Dissertations / Theses on the topic 'Flaviviruses'
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Shomiad, Shueb Rafidah Hanim. "Contribution of different components of innate and adaptive immunity to severity of flavivirus-induced encephalitis in susceptible and resistant hosts." University of Western Australia. School of Biomedical, Biomolecular and Chemical Sciences, 2008. http://theses.library.uwa.edu.au/adt-WU2008.0199.
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[Truncate abstract] Flaviviruses are small, positive-stranded RNA viruses belonging to the family Flaviviridae. Flavivirus infection in humans could cause diseases ranging from febrile illnesses to fatal encephalitis. Mice provide a useful small animal model to study flavivirus-induced encephalitis in humans since mice also develop encephalitis during flavivirus infection. Some strains of mice have been shown to be resistant to flavivirus challenge and this resistance is conferred by a single autosomal dominant gene, designated as Flvr. Recently, OAS1b gene has been identified to be a gene candidate for Flvr. Several congenic resistant mouse strains have been developed by introducing resistance genes from outbred or wild mice onto the genetic background of susceptible C3H mice. These new resistant strains that carry different allelic variants at the Flv locus include C3H/PRI-Flvr (RV), C3H.MOLD-Flvmr (MOLD) and C3H.M.domesticus-Flvr-like (DUB), the latter two being developed in the same laboratory in which the work described in this thesis was accomplished. Preliminary studies in this laboratory found that flavivirus resistant mice are vulnerable to certain flavivirus infections, particularly when challenged by intracerebral (i.c.) route. Intracerebral (i.c.) challenge with flaviviruses such as West Nile virus (WNV) Sarafend strain and Kunjin virus (KUNV) MRM16 strain were found to induce high mortality in flavivirus resistant mice while infection with Murray Valley encephalitis virus (MVEV) OR2 strain did not cause any apparent disease in the same mice. ... Thus, it can be concluded that CD8+ T cells exerted harmful effect to resistant DUB mice during KUNV i.c. infection by producing excessive IFN[gamma] that could be toxic, causing functional loss of the CNS cells. It was shown from in vitro studies that WNV had the highest tropism for macrophages and dendritic cells, followed by KUNV. MVEV however did not replicate well in these cells. This combined with the data from the in vivo studies indicates that macrophages might be involved in the pathogenesis of intraperitoneal (i.p.) infection of WNV but not KUNV and MVEV. The reason for this could be that the production of KUNV in macrophages may not be high enough to induce viraemia and subsequent fatal encephalitis in mice. In contrast, MVEV appears to use different mechanism or cells for virus dissemination. Although macrophages may not be involved in KUNV pathogenesis after i.p. infection, the fact that macrophages support KUNV replication in vitro may indicate the possibility that blood-borne macrophages were recruited to the brain where they can get infected with KUNV during i.c. infection and therefore could participate in KUNV pathogenesis in DUB mice. This study provides evidence for the first time on the detrimental effect of host antiviral immunity and inflammatory mediators during flavivirus i.c. infection in resistant mice. However, it also launches a new question on the selective cell tropism of KUNV versus MVEV responsible for inducing different pattern of immune responses and consequently leading to different outcomes of infection in resistant mice.
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Klitting, Bottero Raphaëlle. "Attenuation of viscerotropic flaviviruses." Thesis, Aix-Marseille, 2017. http://www.theses.fr/2017AIXM0657/document.
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Avec plus de 20% de morts annuels dus aux maladies infectieuses, celles-ci restent un sujet majeur de santé publique. Des maladies d’origine virale (ré)émergent suite aux changements environnementaux, climatiques et sociétaux : le virus Ebola, la Dengue ou, plus récemment, le virus Zika. Dans ce contexte, il est donc aujourd’hui crucial de développer des vaccins efficaces et sûrs contre les infections virales émergentes. Ce projet de thèse vise à mettre en place une nouvelle stratégie de production de vaccins vivants atténués ciblant les virus à ARN en travaillant sur le virus de la fièvre jaune (genre Flavivirus). Après une analyse génomique qui a permis d’approfondir une technique de modification des virus appelée « ré-encodage », des mutants de la fièvre jaune ont été produits puis caractérisés in vitro et in vivo. En parallèle, un modèle rongeur de la fièvre jaune a été développé et a permis de tester in vivo à la fois l’innocuité et l’efficacité vaccinale des virus ré-encodésDespite recent considerable improvements, infectious diseases remain a major issue for public health, with an estimated 20% of annual deaths caused by infections. Among them, viral diseases (re)emerge following environmental, climatic and societal changes: Ebola, Dengue and Zika viruses have recently been the object of special attention. The development of safe and efficient vaccines against emerging viruses is a major challenge for global public health. This thesis work is in line with this issue. Using the yellow fever virus (YFV, genus Flavivirus) as a model, we tried to define new strategies for the design of live-attenuated vaccines for viral infections prevention. After a genomic analysis that allowed to go further into a procedure for virus modification named “re-encoding”, we generated and characterised both in vitro and in vivo mutant strains of YFV. In parallel, a rodent model was set up to test in vivo both the safety and the protective efficiency of the re-encoded viruses
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Uzcategui, Cuello Nathalie Yumari. "Evolution and dispersal of mosquito-borne flaviviruses." Thesis, University of Oxford, 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.288520.
