[Woman physicians in child neurology--their opinions and dreams (discussion)].
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Child↗
Biomedical subjects
A Ishizaki
Publications and source records attributed to A Ishizaki.
Identification and production of a bacteriocin from Enterococcus mundtii QU 2 isolated from soybean.
AIMS: Identification of the bacteriocin produced by Enterococcus mundtii QU 2 newly isolated from soybean and fermentative production of the bacteriocin. METHODS AND RESULTS: The bacteriocin produced by Ent. mundtii QU 2 inhibited the growth of various indicator strains, including Enterococcus, Lactobacillus, Leuconostoc, Pediococcus and Listeria. The bacteriocin activity was stable at wide pH range and against heat treatment, but completely abolished by proteolytic enzymes. The bacteriocin was purified from the culture supernatant by the three-step chromatographic procedure. Mass spectrometry, amino acid sequencing and DNA sequencing revealed that the bacteriocin was similar to class IIa bacteriocins produced by other Ent. mundtii strains. The bacteriocin production decreased in the absence of glucose, nitrogen sources, or Tween 80 in MRS medium. Additionally, it was strongly suppressed by addition of Ca(2+) (CaCO(3) or CaCl(2)). In pH-controlled fermentations, the highest bacteriocin production was achieved at pH 6.0, whereas the highest cell growth was obtained at pH 7.0. CONCLUSIONS: Ent. mundtii QU 2 produced a class IIa bacteriocin. Some growth factors (e.g. Ca(2+) and pH) influenced the bacteriocin production. SIGNIFICANCE AND IMPACT OF THE STUDY: A new soybean isolate, Ent. mundtii QU 2 was found to be a class IIa bacteriocin producer. Factors influencing the bacteriocin production described herein are valuable for applications of the bacteriocins from Ent. mundtii strains.
Amino Acid Sequence↗
Two different pathways for D-xylose metabolism and the effect of xylose concentration on the yield coefficient of L-lactate in mixed-acid fermentation by the lactic acid bacterium Lactococcus lactis IO-1.
In lactic acid bacteria, pentoses are metabolized via the phosphoketolase pathway, which catalyzes the cleavage of D-xylulose-5-phosphate to equimolar amounts of glyceraldehyde 3-phosphate and acetylphosphate. Hence the yield coefficient of lactate from pentose does not exceed 1.0 mol/mol, while that of Lactococcus lactis IO-1(JCM7638) at high D-xylose concentrations often exceeds the theoretical value. This suggests that, in addition to the phosphoketolase pathway, L. lactisIO-1 may possess another metabolic pathway that produces only lactic acid from xylose. In the present study, the metabolism of xylose in L. lactisIO-1 was deduced from the product formation and enzyme activities of L. lactisIO-1 in batch culture and continuous culture. During cultivation with xylose concentrations above ca. 50 g/l, the yield coefficient of L-lactate exceeded 1.0 mol/mol while those of acetate, formate and ethanol were very low. At xylose concentrations less than 5 g/l, acetate, formate and ethanol were produced with yield coefficients of about 1.0 mol/mol, while L-lactate was scarcely produced. In cells grown at high xylose concentrations, a marked decrease in the specific activities of phosphoketolase and pyruvate formate lyase (PFL), and an increase in those of transketolase and transaldolase were observed. These results indicate that in L. lactisIO-1 xylose may be catabolized by two different pathways, the phosphoketolase pathway yielding acetate, formate and ethanol, and the pentose phosphate (PP)/glycolytic pathway which converts xylose to L-lactate only. Furthermore, it was deduced that the change in the xylose concentration in the culture medium shifts xylulose 5-phosphate metabolism between the phosphoketolase pathway and the PP/glycolytic pathway in L. lactisIO-1, and pyruvate metabolism between cleavage to acetyl-CoA and formic acid by PFL and the reduction to L-lactate by lactate dehydrogenase.
Enzymes↗
Biochemical and genetic evidence for production of enterocins A and B by Enterococcus faecium WHE 81.
