Bacterial networks in Atlantic salmon with Piscirickettsiosis.

An unbalanced composition of gut microbiota in fish is hypothesized to play a role in promoting bacterial infections, but the synergistic or antagonistic interactions between bacterial groups in relation to fish health are not well understood. We report that pathogenic species in the Piscirickettsia, Aeromonas, Renibacterium and Tenacibaculum genera were all detected in the digesta and gut mucosa of healthy Atlantic salmon without clinical signs of disease. Although Piscirickettsia salmonis (and other pathogens) occurred in greater frequencies of fish with clinical Salmonid Rickettsial Septicemia (SRS), the relative abundance was about the same as that observed in healthy fish. Remarkably, the SRS-positive fish presented with a generalized mid-gut dysbiosis and positive growth associations between Piscirickettsiaceae and members of other taxonomic families containing known pathogens. The reconstruction of metabolic phenotypes based on the bacterial networks detected in the gut and mucosa indicated the synthesis of Gram-negative virulence factors such as colanic acid and O-antigen were over-represented in SRS positive fish. This evidence indicates that cooperative interactions between organisms of different taxonomic families within localized bacterial networks might promote an opportunity for P. salmonis to cause clinical SRS in the farm environment.

Thiomicrorhabdus immobilis sp. nov., a mesophilic sulfur-oxidizing bacterium isolated from sediment of a brackish lake in northern Japan.

A novel sulfur-oxidizing bacterium, strain Am19T, was isolated from sediment of a brackish lake. Strain Am19T grew chemolithoautotrophically on inorganic sulfur compounds, and heterotrophic growth was not observed. Cells were rod-shaped with length of 1.1-3.0 µm and diameter of 0.5-0.8 µm. Growth was observed at 5-37 °C with an optimum growth temperature of 30 °C. The pH range for growth was 5.6-8.5 with an optimum pH of 6.6-7.0. Major fatty acids were summed feature 3 (C16: 1ω7c and/or C16: 1ω6c), summed feature 8 (C18: 1ω7c and/or C18: 1ω6c) and C16: 0. The sole respiratory quinone was ubiquinone-8. The complete genome of strain Am19T is composed of a circular chromosome with length of 2.5 Mbp and G + C content of 42.7 mol%. Phylogenetic analysis based on genomic data indicated that strain Am19T belongs to the genus Thiomicrorhabdus but is distinct from any existing species. Analysis of the 16S rRNA gene supported creation of a new species to accommodate strain Am19T. On the basis of genomic and phenotypic characteristics, strain Am19T (= NBRC 114602 T = BCRC 81336 T) is proposed as the type strain of a new species, with name of Thiomicrorhabdus immobilis sp. nov.

Heterologous expression and biochemical comparison of two homologous SoxX proteins of endosymbiotic Candidatus Vesicomyosocius okutanii and free-living Hydrogenovibrio crunogenus from deep-sea environments.

Candidatus Vesicomyosocius okutanii is a currently uncultured endosymbiotic bacterium of Phreagena okutanii, a clam that inhabits deep-sea vent environments. The genome of Ca. V. okutanii encodes a sulfur-oxidizing (Sox) enzyme complex, presumably generating biological energy for the host from inorganic sulfur compounds. Here, Ca. V. okutanii SoxX (VoSoxX), a mono-heme cytochrome c component of the Sox complex, was shown to be phylogenetically related to its homologous counterpart (HcSoxX) from a free-living deep-sea bacterium, Hydrogenovibrio crunogenus. Both proteins were heterologously expressed in Escherichia coli co-expressing cytochrome c maturation genes for comparative biochemical analysis. The VoSoxX recombinant had significantly lower thermal stability than HcSoxX, reflecting the difference in growth conditions of the source bacteria. The endosymbiont inhabits a mild intracellular environment, whereas the free-living bacterium dwells in a harsh environment. This study represents the first successful case of heterologous expression of genes from Ca. V. okutanii, allowing further biochemical studies of the molecular mechanism of sulfur oxidation in deep-sea environments.

