bluuebunny/crossref_metadata_2025 huggingface dataset:
IgE Production after Four Routes of Injections of Japanese Cedar Pollen Allergen without Adjuvant: Crucial Role of Resident Cells at Intraperitoneal or Intranasal Injection Site in the Production of Specific IgE toward the Allergen
Authors: Sakurai Kanji , Takenaka Hiroshi , Yoneda Yukio , Tashiro‐Yamaji Junko , Yamamoto Yumiko , Lee Koutetsu , Yamaguchi Satoko , Miyoshi Masako , Kubota Takahiro , Yoshida Ryotaro
Published: November 2013

The production of specific IgE antibodies directed toward cedar pollen correlates well with the onset of allergic rhinitis; but the mechanisms of allergen recognition as nonself and Ig class switch to IgE by the immune system are still not fully understood. In the present study, we injected cedar pollen into mice through 4 different routes (intranasal (i.n.), intraperitoneal (i.p.), intravenous (i.v.), and subcutaneous (s.c.)) without adjuvant 1 to 3 times, and determined time‐dependent changes in the total and specific serum IgE levels compared with those in the serum levels of other isotype Igs. After an i.p. or i.n. injection of allergen into the mice, they produced a 1.5‐ to 1.7‐fold increase in total IgE, but none in IgG, IgM, or IgA antibodies in their serum, whereas an i.v. or s.c. injection of allergen was inactive as an inducer of total IgE antibodies. Upon a 2nd (s.c.) injection of the allergen into the i.p. or i.n. sensitized mice, a large amount of allergen‐specific IgE antibodies was found in the serum. In the case of i.v. or s.c. sensitized mice, however, they produced total, but not specific, IgE antibodies; and a 3rd (s.c.) injection of the allergen resulted in a large amount of specific IgE antibodies in the serum. These results imply that resident cells at the i.p. or i.n. injection site may play a crucial role in the efficient production of total and specific IgE antibodies toward the allergen.
The Cytotoxin‐Hemolysin Genes of Human and Eel Pathogenic Vibrio vulnificus Strains: Comparison of Nucleotide Sequences and Application to the Genetic Grouping
Authors: Senoh Mitsutoshi , Miyoshi Shin‐ichi , Okamoto Keinosuke , Fouz Belen , Amaro Carmen , Shinoda Sumio
Published: November 2013

Vibrio vulnificus can be divided into two groups on the basis of pathogenesis. Group 1 is pathogenic only to humans, whereas group 2 is pathogenic to eels and occasionally to humans. Although both groups produce a 50‐kDa cytotoxin‐hemolysin (V. vulnificus hemolysin; VVH), the toxins are different. In the present study, the nucleotide sequence of the toxin gene (vvhA) of strain CDC B3547 (a group 2 strain) was determined, and the deduced amino acid sequence was compared to that of strain L‐180 (a group 1 strain). The nucleotide sequence of vvhA of strain CDC B3547 was about 96% identical with that of strain L‐180, which results in a difference of 3 amino acid residues in the C‐terminal lectin domain of VVH. Nevertheless, two primer sets for polymerase chain reaction could be designed to differentiate the toxin gene of each strain. When 27 V. vulnificus clinical isolates were tested, group 1 strains (9 strains) were shown to react only to the primers designed for vvhA of strain L‐180; whereas, the gene of group 2 strains (18 strains) could be amplified with the primers for vvhA of strain CDC B3547. These findings may lead to development of a novel genetic grouping system related to the virulence potential or to the host range.
A Quantitative Analysis of Cedar Pollen‐Specific Immunoglobulins in Nasal Lavage Supported the Local Production of Specific IgE, Not of Specific IgG
Authors: Yoshida Tomoaki , Usui Ayako , Kusumi Taeko , Inafuku Shigeru , Sugiyama Tsuyoshi , Koide Naoki , Yokochi Takashi
Published: November 2013

Many studies have proved the relevance of local immune responses, rather than systemic immunity, to the pathogenesis of allergic rhinitis. Indeed, allergen‐specific B lymphocyte undergoes class switching to IgE in situ. However, the relative contribution of in situ production to the amount in serum is still ambiguous. Here, a quantitative comparison of the local concentration of allergen‐specific IgE with the systemic concentration was explored for the estimation. Among seasonal rhinitis patients, total and Japanese cedar pollen (JCP)‐specific IgE, IgA and IgG antibodies were quantified in nasal lavage fluid (NLF) and serum with the time‐resolved fluorescence immunosorbent assay. Although the total amounts of IgE and IgG classes in the NLF, which were apparently passive discharge from the mucosal tissue, were smaller and variable, the relative proportions of JCP‐specific antibodies could be quantitatively compared between NLF and serum or between subjects. The proportions of specific IgE in the NLF were remarkably higher than in serum (average 13.2‐fold) in most subjects, which strongly supported the predominant in situ production of the specific IgE and subsequent dilutions in the systemic circulations. Similar but smaller values were obtained for IgA (average 3.7‐fold). In contrast, the specific proportions of IgG in the NLF were surprisingly consistent with serum (average 1.0‐fold), suggesting that the specific IgG was mostly produced in the downstream lymphoid organs. The local productions of specific IgE would encourage the topical therapies and the usage of the NLF for the diagnosis of allergic rhinitis.
Characterization of the Genus Bifidobacterium by Automated Ribotyping and 16S rRNA Gene Sequences
Authors: Sakata Shinji , Ryu Chun Sun , Kitahara Maki , Sakamoto Mitsuo , Hayashi Hidenori , Fukuyama Masafumi , Benno Yoshimi
Published: November 2013

