Fluorescent sensors of siderophores produced by bacterial pathogens.

Siderophores are iron-chelating molecules that solubilize Fe3+ for microbial utilization and facilitate colonization or infection of eukaryotes by liberating host iron for bacterial uptake. By fluorescently labeling membrane receptors and binding proteins, we created 20 sensors that detect, discriminate, and quantify apo- and ferric siderophores. The sensor proteins originated from TonB-dependent ligand-gated porins (LGPs) of Escherichia coli (Fiu, FepA, Cir, FhuA, IutA, BtuB), Klebsiella pneumoniae (IroN, FepA, FyuA), Acinetobacter baumannii (PiuA, FepA, PirA, BauA), Pseudomonas aeruginosa (FepA, FpvA), and Caulobacter crescentus (HutA) from a periplasmic E. coli binding protein (FepB) and from a human serum binding protein (siderocalin). They detected ferric catecholates (enterobactin, degraded enterobactin, glucosylated enterobactin, dihydroxybenzoate, dihydroxybenzoyl serine, cefidericol, MB-1), ferric hydroxamates (ferrichromes, aerobactin), mixed iron complexes (yersiniabactin, acinetobactin, pyoverdine), and porphyrins (hemin, vitamin B12). The sensors defined the specificities and corresponding affinities of the LGPs and binding proteins and monitored ferric siderophore and porphyrin transport by microbial pathogens. We also quantified, for the first time, broad recognition of diverse ferric complexes by some LGPs, as well as monospecificity for a single metal chelate by others. In addition to their primary ferric siderophore ligands, most LGPs bound the corresponding aposiderophore with ∼100-fold lower affinity. These sensors provide insights into ferric siderophore biosynthesis and uptake pathways in free-living, commensal, and pathogenic Gram-negative bacteria.

Thermostability enhancement of the α-carbonic anhydrase from Sulfurihydrogenibium yellowstonense by using the anchoring-and-self-labelling-protein-tag system (ASLtag).

Carbonic anhydrases (CAs, EC 4.2.1.1) are a superfamily of ubiquitous metalloenzymes present in all living organisms on the planet. They are classified into seven genetically distinct families and catalyse the hydration reaction of carbon dioxide to bicarbonate and protons, as well as the opposite reaction. CAs were proposed to be used for biotechnological applications, such as the post-combustion carbon capture processes. In this context, there is a great interest in searching CAs with robust chemical and physical properties. Here, we describe the enhancement of thermostability of the α-CA from Sulfurihydrogenibium yellowstonense (SspCA) by using the anchoring-and-self-labelling-protein-tag system (ASLtag). The anchored chimeric H5-SspCA was active for the CO2 hydration reaction and its thermostability increased when the cells were heated for a prolonged period at high temperatures (e.g. 70 °C). The ASLtag can be considered as a useful method for enhancing the thermostability of a protein useful for biotechnological applications, which often need harsh operating conditions.

POSTOPERATIVE ENDOPHTHALMITIS CAUSED BY BOSEA THIOOXIDANS.

PURPOSE: To report an unusual case of postcataract endophthalmitis secondary to Bosea thiooxidans. METHODS: Case report. RESULTS: An 86-year-old man presented with postoperative endophthalmitis 3 weeks after uncomplicated cataract surgery. He complained of a headache and blurry vision but denied having eye pain. Slit-lamp examination demonstrated leukocytes with a fibrinous membrane in the anterior segment and vitritis. A diagnosis of acute postoperative endophthalmitis was made, and the patient was treated with intravitreal antibiotics. Culture of vitreous tap revealed Bosea thiooxidans, a gram-negative rod found in soil and water systems. CONCLUSION: To our knowledge, this report is the first to describe Bosea thiooxidans endophthalmitis and one of a few reports of human infection with this microbial agent.

A one-step procedure for immobilising the thermostable carbonic anhydrase (SspCA) on the surface membrane of Escherichia coli.

