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The germination mutant TM-31 of Bacillus megaterium ATCC 12872, was isolated by transposon Tn917 insertional mutagenesis. Glucose, L-proline, L-leucine and KNO3 germinated TM-31 poorly. The DNA in the region of the Tn917 insertion was cloned, and its nucleotide sequence determined. One major open reading frame was present on the cloned DNA. The hydrophobic protein encoded is presumably membrane-associated. A homology search revealed that the gene encoded in the region of the Tn917 insertion is homologous to napA of Enterococcus hirae. napA codes for the NaH-antiporter. It is hypothesized that transport of cations must play an important role in spore germination in B. megaterium ATCC 12872.

The following is a personal account of how a small number of conference interpreters and others interested in the conference interpreting profession in Japan launched the Interpreting Research Assocation of Japan. The paper also reviews some of the pertinent writings, mainly in Japanese, which discuss various aspects of conference interpreting. It is hoped that this article will help shed some light on a number of issues we face in promoting interpretation research, as well as on the profession itself, in Japan.Resume: Cet article decrit de maniere personnelle le lancement de la Interpreting Research Association of Japan par un petit nombre d'interpretes de conference et par d'autres personnes interessees par la profession. Il passe egalement en revue quelques textes, essentiellement japonais, qui analysent differents aspects de l'interpretation de conference. Nous esperons que cet article apportera quelque lumiere sur les problemes auxquels nous nous heurtons dans nos tentatives de promotion de la recherche sur l'interpretation, ainsi que de la profession d'interprete de conference, au Japon.

From among 84 marine algae, the green alga, Dunaliella primolecta C-525, exhibiting the highest antibiotic activity, was selected. A crude extract from this alga strongly inhibited the growth of Staphylococcus aureus, Bacillus cereus, Bacillus subtilis and Entrobacter aerogenes. From experiments on pH, solvent, and an inhibitor of the crude extract of this alga, the antibiotic substance in the algal cells was observed as a base-stable, nonprotein material. To purify this antibiotic substance, a methanol extract of this alga was eluted successively on a DEAE-Sepharose Cl-6B, silica and Sephadex LH-20 gel column. Three peaks exhibiting antibiotic activity from the final column were observed, indicating peak No. 1 to have a molecular weight of over 1300 and peaks Nos 2 and 3 to have molecular weights of about 300–400. The main fractions X, Y and Z of the three peaks were investigated for temperature stability. The activities of fractions X and Z were stable up to 80°C, whereas the activity of fraction Y was unstable above 40°C. The above results indicate that algal cells of D. primolecta contained a number of different antibiotic substances.

Sustained production of AA and EPA, which were accumulated at high levels without a change of fatty acid composition, was demonstrated by the red alga, Porphyridium purpureum . This alga produced 5.1 mg of AA and 5.7 mg of EPA for 1.0/culture in a one-week operation on a light/dark cycle.

河川環境と細菌フローラとの関係を知るために,大阪府北部を流れる箕面川および猪名川の環境の異なる2地点について水生生物による水質の判定,および菌数計測,GC含量の測定,細菌フローラの解析を行った。その結果,汚染の進んだ桑津では生菌数,全菌数とともに全菌数に対する生菌数の割合(CFU/TDC値)が増加していた。また,汚染の度合いに関わらず河川水中ではPseudomonas菌群が優先種であり,汚染の進んだ地点ではAlcaligenes菌群が優先種であること,および清浄な地点においてはAeromonas菌群が存在しVibrio菌群は存在しないが,汚染の進んだ地点では逆になることが分かった。また,2地点から単離した各50株のGC含量の分布パターンが環境の異なる2地点では異なっていた。

A simulation test method was developed to study biodegradation of chemical compounds by monitoring the decrease in total organic carbon (TOC) of a test medium during incubation with river water microorganisms. Artificial river water containing the test chemical as the sole carbon source with 20 ppm of carbon was used as the growth medium and river water microorganisms which were trapped by membrane filtration (0.45 um) were dispersed in the medium by sonication. The test medium was incubated for 28 d with shaking in the dark. Samples were drawn periodically and biodegradation was determined by measuring the remaining TOC. Biodegradation of 13 chemicals was studied by this TOC die-away method and compared these results with that of cultivation method which was reported earlier. The change in dominating populations during incubation was found to depend significantly on the test chemical when their colony morphology was examined periodically.