Structural coordination between active sites of a CRISPR reverse transcriptase-integrase complex.

CRISPR-Cas systems provide adaptive immunity in bacteria and archaea, beginning with integration of foreign sequences into the host CRISPR genomic locus and followed by transcription and maturation of CRISPR RNAs (crRNAs). In some CRISPR systems, a reverse transcriptase (RT) fusion to the Cas1 integrase and Cas6 maturase creates a single protein that enables concerted sequence integration and crRNA production. To elucidate how the RT-integrase organizes distinct enzymatic activities, we present the cryo-EM structure of a Cas6-RT-Cas1-Cas2 CRISPR integrase complex. The structure reveals a heterohexamer in which the RT directly contacts the integrase and maturase domains, suggesting functional coordination between all three active sites. Together with biochemical experiments, our data support a model of sequential enzymatic activities that enable CRISPR sequence acquisition from RNA and DNA substrates. These findings highlight an expanded capacity of some CRISPR systems to acquire diverse sequences that direct CRISPR-mediated interference.

Thiomicrorhabdus sediminis sp. nov. and Thiomicrorhabdus xiamenensis sp. nov., novel sulfur-oxidizing bacteria isolated from coastal sediments and an emended description of the genus Thiomicrorhabdus.

Two novel Gram-strain-negative and rod-shaped bacteria, designated strain G1T and G2T, were isolated from sediment samples collected from the coast of Xiamen, PR China. The cells were motile by a single polar flagellum. Growth of strain G1T occurred at 10-40 °C (optimum, 30 °C), at pH 6.0-9.0 (optimum, pH 7.5) and with 5-1530 mM NaCl (optimum, 510 mM), while the temperature, pH and NaCl concentration ranges for G2T were 4-45 °C (optimum, 28 °C), pH 5.5-8.0 (optimum, pH 6.5) and 85-1530 mM NaCl (optimum, 340 mM). The two isolates were obligate chemolithoautotrophs capable of using thiosulfate, sulfide, elemental sulphur or tetrathionate as an energy source. Strain G1T used molecular oxygen or nitrite as an electron acceptor, while strain G2T used molecular oxygen as the sole electron acceptor. The dominant fatty acids of G1T and G2T were summed feature 3 (C16:1 ω7c and/or C16:1 ω6c), C16 : 0 and summed feature 8 (C18:1 ω7c and/or C18:1 ω6c). The DNA G+C content of G1T and G2T were 45.1 and 48.3 mol%, respectively. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain G1T and G2T were members of the genus Thiomicrorhabdus, and most closely related to Thiomicrorhabdus hydrogeniphila MAS2T (96.0 %) and Thiomicrorhabdus indica 13-15AT (95.4 %), respectively. The 16S rRNA gene sequence similarity between strains G1T and G2T was 95.8 %. Based on the phylogenetic, genomic and phenotypic data presented here, the isolate strains represent novel species of the genus Thiomicrorhabdus, for which the names Thiomicrorhabdus sediminis sp. nov. (type strain G1T=MCCC 1A14511T=KCTC 15841T) and Thiomicrorhabdus xiamenensis sp. nov. (type strain G2T=MCCC 1A14512T=KCTC 15842T) are proposed.

Thiosulfativibrio zosterae gen. nov., sp. nov., and Thiosulfatimonas sediminis gen. nov., sp. nov.

