Methylomarinum vadi gen. nov., sp. nov., a methanotroph isolated from two distinct marine environments.

Two aerobic methane-oxidizing bacterial strains were isolated from distinct marine environments in Japan. Strains IT-4(T) and T2-1 were Gram-stain-negative, aerobic, motile, plump short rods or oval-shaped bacteria with a single polar flagellum and type I intracytoplasmic membranes. They were obligate methanotrophs that grew only on methane or methanol. Each strain possessed the particulate methane monooxygenase (pMMO). The ribulose monophosphate pathway was operative for carbon assimilation. The strains grew best at 37 °C, and did not grow at 45 °C. NaCl was required for growth within a concentration range of 1-8 % (w/v). The major phospholipid fatty acids were C16 : 0, C16 : 1ω7c, and C16 : 1ω5t. The major isoprenoid quinone was MQ-8. The DNA G+C content was 50.9-51.7 mol%. The 16S rRNA gene sequences of the strains showed 99.4 % similarity to each other, and DNA-DNA hybridization analysis indicated that the strains were representatives of the same species. The 16S rRNA gene sequences were highly similar to some marine environmental sequences (94.0-97.7 % similarity), but did not show similarities more than 94 % with sequences of members of other related genera, such as Methylomicrobium, Methylobacter, Methylomonas and Methylosarcina. Phylogenies based on 16S rRNA gene sequences and deduced partial PmoA sequences, and the physiological and chemotaxonomic characteristics revealed that strains IT-4(T) and T2-1 represent a novel species of a new genus in the family Methylococcaceae, for which the name Methylomarinum vadi gen. nov., sp. nov. is proposed. The type strain is IT-4(T) ( = JCM 13665(T) = DSM 18976(T)).

Complete Genome Sequence of a Chemolithoautotrophic Iron-Oxidizing Bacterium, Acidithiobacillus ferrooxidans Strain NFP31, Isolated from Volcanic Ash Deposits on Miyake-Jima, Japan.

The genome sequence of Acidithiobacillus ferrooxidans strain NFP31, which is a chemolithoautotrophic iron-oxidizing bacterium that inhabits acidified volcanic deposits on Mount Oyama, Miyake Island (Miyake-jima), Japan, was determined to identify the genetic characteristics associated with pioneer microbes in newly placed pyroclastic deposits.

Altered expression of CDX-2, PDX-1 and mucin core proteins in "Ulcer-associated cell lineage (UACL)" in Crohn's disease.

The ulcer-associated cell lineage (UACL) induced at the site of ileac chronic ulceration in Crohn's disease has been reported to show histological differentiation resembling gastric pyloric mucosa. To clarify the significance of homeobox gene-encoded transcription factors in the formation of the UACL in Crohn's disease, we investigated the immunohistochemical expression of gastrointestinal mucins (MUC5AC for gastric surface mucous cells; MUC6 for gastric gland mucous cells, and MUC2 for intestinal goblet cells) and homeobox gene-encoded transcription factors (CDX-2 for intestinal mucosa and PDX-1 for pyloric mucosa) in the UACL. The analysis was undertaken on ileal mucosa obtained from ileal resections performed in 19 patients with active Crohn's disease of the small bowel. The UACL was observed in nine patients. In the UACL, expression of mucous cells with a foveolar-structure showed immunoreactivity to MUC5AC, and the mucous cells with a glandular structure showed immunoreactivity to MUC6, and the expression of MUC2 was decreased. In addition, we detected the decreased expression of CDX-2 along with the increased expression of PDX-1 in the UACL. The UACL showed histological differentiation simulating gastric pylori mucosa. The down-regulation of CDX-2 and the up-regulation of PDX-1 could be an important mechanism in the induction of the UACL.

Soluble and particulate methane monooxygenase gene clusters in the marine methanotroph Methylomicrobium sp. strain NI.