In order to characterize the genus Bifidobacterium, ribopatterns and approximately 500 bp (Escherichia coli positions 27 to 520) of 16S rRNA gene sequences of 28 type strains and 64 reference strains of the genus Bifidobacterium were determined. Ribopatterns obtained from Bifidobacterium strains were divided into nine clusters (clusters I–IX) with a similarity of 60%. Cluster V, containing 17 species, was further subdivided into 22 subclusters with a similarity of 90%. In the genus Bifidobacterium, four groups were shown according to Miyake et al.: (i) the Bifidobacterium longum infantis‐longum‐suis type group, (ii) the B. catenulatum‐pseudocatenulatum group, (iii) the B. gallinarum‐saeculare‐pullorum group, and (iv) the B. coryneforme‐indicum group, which showed higher than 97% similarity of the 16S rRNA gene sequences in each group. Using ribotyping analysis, unique ribopatterns were obtained from these species, and they could be separated by cluster analysis. Ribopatterns of six B. adolescentis strains were separated into different clusters, and also showed diversity in 16S rRNA gene sequences. B. adolescentis consisted of heterogeneous strains. The nine strains of B. pseudolongum subsp. pseudolongum were divided into five subclusters. Each type strain of B. pseudolongum subsp. pseudolongum and B. pseudolongum subsp. globosum and two intermediate groups, which were suggested by Yaeshima et al., consisted of individual clusters. B. animalis subsp. animalis and B. animalis subsp. lactis could not be separated by ribotyping using EcoRI. We conclude that ribotyping is able to provide another characteristic of Bifidobacterium strains in addition to 16S rRNA gene sequence phylogenetic analysis, and this information suggests that ribotyping analysis is a useful tool for the characterization of Bifidobacterium species in combination with other techniques for taxonomic characterization.
Genotype Analysis of the Variable Internal Repeat Region in the rRNA Gene of Trichophyton tonsurans Isolated from Japanese Judo Practitioners
Authors: Sugita Takashi , Shiraki Yumi , Hiruma Masataro
Published: November 2013

Tinea capitis due to Trichophyton tonsurans is currently epidemic among Japanese Judo practitioners. T. tonsurans has seven genotypes in a variable internal repeat (VIR) region of the rRNA gene. All 101 isolates obtained from Japanese Judo practitioners had the identical genotype. This suggests that a specific genotype strain occurs throughout Japan.
Genetic Characterization of Escherichia coli O157:H7/– Strains Carrying the stx2 Gene but Not Producing Shiga Toxin 2
Authors: Koitabashi Tsutomu , Vuddhakul Varaporn , Radu Son , Morigaki Tadaaki , Asai Norio , Nakaguchi Yoshitsugu , Nishibuchi Mitsuaki
Published: November 2013

Nine Escherichia coli O157:H7/– strains isolated primarily from non‐clinical sources in Thailand and Japan carried the stx2 gene but did not produce Stx2 toxin in a reversed passive latex agglutination (RPLA) assay. A strain (EDL933) bearing a stx2 phage (933W) was compared to a strain (Thai‐12) that was Stx2‐negative but contained the stx2 gene. To study the lack of Stx2 production, the Thai‐12 stx2 gene and its upstream nucleotide sequence were analyzed. The Thai‐12 stx2 coding region was intact and Stx2 was expressed from a cloned stx2 gene using a plasmid vector and detected using RPLA. A lacZ fusion analysis found the Thai‐12 stx2 promoter non‐functional. Because the stx2 gene is downstream of the late promoter in the stx2 phage genome, the antitermination activity of Q protein is essential for strong stx2 transcription. Thai‐12 had the q gene highly homologous to that of Φ21 phage but not to the 933W phage. High‐level expression of exogenous q genes demonstrated Q antitermination activity was weak in Thai‐12. Replication of stx2 phage was not observed in Stx2‐negative strains. The q‐stx2 gene sequence of Thai‐12 was well conserved in all Stx2‐negative strains. A PCR assay to detect the Thai‐12 q‐stx2 sequence demonstrated that 30% of O157 strains from marketed Malaysian beef carried this sequence and they produced little or no Stx2. These results suggest that stx2‐positive O157 strains that produce little or no Stx2 may be widely distributed in the Asian environment.
Epithelial Cells Secrete Interleukin‐8 in Response to Adhesion and Invasion of Diffusely Adhering Escherichia coli Lacking Afa/Dr Genes
Authors: Meraz Ismail Mustafa , Arikawa Kentaro , Ogasawara Jun , Hase Atsushi , Nishikawa Yoshikazu
Published: November 2013

Escherichia coli that sparsely adhere to human epithelial cells are known as diffusely adherent E. coli (DAEC), and the role of the Afa/Dr family of adhesins is now understood. Strains that do not possess Afa/Dr, however, comprise another group of DAEC, of which the pathogenicity remains unknown. The ability to induce interleukin‐8 (IL‐8) secretion from intestinal epithelial cells might be a feature of enterovirulent bacteria. We previously found that some Afa/Dr DAEC strains induce IL‐8 by stimulating epithelial cells with flagella. The present study examines whether non‐Afa/Dr DAEC can induce IL‐8 in epithelial cells (HEp‐2, INT407, and T84). Among 21 strains, 11 (52%; 11/21) induced as much IL‐8 as high inducer strains of Afa/Dr DAEC. Adhesion did not significantly differ between high and low inducers; therefore diffuse adhesion alone is probably insufficient to induce IL‐8. It was shown that IL‐8 induction and the number of intracellular bacteria directly correlated. Wortmannin, an inhibitor of the phosphatidylinositol‐3‐phosphate kinase, reduced both intracellular bacteria and IL‐8 secretion. Motile strains were significantly more prevalent among high (10/11) than low (4/10) inducers. However, 4 low invasive strains hardly induced IL‐8 despite their motility. In conclusion, some non‐Afa/Dr DAEC invoke the induction of high levels of inflammatory cytokines. Unlike Afa/Dr DAEC, however, non‐Afa/Dr strains may require invasion to cause strong induction. These non‐Afa/Dr high inducers can be enteropathogenic for the cytokine‐inducing properties.
Fusion of Doppel to Octapeptide Repeat and N‐Terminal Half of Hydrophobic Region of Prion Protein Confers Resistance to Serum Deprivation
Authors: Lee Deug‐chan , Sakudo Akikazu , Kim Chi‐kyeong , Nishimura Takuya , Saeki Keiichi , Matsumoto Yoshitsugu , Yokoyama Takashi , Chen Shu G. , Itohara Shigeyoshi , Onodera Takashi
Published: November 2013

