DNA Hybridization Experiments Research Articles

Phenylobacterium glaciei sp. nov., isolated from Vestrebroggerbreen, a valley glacier in Svalbard, Arctic.

A Gram-stain-negative, rod-shaped bacterial strain designated as 20VBR1T was isolated from a valley glacier (Vestrebroggerbreen) snout ice sample from Ny-Ålesund, Svalbard, Arctic. The colonies were smooth, circular and light creamish on half-strength R2A agar and grew at 10-35 °C (optimum, 20 °C), at pH 6.5-8.0 (optimum, 7.0) and with 0-2.5 % (w/v) NaCl (optimum, 0.5 %). 16S rRNA gene sequence analysis revealed that strain 20VBR1T belonged to the genus Phenylobacterium and was most closely affiliated to Phenylobacterium aquaticum W2-3-4T (97.65 % similarity), Phenylobacterium haematophilum LMG 11050T (97.57 %) and Phenylobacterium koreense Slu-01T (96.91 %). 20VBR1T has a genome size of 4.24 Mb, comprising 4185 predicted genes with a DNA G+C content of 67.86 mol%. DNA-DNA hybridization experiments indicated that the DNA-DNA relatedness between strain 20VBR1T and P. aquaticum KACC 18306T was 41.95±4.36 %, well below the threshold (<70 %) to delineate bacterial species. Genome relatedness indexes revealed that the average nucleotide identity and digital DNA-DNA hybridization values between 20VBR1T and its closest phylogenomic relative, P. aquaticum KACC 18306T, were 78.97 and 22.10 %, respectively. The predominant isoprenoid quinone was ubiquinone (Q-10) and the major polar lipids were phosphatidylglycerol, one unknown phospholipid, one unknown glycolipid and four unidentified polar lipids. The major fatty acids (>10 %) of strain 20VBR1T were summed feature 8 (comprising C18 : 1 ω7c and/or C18 : 1 ω6c), summed feature 3 (comprising C16 : 1 ω6c and/or C16 : 1 ω7c) and C16 : 0. Based on the physiological, biochemical, chemotaxonomic, phylogenetic and phylogenomic analyses, isolate 20VBR1T is considered to represent a novel species of the genus Phenylobacterium, for which the name Phenylobacterium glaciei sp. nov. is proposed. The type strain is 20VBR1T (=JCM 33227 T=DSM 111428 T=MCC 4220 T).

DNA hybridisation kinetics using single-molecule fluorescence imaging.

Deoxyribonucleic acid (DNA) hybridisation plays a key role in many biological processes and nucleic acid biotechnologies, yet surprisingly there are many aspects about the process which are still unknown. Prior to the invention of single-molecule microscopy, DNA hybridisation experiments were conducted at the ensemble level, and thus it was impossible to directly observe individual hybridisation events and understand fully the kinetics of DNA hybridisation. In this mini-review, recent single-molecule fluorescence-based studies of DNA hybridisation are discussed, particularly for short nucleic acids, to gain more insight into the kinetics of DNA hybridisation. As well as looking at single-molecule studies of intrinsic and extrinsic factors affecting DNA hybridisation kinetics, the influence of the methods used to detect hybridisation of single DNAs is considered. Understanding the kinetics of DNA hybridisation not only gives insight into an important biological process but also allows for further advancements in the growing field of nucleic acid biotechnology.

Streptomyces smaragdinus sp. nov., isolated from the gut of the fungus growing-termite Macrotermes natalensis.

The taxonomic position of a novel aerobic, Gram-positive actinobacteria, designated strain RB5T, was determined using a polyphasic approach. The strain, isolated from the gut of the fungus-farming termite Macrotermes natalensis, showed morphological, physiological and chemotaxonomic properties typical of the genus Streptomyces . Based on 16S rRNA gene sequence analysis, the closest phylogenetic neighbour of RB5T was Streptomyces polyrhachis DSM 42102T (98.87 %). DNA–DNA hybridization experiments between strain RB5T and S. polyrhachis DSM 42102T resulted in a value of 27.4 % (26.8 %). The cell wall of strain RB5T contained ll-diaminopimelic acid as the diagnostic amino acid. Mycolic acids and diagnostic sugars in whole-cell hydrolysates were not detected. The strain produced the following major phospholipids: diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol, phosphatidylinositol-mannoside and phosphatidylserine. The menaquinone profile showed hexa- and octahydrogenated menaquinones containing nine isoprene units [MK-9(H6) and MK-9(H8)]. The strain exhibited a fatty acid profile containing the following major fatty acids: 12-methyltridecanoic acid (iso-C14 : 0) 12-methyltetradecanoic acid (anteiso-C15 : 0), 13-methyltetradecanoic acid (iso-C15 : 0) and 14-methylpentadecanoic acid (iso-C16 : 0). Here, we propose a novel species of the genus Streptomyces – Streptomyces smaragdinus with the type strain RB5T (=VKM Ac-2839T=NRRL B65539T).