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COSTA, Sónia Fernandes da. "Flaviviruses in mosquitoes from Southern Portugal, 2009-2010." Master's thesis, Instituto de Higiene e Medicina Tropical, 2011. http://hdl.handle.net/10362/7156.
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Os flavivírus são vírus pertencentes à família Flaviviridae, género Flavivirus. Estes formam um grande grupo caraterizado pela sua ampla distribuição e diversidade genética. Os flavivírus são, na sua maioria, transmitidos por artrópodes vectores incluíndo agentes patogénicos para humanos e animais que podem potencialmente provocar grandes epidemias e causar elevadas taxas de mortalidade e morbidade. Nos últimos anos, tem-se registado uma grande expansão a nível da distribuição geográfica dos flavivirus e diversidade dos seus hospedeiros. O vírus do Nilo Ocidental tem sido continuamente detectado em toda a Europa recentemente, e também isolado de mosquitos colhidos no Sul de Portugal, onde já foram registados casos humanos e animais.
O principal objectivo deste trabalho é o rastreio de flavivírus em mosquitos colhidos em duas regiões do Sul de Portugal, onde os mesmos foram anteriormente detectados.
As colheitas de mosquitos foram realizadas em 24 locais em zonas húmidas nos districtos de Faro e Setúbal, através de armadilhas luminosas tipo CDC com CO2 e aspiradores mecânicos manuais para colheita de mosquitos em repouso em abrigos de animais.
Os mosquitos colhidos foram agrupados por lotes contendo aproximadamente 50 espécimens cada, e rastreados para a presença de flavivírus por heminested RT-PCR, direccionado à amplificação de um pequeno fragmento do gene NS5 usando oligonucleótidos degenerados específicos para flavivírus.
Entre Abril e Outubro de 2009 e 2010 foram colhidos no total 36273 mosquitos pertencentes às seguintes espécies: Anopheles algeriensis, An.atroparvus, Aedes berlandi, Ae.caspius, Ae.detritus, Coquillettidia richiardii, Culex laticinctus, Cx.pipiens, Cx.theileri, Cx.univittatus, Culiseta annulata, Cs.longiareolata, Cs.subochrea, e Uranotaenia unguiculata. As espécies mais abundantes foram Ae.caspius, Cx.theileri e Cx.pipiens, respectivamente. Contudo, as densidades de mosquitos foram variáveis de acordo com o método de colheita e área de amostragem. As densidades de mosquitos colhidos em 2010 foram quatro vezes superior às registadas no ano anterior. No total foram analisados 745 lotes dos quais 31% testaram positivos para a presença de sequências de flavivirus.
As espécies que apresentaram taxas de positividade mais elevadas foram: An.algeriensis com uma Taxa Mínima de Infecção (TMI) de 56/1000 no Algarve em 2009, Cs.annulata TMI =22/1000 no Algarve em 2010, Cx.theileri e Cx.pipiens em Setúbal em 2010, TMI =20/1000. An. atroparvus, Ae. caspius, Ae. detritus e Cx. univittatus também produziram lotes positives. No geral, a positividade foi maior no Algarve.
Análise das sequências virais obtidas revelou homologia das nossas sequências virais com sequências de referência de flavivírus específicos de mosquitos depositadas em bases de dados de acesso livre. A análise filogenética reflectiu a variabilidade genética dos flavivírus e revelou a relação genética das nossas sequências com as de outros flavivírus, especialmente os específicos de insectos.
