Widespread soil bacterium that oxidizes atmospheric methane.

The global atmospheric level of methane (CH4), the second most important greenhouse gas, is currently increasing by ∼10 million tons per year. Microbial oxidation in unsaturated soils is the only known biological process that removes CH4 from the atmosphere, but so far, bacteria that can grow on atmospheric CH4 have eluded all cultivation efforts. In this study, we have isolated a pure culture of a bacterium, strain MG08 that grows on air at atmospheric concentrations of CH4 [1.86 parts per million volume (p.p.m.v.)]. This organism, named Methylocapsa gorgona, is globally distributed in soils and closely related to uncultured members of the upland soil cluster α. CH4 oxidation experiments and 13C-single cell isotope analyses demonstrated that it oxidizes atmospheric CH4 aerobically and assimilates carbon from both CH4 and CO2 Its estimated specific affinity for CH4 (a0s) is the highest for any cultivated methanotroph. However, growth on ambient air was also confirmed for Methylocapsa acidiphila and Methylocapsa aurea, close relatives with a lower specific affinity for CH4, suggesting that the ability to utilize atmospheric CH4 for growth is more widespread than previously believed. The closed genome of M. gorgona MG08 encodes a single particulate methane monooxygenase, the serine cycle for assimilation of carbon from CH4 and CO2, and CO2 fixation via the recently postulated reductive glycine pathway. It also fixes dinitrogen and expresses the genes for a high-affinity hydrogenase and carbon monoxide dehydrogenase, suggesting that atmospheric CH4 oxidizers harvest additional energy from oxidation of the atmospheric trace gases carbon monoxide (0.2 p.p.m.v.) and hydrogen (0.5 p.p.m.v.).

Facultative methanotrophs are abundant at terrestrial natural gas seeps.

BACKGROUND: Natural gas contains methane and the gaseous alkanes ethane, propane and butane, which collectively influence atmospheric chemistry and cause global warming. Methane-oxidising bacteria, methanotrophs, are crucial in mitigating emissions of methane as they oxidise most of the methane produced in soils and the subsurface before it reaches the atmosphere. Methanotrophs are usually obligate, i.e. grow only on methane and not on longer chain alkanes. Bacteria that grow on the other gaseous alkanes in natural gas such as propane have also been characterised, but they do not grow on methane. Recently, it was shown that the facultative methanotroph Methylocella silvestris grew on ethane and propane, other components of natural gas, in addition to methane. Therefore, we hypothesised that Methylocella may be prevalent at natural gas seeps and might play a major role in consuming all components of this potent greenhouse gas mixture before it is released to the atmosphere. RESULTS: Environments known to be exposed to biogenic methane emissions or thermogenic natural gas seeps were surveyed for methanotrophs. 16S rRNA gene amplicon sequencing revealed that Methylocella were the most abundant methanotrophs in natural gas seep environments. New Methylocella-specific molecular tools targeting mmoX (encoding the soluble methane monooxygenase) by PCR and Illumina amplicon sequencing were designed and used to investigate various sites. Functional gene-based assays confirmed that Methylocella were present in all of the natural gas seep sites tested here. This might be due to its ability to use methane and other short chain alkane components of natural gas. We also observed the abundance of Methylocella in other environments exposed to biogenic methane, suggesting that Methylocella has been overlooked in the past as previous ecological studies of methanotrophs often used pmoA (encoding the alpha subunit of particulate methane monooxygenase) as a marker gene. CONCLUSION: New biomolecular tools designed in this study have expanded our ability to detect, and our knowledge of the environmental distribution of Methylocella, a unique facultative methanotroph. This study has revealed that Methylocella are particularly abundant at natural gas seeps and may play a significant role in biogeochemical cycling of gaseous hydrocarbons.

Qingshengfania soli Zhang et al. 2015 is a later heterotypic synonym of Pseudochelatococcus lubricantis Kämpfer et al. 2015.

