A mycofactocin-associated dehydrogenase is essential for ethylene glycol metabolism by Rhodococcus jostii RHA1.

Ethylene glycol is an industrially important diol in many manufacturing processes and a building block of polymers, such as poly(ethylene terephthalate). In this study, we found that a mycolic acid-containing bacterium Rhodococcus jostii RHA1 can grow with ethylene glycol as a sole source of carbon and energy. Deletion of a putative glycolate dehydrogenase gene (RHA1_ro03227) abolished growth with ethylene glycol, indicating that ethylene glycol is assimilated via glycolate in R. jostii RHA1. Transcriptome sequencing and gene deletion analyses revealed that a gene homologous to mycofactocin (MFT)-associated dehydrogenase (RHA1_ro06057), hereafter referred to as EgaA, is essential for ethylene glycol assimilation. Furthermore, egaA deletion also negatively affected the utilization of ethanol, 1-propanol, propylene glycol, and 1-butanol, suggesting that EgaA is involved in the utilization of various alcohols in R. jostii RHA1. Deletion of MFT biosynthetic genes abolished growth with ethylene glycol, indicating that MFT is the physiological electron acceptor of EgaA. Further genetic studies revealed that a putative aldehyde dehydrogenase (RHA1_ro06081) is a major aldehyde dehydrogenase in ethylene glycol metabolism by R. jostii RHA1. KEY POINTS: • Rhodococcus jostii RHA1 can assimilate ethylene glycol via glycolate • A mycofactocin-associated dehydrogenase is involved in the oxidation of ethylene glycol • An aldehyde dehydrogenase gene is important for the ethylene glycol assimilation.

A functionally uncharacterized type-2 malate/L-lactate dehydrogenase family protein from Thermus thermophilus HB8 catalyzes stereospecific reduction of 2-keto-3-deoxy-D-gluconate.

2-Keto-3-deoxy- D-gluconate (KDG) is an important intermediate found in various sugars, sugar acids and polysaccharide catabolic pathways. Here, we report that a functionally uncharacterized type-2 malate/L-lactate dehydrogenase family protein (TTHB078) from Thermus thermophilus HB8 catalyzes a novel reaction, NAD(P)H-dependent reductase activity on KDG. This enzyme, designated KdgG, utilizes both NADH and NADPH as electron donors, but higher activity was observed with NADH. Analysis of the reaction product revealed that KdgG catalyzes reversible reduction of KDG to form 3-deoxy-D-mannonate. Molecular phylogenetic analysis indicated that KdgG and its homologs distributed in the genus Thermus form a novel clade among type-2 malate/L-lactate dehydrogenase family proteins.

Construction of a Rhodobacter sphaeroides Strain That Efficiently Produces Hydrogen Gas from Acetate without Poly(ß-Hydroxybutyrate) Accumulation: Insight into the Role of PhaR in Acetate Metabolism.

The purple nonsulfur phototrophic bacterium Rhodobacter sphaeroides produces hydrogen gas (H2) from acetate. An approach to improve the H2 production is preventing accumulation of an intracellular energy storage molecule known as poly(ß-hydroxybutyrate) (PHB), which competes with H2 production for reducing power. However, disruption of PHB biosynthesis has been reported to severely impair the acetate assimilation depending on the genetic backgrounds and/or culture conditions. To solve this problem, we analyzed the relationship between PHB accumulation and acetate metabolism in R. sphaeroides. Gene deletion analyses based on the wild-type strain revealed that among the two polyhydroxyalkanoate synthase genes in the genome, phaC1, but not phaC2, is essential for PHB accumulation, and the phaC1 deletion mutant exhibited slow growth with acetate. On the other hand, a strain with the deletion of phaC1 together with phaR, which encodes a transcriptional regulator capable of sensing PHB accumulation, exhibited growth comparable to that of the wild-type strain despite no accumulation of PHB. These results suggest that PHB accumulation is required for normal growth with acetate by altering the expression of genes under the control of phaR. This hypothesis was supported by a transcriptome sequencing (RNA-seq) analysis revealing that phaR is involved in the regulation of the ethylmalonyl coenzyme A pathway for acetate assimilation. Consistent with these findings, deletion of phaC1 in a genetically engineered H2-producing strain resulted in lower H2 production from acetate due to growth defects, whereas deletion of phaR together with phaC1 restored growth with acetate and increased H2 production from acetate without PHB accumulation. IMPORTANCE This study provides a novel approach for increasing the yield of photofermentative H2 production from acetate by purple nonsulfur phototrophic bacteria. This study further suggests that polyhydroxyalkanoate is not only a storage substance for carbon and energy in bacteria, but may also act as a signaling molecule that mediates bacterial metabolic adaptations to specific environments. This notion will be helpful for understanding the physiology of polyhydroxyalkanoate-producing bacteria, as well as for their metabolic engineering via synthetic biology.