Tendo em consideração os anteriores isolamentos do vírus do Nilo Ocidental, o aumento acentuado nas densidades de mosquitos, o aumento de temperaturas que se tem vindo a registar, os casos recentes de transmissão de flavivírus por toda a Europa e o padrão desconhecido e imprevisível dos surtos destes vírus, os programas contínuos de vigilância epidemiológica têm-se revelado uma ferramenta indispensável para a Saúde Pública.Flaviviruses are viruses belonging to the Flaviviridae family, genus Flavivirus. They comprise a large group of widely spread and genetically diverse arthropod-borne viruses including human and animal pathogens that can potentially cause large-scale epidemics and high mortality and morbidity. In the past few years, flaviviruses have largely expanded their geographical distribution and host range. West Nile virus has been continuously detected throughout Europe lately and has been isolated from mosquitoes in Southern Portugal, where human and animal cases have been reported. The main aim of this work was to search for flaviviruses in mosquitoes collected from two areas in Southern Portugal where West Nile virus and other flaviviruses have previously been detected. Mosquito surveys were carried out in 24 locations in the wetlands of the Faro and Setúbal districts, by CDC-CO2 light-traps and indoors resting collections. Pools containing approximately 50 mosquitoes were screened for flaviviruses by heminested RT-PCR, directed at the amplification of a small fragment of the viral NS5 gene, using degenerated flavivirus-specific primers. A total of 36273 mosquitoes were collected during 2009 and 2010 from April through October, from the following species: Anopheles algeriensis, An.atroparvus, Aedes berlandi, Ae. caspius, Ae. detritus, Coquillettidia richiardii, Culex laticinctus, Cx. pipiens, Cx. theileri, Cx. univittatus, Culiseta annulata, Cs. longiareolata, Cs. subochrea, and Uranotaenia unguiculata. Most abundant species were Ae. caspius Cx. theileri and Cx. pipiens, respectively. However, mosquito densities varied according to collection method and sampling area. A fourfold increase in mosquito density was registered in 2010 compared to 2009. A total of 745 pools were analysed of which 31% tested positive for flaviviral sequences. The species with higher positivity rates were An. algeriensis with Minimum infection rate (MIR) of 56/1000 in the Algarve 2009, Cs. annulata MIR =22/1000 in the Algarve 2010, Cx.theileri and Cx.pipiens in Setúbal 2010, MIR =20/1000. An. atroparvus, Ae. caspius, Ae. detritus and Cx. univittatus also yielded positive pools. Overall, positivity was higher in the Algarve. Viral sequences obtained from positive pools showed homology with insect-specific flavivirus (ISF) sequences deposited in free access public databases. Phylogenetic analysis reflected the genetic variability of flaviviruses and revealed the relatedness of our sequences with other known flaviviruses, especially the insect-specific. In view of previous WNV isolations and assessing from the four-fold increase in mosquito density, the increasing temperatures, the recent cases throughout Europe and the unknown and unpredictable pattern of flaviviruses outbreaks, continuous epidemiological surveillance programmes are quickly becoming indispensable tools for Public Health.
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Jenkins, Gareth. "Determinants of the molecular evolution of RNA viruses." Thesis, University of Oxford, 2001. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.365413.
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Zanotto, Paolo Marinho de Andrade. "Aspects of the molecular evolution of baculoviruses and flaviviruses." Thesis, University of Oxford, 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.318444.
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Proutski, Vitali. "RNA secondary structure of the 3'-UTR of flaviviruses." Thesis, University of Oxford, 1998. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.299156.
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Gao, George Fu. "Molecular biological and immunological studies of tick-borne flaviviruses." Thesis, University of Oxford, 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.296917.
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Cook, Shelley. "The molecular evolution of the flaviviruses and their vectors." Thesis, University of Oxford, 2005. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.427883.
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Izuogu, Adaeze O. Izuogu. "Restriction of tick-borne flaviviruses in the white-footed mouse." University of Toledo Health Science Campus / OhioLINK, 2017. http://rave.ohiolink.edu/etdc/view?acc_num=mco1501786858639212.
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Dunster, Lee Martin. "Studies on the attenuation of flaviviruses following passage in HeLa cells." Thesis, University of Surrey, 1990. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.276337.
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Matusan, Anita Esther 1973. "Mutational analysis of the proteinase and helicase regions of the Dengue virus type 2 NS3 protein." Monash University, Dept. of Microbiology, 2001. http://arrow.monash.edu.au/hdl/1959.1/8339.
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Henriques, Dyana Alves. "Caracterização molecular de arbovírus isolados da fauna diptera nematocera do Estado de Rondônia (Amazônia ocidental brasileira)." Universidade de São Paulo, 2008. http://www.teses.usp.br/teses/disponiveis/42/42132/tde-27032009-124003/.