Actinomadura roseirufa sp. nov., producer of semduramicin, a polyether ionophore.

The taxonomic position of 'Actinomadura roseorufa' LMG 30035T, a semduramicin-producing mutant of strain ATCC 53666P, which was isolated from a soil sample collected in Yamae Village, Kamamoto, Japan, was clarified in the present study using a polyphasic approach. This Gram-positive, aerobic actinomycete formed a well-developed, extensively branched, non-fragmenting substrate and aerial mycelia which differentiated into single, smooth-appearing spores. Based on analysis of nearly complete 16S rRNA gene sequence, strain LMG 30035T was found to be closely related to the type strains of Actinomadura fibrosa ATCC 49459T (98.88 %) and Actinomadura formosensis JCM 7474T (98.82 %) (pairwise similarity values in parentheses). Digital DNA-DNA hybridisation experiments revealed unambiguously that strain LMG 30035T represents a novel Actinomadura species (OrthoANIu values less than 83.1 %; dDDH values less than 27.2 % with type strains of validly named Actinomadura species). Analysis of the cell wall revealed the presence of meso-diaminopimelic acid in the peptidoglycan. The whole-cell sugars were glucose, madurose, galactose, ribose and rhamnose. The major polar lipids included phosphatidylinositol and diphosphatidylglycerol. The predominant menaquinones were MK-9(H6), MK-9(H8), MK-9(H4) and MK-9(H2). The major fatty acids were C16 : 00, 10-methyl C18 : 0, C18 : 1 ω9c and C18 : 00. The DNA G+C content of its genome was 72.5 mol%. In summary, these characteristics distinguish strain LMG 30035T from validly named species of the genus Actinomadura, and therefore, we propose to classify this strain formally as the novel species Actinomadura roseirufa sp. nov. with LMG 30035T (=CECT 9808T,=ATCC 53664T) as the type strain.

Lutibaculum pontilimi sp. nov., isolated from a tidal mudflat and emended description of the genus Lutibaculum.

A novel Gram-reaction-negative bacterium, designated strain GH1-34T, was isolated from a sample of tidal mudflat collected at the seashore of Gangwha Island, Republic of Korea. Cells of the bacterium were strictly aerobic, catalase- and oxidase-positive, motile by means of a polar flagellum and rod shaped. It was found to grow at 0-5 % (w/v) NaCl, 20-45 °C and pH 6-10. The major isoprenoid quinone was Q-10. The polar lipids were phosphatidylcholine, phosphatidylethanolamine, phosphatidylglycerol, an unidentified aminolipid and an unidentified glycolipid. The predominant fatty acids were C18 : 1ω7c and C19 : 0cyclo ω8c. The G+C content of the DNA was 70.9 mol%. Comparative 16S rRNA gene sequence analysis revealed that strain GH1-34T formed a tight cluster with the type strain of Lutibaculum baratangense with 98.3 % sequence similarity; levels of the 16S rRNA gene sequence similarity between the novel strain and other representatives of the order 'Rhizobiales' were <95.0 %. DNA-DNA relatedness between the organism and L. baratangense KCTC 22669T was 33 %, Based on the results of phenotypic analysis and DNA-DNA hybridization experiments, strain GH1-34T (=KCTC 52847T=NBRC 113277T) represents a novel species of the genus Lutibaculum, for which the name Lutibaculum pontilimi sp. nov. is proposed.

Bradyrhizobium ripae sp. nov., a nitrogen-fixing symbiont isolated from nodules of wild legumes in Namibia.

