Methylobacterium flocculans sp. nov., a Floc-Forming Bacterium Isolated from Aquaculture Ponds.

Strain FF17T, a Gram-negative, obligate aerobic, motile, pink-pigmented, and methylotrophic bacterium, was selected for a polyphasic taxonomic investigation due to its capacity for aggregation, or floc formation. The predominant respiratory quinone observed was Q-10, accounting for 83.36% of the total, while the major fatty acids were summed feature 8 (18:1 w6c and/or 18:1 w7c). The major polar lipids included Diphosphatidylglycerol (DPG), phosphatidylglycerol, phosphatidylethanolamine (PE), phosphatidylinositol (PI), and one unknown polar lipid. Phylogenetic analysis showed that strain FF17T was hithermost related to Methylobacterium goesingense iEII3T (99.86%), M. gossipiicola Gh-105 T (99.22%), M. adhaesivum AR27T (98.92%), and M. iners 5317S-33 T (97.27%) based on 16S rRNA gene sequence similarity. A 5,735,273-bp chromosome and six plasmids make up the genome, making it larger than the genomes of the other four Methylobacterium species described above. The digital DNA-DNA hybridization and average nucleotide identity values between strain FF17T and the reference strains were 21.90-28.70 and 77.39-85.04%, respectively. Strain FF17T had a genome DNA G + C content of 68.5 mol%. The analysis of genomes indicated that cellulose apparently plays an important character in the aggregation of Methylobacterium species. Genome annotation revealed the presence of genes involved in assimilatory/dissimilatory nitrate reduction and ammonia assimilation. In conclusion, Strain FF17T is identified as a new species in the Methylobacterium genus, based on analyses of genomics, phylogeny, biochemistry, and fatty acids, and the name Methylobacterium flocculans sp. nov. is proposed. The type strain is FF17T (= MCCC 1K08738T = KCTC 8320 T).

Methylobacterium amylolyticum sp. nov. and Methylobacterium ligniniphilum sp. nov., isolated from soil.

Two pink-pigmented bacteria, designated strains NEAU-140T and NEAU-KT, were isolated from field soil collected from Linyi, Shandong Province, PR China. Both isolates were aerobic, Gram-stain-negative, rod-shaped, and facultatively methylotrophic. 16S rRNA gene sequences analysis showed that these two strains belong to the genus Methylobacterium. Strain NEAU-140T exhibited high 16S rRNA gene sequence similarities to Methylobacterium radiotolerans NBRC 15690T (97.43 %) and Methylobacterium phyllostachyos NBRC 105206T (97.36 %). Strain NEAU-KT exhibited high 16S rRNA gene sequence similarities to M. phyllostachyos NBRC 105206T (99.00 %) and Methylobacterium longum DSM 23933T (98.72 %). A phylogenetic tree based on 16S rRNA gene sequences showed that strain NEAU-140T formed a clade with Methylobacterium aerolatum (95.94 %), Methylobacterium persicinum (95.66 %) and Methylobacterium komagatae (96.87 %), and strain NEAU-KT formed a cluster with M. phyllostachyos and M. longum. The predominant fatty acid in both strains was C18 : 1 ω7c. Both strains contained ubiquinone Q-10 as the only respiratory quinone. The polar lipid profiles of both strains contained diphosphatidylglycerol, phosphatidylethanolamine, and phosphatidylcholine. Whole-genome phylogeny showed that strains NEAU-140T and NEAU-KT formed a phyletic line with M. aerolatum, M. persicinum, Methylobacterium radiotolerans, Methylobacterium fujisawaense, Methylobacterium oryzae, Methylobacterium tardum, M. longum and M. phyllostachyos. The orthologous average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values between strain NEAU-140T and its closely related strains were lower than 82.62 and 25.90  %, respectively. The ANI and dDDH values between strain NEAU-KT and its closely related strains were lower than 86.29 and 31.7 %, respectively. The genomic DNA G+C contents were 71.63 mol% for strain NEAU-140T and 69.08 mol% for strain NEAU-KT. On the basis of their phenotypic and phylogenetic distinctiveness and the results of dDDH and ANI hybridization, these two isolates represent two novel species within the genus Methylobacterium, for which the names Methylobacterium amylolyticum sp. nov. (type strain NEAU-140T=MCCC 1K08801T=DSM 110568T) and Methylobacterium ligniniphilum sp. nov. (type strain NEAU-KT=MCCC 1K08800T=DSM 110567T) are proposed.