Enterococcus faecium WHE 81, isolated from cheese, has been reported to produce a bacteriocin called "enterocin 81" [J. Appl. Microbiol. 85 (1998) 521.]. Purification of "enterocin 81" was carried out using ammonium sulfate precipitation, desalting on ODP-90 reverse-phase column, and purification through SP Sepharose HP cation exchange and C2/C18 reverse-phase chromatographies. The antimicrobial was eluted from the C2/C18 column as four individually active fractions, designated A81, B81, C81 and D81. The purification procedure used proved particularly efficient for the bacteriocin in fraction D81, with a yield of 46%, while only 3.8% the bacteriocin in fraction B81 could be collected. MALDI-TOF mass spectrometry of the bacteriocins in fractions B81 and D81 showed respective masses of 4,833.0 and 5,462.2 Da. Amino acid sequencing of the two peptides revealed two class-II bacteriocins whose sequences were similar to those of enterocin A and enterocin B, respectively. Using proper primers, chromosomal fragments of 212 and 216 bp enclosing bacteriocin structural genes were PCR-amplified. Cloning of the amplicons and their sequencing revealed two genes with sequences identical to the structural genes of enterocins A and B, respectively. It was therefore clearly established that E. faecium WHE 81 produces bacteriocins respectively identical to enterocins A and B. Our results, combined with data from previous reports, suggest that the two bacteriocins may be widespread among enterococcal strains and may play an important role in controlling the growth of pathogens and other undesirable bacteria in certain fermented food products.
Amino Acid Sequence↗
The effect of osmotic stress on the production of nukacin ISK-1 from Staphylococcus warneri ISK-1.
The effects of several additives on the production of a lantibiotic, nukacin ISK-1, from Staphylococcus warneri ISK-1, in batch fermentation were studied. NaCl, KCl and sorbitol stimulated nukacin ISK-1 production. The addition of 1.4 M NaCl increased nukacin ISK-1 activity 1.5-fold over the control, while cell growth and glucose consumption were inhibited. Nukacin ISK-1 production increased with increasing osmolarity of the medium up to about 3 osmol/kg; however, further increases in osmolarity diminished productivity, irrespective of the kind of additive. Northern blot analysis showed that transcription of the nukacin ISK-1 structural gene (nukA) was activated in the presence of 1.4 M NaC1. These data indicate that the stimulation effect was due to osmotic stress, which acted, at least in part, at the transcriptional level on the nukA gene.
Bacterial Proteins↗
Microbial production of poly-D-3-hydroxybutyrate from CO2.
This short review covers the biotechnological aspects of the production of poly-D-3-hydroxybutyric acid, P(3HB), from H2, O2 and CO2 by autotrophic culture of the hydrogen-oxidizing bacterium, Ralstonia eutropha. Considering the efficiency of utilization of a gas mixture as substrate, a practical fermentation process using R. eutropha for the mass production of P(3HB) from CO2 should be designed on the basis of a recycled-gas, closed-circuit culture system. Also, maintaining the O2 concentration in a gas phase lower than 6.9% (v/v) is essential to prevent the gas mixture from exploding. Our study, using an explosion-proof fermentation bench plant and a two-stage culture system with a newly designed air-lift fermenter, demonstrated that very high P(3HB) yield and productivity could be obtained while the O2 concentration was maintained below 6.9%. However, a study on the continuous production of P(3HB) from CO2 by chemostat culture of R. eutropha revealed that the productivity and content of P(3HB) in the cells was considerably lower than by fed-batch culture. It is deduced that the use of the hydrogen-oxidizing bacterium, Alcaligenes latus, which accumulates P(3HB) even in the exponential growth phase, will be useful for the effective production of P(3HB) from CO2.
Bioreactors↗
Development of a novel method for feeding a mixture of L-lactic acid and acetic acid in fed-batch culture of Ralstonia eutropha for poly-D-3-hydroxybutyrate production.
Fermentative production of poly-D-3-hydroxybutyrate [P(3HB)] from a mixture of L-lactic acid and acetic acid by Ralstonia eutropha was investigated. For fed-batch culture with cell density, it is necessary to control the concentration of these organic acids in the culture medium below the inhibitory level for cell growth. Therefore, a novel feeding method, termed the computer-controlled pH-stat substrate feeding method, was developed using the rate of increase of the pH (pH-increasing rate) of the culture medium as an indicator for feed control. The pH-increasing rate, which was calculated every minute by a pH meter-linked computer, represented secondary information regarding substrate consumption by cells. When the pH-increasing rate decreased to 5% of the maximum increasing rate, acidic substrate solution was fed into the fermentor until the pH was reduced to 7.00. Using this feeding strategy, the cell concentration and PHA content obtained in 42 h were 75.0 g/l and 73.1% (w/w), respectively, resulting in a high P(3HB) productivity of 1.30 g/l.h.
