bluuebunny/crossref_metadata_2025 huggingface dataset:
Biochemical Properties of a Penicillinase from Escherichia coli Carrying Rms 298
Authors: Sawada Yosuke , Tai Masaru , Mitsuhashi Susumu
Published: November 2013

We obtained two R plasmids, i.e., Rms195 and Rms298, from a clinical isolate, E. coli GN5503. Penicillin β‐lactamase (PCase) was extracted from ML1410 Rms195+ and Rms298+, and was purified by chromatography. Rms195 PCase was identical to the type I PCase mediated by R‐TEM, RI and Rms212. The isoelectric point of Rms298 PCase was 5.9 and its molecular weight was 21,000 ± 1,000. The substrate profile and physicochemical properties indicate that Rms298 PCase belongs to the type IV PCase mediated by Rms139 isolated from Pseudomonas aeruginosa.
Experimental Pneumonitis: Changes in the Phospholipid Metabolism of the Lung of Guinea Pigs and Rats
Authors: Ito Masami , Takeuchi Nozomu , Masuno Tomiya , Kikui Masanori , Yamamura Yuichi
Published: November 2013

Changes of the metabolism of lung phospholipids and alveolar wash lipids were studied in guinea pigs and rats with experimental pneumonitis produced by the injection of complete Freund's adjuvant. The amount of total lipid obtained by lung lavage decreased significantly in contrast to a marked increase in the content of total protein in treated animals. Among the lipids in the pulmonary washings, only phospholipids were strikingly reduced, and free fatty acids were increased somewhat. However, the composition of phospholipids was not affected by the treatment. The in vitro incorporation of 1‐14C‐acetate into the total lipid and phospholipids in the lung with pneumonitis was slightly lower compared to controls, but the difference was not significant. The in vitro 1‐14C‐palmitic acid incorporation into phospholipids of the lesioned lung, was also similar to that in controls. Thus, phospholipid biosynthesis in the lung was not affected by pneumonitis. On the other hand, the in vitro activity of total phospholipase in rat lung with experimental pneumonitis was enhanced significantly. These results suggest that phospholipase activity is increased in the diseased lung and this may participate in the process of the inflammatory reaction. The possible role of activated macrophages in the inflammatory response of the lung is discussed.
The Host‐Dependent Restriction of Growth of an RNA Coliphage FI
Authors: Hirashima Akikazu , Furuse Kohsuke , Watanabe Itaru
Published: November 2013

Phage FIC is a spontaneous host‐dependent mutant of phage FI which is classified into the fourth group of RNA Escherichia coli phages (RNA coliphages). The mutant phage (FIC) grows normally in E. coli strain Q13 (permissive host), but poorly in strain A/λ (non‐permissive host) (9). Attempts to elucidate the regulatory mechanism of growth of the mutant phage in the non‐permissive host revealed the following: (a) growth of the mutant phage was specifically restricted in E. coli strains that have certain suppressor genes for amber mutation; (b) the mutant phage RNA (FIC‐RNA) could not produce progeny in the spheroplasts of the non‐permissive host; (c) adsorption of the mutant phage to, and penetration of the mutant phage RNA into, the non‐permissive host were normal; and (d) biosynthesis of the phage‐specific late protein and RNA did not occur in the non‐permissive host. Based on these results we conclude that phage FIC is a spontaneous azure‐type mutant of the fourth group of RNA coliphage FI.
A Simple and Efficient Microassay Method for Titration of Interferon
Authors: Ishitsuka Hideo , Nomura Yasuko , Takano Kouichi
Published: November 2013

A simple and efficient microassay method for the titration of interferon was developed by the use of microtest plates for handling a large number of samples. L929 cells pretreated with interferon were infected with vesicular stomatitits virus (VSV) and cultured in the presence of 3H‐uridine. The activity was expressed by the reduction of extracellular radioactive RNA released after destruction of the infected cells, which was measured in terms of the radioactivity incorporated into cold TCA‐insoluble materials in the culture fluid. The interferon titer determined by this method was in the same order as that by the plaque reduction method. The activity by this method was parallel to, but lower than that expressed by the yield reduction of infectious viruses. This method requires only 0.025 ml of each test sample with higher than 1 NIH ref. unit/ml to detect its interferon activity and takes 2 to 3 days for assaying hundreds of samples.
Effect of Capsular Polysaccharide of Klebsiella pneumoniae on the Differentiation and Functional Capacity of Macrophages Cultured In Vitro
Authors: Yokochi Takashi , Nakashima Izumi , Kato Nobuo
Published: November 2013

The effect of the capsular polysaccharide of Klebsiella pneumoniae (CPS‐K) on the differentiation and functional capacity of macrophages cultured in vitro from various lymphoid tissues was investigated. In cultures of peritoneal cells, the number of macrophages did not change throughout incubation periods of from 1 hr to 3 days, and the addition of CPS‐K had no affect. It appears therefore that CPS‐K does not exhibit cytotoxic effects on macrophages. In cultures of spleen cells, only a small number of macrophages appeared within 1 hr, but the number of macrophages increased during further incubation. The addition of CPS‐K to cultures of spleen cells at the start of incubation suppressed markedly the increase in the numbers of macrophage. This finding indicates that CPS‐K blocks the process of the generation of macrophages, probably from their precursor cells in cultures of spleen cells. Only a small number of macrophages appeared in cultures of thymocytes or lymph node cells either with or without CPS‐K. The phagocytic capacity of either peritoneal macrophages or macrophages generated in cultures of spleen cells was activated during incubation in vitro. Macrophages cultured in the presence of CPS‐K for 24 hr or longer appeared to have an enhanced phagocytic activity, although the enhancement of their phagocytic activity by the addition of CPS‐K was less marked in cultures of spleen cells than in those of peritoneal macrophages. Morphologically, macrophages in both cultures of peritoneal cells and spleen cells incubated in the presence of CPS‐K for 4 days possessed much longer cytoplasmic processes than those incubated in the absence of CPS‐K. From the present study, it appears that CPS‐K exhibits dual effects on macrophage precursor cells and macrophages, a blocking effect on the differentiation from the former to the latter and an enhancing effect on the functional capacity of the latter.
Failure of the Reaction of β2‐Microglobulin with Staphylococcal Protein A
Authors: Ochi Yukio , Hachiya Takashi , Kajita Yoshihiro
Published: November 2013