Root-nodule bacteria were isolated from wild legumes growing in the Kavango region, Namibia. Using a polyphasic approach, four strains belonging to the genus Bradyrhizobium (WR4T, WR87, T10 and T12) were further characterized to clarify the taxonomic status of this group. On the basis of 16S rRNA gene sequences, the four strains showed highest similarity to Bradyrhizobium elkanii USDA 76T (99.9 %), Bradyrhizobium pachyrhizi PAC48T (identical) and to Bradyrhizobiumbrasilense UFLA03-321T (identical). Multilocus sequence analysis of concatenated glnII-recA-gyrB-dnaK-rpoB sequences and comparison of the intergenic transcribed spacer (ITS) sequences confirmed that the novel group belongs to a distinct lineage of the genus Bradyrhizobium, with <96.7 % (MLSA) and 97.25 % (ITS) nucleotide identity with B. elkanii USDA 76T. Results from the sequence-based analysis were validated by DNA-DNA hybridization experiments and suggested a novel species. Several phenotypic features including carbon compound utilization and growth characteristics supported the phylogenetic data, thus it is concluded that the strains represent a novel species, for which the name Bradyrhizobium ripae sp. nov. is proposed, with type strain WR4T [LMG 30283, DSM 105795, NTCCM 0019 (Windhoek)].

Hymenobacter rufus sp. nov., a bacterium isolated from soil.

A bacterial strain, S1-2-2-6T, was isolated from a soil sample collected in Jeollabuk-do province, Republic of Korea. Cells of this strain were observed to be Gram-stain-negative, short and rod-shaped, and colonies were red to pink in colour. Analysis of 16S rRNA gene sequences identified this strain as representing a member of the genus Hymenobacter in the family Cytophagaceae, with the highest levels of sequence similarity being observed in relation to Hymenobacter terrae DG7AT (98.2 %), Hymenobacter rubidus DG7BT (97.9 %), Hymenobacter soli PB17T (97.7 %), and Hymenobacter daeguensis 16F3Y-2T (97.3 %). Growth of S1-2-2-6T was observed at 4-30 °C, pH 6-8 and in the presence of 0-0.5 % NaCl. The predominant respiratory quinone of this strain was menaquinone-7, the major fatty acids were C15 : 0 iso, C15 : 0 anteiso, and Summed feature 3 (C16 : 1ω7c/C16 : 1ω6c), and the major polar lipid was phosphatidylethanolamine. The genomic DNA G+C content of S1-2-2-6T was 60.7 mol%. DNA-DNA hybridization experiments with H. terrae, H. rubidus, H. soli and H. daeguensisresulted in relatedness values of 35.9 and 38.4 %, 34.2 and 30.4 %, 28.3 and 33.1 %, and 23.5 and 27.9 %, respectively. These DNA-DNA hybridization results, in addition to some differentiating phenotypic properties, clearly indicate that S1-2-2-6T is a representative of a novel species of the genus Hymenobacter, for which the name Hymenobacter rufus sp. nov. is proposed. The type strain is S1-2-2-6T (=KCTC 52736T=JCM 32196T).

Lactobacillus terrae sp. nov., a novel species isolated from soil samples in the Republic of Korea.

A novel strain, designated NIBRBAC000499792T, was isolated from a soil sample collected at Jukgye, Dongnam, Cheonan, Republic of Korea. Cells were Gram-positive, non-motile, non-spore-forming, rod-shaped, oxidase-negative and catalase-negative. Colonies grown on de Man, Rogosa and Sharpe agar were white, circular, raised and entire. Analysis of the 16S rRNA gene sequence analysis revealed that strain NIBRBAC000499792T belongs to the genus Lactobacillus (family Lactobacillaceae) and is most closely related to Lactobacillus nodensis DSM 19682T (96.1 % similarity) and Lactobacillus tucceti KCTC 21005T (96.7 %). The results of DNA-DNA hybridization experiments demonstrated that strain NIBRBAC000499792T represents a novel species. Major fatty acids are C18 : 1ω9c, C16 : 0 and unidentified 18.846 and/or C19 : 1ω6c and/or C19 : 0cyclo. The predominant respiratory quinones are menaquinone-8 and menaquinone-9. The major polar lipids are phosphatidylglycerol and diphosphatidylglycerol. The minor polar lipids are one unidentified aminophospholipid, one unidentified phospholipid, and four unidentified lipids. Next-generation sequencing analysis of strain NIBRBAC000499792T indicated that the total genome size was 1 548 794 bp with a G+C content of 33.1 mol%, 1586 coding sequences, 50 tRNAs and nine rRNAs. The most closely related genomes belonged to Lactobacillus species. Most metabolic pathways were related to carbon metabolism and carbon fixation. Based on this polyphasic analysis, strain NIBRBAC000499792T represents a novel species of the genus Lactobacillus, for which the name Lactobacillus terrae sp. nov. is proposed, with the type strain NIBRBAC000499792T (=KCTC 21093T=JCM 32269T).

