bluuebunny/crossref_metadata_2025 huggingface dataset:
Crystallization of an R‐Form Lipopolysaccharide from Klebsiella pneumoniae
Authors: Kato Nobuo , Ohta Michio , Arakawa Yoshichika , Naito Setsuko , Sugiyama Tsuyoshi , Ito Hideo , Kido Nobuo , Sasaki Kyoyu , Asai Junpei
Published: November 2013

An R‐form lipopolysaccharide (LPS) from Klebsiella pneumoniae strain LEN‐111 (O3–:K1–) formed crystals, whose shapes were elongated hexagonal plates, trapezoid plates, and rhomboid plates, and whose greatest dimensions were 3.1 × 0.8 μm, when it was suspended in 50 mM Tris buffer at pH 8.5 containing 5 mM MgCl2 and kept at 4 C for as long as 870 days. K. pneumoniae LEN‐111 synthesized LPS molecules possessing incomplete repeating units of the O‐antigenic polysaccharide portion besides the R‐form LPS because of a leaky characteristic, but crystals consisted exclusively of the R‐form LPS. Although the size of crystals was not large enough for X‐ray analysis and limited crystallographic information was available, it was suggested that the crystals consist of hexagonal lattices with an a axis of 4.62 Å and c axis of 79.8 ±2.6 Å. The present results showed that R‐form LPS lacking the O‐antigenic polysaccharide portion tends to form crystals during long‐term incubation in Tris buffer at pH 8.5 containing MgCl2 at 4 C.
Bradykinin Generation Triggered by Pseudomonas Proteases Facilitates Invasion of the Systemic Circulation by Pseudomonas aeruginosa
Authors: Sakata Yoshifumi , Akaike Takaaki , Suga Moritaka , Ijiri Sumiko , Ando Masayuki , Maeda Hiroshi
Published: November 2013

To elucidate the mechanism of bacterial exoprotease in promotion of the intravascular dissemination of Pseudomonas aeruginosa, we examined the possible involvement of bradykinin (whose generation is induced by pseudomonal proteases in septic foci) in the invasion by bacteria, and in access of bacterial toxins to systemic blood circulation. P. aeruginosa 621 (PA 621), which produces very little protease, was injected intraperitoneally into mice together with pseudomonal exoproteases (elastase/alkaline protease). Dissemination of bacteria from the peritoneal septic foci to the blood was assessed by counting viable bacteria in the blood and spleen by use of the colony‐forming assay. The results showed that pseudomonal proteases markedly enhanced (10‐ to 100‐fold) intravascular dissemination of bacteria in mice. This enhancement was induced not only by pseudomonal proteases but also by bradykinin. More importantly, the increased spread of PA 621 induced by pseudomonal protease and bradykinin was significantly augmented by the addition of kininase inhibitors, indicating the direct involvement of bradykinin in bacterial dissemination. Similarly, bradykinin caused effective dissemination of pseudomonal toxins such as endotoxin (lipopolysaccharide) and exotoxin A when the toxins were injected into the peritoneal cavity with bradykinin. Furthermore, the lethality of the infection with PA 621 was strongly enhanced by pseudomonal proteases given i.p. simultaneously with PA 621. On the basis of these results, it is strongly suggested that pseudomonal proteases as well as bradykinin generated in infectious foci are involved in facilitation of bacterial dissemination in vivo.
Presence of Common Antigenic Epitope in Outer Surface Protein (Osp) A and OspB of Japanese Isolates Identified as Borrelia garinii
Authors: Masuzawa Toshiyuki , Kaneda Kazuhide , Suzuki Hiroyuki , Wang Jianhui , Yamada Kazuto , Kawabata Hiroki , Johnson Russell C. , Yanagihara Yasutake
Published: November 2013

Japanese Borrelia strains FujiP2, AP83, NT24, NT29 and HT2 which had a 31‐kilodalton protein non‐reactive with monoclonal antibody (MAb) H5332 to outer surface protein A (OspA) were identified as B. garinii by the DNA hybridization method. MAb P3134 raised to strain NT24 reacted with OspA and the OspB‐ranging protein of these isolates and cross‐reacted with the OspB‐ranging protein of some other isolates. Since the reactive protein was extracted by the Triton X‐114 phase partitioning method, the MAb recognized the common epitope present in OspA and OspB. To our knowledge, this is the first report of an MAb reactive to both OspA and OspB.
Possible Correlation between Borna Disease Virus Infection and Japanese Patients with Chronic Fatigue Syndrome
Authors: Kitani Teruo , Kuratsune Hirohiko , Fuke Isao , Nakamura Yurie , Nakaya Takaaki , Asahi Sayumi , Tobiume Minoru , Yamaguti Kouzi , Machii Takashi , Inagi Reiko , Yamanishi Koichi , Ikuta Kazuyoshi
Published: November 2013

Borna disease virus (BDV) is a neurotropic, as yet unclassified, non‐segmented, negative‐sense, single‐strand RNA virus. Natural infection with this virus has been reported to occur in horses and sheep. In addition, antibodies to BDV in plasma or BDV RNA in peripheral blood mononuclear cells (PBMCs) were also found in patients with neuropsychiatric diseases. We describe here the possible link between the patients with chronic fatigue syndrome (CFS) and infection with BDV.
Effect of Macrolide Antibiotics on Macrophage Functions
Authors: Xu Guang , Fujita Jiro , Negayama Kiyoshi , Yuube Koichi , Hojo Satoko , Yamaji Yasufumi , Kawanishi Koichi , Takahara Jiro
Published: November 2013

