- Volume 57, Issue 3, 2007

Two bacterial strains, designated GH34-4T and GH41-7T, were isolated from greenhouse soil cultivated with cucumber. The bacteria were strictly aerobic, Gram-negative, rod-shaped and oxidase- and catalase-positive. 16S rRNA gene sequence analysis indicated that these strains belong to the genus Lysobacter within the Gammaproteobacteria. Strain GH34-4T showed highest sequence similarity to Lysobacter yangpyeongensis GH19-3T (97.5 %) and Lysobacter koreensis Dae16T (96.4 %), and strain GH41-7T showed highest sequence similarity to Lysobacter antibioticus DSM 2044T (97.5 %), Lysobacter enzymogenes DSM 2043T (97.5 %) and Lysobacter gummosus ATCC 29489T (97.4 %). Levels of DNA–DNA relatedness indicated that strains GH34-4T and GH41-7T represented species clearly different from L. yangpyeongensis, L. antibioticus, L. enzymogenes and L. gummosus. The major cellular fatty acids of strains GH34-4T and GH41-7T were iso-C16 : 0, iso-C15 : 0 and iso-C17 : 1 ω9c, and the major isoprenoid quinone was Q-8. The DNA G+C contents of GH34-4T and GH41-7T were 62.5 and 66.6 mol%, respectively. On the basis of the polyphasic taxonomic data presented, it is evident that each of these strains represents a novel species of the genus Lysobacter, for which the names Lysobacter niabensis sp. nov. (type strain GH34-4T=KACC 11587T=DSM 18244T) and Lysobacter niastensis sp. nov. (type strain GH41-7T=KACC 11588T=DSM 18481T) are proposed.

Three strains, MAFF 210191T, G24103T and G24116, assumed to be members of two novel species, were isolated from several rhizosphere habitats in different parts of Japan. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the isolates formed a distinct monophyletic group together with the two known species of the genus Labrys, suggesting that the isolates have a close affiliation with this genus. In this study, a polyphasic approach was used to characterize and compare the three isolates with the two species of the genus Labrys, Labrys monachus and Labrys methylaminiphilus. All three isolates were aerobic, Gram-negative, motile and non-sporulating and they ranged in shape from spherical to short rods. The cells multiplied by budding and utilized a wide variety of monosaccharides, disaccharides and sugar alcohols as sole carbon and energy sources, but they did not utilize C1 compounds, salicin or d-melezitose. The strains were inhibited by dl-α-alanine and glycine (both at 10 mM). The major cellular fatty acids were C19 : 0 cyclo ω8c, C16 : 0, C18 : 0 and C18 : 1 ω7c. The three isolates shared <12 % and <11 % DNA–DNA relatedness with L. monachus DSM 5896T and L. methylaminiphilus DSM 16812T, respectively. The G+C content of the isolates (61–62 mol%) was also significantly lower than those of the two previously characterized species. In spite of many morphological, physiological and chemotaxonomic similarities among the three isolates, strain MAFF 210191T could be differentiated from strains G24103T and G24116 on the basis of 16S rRNA gene sequence divergence, DNA–DNA relatedness (<46 %) and gelatin hydrolysis. Two novel species are therefore proposed, namely Labrys okinawensis sp. nov., with the type strain MAFF 210191T (=DSM 18385T), and Labrys miyagiensis sp. nov., with the type strain G24103T (=NBRC 101365T=NCIMB 14143T) and also including strain G24116 (=NBRC 101366=NCIMB 14144). Emended descriptions of the genus Labrys and Labrys monachus are also presented.

