bluuebunny/crossref_metadata_2025 huggingface dataset:
Detection of Specific Antibodies against Fimbriae and Membrane Proteins from the Oral Anaerobe Bacteroides gingivalis in Patients with Periodontal Diseases
Authors: Yoshimura Fuminobu , Sugano Toshikazu , Kawanami Masamitsu , Kato Hiroshi , Suzuki Takeshi
Published: November 2013

Sera from a number of patients with periodontal diseases were shown to have specific immunoglobulin G (IgG) antibodies against fimbriae and membrane proteins of Bacteroides gingivalis, a suspected pathogen, by using Western blotting analysis. The sera had a strong tendency to react with fimbriae, or exactly oligomeric structures of fimbriae with a native β‐structure rich‐conformation. However, the sera did not react with fimbrilin, a constituent protein of fimbriae, which is denatured by sodium dodecyl sulfate.
Adhesion of Enterotoxigenic (ETEC) and Bovine Mastitis Escherichia coli Strains to Rat Embryonic Fibroblasts: Role of Amino‐Terminal Domain of Fibronectin in Bacterial Adhesion
Authors: Faris Ahmed , Fröman Gunnar , Switalski Lech , Höök Magnus
Published: November 2013

Adherence of human enterotoxigenic and bovine mastitis Escherichia coli to rat embryonic fibroblasts was studied. Adhesion of E. coli strains B34289c (human) and 1407 (bovine) was rapid and reached maximum after 30–40 min. Strain 1410 (bovine), which binds fibronectin but not its 29K amino‐terminal fragment, did not adhere to the fibroblasts. Strain B34289c grown at 25 C or below and at 40 C or above lost its binding and adhesive properties simultaneously. Maximum binding and adhesion for this strain was achieved when it was grown at 33 C. Strains grown at this temperature adsorbed to fibronectin‐, 29K fragment‐, and Octyl Sepharose, with the exception of bovine strain 1410, which did not adsorb to 29K‐Sepharose as expected. None of the strains adsorbed to cross‐linked Sepharose 4B. 29K‐IgG and Fab fragments thereof specifically blocked both binding (max 55%) and adhesion (>95%). Sonicated and trypsin‐treated bacteria were no longer able to bind or adhere. The supernatant of sonicated bacteria inhibited both binding and adhesion. Penicillin G at 0.5 μg/ml (1/5 minimal inhibitory concentration: MIC) and tetracycline at 0.2 μg/ml (1/5 MIC), when included in the growth medium, suppressed the cell surface components responsible for fibronectin binding and fibroblast adhesion. The presence of fibronectin was demonstrated in the fibroblast extracellular matrix by immunofluorescens with 29K‐IgG antibodies.
Effects of Panose on Glucan Synthesis and Cellular Adherence by Streptococcus mutans
Authors: Koga Toshihiko , Horikoshi Toshio , Fujiwara Taku , Hamada Shigeyuki
Published: November 2013

The effects of panose on glucan synthesis and sucrose‐dependent cellular adherence by Streptococcus mutans were investigated. Panose effectively inhibited glucan synthesis from sucrose by glucosyltransferases from S. mutans strain 6715, but increasing amounts of panose increased the release of fructose from sucrose by the enzymes. On the other hand, production of a series of oligosaccharides of increasing size by the enzymes was markedly enhanced in the presence of panose. These results indicate that panose activates the enzymes and that the inhibition of glucan synthesis by panose is due to the transfer of the glucosyl group of sucrose to panose. Sucrose‐dependent adherence of cells of various S. mutans strains to a glass surface was also inhibited by panose.
Pathogenesis of Lupus Dermatoses in Autoimmune Mice
Authors: Furukawa Fukumi , Taniguchi Shinkichi , Tachibana Takao , Horiguchi Yuji , Kanauchi Hideo , Ohshio Gakuji , Hamashima Yoshihiro , Imamura Sadao
Published: November 2013

We measured histamine concentration and its metabolizing enzymes in the skin of MRL/Mp‐lpr/lpr (MRL/l) and BXSB mice to clarify the contribution of histamine metabolism to the mechanisms of the development of lupus dermatoses. The concentration of histamine seemed to differ with the mouse strain. The activity of histamine‐N‐methyltransferase (HMT), one of two major metabolizing enzymes, was significantly lower in the tail and back skin of MRL/l mice at the age of 5 months than in the control MRL/Mp‐+/+ (MRL/n) mice, although there were no characteristic differences among several mouse strains of 1 mo of age. In the back skin of MRL/l mice, an age‐dependent decrease of HMT activity was observed along with a corresponding decrease in histamine concentration, whereas an age‐dependent increase of both HMT activity and histamine concentration was demonstrated in BXSB mice and other control mouse strains. Autoimmune‐prone male BXSB mice and non‐autoimmune female BXSB mice at 5 mo of age showed similar HMT activity. Corticosteroid treatment restored HMT activity in the skin of MRL/l mice but not in MRL/n mice. In addition, the change in HMT activity in MRL/l mice treated with corticosteroid appeared earlier than changes in clinicopathological examinations including skin eruptions, dermatopathology and proteinuria. Diamine oxidase (DAO) activity, another major metabolizing enzyme, was not detected in the skin of any autoimmune or control mouse strains. These findings suggest that the low activity of HMT in the skin of MRL/l mice plays a significant pathological role in the development of spontaneous lupus‐like eruption. In other mouse strains, it is assumed that HMT activity is regulated by genetic factors.
Nature of Monocyclic β‐Lactam Antibiotic Nocardicin A to β‐Lactamases
Authors: Kojo Hitoshi , Mine Yasuhiro , Nishida Minoru , Goto Sachiko , Kuwahara Shogo
Published: November 2013