Effect of Sucrose on Amino Acid Absorption of Whey: A Randomized Crossover Trial.

Protein intake has been reported to secrete insulin and lower glucose levels, but the effect of carbohydrate and protein co-ingestion on amino acid absorption has not been well documented. A randomized, placebo-controlled, single-blinded, crossover trial was conducted to evaluate the effect of sucrose on blood amino acid levels. Eleven volunteers (both sexes aged 20-60 years with body mass index 21.4 ± 2.4 kg/m2) randomly received one of four test solutions: water (P-group), 10 g sucrose (S-group), 10 g whey protein (W-group), or 10 g whey protein + 10 g sucrose (W-S-group), and blood amino acid concentration, glucose levels, and insulin levels were monitored over 180 min. Following the wash-out period, randomized treatment and blood parameter monitoring were repeated. Consequently, amino acid concentration was significantly lower in the S-group than in the P-group, showing that single ingestion of sucrose decreased blood amino acid levels in a fasted state. However, there was no significant difference between blood amino acid levels of the W- and W-S-groups, suggesting that co-ingestion of sucrose does not affect blood amino acid concentration. Insulin levels were significantly higher in the W-S than in the S-group, and glucose levels were significantly lower in the W-S- than in the S-group, suggesting positive impact on glycotoxicity by reducing blood glucose levels. Therefore, whey protein co-ingestion with sucrose suppresses glucose levels and increases insulin levels as opposed to the sucrose ingestion, but does not affect amino acid absorption of whey protein, indicating that this co-ingestion may not be a problem for protein supplementation.

Complete Genome Sequence of Kaistia sp. Strain 32K, Isolated from Soil as a Mixed Single Colony with Methylobacterium sp. Strain ME121.

Kaistia sp. strain 32K, an aerobic Gram-negative bacterium, was isolated from soil in Japan. Here, we report the complete genome sequence of this bacterium, which has a 5.4-Mbp genome sequence, containing 4,919 protein-coding sequences.

Metabolites Produced by Kaistia sp. 32K Promote Biofilm Formation in Coculture with Methylobacterium sp. ME121.

Previously, we reported that the coculture of motile Methylobacterium sp. ME121 and non-motile Kaistia sp. 32K, isolated from the same soil sample, displayed accelerated motility of strain ME121 due to an extracellular polysaccharide (EPS) produced by strain 32K. Since EPS is a major component of biofilms, we aimed to investigate the biofilm formation in cocultures of the two strains. The extent of biofilm formation was measured by a microtiter dish assay with the dye crystal violet. A significant increase in the amount of biofilm was observed in the coculture of the two strains, as compared to that of the monocultures, which could be due to a metabolite produced by strain 32K. However, in the coculture with strain 32K, using Escherichia coli or Pseudomonas aeruginosa, there was no difference in the amount of biofilm formation as compared with the monoculture. Elevated biofilm formation was also observed in the coculture of strain ME121 with Kaistia adipata, which was isolated from a different soil sample. Methylobacterium radiotolerans, isolated from another soil sample, showed a significant increase in biofilm formation when cocultured with K. adipata, but not with strain 32K. We also found that the culture supernatants of strains 32K and K. adipata accelerated the motility of strains ME121 and M. radiotolerans, wherein culture supernatant of K. adipata significantly increased the motility of M. radiotolerans, as compared to that by the culture supernatant of strain 32K. These results indicated that there was a positive relationship between accelerated motility and increased biofilm formation in Methylobacterium spp. This is the first study to report that the metabolites from Kaistia spp. could specifically modulate the biofilm-forming ability of Methylobacterium spp. Methylobacterium spp. biofilms are capable of inhibiting the biofilm formation of mycobacteria, which are opportunistic pathogens that cause problems in infectious diseases. Thus, the metabolites from the culture supernatant of Kaistia spp. have the potential to contribute to the environment in which increased biofilm production of Methylobacterium is desired.