The carbonic anhydrase superfamily (CA, EC 4.2.1.1) of metalloenzymes is present in all three domains of life (Eubacteria, Archaea, and Eukarya), being an interesting example of convergent/divergent evolution, with its seven families (α-, ß-, γ-, δ-, ζ-, η-, and θ-CAs) described so far. CAs catalyse the simple, but physiologically crucial reaction of carbon dioxide hydration to bicarbonate and protons. Recently, our groups characterised the α-CA from the thermophilic bacterium, Sulfurihydrogenibium yellowstonense finding a very high catalytic activity for the CO2 hydration reaction (kcat = 9.35 × 105 s-1 and kcat/Km = 1.1 × 108 M-1 s-1) which was maintained after heating the enzyme at 80 °C for 3 h. This highly thermostable SspCA was covalently immobilised within polyurethane foam and onto the surface of magnetic Fe3O4 nanoparticles. Here, we describe a one-step procedure for immobilising the thermostable SspCA directly on the surface membrane of Escherichia coli, using the INPN domain of Pseudomonas syringae. This strategy has clear advantages with respect to other methods, which require as the first step the production and the purification of the biocatalyst, and as the second step the immobilisation of the enzyme onto a specific support. Our results demonstrate that thermostable SspCA fused to the INPN domain of P. syringae ice nucleation protein (INP) was correctly expressed on the outer membrane of engineered E. coli cells, affording for an easy approach to design biotechnological applications for this highly effective thermostable catalyst.

Production and covalent immobilisation of the recombinant bacterial carbonic anhydrase (SspCA) onto magnetic nanoparticles.

Carbonic anhydrases (CAs; EC 4.2.1.1) are metalloenzymes with a pivotal potential role in the biomimetic CO2 capture process (CCP) because these biocatalysts catalyse the simple but physiologically crucial reaction of carbon dioxide hydration to bicarbonate and protons in all life kingdoms. The CAs are among the fastest known enzymes, with kcat values of up to 106 s-1 for some members of the superfamily, providing thus advantages when compared with other CCP methods, as they are specific for CO2. Thermostable CAs might be used in CCP technology because of their ability to perform catalysis in operatively hard conditions, typical of the industrial processes. Moreover, the improvement of the enzyme stability and its reuse are important for lowering the costs. These aspects can be overcome by immobilising the enzyme on a specific support. We report in this article that the recombinant thermostable SspCA (α-CA) from the thermophilic bacterium Sulfurihydrogenibium yellowstonense can been heterologously produced by a high-density fermentation of Escherichia coli cultures, and covalently immobilised onto the surface of magnetic Fe3O4 nanoparticles (MNP) via carbodiimide activation reactions. Our results demonstrate that using a benchtop bioprocess station and strategies for optimising the bacterial growth, it is possible to produce at low cost a large amount SspCA. Furthermore, the enzyme stability and storage greatly increased through the immobilisation, as SspCA bound to MNP could be recovered from the reaction mixture by simply using a magnet or an electromagnetic field, due to the strong ferromagnetic properties of Fe3O4.

Spatial homogeneity of bacterial and archaeal communities in the deep eastern Mediterranean Sea surface sediments

The diversity of microorganisms inhabiting the deep sea surface sediments was investigated in 9 stations (700-1900 m depth) in the Levantine basin by 454 massive tag sequencing of the 16S rDNA V4 region using universal primers. In total, 108,811 reads (an average of 10,088 per sample) were assigned to 5014 bacterial and 966 archaeal operational taxonomic units (OTUs; at 97% cut off). The 55% of the reads were of archaea, indicating dominance of archaea over bacteria at eight of the stations. The diversity and estimated richness values were high (e.g., H´ ranged from 5.66 to 7.41 for bacteria). The compositions of the microorganisms at all stations were remarkably similar, with Bray-Curtis similarities of 0.53-0.91 and 0.74-0.99 for bacterial and archaeal orders respectively. At two stations, very high abundances of only a few genera (Marinobacterium, Bacillus, Vibrio, Photobacterium) were accountable for the dissimilarities documented compared to the other deep sea stations. Half of the bacterial reads (51%) belonged to the phylum Proteobacteria, comprising mainly Gammaproteobacteria (41-72% of the proteobacterial reads per sample), Deltaproteobacteria (12-29%), Alphaproteobacteria (7-18%) and Betaproteobacteria (3-14%). The most abundant bacterial family was Sinobacteraceae (order Xanthomonadales) with 5-10% of total bacterial reads per sample. Most abundant reads (15.4% of all microbial reads) were affiliated with Marine Group 1 archaea, putatively capable of ammonia oxidation (213 OTUs), and bacteria involved in nitrification were found in all samples. The data point to the significant role that chemolithotrophic carbon assimilation and nitrification fill in the oligotrophic deep sea Levant sediments (AU)

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[Informative indices of the biocorrosion activity for the determination of the character of the aggression ground].

