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NtrC protein of piezophilic Shewanella violacea was overexpressed and purified, to confirm the protein-DNA interaction. An electrophoretic mobility shift assay demonstrated that the NtrC recognizes the sequence for NtrC binding within the region upstream of the glnA operon. Western blot analysis also showed that the NtrC is expressed at a higher level under high-pressure conditions than under atmospheric pressure conditions.

In PUREX process for spent fuel reprocessing plants, nitric acid solutions are treated in many components. Particularly, heating portions in the components are severely corroded in the boiling solution under heat transfer (HT) conditions. Although corrosion behavior under the conditions has been investigated by many researchers, corrosion mechanism has not been sufficiently understood. Consequently, because of its safety, we need to study dominant factors of corrosion mechanism and to develop a method to predict corrosion progress.Surface temperature, heat flux, concentration and compositions of the solution have been previously considered as important factors for corrosion rate (CR). In this paper, authors have focused on the effects of surface temperature and heat flux on CRs of stainless steels in boiling nitric acid under HT conditions. We performed experimental study in consideration of dissolvers and concentrators. Two types of cells for HT and immersion conditions were applied for corrosion tests and the effects were compared. Test solution used was 33 mol·m−3 (0.033 M) vanadium added to 3 kmol·m−3 (3 M) nitric acid solution. The solution was heated at boiling temperature under atmospheric pressure. Additionally, a boiling curve which was indicated the relation between heat flux and degree of superheating was investigated experimentally. Surface temperatures during corrosion tests were estimated from a boiling curve. The relationship among measured CRs, surface temperature and heat flux were studied.The results showed that CR did not depend on heat flux, but depended on surface temperature. Arrhenius plots on CRs indicated that CR was accelerated by solution boiling against non-boiling. The cause of the acceleration of CR under boiling nitric acid solution was discussed.Copyright © 2012 by ASME

All the self-reproducing cellular organisms so far examined have DNA as the genome. However, a DNA-less organism carrying an RNA genome is suggested by the fact that many RNA viruses exist and the widespread view that an RNA world existed before the present DNA world. Such a possibility is most plausible in the microbial world where biological diversity is enormous and most organisms have not been identified. We have developed experimental methodology to search DNA-less microorganisms, which is based on cultivation with drugs that inhibit replication or expression of DNA, detection of DNA in colonies with a fluorescent dye and double staining for DNA and RNA at a cellular level. These methods have been applied for about 100 microbial samples from various waters including hot springs, soils including deep sea sediments, and organisms. We found many colonies and cells which apparently looked DNA-less and examined them further. So far, all such colonies that reformed colonies on isolation were identified to be DNA-positive. However, considering the difficulty in cultivation, we think it possible for DNA-less microorganisms to live around us. We believe that our ideas and results will be of interest and useful to discover one in the future.

Cytochrome c₅ of pressure-sensitive Shewanella livingstonensis (SL cytc₅) exhibits lower thermal stability than a highly homologous counterpart of pressure-tolerant Shewanella violacea. This stability difference is due to an enthalpic effect that can be attributed to the amino acid residue at position 50 (Leu or Lys). These cytc₅ proteins are appropriate materials for understanding the protein stability mechanism.

Shewanella violacea DSS12 is a psychrophilic piezophile that optimally grows at 30 MPa. It contains a substantial amount of eicosapentaenoic acid (EPA) in the membrane. Despite evidence linking increased fatty acid unsaturation and bacterial growth under high pressure, little is known of how the physicochemical properties of the membrane are modulated by unsaturated fatty acids in vivo. By means of the newly developed system performing time-resolved fluorescence anisotropy measurement under high pressure (HP-TRFAM), we demonstrate that the membrane of S. violacea is highly ordered at 0.1 MPa and 10 °C with the order parameter S of 0.9, and the rotational diffusion coefficient Dw of 5.4 μs–1 for 1-[4-(trimethylamino)pheny]-6-phenyl-1,3,5-hexatriene in the membrane. Deletion of pfaA encoding the omega-3 polyunsaturated fatty acid synthase caused disorder of the membrane and enhanced the rotational motion of acyl chains, in concert with a 2-fold increase in the palmitoleic acid level. While the wild-type membrane was unperturbed over a wide range of pressures with respect to relatively small effects of pressure on S and Dw, the ΔpfaA membrane was disturbed judging from the degree of increased S and decreased Dw. These results suggest that EPA prevents the membrane from becoming hyperfluid and maintains membrane stability against significant changes in pressure. Our results counter the generally accepted concept that greater fluidity is a membrane characteristic of microorganisms that inhabit cold, high-pressure environments. We suggest that retaining a certain level of membrane physical properties under high pressure is more important than conferring membrane fluidity alone.

The facultative piezophile Shewanella violacea DSS12 is known to alter its respiratory components under the influence of hydrostatic pressure during growth, suggesting that it has a respiratory system that functions in adaptation to high pressure. We investigated the pressure- and temperature-dependencies of the respiratory terminal oxidase activity of the membrane of S. violacea relative to non-piezophilic Shewanella species. We observed that the activity in the membrane of S. violacea was more resistant to high pressure than those of non-piezophilic Shewanella even though DSS12 was cultured under atmospheric pressure. On the other hand, the temperature dependency of this activity was almost the same for all of the tested strain regardless of optimal growth temperature. Both high pressure and low temperature are expected to lower protein flexibility, causing a decrease in enzyme activity, but the results of this study suggest that the mechanism maintaining enzyme activity under high hydrostatic pressure is different from that at low temperature. Additionally, the responses of the activity to the pressure- and temperature-changes were independent of membrane lipid composition. Therefore, the piezotolerance of the respiratory terminal oxidases of S. violacea is perhaps dependent on the properties of the protein itself and not on the lipid composition of the membrane. Our observations suggest that S. violacea constitutively express piezotolerant respiratory terminal oxidases that serve adaptation to the deep-sea environment.