Macrolide antibiotics have a variety of actions other than antimicrobial activities. Recently, it has been suggested that macrolide antibiotics act as immunomodulators. In this study, we evaluated the effects of macrolide antibiotics on macrophage functions. For the macrophage, we used the mouse macrophage cell line J774.1. The following effects of macrolide antibiotics on macrophage functions were evaluated: the effect of macrolide antibiotics on macrophage growth; the phagocytosis of beads; cytocidal activity against Candida albicans; and chemotaxis to lipopolysaccharide (LPS). Macrolide antibiotics except for azithromycin significantly stimulated the growth of the macrophage. In addition, pretreatment with macrolide antibiotics except for roxithromycin significantly stimulated the macrophage phagocytosis of beads, macrophage chemotaxis to LPS, and macrophage cytocidal activity against Candida albicans. These results suggest that macrolide antibiotics stimulate macrophage functions.
Adherence of Peripheral Blood Leukocytes to Cytomegalovirus‐Infected Fibroblasts
Authors: Watanabe Masahiro , Ito Masahiro , Kamiya Hitoshi , Sakurai Minoru
Published: November 2013

Peripheral blood monocytes and B cells adhered to cytomegalovirus (CMV)‐infected fibroblasts, whereas T cells and polymorphonuclear leukocytes did not adhere to either CMV‐infected or uninfected fibroblasts. When T cells were activated with anti‐CD3 antibody, activated T cells demonstrated adherence and cytotoxicity to both CMV‐infected and uninfected fibroblasts. Adherence of peripheral blood mononuclear cells (PBMC) and cytotoxicity mediated by adherent activated T cells were blocked by treatment of CMV‐infected fibroblasts with anti‐ICAM‐1 antibody and by treatment of leukocytes with anti‐LFA‐1 antibody. These data suggest that an interaction of ICAM‐1 and LFA‐1 is responsible for the adherence of leukocytes and for adherent activated T cell‐mediated cytotoxicity against CMV‐infected fibroblasts.
Genotype Analysis of Hepatitis C Virus among Blood Donors and Inmates in Metro Manila, The Philippines
Authors: Katayama Yuko , Barzaga Nina G. , Alipio Arlene , Soetjipto  , Doi Hisaya , Ishido Satoshi , Hotta Hak
Published: November 2013

Antibodies against hepatitis C virus (HCV) were detected in 18 (2.3%) of 800 sera from commercial blood donors and 23 (4.6%) of 502 sera from inmates in Metro Manila, the Philippines. The difference in the antibody prevalence between the two groups was statistically significant (P<0.05). HCV RNA was detected in 14 (78%) of the 18 antibody‐positive sera from blood donors and 19 (83%) of the 23 antibody‐positive sera from inmates. Genotype analysis revealed that HCV‐1b (50%) was most common among blood donors, followed by HCV‐1a (36%) and HCV‐2a (7%). Among inmates, on the other hand, HCV‐1a (68%) was most common, followed by HCV‐1b (11%), HCV‐2a (5%) and HCV‐2b (5%). Overall, HCV‐1a and HCV‐1b appeared to be predominant among them. Thus, the genotype prevalence in the Philippines was distinct from those in other Southeast Asian countries such as Thailand, Vietnam and Indonesia, and also distinct from those in the Far East including Taiwan, Mainland China and Japan.
Comparison of OspA Serotypes for Borrelia burgdorferi Sensu Lato from Japan, Europe and North America
Authors: Masuzawa Toshiyuki , Wilske Bettina , Komikado Tetsuro , Suzuki Hiroyuki , Kawabata Hiroki , Sato Nanao , Muramatsu Koichi , Sato Nobutake , Isogai Emiko , Isogai Hiroshi , Johnson Russell C. , Yanagihara Yasutake
Published: November 2013

Sixty‐one Borrelia burgdorferi sensu lato strains from various sources (ticks, human, and wild animals) in Japan and two strains from ticks in Far Eastern Russia were classified on the basis of reactivity with 16 monoclonal antibodies (mAb) to outer surface protein A (OspA) and by DNA‐DNA hybridization assay. Eleven OspA serotypes (J1 to J11) were recognized among the Japanese and the Far East Russian isolates (serotypes J1 to J9 were identified as B. garinii, serotype J10 was identified as B. afzelii, and serotype J11 corresponded to B. japonica), whereas 7 OspA serotypes for North American and European isolates previously reported (Bettina Wilske et al, J. Clin. Microbiol. 31:340‐350, 1993) were not observed except for OspA serotype 2 which showed identical reactivity with OspA serotype J10. This finding provides helpful information for understanding the geographical distribution of Lyme disease borrelia and the development of vaccine and diagnostic tests. In conclusion: 1. B. burgdorferi sensu stricto has not been observed in Japan, 2. Japanese B. afzelii isolates are closely related to those from Europe, 3. B. garinii isolates from Japan are highly heterogeneous and apparently different from European B. garinii isolates.
Continuous Isolation and Characterization of Chlamydia pneumoniae from a Patient with Diffuse Panbronchiolitis
Authors: Miyashita Naoyuki , Matsumoto Akira , Kubota Yoshifumi , Nakajima Masamitsu , Niki Yoshihito , Matsushima Toshiharu
Published: November 2013

We succeeded in isolating Chlamydia pneumoniae organisms continuously from a 70‐year‐old man who had received chemotherapeutic treatment with low dosages of erythromycin for five years to improve diffuse panbronchiolitis (DPB). He had two episodes of acute exacerbation of DPB and a total of six strains of C. pneumoniae were isolated at different stages during the past 13 months. The morphological properties and protein profiles of the elementary bodies of all the C. pneumoniae isolates were similar to each other. Interestingly, his serological response against C. pneumoniae in immunoblotting tests was differed between two episodes. The sera collected during the first episode reacted weakly to the major outer membrane protein (MOMP), whereas those collected during the second episode reacted strongly to the 60‐kDa protein and weakly to MOMP. These facts suggest that the two different episodes occurred as a result of different mechanisms. Additionally, in spite of the low antibody titer by micro‐immunofluorescence test in the second episode as compared with that of the first episode, the immune response against 60‐kDa immunodominant protein increased markedly in the second episode, and we suspect that the second episode was due to an allergic reaction caused by this 60‐kDa protein. These findings suggest that repeated or prolonged exposure to C. pneumoniae may be associated with acute exacerbations of chronic obstructive pulmonary disease, and that the patient should be noted as a possible source of C. pneumoniae infection.
Genetic Analysis of Borrelia burgdorferi Sensu Lato in Korea Using Genomic Hybridization and 16S rRNA Gene Sequence Determination
Authors: Kee Sun‐Ho , Yoon Jung‐Hoon , Oh Hee‐Bok , Park Yong‐Ha , Kim Yoon‐Won , Cho Min‐Kee , Park Kyung‐Seok , Chang Woo‐Hyun
Published: November 2013