Underground corrosion is referred to the most difficult types of corrosion in connection with that it is multifactorial and differs in progressive dynamics of the participation of each parameter in the process of destruction of the metal. With the aim of the evaluation of the informativeness of the index of the biocorrosion activity caused by the influence of various factors to determine the character of the soil aggressiveness in the district of pipeline laying there was studied the complex of microbiological and physical-chemical indices). There was determined the amount of sulfur cycle bacteria (autotrophic thiobacteria and sulphate-reducing bacteria), the total concentration of sulfur and iron in the soil samples adjacent to the surface of the underground pipelines in the territory of the Khanty-Mansi Autonomous District of Yugra, and the ratio of these indices with a specific electrical resistance of the soil. There was established the predominance ofsamples with weak aggressiveness of the soil (55.17% of cases), with the criterion ofbiocorrosion soil activity of 2,44 ± 0,19. The results show significant differences in the thiobacteria content and mobile iron in the studied soil-ground samples. There was revealed a direct correlation of the average force of concentrations of identified bacteria and iron content in the soil. There was shown the necessity of the implementation of dynamic control and the development of methods of protection of metal structures to prevent biocorrosion in the design and in the process of the operation of the pipeline.

Comparative community gene expression analysis of Aquificales-dominated geothermal springs.

Members of Sulfurihydrogenibium are often observed as visible filamentous biomass in circumneutral hot springs and play roles in sulfur-cycling, hydrogen oxidation and iron mineralization. To gain insight into the ecophysiology of Sulfurihydrogenibium populations, we conducted preliminary metatranscriptomic analysis of three distinct thermal springs; Calcite Springs (YNP-CS) and Mammoth Springs (YNP-MHS) in Yellowstone National Park, USA, and Furnas Springs (AZ) in Azores, Portugal. Genes to which transcripts were assigned revealed commonly expressed functions among the sites, while several differences were also observed. All three sites, Sulfurihydrogenibium spp. dominate and are obtaining energy via metabolism of sulfur compounds under microaerophilic conditions. Cell motility was one of the expressed functions in two sites (YNP-CS and AZ) with slower stream flow rates and thicker well-formed biofilms. The transcripts from YNP-CS and -MHS exhibited varying levels of sequence divergence from the reference genomes and corresponding metagenomes, suggesting the presence of microdiversity among Sulfurihydrogenibium populations in situ. Conversely, the majority of the AZ transcripts were identical to the S. azorense genome. Our initial results show that the metatranscriptomes in these similar Aquificales-dominated communities can reveal community-level gene function in geochemically distinct thermal environments.

The first activation study of a bacterial carbonic anhydrase (CA). The thermostable α-CA from Sulfurihydrogenibium yellowstonense YO3AOP1 is highly activated by amino acids and amines.

The α-carbonic anhydrase (CA, EC 4.2.1.1) from the newly discovered thermophilic bacterium Sulfurihydrogenibium yellowstonense YO3AOP1 (SspCA) was investigated for its activation with a series of amino acids and amines. D-His, L-Phe, L-Tyr, L- and D-Trp were the most effective SspCA activators, with activation constants in the range of 1-12 nM, whereas L-His, L/D-DOPA, D-Tyr, and several biogenic amines/catecholamines were slightly less effective activators (K(A) in the range of 37 nM-0.97 µM). The least effective SspCA activator was d-Phe (K(A) of 5.13 µM). The thermal stability, robustness and very high catalytic activity of SspCA make this enzyme an ideal candidate for biomimetic CO(2) capture processes.

Anion inhibition studies of an α-carbonic anhydrase from the thermophilic bacterium Sulfurihydrogenibium yellowstonense YO3AOP1.