Because β2‐microglobulin is structurally similar to IgG, the reaction of β2‐microglobulin with Staphylococcal Protein A, which is known to react with the Fc region of IgG, was examined. 125I‐β2‐microglobulin did not bind to Protein A. This may due to the difference in the amino acid sequence between β2‐microglobulin and the Fc region of IgG.
An Endotoxin Induced Serum Factor That Causes Enhancement of Antibody Response to Heterologous Erythrocytes
Authors: Tanabe Masao J. , Tsurumi Yoshiko , Nakano Masayasu
Published: November 2013

The sera obtained from blood of the mice, which had been intravenously injected with LPS several hours in advance, contained some active substance capable of enhancing anti‐sheep red blood cell (SRBC) antibody responses in mice. Activity of the sera was still retained after passage through a rabbit anti‐LPS antibody‐coated Sepharose 4B column, but greatly reduced by passage through a rabbit anti‐mouse thymocyte antibody‐coated Sepharose 4B column. The active substance in the sera was eluted through a Sephadex G‐200 column at the same position as the serum albumin.

The addition of this substance to B cell rich spleen cell cultures in vitro in the presence of SRBC generated tremendous numbers of antibody forming cells 4 days after the incubation, suggesting that this substance was able to take over the helper function of T cells in thymus dependent antibody responses. However, this substance was not capable of stimulating 3H‐thymidine‐uptake into cultured spleen cells. The possible role of this substance in the adjuvant effect of LPS is discussed.
Studies on the Toxin of Aspergillus fumigatus
Authors: Yokota Katsushi , Shimada Hideaki , Kamaguchi Arihide , Sakaguchi Osamu
Published: November 2013

A hemolytic toxin has been obtained from mycelia and culture filtrates of Aspergillus fumigatus by the procedures that included precipitation with ammonium sulfate, chromatography of DEAE‐Sephadex, affinity chromatography on Concanavalin A‐Sepharose and gel filtration on Sephadex G‐50, G‐100 and G‐150. The purified homolytic toxin was homogeneous on immunological and disk electrophoretic analysis, and the toxin from culture filtrates was identical with that from mycelia by the immunodiffusion technique. The hemolytic toxin was obtained for the first time from fungi and designated as Asp‐hemolysin. The molecular weight of Asp‐hemolysin was estimated to be approximately 30,000 by the gel‐filtration technique and its isoelectric point was found to be around pH 4.0. This Asp‐hemolysin contained large amounts of protein and very small amounts of carbohydrate. The UV absorption spectrum of Asp‐hemolysin showed a maximum absorption at 280 nm and minimum absorption at 251 nm. The extinction coefficient at 280 nm, E 1%1cm, was 12.4 and the ratio of absorbance at 280 nm to that at 260 nm was 2.3. The optimum pH for the hemolytic activity of the toxin toward chicken erythrocytes was 5.0 at room temperature and it was active in the pH range of 3.5 to 10.5. The optimum temperature was 21 C and about 50% of the activity was lost by incubation at 50 C for 5 min or 45 C for 23 min. The hemolytic activity was remarkably inhibited by Hg2+, Cu2+, Fe2+, Ag1+, iodine and p‐CMB, but enhanced slightly by Zn2+ and Co2+.
Common Antigen between Coxsackievirus A 16 and Enterovirus 71
Authors: Hagiwara Akio , Tagaya Isamu , Yoneyama Tetsuo
Published: November 2013

Cross immunofluorescence revealed that coxsackievirus A 16 (CA 16) shared a common antigen with enterovirus 71 (E 71). The cross reactivity of these two serotypes was also examined by complement fixation test with purified virus preparations fractionated by sucrose density gradient centrifugation and two peaks of antigenicity were detected, one being type‐specific and the other cross‐reacting. The common antigen was heat‐stable and attributable to empty capsids. Immunodiffusion also revealed the common antigen. Infants without antibody to E 71 developed complement fixing and precipitin antibody to E 71 after recovery from hand, foot and mouth disease caused by CA 16.
Thymidine Metabolism in Cells Treated with DNA‐Suppressing Factor (DSF)
Authors: Fujii Nobuhiro , Minagawa Tomonori , Kato Tsuneo , Iida Hiroo
Published: November 2013

Inhibition of thymidine incorporation into DNA in cells treated with DNA‐suppressing factor (DSF) has been studied. After 16 hr treatment with DSF, transport of labeled thymidine across the cell membrane was not inhibited, since equilibrium of labeled thymidine with the acid‐soluble pool occurred at the same rate and the radioactivity was at the same level as in untreated cells. The values of Vmax and Km in the kinetics of transport of exogenous thymidine were not changed by DSF. Phosphorylation of labeled thymidine to deoxythymidine triphosphate (dTTP) was not inhibited by DSF. After a chase of labeled thymidine, radioactivity of the acid‐soluble fraction in DSF‐treated cells decreased more rapidly but that of the acid‐insoluble fraction remained at a lower level than in untreated cells. It was assumed that DSF might block the entry of dTTP into DNA.
R Plasmids among Gram‐Negative Bacteria with Multiple Drug Resistance Isolated in a General Hospital
Authors: Suzuki Satoshi , Miyoshi Yasushi , Nakaya Rintaro
Published: November 2013