Nocardicin A is the antibiotic which was first found to possess a monocyclic β‐lactam ring. This antibiotic was inactivated by the cleavage of its β‐lactam ring. The direct spectrophotometric assay was applied to measure the rate of enzymatic hydrolysis of Nocardicin A. Nocardicin A was highly stable to both chromosomal and plasmid‐mediated β‐lactamases. Of the nine β‐lactam antibiotics including cefoxitin and cefuroxime, Nocardicin A was the most stable to the β‐lactamases tested excluding those from Klebsiella oxytoca and Proteus vulgaris. The latter broad‐spectrum β‐lactamases hydrolyzed Nocardicin A rather intensively. Extreme stability of Nocardicin A to β‐lactamases was suggested to be due to the combination of its low affinity to the enzymes and stabilization of its monocyclic β‐lactam ring. Nocardicin A was shown to have inducing ability toward β‐lactamases.
Simplified Method for Preparation of Concentrated Exoproteins Produced by Staphylococcus aureus Grown on Surface of Cellophane Bag Containing Liquid Medium
Authors: Ikigai Hajime , Seki Keiko , Nishihara Shoko , Masuda Shogo
Published: November 2013

A simplified method for preparation of concentrated exoproteins including protein A and α‐toxin produced by Staphylococcus aureus was successfully devised. The concentrated proteins were obtained by cultivating S. aureus organisms on the surface of a liquid medium‐containing cellophane bag enclosed in a sterilized glass flask. With the same amount of medium, the total amount of proteins obtained by the method presented here was identical with that obtained by conventional liquid culture. The concentration of proteins obtained by the method, however, was high enough to observe their distinct bands stained on polyacrylamide gel electrophoresis. This method was considered quite useful not only for large‐scale cultivation for the purification of staphylococcal proteins but also for small‐scale study using the proteins. The precise description of the method was presented and its possible usefulness was discussed.
A Seroepidemiologic Study of Hepatitis B Virus Infection among Barbers in Huangshi City, Hubei, China
Authors: Shaoliang She , Luyuan Shi , Yuanjian Wu , Zhanzhao Li , Cuanzheng Zheng , Yingping Wu , Xinghai Yu
Published: November 2013

Between March and August 1986 in Huangshi City, serum samples were collected from 316 apparently healthy barbers as a study group, as well as from 361 healthy employees of department stores as a control group. They were tested for hepatitis B surface antigen (HBsAg), antibody to HBsAg (anti‐HBs) and antibody to hepatitis B core antigen (anti‐HBc) by enzyme‐linked immunoadsorbent assays. Barbers showed a prevalence higher than that in controls for HBsAg (16.8 vs. 9.2%, P<0.01), anti‐HBs (67.1 vs. 45.9%, P<0.001), and anti‐HBc (39.2 vs. 21.2%, P<0.001). The prevalence of at least one marker of hepatitis B virus (HBV) infection was significantly higher in barbers than in controls (86.1 vs. 61.7%, P<0.001). Although the socioeconomic status and education level did not correlate with the frequency of HBV markers, the prevalence of HBsAg increased in parallel with the duration of practice. Because of their high risk for HBV infection, barbers need to be screened for markers of HBV infection on a routine basis, and are prime candidates for immunoprophylaxis with hepatitis B vaccine.
Physical and Functional Map of an Amikacin‐Resistance Plasmid Isolated from a Multiresistant Strain of Serratia marcescens
Authors: Kanno Rieko
Published: November 2013

pTK159, a multiresistance 40‐kilobase (kb) plasmid, was isolated from a clinical strain of Serratia marcescens. pTK159 was nonconjugative and carried determinants for resistance to amikacin, streptomycin/spectinomycin, sulfamethoxazole and ampicillin. A physical and functional map of pTK159 was constructed. By cloning various fragments of pTK159 in pACYC184 or pBR322, genes for resistance to amikacin, streptomycin/spectinomycin, and sulfamethoxazole were found to be located on a 2.0‐kb BamHI‐HindIII fragment, a 1.4‐kb HindIII fragment and a 2.1‐kb HindIII fragment, respectively. The map of pTK159 was compared with published maps of amikacin‐resistance determinants and transposons.
Regulation of in vivo Immunological Reactions by Monoclonal Antibody against Lymphocyte Activation Antigen
Authors: Yuasa Hiroo , Uede Toshimitsu , Wada Takuro , Yamaguchi Akira , Murakami Toshiya , Osawa Hisao , Diamantstein Tibor , Kikuchi Kokichi
Published: November 2013

In vivo effects of a monoclonal antibody that recognizes rat lymphocyte activation antigen were studied. Spleen cells obtained from sheep red blood cell (SRBC)‐immunized rats developed strong PFC response against SRBC. However, the 5C6‐F4 treatment resulted in the inhibition of subsequent development of PFC response. The suppression of PFC response was due to the inhibition of generation of helper T cells, but not due to the preferential induction of suppressor cells. In addition, 5C6‐F4 antibody was also found to inhibit the clinical expression of collagen‐induced rat arthritis and the synovial inflammation in collagen‐induced arthritis rats. Furthermore, the in vivo generation of cytotoxic cells against syngeneic tumor cells was also inhibited by 5C6‐F4 antibody. The in vivo administration of 5C6‐F4 antibody did not cause any pathological changes in brain, lung, liver, kidney, spleen, thymus, and lymph nodes.
Cytopathic Effect of Heat‐Labile Toxin of Bordetella parapertussis on Aortic Smooth Muscle Cells from Pigs or Guinea Pigs
Authors: Endoh Masahiko , Nagai Masaaki , Ueda Tetsuro , Yoshida Yoji , Nakase Yasukiyo
Published: November 2013

The in vitro effect of the heat‐labile toxin (HLT) of Bordetella parapertussis on HeLa, baby hamster kidney (BHK), chinese hamster ovary (CHO), myeloma (P3‐NS‐1), human embryonic lung (HEL‐R66) cells, erythrocytes, adipocytes and lymphocytes from guinea pigs, mice or rats, or aortic smooth muscle cells from pigs or guinea pigs was examined. Within 8, 6, 4, 2, and 2 hr after the exposure to 1, 3, 10, 30, and 100 MNDs/ml of HLT, respectively, the cultured smooth muscle cells only showed a cytopathic change. When the cells exposed to HLT were washed out within 60 min post‐exposure, the change could be induced with an extend period of lag. Histamine, KCl or norepinephrine caused similar change in the cells, but the period of lag was within 30 min. The HLT activity was neutralized by an anti‐B. parapertussis‐ or B. bronchiseptica‐HLT guinea pig IgG. HLT had no effects on any other cells tested.
Electron Microscopic Observations on Tracheal Epithelia of Mice Infected with Bordetella bronchiseptica
Authors: Sekiya Kachiko , Futaesaku Yutaka , Nakase Yasukiyo
Published: November 2013