Qingshengfania soli DSM 103870T was compared with Pseudochelatococcus lubricantis MPA 1113T to clarify the taxonomic relationship of both species because of their high phylogenetic relationship. 16S rRNA gene sequence comparisons demonstrated that these species share 100  % sequence similarity. Investigation of fatty acid patterns, substrate utilization, and matrix-assisted laser desorption/ionization-time-of-flight (MALDI-TOF) profiles displayed no striking differences between the type strains of both species. DNA-DNA hybridization between both strains showed a 95 % (reciprocal 82 %) similarity, which clearly demonstrated that both strains are members of the same species. Due to priority of publication and validation of the name, Qingshengfania soli is reclassified as Pseudochelatococcus lubricantis, based on the estimated phylogenetic position derived from 16S rRNA gene sequence data, fatty acid, biochemical data, MALDI-TOF, and DNA-DNA hybridization results.

Chelatococcus composti sp. nov., isolated from penicillin fermentation fungi residue with pig manure co-compost.

A novel Gram-stain-negative bacterium, designated strain PC-2T, was isolated from penicillin fermentation fungi residue with pig manure co-compost in China. Phylogenetic analysis, based on 16S rRNA gene sequence comparisons, revealed that strain PC-2T should be assigned to the genus Chelatococcus and that it had 98.9 % similarity with Chelatococcus daeguensis, 98.8 % with Chelatococcus sambhunathii, 98.4 %, with Chelatococcus caeni and 96.0 % with Chelatococcus asaccharovorans. The G+C content of genomic DNA was 70.9 mol%. On the basis of the phylogenetic analysis, DNA-DNA relatedness values, phenotypic characteristics and chemotaxonomic data, strain PC-2 T represents a novel species of the genus Chelatococcus, for which the name Chelatococcus composti sp. nov. is proposed. The type strain is PC-2T (=DSM 101465T=CGMCC 1.15283T).

Chelatococcus reniformis sp. nov., isolated from a glacier.

A Gram-stain-negative, non-motile, reniform bacterial strain, B2974T, was isolated from an ice core of the Muztagh Glacier, on the Tibetan Plateau, China. Strain B2974T grew optimally at pH 7.0-7.5 and 25-30 °C in the presence of 0-2.0 % (w/v) NaCl. 16S rRNA gene sequence similarity analysis indicated that strain B2974T was closely related to Chelatococcus asaccharovorans LMG 25503T at a level of 97.1 %. The major quinone of strain B2974T was ubiquinone Q10. The predominant fatty acids were summed feature 8 (C18 : 1ω7c and/or C18 : 1ω6c) and C19 : 0 cyclo ω8c. sym-Homospermidine was the major polyamine. The genomic DNA G+C content of the strain was 64 mol%. In DNA-DNA hybridization tests, strain B2974T shared 49.32 % DNA-DNA relatedness with the type strain of Chelatococcus asaccharovorans LMG 25503T. Based on the results of phenotypic and chemotaxonomic characteristics, strain B2974T was considered as a novel species of the genus Chelatococcus, for which the name Chelatococcus reniformis sp. nov. is proposed. The type strain is B2974T (=JCM 30308T=CGMCC 1.12919T).

Emended description of the family Beijerinckiaceae and transfer of the genera Chelatococcus and Camelimonas to the family Chelatococcaceae fam. nov.