Aerobic, Gram-stain-negative, obligately chemolithoautotrophic thiosulfate-oxidizing bacteria, strains AkT22T and aks77T were isolated from a brackish lake in Japan. Strains AkT22T and aks77T were isolated from samples of eelgrass and sediment, respectively. Growth on sulfide, tetrathionate, elemental sulfur, and organic substrates was not observed for both strains. Growth of the strains was observed at 5 °C or higher temperature, with optimum growth at 22 °C. Strain AkT22T grew at a pH range of 5.8-8.0, with optimum growth at pH 6.7-7.8. Strain aks77T grew at a pH range of 5.8-8.5, with optimum growth at pH 7.0-7.9. Major cellular fatty acids (> 10% of total) of strain AkT22T were C16:1, C18:1, and C16:0. The sole respiratory quinone was ubiquinone-8 in both strains. The genome of strain AkT22T consisted of a circular chromosome, with size of approximately 2.6 Mbp and G + C content of 43.2%. Those values of the genome of strain aks77T were ca. 2.7 Mbp and 45.5%, respectively. Among cultured bacteria, Thiomicrorhabdus aquaedulcis HaS4T showed the highest sequence identities of the 16S rRNA gene, to strains AkT22T (94%) and aks77T (95%). On the basis of these results, Thiosulfativibrio zosterae gen. nov., sp. nov. and Thiosulfatimonas sediminis gen. nov., sp. nov. are proposed, with type strains of AkT22T (= BCRC 81184T = NBRC 114012T = DSM 109948T) and aks77T (= BCRC 81183T = NBRC 114013T), respectively.

Host-Endosymbiont Genome Integration in a Deep-Sea Chemosymbiotic Clam.

Endosymbiosis with chemosynthetic bacteria has enabled many deep-sea invertebrates to thrive at hydrothermal vents and cold seeps, but most previous studies on this mutualism have focused on the bacteria only. Vesicomyid clams dominate global deep-sea chemosynthesis-based ecosystems. They differ from most deep-sea symbiotic animals in passing their symbionts from parent to offspring, enabling intricate coevolution between the host and the symbiont. Here, we sequenced the genomes of the clam Archivesica marissinica (Bivalvia: Vesicomyidae) and its bacterial symbiont to understand the genomic/metabolic integration behind this symbiosis. At 1.52 Gb, the clam genome encodes 28 genes horizontally transferred from bacteria, a large number of pseudogenes and transposable elements whose massive expansion corresponded to the timing of the rise and subsequent divergence of symbiont-bearing vesicomyids. The genome exhibits gene family expansion in cellular processes that likely facilitate chemoautotrophy, including gas delivery to support energy and carbon production, metabolite exchange with the symbiont, and regulation of the bacteriocyte population. Contraction in cellulase genes is likely adaptive to the shift from phytoplankton-derived to bacteria-based food. It also shows contraction in bacterial recognition gene families, indicative of suppressed immune response to the endosymbiont. The gammaproteobacterium endosymbiont has a reduced genome of 1.03 Mb but retains complete pathways for sulfur oxidation, carbon fixation, and biosynthesis of 20 common amino acids, indicating the host's high dependence on the symbiont for nutrition. Overall, the host-symbiont genomes show not only tight metabolic complementarity but also distinct signatures of coevolution allowing the vesicomyids to thrive in chemosynthesis-based ecosystems.

Engineering a Plant Viral Coat Protein for In Vitro Hybrid Self-Assembly of CO2-Capturing Catalytic Nanofilaments.

Plant virus-based nanoparticles are used as self-assembled protein scaffolds for the construction of enzyme nanocarriers. To date, one-pot production and coupling of both enzymes and scaffolds by genetic conjugation have been demonstrated only in plants. Herein, we report bacterial production and in vitro self-assembly of nanofilaments for CO2 capture. Filamentous virus-like particles (VLPs) were successfully formed by genetically fusing carbonic anhydrase from Hydrogenovibrio marinus (hmCA) to the N terminus of the coat protein (CPPVY) of potato virus Y with a flexible linker. The instability of VLPs against proteolytic degradation was circumvented by the periplasmic export of the fusion protein. The truncated form of CPPVY coexpressed by internal translation was crucial for the successful formation of long filamentous VLPs by alleviating steric hindrance via hybrid assembly. The fast and economic bottom-up fabrication of highly active nanobiocatalyst allows the nanofilaments to be efficiently used and recovered in potential biocatalytic and biosensor systems.

Gammaproteobacteria mediating utilization of methyl-, sulfur- and petroleum organic compounds in deep ocean hydrothermal plumes.