The newly discovered thermophilic bacterium Sulfurihydrogenibium yellowstonense YO3AOP1 encodes an α-carbonic anhydrases (CAs, EC 4.2.1.1) which is highly catalytically active and thermostable. Here we report the inhibition of this enzyme, denominated SspCA, with inorganic and complex anions and other molecules interacting with zinc proteins. SspCA was inhibited in the micromolar range by diethyldithiocarbamate, sulfamide, sulfamic acid, phenylboronic and phenylarsonic acid, trithiocarbonate and selenocyanide (K(I)s of 4-70 µM) and in the submillimolar one by iodide, cyanide, (thio)cyanate, hydrogen sulfide, azide, nitrate, nitrite, many complex anions incorporating heavy metal ions and iminodisulfonate (K(I)s of 0.48-0.86 mM). SspCA was not substantially inhibited by bicarbonate and carbonate, hydrogensulfite and peroxidisulfate (K(I)s in the range of 21.1-84.6mM). The exceptional thermostability and lack of strong affinity for hydrogensulfide, bicarbonate, and carbonate make this enzyme an interesting candidate for biotechnological applications of enzymatic CO(2) fixation.

Biochemical properties of a novel and highly thermostable bacterial α-carbonic anhydrase from Sulfurihydrogenibium yellowstonense YO3AOP1.

A new carbonic anhydrase (CA, EC 4.2.1.1) from the thermophilic bacterium Sulfurihydrogenibium yellowstonense YO3AOP1 was identified and characterized. The bacterial carbonic anhydrase gene was expressed in Escherichia coli yielding an active enzyme, which was purified in large amounts. The recombinant protein (SspCA) was found to belong to the α-CA class and displays esterase activity. The kinetic parameters were determined by using CO(2) and p-nitrophenylacetate (p-NpA) as substrates. The bacterial enzyme presented specific activity comparable to that of bovine carbonic anhydrase (bCA II) but it showed biochemical properties never observed for the mammalian enzyme. The thermophilic enzyme, in fact, was endowed with high thermostability and with unaltered residual activity after prolonged exposure to heat up to 100°C. SspCA and the bovine carbonic anhydrase (bCA II) were immobilized within a polyurethane (PU) foam. The immobilized bacterial enzyme was found to be active and stable at 100°C up to 50 h.

Thermosulfurimonas dismutans gen. nov., sp. nov., an extremely thermophilic sulfur-disproportionating bacterium from a deep-sea hydrothermal vent.

An extremely thermophilic, anaerobic, chemolithoautotrophic bacterium (strain S95(T)) was isolated from a deep-sea hydrothermal vent chimney located on the Eastern Lau Spreading Center, Pacific Ocean, at a depth of 1910 m. Cells of strain S95(T) were oval to short Gram-negative rods, 0.5-0.6 µm in diameter and 1.0-1.5 µm in length, growing singly or in pairs. Cells were motile with a single polar flagellum. The temperature range for growth was 50-92 °C, with an optimum at 74 °C. The pH range for growth was 5.5-8.0, with an optimum at pH 7.0. Growth of strain S95(T) was observed at NaCl concentrations ranging from 1.5 to 3.5% (w/v). Strain S95(T) grew anaerobically with elemental sulfur as an energy source and bicarbonate/CO(2) as a carbon source. Elemental sulfur was disproportionated to sulfide and sulfate. Growth was enhanced in the presence of poorly crystalline iron(III) oxide (ferrihydrite) as a sulfide-scavenging agent. Strain S95(T) was also able to grow by disproportionation of thiosulfate and sulfite. Sulfate was not used as an electron acceptor. Analysis of the 16S rRNA gene sequence revealed that the isolate belongs to the phylum Thermodesulfobacteria. On the basis of its physiological properties and results of phylogenetic analyses, it is proposed that the isolate represents the sole species of a new genus, Thermosulfurimonas dismutans gen. nov., sp. nov.; S95(T) (=DSM 24515(T)=VKM B-2683(T)) is the type strain of the type species. This is the first description of a thermophilic micro-organism that disproportionates elemental sulfur.

Isolation and characterization of a moderately thermophilic nitrite-oxidizing bacterium from a geothermal spring.