Shewanella violacea DSS12, a deep-sea bacterium, produces eicosapentaenoic acid (EPA) as a component of membrane phospholipids. Although various isolates from the deep sea, such as Photobacterium profundum SS9, Colwellia psychrerythraea 34H and various Shewanella strains, produce EPA- or docosahexaenoic acid-containing phospholipids, the physiological role of these polyunsaturated fatty acids remains unclear. In this article, we illustrate the physiological importance of EPA for high-pressure adaptation in strain DSS12 with the help of an EPA-deficient mutant (DSS12(pfaA)). DSS12(pfaA) showed significant growth retardation at 30 MPa, but not at 0.1 MPa. We also found that DSS12(pfaA) grown at 30 MPa forms filamentous cells. When an EPA-containing phospholipid (sn-1-oleoly-sn-2-eicosapentaenoyl phosphatidylethanolamine) was supplemented, the growth retardation and the morphological defect of DSS12(pfaA) were suppressed, indicating that the externally added EPA-containing phospholipid compensated for the loss of endogenous EPA. In contrast, the addition of an oleic acid-containing phospholipid (sn-1,2-dioleoyl phosphatidylethanolamine) did not affect the growth and the morphology of the cells. Immunofluorescent microscopic analysis with anti-FtsZ antibody revealed a number of Z-rings and separated nucleoids in DSS12(pfaA) grown at 30 MPa. These results demonstrate the physiological importance of EPA for the later step of Z-ring formation of S. violacea DSS12 under high-pressure conditions.

Crevice-corrosion tests were performed in gamma-ray irradiated high-temperature water of 288 °C on Type 316L stainless steel. The gamma-ray dose rate was about 30 kGy h−1. Tested specimen surfaces were analyzed using SEM, laser Raman spectroscopy and TEM/EDX. Experimental data were presented in order to show the differences made by the irradiation and crevice-shape simulated structure. Both gamma-ray irradiation and crevice-shape simulated structure changed the corrosion phenomena. On the gamma-ray irradiated crevice-shape simulated surface, α-Fe2O3 particles more than 5 μm in diameter were observed. It suggested that corrosion environment on the crevice-shape simulated surface became severer by gamma-ray irradiation.

Submonolayer Bi-induced structures at the surfaces of 20-monolayer (ML)-thick Ag(111) films on $\text{Si}(111)7\ifmmode\times\else\texttimes\fi{}7$ substrates were investigated using scanning tunneling microscopy. Bi atoms initially adsorbed randomly in the region of Bi coverage $({\ensuremath{\theta}}_{\text{Bi}})$ below 1/3 ML. However, local $\sqrt{3}\ifmmode\times\else\texttimes\fi{}\sqrt{3}$ ordering was extended with increasing ${\ensuremath{\theta}}_{\text{Bi}}$ and $\sqrt{3}\ifmmode\times\else\texttimes\fi{}\sqrt{3}$ reconstruction was completed at ${\ensuremath{\theta}}_{\text{Bi}}=1/3\text{ }\text{ML}$. At larger ${\ensuremath{\theta}}_{\text{Bi}}$ , the surface was covered with a striped structure. Bi atoms were arranged with centered rectangular unit cells, although the contrast of the cell changed periodically along the ${01\overline{1}}$ direction. As a result, bright and dark stripes along the ${\overline{2}11}$ direction appeared alternatively at the surfaces. A close-packed Bi-induced $(\genfrac{}{}{0}{}{3\text{ }0}{1\text{ }2})$ structure is proposed as a model for the striped surface structure.

To evaluate the effect of post-transplant imatinib administration, 34 Philadelphia chromosome-positive acute lymphoblastic leukaemia (Ph(+)ALL) patients were retrospectively analysed, with 7 receiving post-transplant imatinib administration. Overall survival was significantly better in patients with post-transplant administration (66.7% vs. 29.6% at 3 years, p=0.03), with no significant difference in leukaemia-free survival (0% vs. 29.6% at 3 years, p=0.29). The median duration of negative minimal residual disease (MRD) in patients with post-transplant imatinib administration was 6 months in the pre-emptive administration group, where imatinib was administered upon detecting MRD after allogeneic stem cell transplantation (allo-SCT). In the prophylactic administration group, imatinib was administered as soon as possible after allo-SCT, and the median duration of MRD was 12 months. In all patients whose observation periods were longer than one year, MRD became positive in both groups leading to haematological relapse. It is therefore concluded that post-transplant imatinib administration is not an ideal treatment for Ph(+)ALL patients whose MRD is positive at allo-SCT.