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Rondônia apresenta área com rica diversidade de artrópodes, porém pouco se conhece sobre a transmissão de arbovírus por estas espécies. O presente trabalho visou detectar arbovírus, por meio da RT-PCR e da Duplex RT-PCR, nas espécies de dipteros coletados no Estado, bem como caracterizá-los pela reação de sequenciamento. A RT-PCR e a Duplex RT-PCR detectaram as suspensões dos vírus Mayaro e Oropouche até 104 e 101 TCID50/mL, respectivamente, porém o vírus Dengue 2 em pools contendo menos de três mosquitos infectados foi negativa. O controle endógeno foi detectado em 66,8 % das amostras, sendo que, em pools contendo entre um e três mosquitos, a detecção foi aproximadamente metade da detecção nos pools contendo entre 11 e 15. Em 0,66 % dos pools foi encontrado o vírus Oropouche e em outros 0,66 %, o vírus Cacipacoré. O vírus Oropouche foi detectado em Coquillettidia sp. e Deinocerites sp., enquanto o vírus Cacipacoré foi encontrado em Anopheles sp. e Culex sp. As técnicas possibilitaram a detecção dos arbovírus pesquisados nos pools coletados em Rondônia.The Rondônia state has an area with rich arthropods diversity although the knowledge about the arboviruses transmition for these species is poor. The present work aimed to detect arboviruses through RT-PRC and RT-PCR Duplex in the diptera species collected in the region as well as their characterization through the sequence reaction. The RT-PRC and RT-PCR Duplex detected the Mayaro and Oropouche virus suspensions until 104 e 101 TCID50/mL respectively, although it was negative for the Dengue 2 virus in pools containing less than three infected mosquitoes. The endogenous control was detected in 66,8 % of samples and from pools containing from one to three mosquitoes the detection rate was approximately half from that obtained from pools with 11 to 15 mosquitoes. Oropouche virus was found in 0,66 % of pools and Cacipacore virus also in 0,66 % of pools. Oropouche virus was detected in Coquillettidia sp. and Deinocerites sp. while Cacipacoré virus was found in Anopheles sp. and Culex sp. The techniques allowed the detection of examined arboviruses in the pools collected from Rondonia.
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Grant, Alesha. "Functional Conservation of Interferon Antagonism among Flaviviruses| Zika Virus Targets Human STAT2." Thesis, Icahn School of Medicine at Mount Sinai, 2017. http://pqdtopen.proquest.com/#viewpdf?dispub=10270604.
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Flaviviruses are a diverse group of emerging arboviruses capable of infecting an extraordinarily broad range of vertebrate and invertebrate hosts. Nearly half of the viruses in this rapidly expanding genus have been reported to be pathogenic for humans, as well as other vertebrates. The spectrum of human disease includes asymptomatic and febrile illnesses, rash, arthralgia, encephalitis and hemorrhagic fever. The recent outbreak of Zika virus (ZIKV) has uncovered pathology in the form of microcephaly and Guillain-Barré syndrome, cementing the importance of flaviviruses as emerging human pathogens. All vector-borne flaviviruses studied thus far have to overcome type I interferon (IFN) antiviral responses in order to replicate and cause disease in vertebrates. The non-structural protein NS5 is a potent and specific antagonist of IFN signaling for human pathogenic flaviviruses such as dengue virus (DENV), yellow fever virus (YFV), West Nile virus (WNV), and tick-borne encephalitis viruses (TBEVs). Intriguingly, each of these viruses exhibits different mechanisms of IFN antagonism, highlighting the complicated evolutionary nature of flaviviruses. This thesis work presents novel insights into the NS5-mediated antagonism of IFN signaling for several underexamined flaviviruses. Notably, all NS5 proteins examined were able to inhibit IFN-induced gene expression in a mammalian system, indicating a functional conservation of IFN antagonism for flavivirus NS5 proteins. However, mechanistically NS5 function was diverse. Of great interest, ZIKV NS5 bound to the human, but not mouse, IFN-regulated transcriptional activator STAT2 and targeted it for proteasomal degradation. This phenomenon may explain the requirement for IFN deficiency in order to observe ZIKV pathogenesis in mice. Furthermore, the mechanism of ZIKV NS5 resembles that of DENV NS5, but not that of its closer relative Spondweni virus (SPOV). However, unlike DENV NS5, ZIKV NS5 did not require the E3 ubiquitin ligase UBR4 to induce STAT2 degradation. Consequently, flavivirus NS5 proteins exhibit a remarkable functional convergence in IFN antagonism, albeit by virus-specific mechanisms. The potent antagonism of human IFN responses by neglected flaviviruses such as SPOV and Usutu virus (USUV), coupled with similar ecologies to that of known human flavivirus pathogens, suggests their potential for broad emergence into the human population.