Two Gram-negative, facultatively anaerobic bacterial strains, MANO22DT and MANO22P, were isolated from a tidal flat area of Dae-Chun, Chung-Nam, Korea. The isolates were rod-shaped and were motile by means of one or more polar flagella. They grew at 1–12 % NaCl, 4–45 °C and pH 4.1–8.8 and were oxidase-positive, arginine dihydrolase-negative and sensitive to the vibriostatic agent O/129. The isolates required Na+ for growth, produced acid, but no gas, from d-glucose under anaerobic conditions and utilized a wide range of compounds as sole carbon and energy sources. A phylogenetic analysis based on 16S rRNA gene sequences revealed that the strains belong to the Gammaproteobacteria and are specifically related to Vibrio species. They were most closely related to Vibrio rumoiensis FERM P-14531T, with which they were found to share 98.65 % 16S rRNA gene sequence similarity. In the phylogenetic tree, the two novel strains comprised a relatively long subline of descent, sharing a branching point with the outlying species V. rumoiensis, and were found to occupy a phylogenetically distant position on the main Vibrio branch. The levels of DNA–DNA hybridization with respect to V. rumoiensis FERM P-14531T, which is their most closely related phylogenetically related Vibrio species, were 7.4 % (MANO22DT) and 3.9 % (MANO22P). Thus, the two novel isolates appear to represent a novel species within the genus Vibrio, for which the name Vibrio litoralis sp. nov. is proposed. The type strain is MANO22DT (=KCTC 12520T=DSM 17657T).

A novel facultatively methylotrophic, Gram-negative, rod-shaped bacterium, designated strain S2R03-9T, was isolated from jeotgal, a traditional Korean fermented seafood. The organism was strictly aerobic, motile by means of a single polar flagellum, non-sporulating and catalase- and oxidase-positive. Strain S2R03-9T grew in the presence of 0–1 % (w/v) NaCl and at pH 6.0–10.0, with optimum growth in the absence of NaCl and at pH 7.0. It grew at temperatures in the range 20.0–30.0 °C, with optimum growth at 30 °C. Colonies grown on R2A medium were non-pigmented, opaque and creamy white. Phylogenetic analysis based on 16S rRNA gene sequences indicated that it was most closely related to Methylobacterium organophilum JCM 2833T (96.6 % similarity) and the phylogenetic similarities to all other Methylobacterium species with validly published names were less than 95.0 %. The DNA G+C content was 64.9 mol%. The phylogenetic analysis, the phenotypic assessment and the chemotaxonomic data (major ubiquinone, Q-10; major fatty acids, C18 : 1 and C18 : 0) showed that S2R03-9T represents a novel species within the genus Methylobacterium in the class Alphaproteobacteria, for which the name Methylobacterium jeotgali sp. nov. is proposed. The type strain is S2R03-9T (=KCTC 12671T=LMG 23639T).

The taxonomic position of Pseudomonas sp. B13T, isolated as a 3-chlorobenzoate-degrading organism and used for several groundbreaking studies on the enzymology and genetics of the degradative pathway for haloaromatic compounds, was studied in detail. The previously performed physiological studies, the detection of ubiquinone Q-9, the polyamine pattern with putrescine and spermidine as major polyamines, a fatty acid profile with C18 : 1 ω7c, summed feature 3 and C16 : 0 as quantitatively the most important constituents and the 16S rRNA gene sequence demonstrated that Pseudomonas sp. B13T indeed belongs to the genus Pseudomonas. The sequence of the Pseudomonas sp. B13T 16S rRNA gene demonstrated a high degree of similarity with that of Pseudomonas citronellolis DSM 50332T (98.9 %), Pseudomonas nitroreducens DSM 14399T (98.7 %), Pseudomonas jinjuensis DSM 16612T (98.1 %) and Pseudomonas multiresinivorans DSM 17553T (98.7 %). Thus it was shown that strain Pseudomonas sp. B13T can be distinguished from related species by the ability/inability to assimilate N-acetylgalactosamine, d-galactose, putrescine, trans-aconitate and mesaconate and some differences in the fatty acid profile. The positioning of Pseudomonas sp. B13T as a separate taxon was finally verified by DNA hybridization, which demonstrated less than 45 % DNA–DNA similarity between strain Pseudomonas sp. B13T and the reference strains. On the basis of these results, Pseudomonas sp. B13T represents a novel species for which the name Pseudomonas knackmussii sp. nov. is proposed. The type strain is B13T (=DSM 6978T=LMG 23759T).