Microorganisms are globally dispersed and are able to proliferate in any habitat that supports their lifestyles, which, however, has not yet been explored in any specific microbial taxon. The social myxobacteria are considered typical soil bacteria because they have been identified in various terrestrial samples, a few in coastal areas, but none in other oceanic environments. To explore the prevalence of marine myxobacteria and to investigate their phylogenetic relationships with their terrestrial counterparts, we established myxobacteria-enriched libraries of 16S rRNA gene sequences from four deep-sea sediments collected at depths from 853 to 4675 m and a hydrothermal vent at a depth of 204 m. In all, 68 different myxobacteria-related sequences were identified from randomly sequenced clones of the libraries of different samples. These myxobacterial sequences were diverse but phylogenetically similar at different locations and depths. However, they were separated from terrestrial myxobacteria at high levels of classification. This discovery indicates that the marine myxobacteria are phylogeographically separated from their terrestrial relatives, likely because of geographic separation and environment selection.

Remineralization of organic matter in deep-sea sediments is important in oceanic biogeochemical cycles, and bacteria play a major role in this process. Shewanella violacea DSS12 is a psychrophilic and piezophilic gamma-proteobacterium that was isolated from the surface layer of deep sea sediment at a depth of 5110 m. Here, we report the complete genome sequence of S. violacea and comparative analysis with the genome of S. oneidensis MR-1, isolated from sediments of a freshwater lake. Unlike S. oneidensis, this deep-sea Shewanella possesses very few terminal reductases for anaerobic respiration and no c-type cytochromes or outer membrane proteins involved in respiratory Fe(iii) reduction, which is characteristic of most Shewanella species. Instead, the S. violacea genome contains more terminal oxidases for aerobic respiration and a much greater number of putative secreted proteases and polysaccharases, in particular, for hydrolysis of collagen, cellulose and chitin, than are encoded in S. oneidensis. Transporters and assimilatory reductases for nitrate and nitrite, and nitric oxide-detoxifying mechanisms (flavohemoglobin and flavorubredoxin) are found in S. violacea. Comparative analysis of the S. violacea genome revealed the respiratory adaptation of this bacterium to aerobiosis, leading to predominantly aerobic oxidation of organic matter in surface sediments, as well as its ability to efficiently use diverse organic matter and to assimilate inorganic nitrogen as a survival strategy in the nutrient-poor deep-sea floor.

We are engaged in international standardization activities for 3D displays. We consider that for a sound development of 3D displays' market, the standards should be based on not only mechanism of 3D displays, but also human factors for stereopsis. However, we think that there is no common understanding on what the 3D display should be and that the situation makes developing the standards difficult. In this paper, to understand the mechanism and human factors, we focus on a double image, which occurs in some conditions on an autostereoscopic display. Although the double image is generally considered as an unwanted effect, we consider that whether the double image is unwanted or not depends on the situation and that there are some allowable double images. We tried to classify the double images into the unwanted and the allowable in terms of the display mechanism and visual ergonomics for stereopsis. The issues associated with the double image are closely related to performance characteristics for the autostereoscopic display. We also propose performance characteristics, measurement and analysis methods to represent interocular crosstalk and motion parallax.

Imatinib mesylate has significantly improved the outcome of patients with CML. In the IRIS trial, major molecular response (MMR), which is defined as the achievement ofor =3 log reduction in bcr-abl mRNA from the standardized baseline, was observed in 40% of CML patients by 12 months. Achievement of an MMR at 18 months is associated with 100% probability of transformation-free survival at 60 months, and MMR is an important goal of therapy. The nucleic acid quantitative "DNA probe FR Amp-CML" kit based on the transcription-mediated amplification method, can measure major bcr-abl mRNA in peripheral blood leukocytes. In this study, we studied the clinical usefulness of Amp-CML for monitoring minimum residual disease by comparison with the European standard nucleic acid quantitative method and real-time quantitative PCR (RQ-PCR) with GAPDH as an internal control, using peripheral leukocytes obtained from patients receiving imatinib treatment. The results indicated that Amp-CML had a significant correlation with Fusion Quant M-BCR (R0.971, P0.01), a standard nucleic acid quantitative method used in Europe and RQ-PCR (R0.974, P0.01), especially in samples with more than 100 copies/microg RNA of major bcr-abl mRNA. These data suggest that Amp-CML is reliable for monitoring major bcr-abl mRNA in patients having achieved an MMR.

The deep-sea denitrifier Pseudomonas sp. strain MT-1 has two distinct gene clusters encoding dissimilatory nitrate reductases, periplasmic nitrate reductase (Nap) and membrane-bound nitrate reductase (Nar). In order to investigate the physiological roles of these enzymes, we determined the nitrate reductase activity of the soluble and membrane fractions from MT-1 and the type strain of Pseudomonas stutzeri (closely related with MT-1) grown under various conditions. In MT-1, the activities of both fractions were highest when the cells were grown anaerobically in the presence of nitrate under atmospheric pressure. However, the activity of the soluble fraction decreased when the cells were grown under high pressure, whereas that of membrane fraction remained constant. Further, the activity of the soluble fraction decreased when the enzyme reaction was performed at low temperature, although that of membrane fraction was not similarly affected. Additionally, the results of RT-PCR showed that expression of the nar genes was strongly induced under high pressure. In contrast, P. stutzeri(T) showed no such response following a shift in growth pressure. These results suggest that MT-1 possesses a special mechanism for adaptation to the low-temperature and high-pressure environments of the deep sea, and that Nar is the main dissimilatory nitrate reductase in MT-1 in such environments.