Methylobacterium nigriterrae sp. nov., isolated from black soil.

An aerobic, Gram-stain-negative, motile rod bacterium, designated as SYSU BS000021T, was isolated from a black soil sample in Harbin, Heilongjiang province, China. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the isolate belongs to the genus Methylobacterium, and showed the highest sequence similarity to Methylobacterium segetis KCTC 62267 T (98.51%) and Methylobacterium oxalidis DSM 24028 T (97.79%). Growth occurred at 20-37℃ (optimum, 28 °C), pH 6.0-8.0 (optimum, pH 7.0) and in the presence of 0% (w/v) NaCl. Polar lipids comprised of phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, one unidentified aminolipid and one unidentified polar lipid. The major cellular fatty acids (> 5%) were C18:0 and C18:1 ω7c and/or C18:1 ω6c. The predominant respiratory quinone was Q-10. The genomic G + C content was 68.36% based on the whole genome analysis. The average nucleotide identity (≤ 83.5%) and digital DNA-DNA hybridization (≤ 27.3%) values between strain SYSU BS000021T and other members of the genus Methylobacterium were all lower than the threshold values recommended for distinguishing novel prokaryotic species. Based on the results of phenotypic, chemotaxonomic and phylogenetic analyses, strain SYSU BS000021T represents a novel species of the genus Methylobacterium, for which the name Methylobacterium nigriterrae sp. nov. is proposed. The type strain of the proposed novel species is SYSU BS000021T (= GDMCC 1.3814 T = KCTC 8051 T).

Methylobacterium radiodurans sp. nov., a novel radiation-resistant Methylobacterium.

A Gram-negative, aerobic, flagellated, rod-shaped, and pink-pigmented bacterium, strain 17Sr1-43 T, was isolated from a soil sample collected in Nowongu, Seoul, Korea. The isolate could grow at 18-37 °C (optimum, 28-30 °C), pH 6.0-8.0 (optimum, pH 7.0) and in the presence of 0-1.0% (w/v) NaCl (optimum, 0%) with aeration. The major cellular fatty acids were summed feature 8 (C18:1 ω7c and/or C18:1 ω6c) and summed feature 2 (iso-C16:1 I and/or C14:0 3-OH). The predominant respiratory quinone was Q-10 and the major polar lipids were phosphatidylethanolamine, phosphatidylglycerol, phospholipid, and diphosphatidylglycerol. The G + C content of genomic DNA was 69.1 mol%. Strain 17Sr1-43 T was closely related to Methylobacterium gregans KACC 14808 T (98.4% 16S rRNA gene sequence similarity), Methylobacterium hispanicum KACC 11432 T (97.9%), and Methylobacterium phyllosphaerae CBMB27T (96.1%). The complete genome of strain 17Sr1-43 T contains essential genes related to DNA repair processes including bacterial RecBCD dependent pathway and UmuCD system. Based on the phenotypic, genotypic, and chemotaxonomic characteristics, strain 17Sr1-43 T represents a novel species in the genus Methylobacterium, for which the name Methylobacterium radiodurans sp. nov. is proposed. The type strain is strain 17Sr1-43 T (= KCTC 52906 T = NBRC 112875 T).

Methylobacterium nonmethylotrophicum sp. nov., isolated from tungsten mine tailing.

A novel pink-pigmented strain, designated 6HR-1T, was isolated from tungsten mine tailings in Jiangxi Province, PR China. Cells were Gram-stain-negative, aerobic, non-spore-forming, rod-shaped and motile with a polar flagellum (monotrichous). It could not utilize methanol, methylamine, formaldehyde or formate as a sole carbon source. The methanol dehydrogenase mxaF gene was absent but the xoxF gene was present. Phylogenomic and 16S rRNA gene phylogenetic analyses clearly showed that strain 6HR-1T was affiliated to the genus Methylobacterium and closely related to 'Methylobacterium terrae' 17Sr1-28T (98.6 %), Methylobacterium platani JCM 14648T (97.7 %), Methylobacterium variabile DSM 16961T (97.7 %) and Methylobacterium currus KACC 19662T (97.4 %). The average nucleotide identity and digital DNA-DNA hybridization values between strain 6HR-1T and its closely related type species were 87.4-88.7 and 33.2-36.3 %, respectively. It had summed feature 8 (C18 : 1 ω7c and/or C18 : 1 ω6c) as the major fatty acid and ubiquinone 10 as the predominant respiratory quinone. Polyphasic characterization supported that strain 6HR-1T represents a novel species of the genus Methylobacterium, for which the name Methylobacterium nonmethylotrophicum sp. nov. is proposed with the type strain 6HR-1T (=GDMCC 1.662T=KCTC 42760T).