Deep-sea hydrothermal plumes are considered natural laboratories for understanding ecological and biogeochemical interactions. Previous studies focused on interactions between microorganisms and inorganic, reduced hydrothermal inputs including sulfur, hydrogen, iron, and manganese. However, little is known about transformations of organic compounds, especially methylated, sulfur-containing compounds, and petroleum hydrocarbons. Here, we reconstructed nine gammaproteobacterial metagenome-assembled genomes, affiliated with Methylococcales, Methylophaga, and Cycloclasticus, from three hydrothermal ecosystems. We present evidence that these three groups have high transcriptional activities of genes encoding cycling of C1-compounds, petroleum hydrocarbons, and organic sulfur in hydrothermal plumes. This includes oxidation of methanethiol, the simplest thermochemically-derived organic sulfur, for energy metabolism in Methylococcales and Cycloclasticus. Together with active transcription of genes for thiosulfate and methane oxidation in Methylococcales, these results suggest an adaptive strategy of versatile and simultaneous use of multiple available electron donors. Meanwhile, the first near-complete MAG of hydrothermal Methylophaga aminisulfidivorans and its transcriptional profile point to active chemotaxis targeting small organic compounds. Petroleum hydrocarbon-degrading Cycloclasticus are abundant and active in plumes of oil spills as well as deep-sea vents, suggesting that they are indigenous and effectively respond to stimulus of hydrocarbons in the deep sea. These findings suggest that these three groups of Gammaproteobacteria transform organic carbon and sulfur compounds via versatile and opportunistic metabolism and modulate biogeochemistry in plumes of hydrothermal systems as well as oil spills, thus contributing broad ecological impact to the deep ocean globally.

Crystal Structure of Cytochrome cL from the Aquatic Methylotrophic Bacterium Methylophaga aminisulfidivorans MPT.

Cytochrome cL (CytcL) is an essential protein in the process of methanol oxidation in methylotrophs. It receives an electron from the pyrroloquinoline quinone (PQQ) cofactor of methanol dehydrogenase (MDH) to produce formaldehyde. The direct electron transfer mechanism between CytcL and MDH remains unknown due to the lack of structural information. To help gain a better understanding of the mechanism, we determined the first crystal structure of heme c containing CytcL from the aquatic methylotrophic bacterium Methylophaga aminisulfidivorans MPT at 2.13 Å resolution. The crystal structure of Ma-CytcL revealed its unique features compared to those of the terrestrial homologues. Apart from Fe in heme, three additional metal ion binding sites for Na+ , Ca+ , and Fe2+ were found, wherein the ions mostly formed coordination bonds with the amino acid residues on the loop (G93-Y111) that interacts with heme. Therefore, these ions seemed to enhance the stability of heme insertion by increasing the loop's steadiness. The basic N-terminal end, together with helix α4 and loop (G126 to Y136), contributed positive charge to the region. In contrast, the acidic C-terminal end provided a negatively charged surface, yielding several electrostatic contact points with partner proteins for electron transfer. These exceptional features of Ma-CytcL, along with the structural information of MDH, led us to hypothesize the need for an adapter protein bridging MDH to CytcL within appropriate proximity for electron transfer. With this knowledge in mind, the methanol oxidation complex reconstitution in vitro could be utilized to produce metabolic intermediates at the industry level.

Stability-Controllable Self-Immobilization of Carbonic Anhydrase Fused with a Silica-Binding Tag onto Diatom Biosilica for Enzymatic CO2 Capture and Utilization.