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Osório, Hugo Costa. "West nile virus in Portugal: vector population, host interaction and detection of new flaviviruses." Doctoral thesis, Universidade de Évora, 2013. http://hdl.handle.net/10174/9043.
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This thesis describes the studies undertaken in the epidemiology of West Nile virus (WNV) in Portugal exploring several key components of its natural cycle: identification of mosquito vector-populations – geographical and seasonal distribution; analysis of genetic and environmental determinants on feeding patterns of mosquito vectors; experimental transmission of WNV; identification of flaviviruses detected in wild mosquitoes; serological survey of wild birds potentially involved in the epizootic cycle of WNV; finally, a description of a clinical human case of WNV infection reported in 2010.
Culex pipiens was one of the most abundant mosquito species demonstrating ecological and intrinsic competence to transmit WNV. No positive mosquito pools for WNV were detected, but new mosquito-only flaviviruses were identified. The presence of antibodies anti-WNV in a juvenile turtle-dove suggests local virus circulation.
In conclusion, Portugal meets suitable conditions for epizootic circulation of WNV and for the occurrence of accidental human infections; RESUMO
O vírus West Nile em Portugal
Vetores, interação com hospedeiros e deteção de novos flavivírus
Esta tese descreve os estudos desenvolvidos na epidemiologia do vírus West Nile (VWN) em Portugal explorando várias componentes do seu ciclo natural: identificação das populações de mosquitos vetores – distribuição geográfica/ sazonal; análise de determinantes genéticos/ ambientais na preferência de hospedeiro de mosquitos vetores; transmissão experimental do VWN; identificação de flavivírus detectados em mosquitos selvagens; estudo serológico em aves potencialmente reservatórios do vírus; finalmente é descrito um caso clínico humano de infecção por VWN detetado em 2010.
Culex pipiens foi uma das espécies de mosquito mais abundantes e demonstrou competência ecológica e intrínseca para transmitir o VWN. Não foram detectados pools de mosquitos positivos para o VWN, mas foram identificados novos flavivírus específicos de insetos. A presença de anticorpos anti-WNV numa rola juvenil sugere circulação local do vírus.
Em conclusão, Portugal reúne condições adequadas para a circulação epizoótica do VWN e para a ocorrência de infecções humanas acidentais.
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Li, Dongsheng. "The role of HCV core protein in the regulation of HCV replication /." St. Lucia, Qld, 2003. http://www.library.uq.edu.au/pdfserve.php?image=thesisabs/absthe18009.pdf.
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Grillo, Elena. "Presence of Haemosporidia and Flaviviruses in Breeding Prothonotary Warblers (Protonotaria citrea): An Analysis of Spatial and Temporal Trends in Infection Prevalence and Associations with Reproductive Success." VCU Scholars Compass, 2009. http://scholarscompass.vcu.edu/etd/1857.
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As Neotropical migratory birds, Prothonotary Warblers are exposed to parasites in both tropical and temperate regions and may act as dispersal agents between geographic areas. This study identifies the prevalence of Haemosporidia, West Nile Virus (WNV), and St. Louis Encephalitis virus (SLEV) in this species. A total of 71.6% of captured Prothonotary Warblers were infected with Haemosporidia during the 2008 breeding season, and infection prevalence increased throughout the season. This temporal change in prevalence is likely due to infection relapse and transmission of new infections. No correlations between reproductive effort and infection status were observed, nor were any associations between infection prevalence and nest box location identified. WNV and SLEV were present in 37.5% and 6.3% of sampled Prothonotary Warblers, respectively. These results warrant more detailed analyses of pathogen transmission dynamics in this population, physiological mechanisms that affect infection susceptibility, and spatial and temporal trends in infection that may exist.
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Davis, William G. "Protein binding sites and cis-acting sequences on the West Nile Virus 3' (+) SL RNA." unrestricted, 2007. http://etd.gsu.edu/theses/available/etd-07262007-134423/.