The incidence of conjugative R plasmids in multiple drug‐resistant strains of gram‐negative bacteria isolated in 1973 from patients in a 700‐bed general hospital in Tokyo and some properties of the R plasmids isolated are described. Conjugative R plasmids were found in 52 of the 96 strains (54%), from which 74 R plasmids were demonstrated. It is remarkable that the isolation frequency of R plasmids mediating quadruple‐ or five‐drug resistance was rather low, and the complete pattern of multiple resistance of the original isolates was only rarely transferred by conjugation. These results revealed the existing state of the distribution of R plasmids among hospital strains with multiple drug‐resistance.
Further Studies on Amplification of Cell‐Associated Immunological Memory by Secondary Antigenic Stimulus
Authors: Nakashima Izumi , Kato Nobuo
Published: November 2013

Using the hapten‐carrier system in which the dinitrophenyl group (DNP) served as a B cell reactive hapten and bovine serum albumin (BSA) or human gammaglobulin (HGG) as a T cell reactive carrier, changes in the hapten‐specific memory (B cell‐associated memory) and the carrier‐specific memory (T cell‐associated memory) after a secondary antigenic stimulus were analyzed in mice. Since an immunological adjuvant was indispensable in the induction of the primary increase in memory, antigen used for the primary antigenic stimulus was injected together with the capsular polysaccharide of Klebsiella pneumoniae (CPS‐K) which has already been shown to exhibit a potent adjuvant effect. With the cell‐transfer technique, it was found that the cell‐associated hapten‐specific memory for anti‐DNP antibody response to DNP‐BSA was truly amplified by the secondary injection of DNP‐HGG into mice primed with DNP‐HGG, and that the cell‐associated carrier‐specific memory as judged by the helper effect on anti‐DNP response to DNP‐BSA was also truly amplified by the secondary injection of BSA into mice primed with BSA. However, when memory was assessed in actively immunized mice, the secondary injection of BSA into mice primed with DNP‐BSA and HGG decreased anti‐DNP responsiveness to the tertiary injection of DNP‐BSA, whereas the secondary injection of DNP‐HGG secondarily increased anti‐DNP responsiveness. In mice primed with DNP‐BSA the titers of serum antibodies to BSA increased after the secondary injection of DNP‐BSA or BSA. From these results it has been concluded that, like B cell‐associated memory, T cell‐associated memory is also amplified by a secondary antigenic stimulus, although its expression is inhibited in actively immunized mice through negative control by their antibodies.
Effects of Homologous O‐Antibody on Host Responses to Lipopolysaccharide from Yersinia enterocolitica: Neutralization of Its Pyrogenicity
Authors: Ogata Sachio , Kanamori Masato
Published: November 2013

The pyrogenic activity of lipopolysaccharide (LPS) obtained from Yersinia enterocolitica was neutralized by the homologous O‐antiserum at the optimal antigen/antibody ratio. The level of neutralization in either antigen excess or extreme antibody excess was significantly lower than that at the optimal ratio. These facts suggested that the structure of the so‐called “lattice” of LPS/antibody complex might influence the neutralization of pyrogenic activity. Moreover, when LPS was gradually added to the antiserum to reach the optimal antigen/antibody ratio, the pyrogenic activity of LPS was only slightly neutralized by the antibody. The similar event to Danysz phenomenon that has been known in the diphtheria toxin‐antitoxin system was also observed in the LPS‐anti‐LPS system.
Microbial Adjuvant and Autoimmunity
Authors: Yokochi Takashi , Nakashima Izumi , Kato Nobuo , Asai Junpei
Published: November 2013

Experimental autoimmune thyroiditis could be produced in SMA, C3H/He and C57BL mice by repeated injection at intervals of 30 days of syngeneic thyroid extract mixed with the capsular polysaccharide of Klebsiella pneumoniae (CPS‐K) as a powerful adjuvant. The sequence of the development of autoimmune thyroiditis was histologically followed using SMA strain of mice, in which the thyroid lesions were most marked. A single injection of the thyroid extract mixed with CPS‐K induced interstitial infiltration of small lymphocytes and other mononuclear cells, and follicular architecture was partially damaged. At early times (5 to 12 days) after the secondary injection of the material, there were aggregation of polymorphonuclear neutrophilic leukocytes with formation of microabscesses in the follicles and hyaline degeneration of small vessels in the thyroid glands, suggesting that this stage of the thyroid lesions was expression of an Arthus reaction. The maximal severity of the thyroid lesions was reached at 30 days after the secondary injection. At this time, the thyroid lesions consisted of extensive infiltration of small lymphocytes, plasma cells and other mononuclear cells with complete loss of follicular architecture and mild proliferation of fibrous connective tissues. Even at 200 days after the secondary injection, there was no evidence of spontaneous regression and resolution of the thyroid lesions. Based on the histological findings, it seems likely that in our system cell‐mediated immunity plays a dominant role in initiation and maintenance of thyroiditis and humoral antibodies do so in its aggravation.
Separation of Four Components of the Phosphoenolpyruvate: Glucose Phosphotransferase System in Vibrio parahaemolyticus
Authors: Kubota Yoneo , Iuchi Shiro , Fujisawa Asako , Tanaka Shuji
Published: November 2013

