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Higo-Moriguchi K.,Aichi University | Shirato H.,Japan National Institute of Infectious Diseases | Someya Y.,Japan National Institute of Infectious Diseases | Kurosawa Y.,Health Science University | And 3 more authors.
Journal of Medical Virology | Year: 2014

In order to identify the repertoire of antibodies generated on natural infection of norovirus (NoV) in humans, and to characterize the human monoclonal antibodies against NoV, three phage-displayed antibody libraries originating from healthy person(s) were screened using purified virus-like particles (VLPs) of strain Narita 104 (r104, genogroup II, genotype 4) or strain Chiba 407 (rCV, genogroup I, genotype 4) as antigens. On screening with r104, 62 clones were isolated. Among these antibodies, two clones, 12A11 and 12B10, showed intra-genogroup cross-reactivity to genotypes 1, 3-7, 12, and 14, and genotypes 1, 4, 6, and 7 of genogroup II, respectively. In addition, antibodies belonging to the same group were isolated from two different libraries. On screening with rCV, five clones were isolated, two of which were cross-reactive. One, CV-2F5, reacted to genotypes 1-4, and 8 of genogroup I, and the other, CV-1A5, showed inter-genogroup cross-reactivity to all the VLPs employed in this study. The blocking activities of the monoclonal antibodies against the interaction of homotypic VLPs (VLPs used in the panning procedure) with histo-blood group antigens were also assessed as an alternative to neutralization assay. Although the blocking activity of 12A11 was partially limited 12B10 prevented the binding of r104 to histo-blood group antigens that had been reported to bind r104. The blocking activity of CV-2F5 against the attachment of rCV to suitable histo-blood group antigens was weak, but the blocking activity of CV-1A5 was well recognized. Thus, 12B10 and CV-1A5 were suggested to be cross-reactive monoclonal antibodies with neutralizing activity. © 2013 Wiley Periodicals, Inc. Source

Kanai Y.,Rakuno Gakuen University | Kanai Y.,Thailand Japan Research Collaboration Center on Emerging and Re emerging Infections | Tsujikawa M.,Benesis Corporation | Yunoki M.,Benesis Corporation | And 3 more authors.
Journal of Medical Virology | Year: 2010

Pigs are presumed reservoirs for hepatitis E virus (HEV) transmission to humans. To examine infection kinetics, two litters of domestic pigs (A and B, each containing 10 piglets) infected naturally with HEV were studied until pigs were 6 months old. Maternal IgG and IgA antibodies were detected in litter A piglets, but not in litter B ones. All pigs shed HEV in feces when they were 30-110 days old, and 17 developed viremia at 40-100 days of age. Phylogenetic analysis revealed a highly close sequence of HEV genotype 3 in all pigs. The serum levels of specific IgG and IgA were similar in all pigs, although IgA was not detected in the feces. Interestingly, the onset of both viremia and seroconversion was delayed significantly in litter A pigs. The kinetics of fecal virus shedding was similar in both litters; shedding was not detected after the pigs were 120 days old. The differences in the infection kinetics between litters A and B suggested that maternal antibodies delayed the onset of viremia and seroconversion. Quantitative realtime reverse transcriptase-polymerase chain reaction revealed that HEV RNA in feces peaked 10 days after initial shedding of approximately 106.0 copies/g. The viral load was much lower in the serum than in the feces. At 200 days of age, HEV RNA was found in the internal organs of 3 out of 13 pigs. These study findings improve the understanding of the dynamics of natural HEV transmission in pigs, which could help in controlling virus transmission from pigs to humans. © 2009 Wiley-Liss, Inc. Source

Okura M.,Japan National Agriculture and Food Research Organization | Takamatsu D.,Japan National Agriculture and Food Research Organization | Takamatsu D.,Gifu University | Maruyama F.,Tokyo Medical and Dental University | And 11 more authors.
Applied and Environmental Microbiology | Year: 2013

Streptococcus suis strains are classified into 35 serotypes on the basis of the antigenicity of their capsular polysaccharides (CPs). CP synthesis genes are known to be clustered on the chromosome (cps gene cluster). The entire cps gene clusters of S. suis have so far been sequenced in 15 serotypes and found to be located between orfZ and aroA. In this study, to provide comprehensive information about S. suis CPs, we sequenced the entire cps gene clusters of the remaining serotypes and analyzed the complete set of S. suis cps gene clusters. Among the 35 cps gene clusters, 22 were located between orfZ and aroA, whereas the other 13 were flanked by other gene(s) on the chromosomes, and the chromosomal locus was classified into five patterns. By clustering analysis, the predicted products of cps genes found in the 35 serotypes were assigned into 291 homology groups, and all serotypes possessed a serotype-specific gene, except for serotypes 1, 2, 1/2, and 14. Because of the presence of genes encoding flippase (wzx) and polymerase (wzy), CPs of all serotypes were thought to be synthesized by the Wzx/Wzy pathway. Our data also implied the possibility of the transfer of the entire or partial cps gene clusters among S. suis strains, as well as the influence of spontaneous mutations in a single gene or a few genes on the antigenicity of some serotypes. Accumulation of these gene transfers and smallscale mutations may have generated the antigenic diversity of S. suis CPs. © 2013, American Society for Microbiology. Source

