- Volume 59, Issue 6, 2009

The taxonomic position of a group of four strains, isolated from the phyllosphere of grasses, within the species Pseudomonas cedrina was investigated. The isolates formed a separate cluster through ribotyping and MALDI-TOF MS, which could be clearly differentiated from the type strain of P. cedrina. The differences found between the patterns of the type strain of P. cedrina and the novel isolates were more distinct than those between the type strain and recognized species of the genus Pseudomonas, which were phylogenetically related by 16S rRNA gene sequence analysis. Physiological characterization also revealed significant differences between the novel grass isolates and the type strain of P. cedrina. Siderotyping of the pyoverdines revealed identical pyoverdine-isoelectrofocusing patterns for the novel isolates and the type strain of P. cedrina. However, pyoverdine-mediated 59Fe cross uptake studies indicated differences in the siderotype. In contrast, phylogenetic analysis based on 16S rRNA gene sequence analysis and DNA–DNA hybridization studies (reassociation value 76.4 %) supported the affiliation of the novel isolates to the species P. cedrina. As a consequence of these observations, the splitting of the species P. cedrina into two novel subspecies Pseudomonas cedrina subsp. cedrina subsp. nov. (type strain CFML 96-198T=CIP 105541T=DSM 17516T) and Pseudomonas cedrina subsp. fulgida subsp. nov. (type strain P 515/12T=DSM 14938T=LMG 21467T) is proposed.

A novel bacterium (strain K4T) belonging to the genus Sphingomonas was isolated from tidal flat sediment in Korea. Its morphology, physiology, biochemical features and 16S rRNA gene sequence were characterized. Colonies of this strain are yellow in colour and the cells are rod-shaped, exhibiting negative Gram staining. The strain grows at 0–5 % (w/v) NaCl and 20–35 °C, with optimal growth occurring at 0 % (w/v) NaCl and 30 °C. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain K4T is associated with the genus Sphingomonas. Within the phylogenetic tree, this novel strain shares a branching point with Sphingomonas asaccharolytica Y-345T, with which it shares 97.3 % 16S rRNA gene sequence similarity. The polyamine pattern predominantly contains the Sphingomonas-specific triamine sym-homospermidine. Combined analysis of 16S rRNA gene sequences, DNA–DNA relatedness, physiological and biochemical test results identified genotypic and phenotypic differences between strain K4T and other Sphingomonas species. On the basis of these differentiating features, it is concluded that strain K4T (=KCTC 22050T=DSM 19475T) represents a novel species of the genus Sphingomonas, for which the name Sphingomonas aestuarii sp. nov. is proposed.

The name ‘Acinetobacter venetianus’ has been used previously to designate three marine hydrocarbon-degrading Acinetobacter strains, of which strain RAG-1 (=ATCC 31012) has industrial applications for the production of the bioemulsifier emulsan. However, to date, the name of this taxon has not been validly published. In this study, five strains were examined to corroborate the delineation of this taxon by means of phenotypic characterization, DNA–DNA hybridization, selective restriction fragment amplification (AFLP), amplified rDNA restriction analysis (ARDRA), rpoB gene sequence analysis and tRNA intergenic spacer length polymorphism analysis (tDNA-PCR) and to emend the description of ‘Acinetobacter venetianus’ (ex Di Cello et al. 1997 ). AFLP analysis showed that the five strains formed a tight cluster at 56.8±5.0 % genomic relatedness that was separated from strains of other haemolytic species of the genus Acinetobacter and from the type and reference strains of other Acinetobacter species at ≤27 % relatedness, indicating the distinctiveness of the novel strains. The strains were haemolytic and able to grow on citrate (Simmons), l-histidine and malonate. The strains did not oxidize d-glucose or utilize dl-lactate or l-aspartate. The G+C contents of strains RAG-1 and of VE-C3 were 43.9 % and 43.6 mol%, respectively. The novel strains could be recognized by a characteristic ARDRA pattern (CfoI 1, AluI 3, MboI 2, RsaI 2, MspI 3). The consensus tDNA-PCR pattern for the five strains consisted of amplified fragments of 87.9, 100.2, 134.6 and 248.5 bp and was indistinguishable from that of strains of Acinetobacter genomic species 14BJ. The five strains represent a novel species for which the name Acinetobacter venetianus sp. nov. is proposed. The type strain is RAG-1T (=ATCC 31012T=CCUG 45561T=LMG 19082T=LUH 3904T=NIPH 1925T).