Methylobacterium terricola sp. nov., a gamma radiation-resistant bacterium isolated from gamma ray-irradiated soil.

A gamma radiation-resistant and pink-pigmented bacterial strain, designated as 17Sr1-39T, was isolated from a gamma ray-irradiated soil sample collected in the Republic of Korea. Cells were Gram-stain-negative, strictly aerobic, flagellated, asporogenous, rod-shaped and methylotrophic. Results of 16S rRNA gene sequence analysis showed that strain 17Sr1-39T was phylogenetically related to Methylobacterium currus PR1016AT (97.3 %), Methylobacterium aquaticum DSM 16371T (97.2 %), Methylobacterium platani PMB02T (97.0 %), Methylobacterium frigidaeris IER25-16T (96.6 %), Methylobacterium terrae 17Sr1-28T (96.6 %) and Methylobacterium organophilum JCM 2833T (93.4 %). The G+C content calculated based on the genome sequence was 70.4 mol%. The average nucleotide identity and in silico DNA-DNA hybridization values between strain 17Sr1-39T and M. currus, M. aquaticum, M. platani, M. frigidaeris, M. terrae and M. organophilum were 77.3-89.9 and 22-38.2 %, respectively. The predominant fatty acids were summed feature 8 (C18 : 1ω7c and/or C18 : 1ω6c) and summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c). The predominant quinone was ubiquinone 10 and the major polar lipids were diphosphatidylglycerol, phosphatidylglycerol and phosphatidylethanolamine. Based on the data from phenotypic tests and genotypic differences between strain 17Sr1-39T and its close phylogenetic relatives, strain 17Sr1-39T represented a new species belonging to the genus Methylobacterium, for which the name Methylobacterium terricola sp. nov. (=KACC 52905T=NBRC 112874T) is proposed.

Methylobacterium terrae sp. nov., a radiation-resistant bacterium isolated from gamma ray-irradiated soil.

A Gram-stain-negative, asporogenous, aerobic rods, motile by means of a single polar flagellum, catalase- and oxidase-positive, methylotrophic bacterium, designated 17Sr1-28T, was isolated from gamma ray-irradiated soil. The 16S rRNA gene sequence analysis showed that strain 17Sr1-28T was phylogenetically related to Methylobacterium currus PR1016AT (96.8%), Methylobacterium platani PMB02T (96.2%), Methylobacterium aquaticum DSM 16371T (96.3%), Methylobacterium tarhaniae N4211T (96.4%), Methylobacterium frigidaeris IER25-16T (95.8%), and Methylobacterium organophilum JCM 2833T (92.7%). The G+C content calculated based on genome sequence was 71.6%. The average nucleotide identity and in silico DNA-DNA hybridization values between strain 17Sr1- 28T and M. currus, M. platani, M. aquaticum, M. tarhaniae, M. frigidaeris, and M. organophilum were 77.7-90.4% and 22-39.6%, respectively. The major fatty acids of strain 17Sr1-28T were summed feature 8 (C18:1ω7c and/or C18:1ω6c), and summed feature 3 (C16:1ω7c and/or C16:1ω6c). The predominant quinone was ubiquinone 10 and the major polar lipids were diphosphatidylglycerol, phosphatidylcholine, phosphatidylethanolamine, and phosphatidylglycerol. On the basis of the data from phenotypic tests and genotypic differences between strain 17Sr1-28T and its close phylogenetic relatives, strain 17Sr1-28T represents a new species belonging to the genus Methylobacterium, for which the name Methylobacterium terrae sp. nov. (= KCTC 52904T = NBRC 112873T) is proposed.

Distribution of 1-aminocyclopropane-1-carboxylate deaminase and D-cysteine desulfhydrase genes among type species of the genus Methylobacterium.