A bacterium designated strain Liujia-146T was isolated in the Tainan area of southern Taiwan from root nodules of the aquatic legume Neptunia oleracea. 16S rRNA gene sequence analysis indicated that strain Liujia-146T was highly similar to Labrys monachus VKM B-1479T (97.8 %) and Labrys methylaminiphilus JLW10T (95.5 %) and belonged to the order Rhizobiales in the Alphaproteobacteria. On the basis of phylogenetic analysis, DNA–DNA hybridization data, physiological and biochemical characteristics and fatty acid compositions, the organism was shown to belong to the genus Labrys whilst representing a novel species within this genus. We propose to classify strain Liujia-146T (=BCRC 17578T=LMG 23578T) as the type strain of Labrys neptuniae sp. nov.

A polyphasic study was performed to determine the taxonomic position of two Aeromonas strains, 665N and 868ET, isolated from bivalve molluscs, that could not be identified at the species level in a previous numerical taxonomy study. The DNA G+C content of these isolates was 62.3 and 62.6 mol%, respectively. Sequence analysis of the 16S rRNA gene showed that the two new strains were closely related to members of the genus Aeromonas. Fluorescence amplified fragment length polymorphism fingerprinting revealed that strains 665N and 868ET clustered together with a similarity of 78 % but did not cluster with any of the Aeromonas genomospecies. DNA–DNA hybridization experiments revealed a high level of relatedness between the two new isolates (76 %) but low levels of relatedness between these and phylogenetically most closely related Aeromonas genomospecies (30–44 %). Useful tests for the phenotypic differentiation of strains 665N and 868ET from other mesophilic Aeromonas species included those for gas from glucose, lysine decarboxylase, Voges–Proskauer reaction, acid from l-arabinose, hydrolysis of aesculin and utilization of l-lactate. On the basis of genotypic and phenotypic evidence, strains 665N and 868ET are considered to represent a novel species of the genus Aeromonas, for which the name Aeromonas bivalvium sp. nov. is proposed. The type strain is 868ET (=CECT 7113T=LMG 23376T).

Psychrotolerant (0–25 °C), chemolithotrophic Gram-negative cocci were isolated from Alaskan forest soil. The novel isolate was found to grow autotrophically on H2 : CO2 mixtures and to switch to heterotrophic growth on media containing organic substrates. The novel strain utilized a wide range of organic acids, some simple sugars and alcohols. Naphthalene vapour did not support growth. On the basis of 16S rRNA gene sequence similarity, the novel strain is affiliated to the genus Polaromonas, of the class Betaproteobacteria, and is related to Polaromonas naphthalenivorans (99.6 % gene sequence similarity), Polaromonas aquatica (97.4 %) and Polaromonas vacuolata (96.1 %). The membrane phospholipids contained 16 : 1ω7c/16 : 1ω6c, 16 : 0 and 18 : 1ω7c, similar to the fatty acids found for P. naphthalenivorans, P. aquatica and P. vacuolata. On the basis of DNA–DNA hybridization, physiological and biochemical properties, the hydrogen-oxidizing mixotrophic isolate represents a novel species, for which the name Polaromonas hydrogenivorans sp. nov. is proposed. The type strain is DSM 17735T (=NRRL B-41369T).

A free-living diazotrophic strain, DS2T, was isolated from corn rhizosphere. Polyphasic taxonomy was performed including morphological characterization, Biolog analysis, and 16S rRNA, cpn60 and nifH gene sequence analyses. 16S rRNA gene sequence analysis indicated that strain DS2T was closely related to the genus Azospirillum (96 % similarity). Chemotaxonomic characteristics (DNA G+C content 67.9 mol%; Q-10 quinone system; major fatty acid 18 : 1ω7c) were also similar to those of the genus Azospirillum. In all the analyses, including phenotypic characterization using Biolog analysis and comparison of cellular fatty acids, this isolate was found to be different from the closely related species Azospirillum lipoferum, Azospirillum oryzae and Azospirillum brasilense. On the basis of these results, a novel species is proposed for this nitrogen-fixing strain. The name Azospirillum canadense sp. nov. is suggested with the type strain DS2T (=NCCB 100108T=LMG 23617T).