Exploiting carbonic anhydrase (CA), an enzyme that catalyzes the hydration of CO2, is a powerful route for eco-friendly and cost-effective carbon capture and utilization. For successful industrial applications, the stability and reusability of CA should be improved, which necessitates enzyme immobilization. Herein, the ribosomal protein L2 (Si-tag) from Escherichia coli was utilized for the immobilization of CA onto diatom biosilica, a promising renewable support material. The Si-tag was redesigned (L2NC) and genetically fused to CA from the marine bacterium Hydrogenovibrio marinus (hmCA). One-step self-immobilization of hmCA-L2NC onto diatom biosilica by simple mixing was successfully achieved via Si-tag-mediated strong binding, showing multilayer adsorption with a maximal loading of 1.4 wt %. The immobilized enzyme showed high reusability and no enzyme leakage even under high temperature conditions. The activity of hmCA-L2NC was inversely proportional to the enzyme loading, while the stability was directly proportional to the enzyme loading. This discovered activity-stability trade-off phenomenon could be attributed to macromolecular crowding on the highly dense surface of the enzyme-immobilized biosilica. Collectively, our system not only facilitates the stability-controllable self-immobilization of enzyme via Si-tag on a diatom biosilica support for the robust, facile, and green construction of stable biocatalysts, but is also a unique model for studying the macromolecular crowding effect on surface-immobilized enzymes.

Shift of bacterial community and denitrification functional genes in biofilm electrode reactor in response to high salinity.

High salinity suppresses denitrification by inhibiting microorganism activities. The shift of microbial community and denitrification functional genes under salinity gradient was systematically investigated in a biofilm electrode reactor (BER) and biofilm reactor (BR) systems. Denitrification efficiency of both BER and BR was not significantly inhibited during the period of low salinity (0-2.0%). As the salinity increased to 2.5%, BER could overcome the impact of high salinity and maintained a relatively stable denitrification performance, and the effluent NO3--N was lower than 1.5 mg/L. High salinity (>2.5%) impoverished microbial diversity and altered the microbial community in both BER and BR. However, two genera Methylophaga and Methyloexplanations were enriched in BER due to electrochemical stimulation, which can tolerate high salinity (>3.0%). The relative abundance of Methylophaga in BER was almost 10 times as much as in BR. Paracoccus is a hydrogen autotrophic denitrifier, which was obviously inhibited with 1.0% NaCl. The hetertrophic denitrifiers were primarily responsible for the nitrate removal in the BER compared to the autotrophic denitrifiers. The abundance and proportion of denitrifying functional genes confirmed that main denitrifiers shift to salt-tolerant species (nirK-type denitrifiers) to reduce the toxic effects. The napA (2.2 × 108 to 6.5 × 108 copies/g biofilm) and nosZ (2.2 × 107 to 4.4 × 107 copies/g biofilm) genes were more abundant in BER compared to BR's, which was attributed to the enrichment of Methylophaga alcalica and Methyloversatilis universalis FAM5 in the BER. The results proved that BER had greater denitrification potential under high salinity (>2.0%) stress at the molecular level.

Thiomicrorhabdus indica sp. nov., an obligately chemolithoautotrophic, sulfur-oxidizing bacterium isolated from a deep-sea hydrothermal vent environment.

A new mesophilic bacterium, designated strain 13-15AT, was isolated from the deep-sea water from the Carlsberg Ridge, northwestern Indian Ocean. Cells were short rods and motile with a single polar flagellum. Growth was observed in the presence of 85-1700 mM NaCl (optimum 680 mM), at 10-45 °C (optimum, 28 °C) and pH 5.0-9.0 (optimum, pH 7.0). The isolate was an obligate chemolithoautotroph capable of growth using thiosulfate, sulfide, elemental sulfur or tetrathionate as the sole energy source, carbon dioxide as the sole carbon source, and molecular oxygen as the sole electron acceptor. Molecular hydrogen did not support growth. The major fatty acids were C16 : 1 (45.0 %), C18 : 1 (22.5 %) and C16 : 0 (20.1 %). The G+C content of the genomic DNA was 41.6 mol%. The results of phylogenetic analysis based on 16S rRNA gene sequences showed that the novel isolate belonged to the genus Thiomicrorhabdus and was most closely related to Thiomicrorhabdus hydrogeniphila MAS2T (94.8 % sequence similarity). On the basis of the taxonomic data obtained in this study, strain 13-15AT represents a novel species of the genus Thiomicrorhabdus, for which the name Thiomicrorhabdus indica sp. nov. is proposed, with the type strain 13-15AT (=MCCC 1A13986T=KCTC 15750T).

Engineering the genetic components of a whole-cell catalyst for improved enzymatic CO2 capture and utilization.