The extracellular glucosyltransferases (GTFs) of Streptococcus mutans are not secreted into the periplasmic space of Escherichia coli when the corresponding gtf genes are isolated in the latter organism. The utilization of both deletion analysis and gtfB: phoA fusions indicate that the signal sequences of the GTFs are functional in E. coli. However, these results further suggest that amino acid sequences present in the carboxyl terminus of the GTFs inhibit secretion through the cytoplasmic membrane in E. coli.

The expression of Streptococcus mutans mutant glucosyltransferase-I enzymes in S. sanguis and S. milleri suggests that cell-associated glucosyltransferase activity is dependent upon both glucan synthesis and glucan binding by the carboxyl-terminal repeating units of the enzyme. Mutant enzymes lacking these repeating units were only present in the extracellular fluids of these transformed streptococcal strains.

We obtained DNA fragments encoding putative aminotransferases possibly involved in the biosynthesis of aminoglycoside antibiotics from deep-sea sediments of the northwest Pacific Ocean by nested PCR, and 34 individual genes (total 89 clones) were identified. About half of the deep-sea sequences showed similarity with genes of known aminoglycoside-producers, but others were deep-sea specific genes. Furthermore, we found that temperature-gradient gel electrophoresis (TGGE) can be an effective tool in the analysis of these DNA fragments.

Austenitic stainless steels suffer intergranular attack in boiling nitric acid with oxidants. The intergranular corrosion is mainly caused by the segregation of impurities to grain. An extra high purity austenitic stainless steel (EHP alloys) was developed with conducting the new multiple refined melting technique in order to suppress the total harmful impurities less than 100ppm. The basically corrosion behavior of type 310 EHP alloy with respect to nitric acid solution with highly oxidizing ions was investigated. The straining, aging and recrystallizing (SAR) treated type 310 EHP alloy showed superior corrosion resistance for intergranular attack. The segregated boron along the grain boundaries was one of main factor of intergranular corrosion from fission track etching results. The SAR treatment was effective to restrain the intergranular attack for type 310 EHP alloy with B less than 7ppm.Copyright © 2008 by ASME

Summary: The nucleotide sequence of the Streptococcus mutans GS-5 gtfD gene coding for the glucosyltransferase which synthesizes water-soluble glucan (GTF-S) has been determined. The complete gene contains 4293 base pairs and the unprocessed protein is composed of 1430 amino acids with a molecular mass of 159814 Da. The amino terminus of the unprocessed protein resembles the signal sequences of other extracellular proteins secreted by S. mutans and that of the GTF-I secreted by Streptococcus downei. In addition, the GTF-S protein exhibits high amino acid similarity with the strain GS-5 enzymes responsible for insoluble glucan synthesis (GTF-I, GTF-SI) previously isolated and sequenced in this laboratory. These results indicate that all three gtf genes evolved from a common ancestral gene.

To understand reductive genome evolution (RGE), we comparatively analyzed the recently reported small genomes of two chemoautotrophic, intracellular symbionts of deep-sea clams, Calyptogena okutanii and C. magnifica. Both genomes lack most genes for DNA recombination and repair such as recA and mutY. Their genome architectures were highly conserved except one inversion. Many deletions from small (

Shewanella violacea DSS12 is facultative piezophile isolated from the deep-sea. The expression of cydDC genes (required for d-type cytochrome maturation) of the organism is regulated by hydrostatic pressure. In this study, we analyzed the nucleotide sequence upstream of cydDC in detail and found that there are putative binding sites for the NarL protein which is part of a two-component regulatory system also containing the sensor protein NarX. Furthermore, we identified the narQP genes (homologues of narXL) from S. violacea DSS12 and demonstrated the heterologous expression of narP in Escherichia coli. These results will be helpful in examining pressure regulation of gene expression in S. violacea at the molecular level.

The nap gene cluster encoding periplasmic nitrate reductase was identified from Pseudomonas sp. strain MT-1, a deep-sea denitrifier isolated from the Mariana Trench. The ORFs identified were highly homologous with those of Pseudomonas stutzeri, but the cluster included only four ORFs (napDABC), less than those in other organisms. For other bacteria, some additional small ORFs (such as napE, napF, napG, napH, and napK) are found in the nap gene cluster, although their physiological function is still unclear. The soluble fraction of MT-1 grown under denitrifying condition showed significant nitrate reductase activity. This observation suggests that the periplasmic nitrate reductase encoded by the gene cluster identified in this study is functional. The activity was highest when the organism was grown under denitrifying conditions, suggesting that the enzyme participates in dissimilatory nitrite reduction.

An obligately piezophilic bacterium was isolated from sediment collected from the bottom of the Japan Trench at a depth of 7542 m. The isolated strain, designated K41GT, was closely affiliated with members of the genus Psychromonas on the basis of 16S rRNA gene sequence analysis. Levels of DNA–DNA relatedness between strain K41GT and Psychromonas reference strains were significantly lower than that accepted as the phylogenetic definition of a species. The optimal temperature and pressure for growth of strain K41GT were 6 °C and 60 MPa, respectively. The DNA G+C content was 39.1 mol%. Whole-cell fatty acids consisted of significant amounts of unsaturated fatty acids C16 : 1 (37 %) and C14 : 1 (17 %), saturated fatty acid C16 : 0 (31 %) and polyunsaturated fatty acid C22 : 6 (8 %). Based on the taxonomic differences observed, strain K41GT is considered to represent a novel obligately piezophilic Psychromonas species. The name Psychromonas hadalis (type strain, K41GT=JCM 11830T=ATCC BAA-638T) is proposed. This is the second species of obligately piezophilic bacteria to be proposed in the genus Psychromonas.