Sphaerisporangium rhizosphaerae sp. nov., an actinomycete isolated from the rhizosphere soil of a rubber tree (Hevea brasiliensis Muell. Arg).

A novel actinomycete, designated strain NEAU-mq3T, was isolated from the rhizosphere soil of a rubber tree (Hevea brasiliensis Muell. Arg) collected from Xianglu Mountain in Heilongjiang Province, north-east China, and characterized by using a polyphasic approach. Phylogenetic analysis based on the 16S rRNA gene sequence indicated that the organism should be assigned to the genus Sphaerisporangium and that it forms a monophyletic clade with its closest relatives 'Sphaerisporangium dianthi' NEAU-CY18T (99.2 % 16S rRNA gene sequence similarity) and Sphaerisporangium cinnabarinum JCM 3291T (98.8 %). Morphological and chemotaxonomic properties of strain NEAU-mq3T were also consistent with the description of the genus Sphaerisporangium. The whole-cell sugars were madurose, mannose, ribose and glucose. The menaquinones were MK-9(H2), MK-9(H4), MK-9(H0) and MK-9(H6). The diagnostic diamino acid of the peptidoglycan was meso-diaminopimelic acid. The phospholipid profile consisted of diphosphatidylglycerol, phosphatidylmethylethanolamine, phosphatidylethanolamine, phosphatidylinositol mannoside, an unidentified polar lipid and an unidentified phospholipid. The major fatty acids were identified as iso-C16 : 0, 10-methyl C17 : 0, C16 : 1ω7c and C17 : 1ω7c. DNA-DNA hybridization experiments and phenotypic tests were carried out between strain NEAU-mq3T and its most closely related strains, which further clarified their relatedness and demonstrated that NEAU-mq3T could be distinguished from these strains. Therefore, it is concluded that strain NEAU-mq3T represents a novel species of the genus Sphaerisporangium, for which the name Sphaerisporangium rhizosphaerae sp. nov. is proposed. The type strain is NEAU-mq3T (=CGMCC 4.7429T=JCM 32389T).

Exonuclease I-Hydrolysis Assisted Electrochemical Quantitation of Surface-Immobilized DNA Hairpins and Improved HIV-1 Gene Detection.

The complete formation of stem-loop (i.e., hairpin) configuration on chip surface is of particular importance for the application of hairpin DNA (hpDNA) in building biosensors for various analytes with optimized performance. We report herein a convenient electrochemical protocol for evaluating the yield of hairpin DNA conformations upon self-assembly on electrode surface. As of the different hydrolysis capability of Exonuclease I (Exo I) toward single-stranded DNA (ssDNA) and hpDNA, we can selectively remove ssDNA from electrode but retain hpDNA strands; based on the changes in the cyclic voltammetric (CV) responses using [Ru(NH3)6]3+ as redox indicators, we can then determine the fraction of hairpin configurations in mixed DNA self-assembled monolayers (SAMs). It was discovered that the molar fraction of hairpin configuration formed on the surface is considerably lower than that in the binary deposition solution (containing both ssDNA and hpDNA). The accuracy of the Exo I-assisted electrochemical quantitative protocol has been validated by standard DNA hybridization experiments; the relationship between the overall DNA packing density and the yield of hairpin configurations was also evaluated. More importantly, taking HIV-1 gene detection as a trial system, the hpDNA-based biosensor shows significantly improved detection limit and broadened response range upon the background reduction by Exo I-catalyzed hydrolysis.