Our previous studies have shown an essential role played by the octapeptide repeat region (OR) and the N‐terminal half of hydrophobic region (HR) in the anti‐apoptotic activity of prion protein (PrP). As PrP‐like protein Doppel (Dpl), which structurally resembles an N‐terminally truncated PrP, did not show any anti‐apoptotic activity, we examined apoptosis of HpL3–4 cells expressing Dpl fused to various lengths of the N‐terminal region of PrP to investigate whether the PrP/Dpl fusion proteins retain anti‐apoptotic function. HpL3–4 cells expressing Dpl fused to PrP(1–124) with the OR and N‐terminal half of HR of PrP showed anti‐apoptotic function, whereas Dpl fused to PrP(1–95) with OR did not rescue cells from apoptotic cell death induced by serum deprivation. These results indicate that the OR and N‐terminal half of HR of PrP retains anti‐apoptotic activity similar to full‐length PrP.
Mosquito Cells Infected with Japanese Encephalitis Virus Release Slowly‐Sedimenting Hemagglutinin Particles in Association with Intracellular Formation of Smooth Membrane Structures
Authors: Ishikawa Tomohiro , Konishi Eiji
Published: November 2013

Arthropod‐borne flaviviruses can grow in both arthropod and mammalian cells. Virion morphogenesis, though well studied in mammalian cells, is still unclear in arthropod cells. Here, we compared a mosquito cell line C6/36 and a mammalian cell line Vero in extracellular virus particles and intracellular ultrastructures triggered by infection with Japanese encephalitis virus (JEV). Sedimentation analyses of virion and slowly‐sedimenting hemagglutinin (SHA) particles released by infection with the Nakayama strain revealed that C6/36 cells produced higher envelope (E) antigen levels in the SHA than the virion fraction in contrast to Vero cells that showed the opposite pattern. Specific infectivities per ng of E were similar in both cells, whereas specific hemagglutinating activities in the SHA fraction were lower in C6/36 than Vero cells. The precursor membrane protein was less efficiently cleaved to the membrane protein in SHA particles released from C6/36 than Vero cells. Ultrastructural studies showed more remarkable production of smooth membrane structures (SMSs) in C6/36 than in Vero cells. The differences in sedimentation patterns of extracellular virus particles between Nakayama‐infected C6/36 and Vero cells were consistently observed in 5 other strains (Beijing P1, Beijing P3, JaTH‐160, KE‐093 and JaGAr‐O1), except for KE‐093‐infected C6/36 cells which exhibited the Vero‐type sedimentation profile under conditions of open cultivation. By electron microscopy, the production of SMSs from KE‐093‐infected C6/36 cells under open conditions was markedly less than that under closed conditions where the cells exhibited the C6/36‐type sedimentation profile. Thus, intracellular SMS formations were associated with extracellular SHA production in JEV‐infected mosquito cells.
A Legionellosis Case Due to Contaminated Spa Water and Confirmed by Genomic Identification in Taiwan
Authors: Su Hsun‐Pi , Tseng Lei‐Ron , Tzeng Shin‐Chan , Chou Chen‐Ying , Chung Tung‐Ching
Published: November 2013

Tracing the source of a legionellosis (LG) case revealed that the Legionella pneumophila (LP) strain isolated from patient's sputum shared the same serogroup (SG) and PFGE‐type with 4 LP strains obtained from a spa center. With a high LP‐contamination rate (81.2%, 13/16) in all of its 16 basins, this spa center was also found to have a multi‐genotypic distribution among its 13 LP isolates, which can be categorized into 5 PFGE‐types. Despite such a serious contamination in the spa center, which usually had ca. 100 visitors per day, this male patient, bearing LG‐risk factors of long‐term heavy smoking and alcoholism, was the only case identifiable after an active investigation. To explore the possible reason for this sporadic infection, all 5 PFGE‐types of LP isolated were assayed for their presence of two important virulent genes (lvh and rtxA) and were identified as either less‐virulent (lvh+, rtxA–) or non‐virulent (lvh–, rtxA–) types. The strong virulent type (lvh+, rtxA+) usually seen in clinical strains elsewhere was not found here. Moreover, the LG‐causative type in this infection was the only one to be classified as the less‐virulent type, with the presence of lvh gene indicating its relatively more virulent potential than other 4 PFGE‐types. Accordingly, mutual interaction between LP's virulent potential and patient's health‐status was suggested to be the force directing the opportunistic infection of this sporadic case. This is the first spa‐associated infection caused by SG 2 of LP.
Vaccination with Live Escherichia coli Expressing Brucella abortus Cu/Zn Superoxide‐Dismutase
Authors: Andrews Edilia , Salgado Paulina , Folch Hugo , Oñate Angel
Published: November 2013

Previously we reported that immunization with Escherichia coli DH5α‐expressing Brucella abortus Cu/Zn superoxide dismutase [E. coli (pBSSOD)] induces a protective immune response in BALB/c mice. Here we studied the type of immune defense that the recombinant E. coli induces in mice using as our experimental model Brucella superoxide dismutase Cu/Zn presented by J744.A1 to sensitized lymphocytes as the target of specific lysis or as cytokine inductors. The results indicate that E. coli carrying the Cu/Zn gene was able to induce specific cytotoxic T cells, mainly from CD8+ subpopulation and IFN‐γ‐producing cells belonging in their vast majority to the CD4+ subpopulation.
Candida pseudohaemulonii Sp. Nov., an Amphotericin B‐ and Azole‐Resistant Yeast Species, Isolated from the Blood of a Patient from Thailand
Authors: Sugita Takashi , Takashima Masako , Poonwan Natteewan , Mekha Nanthawan
Published: November 2013