Carbonic anhydrase (CA) is a diffusion-limited enzyme that rapidly catalyzes the hydration of carbon dioxide (CO2 ). CA has been proposed as an eco-friendly yet powerful catalyst for CO2 capture and utilization. A bacterial whole-cell biocatalyst equipped with periplasmic CA provides an option for a cost-effective CO2 -capturing system. However, further utilization of the previously constructed periplasmic system has been limited by its relatively low activity and stability. Herein, we engineered three genetic components of the periplasmic system for the construction of a highly efficient whole-cell catalyst: a CA-coding gene, a signal sequence, and a ribosome-binding site (RBS). A stable and halotolerant CA (hmCA) from the marine bacterium Hydrogenovibrio marinus was employed to improve both the activity and stability of the system. The improved secretion and folding of hmCA and increased membrane permeability were achieved by translocation via the Sec-dependent pathway. The engineering of RBS strength further enhanced whole-cell activity by improving both the secretion and folding of hmCA. The newly engineered biocatalyst displayed 5.7-fold higher activity and 780-fold higher stability at 60°C compared with those of the previously constructed periplasmic system, providing new opportunities for applications in CO2 capture and utilization.

Caligus rogercresseyi infestation is associated with Piscirickettsia salmonis-attributed mortalities in farmed salmonids in Chile.

Caligidosis and Piscirickettsiosis are currently the most important sanitary challenges for the Chilean salmon industry. Caligidosis is caused by the ectoparasite, Caligus rogercresseyi and Piscirickettsiosis is caused by the intracellular bacterium, Piscirickettsia salmonis. Both diseases are highly prevalent and widely distributed in farming areas in Chile. The co-occurrence of the two diseases is frequently reported on salmon farms. However, there is little epidemiological evidence as to whether these two diseases are associated and generate interactive effects. This study was undertaken to evaluate the potential effects of C. rogercresseyi infestation on P. salmonis-attributed mortalities in farmed salmonids in Chile. Using a linear regression model, the potential association between the mean abundance of adult C. rogercresseyi in a period of 10 weeks and Piscirickettsiosis cumulative mortalities observed in the following 10 weeks was evaluated, while controlling for important confounders. These two 10-week windows were set around the time-point at which Piscirickettsiosis weekly mortality exceeded 0.1% for the first time in a production cycle. We found that the mean abundance of adult C. rogercresseyi was significantly associated with the Piscirickettsiosis cumulative mortality, suggesting the two diseases have a synergistic relationship. This relationship was of the same intensity in Atlantic salmon and rainbow trout. Our findings highlight the importance of taking effective control measures for C. rogercresseyi as a part of the strategies in place to reduce P. salmonis-attributed mortalities on salmon farms in Chile.

Thiomicrorhabdus aquaedulcis sp. nov., a sulfur-oxidizing bacterium isolated from lake water.

Strain HaS4T is an aerobic sulfur-oxidizing bacterium isolated from water of Lake Harutori in Japan. It was isolated and partially characterized in a previous study, but its taxonomic status has not been determined. The previous study revealed that the strain is an obligate chemolithoautotroph which grows at temperatures ranging from 0 to 25 °C (optimum, 22 °C) and pH from pH 6.2 to 8.8 (optimum, pH 6.6-7.4). In this study, further characterization of the strain was made to describe it as representative of a novel species. Cells of strain HaS4T are rod-shaped, 1.6-2.5 µm long, 0.7-0.9 µm wide and Gram-stain-negative. Major cellular fatty acids were summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c), summed feature 8 (C18 : 1ω7c and/or C18 : 1ω6c) and C16 : 0. Phylogenetic analysis based on the 16S rRNA gene indicated that the strain is related to the genus Thiomicrorhabdus, but phylogenetically distinct from the type strains of existing species in the genus. On the basis of its phylogenetic and phenotypic properties, strain HaS4T (=NBRC 112315T=BCRC 81110T) is proposed as type strain of a new non-marine species of the genus Thiomicrorhabdus with the name Thiomicrorhabdus aquaedulcis sp. nov.

