bluuebunny/crossref_metadata_2025 huggingface dataset:
A Highly Sensitive Method for Detection of 32P Incorporation into Acid‐Soluble Compounds and Its Application to Evaluate ATP‐Forming Ability of Bacillus megaterium QM B1551 Spores at the Very Early Stage of Germination
Authors: Otani Mieko , Umezawa Chisae , Sano Keiji
Published: November 2013

A method for specific removal of [32P]orthophosphate (Pi) as phosphomolybdate‐triethylamine complex was slightly modified by repeating the Pi precipitation procedures in the presence of unlabeled Pi, which resulted in a complete removal of 32Pi (>99.98%). Using this modified method, we determined 32P incorporation into acid‐soluble compounds in order to evaluate the ATP‐forming ability of Bacillus megaterium spores at the very early stage of germination. Addition of fructose as a substrate started the 32P incorporation later than a few min after triggering germination. This delay of a few min was well coincident with the onset of 3‐phosphoglycerate (3PGA) breakdown, indicating that fructose metabolism and the accompanying aerobic ATP formation were initiated only after fructose phosphorylation by the ATP derived from anaerobic breakdown of endogenous 3PGA. In contrast, addition of glucose started incorporation of 32P into acid‐soluble compounds immediately after germination. In the latter case, NADH generated by glucose oxidation to gluconate (catalyzed by glucose dehydrogenase) might serve as an initial ATP source without depending on 3PGA breakdown and glucose metabolism via the Embden‐Meyerhof pathway.
Effect of Antibiotics on Immediate Hypersensitivity Reactions in vitro: Suppression of IgE‐Mediated Histamine Release from Peripheral Blood Basophils by Fosfomycin
Authors: Ida Shiroh , Shindoh Yuriko , Takishima Tamotsu
Published: November 2013

The effect of antibiotics on allergic reactions was studied in vitro using the release of histamine from human peripheral blood leukocytes (basophils) after incubation with anti‐IgE. For the several antibiotics we tested, including beta‐lactams and aminoglycosides, none had the capacity to enhance antigen‐induced histamine release, but some of them (minocycline, polymyxin B, and fosfomycin) suppressed the release of histamine in a dose‐dependent manner. Since fosfomycin has proved to be capable of suppressing IgE‐mediated histamine release non‐cytotoxically, the effect of fosfomycin on histamine release induced by other secretagogues was further studied. The suppression of histamine release was also demonstrated when the leukocytes, preincubated with fosfomycin, were challenged with either Ca ionophore A 23187 or a synthetic peptide, formyl‐methionyl‐leucyl‐phenylalanine (FMLP). We concluded that some antibiotics, particularly fosfomycin, have the capacity to suppress histamine release mediated by various secretagogues, suggesting they may possess an anti‐allergic property as well as a bactericidal activity.
A Simple Method for Quantitative Determination of Bacterial Adherence to Human and Animal Epithelial Cells
Authors: Valentin‐Weigand Peter , Chhatwal Gursharan S. , Blobel Hans
Published: November 2013

A simple quantitative method for determining bacterial adherence to epithelial cells was devised. The method involved incubation of fluorescein‐labeled bacteria with oral epithelial cells. Non‐adherent bacteria were subsequently removed by Percoll density centrifugation. The degree of bacterial adherence was then determined fluorospectrophotometrically by measuring the fluorescein extracted from the adherent bacteria. The degree of adherence corresponded well with the microscopic counts of adherent bacteria. The method proved to be suitable for assaying adherence of both gram‐positive and gram‐negative bacteria to human as well as animal epithelial cells.
Low Molecular Weight Factors Displaying Augmenting Activity for Human Antibody Production in vitro
Authors: Haraguchi Soichi , Hongo Teruaki , Ishihara Hideki , Matsuo Tetsumichi , Yoshida Takato O.
Published: November 2013

Dialyzable low molecular weight antibody‐augmenting factors (LMAAF) were found in the culture supernatant of human tonsillar lymphocytes which were not stimulated by antigen and/or mitogen in vitro. Phagocyte‐depleted nylon wool‐adherent lymphocytes (M−Ny+ cells) were responsible for the release of the LMAAF. Marbrook's culture system was adopted to assay for the LMAAF. The M−Ny+ cells, which were cultured without antigen and/or without mitogen in the reservoir of Marbrook's diffusion culture vessel, released the LMAAF, which diffused across a dialysis membrane and significantly augmented the pokeweed mitogen (PWM)‐induced plaque‐forming cell (PFC) response of phagocyte‐depleted lymphocytes (M− cells) cultured in the inner vessel. Phagocyte‐depleted nylon wool‐passed lymphocytes (M−Ny− cells) cultured in the reservoir could not augment the PWM‐induced PFC response of the M− cells cultured in the inner vessel. The exuded fluid, which was the dialysate of the culture supernatant of the M−Ny+ cells ultrafiltrated with dialysis tubing, also enhanced the PFC response of M− cells cultured in 24‐well multi plates. The exuded fluid also augmented the total IgM and IgG production of human tonsillar and peripheral blood lymphocytes measured by enzyme‐linked immunosorbent assay (ELISA) systems. Gel filtration chromatography on Sephadex G‐25 Superfine column showed that the LMAAF activity was demonstrated in the fractions corresponding to a molecular weight (m.w.) of 362 to 1,355 and a m.w. of 3,560 to 5,700, with a peak activity at about 4,500 dalton. The LMAAF were inactivated by treatment with proteinase K, but not by trypsin, α‐chymotrypsin, RNase, and DNase, and were stable when treated at 56 C for 60 min. The dialysates of culture supernatants from two out of seven Epstein‐Barr virus (EBV)‐transformed M−Ny+ cell lines showed LMAAF‐like activity. These results indicate that phagocyte‐depleted nylon wool‐adherent lymphocytes, possibly B cells, release low molecular weight factors displaying augmenting activity for human antibody production in vitro.
The Interaction of Recombinant IL‐2 with Human Resting Lymphocytes
Authors: Iseki Rieko , Koide Yukio , Ueda Ryuzo , Kondo Nobuo , Hamuro Junji , Yoshida Takato O.
Published: November 2013