A Factor Produced by Kaistia sp. 32K Accelerated the Motility of Methylobacterium sp. ME121.

Motile Methylobacterium sp. ME121 and non-motile Kaistia sp. 32K were isolated from the same soil sample. Interestingly, ME121 was significantly more motile in the coculture of ME121 and 32K than in the monoculture of ME121. This advanced motility of ME121 was also observed in the 32K culture supernatant. A swimming acceleration factor, which we named the K factor, was identified in the 32K culture supernatant, purified, characterized as an extracellular polysaccharide (5-10 kDa), and precipitated with 70% ethanol. These results suggest the possibility that the K factor was directly or indirectly sensed by the flagellar stator, accelerating the flagellar rotation of ME121. To the best of our knowledge, no reports describing an acceleration in motility due to coculture with two or more types of bacteria have been published. We propose a mechanism by which the increase in rotational force of the ME121 flagellar motor is caused by the introduction of the additional stator into the motor by the K factor.

A suicide enzyme catalyzes multiple reactions for biotin biosynthesis in cyanobacteria.

In biotin biosynthesis, the conversion of pimeloyl intermediates to biotin is catalyzed by a universal set of four enzymes: BioF, BioA, BioD and BioB. We found that the gene homologous to bioA, the product of which is involved in the conversion of 8-amino-7-oxononanoate (AON) to 7,8-diaminononanoate (DAN), is missing in the genome of the cyanobacterium Synechocystis sp. PCC 6803. We provide structural and biochemical evidence showing that a novel dehydrogenase, BioU, is involved in biotin biosynthesis and functionally replaces BioA. This enzyme catalyzes three reactions: formation of covalent linkage with AON to yield a BioU-DAN conjugate at the ε-amino group of Lys124 of BioU using NAD(P)H, carboxylation of the conjugate to form BioU-DAN-carbamic acid, and release of DAN-carbamic acid using NAD(P)+. In this biosynthetic pathway, BioU is a suicide enzyme that loses the Lys124 amino group after a single round of reaction.

Engineering the transcriptional activator NifA for the construction of Rhodobacter sphaeroides strains that produce hydrogen gas constitutively.

Purple non-sulfur photosynthetic bacteria such as Rhodobacter sphaeroides and Rhodopseudomonas palustris produce hydrogen gas (H2) via proton reduction, which is catalyzed by nitrogenase. Although the expression of nitrogenase is usually repressed under nitrogen-sufficient conditions, a partial deletion of nifA, which encodes a transcriptional activator of nitrogen-fixation genes, has been reported to enable the constitutive expression of nitrogenase in R. palustris. In this study, we evaluated the effects of a similar mutation (nifA* mutation) on H2 production during the photoheterotrophic growth of R. sphaeroides, based on the notion that H2 production by nitrogenase compensates for the loss of CO2 fixation via the Calvin cycle, thereby restoring the redox balance. The chromosomal nifA* mutation resulted in the slight restoration of the photoheterotrophic growth of a mutant strain lacking ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO), the key enzyme of the Calvin cycle, when the strain was cultured in van Niel's yeast medium. In addition, the strain with the nifA* mutation produced detectable levels of H2 during photoheterotrophic growth with acetate and ammonium; however, the H2 production was considerably lower than that observed during the photoheterotrophic growth of the strain with acetate and L-glutamate, where L-glutamate serves as a poor nitrogen source, thereby causing nitrogenase derepression. On the other hand, introduction of a multicopy plasmid harboring nifA* markedly restored the photoheterotrophic growth of the RubisCO-deletion mutant in van Niel's yeast medium and resulted in efficient H2 production during the photoheterotrophic growth with acetate and ammonium.