The presence of 1-aminocyclopropane-1-carboxylate (ACC) deaminase determines the ability of bacteria to increase the resistance of plants to various types of stress. The genes of ACC deaminase (acdS) and the closely related enzyme D-cysteine desulfhydrase (dcyD) were searched in type strains of various representatives of the genus Methylobacterium. Using PCR screening and in silico searching in the available complete genome sequences of type strains, the genes were found in 28 of 48 species of the genus. Phylogenetic analysis of amino acid sequences of proteins revealed two large groups of sequences of the AcdS protein and one of the DcyD protein. The distribution of these groups correlates well with the phylogenetic tree based on the sequences of the 16S rRNA genes, which apparently indicates a different evolutionary adaptation to association with plants in the representatives of these groups. For the first time for aerobic methylotrophs it was demonstrated that the gene dcyD encodes D-cysteine desulfhydrase by cloning and recombinant protein characterization.

Methylobacterium populi VP2: plant growth-promoting bacterium isolated from a highly polluted environment for polycyclic aromatic hydrocarbon (PAH) biodegradation.

The use of microorganisms to accelerate the natural detoxification processes of toxic substances in the soil represents an alternative ecofriendly and low-cost method of environmental remediation compared to harmful incineration and chemical treatments. Fourteen strains able to grow on minimal selective medium with a complex mixture of different classes of xenobiotic compounds as the sole carbon source were isolated from the soil of the ex-industrial site ACNA (Aziende Chimiche Nazionali Associate) in Cengio (Savona, Italy). The best putative degrading isolate, Methylobacterium populi VP2, was identified using a polyphasic approach on the basis of its phenotypic, biochemical, and molecular characterisation. Moreover, this strain also showed multiple plant growth promotion activities: it was able to produce indole-3-acetic acid (IAA) and siderophores, solubilise phosphate, and produce a biofilm in the presence of phenanthrene and alleviate phenanthrene stress in tomato seeds. This is the first report on the simultaneous occurrence of the PAH-degrading ability by Methylobacterium populi and its multiple plant growth-promoting activities. Therefore, the selected indigenous strain, which is naturally present in highly contaminated soils, is good candidate for plant growth promotion and is capable of biodegrading xenobiotic organic compounds to remediate contaminated soil alone and/or soil associated with plants.

Methylobacterium pseudosasicola sp. nov. and Methylobacterium phyllostachyos sp. nov., isolated from bamboo leaf surfaces.

Two strains of Gram-negative, methylotrophic bacteria, isolated because of their abilities to promote plant growth, were subjected to a polyphasic taxonomic study. The isolates were strictly aerobic, motile, pink-pigmented, facultatively methylotrophic, non-spore-forming rods. The chemotaxonomic characteristics of the isolates included the presence of C18 : 1ω7c as the major cellular fatty acid. The DNA G+C contents of strains BL36(T) and BL47(T) were 69.4 and 69.8 mol%, respectively. 16S rRNA gene sequence analysis of strains BL36(T) and BL47(T) placed them under the genus Methylobacterium, with the pairwise sequence similarity between them and the type strains of closely related species ranging from 97.2 to 99.0%. On the basis of their phenotypic and phylogenetic distinctiveness and the results of DNA-DNA hybridization analysis, the isolates represent two novel species within the genus Methylobacterium, for which the names Methylobacterium pseudosasicola sp. nov. (type strain BL36(T) = NBRC 105203(T) = ICMP 17621(T)) and Methylobacterium phyllostachyos sp. nov. (type strain BL47(T) = NBRC 105206(T) = ICMP 17619(T)) are proposed.

Methylobacterium tarhaniae sp. nov., isolated from arid soil.

A reddish-orange-pigmented, Gram-stain-negative, aerobic, facultatively methylotrophic strain, N4211(T), isolated from arid soil, collected from Abuja, Nigeria, was analysed by using a polyphasic approach. Phylogenetic analysis, based on 16S rRNA gene sequences, showed that strain N4211(T) belonged to the genus Methylobacterium. Strain N4211(T) was most closely related to Methylobacterium aquaticum GR16(T) (98.56 %), Methylobacterium platani PMB02(T) (97.95 %) and Methylobacterium variabile GR3(T) (97.2 %), and the phylogenetic similarities to all other species of the genus Methylobacterium with validly published names were less than 97.0 %. The major ubiquinones detected were Q-10. The major fatty acids were summed feature 7 (C18 : 1 cis11/t9/t6). The DNA G+C content was 67.3 mol%. DNA-DNA relatedness of strain N4211(T) and the most closely related strains M. aquaticum DSM 16371(T) and M. platani KCTC 12901(T) were 60.0 and 48.2 %, respectively. On the basis of phenotypic, phylogenetic and DNA-DNA hybridization data, strain N4211(T) is assigned to a novel species of the genus Methylobacterium for which the name Methylobacterium tarhaniae sp. nov. is proposed. The type strain is N4211(T)( = KCTC 23615(T) = DSM 25844(T)).