Nine Borrelia burgdorferi sensu lato isolated in Korea were subjected to genomic hybridization using 16S rRNA gene probe and specific restriction patterns (HindIII and EcoRV) led these nine Borrelia into five subtypes. The evolutionary relationships of the five isolates corresponding to five RFLP groups were measured through the sequence determination of 16S rRNA gene and phylogenetic analysis. The isolates 935T (group I), 934U and 17Y (Group IIa, IIb) were well clustered with B. garinii and B. afzelii. 5MT and 9MT strains (Group IIIa and Group IIIb) formed a common branch shared with B. afzelii cluster although the evolutionary distance was rather long. So, most of B. burgdorferi sensu lato in Korea was B. afzelii or B. afzelii‐related group and some minor group such as B. garinii also existed.
No Proof of HTLV‐I/II in Intravenous Drug Abusers with a High Rate of HIV‐1 Infection in Central Thailand
Authors: Saguanwongse Suranga , Muangpom Amorntip , Ruchusatsawat Nuangjun , Wongcheree Suthon , Warachit Paijit , Miyamura Kikuko , Yamazaki Shudo , Honda Mitsuo
Published: November 2013

Serum specimens of 1,074 intravenous drug abusers (IVDA) were examined for infection with HIV‐1, HTLV‐I and HTLV‐II in central Thailand. Three hundred and sixty‐two of the specimens were seropositive for HIV‐1 (33.7%). The HIV‐1 seropositive IVDAs exhibited increased seropositivity with age through group 40–44 and significantly decreased seropositivity over the age of 45. In contrast, no seropositivity to either HTLV‐I or ‐II was detected in the samples tested by a particle‐agglutination assay for HTLV followed by type‐specific Western blotting for HTLV. Reference to previous reports suggested that the rate of HIV infection in IVDAs has decreased with no HTLV‐I or HTLV‐II in Thailand when compared with that of the HIV infection in 1992.
Effects of Nerve Growth Factor and Phorbol Derivative on Reactivation of Herpes Simplex Virus Type 1 in Cultured Cells of Latently Infected Adult Mouse Trigeminal Ganglia
Authors: Kimura Susumu
Published: November 2013

Reactivation of herpes simplex virus type 1 (HSV‐1) occurred rapidly in cells of latently infected adult mouse trigeminal ganglia which were cultured in serum‐free medium in the presence of sufficient nerve growth factor (NGF). However, HSV‐1 reactivation was delayed significantly in ganglionic cultures in the absence of exogenous NGF or in cultures treated with 2‐aminopurine in the presence of NGF. The delayed viral reactivation in ganglionic cultures without NGF was accelerated by treatment with phorbol myristate acetate or dibutyryl cyclic AMP. Culture conditions which affected HSV‐1 reactivation did not affect replication of HSV‐1 in normal ganglionic cultures.
Augmentation of Interferon Production after Cell‐Differentiation of U937 Cells by TPA
Authors: Indoh Tomokazu , Shirakawa Sachiko , Kubota Tohru , Yashiki Teruo , Isogai Emiko , Fujii Nobuhiro
Published: November 2013

Persistent infections with mumps virus were established in several human lymphoid cells of T‐cell origin (Molt‐4, TALL‐1, and CCRF‐CEM) and human monocyte cells (U937 and THP‐1). 2′,5′‐Oligoadenylate synthetase (2–5AS) activity was demonstrated to be only slightly induced by interferon (IFN) or TPA (12‐O‐tetradecanoyl‐phorbol‐13‐acetate) treatment in these cells. Treatment of the persistently infected cells with IFN or TPA did not stimulate an increase in the amount of synthetase mRNA. Induction of cell differentiation and augmentation of IFN production by TPA were demonstrated in U937 cells persistently infected with mumps virus (U937‐MP). Similar results for IFN production were obtained from differentiated U937 cells. It is suggested that cell differentiation of U937 cells might be associated with the development of IFN inducibility.
Acid‐Fastness of Nocardia asteroides GUH‐2 Cultured in Brain Heart Infusion Broth Supplemented with Paraffin
Authors: Kohbata Shunro
Published: November 2013

Nocardia asteroides GUH‐2 was not acid‐fast when grown in brain heart infusion (BHI) broth. When grown in BHI broth supplemented with paraffin, many filamentous cells showed acid‐fastness after treatment with 1% acid‐alcohol as the decolorizing agent. When treated with 3% acid‐alcohol, filamentous cells were not acid‐fast. In addition to the acid‐fast filamentous cells of nocardiae, unknown acid‐fast spherical bodies were observed in the paraffin‐supplemented BHI broth cultures.
Pentoxifylline Pretreatment Enhances the Oxidative Burst Activity of Human Peripheral Blood Mononuclear Cells
Authors: Chang Feng‐Yee , Shaio Men‐Fang
Published: November 2013

The effect of pentoxifylline pretreatment on the lucigenin‐augmented chemiluminescence and dismutase‐inhibitable superoxide production of human neutrophils and mononuclear cells (MNCs) was studied. Pentoxifylline at 20–2,000 μg/ml enhanced the lucigenin‐augmented chemiluminescence (118–165% of the control, P < 0.01) of phorbol myristate acetate (PMA)‐stimulated MNC. Pentoxifylline at 20–2,000 μg/ml increased the MNC superoxide production, i.e., 142–171% of the control (P < 0.05) using PMA stimulation and 145–159% of the control (P < 0.01) using opsonized zymosan stimulation. In contrast, pentoxifylline (up to 2,000 μg/ml) did not influence the lucigenin‐augmented chemiluminescence and superoxide production of human neutrophils, stimulated by either PMA or opsonized zymosan. These results suggest that pentoxifylline is an immunomodulator and may have potential usefulness in the enhancement of immune defenses in compromised hosts.
Utilization of Iron Sources and Its Possible Roles in the Pathogenesis of Vibrio parahaemolyticus
Authors: Wong Hin‐Chung , Liu Chi‐Chang , Yu Chung‐Ming , Lee Yeong‐Sheng
Published: November 2013