The taxonomic position of Actinobacillus capsulatus, a member of the family Pasteurellaceae found in rabbits, hares and hamsters, has been challenged. 16S rRNA gene (rrs) sequence data show the species to be heterogeneous. Using a polyphasic approach, 23 strains that were identified previously as belonging, or closely related, to A. capsulatus were analysed. Eighty characters were included in the phenotypic analysis. Phylogenetic analysis was done based on rrs, rpoB, infB and recN sequences. In addition, the recN sequence similarities were used to calculate the whole-genome sequence relatedness of all strains investigated as well as that with other members of the family Pasteurellaceae. The phenotypic analysis allowed identification of five groups. The major group of 17 strains could be classified as A. capsulatus. Two hamster isolates were closely related to A. capsulatus but differed in a few characters. Single isolates from a rabbit and snowshoe-hare were phenotypically related to Actinobacillus suis. One rabbit isolate was related to the genus Mannheimia, while another isolate could not be classified phenotypically with known taxa. The phylogenetic analysis confirmed the phenotypic grouping. In contrast to the rrs-based tree, the A. capsulatus strains clustered unambiguously with the type species and related species of the genus Actinobacillus in the rpoB-, infB- and recN-based trees. Genome similarity comparison using recN finally confirmed the high genomic relationship of the A. capsulatus strains with the type species and related species of the genus Actinobacillus and allowed a clear assignment of the other unrelated strains to the phenotypic and phylogenetic clusters outlined. The present findings allow the description of A. capsulatus to be emended and separate it more clearly from other species, both phenotypically and genotypically. The type strain of A. capsulatus is CCUG 12396T (=Frederiksen 243T=ATCC 51571T=NCTC 11408T=CIP 103283T).

Two strains, designated WB 3.4-79T and WB 3.3-25, were isolated from a hard-water sample collected from the Westerhöfer Bach, Lower Saxony, Germany. The strains shared 100 % DNA–DNA relatedness, indicating membership of the same genospecies. This close relationship was supported by identical 16S rRNA gene sequences and high similarities in fatty acid composition and biochemical characteristics. The G+C content of the genomic DNA of strain WB 3.4-79T was 48.5 mol% and the predominant ubiquinone was Q-8. Major polar lipids were phosphatidylethanolamine and phosphatidylglycerol. Major fatty acids (>10 %) were C16 : 0 and C16 : 1 ω7c. Polyhydroxybutyrate and polyphosphate granules as well as unidentified enterosomes and a polar organelle are visible by electron microscopy. Comparative 16S rRNA gene sequence analysis indicated that the isolates were placed within the class Betaproteobacteria, remotely related to Chitinibacter tainanensis DSM 15459T, Silvimonas terrae KCTC 12358T, Formivibrio citricus DSM 6150T and Iodobacter fluviatilis DSM 3764T. On the basis of phylogenetic and phenotypic distinctness, we propose a novel genus, Deefgea gen. nov., with Deefgea rivuli sp. nov. as the type species. The type strain of Deefgea rivuli is strain WB 3.4-79T (=DSM 18356T=CIP 109326T).

A Gram-negative, strictly aerobic, non-spore-forming bacterium, motile by means of single polar flagellum and rod-shaped, designated strain GR12-1T, was isolated from soil of a ginseng field in Yeongju region, Korea. Phylogenetic analysis based on 16S rRNA gene sequences indicated that this strain is related to members of the genus Pseudoxanthomonas, showing sequence similarity values ranged from 92.3 to 96.2 %. This organism grew at 5–33 °C, with optimum growth at 28 °C. Strain GR12-1T grew optimally in the presence of 0–2 % NaCl. The whole-cell fatty acid profile included iso-C15 : 0, iso-C17 : 1 ω9c, iso-C16 : 0, iso-C11 : 0 3-OH and iso-C17 : 0 as major components. The only isoprenoid quinone was ubiquinone 8 (Q-8). The DNA G+C content was 63.4 mol%. On the basis of phenotypic, genetic and phylogenetic data, strain GR12-1T should be classified as a member of a novel species of the genus Pseudoxanthomonas, for which the name Pseudoxanthomonas yeongjuensis sp. nov. is proposed, with strain GR12-1T (=KACC 11580T=DSM 18204T) as the type strain.