Several lines of evidence suggest that subsets of resting lymphocytes naturally express interleukin‐2 receptors (IL‐2·R). Recombinant IL‐2 (rIL‐2) induced the enhancement of natural killer (NK) activity, the generation of activated killer (AK) cells, the proliferation of resting lymphocytes, and the production of interferon‐γ (IFN‐γ) in lymphocyte cultures. The subsets of lymphocytes mediating these responses appeared to be heterogeneous, but reside predominantly in nylon wool‐passed non‐T, non‐B cells (“lnull cells” or T3‐ cells); in response to rIL‐2, only Leul 1+T3‐ cells showed enhanced NK activity, and both Leul 1+T3‐ and Leul 1‐T3‐ cells showed predominantly AK activity, proliferation and production of IFN‐γ. These findings suggest that the T3‐ fraction (null cell fraction) contains predominantly cells expressing IL‐2·R at the resting state. Unlike the case with activated T cells, however, none of these responses was blocked by any of three monoclonal antibodies to IL‐2·R, including anti‐Tac antibody at any dilution. These results indicate that IL‐2·R on the resting T3‐ cells possess unique biological features compared to those on activated T or B cells. A most likely explanation is that T3‐ cells possess higher affinity IL‐2·R than activated T or B cells. Other possibilities are also discussed.
Effect of Cytolytic Infection on Maintenance of Resistance to HVJ (Sendai Virus) in an Altered BHK Cell Culture
Authors: Yokoo Junko , Miyadai Toshiaki , Yokochi Takashi , Kimura Yoshinobu
Published: November 2013

Altered baby hamster kidney (BHK‐R) cells were serially cultured in the continuous presence of hemagglutinating virus of Japan (HVJ). These cells showed a distinct resistance to superinfection with the homologous HVJ. This resistance of BHK‐R cells gradually disappeared after serial passages in the presence of ultraviolet‐irradiated HVJ particles which lost infectivity but still preserved hemagglutinating and neuraminidase activities. When BHK‐R cells were serially cultured in the presence of a temperature‐sensitive mutant of HVJ at non‐permissive temperature, the cells also lost the resistance. The resistance of BHK‐R cells remained unchanged, even after prolonged incubation in virus‐free maintenance medium under the conditions of no cell division. It was suggested that killing of virus‐sensitive cells, which were generated during cell proliferation, was required for maintenance of the resistance.
Physicochemical Characterization of A and B Subunits of Shiga Toxin and Reconstitution of Holotoxin from Isolated Subunits
Authors: Yutsudo Takashi , Honda Takeshi , Miwatani Toshio , Takeda Yoshifumi
Published: November 2013

The A and B subunits of Shiga toxin were isolated by high performance liquid chromatography and their physicochemical properties were examined. The A subunit of Shiga toxin purified from culture supernatant was not nicked, but it could be nicked in vitro by trypsin. The isoelectric points of the A and B subunits were determined to be 8.2 and 5.8, respectively. Amino acid compositions of the two subunits were also determined. The isolated A and B subunits were reconstituted to form active holotoxin which showed lethal activity to mice which was similar to that of native Shiga toxin.
Possible Existence of a Novel Amphipathic Immunostimulator in the Phenol‐Water Extracts of Mycobacteriaceae
Authors: Ikeda‐Fujita Takako , Kotani Shozo , Tsujimoto Masachika , Ogawa Tomohiko , Takahashi Ichiro , Takada Haruhiko , Shimauchi Hidetoshi , Nagao Shigeki , Kokeguchi Susumu , Kato Keijiro , Yano Ikuya , Okamura Haruki , Tamura Toshishide , Harada Kazuhiro , Usami Hiroko , Yamamoto Akihiko , Tanaka Shigenori , Kato Yoshiko
Published: November 2013

The extracts having diverse immunostimulating activities were obtained as a water‐phase fraction from four bacterial species representing the 4 genera (Mycobacterium, Nocardia, Gordona, and Rhodococcus) of Mycobacteriaceae by the phenol‐water method, which is commonly used for extraction of endotoxic lipopolysaccharides (LPS) from gram‐negative bacteria and amphipathic substances from gram‐positives. These fractions, especially those of G. aurantiaca and R. terrae, showed strong stimulatory effects on murine splenocytes, macrophages of mice and guinea pigs, the immunoadjuvant activities in guinea pigs and mice, and the distinct activities inducing a tumor necrosis factor and interferons α/β and γ in primed mice. The fractions from G. aurantiaca and R. terrae exhibited potent pyrogenicity and the ability to activate the clotting enzyme cascade of the horseshoe crab (Tachypleus tridentatus). Some of these biological activities were not very different from the potency of the reference endotoxic LPS derived from Escherichia coli or Fusobacterium nucleatum. But the test fractions neither showed the activity to prepare rabbit skin to the local Shwartzman reaction, nor reacted with anti‐lipid A conventional and monoclonal antibodies. Furthermore, unlike LPS, these fractions stimulated the splenocytes of C3H/HeJ mice (LPS‐Nonresponder). Although the fractions showing the above biological activities have not yet been adequately purified, they contained polysaccharides, whose main constituent sugar is mannose with a smaller amount of arabinose, fatty acids consisting primarily of palmitic, stearic, and tuberculostearic acids, and small amounts of peptides and amino sugars. Since components characteristic of known immunomodulators of bacterial origin, namely endotoxins (lipid A's), cell wall peptidoglycans, lipoteichoic acids, cord factors (trehalose dimycolates), or deoxyribonucleic acids, were practically not detected in these fractions, the agent responsible for the above bioactivities is considered to be a novel substance different from the known, bacterial immunomodulators.
Antigenic and Biochemical Characterization of Poliovirus Type 1 Isolates
Authors: Hara Minoru , Arita Mineo , Yamazaki Zenichi , Hagiwara Akio , Saito Yoshiko
Published: November 2013