Journal Article↗
[Methylphenidate therapy in 141 patients with hyperkinetic disorder or with pervasive developmental disorder and hyperkinesia].
The efficacy and adverse reactions produced by methylphenidate (MPD) therapy were evaluated in 141 patients with hyperkinetic disorder or pervasive developmental disorder (PDD) with hyperkinesia. Ninety-nine patients were followed for 1 to 5 years to determine if the treatment could be continued and if the patients' adaptation to their environment improved. The results showed that the MPD therapy was effective in 93% of patients whose IQ was > 80 and in 70% whose IQ was < or = 80. The efficacy was not significantly different between patients with PDD and those without PDD. Of the patients in whom the MPD therapy was effective, the majority received a MPD dosage of 0.3 mg/kg once every morning. Adverse reactions, such as excitability, nausea or anorexia, and insomnia were reported in 23% of the patients. Although this figure was not negligible, no serious events occurred. Seizure induction was suspected in 2 patients. Many of the patients (53/83) in whom the MPD treatment was effective continued to receive the treatment throughout the follow-up period. By the time that the conditions were alleviated to the extent that the treatment could be stopped, the patients had become well adapted to their environment. However, in many other cases, adaptation was unsatisfactory. In these cases, psycosocial interventions were necessary, even if the MPD therapy was effective.
Adolescent↗
Molecular cloning of two (R)-specific enoyl-CoA hydratase genes from Pseudomonas aeruginosa and their use for polyhydroxyalkanoate synthesis.
Two Pseudomonas aeruginosa genes, termed phaJ1(Pa) and phaJ2(Pa), homologous to the Aeromonas caviae (R)-specific enoyl-CoA hydratase gene (phaJ(Ac)) were cloned using a PCR technique to investigate the monomer-supplying ability for polyhydroxyalkanoate (PHA) synthesis from beta-oxidation cycle. Two expression plasmids for phaJ1(Pa) and phaJ2(Pa) were constructed and introduced into Escherichia coli DH5alpha strain. The recombinants harboring phaJ1(Pa) or phaJ2(Pa) showed high (R)-specific enoyl-CoA hydratase activity with different substrate specificities, that is, specific for short chain-length enoyl-CoA or medium chain-length enoyl-CoA, respectively. In addition, co-expression of these two hydratase genes with PHA synthase gene in E. coli LS5218 resulted in the accumulation of PHA up to 14-29 wt% of cell dry weight from dodecanoate as a sole carbon source. It has been suggested that phaJ1(Pa) and phaJ2(Pa) products have the monomer-supplying ability for PHA synthesis from beta-oxidation cycle.
Acyltransferases↗
Disposition of bupivacaine and its metabolites in the maternal, placental, and fetal compartments in rats.
BACKGROUND: This study was designed to determine the disposition of bupivacaine and its metabolites in the maternal, placental, and fetal compartments, using multiple sampling time points in chronically prepared awake pregnant rats. METHODS: All animals received an intravenous infusion of bupivacaine at a rate of 0.33 mg. kg-1. min-1 over a period of 15 min. The fetuses were delivered either at the end of infusion or at 2 or 4 h after dosing. Maternal and fetal blood and tissue samples were obtained for the assays of bupivacaine and its metabolites using capillary gas chromatography-mass spectrometry. RESULTS: The elimination half-life of bupivacaine was 37.7 min. The major metabolite was 3'-hydroxybupivacaine. Bupivacaine and 3'-hydroxybupivacaine were present in all samples at the end of administration. The fetal to maternal concentration ratio of bupivacaine in plasma was 0.29, and in the placenta was 0.63. The amnion contained the highest bupivacaine concentration: threefold higher in the maternal and 11-fold higher than in the fetal plasma. At 4 h after dosing, bupivacaine was no longer detectable in any maternal and fetal samples, whereas 3'-hydroxybupivacaine was still present in all tissues except the fetal plasma and heart. CONCLUSIONS: These data indicate that a considerable amount of bupivacaine is taken up by both sides of the placenta, as well as the amnion and myometrium. 3'-Hydroxybupivacaine was present in all tissues except the fetal plasma and heart samples, even after the parent compound became no longer detectable. Whether this slow elimination of 3'-hydroxybupivacaine causes any adverse effects on the fetus-newborn needs to be explored.