INTRODUCTION Allogeneic hematopoietic cell transplantation (HCT) is an effective therapeutic option for adults with acute myelogenous leukemia (AML) and myelodysplastic syndrome (MDS) after the conventional cyclophosphamide and total-body irradiation (CY/TBI) regimen, but post-transplant relapse is still of high importance. High-dose cytarabine (HDCA) can be added to CY/TBI for an intensified regimen; however, its additional effects have not yet been completely elucidated. METHODS We conducted a cohort study to compare the prognosis of HDCA/CY/TBI and CY/TBI in HCT for AML/MDS, using a Japanese transplant registry database. Inclusion criteria are adult patients (aged 16 years or older) who underwent a single-unit cord blood transplantation (CBT) or bone marrow or peripheral blood stem cell transplantation (BMT/PBSCT) as a first HCT after CY/TBI (CY, 60 mg/kg for 2 days; TBI, 10-12 Gy divided into 4-6 fractions) or CY/TBI plus HDCA (HDCA/CY/TBI) between January 2000 and December 2012. HDCA was defined as cytarabine administration of 2 to 3 g/m2 twice a day for 2 to 3 days (total dose, 6-12 g/m2) before HCT. Overall survival (OS) was calculated with the Kaplan-Meier method and compared using log-rank tests for each covariant related to pre-transplant patient characteristics. Factors with significance or borderline significance (p < 0.1) in the univariate analysis were subjected to a multivariate analysis using the Cox proportional hazards model. The Fine-Gray proportional hazards model was used in multivariate analyses for tumor-related mortality and non-relapse mortality (NRM). Our protocol complied with the Declaration of Helsinki, and it was approved by the Registry Data Management Committee and by the Ethics Committee of Kyoto University, where the study was performed. Written informed consent was obtained from each patient at each institution. RESULTS We analyzed data on 3,031 patients in total, composed of the HDCA/CY/TBI group (n = 1,052) and the CY/TBI group (n = 1,979) after CBT (n = 929) and BMT/PBSCT (n= 2,102). The median age was 38 years, and 86.1% of the patients had AML; high-risk disease was observed in 55.5% of the patients. In BMT/PBSCT, unrelated donors comprised 54.6%, and 63.9% of donors were HLA-matched. The median follow-up period after HCT was approximately 3.2 years. In CBT, OS was significantly superior in the HDCA/CY/TBI group compared to the CY/TBI group (adjusted hazard ratio [HR], 0.56; 95% confidence interval [CI], 0.45-0.69; p < 0.01), while in BMT/PBSCT improvement of OS was not observed in the HDCA/CY/TBI group (HR, 1.14; p = 0.13) (refer to Figure). Tumor-related mortality was lower in HDCA/CY/TBI than CY/TBI after CBT (HR, 0.50; p < 0.01); in BMT/PBSCT, however, post-transplant relapse and tumor-related mortality were not suppressed by the addition of HDCA (HR, 0.89 and 0.90; p = 0.58 and 0.34, respectively). The incidence of grade II to IV acute graft-versus-host disease (GVHD) and chronic GVHD was significantly higher in the HDCA/CY/TBI group than the CY/TBI group after CBT (HR, 1.33 and 2.30; p = 0.02 and < 0.01, respectively), whereas there were no differences in BMT/PBSCT (HR, 0.95 and 1.04; p = 0.54 and 0.64, respectively). Incidence of infectious episodes (bacterial, viral, and fungal) showed no significant differences between regimens in both donor sources. Hemorrhagic cystitis and thrombotic microangiopathy increased in HDCA/CY/TBI compared to CY/TBI after BMT/PBSCT (HR, 1.47 and 1.60; p = 0.06 and 0.04, respectively), leading to significantly higher NRM in this group (HR, 1.48, p < 0.01). On the other hand, NRM was not increased by the addition of HDCA after CBT (HR, 0.94; p = 0.73). CONCLUSION This study is the first to show the differences between HDCA/CY/TBI and CY/TBI after HCT for AML/MDS using a large cohort. Superiority of HDCA/CY/TBI to CY/TBI in CBT was clearly indicated, while these merits were not applied to BMT/PBSCT. The higher incidence of GVHD and stronger graft-versus-leukemia (GVL) effects may be involved in these differences between donor sources (BMT/PBSCT compared to CBT). A large-scale prospective study is warranted to validate these results and to establish the suitable conditioning regimens including HDCA administration. Figure 1. Figure 1. Disclosures No relevant conflicts of interest to declare.