By the introduction of Sabin oral poliovirus vaccine, the circulation of wild type polioviruses has virtually disappeared in Japan. However, an outbreak of poliomyelitis associated with sporadic transmission of type 1 wild strain occurred in Nagano in 1980. Furthermore, we found that some type 1 wild strains were introduced into Japan from abroad in 1981. In recent surveys, the two poliovirus type 1 isolates which have non‐vaccine‐like antigenic character were detected in Aichi. Then, an investigation to trace the origin of these strains was performed, by using intratypic serodifferentiation and biochemical techniques. Electrophoretic migration patterns of their structural polypeptides were quite different from the vaccine virus. In the oligonucleotide mapping, however, one of them gave patterns very similar to those of the vaccine virus. We could conclude that one originated most probably from wild strains, and the other was an antigenic variant derived from the vaccine virus. It showed that oligonucleotide mapping was a very useful method for identification of antigenic modified Sabin type 1 derivatives.
Inhibitory Effect of Capsular Antigen of Vibrio vulnificus on Bactericidal Activity of Human Serum
Authors: Shinoda Sumio , Kobayashi Masakazu , Yamada Hiroaki , Yoshida Shin‐Ichi , Ogawa Midori , Mizuguchi Yasuo
Published: November 2013

Opaque (Op) and translucent (Tr) colonial variants were isolated from Vibrio vulnificus strains. Op‐type variants were more resistant than the isogenic Tr‐type variants, but the survival rate of the Op‐type variants varied with the strains. Antisera were prepared by immunizing rabbit with whole cells of Op and Tr variants of some strains, in which the difference of the sensitivity between Op and Tr cells was remarkable. Then agglutination tests with their living and heat‐killed cells were carried out. The results suggested the presence of capsular antigen in Op cells and its absence in Tr cells, with the exception of the existence of a slight amount of capsular material in Tr variants of strain L‐180. The thin capsular layer of Tr cells of strain L‐180 was also demonstrated electron microscopically, but the layer was thinner than that of the isogenic Op cells. Results of determination of sugar content in the extracted capsular fraction also showed that Op to Tr transformation was due to loss of capsular antigen of the cells. These results confirmed the morphological studies previously reported which suggested the prevention of host defense system by the capsular material of the vibrio.
Effects of a New Antibiotic, Isohematinic Acid, on the Resistance of Mice to Experimental Infections
Authors: Shiraishi Akio , Katayama Toshiaki , Matsuda Tomoko , Itoh Yasuhiro , Haneishi Tatsuo
Published: November 2013

Isohematinic acid, an antibiotic newly isolated from the culture broth of Actinoplanes philippinensis SANK 61681, was assessed for its ability to enhance nonspecific resistance to bacterial infections against Escherichia coli and Pseudomonas aeruginosa in mice. This agent, as well as BM 12,531 (Azimexon), was found to prolong the survival of normal mice infected with E. coli and also of compromised mice infected with either E. coli or P. aeruginosa, whose defense system had been deteriorated by treatment with carboquone, an alkylating agent. Like BM 12,531, isohematinic acid administered to normal mice significantly increased the nitroblue tetrazolium reducing potency of polymorphonuclear leucocytes (PMN), indicating that the microbicidal activity of PMN was enhanced by these agents. In addition, in the compromised mice these agents were able to restore the number of peripheral blood leucocytes, which had been reduced to about 30% of the normal level by carboquone. These results suggest that isohematinic acid, like BM 12,531, enhances nonspecific resistance to these bacterial infections by stimulating the microbicidal activity of PMN and inducing leucocytosis.
Oral Immunization in Adult Mice to Live and Heat‐Killed Vibrio cholerae
Authors: Fujisawa Haruo , Tsuru Sumiaki , Taniguchi Mariko , Zinnaka Yutaka
Published: November 2013

Effects of systemic and intestinal local immune responses in mice fed ad libitum and forcedly with live or heat‐killed Vibrio cholerae on the elimination of vibrios from the intestine were investigated. In mice fed with live vibrios, ad libitum feeding could induce potential delayed‐type hypersensitivity and rapid production of vibriocidal antibody in the serum whereas forcedly feeding suppressed the delayed‐type hypersensitivity and retarded the antibody production. In contrast, when killed microorganisms were used as antigens, significant delayed‐type hypersensitivity and rapid response of vibriocidal antibody were induced in forcedly fed mice although ad libitum feeding suppressed the induction of the delayed‐type hypersensitivity and retarded the production of vibriocidal antibody. The elimination of vibrios from the intestine of mice was promoted in both mice groups fed ad libitum and forcedly with live vibrios but not with killed microorganisms. Total IgA in the intestinal contents of mice fed with live vibrios both ad libitum and forcedly were higher than those of mice fed with killed antigens. In addition, when the extracts of intestinal contents were absorbed by live antigens, IgA contents in mice fed with live vibrios were reduced more markedly than those in mice immunized orally by the feeding with killed antigens. These findings suggested that the elimination of vibrios from the organ was closely related to local IgA antibody response to heat‐labile substance of live Vibrio cholerae.
Antigenic Variation of Newcastle Disease Viruses Isolated from Wild Ducks in Japan
Authors: Kawamura Meiko , Nagata‐Matsubara Kayo , Nerome Kuniaki , Yamane Nobuhisa , Kida Hiroshi , Kodama Hiroshi , Izawa Hisao , Mikami Takeshi
Published: November 2013