The effect of O2 and pressure on thiosulfate oxidation by Thiomicrospira thermophila.

Microbial sulfur cycling in marine sediments often occurs in environments characterized by transient chemical gradients that affect both the availability of nutrients and the activity of microbes. High turnover rates of intermediate valence sulfur compounds and the intermittent availability of oxygen in these systems greatly impact the activity of sulfur-oxidizing micro-organisms in particular. In this study, the thiosulfate-oxidizing hydrothermal vent bacterium Thiomicrospira thermophila strain EPR85 was grown in continuous culture at a range of dissolved oxygen concentrations (0.04-1.9 mM) and high pressure (5-10 MPa) in medium buffered at pH 8. Thiosulfate oxidation under these conditions produced tetrathionate, sulfate, and elemental sulfur, in contrast to previous closed-system experiments at ambient pressure during which thiosulfate was quantitatively oxidized to sulfate. The maximum observed specific growth rate at 5 MPa pressure under unlimited O2 was 0.25 hr-1 . This is comparable to the µmax (0.28 hr-1 ) observed at low pH (<6) at ambient pressure when T. thermophila produces the same mix of sulfur species. The half-saturation constant for O2 ( KO2 ) estimated from this study was 0.2 mM (at a cell density of 105 cells/ml) and was robust at all pressures tested (0.4-10 MPa), consistent with piezotolerant behavior of this strain. The cell-specific KO2 was determined to be 1.5 pmol O2 /cell. The concentrations of products formed were correlated with oxygen availability, with tetrathionate production in excess of sulfate production at all pressure conditions tested. This study provides evidence for transient sulfur storage during times when substrate concentration exceeds cell-specific KO2 and subsequent consumption when oxygen dropped below that threshold. These results may be common among sulfur oxidizers in a variety of environments (e.g., deep marine sediments to photosynthetic microbial mats).

Horizontal acquisition of hydrogen conversion ability and other habitat adaptations in the Hydrogenovibrio strains SP-41 and XCL-2.

BACKGROUND: Obligate sulfur oxidizing chemolithoauthotrophic strains of Hydrogenovibrio crunogenus have been isolated from multiple hydrothermal vent associated habitats. However, a hydrogenase gene cluster (encoding the hydrogen converting enzyme and its maturation/assembly machinery) detected on the first sequenced H. crunogenus strain (XCL-2) suggested that hydrogen conversion may also play a role in this organism. Yet, numerous experiments have underlined XCL-2's inability to consume hydrogen under the tested conditions. A recent study showed that the closely related strain SP-41 contains a homolog of the XCL-2 hydrogenase (a group 1b [NiFe]-hydrogenase), but that it can indeed use hydrogen. Hence, the question remained unresolved, why SP-41 is capable of using hydrogen, while XCL-2 is not. RESULTS: Here, we present the genome sequence of the SP-41 strain and compare it to that of the XCL-2 strain. We show that the chromosome of SP-41 codes for a further hydrogenase gene cluster, including two additional hydrogenases: the first appears to be a group 1d periplasmic membrane-anchored hydrogenase, and the second a group 2b sensory hydrogenase. The region where these genes are located was likely acquired horizontally and exhibits similarity to other Hydrogenovibrio species (H. thermophilus MA2-6 and H. marinus MH-110 T) and other hydrogen oxidizing Proteobacteria (Cupriavidus necator H16 and Ghiorsea bivora TAG-1 T). The genomes of XCL-2 and SP-41 show a strong conservation in gene order. However, several short genomic regions are not contained in the genome of the other strain. These exclusive regions are often associated with signs of DNA mobility, such as genes coding for transposases. They code for transport systems and/or extend the metabolic potential of the strains. CONCLUSIONS: Our results suggest that horizontal gene transfer plays an important role in shaping the genomes of these strains, as a likely mechanism for habitat adaptation, including, but not limited to the transfer of the hydrogen conversion ability.

New Zealand rickettsia-like organism (NZ-RLO) and Tenacibaculum maritimum: Distribution and phylogeny in farmed Chinook salmon (Oncorhynchus tshawytscha).