The purpose of this study was to develop novel methods for attachment and cultivation of specifically positioned single yeast cells on a microelectrode surface with the application of a weak electrical potential. Saccharomyces cerevisiae diploid strains attached to an indium tin oxide/glass (ITO) electrode to which a negative potential between −0.2 and −0.4 V vs. Ag/AgCl was applied, while they did not adhere to a gallium-doped zinc oxide/glass electrode surface. The yeast cells attached to the negative potential-applied ITO electrodes showed normal cell proliferation. We found that the flocculin FLO10 gene-disrupted diploid BY4743 mutant strain (flo10Δ /flo10Δ) almost completely lost the ability to adhere to the negative potential-applied ITO electrode. Our results indicate that the mechanisms of diploid BY4743 S. cerevisiae adhesion involve interaction between the negative potential-applied ITO electrode and the Flo10 protein on the cell wall surface. A combination of micropatterning techniques of living single yeast cell on the ITO electrode and omics technologies holds potential of novel, highly parallelized, microchip-based single-cell analysis that will contribute to new screening concepts and applications., Yeast Saccharomyces cerevisiae were selectively attached on the negative potential-applied indium tin oxide/glass electrode. Mechanisms of the yeast cell attachment involve Flocculin Flo10 proteins., Graphical Abstract Figure. Yeast Saccharomyces cerevisiae were selectively attached on the negative potential-applied indium tin oxide/glass electrode. Mechanisms of the yeast cell attachment involve Flocculin Flo10 proteins.

The structural materials of the instruments used in Fukushima Daiichi nuclear power station are exposed to the corrosive condition under the gamma-ray irradiation. In order to determine the effects of the temperature of water under gamma-ray irradiation on the corrosion potential of stainless steel, the measurements of the corrosion potential under gamma-ray irradiation condition was performed. The dissolved oxygen concentration was aerated condition in each temperature in this study. And the corrosive condition under the irradiation was evaluated by the water radiolysis calculation and the measurements of the included hydrogen peroxide in the water after the irradiation. As parameters, the temperature of water and the gamma-ray dose rate were changed. The major results are listed as follows; (1) The corrosion potentials rose about 400 mV by the gamma-ray irradiation in all temperature conditions. (2) The potential was low with higher temperature under irradiation with the aerated condition. It was assumed that the lower dissolved oxygen concentration with higher temperature will cause the dependency of the corrosion potential on the temperature under irradiation. (3) The generation of some ppm of hydrogen peroxide was confirmed by the measurements of hydrogen peroxide concentration. It was indicated that the increase of the corrosion potential by the irradiation was caused by the generation of the high concentration of hydrogen peroxide. (4) It was confirmed that the generation of hydrogen peroxide was lower in higher temperature condition due to the lower dissolved oxygen concentration in high temperature. It was indicated that lower potential in high temperature water was caused by not only lower dissolved oxygen but also lower generation of hydrogen peroxide.

Shewanella violacea DSS12 is a psychrophilic facultative piezophile isolated from the deep sea. In a previous study, we have shown that the bacterium adapted its respiratory components to alteration in growth pressure. This appears to be one of the bacterial adaptation mechanisms to high pressures. In this study, we measured the respiratory activities of S. violacea grown under various pressures. There was no significant difference between the cells grown under atmospheric pressure and a high pressure of 50 MPa relative to oxygen consumption of the cell-free extracts and inhibition patterns in the presence of KCN and antimycin A. Antimycin A did not inhibit the activity completely regardless of growth pressure, suggesting that there were complex III-containing and -eliminating pathways operating in parallel. On the other hand, there was a difference in the terminal oxidase activities. Our results showed that an inhibitor- and pressure-resistant terminal oxidase was expressed in the cells grown under high pressure. This property should contribute to the high-pressure adaptation mechanisms of S. violacea.

Chronic myelogenous leukemia (CML) is a hematological malignancy that is characterized by the chromosome anomaly, t(9;22)(q34;q11). By this chromosomal translocation, a novel activated tyrosine kinase, BCR-ABL chimeric protein, is generated, and the protein is causative of the disease. Recently, Imatinib mesylate targeting to a BCR-ABL chimeric protein has been developed, and shown to achieve complete remission at a high rate. Patients are currently required to receive a fixed dose, 400 mg daily; however, it is possible that some of patients can maintain their remission with reduced doses of imatinib. In this study, we determined levels of BCR-ABL transcript in CML patients by real-time quantitative polymerase chain reaction analysis, and explored the possibility of individualization of therapeutic doses of imatinib. Thirty-five CML patients, including 17 newly diagnosed patients, 16 patients pre-treated with interferon-alpha, and 2 relapsed patients after allogeneic transplantation, were treated with imatinib. Complete cytogenetic response was achieved in 31 (89%) patients. Major molecular response (MMR) was achieved in 21 (60%). Complete molecular response (CMR) was achieved in 7 (20%). Imatinib was discontinued in 2 patients, one patient with MMR due to noncompliance and other patient sustaining CMR, but both patients relapsed 7 and 13 months later, respectively. The doses of imatinib were reduced in 7 patients due to its side effects, but 4 out of the 7 patients have sustained MMR, and 2 of them have sustained CMR for more than 23 months. These results indicate that some patients are able to maintain MMR with low-dose imatinib.

Nitrification and denitrification are bacterial functions, which are important for the global nitrogen cycle. Thus, it is important to study the diversity and distribution of bacteria in the environment, which are involved in the nitrogen cycle on the earth. Ammonia monooxygenase encoded by the amoA gene and nitrite reductase encoded by nirK or nirS are essential enzymes for nitrificaton and denitrification, respectively. These genes can be used as markers for the identification of organisms in the nitrogen cycle. In this study, we identified amoA (42 clones) and nirS (98 clones) genes in parallel from samples recovered from the deep-sea of the Nankai Trough. Genes for nirK could not be amplified from these samples. The obtained amoA sequences were not so closely related to those of amoA genes from previously isolated environmental organisms and those of genes from environmental DNAs. On the other hand, the nirS genes sequenced showed some relationship to some extent with the latter genes. However, some of the newly sequenced genes formed clusters, which contained no previously identified genes on a phylogenetic tree. These are likely present in specific denitrifiers from the deep-sea. The results of this study further suggest that nitrifiers and denitrifiers live in the same area of the Nankai Trough and the nitrogen cycle exists even in the deep-sea.