Four classes of Vibrio parahaemolyticus mutants defective in the phosphoenolpyruvate: glucose phosphotransferase system (PTS) are described. They were phenotypically different, and were defective in different PTS components. The components designated tentatively as II, I, III, and H were separated by gel filtration of a wild‐type extract. Component II, which was specific for glucose and found in the particulate fraction, is probably membrane‐bound, glucose‐specific enzyme II. Both components I and H were soluble proteins, and the latter was relatively heat‐stable. Component I was required for phosphorylation of glucose, trehalose, fructose, mannose, and mannitol. Component H was also required for phosphorylating all the above sugars except fructose. These and some additional findings strongly suggest that components I and H correspond to enzyme I and HPr, respectively. Component III, a soluble heat‐stable protein, may be equivalent to the sugar‐specific factor III found in other organisms, although it seems to participate in phosphorylating two sugars, glucose and trehalose. There were evidences that mutants defective in components I and III were deficient in cyclic adenosine 3′,5′‐monophosphate synthesis under certain conditions.
Lipid Composition of Staphylococcus aureus and Its Derived L‐forms
Authors: Hayami Masaaki , Okabe Akinobu , Kariyama Reiko , Abe Masufumi , Kanemasa Yasuhiro
Published: November 2013

Two strains of Staphylococcus aureus (Newman and Tazaki) and their derived L‐forms were cultured in serum‐containing broth and the differences in their lipid compositions were analyzed. Cardiolipin accounted for more than 50% of the total phospholipid phosphorus in L‐forms, but for less than 25% in parent bacteria. The cardiolipin content of L‐forms was very high through all growth phases, although it increased gradually as growth proceeded. Significant amounts of cholesterol and its esters were present in parent strains and L‐forms, all of which incorporated serum cholesterol into the cell membrane. On the other hand, they could be detected in the L‐forms but not in the parent strains when they were cultured in serum‐free broth. To examine the ability of L‐forms to synthesize cholesterol, the cholesterol content of L‐forms cultured in serum‐free broth was compared with that of the medium. The results indicated that staphylococcal L‐forms could synthesize cholesterol and its esters. These differences in lipid composition suggested that modification of membrane lipids may occur as an adaptation‐al change in response to the disappearance of the cell wall.
Characterization of Bovine Parainfluenza Virus Type 3
Authors: Shibuta Hiroshi , Kanda Tadahito , Adachi Akio , Yogo Yoshiyuki
Published: November 2013

Bovine parainfluenza virus type 3 (PIV‐3) has a buoyant density of 1.197. The RNA of PIV‐3, like that of Sendai virus, is a single continuous chain which lacks polyadenylic acid sequences and tends to self‐anneal to a marked extent. It has a sedimentation coefficient of 42S and a molecular weight of 4.5 × 106, being slightly smaller than Sendai virus RNA (47S, 5.3 × 106).

PIV‐3 has 5 main structural proteins, of which 2 are glycoproteins. The molecular weights of protein1, protein2, protein3, glycoprotein1, and glycoprotein2 were estimated to be 79,000, 68,000, 35,000, 69,000, and 55,000, respectively. Protein2 was suggested to be nucleocapsid protein.
Complement Proteins and Macrophages
Authors: Kawamoto Yasuko , Ueda Masanaga , Ichikawa Hidetaka , Miyama Akio
Published: November 2013

Cobra venom factor was used for the detection of factor B synthesized by mouse peritoneal macrophages. This method was shown to be specific for factor B assay by neutralization by antimouse factor B antibody. The amount of factor B in the culture supernatant, assessed by this method, was found to be dependent on the medium used for cultivation of macrophages. The addition of 25% L cell‐conditioned medium to minimal essential medium (LCM‐MEM) enhanced the production of factor B and also of lysozyme. Kinetic analysis in LCM‐MEM showed that factor B produced by 6 × 104 cells/cm2 increased up to 72 hr and reached a plateau at 96 hr. The amounts of factor B and lysozyme produced in LCM‐MEM depended upon the number of macrophages. Production of factor B was completely inhibited by 1 μg of cycloheximide per ml and was restored by its removal.
Susceptibility of Germ‐free Mice to Infectious Megaenteron
Authors: Itoh Kikuji , Ueda Katsumoto , Fujiwara KÔSaku
Published: November 2013

Germ‐free (GF)‐ICR mice were shown to be less susceptible to oral inoculation with a pathogenic strain of Escherichia coli (E. coli 0115a, c: K(B)) than GF‐CF#1 mice. In GF‐CF#1 mice a large number of organisms were recovered from the intestinal wall from the cecum to the rectum 3 to 7 days after inoculation. Unlike those in GF‐CF#1 mice, lesions in GF‐ICR mice were localized in a part of the cecum and organisms were recovered only from the cecal wall and rarely from organs other than those of the alimentary tract. In both strains of mice, however, organisms were recovered in large number from the intestinal contents. Histopathology and immunofluorescence revealed organisms closely attached to the surface of the cecum, colon and rectal epithelia in GF‐CF#1 mice but only in a part of the cecal epithelium in GF‐ICR mice. After being in contact with conventional CF#1 mice for 21 days and then inoculated orally with the pathogenic E. coli, ex‐GF‐CF#1 mice died within 14 days with severe intestinal lesions, but ex‐GF‐ICR mice survived without lesions.
Induction of Delayed Type Hypersensitivity‐Like Skin Reaction by Peptidoglycans of Bacterial Cell Walls
Authors: Maeda Katsumasa , Koga Toshitaka , Onoue Kaoru , Kotani Shozo , Sumiyoshi Akinobu
Published: November 2013