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Thesis (Ph. D.)--Georgia State University, 2007.Title from file title page. Margo Brinton, committee chair; W. David Wilson, Teryl Frey, committee members. Electronic text (120 p. : ill. (some col.)) : digital, PDF file. Description based on contents viewed Nov. 20, 2008. Includes bibliographical references.
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Holicki, Cora [Verfasser], Martin H. [Akademischer Betreuer] Groschup, Martin H. [Gutachter] Groschup, and Stefanie [Gutachter] Becker. "Mosquito-borne Flaviviruses: Vector and Avian Host Susceptibility for West Nile Virus and Usutu Virus in Germany / Cora Marielle Holicki ; Gutachter: Martin H. Groschup, Stefanie Becker ; Betreuer: Martin H. Groschup." Hannover : Stiftung Tierärztliche Hochschule Hannover, 2020. http://d-nb.info/122488289X/34.
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Pascoal, Jamile de Oliveira. "Patógenos intracelulares em carrapatos do Cerrado e Mata Atlântica: vírus e riquétsias." Universidade Federal de Uberlândia, 2017. https://repositorio.ufu.br/handle/123456789/19614.
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CAPES - Coordenação de Aperfeiçoamento de Pessoal de Nível SuperiorCNPq - Conselho Nacional de Desenvolvimento Científico e Tecnológico
No Brasil, a maioria das viroses transmitidas por artrópodes (arboviroses) pertence ao gênero Flavivirus, no entanto, aqueles transmitidos por carrapatos são menos estudados do que os transmitidos por insetos. Além dos vírus, outros microrganismos possuem grande relevância em saúde pública. Riquétsias são os principais agentes de zoonoses transmitidas por carrapatos no Brasil, especialmente as do Grupo da Febre Maculosa (GFM), que inclui diversas espécies patogênicas e que são transmitidas por carrapatos. Dentre os diversos biomas, dois foram particularmente degradados no Brasil, a Mata Atlântica e o Cerrado. Diante destes motivos, esta tese compõe-se de três capítulos, que tiveram como objetivo identificar carrapatos vetores em áreas do Cerrado e Mata Atlântica adjacentes a áreas com atividade humana, pesquisando por patógenos causadores de zoonoses, Rickettsia spp. e Flavivirus. No primeiro capítulo, a presença de riquétsias foi averiguada em carrapatos de cães e carnívoros da região de Cumari-GO. Pesquisou-se por esse microrganismo em oito espécies de carnívoros silvestres e em cães domésticos. Duas amostras amplificaram fragmentos do gene ompB, presente em riquétsias do GFM, e ao mesmo tempo amplificaram fragmentos específicos para Rickettsia bellii, que não pertence a esse grupo. Isso demonstra uma possível infecção cruzada, podendo o carrapato manter concomitantemente estas duas espécies divergentes. O segundo capítulo, dados de carrapatos de tamanduás (Myrmecophaga tridactyla e Tamandua tetradactyla) referentes a um período de 18 anos foram analisados, 169 amostras foram avaliadas quanto à presença de riquétsias. Quatro destas amostras exibiram sequência de ompA com 100% de identidade com outras sequências de carrapato Amblyomma nodosum, indicando que Rickettsia spp. do grupo da febre maculosa (SFG) em Amblyomma nodosum estava circulando no entorno da cidade de Uberlândia e estado de São Paulo, podendo outros carrapatos, tais como A. sculptum serem infectados, gerando risco para animais domésticos e seres humanos. No último capítulo, carrapatos de seis áreas, uma pertencente à Mata Atlântica e as outras do bioma Cerrado, foram avaliados quanto à presença de Flavivirus. Nestes locais foram pesquisados carrapatos de doze espécies diferentes, sendo onze da família Ixodidae e um da família Argasidae, estes pertenciam às espécies: Amblyomma sculptum, Rhipicephalus sanguineus, Rhipicephalus microplus, Dermacentor nitens, Amblyomma ovale, Amblyomma dubitatum, Amblyomma parvum, Amblyomma rotundatum, Amblyomma incisum, Amblyomma brasiliense, Amblyomma naponense e Ornithodoros sp. Detectou-se a presença de um flavivírus em carrapatos R. microplus coletados na região de Uberlândia. Estas amostras exibiram aproximadamente 98% de identidade com o Vírus do Carrapato Mogiana (MGTV). Todos os carrapatos que exibiram resultados positivos para flavivírus ou riquétsias, foram coletados em áreas antropizadas, confirmando a importância do estudo de vetores hematófagos na epidemiologia de agentes virais e bacterianos de animais domésticos e selvagens.