To clarify the pathogenesis of Bordetella in vivo infection, the tracheal epithelia of mice were examined in detail by electron microscopy at various intervals after intranasal inoculation with graded doses of phase I Bordetella bronchiseptica. In mice infected with a lethal dose (6 to 7 × 107 CFU), a remarkable rupture of the cell membranes of cilia and microvilli of the middle trachea was found on day I postinfection. The rupture of the membrane was observed over the entire tracheal epithelia, on day 2 after infection. The affected cilia were constricted at the transitional region and were broken off. In the ciliated cells the adherence of organisms to ciliary apexes and colonization in the interciliary spaces were also remarkable. In both the ciliated and nonciliated epithelial cells, the cytoplasmic vacuolation and pyknosis or karyorrehexis were also notable. In mice infected with one‐tenth of the lethal dose, similar findings were seen, but appeared more slowly and the bacteria were not seen attaching to ciliary apexes. In mice receiving one‐hundredth of the lethal dose, only mild cilial abnormality such as aggregation of cilia, and slight cytoplasmic vacuolation were found 6 days postinfection. Based on these findings, a possible mechanism of the ciliary damages produced by B. bronchiseptica was postulated.
Luminol‐Dependent Chemiluminescence in Antibody‐Sensitized Neutrophils Stimulated with Protein A‐Bearing Staphylococci
Authors: Nishihara Shoko , Seki Keiko , Ikigai Hajime , Masuda Shogo
Published: November 2013

When mouse polymorphonuclear leukocytes (PMNs) sensitized with rabbit antibody to mouse Ehrlich ascites tumor cells were stimulated by Staphylococcus aureus Cowan I cells, a conspicuous luminol‐dependent chemiluminescence was observed in the absence of opsonin. The profile of the chemiluminescence (CL) response evoked by staphylococcal cells from antibody‐sensitized PMNs had two peaks. An initial peak, observed within 1 min after stimulation, was sharp and high and a second peak, observed about 5 min after stimulation, was low and extended. The CL response of antibody‐sensitized PMNs stimulated by S. aureus Cowan I cells was dose‐dependently blocked by preincubation with soluble SpA. Cells of a mutant derived from S. aureus Cowan I strain with trace amounts of cell‐bound SpA failed to stimulate the antibody‐sensitized PMNs to generate the CL response. The antibody‐sensitized PMNs were found to phagocytize SpA‐bearing S. aureus cells even in the absence of opsonic serum. These results suggest that the observation presented here might provide a useful tool for the investigation of CL response of PMNs.
Priming Activity for Chemiluminescence Reaction of PMN in the Culture Supernatant of Streptococcal Preparation (OK‐432)‐Stimulated Spleen Cells
Authors: Fukase Yoko , Fukase Shigeru , Sendo Fujiro
Published: November 2013

We investigated the effect of supernatant from human spleen cell culture stimulated with a streptococcal preparation, OK‐432 (OK sup), on the luminol‐dependent chemiluminescence (CL) of human polymorphonuclear leukocytes (PMN). The fMLP‐stimulated CL of PMN was markedly enhanced by the pretreatment with OK sup. This result indicates that OK sup contained the factor(s) that primes fMLP‐stimulated CL of PMN. The priming factor(s) in OK sup was partially inactivated by the treatment at 56 C for 30 min and pH 2 or pH 10 treatments. Since the enhancing effect of OK sup on the CL was inhibited by the treatment of sodium azide and the addition of catalase or taurine, it was assumed that OK sup augments the activity of MPO‐H2O2‐HOCl system of fMLP‐stimulated PMN.
Structural Heterogeneity Regarding Local Shwartzman Activity of Lipid A
Authors: Mashimo Jun‐ichi , Tanaka Chiharu , Arata Satoru , Akiyama Yuri , Hata Seiichi , Hirayama Tohru , Egawa Kiyoshi , Kasai Nobuhiko
Published: November 2013

The relation of chemical structure to local Shwartzman activity of lipid A preparations purified by thin‐layer chromatography from five bacterial strains was examined. Two lipid A fractions from E. coli F515—Ec‐A2 and Ec‐A3—exhibited strong activity, similar to that of previous synthetic E. coli‐type lipid A (compound 506 or LA‐15‐PP). The Ec‐A3 fraction contained a component that appeared to be structurally identical to compound 506, and the main component of Ec‐A2 fraction was structurally similar to compound 506 except that it carried a 3‐hydroxytetradecanoyl group at the C‐3′ position of the backbone in place of a 3‐tetradecanoyl‐oxytetradecanoyl group. Free lipid A (12 C) and purified lipid A fractions, Ec‐A2 (12 C) and Ec‐A3 (12 C), respectively, obtained from bacteria grown at 12 C, exhibited activity comparable to Ec‐A2 or Ec‐A3. In these preparations, a large part of the 3‐dodecanoyloxytetradecanoyl group might be replaced by 3‐hexadecenoyloxytetradecanoyl group. Salmonella minnesota R595 free lipid A also contained at least two active lipid A components as seen in E. coli lipid A, but the third component corresponding to the synthetic Salmonella‐type lipid A (compound 516 or LA‐16‐PP) exhibited low activity. A lipid A fraction, Cv‐A4 from Chromobacterium violaceum IFO 12614, which was proposed to have two acyloxyacyl groups at the C‐2 and C‐2′ positions with other acyl groups, exhibited weaker activity than the free lipid A or LPS. The purified lipid A fractions from Pseudomonas diminuta JCM 2788 and Pseudomonas vesicularis JCM 1477 contained an unusual backbone with 2,3‐diamino‐2,3‐dideoxy‐d‐glucose disaccharide phosphomonoester, and these lipid A (Pd‐A3 and Pv‐A3) exhibited strong activity comparable to the E. coli lipid A. Thus, the present results show that the local Shwartzman reaction can be expressed by partly different lipid A structures in both hydrophilic backbone and fatty acyl residues; when they have the same backbone the potency varies markedly depending on the structure of the acyl residues.
The Growth of Compact and Diffuse Variants of Staphylococcus aureus in Bovine Mastitic and Normal Whey
Authors: Mattila Tiina , O'Boyle Denise , Frost Alan J.
Published: November 2013