Vibrio parahaemolyticus is an important enteropathogen in Japan, Taiwan and other coastal regions. The influence of the regulation of iron on the pathogenesis of this pathogen has not been well characterized. The growth of pathogenic and non‐pathogenic strains of V. parahaemolyticus on iron‐limited agar plates was stimulated by ferritin, lactoferrin and transferrin at 30 μM, and also by hemin, hemoglobin and ferric ammonium citrate at 100 μM. Spontaneous iron‐utilizing mutant strains (mutants) were derived from a clinical strain, ST550. Compared with the parent strain, lowered virulence was demonstrated for these mutants, as assayed by adult mouse and suckling mouse models. The in vivo growth and enterotoxigenicity of these mutants were also lower in the suckling mice. Adherence of the mutants to excised mouse intestine was lower as demonstrated by scanning electron microscopy. The iron‐regulated outer membrane protein profile also changed in selected mutants. These results indicate that iron‐regulated outer membrane proteins and other unknown factors associated with iron utilization may have profound influences, besides iron acquisition, on the pathogenesis of V. parahaemolyticus.
Serological Characterization of Trichosporon cutaneum and Related Species
Authors: Ikeda Reiko , Yokota Mariko , Shinoda Takako
Published: November 2013

Recent molecular biological, chemical, physiological and morphological studies indicate that Trichosporon cutaneum and related species should be reclassified. In this study, antigenic characteristics of the species were determined. The results of adsorption experiments revealed that there were at least three serological types: I, II and III. Specific factor sera I, II and III were prepared on the basis of adsorption experiments and isolates were serotyped by cell slide agglutination (CSA). Since the CSA test was difficult to read in some strains, the results of the CSA test were compared with the findings from an enzyme‐linked immunosorbent assay (ELISA). For the ELISA, crude polysaccharide antigens prepared from the culture supernatant were used as the antigen. The types determined by ELISA correlated well with those determined by the CSA test. These data suggest that T. cutaneum and related species have at least three serological types, and that the typing can be done by either CSA or ELISA.
Re‐Speciation of the Original Reference Strains of Serovars in the Citrobacter freundii (Bethesda‐Ballerup Group) Antigenic Scheme of West and Edwards
Authors: Miki Kanji , Tamura Kazumichi , Sakazaki Riichi , Kosako Yoshimasa
Published: November 2013

The antigenic scheme for the Bethesda‐Ballerup group of bacteria established by West and Edwards in 1954 has continued to be applied as a serotyping scheme for Citrobacter freundii. In 1993, however, the classification of the Citrobacter was drastically revised and the species C. freundii redefined by Brenner et al. Accordingly, to judge the propriety to continuously use a single antigenic scheme for the C. freundii complex, the 90 reference strains listed in the antigenic scheme for C. freundii by West and Edwards were characterized phenotypically and specified based on the revised classification. Of these 90 strains, two strains of Hafnia alvei and one of Escherichia coli were found. Among the remaining 87 reference strains, Citrobacter youngae was the predominant species (40 strains), followed by Citrobacter braakii (25 strains), Citrobacter werkmanii (13 strains), and the unnamed Citrobacter genospecies 10 of Brenner et al (six strains). Citrobacter freundii, as redefined, accounted for only three strains and ranked behind the other four species. No overlapping with most of the 42 O‐groups and 82 H‐antigens was recognized between species with few exceptions. O‐groups 1–9 inclusive, which were estimated to represent more than 90% of the former C. freundii strains, occurred in strains of C. youngae and C. braakii; and all nine strains of O‐group 29, formerly known as the Ballerup group, were identified as C. braakii. These findings suggest that further study of the serotyping system is needed for all H2S‐producing Citrobacter species.
Reservoir Competence of the Vole, Clethrionomys rufocanus bedfordiae, for Borrelia garinii or Borrelia afzelii
Authors: Ishiguro Fubito , Takada Nobuhiro , Nakata Keisuke , Yano Yasuhiro , Suzuki Hiroyuki , Masuzawa Toshiyuki , Yanagihara Yasutake
Published: November 2013

In autumn of 1994 and spring of 1995, we examined Borrelia infection among Microtinae voles, Clethrionomys rufocanus bedfordiae, in Hokkaido, Japan. In BSK culturing of the earlobe tissues of 45 C. rufocanus bedfordiae captured, twelve rodents were positive for Borrelia. Eight isolates were used for the polymerase chain reaction (PCR) and the restriction fragment length polymorphism (RFLP) analysis. According to the results, these isolates were classified into B. garinii or B. afzelii. It is considered that a common vole, C. rufocanus bedfordiae, plays a significant role in the transmission and maintenance of B. garinii and B. afzelii, similar to the role of Apodemus speciosus mice.
Different Sensitivity to Streptolysin‐O of Cells in Macrophage Lineage
Authors: Tanigawa Takahiko , Suzuki Jun , Ueta Tsuyoshi , Katsumoto Tetsuo , Tanaka Yoshinori
Published: November 2013

We have surveyed the sensitivity of cells in macrophage lineage to Streptolysin‐O (SLO). SLO had cytotoxic activity on immature myeloid cell lines such as M1 and WEHI‐3BD+. SLO was toxic to the cells after a 2‐hr incubation. However, mature macrophage cell lines such as A640‐BB‐2, J774, and P388D1 were not sensitive to the same dose of SLO. After M1 cells were treated with leukemia inhibitory factor (LIF), a differentiation‐inducer to macrophage, these cells became insensitive to SLO in one day. This cytotoxic action of SLO was inhibited by pretreatment with anti‐Streptolysin‐O antibody or cholesterol. These results indicate that SLO has different effects on macrophage lineage.
Simple Method for Detection of Clostridium botulinum Type A to F Neurotoxin Genes by Ploymerase Chain Reaction
Authors: Takeshi Kouichi , Fujinaga Yukako , Inoue Kaoru , Nakajima Hiroshi , Oguma Keiji , Ueno Tetsuya , Sunagawa Hiroyuki , Ohyama Tohru
Published: November 2013