In Brazil, most arthropod-transmitted viruses (arboviruses) belong to Flavivirus genus, being tick-transmitted less studied than insect-transmitted ones. Other microorganisms have great public health relevance, such as Rickettsia, the main agent of tick-borne diseases in Brazil, specially the Macular Fever (MF) group, which includes several pathogenic species that are transmitted by ticks. Within different biomes, both the Atlantic Forest and the Cerrado were severely degraded in Brazil. This thesis consisted of three chapters, aiming to identify ticks in areas of Cerrado and Atlantic Forest with surrounding human activity, looking for both Rickettsia and Flavivirus infected ticks. In the first chapter, the presence of Rickettsia was investigated in ticks from dogs and other carnivores, being eight wild carnivores species and domestic dogs in Cumari, Goiás state, Brazil. Two samples were positive for ompB gene, common in the MF group, being also positive for specific genes found in Rickettsia bellii, which does not belong to the MF group. This result shows a possible cross-infection, being tick able to maintain these two different species simultaneously. The second chapter, an 18- year-study-data from anteater ticks (Myrmecophaga tridactyla and Tamandua tetradactyla) was analyzed, accounting 169 samples evaluated for the presence of Rickettsia. Four samples were positive for ompA gene, being 100% identical to other tick sequences present in Amblyomma nodosum, indicating that Rickettsia from the MF group in A. nodosum was circulating around the city of Uberlândia and state of São Paulo. Other ticks, such as A. sculptum, could become infected by the Rickettsia presence, generating risk for domestic animals and humans. In the last chapter, ticks from six areas, being one from the Atlantic Forest and the others from the Cerrado biome, were evaluated for the presence of Flavivirus. 12 different species were accounted, in which eleven were from the Ixodidae family and one was from the Argasidae family, having the following species identified: Amblyomma sculptum, Rhipicephalus sanguineus, Rhipicephalus microplus, Dermacentor nitens, Amblyomma ovale, Amblyomma dubitatum, Amblyomma parvum, Amblyomma rotundatum, Amblyomma incisum, Amblyomma brasiliense, Amblyomma naponense and Ornithodoros sp. The presence of a Flavivirus in R. microplus ticks collected in the Uberlândia region was detected. These samples exhibited approximately 98% identity with the Mogiana Tick Virus (MGTV). All ticks that were positive for Flavivirus or Rickettsia were collected in areas with human activities, stating the importance of studies on hematophagous vectors in the epidemiology of both viral and bacterial agents in domestic and wild animals.
Tese (Doutorado)
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Haller, Logan C. "Development of a micropshere-based immunoassay for the detection of IgM antibodies to West Nile virus and St. Louis Encephalitis virus in sentinel chicken sera." [Tampa, Fla] : University of South Florida, 2006. http://purl.fcla.edu/usf/dc/et/SFE0001505.
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Pliego-Zamora, Adriana C. "The Antiviral Properties of Melaleuca alternifolia concentrate (MAC) against West Nile virus." Thesis, Griffith University, 2015. http://hdl.handle.net/10072/367888.
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Many Flaviviruses are mosquito-borne viruses that can causes disease in humans and animals including Dengue virus (DENV), West Nile virus (WNV), yellow fever virus (YFV), Japanese encephalitis virus (JEV) and tick-borne encephalitis virus (TBEV). These viruses are highly pathogenic to humans causing extensive morbidity and mortality and most importantly, their recent re-emergence and global spread is a current growing public health issue. Currently, DENV alone causes an approximate 50 million infection cases annually and WNV infections have been reported in the five continents. Despite the existence of licensed vaccines for YFV, JEV and TBEV, a major challenge remains given there are no specific and effective antiviral therapies against flaviviral infections.
Considering the homologies between flaviviral replication strategies, it may be possible to identify broad-spectrum compounds with both prophylactic and therapeutic activity against different flavivirus strains. Because the steps involved in compound discovery to their clinical use are protracted and expensive, studies should include testing existing clinically approved drugs as antivirals. Melaleuca alternifolia extracts are good candidates to be explored for novel uses as they have been extensively studied and their composition, chemistry, safety and toxicity are well documented. Melaleluca alternifolia concentrate (MAC) is obtained from the further purification of Tea Tree Oil (TTO) and is a new product exhibiting promising antimicrobial and anti-cancer activities.Thesis (PhD Doctorate)
Doctor of Philosophy (PhD)
School of Medical Science
Griffith Health
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Khou, Cécile. "Etude du neurotropisme des Flavivirus neuropathogènes." Thesis, Sorbonne Paris Cité, 2017. http://www.theses.fr/2017USPCC305/document.
