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PXII-1 Assembly of a chimeric hantavirus-like particle containing the araraquara virus nucleoprotein and the andes virus glycoproteins
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Posters, 12th ESCV, Istanbul / Journal of Clinical Virology 46 Suppl. 1 (2009) S15–S61 PXII-2 Phylogenetic analysis of 4 enterovirus 71 strains isolated in Brest, France
Posters XI: Antivirals PXI-1 Oleuropein inhibits rhinovirus infection via interruptions of human rhinovirus attachment to cells H.-J. Choi *, J.-H. Choi, J.-N. Choi, J.-H. Song, K.-S. Park, Y.-S. Jang, Y.-K. Kim. Korea Research Institute of Bioscience and Biotechnology, South Korea Human rhinoviruses (HRV), which are the most frequent causative agents of acute upper respiratory infections, are abundant worldwide. The intercellular adhesion molecule 1 (ICAM-1) is the major group rhinovirus receptor and the molecular pathogenesis of HRV infection is quite completely understood. However, there is no registered clinically effective antiviral agent for treatment of diseases caused by HRVs. In this article, oleuropein (OP) from Ligustrum obtusifolium inhibited PMA-induced ICAM-1/LFA-1 medicated aggregation of HL-60 cells and showed anti-human rhinovirus 3 activity with no cytotoxicity at tested concentration. Furthermore, OP only did inhibit HRV3 infection when it was added prior (−1 h) or co (0 h) infection. However, ribavirin have relative weaker efficacy compared to OP with showing a different trend with OP during HRV3 infection. Collectively, our results suggest that the antiviral activity of OP against HRV3 is correlated with the attachment and entry to host cells. These findings propose that OP may have a possible clinical application in the treatment of disease caused by HRV infection.
Posters XII: Other topics PXII-1 Assembly of a chimeric hantavirus-like particle containing the araraquara virus nucleoprotein and the andes virus glycoproteins F.P. Yeda1 *, S.J. Ontiveros2 , J. Siqueira-Silva1 , M.L. Silva1 , L.T.M. Figueiredo3 , S. St Jeor4 , C.B. Jonsson2 , C.M. H´arsi1 . 1 Department of Microbiology, Institute of Biomedical Sciences, University of S˜ao Paulo, Brazil, 2 Department of Biochemistry and Molecular Biology, Southern Research Institute, Birmingham, AL, USA, 3 Center of Research in Virology, School of Medicine of Ribeir˜ao Preto, University of S˜ao Paulo, Brazil, 4 Department of Microbiology, School of Medicine, University of Nevada, Reno, NV, USA Hantaviruses, are rodent-borne viruses that cause hemorrhagic fever with renal syndrome and hantavirus cardiopulmonary syndrome (HCPS). The S, M and L genomic RNA segments encode the nucleoprotein (N), the glycoproteins G1 and G2, and the RNA polymerase, respectively. The glycoproteins elicit organism to produce neutralizing antibodies. Although nucleoprotein does not elicit high-titer neutralization antibodies, several studies indicate that N does play a role in eliciting protective immunity. Up to now, no effective vaccine has been developed to prevent the HCPS. The aim of this study was to obtain a chimeric hantavirus-like particle containing the Araraquara nucleoprotein and the Andes glycoproteins expressed in baculovirus system. The nucleoprotein and glycoproteins genes were cloned into pFastBacDual. The proteins coexpression assays were done on High five cells and was analyzed by Westernblot. The proteins co-localization was analyzed by immunofluorescence assay (IFA) and confocal laser scanning microscopy (CM).The proteins interaction was analyzed by immunoprecipitation (IP) and density sucrose gradient. The results showed that the proteins were detected in the plasmatic membrane, Golgi and cytoplasmatic fractions. The N-G1 and N-G2 proteins were colocalized in High five cells by IFA and CM. The IP assay showed the interaction between the N, G1 and G2 proteins. Futhermore, the three proteins were detected in the same fraction with density between 0.8 and 1.2 g/cc in sucrose gradient. Our results showed that chimeric hantavirus-like particles were obtained with similar biochemical properties of the hantavirus particle in vivo and it can be used in immunological, structural and functional studies.