The family Beijerinckiaceae was circumscribed in 2005 to accommodate four genera of phylogenetically related alphaproteobacteria: Beijerinckia, Chelatococcus, Methylocella and Methylocapsa. Later, four additional genera, i.e. Methylovirgula, Methyloferula, Methylorosula and Camelimonas, were described and assigned to this family, which now accommodates 21 species with validly published names. Members of this family possess strikingly different lifestyles, including chemoheterotrophy, facultative methylotrophy, obligate methanotrophy and facultative methanotrophy. Levels of 16S rRNA gene sequence similarity among most of these bacteria range from 96 to 98 %, suggesting a common evolutionary origin. The genera Chelatococcus and Camelimonas, however, are not monophyletic with the other described genera based on 16S rRNA gene sequence phylogeny, and instead form a distant cluster more closely related to the Methylobacteriaceae. Physiologically these two genera also lack several properties common to the other Beijerinckiaceae. On the other hand, the genus Rhodoblastus, presently considered a member of the Bradyrhizobiaceae, affiliates with high confidence to the Beijerinckiaceae. Here, we propose to transfer the genera Chelatococcus and Camelimonas to the family Chelatococcaceae fam. nov., and present an emended description of the family Beijerinckiaceae, including the genus Rhodoblastus.

Camelimonas fluminis sp. nov., a cyhalothrin-degrading bacterium isolated from river water.

An aerobic, Gram-stain-negative, short rod-shaped, non-spore-forming, cyhalothrin-degrading bacterial strain, XZ2(T), was isolated from the surface water of Hanjiang River in Wuhan, China. Strain XZ2(T) grew optimally at pH 6.0, 30 °C and in the absence of NaCl. The G+C content of the total DNA was 64.1 mol%. The 16S rRNA gene sequence of strain XZ2(T) showed the highest similarity to that of Camelimonas lactis M 2040(T) (99.1%), followed by Camelimonas abortus UK34/07-5(T) (95.9%) and Chelatococcus daeguensis K106(T) (95.3%). The major cellular fatty acids of strain XZ2(T) were C19 : 0 cyclo ω8c (63.1%), C16 : 0 (15.0%) and C18 : 1ω7c/C18 : 1ω6c (summed feature 8; 8.9%). C18 : 0 3-OH was also detected as the major hydroxylated fatty acid. The respiratory quinone was ubiquinone Q-10. The polar lipid profile included the major compounds phosphatidylcholine and diphosphatidylglycerol, and moderate amounts of phosphatidylethanolamine, phosphatidylglycerol and two unidentified aminolipids. The predominant compound in the polyamine pattern was spermidine. These chemotaxonomic data supported the affiliation of strain XZ2(T) to the genus Camelimonas. The DNA-DNA hybridization value between strain XZ2(T) and Camelimonas lactis M 2040(T) was 43.5 ± 0.6%. DNA-DNA hybridization data as well as biochemical and physiological characteristics strongly supported the genotypic and phenotypic differentiations between strain XZ2(T) and Camelimonas lactis M 2040(T). Therefore, strain XZ2(T) represents a novel species of the genus Camelimonas, for which the name Camelimonas fluminis sp. nov. is proposed. The type strain is XZ2(T) ( = KCTC 42282(T) = ACCC 19738(T)).

Methylocapsa palsarum sp. nov., a methanotroph isolated from a subArctic discontinuous permafrost ecosystem.

An aerobic methanotrophic bacterium was isolated from a collapsed palsa soil in northern Norway and designated strain NE2T. Cells of this strain were Gram-stain-negative, non-motile, non-pigmented, slightly curved thick rods that multiplied by normal cell division. The cells possessed a particulate methane monooxygenase enzyme (pMMO) and utilized methane and methanol. Strain NE2T grew in a wide pH range of 4.1­8.0 (optimum pH 5.2­6.5) at temperatures between 6 and 32 °C (optimum 18­25 °C), and was capable of atmospheric nitrogen fixation under reduced oxygen tension. The major cellular fatty acids were C18 : 1ω7c, C16 : 0 and C16 : 1ω7c, and the DNA G+C content was 61.7 mol%. The isolate belonged to the family Beijerinckiaceae of the class Alphaproteobacteria and was most closely related to the facultative methanotroph Methylocapsa aurea KYGT (98.3 % 16S rRNA gene sequence similarity and 84 % PmoA sequence identity). However, strain NE2T differed from Methylocapsa aurea KYGT by cell morphology, the absence of pigmentation, inability to grow on acetate, broader pH growth range, and higher tolerance to NaCl. Therefore, strain NE2T represents a novel species of the genus Methylocapsa, for which we propose the name Methylocapsa palsarum sp. nov. The type strain is NE2T ( = LMG 28715T = VKM B-2945T).