Soluble methane monooxygenase (sMMO) and particulate methane monooxygenase (pMMO) gene clusters in the marine methanotroph Methylomicrobium sp. strain NI were completely sequenced and analysed. Degenerated primers were newly designed and used to amplify the gene fragments containing intergenic mmoX-Y and mmoD-C regions and a partial pmoC region. Phylogenetic analysis of amino acid sequences deduced from mmoX and pmoA, as well as of 16S rRNA gene sequences, indicated that this strain was most closely related to the halotolerant methanotroph Methylomicrobium buryatense. There were putative sigma(54)- and sigma(70)-dependent promoter sequences upstream of the sMMO and pMMO genes, respectively, and mmoG, which is known to be related to the expression and assembly of sMMO, existed downstream of the sMMO genes. These findings suggest that the major components and regulation of MMOs in this marine methanotroph are quite similar to those in freshwater methane oxidizers, despite the difference in their habitats.

Methylenetetrahydrofolate reductase genotype, vitamin B12, and folate influence plasma homocysteine in hemodialysis patients.

Hyperhomocysteinemia, a well-recognized cardiovascular risk factor, is frequent in hemodialysis (HD) patients. A common polymorphism in the 5,10-methylenetetrahydrofolate reductase (MTHFR) gene, C-->T substitution at nucleotide 677, is associated with homocysteine (Hcy) level elevation. We examined whether three factors involved in the methionine cycle could influence plasma Hcy concentrations in HD patients: MTHFR polymorphism; vitamin B12, an essential cofactor; and folate, the substrate. In a cross-sectional study, serum vitamin B12, folate, and plasma Hcy were measured and MTHFR genotyping was performed in 534 HD patients. Effects of MTHFR genotypes, vitamin B12, and folate on plasma Hcy levels were examined in 450 HD patients not administered vitamin B12 or folate. To examine the effect of vitamin B12 on plasma Hcy concentrations, we compared plasma Hcy concentrations in HD patients with and without vitamin B12 supplementation. To examine whether functional vitamin B12 deficiency exists even in HD patients with normal vitamin B12 concentrations, 15 HD patients (serum vitamin B12 concentrations, 250 to 2,100 pg/mL) were treated with vitamin B12 (mecobalamin, 1.5 mg/d) for 8 weeks. Serum concentrations of methylmalonic acid (MMA) and vitamin B12 were measured. Hcy levels were higher and folate levels were lower in patients with the TT and CT genotypes compared with patients with the CC genotype. Analysis of covariance to determine independent predictors of high Hcy levels identified low serum vitamin B12 and folate levels and high albumin (Alb) levels in CC-genotype patients, low folate levels and high Alb levels in CT-genotype patients, and low folate levels in TT-genotype patients. Plasma Hcy levels were lower in CC- and CT-genotype patients with vitamin B12 supplementation than in those without supplementation. Vitamin B12 supplementation for 8 weeks significantly reduced MMA concentrations in HD patients with normal serum vitamin B12 concentrations. These results indicate that MTHFR genotype influences the correlation of Hcy level with vitamin B12 and folate levels in HD patients. Functional vitamin B12 deficiency may exist, even in HD patients with normal vitamin B12 concentrations. The efficacy of vitamin B12 and folate supplementation on plasma Hcy levels may depend on MTHFR genotype.

Isolation of two novel marine ethylene-assimilating bacteria, Haliea species ETY-M and ETY-NAG, containing particulate methane monooxygenase-like genes.