Utachee P.,Thailand Japan Research Collaboration Center on Emerging and Re emerging Infections | Isarangkura-na-ayuthaya P.,National Institute of Health | Tokunaga K.,Japan National Institute of Infectious Diseases | Ikuta K.,Kobe University | And 5 more authors.
Retrovirology | Year: 2014

Background: The CD4 binding site (CD4bs) of envelope glycoprotein (Env) gp120 is a functionally conserved, important target of anti-human immunodeficiency virus type 1 (HIV-1) neutralizing antibodies. Two neutralizing human monoclonal antibodies, IgG1 b12 (b12) and VRC01, are broadly reactive neutralizing antibodies which recognize conformational epitopes that overlap the CD4bs of Env gp120; however, many CRF01_AE viruses are resistant to neutralization mediated by these antibodies. We examined the mechanism underlying the b12 resistance of the viruses using CRF01_AE Env (AE-Env)-recombinant viruses in this study. Results: Our results showed that an amino acid substitution at position 185 in the V2 region of gp120 played a crucial role in regulating the b12 susceptibility of AE-Env-recombinant viruses by cooperating with 2 previously reported potential N-linked glycosylation (PNLG) sites at positions 186 (N186) and 197 (N197) in the V2 and C2 regions of Env gp120. The amino acid residue at position 185 and 2 PNLG sites were responsible for the b12 resistance of 21 of 23 (>91%) AE-Env clones tested. Namely, the introduction of aspartic acid at position 185 (D185) conferred b12 susceptibility of 12 resistant AE-Env clones in the absence of N186 and/or N197, while the introduction of glycine at position 185 (G185) reduced the b12 susceptibility of 9 susceptible AE-Env clones in the absence of N186 and/or N197. In addition, these amino acid mutations altered the VRC01 susceptibility of many AE-Env clones. Conclusions: We propose that the V2 and C2 regions of AE-Env gp120 contain the major determinants of viral resistance to CD4bs antibodies. CRF01_AE is a major circulating recombinant form of HIV-1 prevalent in Southeast Asia. Our data may provide important information to understand the molecular mechanism regulating the neutralization susceptibility of CRF01_AE viruses to CD4bs antibodies. © 2014 Utachee et al.; licensee BioMed Central Ltd. Source

Okada K.,Thailand Japan Research Collaboration Center on Emerging and Re emerging Infections | Okada K.,Osaka University | Roobthaisong A.,Thailand Japan Research Collaboration Center on Emerging and Re emerging Infections | Nakagawa I.,Tokyo Medical and Dental University | And 3 more authors.
PLoS ONE | Year: 2012

Background: Vibrio cholerae O1 El Tor dominated the seventh cholera pandemic which occurred in the 1960s. For two decades, variants of V. cholerae O1 El Tor that produce classical cholera toxin have emerged and spread globally, replacing the prototypic El Tor biotype. This study aims to characterize V. cholerae O1 isolates from outbreaks in Thailand with special reference to genotypic variations over time. Methods/Findings: A total of 343 isolates of V. cholerae O1 from cholera outbreaks from 2007 to 2010 were investigated, and 99.4% were found to carry the classical cholera toxin B subunit (ctxB) and El Tor rstR genes. Pulsed-field gel electrophoresis (PFGE) differentiated the isolates into 10 distinct pulsotypes, clustered into two major groups, A and B, with an overall similarity of 88%. Ribotyping, multiple-locus variable-number tandem-repeat analysis (MLVA), and PCR to detect Vibrio seventh pandemic island II (VSP-II) related genes of randomly selected isolates from each pulsotype corresponded to the results obtained by PFGE. Epidemiological investigations revealed that MLVA type 2 was strongly associated with a cholera outbreak in northeastern Thailand in 2007, while MLVA type 7 dominated the outbreaks of the southern Gulf areas in 2009 and MLVA type 4 dominated the outbreaks of the central Gulf areas during 2009-2010. Only MLVA type 16 isolates were found in a Thai-Myanmar border area in 2010, whereas those of MLVA types 26, 39, and 41 predominated this border area in 2008. Type 39 then disappeared 1-2 years later as MLVA type 41 became prevalent. Type 41 was also found to infect an outbreak area. Conclusions: MLVA provided a high-throughput genetic typing tool for understanding the in-depth epidemiology of cholera outbreaks. Our epidemiological surveys suggest that some clones of V. cholerae O1 with similar but distinctive genetic traits circulate in outbreak sites, while others disappear over time. © 2012 Okada et al. Source

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