Microbial communities in Calyptogena sediment and microbial mats of Sagami Bay, Japan, were characterized using 16S rRNA gene sequencing and lipid biomarker analysis. Characterization of 16S rRNA gene isolated from these samples suggested a predominance of bacterial phylotypes related to Gammaproteobacteria (57-64%) and Deltaproteobacteria (27-29%). The Epsilonproteobacteria commonly found in cold seeps and hydrothermal vents were only detected in the microbial mat sample. Significantly different archaeal phylotypes were found in Calyptogena sediment and microbial mats; the former contained only Crenarchaeota clones (100% of the total archaeal clones) and the latter exclusively Euryarchaeota clones, including the anaerobic oxidation of methane archaeal groups ANME-2a and ANME-2c. Many of these lineages are as yet uncultured and undescribed groups of bacteria and archaea. Phospholipid fatty acid analysis suggested the presence of sulphate-reducing and sulphur-oxidizing bacteria. Results of intact glyceryl dialkyl glyceryl tetraether lipid analysis indicated the presence of nonthermophilic marine planktonic archaea. These results suggest that the microbial community in the Sagami Bay seep site is distinct from previously characterized cold-seep environments.

A piezotolerant, mesophilic, marine lactic acid bacterium (strain LT20T) was isolated from a deep sub-seafloor sediment core collected at Nankai Trough, off the coast of Japan. Cells were Gram-positive, rod-shaped, non-sporulating and non-motile. The NaCl concentration range for growth was 0–120 g l−1, with the optimum at 10–20 g l−1. The temperature range for growth at pH 7·0 was 4–50 °C, with the optimum at 37–40 °C. The optimum pH for growth was 7·0–8·0. The optimum pressure for growth was 0·1 MPa with tolerance up to 30 MPa. The main cellular phospholipids were phosphatidylglycerols (25 %), diphosphatidylglycerols (34 %) and a group of compounds tentatively identified as ammonium-containing phosphatidylserines (32 %); phosphatidylethanolamines (9 %) were minor components. The fatty acid composition was dominated by side chains of 16 : 0, 14 : 0 and 16 : 1. The G+C content of the genomic DNA was 42 mol%. On the basis of 16S rRNA gene sequence analysis and the secondary structure of the V6 region, this organism was found to belong to the genus Marinilactibacillus and was closely related to Marinilactibacillus psychrotolerans M13-2T (99 %), Marinilactibacillus sp. strain MJYP.25.24 (99 %) and Alkalibacterium olivapovliticus strain ww2-SN4C (97 %). Despite the high similarity between their 16S rRNA gene sequences (99 %), the DNA–DNA hybridization levels were less than 20 %. On the basis of physiological and genetic characteristics, it is proposed that this organism be classified as a novel species, Marinilactibacillus piezotolerans sp. nov. The type strain is LT20T (=DSM 16108T=JCM 12337T).

The genes of cytochrome bd-encoding cydAB were identified from a deep-sea bacterium Shewanella violacea DSS12. These showed significant homologies with known cydAB gene sequences from various organisms. Additionally, highly conserved regions that are important for the enzymatic function were also conserved in cydA of S. violacea. Based on the results, transcriptional analysis of cydAB operon and cydDC operon (required for assembly of cytochrome bd) of S. violacea in microaerobic condition was performed under the growth condition of various pressures. The gene of cydA was expressed even under the condition of atmospheric pressure and its expression was enhanced with pressurization. On the other hand, the expression of cydC was strongly depressed under the condition of atmospheric pressure compared with the case under high pressure. It appeared spectrophotometrically that loss of cytochrome bd in S. violacea under atmospheric pressure shown in previous study is caused mainly by the loss of cydDC. Further, under the growth condition of atmospheric pressure, either less amount or no d-type cytochrome was expressed compared with the case of high-pressure condition even if the organism was grown under alkaline condition or in the presence of uncoupler, which are the inducible condition of d-type cytochrome in Escherichia coli. These results suggested that the significant amount of d-type cytochrome expression is specific event under the growth condition of high pressure.

A novel piezotolerant, mesophilic, facultatively anaerobic, organotrophic, polarly flagellated bacterium (strain LT13aT) was isolated from a deep sediment layer in the Nankai Trough (Leg 190, Ocean Drilling Program) off the coast of Japan. This organism used a wide range of organic substrates as sole carbon and energy sources: pyruvate, glutamate, succinate, fumarate, lactate, citrate, peptone and tryptone. Oxygen, nitrate, fumarate, ferric iron and cystine were used as electron acceptors. Maximal growth rates were observed at a hydrostatic pressure of 10 MPa. Hydrostatic pressure for growth was in the range 0·1–50 MPa. Predominant cellular fatty acids were 16 : 1ω7c, 15 : 0 iso, 16 : 0 and 13 : 0 iso. The G+C content of the DNA was 44·9 mol%. On the basis of 16S rRNA gene sequences, strain LT13aT was shown to belong to the γ-Proteobacteria, being closely related to Shewanella putrefaciens (98 %), Shewanella oneidensis (97 %) and Shewanella baltica (96 %). Levels of DNA homology between strain LT13aT and S. putrefaciens, S. oneidensis and S. baltica were T represents a novel species. Genetic evidence and phenotypic characteristics showed that isolate LT13aT constitutes a novel species of the genus Shewanella. Because of the deep origin of the strain, the name Shewanella profunda sp. nov. is proposed, with LT13aT (=DSM 15900T=JCM 12080T) as the type strain.