The present study was made to characterize the diversity of the aniline-degrading bacteria in river water in respect to taxonomy, substrate specificity of aniline derivatives, and the inducibility and stability of the aniline-degrading activity

Samples of water from two rivers in Osaka, one of which is considered to be unpolluted and the other moderately polluted by man made chemicals were shown to have different capabilities in the biodegradation of aniline. Microorganisms in the unpolluted Mino River water were not able to biodegrade aniline, whereas those in the moderately polluted Ina River water could do so. The microflora differed significantly in the two rivers. The MPN counts of the aniline degrading microorganisms of these two river waters also indicated less chance of aniline biodegradation in the Mino River. This would suggest that aniline degradation in river waters may reveal many facts on degree of adaptation to and alteration of natural populations under pollutant stress.

Y. Gambier, D. Gile e C. Taylor; John Benjamins, Amsterdam.

A new method for transformation of Bacillus megaterium was developed by modification of Chang and Cohen's method. In our method, spore spheroplasts were used as recipient cells instead of the protoplasts of vegetative cells. Longer incubation (60 min) of spore spheroplasts and plasmid DNA before treatment with polyethylene glycol remarkably increased the efficiency of transformation. The frequency of transformation was about 10(4) per microgram of plasmid DNA. A shot-gun-type cloning of chromosome DNA of B. megaterium ATCC 12872 was available in B. megaterium ATCC 19213 strain by this transformation method.

The minimum inhibitory concentrations of ten fatty acids and their methyl esters on Staphylococcus aureus and MRSA #10 were investigated. The highest antibiotic activity was observed for γ-linolenic acid (C18:3), and when the concentration was 500 µg/ml in the medium, the MRSA cells were all killed.

A model experiment was performed to study the behavior of pesticides in foods during cooking treatments, by means of introducing a sample piece of filter paper in stead of food, and it was carried out with 14 pesticides. Most of pesticides used showed little decrease in remainings by washing with or without detergent and the difference between the washing treatments was not distinguishable. Heating in water (boiling) and in oil (frying) reduced the pesticide remainings in the sample due to decomposition and migration into the solvents. During soaking, a certain amount of pesticide was released into the solvents. Residual amounts of pesticides after heating in air (baking) varied from one to another. Behavior of pesticides during cooking treatments was found to be associated with physicochemical properties of individual pesticides, such as molecular weight, boiling point and solubility in water or oil.

Properties of alkali-soluble components from the spore coat of Bacillus megaterium were examined by physicochemical methods. They were composed of acidic polypeptides of various molecular weights with small amounts of phosphorus and sugar. They were allowed to dissociate to unit components by incubation with SDS. The major component was partially purified by gel filtration, and shown to have a mean molecular weight of about 11,000.

Hg- and Cd-spores of Bacillus megaterium QM B1551 were produced in Schaeffer's medium containing mercuric chloride and cadmium chloride respectively. Metals were added to the medium at 9 hr of incubation (Stage V) to give a final concentration of 50 μm. It was found by electron microscopic and biochemical studies that the coats of both Hg- and Cd-spores were thinner than those of control spores. Of the total Hg and Cd in the spores, 77% of the Hg and 63% of the Cd were detected in the spore coats. Hg- and Cd-spores were less resistant to heat and more sensitive to germinants than control spores. Other properties of Hg- and Cd-spores were similar to those of control spores. These results suggest that the spore coat has some relationship to the heat resistance and germinability of spores.

An automated survey system on the chemicals in environments by a gas chromatography-mass spectrometry-computer system has been constructed of Three Dimensional Mass Chromatography, Modified Probability Based Matching method, and Self Training Interpretive and Retrieval method. And it was applied to analyze environmental materials, with interesting results.