Candida haemulonii (types I and II) is rarely isolated from clinical specimens. We isolated a strain that is phylogenetically close to C. haemulonii from the blood of a Thai patient, and named it C. pseudohaemulonii sp. nov. (CBS 10099T=JCM 12453T=DMST 17134T). The new species and C. haemulonii types I and II were resistant to amphotericin B and azole agents but were susceptible to a 1,3‐β‐D‐glucan synthetase inhibitor, micafungin, and 5‐flucytosine. The species were easily distinguished using an ID32 yeast identification kit. The taxonomic description of C. pseudohaemulonii sp. nov. is presented.
Regulation of NF‐κB Signaling by Decoy Oligodeoxynucleotides
Authors: Isomura Iwao , Morita Akimichi
Published: November 2013

The nuclear factor‐κB (NF‐κB) plays a critical role in regulating expression of genes responsible for a wide range of cellular processes, including innate and adoptive immune responses, and pathways related to cell survival and proliferation. Based on its property as a transcription factor, NF‐κB has been considered as a good target for the treatment of inflammatory immune diseases. To regulate gene expression, “decoy” oligodeoxynucleotides (ODNs) are ideal tools to interfere with binding of transcription factors to promoter regions in genes. Herein we review the application of NF‐κB decoy ODNs to control inflammatory disorders and transplantation tolerance. In addition, we provide information about induction of antigen‐specific peripheral tolerance by their topical application. Regulation of NF‐κB signaling by NF‐κB decoy ODNs might be an effective tool to control a variety of disorders caused by inflammatory immune responses.
Transcriptional Downregulator HexS Controlling Prodigiosin and Serrawettin W1 Biosynthesis in Serratia marcescens
Authors: Tanikawa Taichiro , Nakagawa Yoji , Matsuyama Tohey
Published: November 2013

Serratia marcescens has been known as a temperature‐dependent producer of two chemically different exolipids (red pigment prodigiosin and biosurfactant serrawettin W1) in parallel. During genetic investigation of such control mechanisms, mini‐Tn5 insertional mutant Tan1 overproducing these exolipids was isolated. The gene concerning such disregulation was identified as hexS by DNA cloning followed by sequencing and homology analysis of the presumed product with 314 amino‐acids. The product HexS was the homologue of HexA of Erwinia carotovora ssp. carotovora and classified as a transcriptional regulator belonging to LysR family. By RT‐PCR analysis, the hexS mutant was shown to over‐transcribe the pigA gene (the first gene of the pig cluster involved in prodigiosin synthesis) and the swrW gene encoding serrawettin W1 synthetase belonging to the nonribosomal peptide synthetase family. In contrast, transcription of the pswP gene encoding phosphopantetheinyl transferase in Tan1 was in the level of parent strain 274. Purified protein encoded in his6‐hexS demonstrated binding activity to DNA fragments of the upstream region of pigA and swrW genes and not to that of the pswP gene. S. marcescens strain 274 transformed with a low‐copy plasmid carrying hexS demonstrated reduced production of prodigiosin and serrawettin W1, and reduced activity of exoenzymes (protease, chitinase, and DNase) except phospholipase C. Possible generation of virulent S. marcescens by derepression or mutation of the hexS gene in infected tissues or ex vivo environments was suggested.
Peripheral Blood Mononuclear Cells Immunophenotyping in Pulmonary Tuberculosis Patients before and after Treatment
Authors: Barcelos Warly , Martins‐Filho Olindo Assis , Guimarães Tânia Mara Pinto Dabés , Oliveira Márcio Hamilton Prostzner , Spíndola‐de‐Miranda Silvana , Carvalho Beatriz Nascimento , de Toledo Vicente de Paulo Coelho Peixoto
Published: November 2013

Tuberculosis (TB) is a lung disease caused by Mycobacterium tuberculosis. The interaction between the bacillus and the host may lead to a protective cellular immune response. In the present study, we propose the “in vitro” evaluation of this cellular immune response in patients with tuberculosis before and after chemotherapic treatment. Eleven patients with TB and 9 asymptomatic subjects with tuberculin skin test negative (TST–) (purified protein derivative (PPD) ≤10 mm) were evaluated. The peripheral lymphocytes of the subjects were analyzed utilizing the following surface markers: CD3+, CD4+, CD8+, CD19+, CD25+, CD56+, CD14+, CD16+ and HLA‐DR+. At the end of the treatment, symptomatic patients presented a significant predominance (P<0.05) of CD4+ lymphocytes, a significant decrease (P<0.05) in activated CD8+ T cells and a significant increase (P<0.05) in the marker CD19+. A predominance of mature NK cells and a significant decrease (P<0.05) in NKT cells were observed. Also observed was a trend toward decrease in immunoregulatory T cells and a predominance of pro‐inflammatory macrophages. TST– subjects presented a predominance of CD4+ over CD8+ and predominance of CD19+ and of mature NK cells in comparison to the group of patients, both before and after treatment. Thus, several cell types, such as CD4+ and CD8+ T cells, NK cells and their subpopulations, NKT cells and pro‐inflammatory macrophages could act in a synergic way to control the growth and multiplication of M. tuberculosis.
Preliminary Human Study for Possible Alteration of Serum Immunoglobulin E Production in Perennial Allergic Rhinitis with Fermented Milk Prepared with Lactobacillus gasseri TMC0356
Authors: Morita Hirotsugu , He Fang , Kawase Manabu , Kubota Akira , Hiramatsu Masaru , Kurisaki Jun‐Ichi , Salminen Seppo
Published: November 2013