A novel Gram-negative, non-sporulating, moderately halophilic, facultatively alkaliphilic, catalase- and oxidase-positive, obligately aerobic bacterium, strain YIM-Y25T, was isolated from a subterranean brine sample collected from a salt mine in Yunnan, south-west China. Cells were spirilla, motile by monopolar flagella, with meso-diaminopimelic acid in the cell-wall peptidoglycan. Growth occurred with 1–15 % (w/v) NaCl (optimum 5 %), at pH 6.0–10.0 (optimum pH 8.0) and at 15–50 °C (optimum 35–40 °C). Ubiquinone Q-8 was the predominant respiratory quinone. Polar lipids were diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylglycerol, phosphatidylmonomethylamine and an unidentified phospholipid. The major cellular fatty acids were C18 : 1 ω7c, iso-C16 : 0, C16 : 0 and C16 : 1. The genomic DNA G+C content was 58.5 mol%. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain YIM-Y25T was most closely related to the type strain of the sole recognized species of the genus Saccharospirillum, Saccharospirillum impatiens EL-105T (sequence similarity 97.0 %), and these two strains formed a robust lineage in the phylogenetic tree. The level of DNA–DNA relatedness between them was 12.6 %. The combination of phylogenetic analysis, phenotypic differences, chemotaxonomic properties and DNA–DNA hybridization data supported the view that this strain represents a novel species of the genus Saccharospirillum, for which the name Saccharospirillum salsuginis sp. nov. is proposed, with YIM-Y25T (=CCTCC AA 207033T =KCTC 22169T) as the type strain.

Three Arcobacter isolates, recovered from mussels (genus Mytilus), and one isolate from brackish water in Catalonia (north-east Spain) showed a novel pattern using a recently described identification method for members of the genus Arcobacter, 16S rRNA gene RFLP. Enterobacterial repetitive intergenic consensus PCR fingerprinting demonstrated that the three isolates from mussels belonged to two genotypes and that the fourth isolate from water belonged to a third genotype. Analysis of the 16S rRNA and rpoB gene sequences showed that the new isolates formed a separate lineage within the genus Arcobacter. This was also confirmed by the low DNA–DNA relatedness values (16–30 %) of the isolates with the type strains of recognized Arcobacter species. Hydrolysis of indoxyl acetate, a characteristic trait for all species of the genus Arcobacter, was negative for the novel isolates. The susceptibility of the novel isolates to cefoperazone, together with the lack of urease production and nitrate reduction, further enabled them to be differentiated from recognized Arcobacter species based on physiological characteristics. Genotypic and phenotypic characteristics indicated that the new isolates represent a novel species of the genus Arcobacter, for which the name Arcobacter mytili sp. nov. is proposed, with the type strain F2075T (=CECT 7386T =LMG 24559T). The DNA G+C content of strain F2075T was 26.9 mol%.