Rathayibacter oskolensis sp. nov., a novel actinobacterium from Androsace koso-poljanskii Ovcz. (Primulaceae) endemic to the Central Russian Upland.

A rod-shaped, non-endospore-forming and non-motile bacterium, strain DL-329T, was isolated from the above-ground part of a plant, Androsace koso-poljanskii Ovcz. (Primulaceae), at the the State Natural Reserve 'Belogorie', Russia. On the basis of 16S rRNA gene sequence comparisons, the strain clustered with members of the genus Rathayibacter, showing the highest sequence similarity to Rathayibacter tritici (98.89 %), Rathayibacter rathayi (98.82 %) and Rathayibacter festucae (98.82 %). The DNA hybridization experiments demonstrated that strain DL-329T represents a separate genomic species. The results of comparative studies of physiological and chemotaxonomic characteristics, including cell-wall sugar patterns, polar lipid profiles, and the matrix-assisted laser desorption/ionization time-of-flight mass spectra of bacterial cells, allowed clear differentiation of VKM Ac-2121T from the recognized Rathayibacter species at the phenotypic level. Based on the data obtained, a new species, Rathayibacter oskolensis sp. nov., is proposed, with DL-329T (=VKM Ac-2121T=LMG 22542T) as the type strain.

Bacillus fermenti sp. nov., an indigo-reducing obligate alkaliphile isolated from indigo fermentation liquor for dyeing

The indigo-reducing, facultatively anaerobic and obligately alkaliphilic strains Bf-1T, Bf-2 and Bf-4 were isolated from an indigo fermentation liquor used for dyeing, which uses sukumo [composted Polygonum indigo (Polygonum tinctorium Lour.) leaves] as a basic ingredient and was obtained from a craft centre in Date City, Hokkaido, Japan. The 16S rRNA gene sequence analyses indicated that the closest neighbours of strain Bf-1T are Bacillus maritimus DSM 100413T (98.3 % 16S rRNA gene sequence similarity), Bacillus persicus DSM 25386T (98.2 %) and Bacillus rigiliprofundi LMG 28275T (97.7 %). The 16S rRNA gene sequence of strain Bf-1T was almost identical to the sequences of strains Bf-2 and Bf-4 (99.9 %). Cells of strain Bf-1T stained Gram-positive and formed straight rods that achieved motility through a pair of subpolar flagella. Strain Bf-1T grew at temperatures of between 15 and 45 °C with optimum growth at 33‒40 °C. The strain grew in the pH range of pH 8‒12, with optimum growth at pH 10. The isoprenoid quinone detected was menaquinone-7 (MK-7), and the DNA G+C content was 41.7 %. The whole-cell fatty acid profile mainly (>10 %) consisted of iso-C15 : 0 and iso-C16 : 0. Phylogenetically related neighbours, although demonstrating high 16S rRNA gene sequence similarity (>97.6 %) with strain Bf-1T, exhibited less than 9 % relatedness in DNA-DNA hybridization experiments. Based on evidence from this polyphasic study, the isolates represent a novel species, for which the name Bacillus fermenti sp. nov. is proposed. The type strain of this species is Bf-1T (=JCM 31807T=NCIMB 15079T).

Streptomyces capitiformicae sp. nov., a novel actinomycete producing angucyclinone antibiotics isolated from the head of Camponotus japonicus Mayr.

A novel actinomycete, designated strain 1H-SSA4T, was isolated from the head of an ant (Camponotus japonicus Mayr) and was found to produce angucyclinone antibiotics. A polyphasic approach was used to determine the taxonomic status of strain 1H-SSA4T. The DNA G+C content of the draft genome sequence, consisting of 11.4 Mbp, was 70.0 mol%. 16S rRNA gene sequence similarity studies showed that strain 1H-SSA4T belongs to the genus Streptomyces with the highest sequence similarity to Streptomyces hygroscopicus subsp. ossamyceticus NBRC 13983T (98.9 %), and phylogenetically clustered with this species, Streptomyces torulosus LMG 20305T (98.8 %), Streptomyces ipomoeae NBRC 13050T (98.5 %) and Streptomyces decoyicus NRRL 2666T (98.4 %). The morphological and chemotaxonomic properties of the strain were also consistent with those members of the genus Streptomyces. A combination of DNA-DNA hybridization experiments and phenotypic tests were carried out between strain 1H-SSA4T and the above-mentioned strains, which further clarified their relatedness and demonstrated that strain 1H-SSA4T could be distinguished from these strains. Therefore, the strain is concluded to represent a novel species of the genus Streptomyces, for which the name Streptomyces capitiformicae sp. nov. is proposed. The type strain is 1H-SSA4T (=CGMCC 4.7403T=DSM 104537T).