A polymerase chain reaction (PCR)‐based method was established to detect each type of neurotoxin genes of Clostridium botulinum types A to F by employing the oligonucleotide primer sets corresponding to special regions of the light chains of the neurotoxins. In this procedure, the PCR products were easily confirmed by restriction enzyme digestion profiles, and as little as 2.5 pg of template DNAs from toxigenic strains could be detected. The specific PCR products were obtained from toxigenic C. botulinum types A to F, a type E toxin‐producing C. butyricum strain, and a type F toxin‐producing C. baratii strain, but no PCR product was detected in nontoxigenic strains of C. botulinum and other clostridial species. The neurotoxin genes were also detected in food products of a seasoned dry salmon and a fermented fish (Izushi) which had caused type E outbreaks of botulism. Therefore, it is concluded that this PCR‐based detection method can be used for the rapid diagnosis of botulism.
Enhanced Engraftment of HTLV‐I‐Infected Human T Cells in Severe Combined Immunodeficiency Mice by Anti‐asialo GM‐1 Antibody Treatment
Authors: Ishihara Shiro , Okayama Akihiko , Nagatomo Yasuhiro , Murai Koichi , Yamashita Ryozo , Okamoto Masayuki , Shima Takashi , Sasaki Takashi , Mueller Nancy , Tachibana Nobuyoshi , Tsubouchi Hirohito
Published: November 2013

The effects of anti‐asialo GM‐1 antibody (AAGM) treatment on the engraftment of human T‐cell leukemia virus type I (HTLV‐I)‐infected human T cells in severe combined immunodeficiency (SCID) mice were studied. The frequency of tumor formation in an HTLV‐I‐transformed human T‐cell line, MT‐2 cells, at the site of inoculation was significantly higher in AAGM‐treated than untreated mice (P<0.05): 16/18 (89%) and 16/26 (62%), respectively. The promotive effect of AAGM treatment on tumor development was marked in the early stage (less than 3 weeks), suggesting that the immediate reaction of natural killers to the inoculated cells may be important for the prevention of tumor development. The surface phenotypes and clonality of the tumor cells were the same as the MT‐2 cells inoculated. Inoculation of peripheral blood mononuclear cells (PBMC) from one of the 4 adult T‐cell leukemia/lymphoma (ATL) patients resulted in the development of tumors in AAGM‐treated SCID mice. However, the surface phenotypes of the cells from these tumors were a mixture of B cells and T cells, suggesting that these tumors consisted of Epstein‐Barr virus‐transformed B cells and HTLV‐I‐transformed T cells. In addition, HTLV‐I was detected by polymerase chain reaction in various organs of the mice inoculated with PBMC from the ATL patient and the asymptomatic carrier examined. These results suggest that elimination of natural killer function by AAGM treatment is important, although such treatment is not always necessary for the engraftment of HTLV‐I‐infected cells in SCID mice.
Activation of Unusual Mac‐1+CD4− CD8− T Cells Bearing αβ Antigen Receptor in Murine Lungs during Infection with Mycobacterium bovis BCG: Regulation by Interferon‐γ
Authors: Teruya Katsuji , Kawakami Kazuyoshi , Saito Atsushi
Published: November 2013

We have recently (Kawakami et al, Immunol. Lett. 1995;46: 143) demonstrated that unusual Mac‐1+CD4−CD8− T cells bearing αβ antigen receptor (Mac‐1+ αβ T cells) reside in a considerable proportion in murine lungs. The present study was performed to examine the dynamics of accumulation of these cells in the lungs following intravenous administration of Mycobacterium bovis BCG (BCG). Mac‐1+ αβ T cells accumulated rapidly 24 hr after infection, followed by a gradual increase over the observation period of 15 days. Furthermore, the expression of Ia, ICAM‐1 and FcγR II/III on their surface intensified dramatically after BCG infection. The kinetics of enhancement of Ia expression was slower than that of ICAM‐1, with the maximum level attained in one day in the latter molecule but in two weeks in the former. Neutralization of endogenous IFN‐γ by specific mAb completely blocked the augmented expression of Ia on Mac‐1+ αβ T cells after BCG infection, but did not have any significant effect on that of ICAM‐1. In contrast, in vivo administration of IFN‐γ enhanced the expression of ICAM‐1 as well as that of Ia. Our results indicate that accumulation of Mac‐1 αβ T cells within the lung is associated with a differential change in the expression of surface antigens, and suggest that these cells may play a role in the host defense against mycobacterial infection.
Demonstration of a Heat‐Stable 120‐Kilodalton Protein of Rickettsia japonica as a Spotted Fever Group‐Common Antigen
Authors: Uchiyama Tsuneo , Zhao Licheng , Uchida Takahiro
Published: November 2013