Two novel ethylene-assimilating bacteria, strains ETY-M and ETY-NAG, were isolated from seawater around Japan. The characteristics of both strains were investigated, and phylogenetic analyses of their 16S rRNA gene sequences showed that they belonged to the genus Haliea. In C1-4 gaseous hydrocarbons, both strains grew only on ethylene, but degraded ethane, propylene, and propane in addition to ethylene. Methane, n-butane, and i-butane were not utilized or degraded by either strain. Soluble methane monooxygenase-type genes, which are ubiquitous in alkene-assimilating bacteria for initial oxidation of alkenes, were not detected in these strains, although genes similar to particulate methane monooxygenases (pMMO)/ammonia monooxygenases (AMO) were observed. The phylogenetic tree of the deduced amino acid sequences formed a new clade near the monooxygenases of ethane-assimilating bacteria similar to other clades of pMMOs in type I, type II, and Verrucomicrobia methanotrophs and AMOs in alpha and beta proteobacteria.

Hyperplastic polyps and sessile serrated 'adenomas' of the colon and rectum display gastric pyloric differentiation.

The serrated polyp-neoplasia pathway is a novel concept that has been demonstrated to differ from the conventional adenoma-carcinoma pathway. To characterize the phenotypic patterns of differentiation in colorectal serrated polyps, we examined the immunohistochemical expression profile of gastric (MUC5AC, TFF1, MUC6, GlcNAcalpha1 --> 4Gal --> R, and PDX1) and intestinal (MUC2, TFF3, and CDX2) epithelial markers in 15 hyperplastic polyps (HPs), 29 sessile serrated adenomas (SSAs),12 traditional serrated adenomas (TSAs), and 16 conventional adenomas (CAs). MUC5AC and TFF1 were upregulated in the HPs, SSAs, and TSAs. MUC6 was expressed in the HPs and SSAs. GlcNAcalpha1 --> 4Gal --> R was expressed only in the SSAs. Although MUC2 expression was preserved, TFF3 was downregulated in the HPs, SSAs, and TSAs. PDX1 was upregulated in the HPs, SSAs, and TSAs. On the other hand, CDX2 was downregulated in the HPs and SSAs. The colorectal serrated polyps showed higher expression of gastric makers than CAs. The HPs and SSAs showed gastric and intestinal mixed phenotype expression with gastric pyloric organoid differentiation and almost identical, but different from the TSAs, marker profile. PDX1 up-regulation and CDX2 down-regulation could be important for the induction of a gastric pyloric pattern of cell differentiation in colorectal serrated polyps.

Novel and diverse integron integrase genes and integron-like gene cassettes are prevalent in deep-sea hydrothermal vents.

The lack of information about mobile DNA in deep-sea hydrothermal vents limits our understanding of the phylogenetic diversity of the mobile genome of bacteria in these environments. We used culture-independent techniques to explore the diversity of the integron/mobile gene cassette system in a variety of hydrothermal vent communities. Three samples, which included two different hydrothermal vent fluids and a mussel species that contained essentially monophyletic sulfur-oxidizing bacterial endosymbionts, were collected from Suiyo Seamount, Izu-Bonin, Japan, and Pika site, Mariana arc. First, using degenerate polymerase chain reaction (PCR) primers, we amplified integron integrase genes from metagenomic DNA from each sample. From vent fluids, we discovered 74 new integrase genes that were classified into 11 previously undescribed integron classes. One integrase gene was recorded in the mussel symbiont and was phylogenetically distant from those recovered from vent fluids. Second, using PCR primers targeting the gene cassette recombination site (59-be), we amplified and subsequently identified 60 diverse gene cassettes. In multicassette amplicons, a total of 13 59-be sites were identified. Most of these sites displayed features that were atypical of the features previously well conserved in this family. The Suiyo vent fluid was characterized by gene cassette open reading frames (ORFs) that had significant homologies with transferases, DNA-binding proteins and metal transporter proteins, while the majority of Pika vent fluid gene cassettes contained novel ORFs with no identifiable homologues in databases. The symbiont gene cassette ORFs were found to be matched with DNA repair proteins, methionine aminopeptidase, aminopeptidase N, O-sialoglycoprotein endopeptidase and glutamate synthase, which are proteins expected to play a role in animal/symbiont metabolism. The success of this study indicates that the integron/gene cassette system is common in deep-sea hydrothermal vents, an environment type well removed from anthropogenic disturbance.