Strains of Staphylococcus aureus producing either diffuse or compact colonies in serum‐soft agar were grown in bovine normal and mastitic whey. Bacterial growth was followed by automated turbidometry. Compact strains multiplied faster and to higher final numbers in mastitic whey than diffuse strains, whereas diffuse strains grew to higher numbers in normal whey. Nutrients (hemolysed bovine blood, bovine serum, proteose‐peptone) were added to normal whey to enhance bacterial growth as in mastitic whey. The growth‐promoting effect of these nutrients was dose‐dependent for compact strains but not for diffuse strains. The difference in the growth characteristics of diffuse and compact strains in bovine whey explains some aspects of the pathogenesis of bovine mastitis.
Validity of an Enzyme‐Linked Immunosorbent Assay with Serotype‐Specific Monoclonal Antibodies for Serotyping Human Rotavirus in Stool Specimens
Authors: Urasawa Shozo , Urasawa Tomoko , Taniguchi Koki , Morita Yasuyuki , Sakurada Norio , Saeki Yoshito , Morita Osayuki , Hasegawa Sumiyo
Published: November 2013

We recently developed a method for serotyping human rotavirus (HRV) by an enzyme‐linked immunosorbent assay with HRV serotype‐specific neutralizing monoclonal antibodies (ELISA serotyping). In the present study this method was compared with the fluorescent focus neutralization test with serotype‐specific rabbit antisera (NT serotyping) in the sensitivity and specificity of the test. Direct serotyping of HRVs which were contained in stool specimens indicated that while only 37% of the samples were successfully serotyped in NT, 78% of the samples could be serotyped in ELISA. Regarding the samples whose serotype could be determined in the two tests, the assigned serotypes were identical in both tests. The results obtained indicated the utility of ELISA serotyping in clinical and epidemiologic studies of HRV infection.
Evidence Suggesting That the Life Span of Mycobacterium avium Complex is Longer than That of Other Mycobacteria
Authors: Tsukamura Michio
Published: November 2013

Mycobacterium avium complex strains often contain considerably more numbers of viable bacterial units per mg wet weight than other mycobacteria, especially other slowly growing ones. This finding suggests that the life span of M. avium complex strains is often longer than the life span of other mycobacteria. The other mycobacteria, especially slowly growing ones seem to die more rapidly after their multiplication.
Opsonic Effect of Fibronectin on Staphylococcal Phagocytosis by Human Polymorphonuclear Leukocytes: Its Relative Inefficiency in Post‐Phagocytic Metabolic Activities and in Intracellular Killing
Authors: Yonemasu Kunio , Sasaki Takako , Hashimoto Hiratsugu , Kashiba Shuzo
Published: November 2013

The binding of 125I‐labeled human plasma fibronectin (FN) to two strains of live Staphylococcus aureus (S. aureus) (a coagulase‐positive Cowan I and a coagulase‐negative Newman D2C) and the opsonic effect of FN on phagocytosis of these bacteria by human polymorphonuclear leukocytes (PMN) have been studied. 125I‐FN bound to a similar extent in both staphylococcal strains. The 125I‐FN‐binding was significantly inhibited by human fibrinogen as well as unlabeled FN. The FN‐binding was also reduced markedly by trypsinization of these bacteria, but the extent of its decrease did not correlate with their tryptic susceptibility of protein A and clumping factor. FN enhanced the uptake of these bacteria by PMN. However, its binding had no effect on superoxide anion (O2−) generation. The FN‐binding definitely stimulated staphylococcal ingestion and intracellular killing by PMN, but the extent of such promotion was dissimilar between these two strains of bacteria. These results suggest that post‐phagocytic metabolic activities as well as intracellular killing of these Staphylococci may also be greatly influenced by FN‐unrelated factors as are other bacteria having no FN‐receptors.
Comparative Studies of Strains Ictero No. I and RGA as the Type Strain of Leptospira interrogans: Agglutinin Absorption Test, Protein and Antigen Profiles, and Enzyme Activities
Authors: Hata Katsura , Ono Etsuro , Yanagawa Ryo
Published: November 2013

Strain Ictero No. I, the first isolate of Leptospira, isolated by Inada and Ido in 1914, was found to be sufficiently qualified to be the type strain of Leptospira interrogans rather than strain RGA. In an agglutinin absorption test, anti‐Ictero No. I serum was not absorbed completely with strain RGA, and 25% of the homologous titer remained unabsorbed, while anti‐RGA serum was completely absorbed with strain Ictero No. I. Thus, strain Ictero No. I was not serologically identical with strain RGA, and the two strains were considered to be different serovars. A protein band with a molecular weight of approximately 33,000 daltons was detected in strain Ictero No. I but not in strain RGA by SDS‐PAGE. By Western blotting, this protein band was detectable with anti‐Ictero No. I serum but not with anti‐RGA serum. The presence of the 33K protein in strain Ictero No. I, but not in strain RGA, was confirmed by radioimmunoprecipitation using [125I]‐labeled antigens, indicating that the protein antigen was surface‐exposed. Only 8 of the 89 enzymes activities were different between strains Ictero No. I and RGA (line Sapporo). From the above results, we propose that strain Ictero No. I should be designated as the type strain of L. interrogans instead of strain RGA.
Prevalence of Human Immunodeficiency Virus (HIV) Infection among Hemophiliacs in Fukuoka, Japan
Authors: Kashiwagi Seizaburo , Hayashi Jun , Ikematsu Hideyuki , Noguchi Akinori , Ikeda Kiyoshi , Kishida Kunio , Shirakawa Mitsuru , Takenaka Akira , Mori Ryoichi
Published: November 2013