Geothermal environments are a suitable habitat for nitrifying microorganisms. Conventional and molecular techniques indicated that chemolithoautotrophic nitrite-oxidizing bacteria affiliated with the genus Nitrospira are widespread in environments with elevated temperatures up to 55 °C in Asia, Europe, and Australia. However, until now, no thermophilic pure cultures of Nitrospira were available, and the physiology of these bacteria was mostly uncharacterized. Here, we report on the isolation and characterization of a novel thermophilic Nitrospira strain from a microbial mat of the terrestrial geothermal spring Gorjachinsk (pH 8.6; temperature 48 °C) from the Baikal rift zone (Russia). Based on phenotypic properties, chemotaxonomic data, and 16S rRNA gene phylogeny, the isolate was assigned to the genus Nitrospira as a representative of a novel species, for which the name Nitrospira calida is proposed. A highly similar 16S rRNA gene sequence (99.6% similarity) was detected in a Garga spring enrichment grown at 46 °C, whereas three further thermophilic Nitrospira enrichments from the Garga spring and from a Kamchatka Peninsula (Russia) terrestrial hot spring could be clearly distinguished from N. calida (93.6-96.1% 16S rRNA gene sequence similarity). The findings confirmed that Nitrospira drive nitrite oxidation in moderate thermophilic habitats and also indicated an unexpected diversity of heat-adapted Nitrospira in geothermal hot springs.

Genome-wide RNA interference in Drosophila cells identifies G protein-coupled receptor kinase 2 as a conserved regulator of NF-kappaB signaling.

Because NF-kappaB signaling pathways are highly conserved in evolution, the fruit fly Drosophila melanogaster provides a good model to study these cascades. We carried out an RNA interference (RNAi)-based genome-wide in vitro reporter assay screen in Drosophila for components of NF-kappaB pathways. We analyzed 16,025 dsRNA-treatments and identified 10 novel NF-kappaB regulators. Of these, nine dsRNA-treatments affect primarily the Toll pathway. G protein-coupled receptor kinase (Gprk)2, CG15737/Toll pathway activation mediating protein, and u-shaped were required for normal Drosomycin response in vivo. Interaction studies revealed that Gprk2 interacts with the Drosophila IkappaB homolog Cactus, but is not required in Cactus degradation, indicating a novel mechanism for NF-kappaB regulation. Morpholino silencing of the zebrafish ortholog of Gprk2 in fish embryos caused impaired cytokine expression after Escherichia coli infection, indicating a conserved role in NF-kappaB signaling. Moreover, small interfering RNA silencing of the human ortholog GRK5 in HeLa cells impaired NF-kappaB reporter activity. Gprk2 RNAi flies are susceptible to infection with Enterococcus faecalis and Gprk2 RNAi rescues Toll(10b)-induced blood cell activation in Drosophila larvae in vivo. We conclude that Gprk2/GRK5 has an evolutionarily conserved role in regulating NF-kappaB signaling.

Sulfurihydrogenibium rodmanii sp. nov., a sulfur-oxidizing chemolithoautotroph from the Uzon Caldera, Kamchatka Peninsula, Russia, and emended description of the genus Sulfurihydrogenibium.

Four thermophilic, sulfur-oxidizing, chemolithoautotrophic strains with >99 % 16S rRNA gene sequence similarity were isolated from terrestrial hot springs in the Geyser Valley and the Uzon Caldera, Kamchatka, Russia. One strain, designated UZ3-5T, was characterized fully. Cells of UZ3-5T were Gram-negative, motile, slightly oval rods (about 0.7 microm wide and 1.0 microm long) with multiple polar flagella. All four strains were obligately microaerophilic chemolithoautotrophs and could use elemental sulfur or thiosulfate as electron donors and oxygen (1-14 %, v/v) as the electron acceptor. Strain UZ3-5T grew at temperatures between 55 and 80 degrees C (optimally at 75 degrees C; 1.1 h doubling time), at pH 5.0-7.2 (optimally at pH 6.0-6.3) and at 0-0.9 % NaCl (optimally in the absence of NaCl). The G+C content of the genomic DNA of strain UZ3-5T was 35 mol%. Phylogenetic analysis revealed that strain UZ3-5T was a member of the genus Sulfurihydrogenibium, its closest relative in culture being Sulfurihydrogenibium azorense Az-Fu1T (98.3 % 16S rRNA gene sequence similarity). On the basis of its physiological and molecular characteristics, strain UZ3-5T represents a novel species of the genus Sulfurihydrogenibium, for which the name Sulfurihydrogenibium rodmanii sp. nov. is proposed. The type strain is UZ3-5T (=OCM 900T =ATCC BAA-1536T =DSM 19533T).