Two strains of obligately piezophilic bacteria were isolated from sediment collected from the bottom surface of a small canyon on the seaward slope of the Japan Trench at a depth of 6278 m. The isolated strains, Y223GT and Y251E, are closely affiliated with members of the genus Colwellia on the basis of 16S rRNA gene sequence analysis. The G+C contents of both strains were about 39 mol%. DNA–DNA hybridization values between these strains and Colwellia reference strains were significantly lower than those accepted as the phylogenetic definition of a species. The novel strains are Gram-negative, polarly flagellated and facultatively anaerobic. The optimal pressure for growth was 60 MPa at both 4 and 10 °C; the most rapid growth rate was observed at 10 °C and 60 MPa. No growth occurred at 15 °C under any pressure studied. The major isoprenoid quinone is Q-8. The predominant cellular fatty acids are C16 : 0 and C16 : 1. Based on the taxonomic differences observed, the isolated strains appear to represent a novel obligately piezophilic Colwellia species. The name Colwellia piezophila sp. nov. (type strain Y223GT=JCM 11831T=ATCC BAA-637T) is proposed.

Hydrostatic pressure is a well-known physical stimulus, but its effects on cell physiology have not been clarified. To investigate pressure effects on Escherichia coli, we carried out DNA microarray analysis of the entire E. coli genome. The microarray results showed pleiotropic effects on gene expression. In particular, heat- and cold-stress responses were induced simultaneously by the elevated pressure. Upon temperature stress (including both temperature up- and down-shifts) and other environmental stresses, gene expression adjusts to adapt to such environmental changes through regulations by several DNA-binding proteins. An E. coli mutant, which deleted the hns gene encoding one of the regulator proteins, exhibited great pressure sensitivity. The result suggested that the H-NS protein was a possible transcriptional regulator for adaptation of the high-pressure stress.

RNA polymerase was purified from the piezophile Shewanella violacea DSS12, and the transcriptional activity after pressure treatment was compared with that of the mesophile Escherichia coli. Application of pressure at 100 MPa for 30 min reduced the E. coli RNA polymerase activity to 60% of the activity at atmospheric pressure, whereas the S. violacea RNA polymerase maintained full activity, indicating that the S. violacea RNA polymerase is more stable than its E. coli counterpart. This result was supported by the analysis of the strength of subunit interactions of the enzyme from both species, using a high-pressure electrophoresis apparatus, which showed that a pressure of 140 MPa caused dissociation of E. coli RNA polymerase but not that of S. violacea RNA polymerase. On the other hand, the core enzyme of S. violacea RNA polymerase, which lacked the sigma70 factor, was dissociated at 140 MPa. These results suggest that the sigma70 factor is required for stabilization of S. violacea RNA polymerase under high-pressure conditions. In this paper, we provide in vitro evidence for piezoadaptation at the transcriptional level, using purified RNA polymerase from cells of S. violacea and E. coli.

To elucidate the mechanism for the regulation of aspartate kinase (AK) via feedback inhibition, we constructed several chimeric enzymes between Bacillus subtilis AK II, a lysine-sensitive mesophilic enzyme, and Thermus flavus AK, a threonine-sensitive thermostable enzyme, each having the same α 2 β 2 -type tetrameric structure. A chimeric AK, named BTT, composed of the chimeric α subunit that comprises of the N-terminal catalytic region from B. subtilis AK II and the C-terminal region from T. flavus , and the β subunit from T. flavus , was inhibited only by threonine. Another chimeric enzyme, BT, which has a similar structure to that of BTT but lacks the β subunit, having α 2 -type homo-dimeric structure, was also responsive only to threonine. However, the addition of threonine enhanced the activity of BT. These results indicate the regulatory function of C-terminal region and β subunit in AK. BTT showed extremely high thermostability comparable to that of T. flavus , suggesting that the β subunit also contributed to the stability of the AK.

Tryptophan uptake appears to be the limiting factor in growth of tryptophan auxotrophic Saccharomyces cerevisiae strains under the conditions of high hydrostatic pressure and low temperature. When the cells are subjected to a pressure of 25 MPa, tryptophan permease Tat2 is degraded in a manner dependent on ubiquitination by Rsp5. One of the high-pressure growth-conferring genes, HPG2, was shown to be allelic to TAT2. The HPG2-1 (Tat2(E27F)) mutation site is located within the ExKS motif in the N-terminus, and the HPG2-2 (Tat2(D563N)) and HPG2-3 (Tat2(E570K)) mutation sites are located at the KQEIAE sequence in the C-terminus. The HPG2 mutations enhance the stability of Tat2 during high-pressure or low-temperature incubation, leading to cell growth under these stressful conditions. These results suggest that the cytoplasmic tails are involved in Rsp5-mediated ubiquitination of Tat2 under high-pressure or low-temperature conditions.