Pseudochelatococcus lubricantis gen. nov., sp. nov. and Pseudochelatococcus contaminans sp. nov. from coolant lubricants.

Two Gram-negative, rod-shaped, non-spore-forming bacteria, isolated from metal working fluids were investigated to determine their taxonomic positions. On the basis of 16S rRNA gene sequence phylogeny, both strains (MPA 1113(T) and MPA 1105(T)) formed a distinct cluster with 97.7 % sequence similarity between them, which was in the vicinity of members of the genera Methylobacterium, Camelimonas, Chelatococcus, Bosea, Salinarimonas and Microvirga to which they showed low sequence similarities (below 94 %). The predominant compounds in the polyamine pattern and in the quinone system of the two strains were spermidine and ubiquinone Q-10, respectively. The polar lipid profiles were composed of the major compounds: phosphatidylmonomethylethanolamine, phosphatidylglycerol, phosphatidylcholine, major or moderate amounts of diphosphatidylglycerol, two unidentified glycolipids and three unidentified aminolipids. Several minor lipids were also detected. The major fatty acids were either C19 : 0 cyclo ω8c or C18 : 1ω7c. The results of fatty acid analysis and physiological and biochemical tests allowed both, the genotypic and phenotypic differentiation of the isolates from each other, while the chemotaxonomic traits allowed them to be differentiated from the most closely related genera. In summary, low 16S rRNA gene sequence similarities and marked differences in polar lipid profiles, as well as in polyamine patterns, is suggestive of a novel genus for which the name Pseudochelatococcus gen. nov. is proposed. MPA 1113(T) ( = CCM 8528(T) = LMG 28286(T) = CIP 110802(T)) and MPA 1105(T) ( = CCM 8527(T) = LMG 28285(T)) are proposed to be the type strains representing two novel species within the novel genus, Pseudochelatococcus gen. nov., for which the names Pseudochelatococcus lubricantis sp. nov. and Pseudochelatococcus contaminans sp. nov. are suggested, respectively.

Chelatococcus caeni sp. nov., isolated from a biofilm reactor sludge sample.

A polyphasic taxonomic study was carried out on strain EBR-4-1(T), which was isolated from a biofilm reactor in the Republic of Korea. The cells of the strain were Gram-stain-negative, non-spore-forming, motile and rod-shaped. Comparative 16S rRNA gene sequence studies showed a clear affiliation of this strain to the Alphaproteobacteria, and it was most closely related to Chelatococcus daeguensis CCUG 54519(T), Chelatococcus sambhunathii HT4(T), and Chelatococcus asaccharovorans DSM 6462(T) with 16S rRNA gene sequence similarities to the type strains of these species of 98.8 %, 98.7 %, and 96.3 %, respectively. The G+C content of the genomic DNA of strain EBR-4-1(T) was 68.7 mol%. Phenotypic and chemotaxonomic data [Q-10 as the major ubiquinone; C19 : 0cycloω8c, C18 : 1 2-OH, and summed feature 8 (C18 : 1ω7c and/or C18 : 1ω6c) as the major fatty acids] supported the affiliation of strain EBR-4-1(T) to the genus Chelatococcus. On the basis of the polyphasic evidence, it is proposed that strain EBR-4-1(T) should be assigned to a new species, Chelatococcus caeni sp. nov. The type strain is EBR-4-1(T) ( = KCTC 32487(T) = JCM 30181(T)).

The (d)evolution of methanotrophy in the Beijerinckiaceae--a comparative genomics analysis.

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.

Removal of nitric oxide in a biotrickling filter under thermophilic condition using Chelatococcus daeguensis.