Nineteen strains of Newcastle disease virus (NDV) isolated from wild ducks in Japan were placed into 4 distinct antigenic groups on the basis of their reactivities to 8 monoclonal antibodies against the HN molecule of NDV in hemagglutination inhibition tests. The NDV strains of duck origin were antigenically distinct from NDV‐B1 and NDV‐Miyadera originated from chickens, and varied in their virulence to chicken embryos. No apparent correlation was found between the antigenicity of the HN molecule and virulence.
Purification and Immunochemical Properties of a Wall Protein Antigen from Clostridium difficile ATCC 11011
Authors: Takumi Kenji , Takeoka Aya , Kawata Tomio
Published: November 2013

A wall‐surface protein antigen, designated 32K antigen, was extracted from whole cells of Clostridium difficile strain ATCC 11011 with phosphate buffered saline and purified by ion‐exchange chromatography, gel filtration, and chromatofocusing. The 32K antigen preparation was determined to be highly homogeneous by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis. The amino acid composition of the antigen was characteristic in the predominance of the acidic amino acids, the very low contents of methionine and histidine, and the lack of cysteine. A monomeric molecular weight of the 32K antigen was estimated to be 32,000 by SDS‐PAGE and 30,200 by sedimentation equilibrium. The antigen exhibited two isoelectric forms (IP, 4.12 and 3.96). Neither carbohydrate nor phosphorus was detectable in the antigen. The antigen was relatively resistant to trypsin but sensitive to pepsin. Immunoblot analysis of the wall proteins isolated from other strains of C. difficile probed with monospecific antiserum against the antigen from ATCC 11011 showed that the antigenicity of 32K wall protein was common among some of the strains containing 32K wall proteins.
Response of Ligated Rabbit Ileal Loop to Clostridium perfringens Type C Strains and Their Toxic Filtrates
Authors: Yamagishi Takayoshi , Gyobu Yotaku , Sakamoto Kenichi , Ishisaka Shintaro , Saito Kimie , Morinaga Shinichi , Katsuda Shogo , Umei Tamiko , Konishi Kenichi
Published: November 2013

The vegetative cells and toxic filtrates of Clostridium perfringens type C strains were injected into ligated rabbit ileal loops and the responses were observed. Out of 12 strains examined, 2 strains showed positive reaction in this test, when the vegetative cells were injected. One of these 2 strains was an enterotoxigenic and beta‐toxigenic and the other was beta‐ and delta‐toxigenic but not enterotoxigenic. Culture filtrates containing beta or delta toxin also showed fluid accumulation in the rabbit ileal loop. Histological findings of loops injected with culture filtrates containing beta toxin showed separated and effaced villi, hemorrhage in the mucosa, engorged vessels, inflammatory cell infiltration, and hyperplasia of the lymphoid tissue.
Application and Limitations of Differential Hybridization in the Isolation of T Cell‐Specific cDNA Clones
Authors: Shigemoto Kazuhiro , Kanno Masamoto , Sakamoto Tohru , Imai Kenji , Taniguchi Masaru
Published: November 2013

We investigated the limitations and effectiveness of differential hybridization in the cloning of T cell‐specific cDNA (complementary DNA) molecular clones. By using the technique with T cell and B cell cDNA probes, together with Northern blot analysis, we successfully isolated cDNA clones exclusively expressed in T cells from 1 × 104 plaque‐forming units of a T cell hybridoma. These clones represent 0.068% of the mass of the cytoplasmic mRNA. Our result shows that differential hybridization is an effective procedure when used in combination with Northern blot analysis for screening of genes selectively expressed in T cells.
Further Evidence for Heterogeneity of Fcγ‐Receptors on Guinea Pig Splenic B and T Lymphocytes: Analysis Using Monoclonal Antibodies to Two Distinct Types of Fcγ‐Receptor
Authors: Yamashita Toshiyuki , Koyama Jiro
Published: November 2013

In our previous paper, we reported that guinea pig splenic lymphocytes expressed two distinct Fc‐receptors for homologous IgG (FcγRs), one monospecific for IgG2 (Fcγ2R) and the other bispecific for IgG1 and IgG2 (Fcγ1/γ2R), when analyzed by EA‐rosette assay. These FcγRs on the cells were further studied by using two monoclonal antibodies toward the FcγRs on guinea pig peritoneal macrophages (anti‐Fcγ1/γ2R and anti‐Fcγ2R antibody). The anti‐Fcγ1/γ2R antibody completely inhibited the rosette formation of splenic lymphocytes with IgG1‐sensitized sheep erythrocytes [EA(IgG1)]. On the other hand, EA(IgG2)‐rosette formation was inhibited partially by anti‐Fcγ2R but not by anti‐Fcγ1/γ2R antibody. Complete inhibition of the EA (IgG2)‐rosette formation was achieved by simultaneous additions of both anti‐Fcγ2R and anti‐Fcγ1/γ2R antibodies. The binding of IgG2 antibody complexed with ovalbumin to the cells was partially inhibited by either anti‐FcγR antibody, and complete inhibition occurred in the presence of both the antibodies, indicating that two types of FcγR, Fcγ1/γ2R, and Fcγ2R, are expressed on the cells. The determination of these FcγRs on B and T lymphocytes by two‐color flow cytometry showed that about 52% of B lymphocytes expressed Fcγ1/γ2R alone and 32% of the cells expressed both the FcγRs. On the other hand, about 12% of T lymphocytes was found to express Fcγ2R alone and the cells expressing Fcγ1/γ2R were in the minority (3.8%). T lymphocytes expressing both the FcγRs were not detected. These results show that guinea pig B lymphocytes bear two types of FcγRs and are heterogeneous with regard to their FcγRs and that T lymphocytes express Fcγ2R mainly.
Large Plasmid in Shigella boydii Is Responsible for Epithelial Cell Penetration
Authors: Takahashi Kazuya , Yoshida Yoko , Okamura Noboru
Published: November 2013