Introduction of Glyoxylate Bypass Increases Hydrogen Gas Yield from Acetate and l-Glutamate in Rhodobacter sphaeroides.

Rhodobacter sphaeroides produces hydrogen gas (H2) from organic compounds via nitrogenase under anaerobic-light conditions in the presence of poor nitrogen sources, such as l-glutamate. R. sphaeroides utilizes the ethylmalonyl-coenzyme A (EMC) pathway for acetate assimilation, but its H2 yield from acetate in the presence of l-glutamate has been reported to be low. In this study, the deletion of ccr encoding crotonyl-coenzyme A (crotonyl-CoA) carboxylase/reductase, a key enzyme for the EMC pathway in R. sphaeroides, revealed that the EMC pathway is essential for H2 production from acetate and l-glutamate but not for growth and acetate consumption in the presence of l-glutamate. We introduced a plasmid expressing aceBA from Rhodobacter capsulatus encoding two key enzymes for the glyoxylate bypass into R. sphaeroides, which resulted in a 64% increase in H2 production. However, compared with the wild-type strain expressing heterologous aceBA genes, the strain with aceBA introduced in the genetic background of an EMC pathway-disrupted mutant showed a lower H2 yield. These results indicate that a combination of the endogenous EMC pathway and a heterologously expressed glyoxylate bypass is beneficial for H2 production. In addition, introduction of the glyoxylate bypass into a polyhydroxybutyrate (PHB) biosynthesis-disrupted mutant resulted in a delay in growth along with H2 production, although its H2 yield was comparable to that of the wild-type strain expressing heterologous aceBA genes. These results suggest that PHB production is important for fitness to the culture during H2 production from acetate and l-glutamate when both acetate-assimilating pathways are present.IMPORTANCE As an alternative to fossil fuel, H2 is a promising renewable energy source. Although photofermentative H2 production from acetate is key to developing an efficient process of biohydrogen production from biomass-derived sugars, H2 yields from acetate and l-glutamate by R. sphaeroides have been reported to be low. In this study, we observed that in addition to the endogenous EMC pathway, heterologous expression of the glyoxylate bypass in R. sphaeroides markedly increased H2 yields from acetate and l-glutamate. Therefore, this study provides a novel strategy for improving H2 yields from acetate in the presence of l-glutamate and contributes to a clear understanding of acetate metabolism in R. sphaeroides during photofermentative H2 production.

Structural basis of L-glucose oxidation by scyllo-inositol dehydrogenase: Implications for a novel enzyme subfamily classification.

For about 70 years, L-glucose had been considered non-metabolizable by either mammalian or bacterial cells. Recently, however, an L-glucose catabolic pathway has been discovered in Paracoccus laeviglucosivorans, and the genes responsible cloned. Scyllo-inositol dehydrogenase is involved in the first step in the pathway that oxidizes L-glucose to produce L-glucono-1,5-lactone with concomitant reduction of NAD+ dependent manner. Here, we report the crystal structure of the ternary complex of scyllo-inositol dehydrogenase with NAD+ and L-glucono-1,5-lactone at 1.8 Å resolution. The enzyme adopts a homo-tetrameric structure, similar to those of the inositol dehydrogenase family, and the electron densities of the bound sugar was clearly observed, allowing identification of the residues responsible for interaction with the substrate in the catalytic site. In addition to the conserved catalytic residues (Lys106, Asp191, and His195), another residue, His318, located in the loop region of the adjacent subunit, is involved in substrate recognition. Site-directed mutagenesis confirmed the role of these residues in catalytic activity. We also report the complex structures of the enzyme with myo-inositol and scyllo-inosose. The Arg178 residue located in the flexible loop at the entrance of the catalytic site is also involved in substrate recognition, and plays an important role in accepting both L-glucose and inositols as substrates. On the basis of these structural features, which have not been identified in the known inositol dehydrogenases, and a phylogenetic analysis of IDH family enzymes, we suggest a novel subfamily of the GFO/IDH/MocA family. Since many enzymes in this family have not biochemically characterized, our results could promote to find their activities with various substrates.