A novel, Gram-negative bacterium, designated CS5-B2T, was isolated from soil that had been collected from a cliff on Mara Island, Republic of Korea. Phylogenetic analyses based on 16S rRNA gene sequences showed that the strain fell within the radiation of the genus Dyella. The closest relatives were the type strains of Dyella koreensis, Dyella ginsengisoli and Dyella japonica and 16S rRNA gene sequence similarities between strain CS5-B2T and these strains were 99.0, 97.9 and 97.8 %, respectively. The sequence similarities between the novel isolate and other related taxa compared in the phylogenetic analysis were less than 96.7 %. The cells of strain CS5-B2T were aerobic, oxidase-negative, catalase-positive, motile rods. The temperature range for growth was 20–37 °C, with optimal growth at 30–37 °C. Growth occurred at pH 5.1–9.1, with optimal growth at pH 6.1–9.1. NaCl tolerance for growth was from 1  to 2 % (w/v). Ubiquinone-8 was the predominant respiratory lipoquinone. The major fatty acids were iso-C15 : 0 and iso-C17 : 1 ω9c. The G+C content of the DNA was 65.7–66.6 mol%. The level of DNA–DNA relatedness with D. koreensis KCTC 12359T was 20.2 and 29.6 % in duplicate measurements. On the basis of phenotypic features, phylogenetic analysis and DNA–DNA relatedness, a novel species of the genus Dyella is proposed, with the name Dyella marensis sp. nov. The type strain is CS5-B2T (=JCM 14959T =KCTC 22144T).

Nine Gram-negative, rod-shaped, non-spore-forming isolates with identical or very similar repetitive-sequence-based PCR profiles were recovered from an evaporative lagoon in Mexico. Two strains, designated 1NT and 3N, had virtually identical 16S rRNA gene sequences and, on the basis of these sequences, were identified as members of the genus Pseudomonas, with Pseudomonas peli R-20805T as the closest relative. All nine isolates had practically identical whole-cell protein profiles. The major fatty acids [C16 : 0, C18 : 1 ω7c and summed feature a (C16 : 1 ω7 and/or C16 : 1 ω6c)] of strains 1NT and 3N supported their affiliation with the genus Pseudomonas. The DNA–DNA reassociation values with respect to P. peli LMG 23201T and other closely related Pseudomonas species were <15 %. Physiological and biochemical tests allowed phenotypic differentiation of the strains analysed, including strain 1NT, from the five phylogenetically closest Pseudomonas species. On the basis of the data obtained by using this polyphasic taxonomic approach, the nine strains represent a novel species, for which the name Pseudomonas cuatrocienegasensis sp. nov. is proposed. The type strain is 1NT (=LMG 24676T=CIP 109853T).

Two luminous marine bacteria, strains LC2-065T and LC2-102, were isolated from seawater at Sagami Bay in Japan. These bacteria were Gram-negative, oxidase-negative, catalase-positive, motile and coccoid-rods. 16S rRNA gene sequence analysis and multilocus sequence analysis (MLSA) using six loci (ftsZ, gapA, gyrB, mreB, pyrH and topA) and sequence analysis of the alpha subunit of luciferase (luxA) gene revealed that these bacteria were distinct from other species of the genus Photobacterium. These novel strains were most closely related to Photobacterium kishitanii. The DNA–DNA hybridization value between strain LC2-065T and Photobacterium kishitanii ATCC BAA-1194T was 42.1 %. The major fatty acids were C12 : 0, C14 : 0, C16 : 0, C18 : 0 and C15 : 0 iso 2-OH and/or C16 : 1 ω7c (summed feature 3). The DNA G+C contents of strains LC2-065T and LC2-086 were 42.2 and 42.9 mol%, respectively. The phenotypic features of the novel strains were similar to those of P. kishitanii and P. phosphoreum, but there were sufficient physiological differences for the novel strains to be easily differentiated. On the basis of these results, these new strains represent a novel species, for which the name Photobacterium aquimaris sp. nov. is proposed. The type strain is LC2-065T (=NBRC 104633T=KCTC 22356T).