UNLABELLED: The development of a thermophilic biotrickling filter (BTF) system to inoculate a newly isolated strain of Chelatococcus daeguensis TAD1 for the effective treatment of nitric oxide (NO) is described. A bench-scale BTF was run under high concentrations of NO and 8% O2 in thermophilic aerobic environment. A novel aerobic denitrifier Chelatococcus daeguensis TAD1 was isolated from the biofilm of an on-site biotrickling filter and it showed a denitrifying capability of 96.1% nitrate removal rate in a 24 h period in aerobic environment at 50 degrees C, with no nitrite accumulation. The inlet NO concentration fluctuated between approximately 133.9 and 669.6 mg m-3 and kept on a steady NOx removal rate above 80% in an oxygen stream of 8%. The BTF system was able to consistently remove 80-93.7% NO when the inlet NO was 535.7 mg m-3 in an oxygen stream of 2-20%. The biological removal efficiency of NO at 50 degrees C is higher than that at 25 degrees C, suggesting that the aerobic denitrifier TAD1 display well denitrification performance under thermophilic condition. Starvation for 2, 4 and 8 days resulted in the re-acclimation times of Chelatococcus daeguensis TAD1 ranging between 4 and 16 hours. A longer recovery time than that for weekend shutdown will be required when a longer starvation occurs. The results presented here demonstrate the feasibility of biotrickling filter for the thermophilic removal of NOx from gas streams. IMPLICATIONS: A novel denitrifier Chelatococcus daeguensis TAD1 was isolated from an on-site biotrickling filter in aerobic environment at 50 degrees C. To date, C. daeguensis has not been previously reported to be an aerobic denitrifier. In this study, a thermophilic biotrickling filter system inoculated with Chelatococcus daeguensis TADI for treatment of nitric oxide is developed. In coal-fired power plants, influent flue gas stream for nitrogen oxides (NOx) removal typically exhibit temperatures between 50 and 60 degrees C. Traditionally, cooling gases to below 40 degrees C prior to biological treatment is inevitable, which is costly. Therefore, the application ofthermophilic microorganisms for the removal of nitric oxide (NO) at this temperature range would offer great savings and would greatly extend the applicability ofbiofilters and biotrickling filters. Until now there has not been any study published about thermophilic biological treatment of NO under aerobic condition.

Methylorosula polaris gen. nov., sp. nov., an aerobic, facultatively methylotrophic psychrotolerant bacterium from tundra wetland soil.

Three strains of Gram-negative, aerobic, motile bacteria with bipolar flagella were isolated from acidic tundra wetland soils near the city of Vorkuta and from the Chukotka and Yugorsky Peninsulas and designated strains V-022(T), Ch-022 and Ju-022. The cells were rod-shaped, 0.5-0.6 µm in width and 1.3-4.5 µm in length and reproduced by irregular fission. These bacteria were facultative methylotrophs that used methanol, methylamines and a wide range of other sources of carbon and energy such as sugars and polysaccharides, ethanol and amino acids. The isolates used the Calvin-Benson pathway for the assimilation of one-carbon compounds and were unable to fix nitrogen. The new strains were moderately acidophilic and psychrotolerant, capable of growth over a pH range of 4.0 to 7.8, with optimum growth at pH 5.5-6.0. Growth occurred between 4 and 30 °C (optimum 20-25 °C). The principal phospholipid fatty acid was C(18:1)ω7c. The DNA G+C content of strain V-022(T) was 65.2 mol%. Analysis of the 16S rRNA gene sequences revealed that all three isolates V-022(T), Ch-022 and Yu-022 exhibited almost identical 16S rRNA gene sequences (99.9% gene sequence similarity) and formed a new lineage within the class Alphaproteobacteria. The name Methylorosula polaris is suggested to accommodate this new genus and novel species with strain V-022(T) (=DSM 22001(T)=VKM V-2485(T)) as the type strain of the type species.