A large plasmid in a virulent Shigella boydii 5 strain was transferred to plasmid‐cured avirulent strains of S. boydii 5, S. boydii 12, S. sonnei form II, and Escherichia coli K12. The transconjugants acquired the ability to invade tissue culture cells, which indicated that the large plasmid in S. boydii is responsible for epithelial cell invasiveness.
Biological Activities of Lipid A from Vibrio parahaemolyticus: Stimulation of Murine Peritoneal Macrophages
Authors: Bandekar Jayant Ramachandra , Nerkar Devdas Punjaji
Published: November 2013

The toxicity and macrophage stimulating property of Vibrio parahaemolyticus lipid A was studied. The LD50 dose of lipid A in galactosamine‐sensitized mice was found to be 0.6 μg when injected intraperitoneally. Administration of lipid A resulted in stimulation of peritoneal macrophages as evident by increase in their cellular RNA contents and lysosomal enzyme activities. The treatment also caused enhancement in the phagocytic activity of macrophages.
T Cell Subsets Responsible for Clearance of Sendai Virus from Infected Mouse Lungs
Authors: Iwai Hiroshi , Machii Kenji , Otsuka Yuko , Ueda Katsumoto
Published: November 2013

T cell subsets responsible for clearance of Sendai virus from mouse lungs determined by adoptive transfer of immune spleen cell fractions to infected nude mice. T cells with antiviral activity developed in spleens by 7 days after intranasal infection. Spleen cell fractions depleted of Lyt‐2+, Lyt‐1+, or L3T4+ cells showed antiviral activity in vivo, although the degree of the activity was lower than that of control whole spleen cells. The antiviral activity of the Lyt‐2+ cell‐depleted fraction was consistently higher than that of L3T4+ (Lyt‐1+)‐depleted cells. In vitro cytotoxic activity against Sendai virus‐associated, syngeneic lipopolysaccharide‐blast cells was detected in stimulated cells from intraperitoneally immunized mice but was lost after depletion of Lyt‐2+ cells. Multiple injection of anti‐Sendai virus antibody into infected nude mice had no effect on lung virus titer. These results indicate that L3T4+ (Lyt‐1+) and Lyt‐2+ subsets are cooperatively responsible for efficient clearance of Sendai virus from the mouse lung.
An mpo Mutant of Bacillus subtilis Requires a Higher Concentration of Manganese than the Wild Type Strain for Normal Sporulation
Authors: Matsuzaki Shigenobu , Kobayashi Komei , Kobayashi Yasuo
Published: November 2013

mpo‐1 Mutation, which causes the overproduction of at least two membrane and one cytoplasmic proteins, induced high manganese concentration‐dependent sporulation in Bacillus subtilis. Sporulation of the mpo‐1 mutant was blocked at stage 0‐I by a low concentration of manganese.
Persistent Infection with Herpes Simplex Virus Type 1 in an Ia Antigen‐Positive Murine Macrophage Cell Line
Authors: Tang Jie‐liu , Yamamoto Masahiro , Sakuma Shunji , Mori Ryoichi , Nagayama Ariaki
Published: November 2013

The interaction of herpes simplex virus type 1 (HSV‐1) with murine macrophage cell lines was examined. The cell lines appeared to be moderately permissive for HSV‐1 replication, though the yield of the virus was limited compared with that in Vero cells. Furthermore, the murine macrophage cell line SL‐1, bearing Ia antigen, was persistently infected with HSV‐1 for over one year, and was designated SL‐1/KOS. Persistent infection could not be established in an Ia antigen‐negative macrophage cell line, SL‐4. In the SL‐1/KOS culture, there was a small number of infected cells as revealed by infectious center assay. Treatment with monoclonal antibody against HSV‐1 cured the persistent infection. Therefore maintenance of the persistent infection is considered to be due to a carrier culture consisting of a minority of infected cells and a majority of uninfected cells. In the SL‐1/KOS cultures a low level of interferon (IFN) was found. When a large amount of exogenous recombinant murine IFN‐β (105–106 international units/ml) was added to the culture, virus production diminished to undetectable levels. These results suggest that IFN plays an important role in the maintenance of persistent infection. In long‐term persistently infected cultures, syncytium formation appeared and the virus from such cultures had a different DNA structure from that of the virus originally used for infection as revealed by restriction endonuclease analysis.
Effect of Temperature on Antibacterial Activity of Lidocaine to Staphylococcus aureus and Pseudomonas aeruginosa
Authors: Taki Yoshiharu , Seki Keiko , Ikigai Hajime , Nishihara Shoko , Ueno Hiroshi , Murota Kagehisa , Masuda Shogo
Published: November 2013

The effect of temperature on the antibacterial activity of lidocaine to Staphylococcus aureus and Pseudomonas aeruginosa was investigated in vitro. At 10 C at which S. aureus organisms do not grow and might be metabolically inactive, the antibacterial activity of lidocaine to S. aureus was not observed in a concentration of 1%, which was quite antibacterial to S. aureus at 37 C. On the other hand, at 40 C a conspicuously increased antibacterial activity to S. aureus of lidocaine was observed in a concentration of 0.25% which was not antibacterial to S. aureus organisms at 37 C. Similar results were obtained when P. aeruginosa organisms were examined in place of S. aureus, although P. aeruginosa was found to be less susceptible to lidocaine than S. aureus. The clinical significance of the thermal effect on the antibacterial activity of lidocaine was discussed in brief.
Differentiation between Genera Rhodococcus and Nocardia by Susceptibility Testing to Kanamycin and Some Other Antituberculosis Agents
Authors: Tsukamura Michio
Published: November 2013