Structure and function of an ancestral-type ß-decarboxylating dehydrogenase from Thermococcus kodakarensis.

ß-Decarboxylating dehydrogenases, which are involved in central metabolism, are considered to have diverged from a common ancestor with broad substrate specificity. In a molecular phylogenetic analysis of 183 ß-decarboxylating dehydrogenase homologs from 84 species, TK0280 from Thermococcus kodakarensis was selected as a candidate for an ancestral-type ß-decarboxylating dehydrogenase. The biochemical characterization of recombinant TK0280 revealed that the enzyme exhibited dehydrogenase activities toward homoisocitrate, isocitrate, and 3-isopropylmalate, which correspond to key reactions involved in the lysine biosynthetic pathway, tricarboxylic acid cycle, and leucine biosynthetic pathway, respectively. In T. kodakarensis, the growth characteristics of the KUW1 host strain and a TK0280 deletion strain suggested that TK0280 is involved in lysine biosynthesis in this archaeon. On the other hand, gene complementation analyses using Thermus thermophilus as a host revealed that TK0280 functions as both an isocitrate dehydrogenase and homoisocitrate dehydrogenase in this organism, but not as a 3-isopropylmalate dehydrogenase, most probably reflecting its low catalytic efficiency toward 3-isopropylmalate. A crystallographic study on TK0280 binding each substrate indicated that Thr71 and Ser80 played important roles in the recognition of homoisocitrate and isocitrate while the hydrophobic region consisting of Ile82 and Leu83 was responsible for the recognition of 3-isopropylmalate. These analyses also suggested the importance of a water-mediated hydrogen bond network for the stabilization of the ß3-α4 loop, including the Thr71 residue, with respect to the promiscuity of the substrate specificity of TK0280.

Isolated port-site metastasis of hepatocellular carcinoma after laparoscopic liver resection.

Port-site metastasis of hepatocellular carcinoma (HCC) is extremely rare, and only one case has been reported in the English-language literature. Contamination with malignant cells along the needle tract during percutaneous biopsy or radiofrequency ablation is a well-recognized cause of HCC recurrence. Here, we describe a case of port-site metastasis after laparoscopic liver resection of HCC. The patient, who had undergone laparoscopic partial resection of the left lateral segment of the liver 18 months earlier, was diagnosed with HCC. CT showed a nodule in the abdominal wall where the laparoscopic port had been inserted during resection. Local excision was performed, and histological examination revealed HCC consistent with recurrence after laparoscopic resection. The experience described in this report highlights the risk of port-site metastasis of HCC. Imaging for oncologic surveillance after laparoscopic resection must include all port sites.

C-Reactive Protein/Albumin Ratio and Prognostic Nutritional Index Are Strong Prognostic Indicators of Survival in Resected Pancreatic Ductal Adenocarcinoma.

Purpose: We evaluated the clinical importance, such as the occurrence of postoperative pancreatic fistula (POPF) or prognosis, of preoperative serum markers of chronic inflammation, nutrition, and immunity, as well as that of serum tumor markers after curative resection of pancreatic ductal adenocarcinomas (PDACs). Methods: Between 2006 and 2015, 43 patients with PDACs underwent curative resection at Tottori Prefectural Central Hospital. We analyzed which preoperative indicators (i.e., C-reactive protein/albumin ratio [CAR], neutrophil/lymphocyte ratio [NLR], prognostic nutritional index [PNI], carcinoembryonic antigen [CEA], and carbohydrate antigen 19-9 [CA 19-9]) were the most relevant risk factors for occurrence of POPF and poor patient survival. Results: POPF was detected in 8/43 (18.6%) patients. One patient died of pancreatic fistula at 2 months postoperatively. Among nine candidate factors (operative procedure, operation time, tumor stage, preoperative serum amylase, preoperative CAR, NLR, PNI, CEA, and CA 19-9), we did not identify any significant risk factor for the occurrence of POPF. The 5-year overall survival (OS) rate of the 43 patients was 22.4%, and the overall median survival time was 21 months. The multivariate OS analysis demonstrated that high CAR and low PNI were strong preoperative markers of poor prognosis independently of tumor stage. Conclusions: Preoperative CAR and PNI are useful prognostic markers for patients with operable PDACs.