Regulation of the ASC expression in response to LPS stimulation is related to IL-8 secretion in the human intestinal mucosa.

Expression of the caspase-activating adaptor ASC was reported to be associated with the production of IL-8, a primary mediator of mucosal inflammation, in vitro. However, the significance of the ASC-mediated IL-8 production in primary tissues has remained poorly understood. Primary intestinal mucosa isolated from surgically resected ileum or colon was incubated with several concentrations of LPS or left untreated. ASC expression was up-regulated at 2 h after stimulation with low doses of LPS (1-10 ng/ml), and was associated with IL-8 secretion, and then was down-regulated later. In contrast, ASC expression remained at the basal level in mucosal tissue treated with a high dose of LPS (1000 ng/ml). Interestingly, in mucosa from several cases of Crohn's disease, ASC was highly expressed without stimulation, and IL-8 was stably secreted with no regulation by LPS. These findings revealed that ASC expression correlates with IL-8 secretion and may play an important role in maintaining mucosal homeostasis.

A nonconserved carboxy-terminal segment of GroEL contributes to reaction temperature.

The role of the C-terminal segment of the GroEL equatorial domain was analyzed. To understand the molecular basis for the different active temperatures of GroEL from three bacteria, we constructed a series of chimeric GroELs combining the C-terminal segment of the equatorial domain from one species with the remainder of GroEL from another. In each case, the foreign C-terminal segment substantially altered the active temperature range of the chimera. Substitution of L524 of Escherichia coli GroEL with the corresponding residue (isoleucine) from psychrophilic GroEL resulted in a GroE with approximately wild-type activity at 25 degrees C, but also at 10 degrees C, a temperature at which wild-type E. coli GroE is inactive. In a detailed look at the temperature dependence of the GroELs, normal E. coli GroEL and the L524I mutant became highly active above 14 degrees C and 12 degrees C respectively. Similar temperature dependences were observed in a surface plasmon resonance assay of GroES binding. These results suggested that the C-terminal segment of the GroEL equatorial domain has an important role in the temperature dependence of GroEL. Moreover, E. coli acquired the ability to grow at low temperature through the introduction of cold-adapted chimeric or L524I mutant groEL genes.

Fractalkine expression and the recruitment of CX3CR1+ cells in the prolonged mesangial proliferative glomerulonephritis.

BACKGROUND: We established the reversible and the prolonged models of mesangial proliferative glomerulonephritis (GN) with anti-Thy 1 antibody 1-22-3. However, the essential factors leading to the prolonged glomerular alterations have not been identified. METHODS: The expressions of several chemokines and cytokines were compared in the reversible and the prolonged models. Expression of fractalkine and the number of the fractalkine receptor CX3CR1-positive cells in the glomeruli in the prolonged model were significantly higher than those in the reversible model. Then, the localization of fractalkine and the characteristics of CX3CR1+ cells were analyzed in glomeruli. To elucidate the significance of the fractalkine expression, we analyzed the expression in the model treated with angiotensin II receptor antagonist, candesartan. RESULTS: Immunostaining of fractalkine was detected on endothelial cells on the fifth day, and fractalkine staining also was detected in the mesangial area on day 14. Major parts of the CX3CR1+ cells in the glomeruli were macrophages, especially ED3+ cells. Candesartan treatment ameliorated the glomerular morphological findings at six weeks after disease induction. Although the treatment did not ameliorate the morphological finding at two weeks, decreased expression of fractalkine and CX3CR1+ were already detected at two weeks in rats treated with candesartan. CONCLUSIONS: Fractalkine expression and the recruitment of CX3CR1+ cells in glomeruli might play an important role in the development of the prolonged disease. These expressions could be predictors of the prolonged disease of the mesangial proliferative glomerulonephritis.