Sulfurihydrogenibium kristjanssonii sp. nov., a hydrogen- and sulfur-oxidizing thermophile isolated from a terrestrial Icelandic hot spring.

Three thermophilic, aerobic, hydrogen- and sulfur-oxidizing bacteria were isolated from an Icelandic hot spring near the town of Hveragerdi and share >99 % 16S rRNA gene sequence similarity. One of these isolates, designated strain I6628T, was selected for further characterization. Strain I6628T is a motile rod, 1.5-2.5 microm long and about 0.5 microm wide. Growth occurred between 40 and 73 degrees C (optimally at 68 degrees C), at pH 5.3-7.8 (optimally at pH 6.6) and at NaCl concentrations between 0 and 0.5 % (w/v). Strain I6628T grew with H2, S0 or S2O3(2-) as an electron donor with O2 (up to 25 %, v/v; optimally at 4-9 %) as the sole electron acceptor. CO2 and succinate were utilized as carbon sources but no organic compounds, including succinate, could be used as an energy source. The G+C content of the genomic DNA was determined to be 28.1 mol%. Phylogenetic analysis of the 16S rRNA gene sequence indicated that strain I6628T is a member of the genus Sulfurihydrogenibium, the closest cultivated relative being the recently described strain Sulfurihydrogenibium rodmanii UZ3-5T (98.2 % sequence similarity). On the basis of the physiology and phylogeny of this organism, strain I6628T represents a novel species of the genus Sulfurihydrogenibium, for which the name Sulfurihydrogenibium kristjanssonii sp. nov. is proposed. The type strain is I6628T (=DSM 19534T =OCM 901T =ATCC BAA-1535T).

Thermosulfidibacter takaii gen. nov., sp. nov., a thermophilic, hydrogen-oxidizing, sulfur-reducing chemolithoautotroph isolated from a deep-sea hydrothermal field in the Southern Okinawa Trough.

A novel thermophilic, sulfur-reducing chemolithoautotroph, strain ABI70S6(T), was isolated from a deep-sea hydrothermal field at the Yonaguni Knoll IV, Southern Okinawa Trough. Cells of strain ABI70S6(T) were motile rods, 0.9-2.0 microm in length and 0.4-0.8 microm in width. Strain ABI70S6(T) was an obligately anaerobic chemolithotroph, exhibiting hydrogen oxidation coupled with sulfur reduction. Growth was observed at 55-78 degrees C (optimum, 70 degrees C), pH 5.0-7.5 (optimum, pH 5.5-6.0) and 0.5-4.5 % NaCl (optimum, 3.0 % NaCl). H(2) and elemental sulfur were utilized as electron donor and acceptor, respectively. The major fatty acids were C(16 : 0) (40.0 %) and C(20 : 1) (60.0 %). The G+C content of genomic DNA was 44.2 mol%. The physiological attributes of strain ABI70S6(T) are similar to those of species of genera within the family Desulfurobacteriaceae, most of which are thermophilic and chemolithoautotrophic sulfur reducers. However, 16S rRNA gene sequence similarities between the novel isolate and type strains of all species within the family Desulfurobacteriaceae were <87 %, which is close to the similarities found between the novel isolate and members of the family Thermodesulfobacteriaceae (<85 %). Based on physiological and phylogenetic features of the novel isolate, it is proposed that it represents a novel species in a novel genus, Thermosulfidibacter takaii gen. nov., sp. nov., within the phylum Aquificae. The type strain of T. takaii is ABI70S6(T) (=JCM 13301(T)=DSM 17441(T)).

Hydrogenivirga okinawensis sp. nov., a thermophilic sulfur-oxidizing chemolithoautotroph isolated from a deep-sea hydrothermal field, Southern Okinawa Trough.