The fermented milk prepared with Lactobacillus gasseri TMC0356 was administered at 200 ml per day for 4 weeks to 15 subjects with high serum IgE levels and perennial allergic rhinitis. The serum total IgE concentration was significantly reduced after 28 days' exposure to the fermented milk (P<0.05) compared to that before the intervention. The serum IgE specific to Acari and those to Japanese cedar pollen also significantly declined (P<0.05). T helper 1 (Th1) cells in the composition of their peripheral blood mononuclear cells (PBMCs) significantly increased after 14 days (P<0.01) and after 28 days (P<0.05). These results suggest that the fermented milk prepared with L. gasseri TMC0356 may alter serum IgE concentration, at least partly by enhancement of Th1 immune responses of the subjects with high concentration of serum IgE. However, further studies are still necessary to know the underlying mechanisms by which the tested fermented milk could influence the host immunity.
A 4‐Aminofurazan Derivative—A189—Inhibits Assembly of Bacterial Cell Division Protein FtsZ In Vitro and In Vivo
Authors: Ito Hideaki , Ura Atsushi , Oyamada Yoshihiro , Tanitame Akihiko , Yoshida Hiroaki , Yamada Sakuo , Wachi Masaaki , Yamagishi Jun‐ichi
Published: November 2013

Out of 95,000 commercially available chemical compounds screened by the anucleate cell blue assay, 138 selected hit compounds were further screened. As a result, A189, a 4‐aminofurazan derivative was found to inhibit FtsZ GTPase with an IC50 of 80 μg/ml and to exhibit antibacterial activity against Staphylococcus aureus and Escherichia coli. Light scattering demonstrated that A189 inhibited FtsZ assembly in vitro, and microscopic observation of A189‐treated E. coli indicated that A189 perturbed FtsZ ring formation and made bacterial cells filamentous. However, nucleoids staining with DAPI revealed that A189 did not affect DNA replication and chromosome segregation in bacterial filamentous cells. Furthermore, A189 made sulA‐deleted E. coli cells filamentous. Taken together, these findings suggest that A189 inhibits FtsZ GTPase activity, resulting in perturbation of FtsZ ring formation, which leads to bacterial cell death.
Detection of Enteroviruses and Mammalian Reoviruses in Korean Environmental Waters
Authors: Kim Hee Jung , Shin Young Oh , Kim Sang Hyun
Published: November 2013

Using the standard total culturable virus assay‐most probable number (TCVA‐MPN) method, we evaluated a total of 348 samples, including surface water, finished water, and tap water samples, collected from randomly selected water treatment plants in Korea from August 2001 through July 2005 according to the Information Collection Rule. All the TCVA‐positive samples were also subjected to integrated cell culture‐PCR (ICC‐PCR) methods for the detection of enteroviruses, hepatitis A virus, adenoviruses, and reoviruses. The most probable number of infectious units per 100 liters for the environmental water samples ranged from 0.5 to 47.3. Nine of the 13 TCVA‐positive samples (69.2%) were found by ICC‐PCR to be positive for human enteroviruses, which were confirmed to be coxsackievirus type B3, coxsackievirus type B4, coxsackievirus type B6, echovirus type 30, and vaccine strain poliovirus type 3 by direct sequencing. Eleven of the 13 TCVA‐positive samples (84.6%) were found by ICC‐PCR assay to be positive for reoviruses. The serotype of all the reoviruses was the same as reovirus type 1 by direct sequencing. Both enteroviruses and reoviruses were concurrently detected in seven TCVA‐positive samples (53.8%).
Human Herpesvirus‐6‐Specific Interleukin 10‐Producing CD4+ T Cells Suppress the CD4+ T‐Cell Response in Infected Individuals
Authors: Wang Fang , Yao Kun , Yin Quan‐Zhang , Zhou Feng , Ding Chuan‐Lin , Peng Guang‐Yong , Xu Jian , Chen Yun , Feng Dong‐Ju , Ma Chun‐Lin , Xu Wen‐Rong
Published: November 2013

Human herpesvirus‐6 (HHV‐6) infection normally persists for the lifetime of the host and may reactivate with immunosuppression. The mechanism behind HHV‐6 latent infection is still not fully understood. In this study, we observed that decreased proliferation of CD4+ T cells and PBMCs but not CD8+ T cells from HHV‐6‐infected individuals was stimulated with HHV‐6‐infected cell lysates. Moreover, HHV‐6‐stimulated CD4+ T cells from HHV‐6‐infected individuals have suppressive activity on naïve CD4+ T and CD8+ T cells from HHV‐6‐uninfected individuals. However, no increased proportion of CD4+ CD25+ Treg cells from HHV‐6‐infected individuals contributed to the suppressive activity of the HHV‐6‐stimulated CD4+ T cells from HHV‐6‐infected individuals. Transwell experiments, ELISA and anti‐IL‐10 antibody blocking experiment demonstrated that IL‐10 may be the suppressive cytokine required for suppressive activity of CD4+ T cells from HHV‐6‐infected individuals. Results of intracellular interleukin (IL)‐10 and IL‐4 further implicated the HHV‐6‐speciflc IL‐10‐producing CD4+ T cells in the suppressive activity of CD4+ T cells from HHV‐6‐infected individuals. Results of intracellular interferon (IFN)‐γ demonstrated a decreased frequency of HHV‐6‐speciflc IFN‐γ‐producing CD4+ T, but not CD8+ T cells in HHV‐6‐infected individuals, indicating that it was the CD4+ Th1 responses in HHV‐6‐infected individuals that were selectively impaired. Our findings indicated that HHV‐6‐specific IL‐10‐producing CD4+ T cells from HHV‐6‐infected individuals possess T regulatory type 1 cell activity: immunosuppression, high levels of IL‐10 production, with a few cells expressing IFN‐γ, but none expressing IL‐4. These cells may play an important role in latent HHV‐6 infection.
Human Milk Glycoproteins Inhibit the Adherence of Salmonella typhimurium to HeLa Cells
Authors: Bessler Heidi Christina , de Oliveira Inaiara Rosa , Giugliano Loreny Gimenes
Published: November 2013