Four yeast strains (TM1-01T, TM1-07, TM3-47 and TM3-49) were isolated by membrane filtration from estuarine water collected from a mangrove forest in Phang-Nga province, southern Thailand. Analysis of the D1/D2 domains of the large-subunit rDNA sequence revealed that the sequences of the four strains were identical. The closest recognized species in terms of pairwise sequence similarity was Pichia deserticola, but the level of nucleotide substitution (4.8 %) was sufficient to justify the description of a separate species. Phylogenetic analysis demonstrated that the four strains occupy a basal position with respect to Pichia membranifaciens and close relatives. The four strains showed identical phenotypic characteristics, including proliferation by multilateral budding, absence of ascospores, arthrospores and ballistospores and negative reactions for Diazonium blue B and urease. The major ubiquinone was Q-7. On the basis of the above findings, these four strains were assigned to a single novel species of the genus Candida, for which the name Candida thaimueangensis sp. nov. is proposed. The type strain is TM1-01T (=CBS 10360T=NBRC 101967T=BCC 21229T).

A Gram-positive, rod-shaped, spore-forming and moderately halophilic bacterium (strain 8-1T) was isolated from a sediment sample of a neutral salt lake in Xin-Jiang, China. The strain grew optimally at 30 °C, pH 7.0–7.5 and 8–12 % (w/v) NaCl. The major cellular fatty acids were anteiso-C15 : 0, iso-C15 : 0 and anteiso-C17 : 0. The predominant isoprenoid quinone was MK-7 and the cell-wall peptidoglycan contained meso-diaminopimelic acid. Phosphatidylglycerol, diphosphatidylglycerol and two unidentified glycolipids were found to be the major polar lipid components. The genomic DNA G+C content was 43.4 mol%. Phylogenetic analysis based on the 16S rRNA gene sequence revealed that strain 8-1T was closely related to Lentibacillus salarius KCTC 3911T (98.0 % sequence similarity) and other recognized species within the genus Lentibacillus (94.5–95.9 %). The level of DNA–DNA relatedness between strain 8-1T and L. salarius KCTC 3911T was 40 %. Based on the phenotypic and genotypic data presented, strain 8-1T is considered to represent a novel species of the genus Lentibacillus, for which the name Lentibacillus halodurans sp. nov. is proposed. The type strain is 8-1T (=CGMCC 1.3702T=DSM 18342T).

An anaerobic, mesophilic, syntrophic fatty-acid-oxidizing bacterium, designated strain OL-4T, was isolated as a co-culture with Methanobacterium formicicum DSM 1535NT from an anaerobic expanded granular sludge bed reactor used to treat an oleate-based effluent. Strain OL-4T degraded oleate, a mono-unsaturated fatty acid, and straight-chain fatty acids C4 : 0–C18 : 0 in syntrophic association with Methanobacterium formicicum DSM 1535NT. Even-numbered fatty acids were degraded to acetate and methane whereas odd-numbered fatty acids were degraded to acetate, propionate and methane. Branched-chain fatty acids were not degraded. The bacterium could not grow axenically with any other substrate tested and therefore is considered to be obligately syntrophic. Fumarate, sulfate, thiosulfate, sulfur and nitrate could not serve as electron acceptors for strain OL-4T to degrade oleate or butyrate. Cells of strain OL-4T were curved rods, formed spores and showed a variable response to Gram staining. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain OL-4T was most closely related to the fatty-acid-oxidizing, syntrophic bacterium Syntrophomonas sp. TB-6 (95 % similarity), Syntrophomonas wolfei subsp. wolfei DSM 2245T (94 % similarity) and Syntrophomonas erecta DSM 16215T (93 % similarity). In addition to this moderate similarity, phenotypic and physiological characteristics, such as obligate syntrophy, spore formation and utilization of a broader substrate range, differentiated strain OL-4T from these Syntrophomonas species. Therefore strain OL-4T represents a novel species, for which the name Syntrophomonas zehnderi sp. nov. is proposed. The type strain is OL-4T (=DSM 17840T=JCM 13948T).