Water‐soluble glycopeptides isolated from Lactobacillus plantarum and Staphylococcus epidermidis cell walls elicited a delayed type hypersensitivity (DTH)‐like skin reaction in rats previously immunized with Mycobacterium tuberculosis cell walls, but not in unimmunized rats. Histological examination of the skin reaction sites in immunized animals revealed a close similarity of this skin reaction to a typical DTH reaction with respect to the time course of development and the types of cells that infiltrated into the skin reaction sites, which were characterized by a predominant infiltration of mononuclear cells at 48 hr. This DTH‐like reaction was also demonstrated by immunizing the rats with the cell wall peptidoglycans of L. plantarum or S. epidermidis and skin testing them with homologous as well as heterologous peptidoglycans. The DTH‐like reaction appeared to be caused by peptidoglycans that exist in common in the cell walls of phylogenetically distant bacterial species. Furthermore, it was also suggested that the putative antigenic determinants) might include both the glycan chain and part of the peptide moieties of the cell wall peptidoglycan rather than either of the single moieties.
Macrophage Activation by Muramyl Dipeptide as Measured by Macrophage Spreading and Attachment
Authors: Tanaka Atsushi , Nagao Shigeki , Imai Katsuyuki , Mori Ryoichi
Published: November 2013

A synthetic bacterial cell wall constituent, muramyl dipeptide (MDP), was found to induce the enhancement of macrophage spreading and attachment on glass or plastic surfaces. Macrophages exposed to bacterial lipopolysaccharide or lymphokine‐containing cell supernatants showed similar enhancement. This finding supports the view that MDP activates macrophages. MDP was also found to enhance the viability of macrophages but to inhibit 3H‐thymidine incorporation by macrophages.
Role of Humoral Antibodies in Resistance to Mycoplasma pneumoniae Pneumonia in Hamsters
Authors: Hayatsu Eizo , Kawakubo Yasuaki , Yayoshi Masumi , Araake Minako , Yoshioka Morimasa , Nishiyama Yasukazu
Published: November 2013

Golden Syrian hamsters adoptively immunized with hyperimmune Mycoplasma pneumoniae rabbit antiserum, immunoglobulin (Ig) M‐rich (IgM) fraction, IgG‐rich (IgG) fraction, antiserum absorbed with either killed M. pneumoniae or killed Staphylococcus aureus organisms, or antiserum treated with 2‐mercaptoethanol (2‐ME) were examined for resistance against aerosol infection with virulent M. pneumoniae. Significant resistance to the establishment of infection in the respiratory tract was shown in hamsters pretreated with the untreated antiserum, IgG fraction or 2‐ME‐treated antiserum, whereas animals pretreated with the IgM fraction and the antisera absorbed with M. pneumoniae or S. aureus organisms were not significantly resistant. Histopathologically, lung lesions were markedly suppressed in animals with high resistance, but were typically pneumonic in animals with low or no resistance. The efficacy of adoptively administered serum preparations was closely related to their antibody titers. The results indicate that humoral antibody plays an important role in protection against experimental M. pneumoniae pneumonia in hamsters, although the participation of the cell‐mediated immune response was not determined.
Susceptibility of Influenza Viruses to Interferon and to Poly (I) · Poly (C) Determined by the Plaque Reduction Method
Authors: Machida Haruhiko , Kuninaka Akira , Yoshino Hiroshi
Published: November 2013

Susceptibility of eight strains of influenza A and B viruses to interferon and to poly(I) · poly(C) were determined by the plaque reduction method. All strains tested were slightly less susceptible than vesicular stomatitis virus (VSV) in an established line of canine kidney (MDCK) cells. The 50% plaque depression doses (PD50) of poly(I) · poly(C) for influenza A and B viruses were as high as 3.0‐ to 4.5‐fold and 6‐ to 18‐fold that for VSV, respectively. The amounts of interferon required to inhibit plaque formation of influenza A and B viruses by 50% were 3.0–6.2 and 7.3–15.2 units/ml, respectively. The ratio of PD50 of poly(I) · poly(C) for each strain of influenza viruses tested to that for VSV in chick embryo cells was almost the same as in MDCK cells. Furthermore, in chick embryo cells, the strains of influenza virus tested were demonstrated to be much more susceptible to poly (I) · poly(C) than both Newcastle disease virus and vaccinia virus. It is suggested that influenza viruses may be relatively susceptible to interferon and to poly(I) · poly(C).
Catabolite‐Like Repression of Extracellular Enzyme Production in Vibrio parahaemolyticus
Authors: Iuchi Shiro , Tanaka Shuji
Published: November 2013

Production of extracellular amylase and protease in Vibrio parahaemolyticus was repressed by various carbohydrates present in the medium. In addition, the protease production was repressed very strongly by peptones or casamino acids. Cyclic adenosine 3′, 5′‐monophosphate (cyclic AMP) added exogenously could reverse the repression of amylase production, but not that of protease production irrespective of the “repressors” used. Mutants of V. parahaemolyticus, which resembled the reported cya (adenylate cyclase) and crp (cyclic AMP receptor protein) mutants of Escherichia coli and related organisms, were examined for the exoenzyme production. Amylase production in the mutants was defective, while their protease production was not defective, but rather accentuated as compared with that in the parental strain.