The ability of human milk, as well as its protein fractions, to inhibit the adhesion and invasion of Salmonella typhimurium to HeLa cells was investigated. The results revealed that milk secretory immunoglobulin A (sIgA) inhibited neither the adherence nor the bacterial invasion; however, free secretory component and lactoferrin inhibited the bacterial adhesion and interacted with several bacterial proteins. Our data indicated that glycoproteins such as free secretory component and lactoferrin could act as protective compounds against infant enteric diseases, possibly binding to bacterial surface and blocking adhesion, the primordial step of S. typhimurium infection.
Autoimmune Regulator (AIRE) Gene Is Expressed in Human Activated CD4+ T‐Cells and Regulated by Mitogen‐Activated Protein Kinase Pathway
Authors: Nagafuchi Seiho , Katsuta Hitoshi , Koyanagi‐Katsuta Rimiko , Yamasaki Satoshi , Inoue Yasushi , Shimoda Kazuya , Ikeda Yukari , Shindo Mieko , Yoshida Eiko , Matsuo Tomohito , Ohno Yuju , Kogawa Kazuhiko , Anzai Keizo , Kurisaki Hironori , Kudoh Jun , Harada Mine , Shimizu Nobuyoshi
Published: November 2013

The autoimmune regulator (AIRE) gene is a gene responsible for autoimmune polyendocrinopathy‐candidiasis‐ectodermal dystrophy. Here we show that AIRE is expressed in human peripheral CD4‐positive T‐cells, and most highly in antigen‐ and interleukin 2‐stimulated T (IL‐2T) cells. Mitogen‐activated protein kinases (MAPKs), including MAPK kinase (MEK) 1/2 and p38 MAPK, were phosphorylated in IL‐2T cells and the expression of the AIRE gene was inhibited by a specific p38 MAPK inhibitor (SB203580), thereby indicating that AIRE gene expression is controlled by the MAPK pathway in IL‐2T cells. These data suggested the possible significance of the AIRE gene in the peripheral immune system.
In Vitro Infection of Immortalized Primary Hepatocytes by HCV Genotype 4a and Inhibition of Virus Replication by Cyclosporin
Authors: El‐Farrash Mohamed A. , Aly Hussein H. , Watashi Koichi , Hijikata Makoto , Egawa Hiroto , Shimotohno Kunitada
Published: November 2013

Hepatitis C virus (HCV) is a major cause of liver cirrhosis and hepatocellular carcinoma worldwide. We previously reported that cyclosporin A (CsA) inhibits HCV‐1b replication. However, its inhibition of JFH‐1 (HCV‐2a) was much less. Since HCV genotype clearly affects the in vitro and in vivo response to anti‐viral therapy, we wished to examine the effect of CsA and its non‐immunosuppressive derivative NIM811 on HCV genotype 4a replication. We first established an in vitro system supporting HCV‐4a infection and replication using immortalized human hepatocytes, HuS‐E7/DN24 (HuS) cells, and these cells were infected with sera obtained from Egyptian patients with chronic HCV‐4a infection. HuS cells supported more robust HCV‐4a replication than both HuH‐7.5 and PH5CH8 cells, and HCV‐4a infection and replication were completely inhibited by 3 μg/ml CsA and 0.5 μg/ml NIM811. Thus, HuS cells are a good model system supporting the infection and high‐level replication of HCV‐4a, and both CsA and NIM811 effectively inhibit HCV‐4a replication in this system.
The Relationship between Microbiota and Polyamine Concentration in the Human Intestine: A Pilot Study
Authors: Matsumoto Mitsuharu , Benno Yoshimi
Published: November 2013

The fecal microbiota of 10 hospitalized elderly subjects and 14 healthy adults were analyzed by terminal‐restriction fragment length polymorphism (T‐RFLP) analysis using HhaI, MspI, HaeIII, and AluI, as well as fecal polyamine (PA) concentration. The T‐RFLP profiles of the fecal microbiota of the subjects were roughly divided into 2 clusters—I (9 out of 11 were derived from hospitalized elderly subjects) and II (12 out of 13 were derived from healthy adults). The average concentration of putrescine in Cluster II was 5.8 times higher than that of putrescine in Cluster I (P=0.0015). Using a phylogenetic assignment database for T‐RFLP analysis of human colonic microbiota, the terminal‐restriction fragments (T‐RFs) characteristically detected in the case of subjects with high fecal PA concentration were predicted to be derived from bacterial species and phylotypes belonging to Clostridium subcluster XIVa, particularly including Clostridium xylanolyticum, Clostridium saccharolyticum, the uncultured human intestinal bacterium clone JW1H4 (a relative of Desulfotomaculum guttoideum), Roseburia intestinalis, the uncultured bacterium clone 41F10 (a relative of Eubacterium ramulus), Roseburia cecicola, Ruminococcus obeum and its relatives. From these results, we concluded that fecal microbiota may be linked with fecal PA concentration and that some bacterial species belonging to Clostridium subcluster XIVa may play a major role in the control of intestinal PA concentration in humans.
Comparison of Fecal Microbiota and Polyamine Concentration in Adult Patients with Intractable Atopic Dermatitis and Healthy Adults
Authors: Matsumoto Mitsuharu , Kakizoe Kenji , Benno Yoshimi
Published: November 2013