Summary: A spore coat protein of Bacillus megaterium QM B1551 was purified from an SDS-DTT extract of lysozyme-digested spores using Sephadex, DEAE-cellulose chromatography and polyacrylamide disc gel electrophoresis. The molecular weight of the protein was approximately 12000. It constituted about 13% of spore dry weight and was rich in lysine, aspartic acid, glycine and alanine residues. The sum of these four amino acids constituted 53.9% of the total amino acids. Small amounts of phosphorus and sugar were detected, but no amino sugar. The protein was synthesized in the cytosol of sporulating mother cells at t 3, and accumulated on the forespores.

The effect of salts on components of the respiratory chain of marine bacterium 1055-1 has been studied. Sodium ions activated the NADH oxidase system. This was not due to activation of NADH dehydrogenase but resulted from the activation of some other components that were membrane-bound. One of these activated components was involved in the 2-heptyl-4-hydroxyquinoline N-oxide (HOQNO)-sensitive reduction of ubiquinone. In contrast, succinate oxidase was inhibited by high concentrations of salts and it was concluded that the inhibited component was succinate dehydrogenase.

Heavy metal (Hg, Pb, Cd, Cu, Ni, Fe, Zn, and Mn) contents of the samples from 10 males were followed throughout a year. The values were characteristic of the individuals, and some correlationships between the contents of different metals were shown.

It was proved that three spore coat proteins of 48, 36, and 22 kDa (P48, P36, and P22) were the components of the outermost layer (OL) of Bacillus megaterium ATCC 12872 spore by analysis of the isolated OL. And it was indicated that these proteins were deposited not by disulfide bond, but by ionic and/or hydrophobic bonds on the spore. Among them, P36 and P22 were expected to be located on the very surface of the spore by immunological analysis. In the OL deficient mutant of B. megaterium ATCC 12872, MAE05, whose spore was lacking in these OL proteins and galactosamine-6-phosphate polymer, both P36 and P22 were present in the mother cell cytoplasm and deposited on the forespores, but they disappeared with the lysis of mother cells. An OL protein-releasing factor having proteolytic activity was detected in the culture supernatant at the late sporulating stage of both the wild-type and the mutant strains. But the factor could not act on the proteins of the mature spores and the forespores at t10 (tn indicates n hr after the end of exponential growth) of the wild-type strain. Moreover, P36 and P22 were found in the spores of a revertant of MAE05 which could form galactosamine-6-phosphate polymer, suggesting that this sugar polymer played the role in protecting the OL proteins against the protease-like substance after the deposition.

Galactosamine-6-phosphate was identified as a component of the coat of the Bacillus megaterium QM B1551 spore. It was one of the main constituents of the outermost layer of the spore coat, but it was absent from the other integuments including the cortex. These findings suggest that galactosamine-6-phosphate comprises the phosphorus-containing skeleton structure of the spore coat.

Orally administered 3 H-benzo(a)pyrene (BP) was concentrated in liver, kidney and lung of rat. Liver contained BP-diols, BP-quinones and 3-hydroxy-BP, and extremely high amounts of BP-quinones and unmetabolized BP were found in lung until after 7 hr of administration. In kidney, BP-quinones and BP-diols were major metabolites, but not BP or 3-hydroxy-BP. Low level of metabolites as conjugates with glucuronate and more polar compounds were also detected in kidney. These results suggested that an orally administered BP was metabolized to BP-diols and 3-hydroxy-BP mainly in liver, to BP-quinones mainly in lung, and to BP-diols and glucuronides or more polar compounds mainly in kidney.

We analyzed Cd contents in more than 500 samples of foods purchased in Tokyo and the Osaka-Kobe area, and estimated the general contents in foods. The highest value was obtained in seaweeds (0.36 ppm) and high contents were found in shellfish, squid, "Tsukudani" of fish and wheat germ. Other foods contained less than 0.07 ppm. The average uptake of Cd from foods was calculated to be about 40 μg/day/man, by multiplying the values estimated by averaging the general contents in similar foods by the amounts of daily uptake of the various food groups.