A taxonomic study was carried out on strain A-11-3T, which was isolated from an oil-enriched consortia from the surface seawater of Hong-Deng dock in the Straits of Malacca and Singapore. Cells were aerobic, Gram-negative, non-spore-forming irregular rods. The strain was catalase- and oxidase-negative. It grew on a restricted spectrum of organic compounds, including some organic acids and alkanes. 16S rRNA gene sequence comparisons showed that strain A-11-3T was most closely related to the type strains of Alcanivorax jadensis (96.8 % sequence similarity), Alcanivorax borkumensis (96.8 %), Alcanivorax dieselolei (94.8 %), Alcanivorax venustensis (94.2 %) and Alcanivorax balearicus (94.0 %). The predominant fatty acids were C16 : 0 (31.2 %), C18 : 1 ω7c (24.8 %), C18 : 0 (9.6 %), C12 : 0 (8.3 %), C16 : 1 ω7c (8.3 %) and C16 : 0 3-OH (5.1 %). The G+C content of the genomic DNA was 54.7 mol%. Moreover, the strain produced lipopeptides as its surface-active compounds. According to physiological and biochemical tests, DNA–DNA hybridization results and sequence comparisons of the 16S–23S internal transcribed spacer, the gyrB gene and the alkane hydroxylase gene alkB1, strain A-11-3T was affiliated with the genus Alcanivorax but could be readily distinguished from recognized Alcanivorax species. Therefore strain A-11-3T represents a novel species of the genus Alcanivorax for which the name Alcanivorax hongdengensis sp. nov. is proposed. The type strain is A-11-3T (=CGMCC 1.7084T=LMG 24624T=MCCC 1A01496T).

Two novel Psychrobacter-like bacteria, strains KC 40T and KC 65, were isolated from a marine crustacean specimen collected from the Sea of Japan, and were characterized by using a polyphasic approach. Strains were selected on the basis of their ability to produce black–brown diffusible pigments on commonly used organic media, which appears to be a unique characteristic of recognized members of the genus Psychrobacter. Phylogenetic analyses based on both 16S rRNA and gyrB gene sequences showed that the novel isolates formed a separate cluster within the genus Psychrobacter. Strains KC 40T and KC 65 shared highest levels of 16S rRNA gene sequence similarity with Psychrobacter urativorans DSM 14009T (98.0 %), Psychrobacter pulmonis CCUG 46240T (97.9 %), Psychrobacter cibarius JG-219T (97.9 %), Psychrobacter faecalis Iso-46T (97.8 %), Psychrobacter aquimaris SW-210T (97.6 %), Psychrobacter namhaensis SW-242T (97.6 %) and Psychrobacter nivimaris 88/2-7T (97.6 %). DNA–DNA hybridization experiments revealed 84 % DNA–DNA relatedness between strains KC 40T and KC 65 but much lower levels of relatedness (7–35 %) between the novel strains and the type strains of recognized Psychrobacter species, confirming their assignment to a single novel species of the genus Psychrobacter. The two novel strains could be distinguished from recognized species of the genus Psychrobacter based on a combination of physiological and biochemical characteristics. On the basis of phenotypic and molecular properties, strains KC 40T and KC 65 are considered to represent a novel species of the genus Psychrobacter, for which the name Psychrobacter fulvigenes sp. nov. is proposed. The type strain is KC 40T (=KMM 3954T=NRIC 0746T=JCM 15525T).

A mesophilic, aerobic, facultatively chemolithoautotrophic bacterium, designated strain EPR70T, was isolated from hydrothermal fluids from diffuse-flow vents on the East Pacific Rise at  ° 50′ N 10 ° 17′ W. Cells were Gram-negative rods, approximately 0.8–1.0 μm long and 0.3–0.5 μm wide. Strain EPR70T grew at 20–40 °C (optimum 30–35 °C), 1–25 % NaCl (optimum 2.5 %) and pH 5.0–7.5 (optimum pH 5.5). The shortest generation time observed for strain EPR70T was 42 min. Growth occurred under aerobic chemolithoautotrophic conditions in the presence of thiosulfate and CO2. Strain EPR70T grew heterotrophically with acetate or n-alkanes as sole carbon and energy sources, and in complex artificial seawater medium. Nitrate was not used as an electron acceptor. The G+C content of the genomic DNA was 64 mol%. Phylogenetic analysis of the 16S rRNA gene indicated that this organism is a member of the class Gammaproteobacteria, with Salinisphaera shabanensis E1L3AT as its closest relative (94 % sequence similarity). On the basis of phylogenetic analyses based on 16S rRNA, rbcL and alkB genes and physiological analysis, it is proposed that the organism represents a novel species within the genus Salinisphaera, for which the name Salinisphaera hydrothermalis sp. nov. is proposed. The type strain is EPR70T (=DSM 21483T =JCM 15514T).