Fecal microbiota and polyamine concentration obtained from eleven intractable adult‐type atopic dermatitis (AD) patients and thirteen healthy adults were compared. Fecal microbiota were analyzed using terminal‐restriction fragment length polymorphism. The fecal microbiota of volunteers were divided into two clusters, A (n=16) and B (n=8), and the number of AD patients was found to be higher in Cluster B than Cluster A, suggesting that there are relationships between the obstinacy of intractable adult‐type AD and intestinal microbiota in Cluster B. Fecal spermidine concentration in Cluster B were lower than that in Cluster A significantly (P<0.05). Fecal putrescine concentration in Cluster B also tended to be lower than that in Cluster A. Terminal‐restriction fragment (T‐RF) of 122 bp generated by digestion with HhaI, which were predicted as unknown bacteria, were detected characteristically in Cluster A. In contrast, T‐RFs of 368/9 bp generated by digestion with HhaI, which were predicted as Enterobacteriaceae, were detected characteristically in Cluster B. These bacteria are closely associated with intestinal polyamine concentration. These findings raise the possibility that a low concentration of intestinal polyamines produced by intestinal microbiota is one of the important factors in the onset of intractable adult‐type AD.
Expression and Distribution of Very Late Antigen‐5 in Mouse Peritoneal Macrophages upon Ingestion of Fibronectin‐Bound Staphylococcus aureus
Authors: Shinji Hitomi , Kamada Minori , Seki Keiko , Tajima Akiko , Iwase Tadayuki , Masuda Shogo
Published: November 2013

Many pathogens colonize host tissues by binding to the extracellular matrix via their cell surface adhesion molecules, which are called MSCRAMMs (microbial surface components recognizing adhesive matrix molecules). Staphylococcus aureus expresses several of these adhesion molecules, some of which bind to filbronectin. Of these adhesion molecules, filbronectin‐binding proteins play a role in the pathogenicity of S. aureus, although it is not yet clear whether they enhance its virulence. We have previously shown that filbronectin‐bound S. aureus is efficiently phagocytosed by thioglycolate‐induced mouse peritoneal macrophages. Bacterial ingestion is mediated by Very Late Antigen‐5 (VLA‐5; α5β1 integrin) and is accompanied by the formation of adhesion complexes. Here we show that the expression of VLA‐5 is restricted to thioglycolate‐induced inflammatory macrophages and is not found in the resident macrophages. When cells were in suspension, α5 integrin was not expressed on the surface of either resident or inflammatory macrophages, whereas in adherent cells, this integrin was distributed on the surface of inflammatory but not resident macrophages. A high level of this integrin was present in the cytoplasmic region only in inflammatory macrophages. In agreement with this, filbronectin‐mediated phagocytosis of S. aureus was observed only in the inflammatory macrophages. In inflammatory macrophages ingesting fibronectin‐bound S. aureus, α5 integrin was concentrated close to the phagocytosed bacteria. This change in distribution was not found in macrophages ingesting untreated bacteria. Together with our previous work, these results indicate that, upon ingestion of fibronectin‐bound S. aureus, VLA‐5 accumulates in the area of phagocytosis in inflammatory macrophages, where it forms adhesion complexes.
Effect of Interleukin‐10 on the Paracoccidioides brasiliensis Killing by Gamma‐Interferon Activated Human Neutrophils
Authors: Costa Diego Luís , Dias‐Melicio Luciane Alarcão , Acorci Michele Janegitz , Bordon Ana Paula , Tavian Elisandra Garcia , Peraçoli Maria Terezinha Serrão , de Campos Soares Angela Maria Victoriano
Published: November 2013

Paracoccidioidomycosis, a deep mycosis endemic in Latin America, is a chronic granulomatous disease caused by the fungus Paracoccidioides brasiliensis. Phagocytic cells play a critical role against this fungus, and several studies have shown the effects of activator and suppressive cytokines on macrophage and monocyte functions. However, studies on polymorphonuclear neutrophils (PMNs), that are the first cells recruited to the infection sites, are scarcer. Thus, the objective of this paper was to assess whether interleukin‐10 (IL‐10), a potent anti‐inflammatory cytokine, is able to block the activity of IFN‐γ‐activated human PMNs upon P. brasiliensis intracellular killing, in vitro. The results showed that IFN‐γ‐activated PMNs have an effective fungicidal activity against the fungus. This activity was associated with the release of high levels of H2O2, the metabolite involved in phagocytic cells antifungal activities. However, the concomitant incubation of these cells with IFN‐γ and IL‐10 significantly blocked IFN‐γ activation. As a consequence, PMNs killing activity and H2O2 release were inhibited. Together, our results show the importance of PMNs exposure to activator or suppressor cytokines in the early stages of paracoccidioidomycosis infection.
Beneficial Effect of GB Virus C Co‐Infection in Human Immunodeficiency Virus Type 1‐Infected Individuals
Authors: Hattori Junko , Okumura Naoya , Yamazaki Yumiko , Uchiyama Masataka , Hamaguchi Motohiro , Nishiyama Yukihiro , Kaneda Tsuguhiro
Published: November 2013

Several reports have documented a better prognosis for HIV‐1‐infected patients co‐infected with GBV‐C, while other reports have contradicted such findings with the result that this issue remains controversial. We attempted to clarify the complicated status of the effect of GBV‐C co‐infection on HIV‐1‐infected patients. GBV‐C RNA was detected in 37 samples in 182 HIV‐1‐infected patients (20.3%) using RT/nested PCR. Of these, 3 were determined to be GBV‐C genotype 1, 12 were genotype 2, and the remaining 22 were genotype 3. The GBV‐C viral load quantified by real‐time PCR ranged from 7.8 × 103 to 3.3 × 106 copies/ml. Weakly negative correlation was observed between GBV‐C viral load and HIV‐1 viral load in 19 HAART‐naïve patients, indicating that a higher GBV‐C viral load is associated with a greater suppression of HIV‐1 replication. A previously published in vitro study suggested that GBV‐C infection would induce up‐regulation of RANTES, leading to suppression of HIV‐1 replication. However, in our present study, the blood RANTES level was significantly lower in the GBV‐C co‐infected group than in the uninfected group (190–9,959 vs. 264–31,038 pg/ml, P=0.004). Our results suggested that a suppression of HIV‐1 replication by GBV‐C co‐infection is not mediated by up‐regulated RANTES, and thus call for another as yet unknown factor.
A Correlation between Phagocytosis and Apoptosis in THP‐1 Cells Infected with Prevalent Strains of Mycobacterium tuberculosis
Authors: Rajavelu Priya , Das Sulochana D.
Published: November 2013