S. Vallet1,2 *, M.-C. Legrand-Quillien1 , T. Dailland3 , G. Podeur2 , S. Gouriou2 , I. Schuffenecker4 , C. Payan1,2 , P. Marcorelles5 . 1 CHU de Brest, D´epartement de Microbiologie, Brest, France, 2 Universit´e de Brest, Facult´e de M´edecine et des sciences de la Sant´e, LUBEM EA3882, Brest, France, 3 CHU Brest, D´epartement de P´ediatrie, Brest, France, 4 National Reference Center for Enteroviruses, Laboratoire de Virologie, Centre de Biologie et Pathologie EST, Bron, France, 5 CHU Brest, Laboratoire d’Anatomo-pathologie, Brest, France Enterovirus 71 (EV71) induces mostly asymptomatic or benign infections. It is a leading cause of hand-foot-and-mouth disease (HFMD). Since its identification in 1969, outbreaks of HFMD occur regularly in Asia-Pacific region. This neurotropic virus can cause severe complications, including aseptic meningitis, encephalitis and poliomyelitis-like paralysis mainly in children under 5 years. Since 1997, a growing number of pulmonary oedema often fatals have been described. Until now, few cases of symptomatic EV71 infections have been reported in France. Since 2004, four EV71 strains have been isolated in Brest, France. One of these four strains was associated with the first fatal case of EV71 infection reported in France. The four children (between 1 and 23 months of age) were referred to the pediatric emergency ward of the Brest University Hospital with respectively pulmonary oedema which rapidely became fatal, febrile gingivo-stomatitis with encephalitis, acute respiratory syndrom and HFMD). Full length VP1 gene of each isolated viral strains has been sequenced. These sequences were compared by phylogenetic analyses with representative worldwide EV71 strains from all 11 known subgenotypes. The VP1 sequences of the deceased case and of two other children clustered together into subgenotypes C2 while the fourth one clustered with subgenotypes C4. The high nucleotide identity between the three C2 strains, their place and period of circulation, suggest a probable identical ancestral strain. International spread of EV71 outside Asia-Pacific region needs to be monitored, as both the possibility of an outbreak and the unpredictability of virus circulation patterns represent a public health concern. PXII-3 Age-related detection of ’high-risk’ human papilloma virus (HPV) 16 in children’s oral cavity G. Sourvinos1 *, I. Mammas1 , P. Giamarellou2 , C. Michael2 , D.A. Spandidos1 . 1 Department of Clinical Virology, School of Medicine, University of Crete, Greece, 2 Department of Pathology, “P.&A. Kyriakou” Children’s Hospital, Athens, Greece Background and Aim: To date, several studies have evaluated the presence of HPV DNA in the oral cavity in children by studying oral swabs or washings from healthy children. The aim of our study was to investigate the presence of HPV DNA in Greek children’s oral cavity. Methods: Seventy-eight (78) samples of oral mucosa from 78 children, 34 girls and 44 boys (age range: 2 years to 14 years), were tested for the presence of HPV DNA using polymerase chain reaction (PCR) with general primers GP5+/GP6+. HPV typing was performed by PCR with specific primers for HPV 16, 18, 33 and 11. Results: HPV DNA was detected in 4 (5.1%) of the 78 collected specimens. The frequencies of HPV typing were 3/4 (75%) for HPV 16, 0/4 (0%) for HPV 11, for HPV 33 and HPV 18, while one HPV-positive sample remained untyped. No multiple HPV infection was detected. All three HPV 16-positive children were less than 5 years of age. The mean age of children with HPV 16positive specimens was lower than that of HPV-negative children (p < 0.010). No statistical correlation in the prevalence of HPV infection was observed according to children’s sex, origin or residence. Conclusion: Age-related detection of HPV 16 infection in the oral cavity of healthy children has raised questions concerning the mode of HPV transmission in childhood. The possible role of the presence of ‘high-risk’ HPVs in the oral cavity, even at a low frequency, remains to be elucidated.