Brenneria populi subsp. brevivirga subsp. nov. isolated from symptomatic bark of Populus × euramericana canker, and description of Brenneria populi subsp. populi subsp. nov.

Two Gram-stain-negative, facultatively anaerobic, motile bacterial strains isolated from symptomatic bark of Populus × euramericana canker in China were investigated using a polyphasic approach, including 16S rRNA gene sequencing, multilocus sequence analysis, and biochemical and physiological assays. 16S rRNA gene sequencing indicated that these strains belonged to the genus Brenneria, family Pectobacteriaceae, and had the highest sequence similarity with Brenneria populi CFCC 11963T (98 %). DNA-DNA hybridization experiments revealed DNA-DNA relatedness values of 72.1-78.2 % between the new isolates and strains of B. populi, revealing that these strains belonged to the same species. The 16S rRNA gene sequencing and multilocus sequence analysis suggested that the two novel strains and those of B. populi are phylogenetically closely related but form two clearly separated subgroups. Based on the data, the two novel isolates represent a subspecies of B. populi for which the name B. populi subsp. brevivirga subsp. nov. is proposed with D8-10-4-5T (=CFCC 11935T=KCTC 42841T) as the type strain, with the automatic creation of B. populi subsp. populi subsp. nov. (type strain D9-5T=CFCC 11963T=KCTC 42088T).

Herpetosiphon gulosus sp. nov., a filamentous predatory bacterium isolated from sandy soil and Herpetosiphon giganteus sp. nov., nom. rev.

Three filamentous gliding bacteria from the German Collection of Microorganisms and Cell Cultures, Hp g11, Hp g471 and Hp g472, were subjected to a phylogenetic analysis. These organisms had previously been classified as members of the genus Herpetosiphon based on their growth physiology and morphology. However, a taxonomic assignment at the species level had not been carried out. Analysis of 16S rRNA sequences now confirmed the close relationship of strain Hp g472 to Herpetosiphon aurantiacus DSM 785T (98.6 % nucleotide identity) and Herpetosiphon geysericola DSM 7119T (97.7 %). The results of DNA-DNA hybridization experiments further implied that strain Hp g472 should be classified as a distinct species. The DNA G+C content of strain Hp g472 was 49.9 mol%. The major quinone was MK-10 and the predominant cellular fatty acids were C18 : 1, C16 : 1 and C16 : 0. Based on phenotypic, chemotaxonomic and phylogenetic data it was concluded that strain Hp g472 represents a novel species of the genus Herpetosiphon, for which the name Herpetosiphon gulosus sp. nov. is proposed. The type strain is Hp g472T (=DSM 52871T=NBRC 112829T). In contrast to Hp g472T, the strains Hp g11 and Hp g471 exhibited closest 16S rRNA gene sequence similarity (>99 %) with 'Herpetosiphon giganteus' Hp a2. The distinctive genotypic and phenotypic properties of the latter supported the revival of the name as Herpetosiphon giganteus (ex Reichenbach & Golecki, 1975) sp. nov., nom. rev. We propose the previously deposited reference strain DSM 589T=NBRC 112828T as the type strain.

Flaviaesturariibacter luteus sp. nov., isolated from an agricultural floodplain soil, and emended description of the genus Flaviaesturariibacter.