The innate ability of infected macrophages to undergo programmed cell death (apoptosis) and curtail the infection is crucial for the host defense. Although phagocytosis and intracellular killing mechanisms leading to apoptosis in macrophages are highly effective in eliminating the infecting tuberculous bacilli, some Mycobacterium tuberculosis (Mtb) strains have evolved strategies to inhibit this microbicidal function and make use of macrophage for its successful and prolonged survival. Two clinical strains of Mtb (S7 and S10) found to be prevalent and primitive, based on molecular epidemiological studies, were used to study the magnitude in induction of apoptosis in THP‐1 cells at various time points of infection and to correlate it with phagocytosis. The percentage of phagocytosis did not show any strain‐specific association with differentiated THP‐1 cells. But in the phagocytic index, the clinical strains showed a low dose of infection in the 1–10 bacilli category thereby exerting less burden on the cells. The induction of apoptosis was strain dependent The THP‐1 cells infected with H37Ra and S10 showed an increase in apoptosis at all time points while the S7 strain induced minimum apoptosis. A negative correlation between apoptosis and phagocytic index was observed in the 1–10 category and a positive correlation in the >20 category of the phagocytic index. This novel observation indicates that the magnitude of THP‐1 cell apoptosis is a function of the number of internalized mycobacteria. These results indicated a differential mode of infection by clinical strains and their adaptation to different survival strategies that may lead to immune suppression and pathogenesis of the disease.
A New Screening Method to Identify Inhibitors of the Lol (Localization of lipoproteins) System, a Novel Antibacterial Target
Authors: Ito Hideaki , Ura Atsushi , Oyamada Yoshihiro , Yoshida Hiroaki , Yamagishi Jun‐ichi , Narita Shin‐ichiro , Matsuyama Shin‐ichi , Tokuda Hajime
Published: November 2013

As the Lol system, which is involved in localization of lipoproteins, is essential for Escherichia coli growth and widely conserved among gram‐negative bacteria, it is considered to be a promising target for the development of anti‐gram‐negative bacterial agents. However, no high‐throughput screening method has so far been developed to screen for Lol system inhibitors. By combining three assay systems (anucleate cell blue assay, Lpp assay, and LolA‐dependent release inhibition assay) and a drug susceptibility test, we have successfully developed a new screening method for identification of compounds that inhibit the Lol system. Using this new screening method, we screened 23,600 in‐house chemical compounds and found 2 Lol system inhibitors. We therefore conclude that our new screening method can efficiently identify new antibacterial agents that target the Lol system.
DDX1 Promotes Proliferation of the JC Virus through Transactivation of Its Promoter
Authors: Sunden Yuji , Semba Shingo , Suzuki Tadaki , Okada Yuki , Orba Yasuko , Nagashima Kazuo , Umemura Takashi , Sawa Hirofumi
Published: November 2013

Recently, we demonstrated that the DEAD box protein 1 (DDX1), an RNA helicase, and the cleavage stimulation factor (CstF) form a complex that binds to the JC virus transcriptional control region (JCV‐TCR). Here, we examined the function of DDX1, which is expressed at much higher levels in the JCV‐susceptible cell line IMR‐32 than in non‐susceptible cell lines. DDX1 had no effect on the replication efficiency of JCV, but over expression of DDX1 significantly increased transactivation of the JCV promoter. Furthermore, DDX1 enhanced the expression of JCV proteins in JCV infected cells, and knockdown of DDX1 using small interfering (si) RNA suppressed the expression of JCV proteins. Our results clearly demonstrate that DDX1 regulates proliferation of JCV in vitro through transcriptional activation.
Interferon‐Gamma Reverses the Evasion of Birc1e/Naip5 Gene Mediated Murine Macrophage Immunity by Legionella pneumophila Mutant Lacking Flagellin
Authors: Akamine Morikazu , Higa Futoshi , Haranaga Syusaku , Tateyama Masao , Mori Naoki , Heuner Klaus , Fujita Jiro
Published: November 2013

Legionella pneumophila is the etiologic agent of Legionnaires' disease. This bacterium contains a single monopolar flagellum, of which the FlaA subunit is a major protein constituent The murine macrophage resistance against this bacterium is controlled by the Birc1e/Naip5 gene, which belongs to the NOD family. We evaluated the intracellular growth of the flaA mutant bacteria as well as another aflagellated fliA mutant, within bone marrow‐derived macrophages from mice with an intact (C57BL/6, BALB/c) or mutated (A/J) Birc1e/Naip5 gene. The flaA mutant L. pneumophila multiplied within C57BL/6 and BALB/c macrophages while the wild‐type strain did not Cell viability was not impaired until 3 days after infection when the flaA mutant bacteria replicated 102‐3‐fold in macrophages, implying that L. pneumophila inhibited host cell death during the early phase of intracellular replication. The addition of recombinant interferon‐gamma (IFN‐γ) to the infected macrophages restricted replication of the flaA mutant within macrophages; these treated cells also showed enhanced nitric oxide production, although inhibition of nitric oxide production did not affect the IFN‐γ induced inhibition of Legionella replication. These findings suggested that IFN‐γ activated macrophages to restrict the intracellular growth of the L. pneumophila flaA mutant by a NO independent pathway.