Genomic libraries of Rickettsia japonica were cloned into an expression vector λgt11. A clone expressing a protein reactive with antiserum against 120‐kilodalton (kDa) proteins, a mixture of heat‐modifiable and heat‐stable polypeptides, was selected and designated as λRj120‐1. The expressed protein has a molecular mass of 180 kDa. Western immunoblotting demonstrated that the expressed protein was a fusion protein with β‐galactosidase. The antiserum against 120‐kDa proteins was absorbed by the induced lysogen, resulting in the removal of reactivity to the heat‐stable 120‐kDa polypeptide. The antiserum against the expressed protein reacted with heat‐stable 120‐ to 130‐kDa polypeptides of spotted fever group (SFG) rickettsiae in addition to R. japonica. The findings indicated that the protein expressed from the cloned gene of R. japonica possessed the antigenicity group‐common to SFG rickettsiae. Primers designed from the gene coding for R. conorii heat‐stable 120‐kDa protein (Schuenke, K.W., and Walker, D.H., Infect. Immun. 62: 904‐909, 1994) and λgt11 lacZ gene amplified the λRj120‐1 DNA by the polymerase chain reaction (PCR). Analysis of restriction fragment length polymorphism (RFLP) of the PCR‐amplified products revealed that the cloned DNA corresponds to a portion of the gene coding for the heat‐stable 120‐kDa protein of R. conorii with 2,519 nucleotides beginning at nucleotide 190 of the open reading frame. RFLP demonstrated that the cloned gene was highly homologous to the corresponding gene of R. conorii.
Expression and Purification of the Crystalline Surface Layer Protein of Rickettsia typhi
Authors: Hahn Myong‐Joon , Chang Woo‐Hyun
Published: November 2013

The crystalline surface layer (S‐layer) protein (SLP) of Rickettsia typhi is known as the protective antigen against murine typhus. We previously reported a cloning and sequence analysis of the SLP gene of R. typhi (slpT) and showed that the open reading frame of this gene encodes both the SLP and a 32‐kDa protein. To express only the SLP from this gene, the putative signal sequence and the 32‐kDa protein portion were removed from the slpT. This protein was expressed in Escherichia coli as a fusion protein, consisting of the SLP and maltose binding protein. The recombinant protein reacted strongly with polyclonal antiserum of a patient with murine typhus.
ExperimentalChlamydia psittaciInfection of Japanese Quail
Authors: Takashima Ikuo , Hiyoshi Maki , Kariwa Hiroaki , Mukaiya Ryusuke , Hashimoto Nobuo
Published: November 2013

Japanese quail were used for the infection model of avian chlamydiosis. One‐day‐old Japanese quail were highly susceptible to lethal infection by aChlamydia psittacistrain of budgerigar origin upon inoculation via the air sac route with 104.1FFU of the organism, showing an acute and lethal course with chlamydial propagation. In contrast, 7‐day‐old quail developed resistance to the infection as shown by the lack of lethal effect with the same dose. The resistance of 7‐day‐old birds was abolished by immunosuppressive treatment with cyclophosphamide. Upon inoculation with a sublethal dose of 102.1FFU, latent infection was established in 1‐day‐old birds with a minimum number of the organism. The latent infection in the birds was converted to the lethal form by treatment with cyclophosphamide along with chlamydial propagation and suppression of antibody production.
Analysis of Vibrio cholerae O139 Bengal Isolated from Different Geographical Areas Using Macrorestriction DNA Analysis
Authors: Kurazono Hisao , Yamasaki Shinji , Ratchtrachenchai Orn‐anong , Nair G. Balakrish , Takeda Yoshifumi
Published: November 2013

Vibrio cholerae O139 isolated from different countries, as well as from different locations within a country, were examined using macrorestriction DNA analysis to determine the clonality of the O139 strains. NotI digests of genomic DNA of representative strains from Nepal, India, Bangladesh, China, Thailand, and Malaysia revealed very similar but not identical patterns. Examinations of the banding patterns generated by pulsed‐field gel electrophoresis of strains isolated within countries revealed complete homogeneity. These results further reiterate the spread of an identical clone of V. cholerae O139 although it appears that genetic polymorphism among the O139 strains is becoming apparent.
Cryptic Appearance of a New Clone of Vibrio cholerae Serogroup O1 Biotype El Tor in Calcutta, India
Authors: Yamasaki Shinji , Nair G. Balakrish , Bhattacharya S.K. , Yamamoto Shingo , Kurazono Hisao , Takeda Yoshifumi
Published: November 2013

In this study, pulsed‐field gel electrophoresis (PFGE) was applied to determine if the Vibrio cholerae O1 strains which reappeared after being temporarily displaced in Calcutta by the O139 serogroup were different from those isolated before the advent of the O139 serogroup. NotI digestion generated a total of 11 different patterns among the 24 strains of V. cholerae randomly selected to represent different time frames. Among the V. cholerae O1 strains isolated after July 1993, 4 PFGE banding patterns designated as H through K were observed with pattern H dominating. Pattern H was distinctly different from all other patterns encountered in this study including patterns A, B and C of V. cholerae O1 El Tor, which dominated before November 1992, and pattern F, which was the dominant V. cholerae O139 pattern. Further, pattern H was also different from the NotI banding patterns of the representative strains of the 4 toxigenic clonal groups of V. cholerae O1 El Tor currently prevailing in different parts of the world. NotI fragments of the new clone of V. cholerae O1 did not hybridize with an O139 specific DNA probe, indicating that there was no O139 genetic material in the new clone of V. cholerae O1. Hybridization data with an O1‐specific DNA probe again differentiated between the clones of V. cholerae O1 existing before the genesis of the O139 serogroup and the O1 strains currently prevalent.
Location and Ultrastructure of Borrelia japonica in Naturally Infected Ixodes ovatus and Small Mammals
Authors: Yano Yasuhiro , Takada Nobuhiro , Ishiguro Fubito
Published: November 2013

The internal organs of Ixodes ovatus and the ears of wild rodents (Apodemus speciosus, Eothenomys smithii) and an insectivore (Crocidura dsinezumi) were cultured to isolate borreliae; positive samples were examined for the distribution and dissemination of spirochetes in the host tissues using electron microscopy. Seven isolates were derived from the unfed ticks and the three species of mammals. These isolates were identified as Borrelia japonica judging from the outer surface protein profile using sodium dodecyl sulfate‐polyacrylamide gel electrophoresis and reactivity to a B. japonica‐specific monoclonal antibody. Borreliae were found only in the midgut lumen of the tick in close contact with the microvilli on the midgut epithelium; on the other hand, borreliae found in the ears of the mammals existed freely in the collagenous intercellular substances of connective tissues or in close contact with fibrocytes. The ultrastructural disparities between the borreliae in ticks and mammals appeared to correspond to differences in motility. Interestingly, the borrelia which invaded through the perineurium appeared to contact the basement membrane of a Schwann cell that enclosed several nonmyelinated nerve fibers. This may offer important information regarding the involvement of the nervous system in Lyme disease.
Optical Configuration Analysis of Hydroxy Fatty Acids in Bacterial Lipids by Chiral Column High‐Performance Liquid Chromatography
Authors: Nakagawa Yoji , Kishida Kenichi , Kodani Yoshifumi , Matsuyama Tohey
Published: November 2013