Aspartate kinase involved in 4-hydroxy-3-nitrosobenzamide biosynthesis in Streptomyces murayamaensis.

Streptomyces murayamensis carries two aspartate kinase (AK) genes: one for the biosynthesis of lysine, threonine, and methionine, and the other (nspJ) contained in the biosynthetic gene cluster for the secondary metabolite, 4-hydroxy-3-nitrosobenzamide, for catalyzing the first reaction. AKs involved in the biosynthesis of amino acids are often regulated allosterically by the end products. In the present study, we characterized NspJ to investigate whether AKs involved in secondary metabolism were also allosterically regulated. NspJ was in α2ß2 and (α2ß2)2 heterooligomeric forms, and was insensitive to all the compounds tested including lysine, threonine, and methionine. The reduction in the activity following the removal of ammonium sulfate, which induced subunit dissociation, suggests that the ß subunit may be involved in stabilizing the structure of the α subunit in order to exhibit its activity. This study has provided the first example of a feedback-insensitive α2ß2-type AK, which is involved in the secondary metabolism.

Crystal Structure of the LysY·LysW Complex from Thermus thermophilus.

Several bacteria and archaea utilize the amino group-carrier protein, LysW, for lysine biosynthesis, in which an isopeptide bond is formed between the C-terminal Glu of LysW and an amino group of α-aminoadipate (AAA). The resulting LysW-γ-AAA is phosphorylated by LysZ to form LysW-γ-AAA phosphate, which is subsequently reduced to LysW-γ-aminoadipic semialdehyde (LysW-γ-AASA) through a reaction catalyzed by LysY. In this study, we determined the crystal structures of LysY from Thermus thermophilus HB27 (TtLysY) complexed with TtLysW-γ-AASA and TtLysW-γ-AAA, respectively. In both structures, the globular domain of TtLysW was recognized by positively charged residues on helix α9 and the ß11-α10 loop of TtLysY through conformational changes. A mutational analysis confirmed that the interactions observed between TtLysY and TtLysW are important for the function of TtLysY. The extended LysW recognition loop and conserved arginine residue were identified as signatures to discriminate LysY from ArgC, which is involved in arginine biosynthesis. Combined with the previously determined TtLysZ·TtLysW complex structure, TtLysW may simultaneously bind TtLysZ and TtLysY. These structural insights suggest the formation of a TtLysWZY ternary complex, in which the flexible C-terminal extension of TtLysW promotes the efficient transfer of the labile intermediate from the active site of TtLysZ to that of TtLysY during the sequential reactions catalyzed by TtLysZY.

Evaluation of prognostic markers for patients with curatively resected thoracic esophageal squamous cell carcinomas.

The Glasgow Prognostic Score (GPS), neutrophil/lymphocyte ratio (NLR) and prognostic nutritional index (PNI) are prognostic parameters for malignancies. Additionally, serum squamous cell carcinoma antigen (SCC-Ag) and cytokeratin 19 fragments (CYFRA 21-1) are tumor markers for squamous cell carcinoma. In the present study, the prognostic importance of these markers in patients with resectable thoracic esophageal cancer was investigated. In this retrospective study, 84 enrolled patients diagnosed with resectable clinical stage I-III thoracic esophageal squamous cell carcinomas (ESCCs) underwent thoracic esophageal resection and three-field lymph node dissection at Tottori University Hospital between January 2007 and December 2013. The correlations among preoperative patient markers (GPS, NLR, PNI, SCC-Ag and CYFRA 21-1) and the occurrence of postoperative complications and patient survival were analyzed. The operative mortality was 2.4%, and morbidity was 42.9%. Strong correlations between occurrence of postoperative complications and open thoracotomy (P=0.083) and high-serum CYFRA 21-1 (P=0.007) were observed. In 15 patients with high-serum CYFRA 21-1, postoperative complications were detected in 11 of them (73.3%); on the other hand, complications occurred in 25 of 69 (36.2%) with low-serum CYFRA 21-1. The 5-year disease-free survival rate and 5-year overall survival rate of all the patients were 52.2 and 50.8%, respectively. Among the prognostic parameters, preoperative high NLR was determined to be a poor prognostic factor, independent of the tumor stage in the multivariate analysis. These results may indicate that, in patients with preoperative high-serum CYFRA 21-1, more attention should be paid to the occurrence of postoperative complications. Therefore, in such cases, anastomosis between blood vessels of the substitute esophagus and cervical vessels would be recommended. Furthermore, in patients with high preoperative NLR, effective adjuvant chemoradiotherapy should be considered to prolong the patients' survival, even of stage I or II patients.