Amnion↗
Class IIa bacteriocins: biosynthesis, structure and activity.
In the last decade, a variety of ribosomally synthesized antimicrobial peptides or bacteriocins produced by lactic acid bacteria have been identified and characterized. As a result of these studies, insight has been gained into fundamental aspects of biology and biochemistry such as producer self protection, membrane-protein interactions, and protein modification and secretion. Moreover, it has become evident that these peptides may be developed into useful antimicrobial additives. Class IIa bacteriocins can be considered as the major subgroup of bacteriocins from lactic acid bacteria, not only because of their large number, but also because of their activities and potential applications. They have first attracted particular attention as listericidal compounds and are now believed to be the next in line if more bacteriocins are to be approved in the future. The present review attempts to provide an insight into general knowledge available for class IIa bacteriocins and discusses common features and recent findings concerning these substances.
Amino Acid Sequence↗
Purification and identification of a growth-stimulating peptide for Bifidobacterium bifidum from natural rubber serum powder.
Natural rubber serum powder, which is a by-product obtained in the production of latex rubber, has a strong growth-stimulating activity for Bifidobacterium bifidum JCM 1254. The retained fraction obtained by ultrafiltration (molecular weight cutoff 1000) showed a growth-stimulating activity in a dose-dependent manner on B12 assay medium with ammonium sulfate. One of the growth stimulators was purified from the retained fraction by acetone precipitation, solid-phase extraction with a hydrophobic pretreatment column, and multistage reversed-phase HPLC. An increase of 53-fold in the specific activity, and a recovery of 1.3% were obtained. The amino acid composition and N-terminal sequence analysis of this growth stimulator provided the structure of Ala-Thr-Pro-Glu-Lys-Glu-Glu-Pro-Thr-Ala. The molecular mass was 1075 by MALDI-TOF MS analysis. These results showed that this growth stimulator was a decapeptide with the sequence shown above. This is the first report that clarified the structure of an active peptide for the growth of Bifidobacterium.
Amino Acid Sequence↗
A novel lantibiotic, nukacin ISK-1, of Staphylococcus warneri ISK-1: cloning of the structural gene and identification of the structure.
Staphylococcus warneri ISK-1, which we had previously reported as Pediococcus sp. ISK-1, produces a novel bacteriocin, nukacin ISK-1. Edman degradation of the chemically reduced nukacin ISK-1 produced a sequence of 27 amino acids, 7 of which were unidentified. Using single-specific-primer-PCR product as a probe, a 3.6-kb HindIII fragment containing the nukacin ISK-1 structural gene (nukA) was cloned and sequenced. The deduced amino acid sequence of nukacin ISK-1 had 57 amino acids, including a 30-amino acid leader region. The propeptide sequence showed significant similarity to those of lacticin-481 type lantibiotics. In the region upstream of nukA, a part of a long open reading frame (ORF), designated as nukM, encoding a putative modification enzyme was oriented in the opposite direction. In the region downstream of nukA, ORF1 was found in which the sequence of the putative translational product was similar to various response regulatory proteins.
Amino Acid Sequence↗
Cloning of a full-length cDNA encoding ent-kaurene synthase from Gibberella fujikuroi: functional analysis of a bifunctional diterpene cyclase.
We report here the nucleotide sequence of a full-length cDNA encoding ent-kaurene synthase that was isolated by a reverse-transcription polymerase chain reaction from Gibberella fujikuroi (Gcps/ks). This cDNA encodes 952 amino acid residues with a relative molecular mass of 107 kDa. The sequence similarity between Gcps/ks and ent-kaurene synthase of the gibberellin A1-producing fungus, Phaeosphaeria sp. L487, is very high, suggesting that Gcps/ks is also a bifunctional diterpene cyclase. Its recombinant protein expressed in Escherichia coli converted geranylgeranyl diphosphate to copalyl diphosphate and ent-kaurene.
Alkyl and Aryl Transferases↗
Control of acetic acid concentration by pH-stat continuous substrate feeding in heterotrophic culture phase of two-stage cultivation of Alcaligenes eutrophus for production of P(3HB) from CO2, H2, and O2 under non-explosive conditions.