These findings strongly suggest that amylase production is subject to catabolite repression mediated by cyclic AMP, whereas protease production is controlled by a repression mechanism which mimics in part, but may be distinct from catabolite repression.
A Complement Inhibitor Produced by Stachybotrys complementi, nov. sp. K‐76, a New Species of Fungi Imperfecti
Authors: Miyazaki Wasei , Tamaoka Hisashi , Shinohara Masanao , Kaise Hirotsugu , Izawa Taketoshi , Nakano Yoshimasa , Kinoshita Taroh , Hong Kyongsu , Inoue Kozo
Published: November 2013

A complement inhibitor, K‐76, was isolated and purified from the culture supernatant of a fungus, Stachybotrys complementi, nov. sp. K‐76, isolated from soil of Ishigaki Island, Okinawa. K‐76 is a sesquiterpene compound and it can be oxidized to a monocarboxylic derivative (K‐76 COOH), the sodium salt of which is very soluble and much less toxic than K‐76. K‐76 and K‐76 COOH both inhibited complement activation by either the classical or alternative pathway. They inhibited generation of the factor chemotactic to human polymorphonuclear leukocytes from human serum by aggregated immunoglobulin. When sensitized erythrocytes were treated with complement in the presence of K‐76 COOH, the resulting unlysed cells were found to be in the state of EAC1, 4b, 2a, 3b. Thus K‐76 COOH is considered to block mainly the C5 intermediate step. K‐76 COOH did not inhibit any proteases or esterases tested, except when tested at high concentration.
Studies on Streptococci: I. Distribution of Fecal Streptococci in Man
Authors: Watanabe Tsugio , Shimohashi Hirotaka , Kawai Yasuo , Mutai Masahiko
Published: November 2013

To understand the significance to the host of streptococci as part of the intestinal microflora, we first tried to investigate the distribution of human fecal streptococci on the species level. Of the selective media compared, KMN agar was more effective than the other media for the isolation of streptococci from human feces. We made an effort to improve streptococcal classification. Especially we used utilization of 1% pyruvate, 1% arginine, and 1% citrate for differentiation between Streptococcus faecalis and S. faecium. In a tellurite tolerance test, S. faecalis was distinguished more clearly from S. faecium in the medium containing 0.16 or 0.32% tellurite. We devised methods of presumptive identification of fecal streptococci from the results of the characteristics of 1,442 isolates. These methods enabled us to identify many strains rapidly. Different results in the distribution of species of streptococci between children and adults were observed. S. faecalis and S. faecium were isolated constantly from both groups. S. bovis and S. avium were isolated frequently from the feces of children. On the other hand, “viridans” streptococci, e.g. S. salivarius, S. mitis and S. MG‐intermedius were present at a high frequency in, and no S. avium could be isolated from, the feces of adults.
Biological Properties of Plaque‐Size Variants of Sendai Virus
Authors: Sugita Kenji , Maru Morimasa , Sato Kosaburo
Published: November 2013

Large (RL)‐ and small (RS)‐plaque variants of Sendai virus were isolated in cultures of LLCMK2 cells in the presence of trypsin and their biological properties were determined. The RL variant was more virulent to mice than the RS variant. The RL variant had a higher growth rate than the RS variant in multiple‐step growth in the presence of trypsin, but the two variants had an almost equal growth rate in its absence. Restoration of hemolytic activity and cleavage of the F protein of the RL variant were achieved by milder trypsin treatment than was needed for the RS variant.
Regulatory Factors of Lymphocyte‐Lymphocyte Interaction: I. Con A‐Induced Mitogenic Factor Acts on the Late G1 Stage of T‐Cell Proliferation
Authors: Kawano Michio , Namba Yuziro , Hanaoka Masao
Published: November 2013

DNA synthesis in murine lymphocytes was augmented by a soluble factor in the supernatant of serum‐free cultures of syngeneic spleen cells activated with concanavalin A (Con A). This so‐called mitogenic factor (MF), which is probably identical with interleukin II, partially purified by DEAE‐cellulose and Sephadex G‐75 chromatography, is a fairly homogeneous molecule of 17–25 × 103 daltons. By using partially purified MF, the role of MF in lymphocyte proliferation was investigated.

Pretreatment of lymphocytes with Con A for the first 3 hr of culture, which does not in itself induce cell proliferation, markedly augmented the effect of MF. The presence of MF, however, is necessary only in a restricted stage (s) of lymphocyte proliferation. The addition and removal of MF at various times during culture showed that MF exerts its effect on a process which occurs 3–6 hr before the beginning of DNA synthesis. These results strongly suggest that MF regulates the proliferation of mitogen‐stimulated lymphocytes by acting on a restricted stage (s) of the cell cycle.
Correlation between Production of Inflammatory Substances and Generation of Effector T Cells Mediating Delayed‐Type Hypersensitivity in Mice
Authors: Tamura Shin‐Ichi , Kojima Asato , Egashira Yasuyuki
Published: November 2013

In vitro exposure to human serum albumin (HSA) of splenic lymphocytes from mice sensitized for delayed‐type hypersensitivity (DTH) against HSA resulted in the release of substances that could induce a footpad inflammatory reaction with a maximum 6 hr after injection into normal mice. The substances were fractionated mainly in a molecular weight range of 30,000 to 70,000 daltons on Sephadex G‐200. The ability of sensitized lymphocytes to produce the substances was dependent on T cells, was antigen specific, and correlated well with the ability of the lymphocytes to mediate DTH reactions. Moreover, the substances were produced efficiently by the DTH effector cell population generated in the in vitro culture system and also by the effector cell‐enriched fractions on discontinuous bovine serum albumin gradients. These results suggest that the substances are produced by DTH‐effector cells.
Murine Defense Mechanism against Candida albicans Infection: II. Opsonization, Phagocytosis, and Intracellular Killing of C. albicans
Authors: Kagaya Keiko , Fukazawa Yoshimura
Published: November 2013