Camelimonas abortus sp. nov., isolated from placental tissue of cattle.

A gram-negative, rod-shaped, non-spore-forming bacterium, isolated from placental tissue of a cow, was investigated for its taxonomic position. On the basis of 16S rRNA gene sequence similarities, strain UK34/07-5(T) was shown to belong to the class Alphaproteobacteria, closely related to the type strain of Camelimonas lactis (96.0 % sequence similarity). The polyamine pattern showed the major compound spermidine and moderate amounts of putrescine. The major quinone was ubiquinone Q-10. The polar lipid profile was composed of the major compounds phosphatidylethanolamine, phosphatidylcholine, phosphatidylglycerol and phosphatidylmonomethylethanolamine and moderate amounts of diphosphatidylglycerol, three unidentified aminolipids and an unidentified phospholipid. The profile of major fatty acids, consisting of C(19 : 0) cyclo ω8c and C(18 : 1)ω7c, with C(18 : 0) 3-OH as the hydroxylated fatty acid, was very similar to that of C. lactis M 2040(T). The results of DNA-DNA hybridization and physiological and biochemical tests allowed both genotypic and phenotypic differentiation of the isolate from C. lactis. The relatively low 16S rRNA gene sequence similarity of 96.0 % to C. lactis M 2040(T) and marked differences in the polar lipid profiles as well as the results of physiological tests and the DNA-DNA hybridization data support the creation of a novel species, for which the name Camelimonas abortus sp. nov. is proposed, with the type strain UK34/07-5(T) ( = CIP 110303(T)  = CCUG 61094(T)  = DSM 24741(T)  = CCM 7941(T)).

Evaluation of the coal-degrading ability of Rhizobium and Chelatococcus strains isolated from the formation water of an Indian coal bed.

The rise in global energy demand has prompted researches on developing strategies for transforming coal into a cleaner fuel. This requires isolation of microbes with the capability to degrade complex coal into simpler substrates to support methanogenesis in the coal beds. In this study, aerobic bacteria were isolated from an Indian coal bed that can solubilize and utilize coal as the sole source of carbon. The six bacterial isolates capable of growing on coal agar medium were identified on the basis of their 16S rRNA gene sequences, which clustered into two groups; Group I isolates belonged to the genus Rhizobium, whereas Group II isolates were identified as Chelatococcus species. Out of the 4 methods of whole genome fingerprinting (ERIC-PCR, REP-PCR, BOX-PCR, and RAPD), REPPCR showed maximum differentiation among strains within each group. Only Chelatococcus strains showed the ability to solubilize and utilize coal as the sole source of carbon. On the basis of 16S rRNA gene sequence and the ability to utilize different carbon sources, the Chelatococcus strains showed maximum similarity to C. daeguensis. This is the first report showing occurrence of Rhizobium and Chelatococcus strains in an Indian coal bed, and the ability of Chelatococcus isolates to solubilize and utilize coal as a sole source of carbon for their growth.

The status of the species Beijerinckia fluminensis Dobereiner and Ruschel 1958. Request for an Opinion.

In a previous article [Oggerin M., Arahal, D. R., Rubio, V. & Marin, I. (2009). Int J Syst Evol Microbiol 59, 2323-2328], it has been shown that strain Beijerinckia fluminensis UQM 1685(T) and its derived equivalent B. fluminensis CIP 106281(T) do not conform to the description of the type strain of Beijerinckia fluminensis Döbereiner and Ruschel 1958. Indeed, both strains were identified as members of the species Rhizobium radiobacter and exhibited marked phenotypic and genotypic differences with members of the genus Beijerinckia. It was concluded that both strains, and any other equivalents derived from them, do not descend from the nomenclatural type. Since then, our attempts to find older deposits of the type strain, hopefully derived from the original isolate, or other existing strains of Beijerinckia fluminensis that could be proposed as a neotype strain, have been in vain. It is therefore proposed that the Judicial Commission should place the name Beijerinckia fluminensis Döbereiner and Ruschel 1958 on the list of rejected names if a suitable replacement type strain or a neotype cannot be found within two years following the publication of this Request (Rule 18c).