Through the adoption of a chiral stationary phase in high‐performance liquid chromatography and a simple derivatization method for hydroxy fatty acids, it became easy to separate and identify the optical isomers of 2‐ and 3‐hydroxy fatty acids composing several kinds of microbial lipids. The 2‐ and 3‐hydroxy fatty acids were converted with dinitrophenyl isocyanate to their 3, 5‐dinitrophenyl urethane derivatives (DU‐derivatives), which were analyzable by HPLC using a chiral column. By varying the composition of an eluent, separation of the DU‐derivatives of hydroxy fatty acids differing in optical configuration, chain length and position of hydroxyl group was achieved. The general elution orders of these DU‐derivatives were determined with authentic 2‐ and 3‐hydroxy fatty acids. Small amounts (~300 μg) of ornithine‐containing lipids isolated from the Serratia marcescens strains were examined by this method to identify 3‐hydroxy fatty acids of the lipids as D isomers.
Alterations in the Expression of ELAM‐1, ICAM‐1 and VCAM‐1 after In Vitro Infection of Endothelial Cells with a Clinical Isolate of Human Cytomegalovirus
Authors: Shahgasempour Shapour , Woodroffe Salwa B. , Garnett Helen M.
Published: November 2013

Human cytomegalovirus (HCMV) infection of endothelial cells resulted in increased adhesion of the cells to peripheral blood leukocytes. It was demonstrated by flow cytometry that increased adhesiveness parallels the increased expression of cell surface adhesion molecules (ELAM‐1, ICAM‐1, VCAM‐1). The increased adhesion of PMN and T‐lymphocytes was due to upregulation in the expression of ELAM‐1 and ICAM‐1. The upregulation of VCAM‐1 resulted in the increased adhesiveness of monocytes and T‐lymphocytes to HCMV‐infected HUVEC. The increased adhesiveness to leukocytes was caused by HCMV replication since endothelial cells exposed to HCMV‐free supernatants and UV‐inactivated HCMV did not show any increase in adhesiveness to any of the leukocytes tested.
Analysis of the Structural Gene Encoding a Hemolysin in Vibrio mimicus
Authors: Rahman Md. Monzur , Miyoshi Shih‐Ichi , Tomochika Ken‐Ichi , Wakae Hitoshi , Shinoda Sumio
Published: November 2013

An environmental isolate of V. mimicus, strain E‐33, has been reported to produce and secrete a hemolysin of 63 kDa. The hemolysin is enterotoxic in test animals. The nucleotide sequence of the structural gene of the hemolysin was determined. We found a 2,232 bp open reading frame, which codes a peptide of 744 amino acids, with a calculated molecular weight of 83,903 Da. The sequence for the structural gene was closely related to the V. cholerae el tor hlyA gene, coding an exocellular hemolysin. The amino terminal amino‐acid sequence of the 63 kDa hemolysin, purified from V. mimicus, was determined by the Edman degradation method and found to be NH2‐S‐V‐S‐A‐N‐N‐V‐T‐N‐N‐N‐E‐T. This sequence is identical from S‐152 to T‐164 predicted from the nucleotide sequence. So, it seems that the mature hemolysin in V. mimicus is processed upon deleting the first 151 amino acids, and the molecular mass is 65,972 Da. Analyzing the deduced amino‐acid sequence, we also found a potential signal sequence of 24 amino acids at the amino terminal. Our results suggest that, like V. cholerae hemolysin, two‐step processing also exists in V. mimicus hemolysin.
Domain‐Specific Rearrangement between the Two Arg‐Gingipain‐Encoding Genes in Porphyromonas gingivalis: Possible Involvement of Nonreciprocal Recombination
Authors: Nakayama Koji
Published: November 2013

Porphyromonas gingivalis has two functional genes (rgpA and rgpB) encoding an arginine‐specific cysteine proteinase (Arg‐gingipain, RGP). Several RGP‐encoding genes have been cloned and sequenced from various P. gingivalis strains, but all of the genes seem to be essentially equivalent to rgpA. In this study, we cloned and sequenced the second rgp gene (rgpB). A comparison of the rgpB gene and the rgp1 gene, one of the rgpA‐equivalent genes, revealed that their gene structures were very similar to each other, except that the rgpB gene did not possess most of the hemagglutinin domain present in the C‐terminal region of the rgpl gene, and provided strong evidence for homologous recombination between the proteinase domain regions of the two rgp genes. The presence of nonreciprocal recombination in P. gingivalis was experimentally proven using suicide/integration plasmid systems. The results provide one of the hypothetical scenarios of the generation of the two rgp genes; that is, they have been generated through the duplication of an ancestor rgp gene, insertion of the hemagglutinin domain region into one copy of the two resulting rgp genes (or deletion of the region from one rgp) and homologous recombination between the proteinase domain regions of the two rgp genes.
Supernatant from Cultured Intestinal Cells Inhibits the Growth of Gram‐Negative Bacteria
Authors: Kops Sandra Kraeuter , Chao David , West A. Brian
Published: November 2013

Bacterial growth chambers of Transwell units bearing intestinal epithelial monolayers (C2BBe) was consistently observed to be stagnant during the course of transmigration studies with Salmonella typhi. This limitation could not be explained by varying the bacterial load in the inoculum. Conditioned media produced by cultured C2BBe cells would not support bacterial growth. Growth support of the media was restored by heating to 95 C for 10 min. C2BBe conditioned media had bacteriostatic activity for a large variety of gram‐negative, enteropathogenic bacteria but had no effect on gram‐positive bacteria.
Infection of an HIV‐1/SIVmac Chimeric Virus Having HIV‐1 env to Macaque Monkeys via the Vaginal Cavity
Authors: Ido Eiji , Okada Mineyuki , Enose Yoshimi , Kuwata Takeo , Igarashi Tatsuhiko , Sata Tetsutaro , Terao Keiji , Chen Jiangli , Hayami Masanori
Published: November 2013