A-two stage culture method of hydrogen-oxidizing bacterium, Alcaligenes eutrophus, is used to produce poly-D-3-hydroxybutyrate, P(3HB) from CO2, O2, and H2 without using a very high oxygen transfer rate while maintaining the O2 concentration in gas phase below 6.9 (v/v)% to prevent detonation of the gas mixture. The two-stage method consists of a heterotrophic culture using fructose as carbon source for exponential cell growth and an autotrophic culture for P(3HB) accumulation. We investigated the use of acetic acid as a cheaper carbon source than fructose for the heterotrophic culture in the two-stage method. However, the acetate concentration in the culture system must be maintained at 1.0 g. dm-3 since its inhibitory effect on the cell growth is very strong. Then, high cell density cultivation of A. eutrophus was investigated by pH-stat continuous feeding of acetic acid to control acetate concentration. As a result, acetate concentration was automatically maintained around 1.0 g. dm-3 by using a feed with a composition in CH3COOH/CH3COONH4/KH2PO4 molar ratio of 5:1:0.084. Cell concentration increased to 48.6 g. dm-3 after 21 h of cultivation. The cell mass grown in the fed-batch culture on acetic acid was useful for P(3HB) production from CO2 in the subsequent autotrophic culture stage. Copyright 1999 John Wiley & Sons, Inc.
Journal Article↗
Mucin-producing tumors of the pancreas: clinicopathological features, surgical treatment, and outcome.
Mucin-producing tumors (MPTs) of the pancreas are increasingly being recognized. To evaluate the appropriate surgical treatment and predict the prognosis of MPTs, we performed a retrospective clinicopathological study in 51 patients, 27 with benign tumors and 24 with borderline/malignant tumors. Three of the malignant tumors showed stromal invasion and lymph node metastasis on histological examination. Of the 24 patients with borderline/malignant tumors, 2 died of MPTs and 4 died of other diseases. At the last follow-up, 35 patients were alive and well. The 5-year postoperative survival rate was 90% for patients with benign tumors, and 78% of these with borderline/malignant tumors. Five of the patients with borderline/malignant tumors had multicentric tumors. Three of these patients underwent resection of the rest of the pancreas, 5, 6, and 8 years, respectively, after the first operation. Extended radical resection is required for malignant MPT with invasion of the pancreatic stroma. We prefer to perform pancreatogastrostomy or Imanaga's procedure to allow examination of the body and tail of the pancreas by endoscopic retrograde pancreatography after resection of the pancreatic head. Careful follow-up for a long period may be the most prudent approach for detecting multiple MPTs in the residual pancreas after surgical treatment.
Adenocarcinoma, Mucinous↗
Papillary adenoma of the distal common bile duct.
A 73-year-old man with a papillary adenoma located in the distal common bile duct is reported. He underwent pylorus-preserving pancreatoduodenectomy. The lesion in the common bile duct featured papillary proliferation of the epithelium and fibrous elements with diffuse infiltration by inflammatory cells. Positive staining for MIB-1 (Ki-67) and p53 was identified in the nuclei of the proliferative epithelium. These findings suggested the malignant potential of this lesion. Further progress in imaging diagnostic techniques should increase the frequency with which such lesions are discovered. Even now, if mural irregularities and defects are found in the extrahepatic biliary system, especially the distal common bile duct, the possibility of such borderline biliary adenoma should be taken into consideration when making a diagnosis.
Adenoma, Bile Duct↗
Extractive acetone-butanol-ethanol fermentation using methylated crude palm oil as extractant in batch culture of Clostridium saccharoperbutylacetonicum N1-4 (ATCC 13564).
The possibility of employing methylated crude palm oil (CPOE) as an extraction solvent to reduce end-product inhibition and to enhance solvent productivity in acetone-butanol-ethanol (ABE) fermentation was evaluated using oleyl alcohol as the standard butanol extractant. Fermentation was carried out at an initial glucose concentration of 90 g/l. CPOE did not inhibit the growth of the fermentative organism. Without solvent extraction, butanol production ceased after 30 h at a concentration of 15.4 g/l limiting cell growth to 3.98 g/l and glucose consumption to 62%. Applying CPOE as the extraction solvent, about 47% of the total butanol produced was extracted, glucose consumption was increased to 83% and relatively high glucose consumption rates and solvent productivities were obtained. Butanol production increased to 20.9 g/l; total ABE solvents and yield also increased from 21.2 g/l and 38% (in conventional fermentation) to 29.8 g/l and 40.4%, respectively.
Journal Article↗