The phagocytic and intracellular killing activities of normal mouse phagocytes against Candida albicans were studied to elucidate the role of these activities in nonspecific and specific defense mechanisms. In the presence of fresh normal mouse serum, viable C. albicans cells were ingested by mouse peripheral blood leukocytes (PBLs) and peritoneal macrophages (PMPs) at the same rate. Serum‐chelation experiments indicated that the factors involved in the alternative complement pathway are opsonins for C. albicans. PBLs killed intracellular C. albicans more effectively than PMPs. Lymphokine‐activated PMPs manifested marked intracellular killing activity and the occurrence of increased superoxide anion‐ and singlet oxygen production, in the absence of increased myeloperoxidase (MPO) production, suggests that the enhanced, MPO‐independent, oxidative mechanism may play an important role in the candidacidal activity. Specific rabbit antibodies played no role in the phagocytosis and intracellular killing of C. albicans. These results suggest that PMNs and factors involved in the alternative complement pathway, and lymphokine‐activated macrophages play major roles in the protection of mice against C. albicans infection.
Mode of Subacute Sclerosing Panencephalitis (SSPE) Virus Infection in Tissue Culture Cells: III. Neurovirulence of Cell‐Free SSPE Viruses of Niigata‐1, Kitaken‐1, and Biken Strains
Authors: Ohuchi Masanobu , Ohuchi Reiko , Homma Morio
Published: November 2013

Cell‐free viruses recovered from virus‐carrying cultures of the Niigata‐1, Kitaken‐1, and Biken strains of SSPE virus were examined for neurovirulence. The cell‐free viruses were prepared by freezing and thawing or by EDTA treatment of the virus‐carrying cultures and inoculated into adult mice intracerebrally. A considerable number of the inoculated mice showed clinical signs about 1 to 5 weeks after the inoculation. The first symptom was hyperreactivity, which was followed by paresis and myoclonus. All of the affected mice fell in paralysis and finally died. The virus could be recovered from the moribund mice by cocultivation of the brain cells with Vero cells. Immunofluorescence staining of the brain tissue revealed that infected cells containing viral antigens were distributed sparsely. No inflammatory feature, however, was observed in the brain as far as examined and neutralizing antibody against SSPE virus was not detected in sera from the mice inoculated with the cell‐free SSPE viruses.
Pili Mediated Agglutination of Serratia marcescens in Human Urine
Authors: Amako Kazunobu , Ohshima Hisaaki , Yasunaka Kakuko , Kono Kenji
Published: November 2013

Of 51 strains of Serratia marcescens isolated from patients with urinary or respiratory tract infections, 35 agglutinated in human urine. The agglutinating strains possessed numerous pili which were morphologically distinct from common pili or type I pili. The diameter of the pili was 3 nm and the average length was 0.3 μm. Electron microscopic examination showed that 80% or more of the cells of the agglutinating strains and 0 to 8% of the cells of the nonagglutinating strains were piliated. When an agglutinating strain was heated at 55 C for 10 min, it lost its agglutinating capacity and concomitantly its pili. These results suggest that the agglutination might occur because of interactions between the pili and some factors in human urine. The urinary slime appears to contain these agglutinating factors.
Sequential Passages of Human Rotavirus in MA‐104 Cells
Authors: Urasawa Tomoko , Urasawa Shozo , Taniguchi Koki
Published: November 2013

Starting with a small amount of diarrheal feces containing human rotavirus (HRV), we succeeded in propagation of the virus using the roller culture technique with MA‐104 cells. Furthermore, we made a successful adaptation of HRV to a stationary culture and developed a plaque assay for the cell culture‐adapted viruses. The 3 culture‐adapted virus isolates, KU, YO, and 44 produced plaques (about 0.5–1.0 mm in diameter) under the overlay medium consisting of 0.6% purified agar, 3 μg of acetyl trypsin/ml and 50 μg of DEAE‐dex‐tran/ml. Subsequent plaque purification resulted in the formation of clear, larger plaques.

It was shown from the results of cross neutralization tests using the fluorescent focus reduction method that the three culture‐adapted HRV isolates were clearly different antigenically from bovine rotavirus (NCDV) and, further, that a noticeable difference in antigenicity also existed among the HRV isolates.
Immunological Responses of Hamsters in the Acquired Immune State to Mycoplasma pneumoniae Infection
Authors: Hayatsu Eizo , Kawakubo Yasuaki , Yayoshi Masumi , Araake Minako , Wakai Mariko , Yoshida Atsumasa , Yoshioka Morimasa , Nishiyama Yasukazu
Published: November 2013

Protective effects of vaccination of hamsters against Mycoplasma pneumoniae infection, evaluated according to the recovery of mycoplasmas and histopathological changes in the respiratory tract after challenge infection, persisted for at least 6 months after the final vaccination. Serum antibody levels reached a maximum in the second week after the last vaccination and decreased markedly between the first and the third months, but increased again in sera obtained from animals given booster injections. Metabolism‐inhibiting antibodies were detected in bronchial washings of animals showing high resistance obtained by vaccinal or passive immunization. Antiserum transfer was also effective for protection but cell‐mediated immune responses were not demonstrated in any animals up to 6 months after the vaccination. Even after 10 months, suppression of both mycoplasmal proliferation and lung lesions was apparent, and a single dose of the vaccine induced a significant booster effect. These findings suggest that (1) humoral immunity is more important than cell‐mediated immunity in resistance of hamsters to M. pneumoniae pneumonia, and (2) the antibody secreted in the respiratory tract may be involved in the local defense mechanisms.
A Simplified Micro‐Method for Cytotoxicity Testing Using a Flat‐Type Titration Plate for the Detection of H‐2 Antigens
Authors: Shiroishi Toshihiko , Sagai Tomoko , Moriwaki Kazuo
Published: November 2013

A new micro‐method for cytotoxicity testing using a flat‐type titration plate has been developed. This micro‐method is so simple that it involves no washing and staining procedures. In this method, cell viability is judged on the basis of the refractility change in the lymphocytes seen under the phase‐contrast microscope. This micro‐method has higher sensitivity and reproducibility than the dye exclusion method and 51Cr‐release assay. Fixation of cells with glutaraldehyde and tight covering of the plate allow us to read the result even after 3 weeks. These advantages of the new method enable us to deal with a large number of samples at one time.