To determine when the hemophiliacs in Fukuoka prefecture, Japan, first became positive for antibodies, we tested human immunodeficiency virus (HIV) antibodies on serum samples obtained from 1976–1987 stored at −30 C. Fifteen out of 64 hemophilia A patients (23.4%), five out of 11 hemophilia B patients (45.5%), but none of 17 patients with von Willebrand's disease (0%) were positive for HIV antibodies. In this series, two with hemophilia A became positive for HIV antibodies for the first time in 1983, and in 1984 another four with hemophilia A and one with hemophilia B became positive.
Restriction Endonuclease DNA Analysis of Leptospira interrogans Serovars Icterohaemorrhagiae and Copenhageni
Authors: Tamai Tomonori , Sada Eiji , Kobayashi Yuzuru
Published: November 2013

Leptospira interrogans serovar icterohaemorrhagiae strains Ictero No. I and RGA and serovar copenhageni strains M20, Shiromizu and Shibaura were examined by restriction endonuclease DNA analysis. Fifteen endonucleases (AluI, BamHI, BglII, EcoRI, HaeIII, HhaI, HindIII, KpnI, PstI, SacI, SalI, SmaI, StyI, XbaI and XhoI) were used as the digesting enzymes.

Strain Ictero No. I showed endonuclease cleavage patterns which differed from those of the other four strains only when it was digested with enzymes KpnI and HindIII. When digested with KpnI, an extra band of about 5.4 kb was clearly produced, and when digested with HindIII, an extra band of about 25 kb was produced. When the other 13 enzymes were used, no differences were found between the endonuclease cleavage patterns among the five strains. Moreover, strains RGA, M20, Shiromizu and Shibaura could not be distinguished by the restriction endonuclease DNA analysis using all 15 endonucleases. In addition, six newly isolated leptospires from patients with leptospirosis and from Rattus norvegicus were compared with the Ictero No. I and M20 strains, by restriction endonuclease DNA analysis using enzymes KpnI and HindIII. Three leptospires belonging to serovar icterohaemorrhagiae showed the same endonuclease cleavage patterns as the M20 strain. The other three strains, which belong to serovar copenhageni, showed almost the same endonuclease cleavage patterns as the M20 strain; only the Kaijima 702 strain produced an extra band which was not identical to the Ictero No. I‐specific extra band when digested with HindIII. The leptospiral restriction endonuclease DNA analysis has revealed taxonomic structures that are unrecognized by serology alone.
Rapid Preparation of Samples for Compositional Sugar Analysis of the “Degraded Polysaccharide” Fraction of Lipopolysaccharides from Vibrio cholerae
Authors: Kondo Seiichi , Hisatsune Kazuhito
Published: November 2013

A simple and rapid method was devised for direct isolation and fractionation of the “degraded polysaccharide” (DPS) fraction of O‐antigenic (or endotoxic) lipopolysaccharides (LPS) directly from heat‐killed Vibrio cholerae (O1 and non‐O1) cells without separating the LPS. Neither phenol‐water extraction nor ultracentrifuge is needed in this method. V. cholerae NIH 41 was used as standard. The cells (3–5 g wet weight) were heated in 5% acetic acid at 100 C for 1.5 hr. The acetic acid extract obtained as the supernatant by centrifugation was evaporated to dryness in vacuo, and the resultant residue was dissolved in 10 ml of distilled water. The solution was mixed with 2 volumes of acetone, and the supernatant obtained by centrifugation was mixed with 5 volumes of acetone and centrifuged. Fraction Sed. II was recovered as the precipitate, while the supernatant was evaporated to dryness in vacuo, yielding fraction Sup. III. Sed. II had a sugar composition that was identical, at least qualitatively, to that of DPS isolated from LPS of the corresponding strain except for the absence of a fructose component in the case of V. cholerae NIH 41, while instead Sup. III from V. cholerae NIH 41 contained fructose.
Seroepidemiologic Study of Adult T‐Cell Leukemia Virus (ATLV) and Hepatitis B Virus Infection in Okinawa, Japan
Authors: Kashiwagi Seizaburo , Ikematsu Hideyuki , Hayashi Jun , Kajiyama Wataru , Nomura Hideyuki , Noguchi Akinori , Tani Shunichi , Goto Masashi
Published: November 2013

A total of 2,283 serum samples were collected from healthy subjects in three islands of the Yaeyama district of Okinawa, Japan. These sera were tested for the presence of hepatitis B surface antigen (HBsAg), for antibody to hepatitis B core antigen (anti‐HBc), and for antibody to adult T‐cell leukemia‐associated antigen (anti‐ATLA). Correlation between hepatitis B virus infection and adult T‐cell leukemia virus (ATLV) infection was determined by using the prevalence rates for three virus markers. Overall prevalence of HBsAg, anti‐HBc and anti‐ATLA was 6.5%, 57.4%, and 17.9%, respectively. Age‐specific prevalence of anti‐HBc and anti‐ATLA increased with age, but that of HBsAg did not. Sex‐specific prevalence of HBsAg was significantly higher in males than in females, but that of anti‐ATLA was significantly higher in females than in males. Statistical analysis revealed that prevalence of anti‐ATLA was significantly higher in HBsAg‐positive persons and HBsAg‐negative/anti‐HBc‐positive persons than in those negative for HBsAg and anti‐HBc. These data suggest that hepatitis B virus‐infected persons have a significantly higher chance of adult T‐cell leukemia virus infection than those without hepatitis B virus infection in the area studied.
Iron and Transferrin Uptake by Phytohemagglutinin‐Stimulated Human Peripheral Blood Lymphocytes
Authors: Taylor Peter George , Soyano Andrés , Romano Egidio , Layrisse Miguel
Published: November 2013