Cytotoxic (A549) and antimicrobial effects of Methylobacterium sp. isolate (ERI-135) from Nilgiris forest soil, India.

OBJECTIVE: To assess the antimicrobial and cytotoxic effects of Methylobacterium sp. isolated from soil sample of Doddabetta forest, Nilgiris, Western Ghats of Tamil Nadu. METHODS: Isolation of Methylobacterium was performed from soils by serial dilution plate technique. The strain was grown in modified nutrient gulucose agar (MNGA) medium to study the morphology and biochemical characteristics. Methylobacterium sp. was screened for its antimicrobial activity against pathogenic bacteria and fungi. The strain was subjected to 16S rRNA analysis and was identified as Methylobacterium sp. The nucleotide sequence of the 16S rRNA gene of the isolate exhibited close similarity with other Methylobacterium sp. and has been submitted to Genbank. The antibacterial substances were extracted using chloroform and ethyl acetate from MNGA medium in which ERI-135 had grown for 5 d at 30 °C. Cytotoxic effect was also studied. GC-MS analysis was carried out. The antimicrobial activity was assessed using broth micro dilution technique. RESULTS: Ethyl acetate extract showed activity against bacteria such as Bacillus subtilis, Klebsiella pneumoniae (K. pneumoniae), Pseudomonas aeruginosa, Salmonella typhimurium, Shigella flexneri, Enterobacter aerogenes, Staphylococcus aureu and Staphylococcus epidermidis (S. epidermidis) and fungi such as, Candida albicans and Trichophyton rubrum. The lowest minimum inhibitory concentrations were: 250 µg/mL against S. epidermidis and 250µg/mL against K. pneumonia. The isolate had the ability to produce enzymes such as protease. The exyract showed cytotoxic effect in human adenocarcinoma cancer cell line (A549). GC-MS analysis showed the presence of isovaleric acid (3.64%), 2-Methylbutanoic acid (5.03%), isobutyramide (5.05%), N,N-oimethylformamide-di-t-butylacetal (9.79%), benzeneacetamide (15.56%), octyl butyl phthalate (3.59%) and diisooctyl phthalate (5.79) in the extract. CONCLUSIONS: Methylobacterium sp. (ERI-135) showed promising antibacterial and cytotoxic activity. This is the first report in the antimicrobial and cytotoxic effect of Methylobacterium sp.

Methylobacterium bullatum sp. nov., a methylotrophic bacterium isolated from Funaria hygrometrica.

A novel, pink-pigmented aerobic, facultatively methylotrophic bacterial strain (F3.2(T)) isolated from the phyllosphere of Funaria hygrometrica, was analyzed using a polyphasic approach. Cells were Gram-negative, motile rods, strictly aerobic and non-spore-forming and exhibited surface structures varying in quantity, distribution and morphology. The isolate grew at 10-33°C over a pH range of 5.5-8.0 and in the presence of less than 1.0% NaCl. Strain F3.2(T) shared less than 70% DNA-DNA binding to the next type strain of the genus Methylobacterium (M. adhaesivum DSM 17169(T)). In addition to the major cellular fatty acid C(18:1)ω7c (81.7%), present in all Methylobacterium species (and also members of the genus Alphaproteobacteria), a high value (11.7%) of the fatty acids (summed feature) C(16:1)ω7c and/or iso-C(15:0)2OH was determined. Phylogenetic analyses based on 16S rDNA and methanol dehydrogenase gene sequences, DNA-DNA hybridization values, chemotaxonomic and phenotypic characteristics indicate that the strain F3.2(T) represents a novel species within the genus Methylobacterium. We propose the name Methylobacterium bullatum sp. nov. for this species. The type strain is the strain F3.2(T) (DSM 21893(T)=LMG 24788(T)).

Methylobacterium marchantiae sp. nov., a pink-pigmented, facultatively methylotrophic bacterium isolated from the thallus of a liverwort.