The initiating mechanism in the germination of Bacillus thiaminolyticus spores was studied with 14C-L-alanine. A characteristic pattern of incorporation of L-alanine into the spores was observed during the early stages of germination with two incorporation peaks, one occurred just after contact with L-alanine (first incorporation) and the other 5 min later (second incorporation). L-Glutamine, L-valine, or L-serine substituted for the incorporation of L-alanine during the first stage of germination. Although, L-alanine taken up during the first incorporation phase was extractable with trichloroacetic acid (TCA), that taken up during the second incorporation phase was not extractable. The distribution of radioactivity showed that incorporated L-alanine was located in the spore coat, mainly in the paracrystal fraction. The radioactive material which remained in the germination medium or was extractable from the spore coat fraction with TCA treatment or pronase digestion was identified as alanine. Significance of incorporation of L-alanine and its location in the spore in reference to the initiation of germination is discussed.

After intravenous injection of 3-hydroxybenzo (a) pyrene into rat, benzo (a) pyrene-diols were detected in urine, and in vitro experiment using isolated hepatocytes and sliced kidney also indicated the metabolism of 3-hydroxybenzo (a) pyrene to diol derivatives. THese results suggested the presence of unknown metabolic pathway of benzo (a) pyrene-phenols such as 3-hydroxybenzo (a) pyrene to diol derivatives both in vivo and in vitro .

SUMMARY: Cells of Bacillus subtilis NRRL B558 at early stages of sporulation contained considerable amounts of cysteic acid. The cell cysteic acid content began to increase at the onset of sporulation and was maximal when synthesis of dipicolinic acid commenced; it then decreased as sporulation advanced. Cysteic acid was synthesized from cysteine and metabolized to sulpholactate in the mother-cell cytoplasm, and the sulpholactate was then incorporated into the forespore. This metabolic change is considered to be a general phenomenon in spore-formers that contain sulpholactate in their spores.

Rain water (about 140 l), collected in suburban residential and downtown areas, was fractionated by column chromatography on Amberlite XAD-2 resin and silica gel. Trace amounts of organic compounds in rain water were identified by the use of a GC-MS-computer system combined with three-dimensional mass chromatography. Phthalate esters, polycyclic aromatic hydrocarbons, higher fatty acids and their esters, and other aromatic derivatives were identified with high reliability.

Exposure to cadmium induced the nematode Caenorhabditis elegans to synthesize metallothioneinlike proteins, which were isolated and characterized. The supernatant obtained after disruption of cadmium-exposed nematodes was applied to a Sephadex G-75 column, and cadmium was detected in a high-molecular-weight fraction and a lowmolecular-weight fraction, of which the latter corresponded in elution volume to the well-characterized metallothionein. From this low molecular weight cadmium-rich fraction, two major cadmium-binding proteins were separated by DEAE Sephadex A-25 ion exchange chromatography. These two isoproteins showed similar ultraviolet absorption spectra to mammalian metallothioneins, indicating that they can be categorized metallothioneins. The slower-eluted isoform, MT-II, was as homogeneous on polyacrylamide disc gel electrophoresis while the faster-eluted isoform, MT-I, contained some impurity. The amino acid composition of the purified MT-II was determined. The content of cysteic residues was 22.3 % and the next most abundant residues were aspartate, alanine, glutamate and lysine. The content of acidic amino acids was high and that of aromatic amino acids was very small. The properties were similar to those of metallothioneins found in lower phyla of the animal kingdom rather than in mammalian species.

Forespores of Bacillus subtilis 60015 were isolated from sporulating cells at t5 and were incubated with an amino acid mixture containing [14C]phenylalanine. Three species of 14Clabelled cytoplasmic proteins synthesized in the forespores were purified to homogeneity by gel filtration in the presence of detergents and by ion-exchange chromatography. The molecular weights of the purified proteins, called A, B and C, were estimated by SDS-PAGE as 24200, 1 1 500 and 12 700, respectively. Protein B contained remarkably high amounts of tyrosine (2343%) and alanine (22.8%), and proteins A and C contained relatively high amounts of glutamate + glutamine, glycine, and alanine (10.4 to 13.3%). The synthesis of these proteins can provide markers for the control events in the forespore compartment during sporulation.