Iron (Fe) and transferrin (TF) uptake by human peripheral blood lymphocytes stimulated in vitro with phytohemagglutinin was measured. Pulses of 59FeTF or 125I‐TF were added to the cultures either at time 0 or 8 hr before the end of a 72‐hr incubation. In time‐course experiments, peak iron and transferrin uptake coincided with the peak of tritiated thymidine uptake taken as a measure of cellular activation. Iron, but not transferrin, was accumulated by the cells. Non‐linear relationships existed between both iron and transferrin uptake and the degree of activation. Both rose markedly above basal levels only at a level of activation at least 50% of the maximum observed. The results suggest that although iron utilization is related to cellular activity, the uptake mechanism is only activated when an increased iron metabolism has exhausted internal stores.
A New Salmonella Serovar S.III b 58: z10:z53: Rz50
Authors: Venkateswaran Kasthuri , Hashimoto Hideo , Jayasheela Manur , Tamura Kazumichi
Published: November 2013

A new Salmonella serovar S.III b 58: z10:z53: Rz50 was isolated from the water samples of Ashida river, Fukuyama city, Japan. Its antigenic structure is described.
Suppressive Effect of Macrophages on Interferon‐γ Production by Human Peripheral Lymphocytes Stimulated with Mycoplasma pneumoniae
Authors: Nakayama Tetsuo , Urano Takashi , Osano Mitsuru , Maehara Nobutoshi , Sasaki Keiko , Hayatsu Eizo , Makino Satoshi
Published: November 2013

Production of interferon (IFN)‐γ was investigated in human peripheral lymphocytes stimulated with Mycoplasma pneumoniae. Lymphocytes obtained from non‐immune individuals produced no IFN. IFN‐γ was produced by T cells obtained from immune individuals, and the helper/inducer T cells produced two‐ to sixfold higher titer of IFN‐γ than the suppressor/cytotoxic T cells. The addition of macrophages in T cell cultures suppressed the production of IFN‐γ; this differs from the previous result wherein the addition of macrophages enhanced the production of IFN‐γ, when stimulated with mumps virus or measles virus.
Immunoblotting Analysis of Anti‐Rickettsial Antibodies Produced in Patients of Tsutsugamushi Disease
Authors: Ohashi Norio , Tamura Akira , Suto Tsunehisa
Published: November 2013

The reactivity of sera from patients of Tsutsugamushi disease (scrub typhus) with the antigenic polypeptides of Rickettsia tsutsugamushi was analyzed by the immunoblotting method. The reactivity varied greatly among the sera of individual patients tested. IgG and IgM antibodies of most patients reacted with the 54–56 kilodalton (54–56K) polypeptide located on the rickettsial surface, suggesting that this polypeptide is a predominant antigen in the infection. Other polypeptides of 60K, 50–52K, 46–47K, 35K, and 21–25K were reactive with some but not all sera. From the reactivity of these polypeptides, it was suggested that the 54–56K polypeptide is both strain‐specific and group‐specific, the 60K polypeptide is group‐specific, and the 35K and 21–25K polypeptides are subgroup‐specific. IgG antibodies seem to be more cross‐reactive with polypeptides of multiple strains than IgM antibodies and have a tendency of increased cross‐reactivity that was observed in the sera obtained at the later stage of illness.
Effects of Cholera Toxin on Delayed‐Type Hypersensitivity to Sheep Red Blood Cells Inoculated Intranasally into Mice
Authors: Tamura Shin‐ichi , Samegai Yasuo , Kurata Hideki , Kurata Takeshi
Published: November 2013

The effects of cholera toxin (CT) on delayed‐type hypersensitivity (DTH) to sheep red blood cells (SRBC) were studied in mice sensitized by intranasal administration of SRBC. CT (1 μg/mouse), given intranasally together with SRBC (2 × 107/mouse), induced a maximally enhanced DTH response, which reached its peak around 7 days after sensitization, and also induced an accelerated DTH response upon a second administration of SRBC 28 days later. The ability of CT to enhance the DTH to SRBC was lost, either when CT was administered via the intraperitoneal or subcutaneous route, or when CT was introduced into the nasal site from which a large proportion of the SRBC was discharged 2 days after SRBC administration. These results indicate that the cells that are located in the nasal site and participate in the earlier events of DTH response were most affected by CT. The following effects of CT on the earlier events, which occur within 24 hr after the intranasal administration of both CT and SRBC, appeared to be involved in the mechanisms by which CT enhances DTH to SRBC: (i) facilitation of the penetration of the antigen into the nasal tissue; (ii) reinforcement of the migration of immunocompetent cells from the blood to the nasal tissues; (iii) promotion of the ability of Ia‐positive macrophages to present the antigenic determinants to T cells; (iv) facilitation of the differentiation of primed T cells to DTH‐effector T cells.
New Complement Fixation Test with Peroxidase‐Labeled Complement Clq for Direct and Quantitative Determination of Antibodies to Herpes Simplex Virus
Authors: Taguchi Fumiaki
Published: November 2013

An enzyme‐labeled complement fixation (ELISA‐CF) test for the direct and quantitative determination of complement fixing (CF) antibodies has been developed. This paper described the introduction of the ELISA‐CF test that used peroxidase‐labeled Clq component of complement to detect CF antibodies which had reacted with herpes simplex virus (HSV), as a virus model. Equal volumes of heat‐inactivated serum and the peroxidase‐labeled Clq (P*‐Clq) were simultaneously added to wells of microplates which had been coated with HSV CF antigen or with cell control antigen. The enzymatic activities of P*‐Clq bound to the immune complex were determined photometrically. The ELISA‐CF test allows processing of serum specimens in a 3‐hr operation, with procedural simplicity and increased specificity and sensitivity compared with the conventional CF test.
Expression of Antigen Genes of Leptospira interrogans Serovar canicola in Escherichia coli
Authors: Yamaguchi Tsuyoshi , Ono Etsuro , Yanagawa Ryo
Published: November 2013

Leptospira interrogans serovar canicola DNA was cloned into the plasmid pBR322 and introduced into E. coli. Eight out of approximately 10,000 transformants were found to express antigens of canicola by ELISA including colony ELISA blot test using anti‐canicola antiserum. The canicola antigens expressed in the transformants reacted with the antisera against the serovars belonging to Canicola serogroup and other serogroups of L. interrogans. They did not react, however, with the antiserum against L. biflexa (with only one exception) nor with the antiserum against Leptonema illini. Thus, the recombinant DNA technique may provide alternative possibilities for preparing antigens of leptospires.
Adsorption of LLCMK2 Cell‐Grown Sendai Virus onto Human Red Blood Cells and Its Release from the Virus Adsorbed Cells
Authors: Komatsu Hiroyoshi , Tozawa Hideki
Published: November 2013