Comparison of single-stapling and hemi-double-stapling methods for intracorporeal esophagojejunostomy using a circular stapler after totally laparoscopic total gastrectomy.

BACKGROUND: Laparoscopic total gastrectomy is not widely performed because of the difficulty of esophagojejunal reconstruction. This study analyzed complication rates of two different methods for reconstruction by a circular stapler after totally laparoscopic total gastrectomy (TLTG). METHODS: Between 2010 and 2014, clinical data of 19 patients who underwent TLTG for gastric adenocarcinoma were collected retrospectively. There were two methods to fix the anvil of a circular stapler into the distal esophagus: In the single-stapling technique (SST) group, Endo-PSI(II) was used for purse-suturing on the distal esophagus for reconstruction, and in the hemi-double-stapling technique (hemi-DST) group, the esophagus was cut by linear stapler with the entry hole of the anvil shaft opened after inserting the anvil tail. In both groups, surgical procedures were the same, except for the reconstruction. RESULTS: All TLTGs were performed securely without mortality. Intracorporeal laparoscopic esophagojejunal anastomosis was performed successfully for all the patients. In the hemi-DST group, four patients experienced anastomotic stenosis, three of whom required endoscopic balloon dilation. In contrast, no stenosis was seen in the SST group (p = 0.033). CONCLUSIONS: Anastomosis with SST is preferred to that with hemi-DST to minimize postoperative complications.

Draft Genome Sequence of Methylobacterium sp. ME121, Isolated from Soil as a Mixed Single Colony with Kaistia sp. 32K.

Methylobacterium sp. ME121 was isolated from soil as a mixed single colony with Kaistia sp. 32K, and its growth was enhanced by coculture. Here, we report the draft genome sequence of Methylobacterium sp. ME121, which may contribute to the study of the molecular mechanisms underlying this phenomenon.

Feasibility study of alternate-day S-1 as adjuvant chemotherapy for gastric cancer: a randomized controlled trial.

BACKGROUND: The Adjuvant Chemotherapy Trial of S-1 for Gastric Cancer established oral S-1 administration for 1 year as the standard postoperative adjuvant chemotherapy for gastric cancer in Japan. We conducted a multicenter cooperative prospective study comparing daily and alternate-day S-1 administration as postoperative adjuvant therapy for gastric cancer. METHODS: Patients with Stage II or III gastric cancer who underwent curative surgery were randomly assigned to receive standard daily S-1 administration [group A: 80-120 mg/day S-1 depending on body surface area (BSA); days 1-28 every 6 weeks for 1 year] or alternate-day administration (group B: 80-120 mg/day S-1 depending on BSA; alternate days for 15 months). Treatment completion rate was the primary endpoint, and relative dose intensity and safety, overall survival, and relapse-free survival (RFS) were secondary endpoints. RESULTS: Seventy-three patients were enrolled. The treatment completion rate was 72.2 % in group A and 91.8 % in group B; the relative dose intensity was 67.5 % in group A and 81.2 % in group B; and compliance was better in group B. Digestive system adverse effects were less frequent in group B than in group A. Median follow-up time was 2.8 years; 3-year survival rate was 69.6 % in group A and 87.3 % in group B; and 3-year RFS rate was 76.4 % in group A and 73.1 % in group B. CONCLUSIONS: Our data show improved compliance and fewer adverse effects with alternate-day S-1 administration, which appears to be a more sustainable option for adjuvant chemotherapy for Stage II or III gastric cancer.