We also introduce a rapid technique for the preparation of a cell suspension from the lymph node without killing the animal.
Carbohydrate Fermentation by Clostridium difficile
Authors: Nakamura Shinichi , Nakashio Satoshi , Yamakawa Kiyotaka , Tanabe Naomi , Nishida Shoki
Published: November 2013

Biochemical properties of Clostridium difficile were reinvestigated for the practical identification of the organism in clinical laboratories. Bacterial growth in 2% proteose peptone medium supplemented with 0.01% l‐cysteine · HCl and 0.1% agar supported sufficient growth to read the fermentation results just as well as did pre‐reduced anaerobically sterilized medium. Incubation for 2 days was long enough for determining the ability to ferment fructose, glucose, mannitol, mannose, melezitose, and sorbitol. All of the 82 strains liquefied 2% but not 10% gelatin. The significance of mannitol fermentation and gelatin liquefaction is stressed since C. difficile is the only species fermenting mannitol among the gelatin‐liquefying species of Clostridia having subterminal spores.
Characterization of Cells Infiltrating the Lungs of X‐Irradiated and Nude Mice after Influenza Virus Infection
Authors: Shimomura Emiko , Suzuki Fujio , Ishida Nakao
Published: November 2013

After X‐irradiated and nonirradiated mice (C3H/He) as well as athymic nude mice and haired littermates (BALB/c) were infected with influenza A virus (Kumamoto strain, H2N2), they were examined for survival period, the development of consolidation in the lungs and the characteristics of the cells infiltrating the lung tissues. In two different T‐cell deficient groups, there was a definite delay in the development of consolidation compared with their respective controls and this was reflected in prolonged survival periods: 5 days longer for irradiated mice and 6 days longer for nude mice. In both T‐cell deficient and normal groups, about 70% of the cells obtained from consolidated lung tissues after virus infection were found to be small lymphoid cells and there were no morphological differences between the T‐cell deficient and normal groups. None of these small lymphoid cells from the peripheral blood or the spleens of T‐cell deficient mice responded to concanavalin A. In the lungs of both X‐irradiated mice and nude mice, however, a definite increase in cells having natural killer activity was found at the late stages of the influenza infection, suggesting their participation in the development of consolidation.
Keratinolytic Fungi in Egyptian Soils
Authors: Abdel‐Fattah Hassan M. , Moubasher Abdel‐Aal H. , Maghazy Soad M.
Published: November 2013

The frequency of occurrence of keratinolytic fungi in seventy soil samples, collected from different sites in Upper Egypt and in the coastal area of the Mediterranean, was determined by baiting with human and animal hairs and pigeon feathers.

Twenty‐one species, in addition to an unidentified species, which belong to sixteen genera were collected. Chrysosporium indicum, C. tropicum, C. keratinophilum, and Microsporum gypseum were the most frequent fungal species recovered from the baited soils.

The soil samples collected from the salt marshes of the coastal Mediterranean area were completely free from any keratinolytic fungi, whereas the soil samples collected from cultivated soils contributed species, ranging from one to four species.
An Internal View of the Spherical Body of Treponema macrodentium as Revealed by Scanning Electron Microscopy
Authors: Umemoto Toshihiko , Namikawa Isamu , Yoshii Zensaku , Konishi Hisanori
Published: November 2013

The osmium‐dimethyl sulfoxide‐osmium method for clear visualization of intracellular structure was used to observe the detailed inner structure of the spherical bodies produced in vitro by a human oral treponeme. Scanning electron microscopy of the cracked spherical body revealed no morphological differences between the outer and inner surfaces of the spherical body membrane, and that multiple folded or somewhat linear main bodies adhere closely to the inner surface. In addition, axial flagella partially free from the main bodies spread widely within the body to make a network, and a number of blebs ranging from approximately 1 μm to 0.2 μm in diameter were located near the terminal or subterminal areas of the main bodies. The origin of the blebs and the mechanism of spherical body formation are discussed.
Gelation of Limulus Amoebocyte Lysate by Simple Polysaccharides
Authors: Mikami Takeshi , Nagase Toshihiko , Matsumoto Tatsuji , Suzuki Shigeo , Suzuki Masuko
Published: November 2013

The Limulus lysate gelation activity of several simple polysaccharides including yeast mannans and bacterial dextrans was investigated. The mannans from Saccharomyces cerevisiae wild and mutant strains possessing dense branches showed positive gelation activity at concentrations of 1 μg/ml or more regardless of differences in their chemical structure. However, two synthetic mannans possessing linear structures with α1←2 and α1←6 linkages also gave positive reactions at concentrations of 10 μg/ml or more and 500 μg/ml or more, respectively. The dextran from Leuconostoc mesenteroides IAM 1046 consisting of a dense branching moiety displayed reactivity at concentrations of 100 μg/ml or more, while the dextrans devoid of such branches were negative in this reaction. The optimal concentration for Limulus lysate gelation could not be determined for any of the polysaccharides and lipopolysaccharides (LPS) tested in this study. The gelation activity of the polysaccharides was stable to treatment with 100 mm NaOH at 30 C for 72 hr. The minimum concentration for the gelation activity of LPS treated with 100 mm NaOH under the same conditions was reduced from 10−6~−9 μg/ml to 1–10 μg/ml. The above findings demonstrate that the major part of Limulus lysate gelation activity of LPS depends on the alkali‐degradable lipid A moiety, and that such simple polysaccharides are also able to participate in this activity even though the extent of participation is very low.