A total of 777 fish from three growing regions of New Zealand Chinook salmon farms comprising of five sites were tested. Quantitative PCR was used to determine the distribution of New Zealand rickettsia-like organism and Tenacibaculum maritimum. Genetic information from these bacteria were then compared with strains reported worldwide. Using this information, suggested associations of pathogens with clinically affected fish were made. NZ-RLO was detected in two of the three regions, and T. maritimum was detected in all regions. Three strains of NZ-RLO were identified during this study. Based on analysis of the ITS rRNA gene, NZ-RLO1 appears to be part of an Australasian grouping sharing high similarity with the Tasmanian RLO, NZ-RLO2 was shown to be the same as an Irish strain, and NZ-RLO3 was shown be closely related to two strains from Chile. Based on multi-locus sequence typing, the New Zealand T. maritimum was the same as Australian strains. NZ-RLOs were detected more frequently in fish with skin ulcers than fish without skin ulcers. While additional research is required to investigate the pathogenicity of these organisms, this is the first time that NZ-RLOs have been associated with the development of clinical infections in farmed Chinook salmon.

Diversity in CO2-Concentrating Mechanisms among Chemolithoautotrophs from the Genera Hydrogenovibrio, Thiomicrorhabdus, and Thiomicrospira, Ubiquitous in Sulfidic Habitats Worldwide.

Members of the genera Hydrogenovibrio, Thiomicrospira, and Thiomicrorhabdus fix carbon at hydrothermal vents, coastal sediments, hypersaline lakes, and other sulfidic habitats. The genome sequences of these ubiquitous and prolific chemolithoautotrophs suggest a surprising diversity of mechanisms for the uptake and fixation of dissolved inorganic carbon (DIC); these mechanisms are verified here. Carboxysomes are apparent in the transmission electron micrographs of most of these organisms but are lacking in Thiomicrorhabdus sp. strain Milos-T2 and Thiomicrorhabdus arctica, and the inability of Thiomicrorhabdus sp. strain Milos-T2 to grow under low-DIC conditions is consistent with the absence of carboxysome loci in its genome. For the remaining organisms, genes encoding potential DIC transporters from four evolutionarily distinct families (Tcr_0853 and Tcr_0854, Chr, SbtA, and SulP) are located downstream of carboxysome loci. Transporter genes collocated with carboxysome loci, as well as some homologs located elsewhere on the chromosomes, had elevated transcript levels under low-DIC conditions, as assayed by reverse transcription-quantitative PCR (qRT-PCR). DIC uptake was measureable via silicone oil centrifugation when a representative of each of the four types of transporter was expressed in Escherichia coli The expression of these genes in the carbonic anhydrase-deficient E. coli strain EDCM636 enabled it to grow under low-DIC conditions, a result consistent with DIC transport by these proteins. The results from this study expand the range of DIC transporters within the SbtA and SulP transporter families, verify DIC uptake by transporters encoded by Tcr_0853 and Tcr_0854 and their homologs, and introduce DIC as a potential substrate for transporters from the Chr family.IMPORTANCE Autotrophic organisms take up and fix DIC, introducing carbon into the biological portion of the global carbon cycle. The mechanisms for DIC uptake and fixation by autotrophic Bacteria and Archaea are likely to be diverse but have been well characterized only for "Cyanobacteria" Based on genome sequences, members of the genera Hydrogenovibrio, Thiomicrospira, and Thiomicrorhabdus have a variety of mechanisms for DIC uptake and fixation. We verified that most of these organisms are capable of growing under low-DIC conditions, when they upregulate carboxysome loci and transporter genes collocated with these loci on their chromosomes. When these genes, which fall into four evolutionarily independent families of transporters, are expressed in E. coli, DIC transport is detected. This expansion in known DIC transporters across four families, from organisms from a variety of environments, provides insight into the ecophysiology of autotrophs, as well as a toolkit for engineering microorganisms for carbon-neutral biochemistries of industrial importance.