A novel extremely thermophilic sulfur-oxidizing bacterium, strain LS12-2(T), was isolated from a deep-sea hydrothermal field at the Yonaguni Knoll IV, Southern Okinawa Trough. Cells of strain LS12-2(T) were motile rods, 1.5-4.0 microm in length and 0.4-0.5 microm in width. Strain LS12-2(T) was an obligate chemolithoautotroph that could utilize elemental sulfur or thiosulfate as an electron donor and nitrate or oxygen as an electron acceptor. Growth was observed at 65-85 degrees C (optimum 70-75 degrees C), pH 5.8-8.3 (optimum pH 6.9-7.5), 1.0-4.0 % (w/v) NaCl (optimum 2.5 %) and 1.0-7.0 % O(2) in the gas phase (optimum 3.0 %). Fatty acids detected were C(16 : 0) (8.0 %), C(18 : 0) (9.0 %), C(18 : 1) (62.5 %) and C(20 : 1) (20.5 %). The genomic DNA G+C content was 51.3 mol%. 16S rRNA gene sequence analysis indicated that strain LS12-2(T) belonged to the genus Hydrogenivirga. Based on physiological and phylogenetic characteristics of the isolate, it is proposed that this strain represents a novel species in the genus Hydrogenivirga, Hydrogenivirga okinawensis sp. nov. The type strain of Hydrogenivirga okinawensis is LS12-2(T) (=JCM 13302(T)=DSM 17378(T)).

Venenivibrio stagnispumantis gen. nov., sp. nov., a thermophilic hydrogen-oxidizing bacterium isolated from Champagne Pool, Waiotapu, New Zealand.

A novel thermophilic, hydrogen-oxidizing bacterium, designated strain CP.B2(T), was isolated from a terrestrial hot spring in Waiotapu, New Zealand. Cells were motile, slightly rod-shaped, non-spore-forming and Gram-negative. Isolate CP.B2(T) was an obligate chemolithotroph, growing by utilizing H(2) as electron donor and O(2) as corresponding electron acceptor. Elemental sulfur (S(0)) or thiosulfate ( ) was essential for growth. Microbial growth occurred under microaerophilic conditions in 1.0-10.0 % (v/v) O(2) [optimum 4-8 % (v/v) O(2)], between 45 and 75 degrees C (optimum 70 degrees C) and at pH values of 4.8-5.8 (optimum pH 5.4). The DNA G+C content was 29.3 mol%. 16S rRNA gene sequence analysis demonstrated that strain CP.B2(T) belonged to the order Aquificales, with a close phylogenetic relationship to Sulfurihydrogenibium azorense (94 % sequence similarity to the type strain). However, genotypic and metabolic characteristics differentiated the novel isolate from previously described genera of the Aquificales. Therefore, CP.B2(T) represents a novel species in a new genus, for which the name Venenivibrio stagnispumantis gen. nov., sp. nov. is proposed. The type strain of Venenivibrio stagnispumantis is CP.B2(T) (=JCM 14244(T) =DSM 18763(T)).

Physiological and phylogenetic characterization of a novel lithoautotrophic nitrite-oxidizing bacterium, 'Candidatus Nitrospira bockiana'.

A new isolate of a lithoautotrophic nitrite-oxidizing bacterium was obtained from internal corrosion deposits from a steel pipeline of the Moscow heating system. The organism oxidized nitrite as the sole energy source and fixed carbon dioxide as the only carbon source. The cells were extremely pleomorphic: loosely wound spirals, slightly curved and even straight rods were detected, as well as coccoid cells. The highest rate of nitrite consumption (1.5 mM nitrite as substrate) was measured at 42 degrees C, with a temperature range of 28-44 degrees C. In enrichment cultures with Nocardioides sp. as an accompanying organism, optimal oxidation of 5.8 mM nitrite occurred at 45 degrees C, with a range of 28-48 degrees C. Neither pyruvate nor yeast extract stimulated nitrification. Organotrophic growth was not observed. Phylogenetic analysis of 16S rRNA gene sequences revealed that the novel isolate represents a new sublineage of the genus Nitrospira. On the basis of physiological, chemotaxonomic and molecular characteristics, the name 'Candidatus Nitrospira bockiana' is proposed.