The test for susceptibility to kanamycin is useful for differentiating between the genera Rhodococcus and Nocardia. All rhodococci except R. equi, R. erythropolis, and R. aurantiacus are susceptible to kanamycin, whereas all nocardiae except N. otitidis‐caviarum are resistant to kanamycin. Tests for susceptibility to rifampicin, streptomycin, and minocycline also are useful for differentiating among the species of each genus.
Cell‐Surface Properties of Enterotoxigenic and Cytotoxic Salmonella enteritidis and Salmonella typhimurium: Studies on Hemagglutination, Cell‐Surface Hydrophobicity, Attachment to Human Intestinal Cells and Fibronectin‐Binding
Authors: Baloda Suraj B. , Faris Ahmed , Krovacek Karel
Published: November 2013

Thirteen Salmonella enteritidis and S. typhimurium strains with smooth or rough colony morphology were investigated for their surface properties based on hemagglutination (HA), hydrophobicity, and fibronectin‐binding profiles. The strains showed 5 different patterns of HA which was mannose‐sensitive. The rough strains possessed comparatively greater number of fimbriae than the corresponding smooth strains and also attached to human intestinal cells in greater numbers. The Salmonella strains used in this study interacted with fibronectin and its 29‐kDa N‐terminal fragment to varied extents. These properties may be helpful in broadening the prospective interaction capabilities of Salmonella organisms with the host surfaces.
Structural Characterization of Recombinant Human Interferon‐Gammas Derived from Two Different Mammalian Cells
Authors: Sano Emiko , Kamata Koichi , Okano Kiyoshi , Yamazaki Syojiro , Iizuka Masahiko , Kobayashi Sigeyasu
Published: November 2013

Recombinant human interferon‐gammas (rHuIFN‐γs) were obtained from two different mammalian cells (mouse C127 cells and Chinese hamster ovary, CHO, cells) cultured in a microcarrier culture system. Both rHuIFN‐γs were purified using sequential chromatographies for their comparison of structural properties. The peptide maps of HuIFN‐γs digested with V8 protease and Western blot analysis demonstrated that C127 cells yielded mainly about 25kDa component and CHO cells produced about 25kDa and about 20kDa components. By the identification of glycosylated peptides, it was suggested that 20kDa and 25kDa components are glycosylated at one and at two sites, respectively. C‐terminal amino acid sequence analysis indicated that both rHuIFN‐γs consisted of at least six different species lacking 2 to 16 amino acid residues from C‐terminus, so that C‐termini of both rHuIFN‐γs were slightly different from each other.

Amino acid sequence and composition analyses of N‐terminal peptides demonstrated that N‐termini of both rHuIFN‐γs were blocked and were supposed to be identical with that of natural HuIFN‐γ. These results suggested that different molecular heterogeneities of rHuIFN‐γs resulted from the difference of post‐translational modifications of host cells.
Possible Regulation of Autologous Mixed Lymphocyte Reaction by OKM1+ NK Cells
Authors: Goto Makoto , Zvaifler Nathan J.
Published: November 2013

T cells are stimulated by autologous non‐T cells and interleukin 2 (IL‐2) is produced in the conventional autologous mixed lymphocyte reaction (AMLR) in young healthy controls. The role of cells with natural killer (NK) cell markers (OKM1+ cells or Leu 7+ cells) in the AMLR was studied. There were significant inverse correlations between the percentage of input OKM1+ cells minus monocyte (OKM1+ NK cells) and either AMLR proliferation (γ = −0.9, P<0.001) or IL‐2 production (γ = −0.75, P<0.01) in the AMLR cultures after 7 days measured at 7 days. A statistically significant correlation was observed between the percentage of input Leu 7+ cells and AMLR proliferation (γ = −0.64, P<0.05), but not IL‐2 production. These results suggest that the AMLR is controlled by OKM1+ NK, perhaps acting through IL‐2 regulation.
A Case of Lung Infection Caused by an Unusual Strain of Nocardia farcinica
Authors: Tsukamura Michio , Shimoide Hisao , Kaneda Kenji , Sakai Ryoichi , Seino Akio
Published: November 2013

A case of lung infection caused by an unusual strain of Nocardia farcinica is reported. This is the third case of the N. farcinica infection in this country. The strain failed to utilize rhamnose as sole carbon source, but could be identified by a numerical identification method. The mycolic acids contained 1–3 double bonds and the numbers of the carbon atoms of the mycolic acids were 50 to 60, average 56.
Modification of the Genital Herpes Infection in a Guinea Pig Model, by Prior Immunization with Bovine Herpes Mammillitis Virus
Authors: Maragos Constantina , May John T.
Published: November 2013

Bovine herpes mammillitis virus has been shown to partially protect guinea pigs against primary genital herpes simplex virus type 2 infections, further confirming the immunologic cross‐reactivity of these two viruses.
Production and Partial Purification of a Fluid‐Accumulating Factor of Non‐O1 Vibrio cholerae
Authors: Gyobu Yotaku , Kodama Hirohide , Uetake Hisao
Published: November 2013

A fluid‐accumulating factor (FAF in the ligated rabbit ileal loop test) from a strain of non‐O1 Vibrio cholerae not producing cholera toxin‐like enterotoxin (CTLT) was partially purified by ammonium sulfate precipitation, gel filtration with Sephadex G‐100, and DEAE cellulose column chromatography. The preparation thus obtained showed collagenolytic, cytolytic, necrotic, and hemorrhagic activities, but was not lethal to mice nor hemolytic to sheep erythrocytes. Desquamation of epithelial cells, inflammatory edema, and hemorrhage were observed in sections of rabbit intestine after inoculation of partially purified FAF (PPFAF). Biological and enzymatic activities of FAF were completely neutralized with anti‐PPFAF rabbit serum. More than 70% of non‐O1 V. cholerae strains from human diarrheal feces produced FAF in the shake culture of heart infusion broth (Difco). A fluid‐accumulating factor immunologically similar to FAF of non‐O1 V. cholerae was also produced by V. mimicus strains isolated from human diarrheal feces. These results indicate that the FAF produced by CTLT‐negative non‐O1 V. cholerae strains is an entity closely related to a cytolytic and hemorrhagic substance or the like, and that this FAF may play a role in the enteropathogenicity of CTLT‐negative strains.
Distribution of Clostridium botulinum in Japan and in Shinkiang District of China
Authors: Yamakawa Kiyotaka , Kamiya Shigeru , Nishida Shoki , Yoshimura Kiyoto , Yu He , Lu Deyuan , Nakamura Shinichi
Published: November 2013