A novel slow-growing bacterial strain designated as AW305T was isolated from an agricultural floodplain soil located in Mashare, Kavango region, Namibia. Cells stained Gram-negative, were non-motile, non-spore-forming, coccoid to rod-shaped and did not form a capsule. Colonies were yellow-pigmented, but flexirubin-type pigments were not detected. AW305T had an aerobic chemo-organoheterotrophic metabolism, using a narrow spectrum of carbon sources for growth, with preference for complex protein substrates, organic acids and amino acids. AW305T was able to grow at 15-40 °C, pH 5.3-8.3 and in the presence of up to 0.25 % (w/v) NaCl. 16S rRNA gene sequence comparison showed that AW305T belonged to the genus Flaviaesturariibacter (family Chitinophagaceae). Its closest relatives were Flaviaesturariibacter amylovorans GCR0105T (97.0 %), Flavisolibacter ginsengiterrae Gsoil 492T (93.6 %) and Flavisolibacter ginsengisoli Gsoil 643T (93.2 %). DNA-DNA hybridization experiments corroborated that AW305T represents an independent genomospecies. The genomic DNA G+C content was 57.6 mol%. Major fatty acids were iso-C15 : 1 G, iso-C15 : 0, C16 : 1ω5c, iso-C17 : 0 3-OH and summed feature 3 (C16 : 1ω7c and/or iso-C15 : 0 2-OH). The predominant respiratory quinone was MK-7, albeit minor amounts of MK-6 were also detected. The polar lipids comprised major amounts of phosphatidylethanolamine and minor amounts of two unidentified lipids, an unidentified phospholipid, an unidentified glycolipid and an unidentified aminoglycophospholipid. On the basis of the polyphasic characterization, strain AW305T represents a novel species of the genus Flaviaesturariibacter for which the name Flaviaesturariibacter luteus sp. nov. is proposed, with the type strain AW305T (=DSM 100282T=LMG 29416T).

Hymenobacter aquaticus sp. nov., a radiation-resistant bacterium isolated from a river.

A Gram-stain-negative, non-motile, non-spore-forming, rod-shaped, aerobic bacterial strain, designated 16F3PT, was isolated from the Han River, South Korea, and characterized taxonomically using a polyphasic approach. Phylogenetic analysis based on 16S rRNA gene sequences revealed 16F3PT to be within the genus Hymenobacter, and most closely related to Hymenobacterchitinivorans Txc1T (98.62 %) and Hymenobacterelongatus VUG-A112T (98.46 %). The phylogenetic distance from other species of the genus Hymenobacter with validly published names was greater than 4 % (i.e. sequence similarity was less than 96.0 %). Chemotaxonomic data also supported the classification of strain 16F3PT within the genus Hymenobacter. C16 : 0 (19.8 %), summed feature 3 (C16 : 1ω7c/C16 : 1ω6c; 15.4 %) and iso-C15 : 0 (13.0 %) were the major fatty acids, MK-7 was the predominant respiratory quinone, and phosphatidylethanolamine was the major polar lipid. The G+C content of the genomic DNA of strain 16F3PT was 61.9 mol%. DNA-DNA hybridization experiments showed that the values for DNA-DNA relatedness between strain 16F3PT and the phylogenetically closest neighbours were below 19 %. On the basis of its phenotypic properties and phylogenetic distinctiveness, strain 16F3PT represents a novel species of the genus Hymenobacter, for which the name Hymenobacter aquaticus sp. nov. is proposed. The type strain is 16F3PT (=KCTC 52194T=JCM 31653T).

Aquaspirillum soli sp. nov., isolated from a soil sample.

A Gram-stain-negative, smooth, opaque, white-pigmented, helical-shaped, catalase- and oxidase-positive bacterium that was motile by means of bipolar tufts of flagella and grew under microaerophilic conditions, was isolated from a soil sample of a reed pond in Shangqiu, Henan province, PR China. The strain, designated THG-SQE6T, grows well at 25-42 °C, pH 6.5-7.5 and in the presence of 0-1 % (w/v) NaCl. hylogenetic analysis based on 16S rRNA gene sequences showed that strain THG-SQE6T was most closely related to Aquaspirillum serpens IAM 13944T (97.23 % 16S rRNA gene sequence similarity). The DNA G+C content of strain THG-SQE6T was 53.10 mol%. In DNA-DNA hybridization experiments, the DNA-DNA relatedness between strain THG-SQE6T and its closest phylogenetic neighbour was below 62.6 %. The predominant isoprenoid quinone detected in strain THG-SQE6T was ubiquinone-8 (Q-8). The major polar lipids were phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylglycerol, phosphatidylserine, an unidentified phospholipid, an unidentified aminolipid and two unidentified lipids. The major fatty acids were identified as C12 : 0 3-OH, C16 : 0, C18 : 0, summed feature 3 and summed feature 8. These data support the affiliation of strain THG-SQE6T to the genus Aquaspirillum. Based on findings from the phenotypic, genotypic and phylogenetic characterization of strain THG-SQE6T, a novel species of the genus AquaspirillumnamedAquaspirillum soli sp. nov. is proposed. The type strain is THG-SQE6T (=KACC 18846T=CCTCC AB 2016081T).