An early stage of virus adsorption was studied in a system of Sendai virus metabolically labeled with [3H] leucine in LLCMK2 cells and of human red blood cells (RBCs). The efficiency of viral release from the virus‐bound RBCs by incubation at 37 C depended on the number of virus particles which had been used for adsorption onto the RBC at 4 C. When 7.8 × 102 virus particles were previously adsorbed onto the RBC at 4 C, most of the viruses were dissociated from the RBC at 37 C. In the case of adsorption of 3 to 12 virus particles per RBC, however, most of the viruses were not dissociated from the RBC by incubation at 37 C. Such RBC‐bound viruses were released by incubation with various bacterial neuraminidases (Clostridium perfringens, etc.) or with a large number of LLCMK2 cell‐grown Sendai virus (LLCMK2‐Sendai) particles, but not released by treatment with hemagglutinin‐neuraminidase protein (Sendai‐gp) isolated from egg‐grown Sendai virus.
Effective Production of Mouse Monoclonal Antibodies against Influenza Virus by Repeated Intrasplenic Immunization
Authors: Maéno Mitsugu , Shimizu Kazufumi , Shikata Toshio
Published: November 2013

Two immunization techniques that enable production of mouse monoclonal antibodies were evaluated in terms of small quantities of antigen. Various amounts of purified influenza A virus particles were applied either for in vitro sensitization in cultured splenocytes or for intrasplenic immunization, followed by hybridization of the immunized cells with mouse myeloma cells. Hybridomas producing specific antibodies for influenza viral proteins were detected by enzyme‐linked immunosorbent assay when more than 50 μg of antigens was used for the in vitro immunization method, and at least 5 μg was necessary for a single intrasplenic immunization. On the other hand, as little as 60 ng of antigen administered in two intrasplenic injections was sufficient to produce specific hybridomas. Two out of six randomly selected monoclonal antibodies obtained using the repeated intrasplenic immunization method were IgG and the other four were IgM. Immunoprecipitation analyses revealed that the recognized antigens involved a viral inner protein (nucleocapsid protein), as well as an envelope glycoprotein (hemagglutinin). We conclude that immunization by two direct injections of antigen into the spleen is the most effective method for sensitization with nanogram quantities of insoluble antigen such as influenza viruses.
A Role of Secreted Proteinase of Candida albicans for the Invasion of Chick Chorio‐Allantoic Membrane
Authors: Kobayashi Issei , Kondoh Yuzuru , Shimizu Katsumi , Tanaka Kenji
Published: November 2013

The invasion of Candida albicans strains into the chorio‐allantoic membrane (CAM) of a developing chick was studied by light and electron microscopy. A proteinase‐producing strain, NUM961, invaded into intact CAM, but proteinase‐deficient strain NUM678 cells remained on the surface of the CAM with no evidence of damage to the host cells. However, NUM678 cells invaded into the ectoderm‐damaged CAM, or proteinase‐treated one. Electron microscopy revealed that treatment with purified Candida proteinase disorganized the ectoderm tissue by disrupting the intercellular junctions. These results suggest that Candida proteinase damages the CAM surface, which enables the invasion of the growing hyphae.
Characterization of Replicative Form DNA of the Autonomous Parvovirus Mink Enteritis Virus
Authors: Shinagawa Morikazu , Nomura Yasuharu , Kariatumari Tsutomu , Ishiguro Naotaka , Horiuchi Motohiro , Goto Hitoshi
Published: November 2013

Characterization of replicative form (RF) DNA of mink enteritis virus (MEV) was carried out. Most of the RF DNA were bound to terminal protein but some were free from the protein. The protein‐free RF DNA increased about 7 times from 30 to 50 hr post‐infection, while the DNA with protein increased less. The molecules of the replicative intermediate which were partially single‐stranded DNA and bound to terminal protein were present. Two terminal conformations, “extended” and “turnaround,” were observed in both ends of both terminal protein‐bound and protein‐free RF DNA. The 5′ end labeling revealed that 5′ ends of protein‐free RF DNA were not blocked to phosphorylation by an amino acid or an oligopeptide which attaches to 5′ ends of proteolytically deproteinized RF DNA. Restriction analysis of incomplete RF DNA which was partially double‐stranded DNA showed that extended conformation was dominant in such incomplete RF molecules.
A Novel Enzyme Immunoassay for Conidia of Fusarium oxysporum f.sp. cucumerinum F504
Authors: Kitagawa Tsunehiro , Sakamoto Yuichiro , Ogura Hirosuke
Published: November 2013

This study was undertaken to develop a new tool to study fusarial diseases of plants. Micro‐ and macro‐conidia of a strain (F504) of Fusarium oxysporum were isolated and antiserum against the conidia was elicited in rabbits. A highly specific and sensitive competitive‐type enzyme‐linked immunosorbent assay (ELISA) for conidia of the strain was developed using the antiserum with β‐d‐galactosidase‐labeled anti‐rabbit IgG as the secondary antibody and conidia fragments of the strain as antigen attached to Amino‐Dylark solid‐phase balls. The assay was highly specific to conidia of the strain F504, while conidia‐free hypha of the strain F504 as well as all other microorganisms tested including nine other strains of Fusarium species showed little cross‐reactivity. Application of the ELISA to following the growth rates of conidia in hyphae of the strain F504 under several conditions are also reported.
Sugar Composition of the Polysaccharide Portion of Lipopolysaccharides of Vibrio fluvialis, Vibrio vulnificus, and Vibrio mimicus
Authors: Iguchi Takehiro , Kondo Seiichi , Hisatsune Kazuhito
Published: November 2013