Soil specimens obtained from several areas of Japan, which are closely located to or facing the Continental land of China, were examined for the distribution of Clostridium botulinum, especially pertaining to types A and B. A total of 266 specimens of Japan, when cultured, showed no type A or B toxicity, although 30 (11.3%), 4 (1.5%), and 10 (3.8%) of the specimens showed C1, C2, and type E toxicities, respectively. On the contrary, types A and/or B toxicities were shown, by the same method, in 14 of 20 specimens of Shinkiang district, China. The highest number of C. botulinum cells found in one gram of soil specimen was 25 for type A and 10 for type B.
Interferon Production under the Control of Heterologous Inducible Enhancers and Promoters
Authors: Asano Masahide , Nagashima Hitoshi , Iwakura Yoichiro , Kawade Yoshimi
Published: November 2013

In a search for a useful enhancer to control expression of interferon (IFN) gene in mammalian cells, mouse IFN‐β cDNA ligated 3′ downstream to the enhancer‐promoter of either mouse metallothionein‐I (MT‐I) or Drosophila heat shock protein (HSP) was introduced into various cultured cells by calcium‐phosphate precipitation method, and the level of IFN transiently produced was compared. In the case of the MT‐I enhancer‐promoter, low levels of IFN were produced without induction (0–21 IU/ml) and the level increased 5–50 times by heavy metals. In contrast, the basal level of expression of the HSP enhancer‐promoter was very low and its expression was increased several hundred to thousand times by heat shock or arsenite. Thus the HSP enhancer‐promoter appears to be a potent inducible element with very low basal level and high inducibility.
Characterization of L1210 S Cells with Low Sensitivity to Mouse Interferon‐γ
Authors: Iwata Akira , Watanabe Yoshihiko , Sakata Tsuneaki , Sokawa Yoshihiro , Kawade Yoshimi
Published: November 2013

A mouse leukemic cell line L1210 Sg with a low sensitivity to interferon‐γ (IFN‐γ) was described. On the nature of the antiviral action and binding of IFN, L1210 Sg cells were compared with L1210m cell line which is sensitive to IFN‐γ. For a half reduction of the vesicular stomatitis virus‐RNA synthesis, L1210 Sg cells required 500–fold more IFN‐γ than L1210m cells did. However, both cell lines were induced to the antiviral state to the same extent with IFN‐α or ‐β. L1210 Sg and L1210m cells were sensitive to the anti‐proliferative action of IFN‐α and ‐β, but insensitive to IFN‐γ. (2′‐5′)Oligoadenylate synthetase was induced in these cell lines by IFN‐β, but not by IFN‐γ, which suggests that the induction of this synthetase may not be responsible for the antiviral action of IFN‐γ. No substantial difference between L1210 Sg and L1210m cells was found in IFN receptors for IFN‐γ and IFN‐β either in number per cell or in their affinity to corresponding IFN type. However, differences were noted in time course profiles of cell‐associated IFN‐γ at 37 C: in L1210m cells, a rise‐and‐decay profile of IFN‐γ bound to cells was observed at 37 C, but in L1210 Sg cells, rise and slight decay was observed. On the other hand, a similar rise‐and‐decay curve of IFN‐β bound to these cells was observed. These results indicated that the low sensitivity of L1210 Sg cells to IFN‐γ may be due to this slight decay of receptor‐bound IFN‐γ.
The Analysis of Immature Lymphoid Precursors Stored in Longterm Bone Marrow Culture
Authors: Miyazoe Ichiro , Taniguchi Masaru , Takemori Toshitada
Published: November 2013

The long‐term bone marrow culture system developed by Dexter (MBMC) is known to store immature lymphoid precursors capable of differentiating into mature B cells in irradiated or immunodeficient mice. It has been suggested that pre‐B cells are not generated under such culture conditions, but that opinion was not based on any systematic analyses. In the present study under carefully controlled conditions, we observed that pre‐B and pro‐B cells were eliminated from the late stage of primary MBMC, and the former were not generated in recharged MBMC. Under appropriate conditions, these immature precursors in recharged MBMC generated in vitro immunoglobulin‐positive (Ig+) cells to differentiate into antibody‐forming cells upon stimulation with lipopolysaccharide (LPS). LPS‐reactive B cells were observed in every 10th of the Ig+ cells, the frequency being essentially the same as that observed in normal B cells in different tissues. The immature B cell precursors generating LPS reactive cells were expressed in recharged MBMC at the frequency of 4.2 × 10−6. A staining experiment showed that cells bearing AA4.1 were stored at the frequency of 10−4‐10−5. This frequency is thought to be similar to that of lymphoid precursors in recharged MBMC committed to differentiate along B lineage cells. Based on these results, we discussed the stage, nature, and mode of differentiation of immature lymphoid precursors stored in MBMC.
Linkage of K99 Production and STa Activity in a Plasmid of an Escherichia coli Porcine Isolate
Authors: Shimizu Mikio , Sakano Tetsuya , Satoh Shizuo
Published: November 2013