Posters, 12th ESCV, Istanbul / Journal of Clinical Virology 46 Suppl. 1 (2009) S15–S61 PXII-2 Phylogenetic analysis of 4 enterovirus 71 strains isolated in Brest, France
Posters XI: Antivirals PXI-1 Oleuropein inhibits rhinovirus infection via interruptions of human rhinovirus attachment to cells H.-J. Choi *, J.-H. Choi, J.-N. Choi, J.-H. Song, K.-S. Park, Y.-S. Jang, Y.-K. Kim. Korea Research Institute of Bioscience and Biotechnology, South Korea Human rhinoviruses (HRV), which are the most frequent causative agents of acute upper respiratory infections, are abundant worldwide. The intercellular adhesion molecule 1 (ICAM-1) is the major group rhinovirus receptor and the molecular pathogenesis of HRV infection is quite completely understood. However, there is no registered clinically effective antiviral agent for treatment of diseases caused by HRVs. In this article, oleuropein (OP) from Ligustrum obtusifolium inhibited PMA-induced ICAM-1/LFA-1 medicated aggregation of HL-60 cells and showed anti-human rhinovirus 3 activity with no cytotoxicity at tested concentration. Furthermore, OP only did inhibit HRV3 infection when it was added prior (−1 h) or co (0 h) infection. However, ribavirin have relative weaker efficacy compared to OP with showing a different trend with OP during HRV3 infection. Collectively, our results suggest that the antiviral activity of OP against HRV3 is correlated with the attachment and entry to host cells. These findings propose that OP may have a possible clinical application in the treatment of disease caused by HRV infection.
Posters XII: Other topics PXII-1 Assembly of a chimeric hantavirus-like particle containing the araraquara virus nucleoprotein and the andes virus glycoproteins F.P. Yeda1 *, S.J. Ontiveros2 , J. Siqueira-Silva1 , M.L. Silva1 , L.T.M. Figueiredo3 , S. St Jeor4 , C.B. Jonsson2 , C.M. H´arsi1 . 1 Department of Microbiology, Institute of Biomedical Sciences, University of S˜ao Paulo, Brazil, 2 Department of Biochemistry and Molecular Biology, Southern Research Institute, Birmingham, AL, USA, 3 Center of Research in Virology, School of Medicine of Ribeir˜ao Preto, University of S˜ao Paulo, Brazil, 4 Department of Microbiology, School of Medicine, University of Nevada, Reno, NV, USA Hantaviruses, are rodent-borne viruses that cause hemorrhagic fever with renal syndrome and hantavirus cardiopulmonary syndrome (HCPS). The S, M and L genomic RNA segments encode the nucleoprotein (N), the glycoproteins G1 and G2, and the RNA polymerase, respectively. The glycoproteins elicit organism to produce neutralizing antibodies. Although nucleoprotein does not elicit high-titer neutralization antibodies, several studies indicate that N does play a role in eliciting protective immunity. Up to now, no effective vaccine has been developed to prevent the HCPS. The aim of this study was to obtain a chimeric hantavirus-like particle containing the Araraquara nucleoprotein and the Andes glycoproteins expressed in baculovirus system. The nucleoprotein and glycoproteins genes were cloned into pFastBacDual. The proteins coexpression assays were done on High five cells and was analyzed by Westernblot. The proteins co-localization was analyzed by immunofluorescence assay (IFA) and confocal laser scanning microscopy (CM).The proteins interaction was analyzed by immunoprecipitation (IP) and density sucrose gradient. The results showed that the proteins were detected in the plasmatic membrane, Golgi and cytoplasmatic fractions. The N-G1 and N-G2 proteins were colocalized in High five cells by IFA and CM. The IP assay showed the interaction between the N, G1 and G2 proteins. Futhermore, the three proteins were detected in the same fraction with density between 0.8 and 1.2 g/cc in sucrose gradient. Our results showed that chimeric hantavirus-like particles were obtained with similar biochemical properties of the hantavirus particle in vivo and it can be used in immunological, structural and functional studies.