Krasilnikoviella muralis gen. nov., sp. nov., a member of the family Promicromonosporaceae, isolated from the Takamatsuzuka Tumulus stone chamber interior and reclassification of Promicromonospora flava as Krasilnikoviella flava comb. nov.

A Gram-stain-positive, facultatively anaerobic actinomycete, designated strain T6220-5-2bT, was isolated from a sample taken from a mouldy spot on the surface of a mural painting (the white tiger, Byakko) inside the stone chamber of Takamatsuzuka Tumulus in Asuka village, Nara Prefecture, Japan. Based on 16S rRNA gene sequence analysis of the isolate, it was closely related to the genus Promicromonospora, but formed of a novel lineage within the family Promicromonosporaceae. The closest related species to strain T6220-5-2bT was Promicromonospora flava, with which it shared 99.1 % 16S rRNA gene sequence similarity. The isoprenoid quinone systems were menaquinones MK-9(H2), MK-9(H0) and MK-9(H4). The predominant cellular fatty acids for the isolate were anteiso-C15 : 0 and iso-C15 : 0. The peptidoglycan contained glutamic acid, aspartic acid, alanine and lysine, with the last named being the diagnostic diamino acid. The cell-wall acyl type was acetyl. The major polar lipids of the isolate were phosphatidylglycerol, diphosphatidylglycerol, phosphatidylinositol, phosphatidylinositolmannoside, two unknown phospholipids and an unknown phosphoglycolipid. Whole-cell sugars of the isolate were galactose, glucose and ribose. The DNA G+C content of the genomic DNA was 75.2 mol%. Based on the results of phylogenetic, physiological and biochemical analyses and DNA-DNA hybridization experiments, the isolate was considered to represent a novel species of a new genus in the family Promicromonosporaceae, for which the name Krasilnikoviella muralis gen. nov., sp. nov. is proposed. The type strain of Krasilnikoviella muralis is T6220-5-2bT (=JCM 28789T=NCIMB 15040T). The reclassification of Promicromonospora flava as Krasilnikoviella flava comb. nov. is also proposed with the emended description of this species.

Novosphingobium guangzhouense sp. nov., with the ability to degrade 1-methylphenanthrene

A novel Gram-stain-negative, flagellated, rod-shaped, yellow-pigmented aerobic bacterium, strain SA925T, that is capable of degrading 1-methylphenanthrene was isolated from oil-polluted soil collected from a refinery located in Guangzhou, China. Phylogenetic analysis based on the 16S rRNA gene sequence demonstrated that strain SA925T belongs to the genus Novosphingobium and is evolutionarily close to the type strains of Novosphingobium gossypii (98.5 % similarity), Novosphingobium panipatense (98.2 %), Novosphingobium mathurense (98.0 %) and Novosphingobium pentaromativorans (96.5 %). The G+C content of the genomic DNA was 60.2 mol%. DNA-DNA hybridization experiments between strain SA925T and the closest strain, Novosphingobium gossypii JM-1396T, revealed a low level of relatedness (35.5 %). Strain SA925T grew at 10-35 °C, at pH 6.0-8.0 and in the presence of 0-4 % (w/v) NaCl. The major fatty acids were C18 : 1ω7c, C16 : 0 and summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c). The polar lipid profiles mainly consisted of diphosphatidylglycerol, phosphatidylglycerol, phosphatidyldimethylethanolamine, phosphatidylethanolamine and sphingoglycolipid (the characteristic polar lipid). The predominant ubiquinone was Q-10. The major polyamine was spermidine. Based on the phylogenetic, phenotypic and physiological characteristics, strain SA925T was considered to represent a novel species of the genus Novosphingobium, for which the name Novosphingobium guangzhouense sp. nov. is proposed. The type strain is SA925T (=DSM 32207T=GDMCC 1.1110T).