A bacterial strain designated SBKo001T was isolated from a forest soil sample from Mt Makiling in Laguna, Philippines. It shows the general characteristics associated with myxobacteria, such as swarming of Gram-negative, rod-shaped vegetative cells, fruiting body formation and bacteriolytic activity. The strain is mesophilic, strictly aerobic and chemoheterotrophic and also exhibits resistance to various antibiotics. Major fatty acids are iso-C15 : 0, C17 : 1 2-OH and C20 : 4 (arachidonic acid). The G+C content of the genomic DNA is 69.2 mol%. A reference strain, NOSO-1 (=DSM 53757), isolated from the Etosha Basin in Namibia, shares nearly the same characteristics with SBKo001T. The identical 16S rRNA gene sequences of the two strains show 94 % identity to strains of the cellulose-degrading Byssovorax and Sorangium species. Phylogenetic analysis reveals a novel branch diverging from the Polyangiaceae, Sorangiineae, Myxococcales. Their uniqueness in morphological growth stages, unusual fatty acid profile, broad-spectrum antibiotic resistance and branch divergence from the Polyangiaceae imply that strains SBKo001T and NOSO-1 not only represent a novel genus and species, proposed here as Phaselicystis flava gen. nov., sp. nov., but also belong to a new family, Phaselicystidaceae fam. nov. The type strain of Phaselicystis flava is SBKo001T (=DSM 21295T =NCCB 100230T).

Strain 10-1-101T, isolated from a sand sample collected from the desert of Xinjiang, China, was a Gram-negative, aerobic, motile, rod-shaped bacterium. Colonies grown on 0.1× trypticase soy broth agar were circular, convex and light-pink-coloured. The major cellular fatty acids of the novel strain were C18 : 1 ω7c (72.71 %) and C16 : 0 (7.05 %). The DNA G+C content of strain 10-1-101T was 68.8 mol% and Q-10 was the major respiratory quinone. 16S rRNA gene sequence analysis indicated that strain 10-1-101T was related to type strains of the genus Skermanella, with sequence similarity values of 94.07 % with Skermanella aerolata DSM 18479T and 92.74 % with Skermanella parooensis DSM 9527T. On the basis of genotypic, phenotypic and phylogenetic data, this new strain represents a novel species of the genus Skermanella, for which the name Skermanella xinjiangensis sp. nov. is proposed. The type strain is 10-1-101T (=CCTCC AB 207153T=NRRL B-51273T).

Four strains of a novel heterothallic yeast species were isolated from pollen-storing cells of a honeycomb of honeybee (Apis mellifera) in Hungary. Analysis of the D1/D2 domain of the large-subunit (26S) rRNA gene sequences placed the strains in the Trichomonascus clade. The four strains share identical D1/D2 sequences and differ by 24 substitutions and nine indels from the genetically most closely related species, Blastobotrys attinorum. The name Trichomonascus apis sp. nov. is proposed for the novel species. The carbon-source assimilation spectrum of T. apis sp. nov. is rather broad. Unlike B. attinorum, it assimilates sucrose, trehalose, d-glucuronate and succinate and does not grow at 37 °C, thus enabling the two taxa to be distinguished. The type and isotype strains of Trichomonascus apis are NCAIM Y.01848T (=CBS 10922T =NRRL Y-48475T) and NCAIM Y.01849IT (=CBS 10923IT =NRRL Y-48476IT), respectively.