A chemotaxonomic study was carried out on Vibrio fluvialis and V. vulnificus on the basis of the sugar composition of the polysaccharide portion of their lipopolysaccharides (LPS). A previously developed rapid method of preparing samples for compositional sugar analysis was employed. Nineteen O‐serogroups of V. fluvialis were divided into 14 chemotypes while seven O‐serogroups of V. vulnificus were divided also into seven chemotypes since the polysaccharide portion of LPS of each serogroup has a different sugar composition from that of the other serogroups. Close similarities in the sugar composition of the same portion were demonstrated between serologically cross‐reacting non‐O1 group V. cholerae and V. fluvialis, and non‐O1 V. cholerae and V. mimicus.
A Novel Neutrophil Adherence Test Effectively Reflects the Activated State of Neutrophils
Authors: Yakuwa Naoshi , Inoue Tomio , Watanabe Tadashi , Takahashi Keiji , Sendo Fujiro
Published: November 2013

We developed a novel and convenient neutrophil adherence test for ascertaining the activated state of neutrophils. Human peripheral blood neutrophils were placed in wells of a 96‐well flat‐bottom culture plate, and incubated in the presence or absence of neutrophil stimulants for varying periods of time at 37 C in a CO2 incubator (5% CO2, 95% air). After non‐adherent cells were completely removed by vibration on a vortex mixer, residual adherent cells were fixed and stained with crystal violet containing 12% formaldehyde and 10% ethanol. After thorough washing, 1% SDS was added to the plate, and the absorbance of each well was measured at 570 nm. Our results correlate well with those of a previously reported method using 51Cr‐labeled cells.
Computer Analysis between Nucleotide and Amino Acid Sequences of Bean Golden Mosaic Virus and Those of Maize Streak, Wheat Dwarf, Chloris Striate Mosaic, and Beet Curly Top Viruses
Authors: Ikegami Masato
Published: November 2013

Bean golden mosaic virus (BGMV) DNA 1 and 2 have little sequence homology with maize streak virus (MSV), wheat dwarf virus (WDV), and chloris striate mosaic virus (CSMV) DNAs. BGMV DNA 1 and beet curly top virus (BCTV) DNA are closely related, whereas BGMV DNA 2 and BCTV DNA are not related. Direct amino acid homologies of predicted proteins between BGMV ORFs and MSV ORFs, WDV ORFs or CSMV ORFs were 40–50%. BGMV 1L1 and BCTV L1, and BGMV 1L3 and BCTV L4 were highly conserved. The sequence TAATATTAC was detected in the loops of hairpin structures of 5 geminiviruses.
Enhancement of Phagocytosis and Bactericidal Activity of Neutrophils in Miniature Pigs by Dihydroheptaprenol, a Synthetic Polyprenol Derivative
Authors: Araki Seiichi , Suzuki Mamoru , Ogura Kazuaki , Kimura Makoto , Imamura Eisei , Kuniyasu Chikara , Kagaya Keiko , Fukazawa Yoshimura
Published: November 2013

Dihydroheptaprenol (DHP), a synthetic polyprenol derivative, markedly stimulated the generation of peripheral blood neutrophils after intramuscular injection in miniature pigs. The generated neutrophils exhibited enhanced phagocytic activity against latex particles and also enhanced killing activity against Escherichia coli. The effective dose in miniature pigs (1.4 mg/kg) was markedly less than that required in mice (100 mg/kg). These results indicate that DHP induces resistance to some bacterial infections in pigs, suggesting the applicability of DHP for humans.
Productive Ascites Growth of Heterohybridomas between Epstein‐Barr Virus‐Transformed Human B Cells and Murine P3×63Ag8.653 Myeloma Cells
Authors: Matsumoto Mitsuru , Mochizuki Kenichi , Kobayashi Yuzuru
Published: November 2013

Heterohybridomas between Epstein‐Barr virus‐transformed human B cells and murine myeloma P3×63Ag8.653 cells were found to produce sizable quantities of human monoclonal antibodies in nude mouse ascites. Mice injected intraperitoneally with two of such heterohybridomas yielded several milliliters of ascites fluid which contained nearly 100‐fold higher anti‐platelet activities than those in routine culture supernatants.
Monoclonal Antibodies against the Surface Antigens of Shigella flexneri Serotype 1b and Shigella dysenteriae Serotype 1
Authors: Suzuki Kumiko , Takeda Tae
Published: November 2013

Monoclonal antibodies against the surface antigens of Shigella flexneri 1b and S. dysenteriae 1 were prepared. The specificities of the antibodies were evaluated by enzyme‐linked immunosorbent assay (ELISA), and quantitative agglutination using microtiter plate. Monoclonal antibodies against S. flexneri 1b, designated Sf2B2 and Sf2G4, belonged to IgG2a and IgG1 subclass, respectively. The former was specific for S. flexneri 1b, whereas the latter was reactive not only to S. flexneri 1b, but also weakly to 3a and 4b. Monoclonal antibody against S. dysenteriae 1, Sd5E1 (IgM), reacted with S. dysenteriae 1, 3, 6, 7, and S. boydii 2.
Replication of Measles Virus in Vero Cells at Elevated Temperatures
Authors: Ogura Hisashi , Nakamura Shinichi
Published: November 2013

In one‐step growth experiment of measles virus (MV) in Vero cells at 39 C, the appearance of MV infectivity was delayed for 24 hr and the maximum titer was reduced by approximately 1,000‐fold, when compared with those at 35 C. MV infectivity was thermolabile at the high temperature. Penetration was rather enhanced at 39 C. By Northern blot hybridization, viral RNAs including 50S genome‐sized RNA and mRNAs were first detectable 24 hr post‐infection (PI) at 35 C and 36 hr PI at 39 C, respectively. Rapid degradation of viral mRNAs was not observed in the infected cells at 39 C. The synthesis of N, F, and M proteins was relatively reduced at the high temperature and appearance of the other viral protein was delayed, in agreement with the time course of viral RNA synthesis. All these data suggest that less efficient synthesis of viral RNA, restriction of synthesis of N, F, and M proteins at translational level and thermolability of infectivity are all involved in the suppressed MV production in Vero cells at 39 C.