An immunochemical staining technique for the spore coat proteins of Bacillus megaterium ATCC 12872 was developed using colloidal gold as a second antibody. For reducing the non-specific immunogold binding and increasing the specific binding, the affinity-purified IgG was used as a first antibody. In sporulating cells at t10, gold particles were found not only in the spore coat but also in the mother cell cytoplasm, suggesting that some coat proteins were synthesized in the cytoplasm. Use of the specific affinity-purified antibody to 48K-protein demonstrated that this protein was one of the components of the outer coat.

When the spores of Bacillus megaterium QM B1551 (ATCC 12872) were incubated with 5 mM CdCl2 at 30 C, they underwent the early germination events, such as loss of heat resistance and release of calcium dipicolinate, in the same way as when they were germinated by glucose + KNO3. However, germination by CdCl2 caused no increase in the reducing groups in the cortex and no excretion of glucosamine-containing materials due to the hydrolysis of the cortex peptidoglycan. Addition of CdCl2 at any time during germination by glucose + KNO3 inhibited the release of glucosamine-containing materials from the spores, whereas removal of cadmium from the CdCl2-germinated spores by treatment with cysteine restored the hydrolysis of peptidoglycan. These results suggested that CdCl2 caused the early events of spore germination but prevented the spores from undergoing the events following germination by inhibiting the enzymatic lysis of the cortex peptidoglycan. The conclusion from the study is that cortex degradation is not always required for the initiation of germination.

The structure of the skeleton of spores of Bacillus megaterium was examined after ashing in a plasma asher and the elemental composition of the ashed whole spores was determined with an analytical electron microscope. All spores were ashed in situ although they shrank by about 15%. Even P and S, in addition to metals, were recovered well from ashed samples. Ash was rich in the core and the coat, and poor in the cortex. Ca, P, S, and Mg were detected in the core and coat of the spore of B. megaterium QM B1551. Ca in the core was markedly decreased by germination or autoclaving. In the spore of B. megaterium ATCC 19213, almost all of the ash was detected in the core and its elemental composition was similar to that of the core of the strain QM B1551 spore. These results suggest strongly that the core is the site of Ca associated with dipicolinic acid.

A three dimensional mass chromatography (TDMC), which is the extended method of mass chromatography, has been developed for the analysis of environmental chemicals. A considerable performance was suggested by the combination of TDMC to gas chromatography-mass spectrometry-computer system.

Biodegradability of organic compounds in river and sea water was tested according to the simple and rapid method, the cultivation method, which was reported in our previous report. Out of 170 chemicals tested, 103 compounds were judged to be hard-, 43 to be moderate-, and 24 to be easy-of-degradability. In comparison with the MITI method, our method was applicable to wider range of compounds with less expensive equipments and gave more strict marks in 3 d, indicating that our method is a useful screening test method for biodegradability of chemicals in environmental water.

A retrieval system of mass spectra based upon the Probability Based Matching method was studied in order to apply it to survey the chemicals in environments by a gas chromatography-mass spectrometry-computer system. As retrieval indices, peak missing probability and relative confidence value were proposed.

Bacillus megaterium芽胞の発芽にたいするHgCl2の作用について検討した。発芽は1mMのHgCl2の存在下で完全に阻害された。この時芽胞内へのHgの取込みがみられた。HgCl2処理芽胞を肉エキスブイヨン中で保温した場合,発芽の回復がみとめられた。この時,発芽の進行にともなつて,芽胞からのHgの流出量の増加が認められた。未処理芽胞を各種薬剤で処理した時,芽胞からのタンパク質の流出とHgCl2処理芽胞を同様の薬剤で処理した時のHgの流出量とに関する実験から,urea処理時に流出するタンパク質はHg結合タンパク質を含む可能性のあることが推察された。またHgCl2処理芽胞は未処理芽胞と比較して発芽剤にたいする特異性が異なることを認めた。