A pink-pigmented, facultatively methylotrophic bacterium, designated strain JT1(T), was isolated from a thallus of the liverwort Marchantia polymorpha L. and was analysed by using a polyphasic approach. Comparative 16S rRNA gene sequence analysis placed the strain in a clade with Methylobacterium adhaesivum AR27(T), Methylobacterium fujisawaense DSM 5686(T), Methylobacterium radiotolerans JCM 2831(T) and Methylobacterium jeotgali S2R03-9(T), with which it showed sequence similarities of 97.8, 97.7, 97.2 and 97.4 %, respectively. However, levels of DNA-DNA relatedness between strain JT1(T) and these and the type strains of other closely related species were lower than 70 %. Cells of JT1(T) stained Gram-negative and were motile, rod-shaped and characterized by numerous fimbriae-like appendages on the outer surface of their wall (density up to 200 µm(-2)). Major fatty acids were C(18 : 1)ω7c and C(16 : 0). Based on the morphological, physiological and biochemical data presented, strain JT1(T) is considered to represent a novel species of the genus Methylobacterium, for which the name Methylobacterium marchantiae sp. nov. is proposed. The type strain is JT1(T) ( = DSM 21328(T)  = CCUG 56108(T)).

Methylobacterium phyllosphaerae sp. nov., a pink-pigmented, facultative methylotroph from the phyllosphere of rice.

A pink-pigmented, aerobic, facultatively methylotrophic bacterial strain, CBMB27T, isolated from leaf tissues of rice (Oryza sativa L. 'Dong-Jin'), was analysed using a polyphasic taxonomic approach. Comparative 16S rRNA gene sequence-based phylogenetic analysis placed the strain in a clade with the species Methylobacterium oryzae, Methylobacterium fujisawaense and Methylobacterium mesophilicum; strain CBMB27T showed sequence similarities of 98.3, 98.5 and 97.3 %, respectively, to the type strains of these three species. DNA-DNA hybridization experiments revealed low levels (<38 %) of DNA-DNA relatedness between strain CBMB27T and its closest relatives. The sequence of the 1-aminocyclopropane-1-carboxylate deaminase gene (acdS) in strain CBMB27T differed from those of close relatives. The major fatty acid of the isolate was C(18 : 1)omega7c and the G+C content of the genomic DNA was 66.8 mol%. Based on the results of 16S rRNA gene sequence analysis, DNA-DNA hybridization, and physiological and biochemical characterization, which enabled the isolate to be differentiated from all recognized species of the genus Methylobacterium, it was concluded that strain CBMB27T represents a novel species in the genus Methylobacterium for which the name Methylobacterium phyllosphaerae sp. nov. is proposed (type strain CBMB27T =LMG 24361T =KACC 11716T =DSM 19779T).

Methylobacterium persicinum sp. nov., Methylobacterium komagatae sp. nov., Methylobacterium brachiatum sp. nov., Methylobacterium tardum sp. nov. and Methylobacterium gregans sp. nov., isolated from freshwater.

Eight strains, 002-165T, 002-079T, B0021T, Hojyo2, RB603B, RB677T, 002-074T and RB678, isolated from the environment of food-processing factories in Japan, were characterized using a polyphasic approach. The isolates were Gram-negative, strictly aerobic, pink-pigmented, facultatively methylotrophic, non-spore-forming rods. The chemotaxonomic characteristics of these isolates included the presence of C18 : 1omega7c as the major cellular fatty acid and ubiquinone Q-10 as the predominant ubiquinone. The DNA G+C content was 67.1-71.1 mol%. Phylogenetic analyses of 16S rRNA and DNA gyrase B subunit (gyrB) nucleotide sequence confirmed that the eight strains belonged to the Methylobacterium clade. Moreover, a DNA-DNA hybridization analysis showed that the eight isolates represented five novel species. On the basis of their phenotypic and phylogenetic distinctiveness, the isolates represent five novel species within the genus Methylobacterium, for which the names Methylobacterium persicinum sp. nov. (type strain 002-165T =DSM 19562T =NBRC 103628T =NCIMB 14378T), Methylobacterium komagatae sp. nov. (type strain 002-079T =DSM 19563T =NBRC 103627T =NCIMB 14377T), Methylobacterium brachiatum sp. nov. (type strain B0021T =DSM 19569T =NBRC 103629T =NCIMB 14379T), Methylobacterium tardum sp. nov. (type strain RB677T =DSM 19566T =NBRC 103632T =NCIMB 14380T) and Methylobacterium gregans sp. nov. (type strain 002-074T =DSM 19564T =NBRC 103626T =NCIMB 14376T) are proposed.