We previously reported that an HIV‐1/SIVmac chimeric virus (designated as NM‐3rN) having HIV‐1 env efficiently infected macaque monkeys by intravenous inoculation. In this study, this chimeric virus was atraumatically inoculated into the vaginal cavity of two rhesus and one cynomolgus monkeys. Although antibody response and detection of proviral genome by PCR were observed in both rhesus monkeys, virus recovery was only once from PBMC in one of them. In the cynomolgus monkey, no virus was recovered and proviral DNA detection was rare. Thus, vaginal inoculation with NM‐3rN resulted in poor systemic infection, implying the presence of selective pressure while passing through mucosal membranes.
Presence of IgM Antibodies Which Sensitize HIV‐1‐Infected Cells to Cytolysis by Homologous Complement in Long‐Term Survivors of HIV Infection
Authors: Okada Noriko , Wu Xiaoshan , Okada Hidechika
Published: November 2013

Although human cells are resistant to homologous human complement due to the presence of species‐specific membrane inhibitors, a naturally occurring IgM antibody which recognizes an asialo‐oligosaccharide can sensitize HIV‐1‐infected cells for complement‐mediated cytolysis. Therefore, we investigated whether long‐term survivors of HIV‐1 infection harbor such antibodies in their sera. Thirty of 31 sera from HIV‐1 seropositive hemophilia patients who have survived HIV‐1 infection 10 years or more showed appreciable cytolytic activity, while only 2 sera of 10 seropositive patients presumed to have been infected with HIV‐1 (due to sexual contact) more recently showed cytolytic activity. On the other hand, only 7 out of 43 sera from seronegative hemophilia patients showed cytolytic activity. Immunofluorescence staining for IgM on HIV‐L‐infected cells essentially correlated with the cytolytic capacity of the sera. Therefore, naturally occurring IgM antibodies and/or generated IgM antibodies reactive with the HIV‐L‐infected cells in patients might have been responsible for long‐term survival due to complement‐mediated immune cytolysis which may, in conjunction with cytotoxic T lymphocytes, synergistically suppress the infected cells in vivo. Therefore, the transfusion of such IgM antibodies could be effective for the treatment of HIV‐L‐infected individuals.
Cytokines Released by Human Peripheral Blood Mononuclear Cells Inhibit the Production of Early and Late Cytomegalovirus Proteins
Authors: Torigoe Sadayoshi , Campbell Donald E. , Starr Stuart E.
Published: November 2013

Cytomegalovirus‐infected human fibroblasts are susceptible to lysis by natural killer cells and cytotoxic T cells. The purpose of this study was to determine whether non‐lytic mechanisms might also contribute to the control of cytomegalovirus infection. The appearance of cytomegalovirus proteins in infected fibroblasts was determined by flow cytometry. Infected fibroblasts incubated with peripheral blood mononuclear cells for 3 days expressed less early and late proteins than fibroblasts incubated without peripheral blood mononuclear cells. Supernatants generated by the cocultivation of peripheral blood mononuclear cells with cytomegalovirus‐infected fibroblasts inhibited the production of cytomegalovirus early and late proteins. The soluble factors in supernatants which contributed to the inhibitory effect were identified as interferons α, β and γ, and tumor necrosis factors α and β. The ability of supernatants to inhibit the production of cytomegalovirus early protein was mimicked by combinations of corresponding recombinant cytokines. The inhibition of cytomegalovirus protein production by cytokines produced by peripheral blood mononuclear cells may contribute to early containment of cytomegalovirus infection.
Herpes Simplex Virus‐Specific IgM, IgA and IgG Subclass Antibody Responses in Primary and Nonprimary Genital Herpes Patients
Authors: Hashido Madoka , Kawana Takashi
Published: November 2013

To clarify the humoral immunity in herpes simplex virus (HSV) infection, HSV‐specific IgM, IgA and IgG subclass antibody responses were studied in patients with genital herpes: 17 primary, 13 recurrent and 6 nonprimary first episode. A total of 181 serum samples serially collected from the patients, 5 per patient until 213 days after the onset of disease (on average), were analyzed by an enzyme‐linked immunosorbent assay. IgGl, IgG3 and IgA were detected in all patients with primary and nonprimary infections, whereas IgG4 was detected in 74% of only those with nonprimary infections and IgG2 was detected in none. IgM was detected in 100% of the patients with primary infections, but also in 68% of those with nonprimary infections. IgA showed a peak similar to that of IgM in patients with primary infections. No significant difference was observed in the detection rate or pattern of antibody responses between the recurrent and nonprimary first episode infections, nor between the HSV‐1 and HSV‐2 infections. These findings may be useful to improve the diagnostic potential of HSV serology.
Isolation of a New Orientia tsutsugamushi Serotype
Authors: Seong Seung‐yong , Park Sae‐gwang , Kim Hang‐rae , Han Tae‐hee , Kang Jae‐seung , Choi Myung‐sik , Kim Ik‐sang , Chang Woo‐hyun
Published: November 2013

Orientia tsutsugamushi, the etiological agent of scrub typhus, is an antigenically diverse organism and many serologically distinct strains have been identified. The 56 kDa protein of O. tsutsugamushi, a major protein in the outer membrane, has been thought to be responsible for this antigenic variability. A strain of O. tsutsugamushi isolated in Korea cross‐reacted with both Gilliam strain‐specific and Karp strain‐specific monoclonal antibodies. When its 56 kDa protein gene was cloned and analyzed, its sequence showed variation especially between 1,200 and 1,250 bp, showing that this isolate is a new O. tsutsugamushi strain.