S. Vallet1,2 *, M.-C. Legrand-Quillien1 , T. Dailland3 , G. Podeur2 , S. Gouriou2 , I. Schuffenecker4 , C. Payan1,2 , P. Marcorelles5 . 1 CHU de Brest, D´epartement de Microbiologie, Brest, France, 2 Universit´e de Brest, Facult´e de M´edecine et des sciences de la Sant´e, LUBEM EA3882, Brest, France, 3 CHU Brest, D´epartement de P´ediatrie, Brest, France, 4 National Reference Center for Enteroviruses, Laboratoire de Virologie, Centre de Biologie et Pathologie EST, Bron, France, 5 CHU Brest, Laboratoire d’Anatomo-pathologie, Brest, France Enterovirus 71 (EV71) induces mostly asymptomatic or benign infections. It is a leading cause of hand-foot-and-mouth disease (HFMD). Since its identification in 1969, outbreaks of HFMD occur regularly in Asia-Pacific region. This neurotropic virus can cause severe complications, including aseptic meningitis, encephalitis and poliomyelitis-like paralysis mainly in children under 5 years. Since 1997, a growing number of pulmonary oedema often fatals have been described. Until now, few cases of symptomatic EV71 infections have been reported in France. Since 2004, four EV71 strains have been isolated in Brest, France. One of these four strains was associated with the first fatal case of EV71 infection reported in France. The four children (between 1 and 23 months of age) were referred to the pediatric emergency ward of the Brest University Hospital with respectively pulmonary oedema which rapidely became fatal, febrile gingivo-stomatitis with encephalitis, acute respiratory syndrom and HFMD). Full length VP1 gene of each isolated viral strains has been sequenced. These sequences were compared by phylogenetic analyses with representative worldwide EV71 strains from all 11 known subgenotypes. The VP1 sequences of the deceased case and of two other children clustered together into subgenotypes C2 while the fourth one clustered with subgenotypes C4. The high nucleotide identity between the three C2 strains, their place and period of circulation, suggest a probable identical ancestral strain. International spread of EV71 outside Asia-Pacific region needs to be monitored, as both the possibility of an outbreak and the unpredictability of virus circulation patterns represent a public health concern. PXII-3 Age-related detection of ’high-risk’ human papilloma virus (HPV) 16 in children’s oral cavity G. Sourvinos1 *, I. Mammas1 , P. Giamarellou2 , C. Michael2 , D.A. Spandidos1 . 1 Department of Clinical Virology, School of Medicine, University of Crete, Greece, 2 Department of Pathology, “P.&A. Kyriakou” Children’s Hospital, Athens, Greece Background and Aim: To date, several studies have evaluated the presence of HPV DNA in the oral cavity in children by studying oral swabs or washings from healthy children. The aim of our study was to investigate the presence of HPV DNA in Greek children’s oral cavity. Methods: Seventy-eight (78) samples of oral mucosa from 78 children, 34 girls and 44 boys (age range: 2 years to 14 years), were tested for the presence of HPV DNA using polymerase chain reaction (PCR) with general primers GP5+/GP6+. HPV typing was performed by PCR with specific primers for HPV 16, 18, 33 and 11. Results: HPV DNA was detected in 4 (5.1%) of the 78 collected specimens. The frequencies of HPV typing were 3/4 (75%) for HPV 16, 0/4 (0%) for HPV 11, for HPV 33 and HPV 18, while one HPV-positive sample remained untyped. No multiple HPV infection was detected. All three HPV 16-positive children were less than 5 years of age. The mean age of children with HPV 16positive specimens was lower than that of HPV-negative children (p < 0.010). No statistical correlation in the prevalence of HPV infection was observed according to children’s sex, origin or residence. Conclusion: Age-related detection of HPV 16 infection in the oral cavity of healthy children has raised questions concerning the mode of HPV transmission in childhood. The possible role of the presence of ‘high-risk’ HPVs in the oral cavity, even at a low frequency, remains to be elucidated.