Methyloferula stellata gen. nov., sp. nov., an acidophilic, obligately methanotrophic bacterium that possesses only a soluble methane monooxygenase.

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.

Complete genome sequence of Beijerinckia indica subsp. indica.

Beijerinckia indica subsp. indica is an aerobic, acidophilic, exopolysaccharide-producing, N(2)-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.

Isolation and characterization of new poly(3HB)-accumulating star-shaped cell-aggregates-forming thermophilic bacteria.

AIMS: This study aimed at isolating thermophilic bacteria that utilize cheap carbon substrates for the economically feasible production of poly(3-hydroxybutyrate), poly(3HB), at elevated temperatures. METHODS AND RESULTS: Thermophilic bacteria were enriched from an aerobic organic waste treatment plant in Germany, and from hot springs in Egypt. Using the viable colony staining method for hydrophobic cellular inclusions with Nile red in mineral salts medium (MSM) containing different carbon sources, six Gram-negative bacteria were isolated. Under the cultivation conditions used in this study, strains MW9, MW11, MW12, MW13 and MW14 formed stable star-shaped cell-aggregates (SSCAs) during growth; only strain MW10 consisted of free-living rod-shaped cells. The phylogenetic relationships of the strains as derived from 16S rRNA gene sequence comparisons revealed them as members of the Alphaproteobacteria. The 16S rRNA gene sequences of the isolates were very similar (>99% similarity) and exhibited similarities ranging from 93 to 99% with the most closely related species that were Chelatococcus daeguensis, Chelatococcus sambhunathii,Chelatococcus asaccharovorans, Bosea minatitlanensis, Bosea thiooxidans and Methylobacterium lusitanum. Strains MW9, MW10, MW13 and MW14 grew optimally in MSM with glucose, whereas strains MW11 and MW12 preferred glycerol as sole carbon source for growth and poly(3HB) accumulation. The highest cell density and highest poly(3HB) content attained were 4·8g l(-l) (cell dry weight) and 73% (w/w), respectively. Cells of all strains grew at temperatures between 37 and 55°C with the optimum growth at 50°C. CONCLUSIONS: New PHA-accumulating thermophilic bacterial strains were isolated and characterized to produce poly(3HB) from glucose or glycerol in MSM at 50°C. SSCAs formation was reported during growth. SIGNIFICANCE AND IMPACT OF THE STUDY: To the best of our knowledge, this is the first report on the formation of SSCAs by PHA-accumulating bacteria and also by thermophilic bacteria. PHA-producing thermophiles can significantly reduce the costs of fermentative PHA production.

Methylocapsa aurea sp. nov., a facultative methanotroph possessing a particulate methane monooxygenase, and emended description of the genus Methylocapsa.

An aerobic, methanotrophic bacterium, designated KYG(T), was isolated from a forest soil in Germany. Cells of strain KYG(T) were Gram-negative, non-motile, slightly curved rods that multiplied by binary fission and produced yellow colonies. The cells contained intracellular granules of poly-ß-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 KYG(T) 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 C(18 : 1)ω7c (81.5 %) and the DNA G+C content was 61.4 mol%. Strain KYG(T) belonged to the family Beijerinckiaceae of the class Alphaproteobacteria and was most closely related to the obligate methanotroph Methylocapsa acidiphila B2(T) (98.1 % 16S rRNA gene sequence similarity and 84.7 % pmoA sequence similarity). Unlike Methylocapsa acidiphila B2(T), which grows only on methane and methanol, strain KYG(T) 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 KYG(T) differed from Methylocapsa acidiphila B2(T) 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 KYG(T) (=DSM 22158(T) =VKM B-2544(T)).