An Escherichia coli strain ZP118 of porcine origin was shown to harbor a 68‐megadalton (Md) plasmid coding for a colonization factor K99 and heat‐stable enterotoxin (STa), and a self‐transmissible 51‐Md plasmid coding for drug resistance. One of the transconjugants obtained by mating between ZP118 and E. coli C was found to harbor a 90‐Md plasmid coding for K99, STa and drug resistance. Restriction endonuclease analysis suggested that the 90‐Md plasmid could be a recombinant between the 68‐Md plasmid and the 51‐Md plasmid.
Enhancement of Host Resistance against Listeria Infection by Lactobacillus casei: Activation of Liver Macrophages and Peritoneal Macrophages by Lactobacillus casei
Authors: Sato Katsumasa , Saito Hajime , Tomioka Haruaki
Published: November 2013

The functions of liver macrophages and peritoneal macrophages obtained after injection of Lactobacillus casei were examined. Listericidal activity in vivo was enhanced in liver macrophages 13 days after L. casei injection but was somewhat suppressed in the macrophages 2 days after the injection. The listericidal activity in vitro was enhanced in peritoneal macrophages obtained 13 days after L. casei injection but was suppressed in cells obtained 2 days later. The PMA‐triggered respiratory burst in the liver macrophages elicited by L. casei was higher than that of resident macrophages. Alkaline phosphodiesterase activity in the liver macrophages was decreased by L. casei injection, as was also the case with peritoneal macrophages. These observations indicate that L. casei augmented cellular functions of both liver and peritoneal macrophages.
Effects of the Antiserum against a Fraction Enriched in Scrapie‐Associated Fibrils on the Scrapie Incubation Period in Mice
Authors: Ito Masako , Shinagawa Morikazu , Doi Satoshi , Sasaki Satoshi , Isomura Hiroshi , Takahashi Kaname , Goto Hitoshi , Sato Gihei
Published: November 2013

An antiserum against a fraction enriched for scrapie‐associated fibrils (SAF), was examined for its effects on scrapie incubation period by inoculating mice either intraperitoneally or intracerebrally with various dilutions of the serum mixed with scrapie‐infected mouse brain homogenate. After intraperitoneal inoculation the mean time of the incubation period increased with increasing concentrations of the antiserum in a statistically significant fashion, when the serum dilutions were made with phosphate‐buffered saline. After intracerebral inoculation, however, there were no statistically significant differences between the control group and any of the antiserum‐groups. When the antiserum dilutions were made with pre‐immune serum, the mice inoculated intraperitoneally also showed no significant differences between the two groups. These results indicate that the specific antibodies to SAF have no effect on the scrapie infectivity.
Affinity Constants of Anti‐Leptospira Monoclonal Antibodies, Numbers of Antigenic Determinants on Leptospiras, and Their Influence on the Microscopic Agglutination Test and ELISA
Authors: Takimoto Toru , Ono Etsuro , Ito Toshihiro , Yanagawa Ryo
Published: November 2013

The affinity constants of anti‐canicola and anti‐hebdomadis monoclonal antibodies and the numbers of antigenic determinants per organism of each serovar of leptospira were determined, and their influence on the results of microscopic agglutination test (MAT) and ELISA were examined. In the combinations of a monoclonal antibody and some serovar which showed higher affinity constants at levels of 107 to 108/m order, both MAT and ELISA were positive except for one combination, while in the combinations which gave lower affinity constants at levels of 106/m order, MAT was positive but ELISA was negative. The number of antigenic determinants seemed to have no significant influence on the results. The disagreement due to the difference in the threshold affinities of MAT and ELISA should be considered in the use of monoclonal antibodies in the taxonomy of leptospira.
Development of a Highly Sensitive Bead‐ELISA to Detect Bacterial Protein Toxins
Authors: Oku Yuichi , Uesaka Yoshihiko , Hirayama Toshiya , Takeda Yoshifumi
Published: November 2013

A highly sensitive sandwich enzyme‐linked immunosorbent assay to detect bacterial toxins was developed. Fab of anti‐toxin IgG was conjugated with horseradish peroxidase by the maleimide method and tetramethylbenzidine was used as substrate. As the solid phase, a 6.5 mm diameter polystyrene bead was used and this was coated with the anti‐toxin IgG. The entire assay could be completed within 3.5 hr. The sensitivity of this bead‐ELISA was found to be quite high with various bacterial toxins: less than 20 pg/ml for thermostable direct hemolysin of Vibrio parahaemolyticus, less than 60 pg/ml for Shiga toxin, less than 20 pg/ml for VT2 (Shiga‐like toxin II) of Escherichia coli, less than 200 pg/ml for heat‐labile enterotoxin of E. coli, and less than 6 pg/ml for cholera enterotoxin.
Genomic Comparison of Adenovirus Type 3 Isolates from Patients with Acute Conjunctivitis in Japan, Australia, and the Philippines
Authors: Guo Deng Fu , Shibata Riri , Shinagawa Morikazu , Sato Gihe , Aoki Koki , Sawada Harumi
Published: November 2013

A total of 53 Ad3 isolates obtained from patients with acute conjunctivitis in Japan, Australia, and the Philippines during 1973 to 1986 were analyzed for their genome types with 4 restriction endonucleases, BamHI, BglII, HindIII, and SmaI. Two new genome types designated tentatively as Ad3f and Ad3g were identified by combination of BamHI and BglII in the isolates. The changes of restriction sites and sizes of restriction fragments in newly recognized Ad3f and Ad3g were located at the similar regions reported in other Ad3 genome types by O'Donnell et al (1986) on physical maps of the Ad3 prototype strain GB genome. In Japan, 46 Ad3 isolates obtained from 1983 to 1986 were either Ad3f or Ad3g. Yearly alternation of predominance of both genome types were observed in the northern part of Japan during the period. In Australia, two genome types Ad3p and Ad3f were found in 6 isolates, and the former was observed in the 3 isolates obtained before 1981 and the remaining 3 isolates were obtained after 1983. In the Philippines, the only isolate obtained in 1984 was Ad3p.