A novel, moderately halophilic, Gram-positive coccus, designated strain S2R53-5T, was isolated from jeotgal, a traditional Korean fermented seafood. The organism was strictly aerobic, non-motile, non-sporulating and catalase- and oxidase-positive. Strain S2R53-5T grew in the presence of 0.5–15 % (w/v) NaCl and at pH 6.5–11.0, with optimum growth at 5 % (w/v) NaCl and pH 7.0. The temperature range for growth was 20.0–30.0 °C, with an optimum temperature of 30 °C. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain S2R53-5T belongs to the family Staphylococcaceae and was most closely related to Salinicoccus roseus DSM 5351T (96.8 % gene sequence similarity), Salinicoccus hispanicus DSM 5352T (96.1 %), Salinicoccus alkaliphilus T8T (95.2 %) and Jeotgalicoccus halotolerans YKJ-101T (95.1 %). The genomic DNA G+C content was 47.0 mol%, which is in the range of 46–51 mol% that is characteristic for the genus Salinicoccus. Levels of DNA–DNA relatedness between strain S2R53-5T and S. roseus DSM 5351T, S. hispanicus DSM 5352T and S. alkaliphilus KCTC 13928T were 32.2, 15.4 and 4.6 %, respectively. Chemotaxonomic data (major menaquinone, MK-6; major fatty acids, iso-C15 : 0 and anteiso-C15 : 0; cell-wall murein type, Lys and Gly) and 16S rRNA gene sequence analysis supported the affiliation of strain S2R53-5T with the genus Salinicoccus. The combined evidence from the low DNA–DNA relatedness, physiological, biochemical and other genotypic data indicate that strain S2R53-5T clearly represents a novel species of the genus Salinicoccus, for which the name Salinicoccus jeotgali sp. nov. is proposed. The type strain is S2R53-5T (=KCTC 13030T=LMG 23640T).

Multilocus sequence analysis (MLSA) was performed on representatives of Ensifer (including species previously assigned to the genus Sinorhizobium) and related taxa. Neighbour-joining (NJ), maximum-parsimony (MP) and maximum-likelihood (ML) phylogenies of dnaK, gltA, glnA, recA, thrC and 16S rRNA genes were compared. The data confirm that the potential for discrimination of Ensifer species is greater using MLSA of housekeeping genes than 16S rRNA genes. In incongruence-length difference tests, the 16S rRNA gene was found to be significantly incongruent with the other genes, indicating that this gene should not be used as a single indicator of relatedness in this group. Significant congruence was detected for dnaK, glnA and thrC. Analyses of concatenated sequences of dnaK, glnA and thrC genes yielded very similar NJ, MP and ML trees, with high bootstrap support. In addition, analysis of a concatenation of all six genes essentially produced the same result, levelling out potentially conflicting phylogenetic signals. This new evidence supports the proposal to unite Ensifer and Sinorhizobium in a single genus. Support for an alternative solution preserving the two genera is less strong. In view of the opinions expressed by the Judicial Commission, the name of the genus should be Ensifer, as proposed by Young [ Young, J. M. (2003) . Int J Syst Evol Microbiol 53, 2107–2110]. Data obtained previously and these new data indicate that Ensifer adhaerens and ‘Sinorhizobium morelense’ are not heterotypic synonyms, but represent separate species. However, transfer to the genus Ensifer is not possible at present because the species name is the subject of a pending Request for an Opinion, which would affect whether a novel species in the genus Ensifer or a new combination based on a basonym would be created.