria; however, their physiological function is not known. Here, we report the identification and characterization of trimethylamine (TMA) monooxygenase, termed Tmm, from Methylocella silvestris, using a combination of proteomic, biochemical, and genetic approaches. This bacterial FMO contains the FMO sequence motif (FXGXXXHXXXF/Y) and typical flavin adenine dinucleotide and nicotinamide adenine dinucleotide phosphate-binding domains. The enzyme was highly expressed in TMA-grown M. silvestris and absent during growth on methanol. The gene, tmm, was expressed in Escherichia coli, and the purified recombinant protein had high Tmm activity. Mutagenesis of this gene abolished the ability of M. silvestris to grow on TMA as a sole carbon and energy source. Close homologs of tmm occur in many Alphaproteobacteria, in particular Rhodobacteraceae (marine Roseobacter clade, MRC) and the marine SAR11 clade (Pelagibacter ubique). We show that the ability of MRC to use TMA as a sole carbon and/or nitrogen source is directly linked to the presence of tmm in the genomes, and purified Tmm of MRC and SAR11 from recombinant E. coli showed Tmm activities. The tmm gene is highly abundant in the metagenomes of the Global Ocean Sampling expedition, and we estimate that 20% of the bacteria in the surface ocean contain tmm. Taken together, our results suggest that Tmm, a bacterial FMO, plays an important yet overlooked role in the global carbon and nitrogen cycles.
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A comparison of labeling and label-free mass spectrometry-based proteomics approaches.
Journal of proteome research, 2009Co-Authors: Vibhuti J. Patel, Andrew T Crombie, Konstantinos Thalassinos, Susan E. Slade, Joanne B. Connolly, James H. ScrivensAbstract:The proteome of the recently discovered bacterium Methylocella silvestris has been characterized using three profiling and comparative proteomics approaches. The organism has been grown on two different substrates enabling variations in protein expression to be identified. The results obtained using the experimental approaches have been compared with respect to number of proteins identified, confidence in identification, sequence coverage and agreement of regulated proteins. The sample preparation, instrumental time and sample loading requirements of the differing experiments are compared and discussed. A preliminary screen of the protein regulation results for biological significance has also been performed.