Methylobacterium iners sp. nov. and Methylobacterium aerolatum sp. nov., isolated from air samples in Korea.

Two bacterial strains isolated from air samples, 5317S-33(T) and 5413S-11(T), were characterized by determining their phenotypic characteristics, cellular fatty acid profiles and phylogenetic positions based on 16S rRNA gene sequence analysis. 16S rRNA gene sequence analysis showed that these isolates belonged to the genus Methylobacterium. Strain 5317S-33(T) was most closely related to Methylobacterium adhaesivum AR27(T) (97.9% sequence similarity). Strain 5413S-11(T) was most closely related to Methylobacterium fujisawaense DSM 5686(T) (97.3% sequence similarity), Methylobacterium oryzae CBMB20(T) (97.1% similarity) and Methylobacterium radiotolerans JCM 2831(T) (97.0% similarity). Cells of both strains were strictly aerobic, Gram-negative, motile and rod-shaped. The major fatty acid was C(18:1)omega7c. The G+C contents of the genomic DNA were 68.0 mol% for strain 5317S-33(T) and 73.2 mol% for strain 5413S-11(T). According to DNA-DNA hybridization data, strain 5317S-33(T) showed a level of DNA-DNA relatedness of 33 % with M. adhaesivum DSM 17169(T), and strain 5413S-11(T) showed low levels of DNA-DNA relatedness (<35%) with M. fujisawaense DSM 5686(T), M. oryzae CBMB20(T) and M. radiotolerans DSM 1819(T). On the basis of this polyphasic analysis, it was concluded that strains 5317S-33(T) and 5413S-11(T) represent two novel species within the genus Methylobacterium, for which the names Methylobacterium iners sp. nov. (type strain 5317S-33(T) =KACC 11765(T) =DSM 19015(T)) and Methylobacterium aerolatum sp. nov. (type strain 5413S-11(T) =KACC 11766(T) =DSM 19013(T)) are proposed.

Methylobacterium oryzae sp. nov., an aerobic, pink-pigmented, facultatively methylotrophic, 1-aminocyclopropane-1-carboxylate deaminase-producing bacterium isolated from rice.

A pink-pigmented, facultatively methylotrophic bacterium, strain CBMB20T, isolated from stem tissues of rice, was analysed by a polyphasic approach. Strain CBMB20T utilized 1-aminocyclopropane 1-carboxylate (ACC) as a nitrogen source and produced ACC deaminase. It was related phylogenetically to members of the genus Methylobacterium. 16S rRNA gene sequence analysis indicated that strain CBMB20T was most closely related to Methylobacterium fujisawaense, Methylobacterium radiotolerans and Methylobacterium mesophilicum; however, DNA-DNA hybridization values were less than 70 % with the type strains of these species. The DNA G+C content of strain CBMB20T was 70.6 mol%. The study presents a detailed phenotypic characterization of strain CBMB20T that allows its differentiation from other Methylobacterium species. In addition, strain CBMB20T is the only known member of the genus Methylobacterium to be described from the phyllosphere of rice. Based on the data presented, strain CBMB20T represents a novel species in the genus Methylobacterium, for which the name Methylobacterium oryzae sp. nov. is proposed, with strain CBMB20T (=DSM 18207T=LMG 23582T=KACC 11585T) as the type strain.

Formaldehyde uptake by Methylobacterium sp. MF1 and Acidomonas methanolica MB 58 with the different formaldehyde assimilation pathways.

Methylobacterium sp. MF1 (an obligate methylotrophic bacterium isolated newly by the authors) and Acidomonas methanolica MB58 (a facultative methylotrophic bacterium) uptake formaldehyde similarly. It was found that the former assimilated formaldehyde via the serine pathway whereas the latter did so via the ribulose-monophosphate pathway from the measurement of the key enzyme activities in each assimilation pathway. That is, hydroxy pyruvate reductase was detected in only the above-mentioned MF1 strain, but hexulose phosphate synthase (HPS) was not. The efficiencies of formaldehyde consumption by both strains under a continuous chemostat cultivation in the steady state were almost the same in spite of their different assimilation pathways. That is, the consumption efficiencies of the MF strain and the MB58 strain were ca. 1.2 g/L/d and ca. 1.8 g/L/d, respectively, under the experimental conditions. In the future, optimal continuous operating conditions will be investigated.