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Les Flavivirus neuropathogènes, tels que le virus de l’encéphalite japonaise (JEV), le virus West Nile (WNV), le virus de la fièvre jaune (YFV) et le virus Zika (ZIKV) causent des maladies neurologiques. Ces maladies sont dues à une infection des cellules du système nerveux central (CNS) par ces virus. Le CNS est un organe privilégié, isolé des agents pathogènes par une barrière entre le sang et le cerveau, appelée barrière hémato-encéphalique (BBB). Les Flavivirus neuropathogènes capables de traverser cette BBB afin d’atteindre leurs cellules cibles, localisées dans le CNS, sont neuroinvasifs. Le but de cette étude est de comprendre les mécanismes cellulaires permettant aux Flavivirus de traverser la BBB et les effets de l’infection par les virus ZIKV et WNV des cellules du CNS sur le développement de celles-ci.Le YFV est un virus hépatotrope, infectant majoritairement le foie et les reins. Deux vaccins vivants atténués dirigés contre le YFV, le vaccin FNV (pour French Neurotropic Virus) et le vaccin 17D, ont été obtenus empiriquement par passages successifs de souches virulentes de YFV sur cerveaux de souriceaux. Ces vaccins ne causent plus de maladies touchant les reins et le foie, mais peuvent parfois causer des encéphalites post-vaccinales. Ces cas d’encéphalites démontrent que ces souches vaccinales sont devenues neurovirulentes mais aussi neuroinvasives car les virus ont pu franchir la BBB. A cause d’une incidence trop élevée d’encéphalites post-vaccinales par rapport au vaccin 17D, le vaccin FNV a été retiré du marché dans les années 1980.Le JEV est un virus neurotrope, causant des encéphalites graves en Asie du Sud-Est. A ce jour, il existe un vaccin vivant atténué, le JEV SA14-14-2, obtenu empiriquement par passages successifs d’une souche virulente sur cellules de hamster. Ce vaccin est moins neurovirulent et moins neuroinvasif que les souches virulentes de JEV en modèle de souris, et protège contre des infections humaines par le JEV. Cependant, des cas d’encéphalites ont été rapportés après injection de ce vaccin. Il apparait donc que, dans certains cas, la souche vaccinale JEV SA14-14-2 est capable de traverser la BBB et d’infecter les cellules neuronales. Les dernières épidémies à virus ZIKV en Polynésie Française et en Amérique du Sud ont induit une augmentation de cas de malformations congénitales dans les zones touchées. Cela a soulevé de nouvelles questions quant à la capacité d’un Flavivirus à provoquer des malformations congénitales du CNS. Dans cette étude, nous avons identifié les mécanismes cellulaires permettant aux Flavivirus de traverser la BBB et les effets de l’infection par les virus ZIKV et WNV des cellules du CNS sur le développement de celles-ci.Nous avons utilisé deux systèmes in vitro permettant d’étudier le développement du CNS et la neuroinvasion des Flavivirus. Un premier système consiste en l’infection de coupes de cerveaux d’embryon de souris. En utilisant ce système, nous avons montré que le ZIKV a un tropisme préférentiel pour les cellules progénitrices de neurones, alors que le WNV a un tropisme préférentiel pour les neurones. Nous avons également montré que l’infection des progéniteurs neuronaux par le ZIKV induit un arrêt de la mitose cellulaire, alors que l’infection par le WNV n’a aucun effet sur la mitose. L’étude sur l’effet apoptotique de l’infection par les deux virus WNV et ZIKV n’a montré aucune différence entre les deux virus à des temps précoces d’infection.Un deuxième système a été mis au point pour l’étude de la neuroinvasion par les Flavivirus neuropathogènes. Ce système est composé de cellules endothéliales hCMEC/D3 pouvant former des jonctions serrées. Ces cellules ont été cultivées sur filtres d’insert de puits de culture cellulaire Transwell, placés au-dessus de cellules neuronales humaines. A l’aide de ce système, nous avons comparé la capacité à traverser la BBB de plusieurs FlavivirusNeuropathogenic Flaviviruses, such as Japanese encephalitis virus (JEV), West Nile virus (WNV), yellow fever virus (YFV) and Zika virus (ZIKV), cause neurological diseases. These diseases are due to viral infection of central nervous system (CNS) cells. The CNS is a privileged organ, isolated from pathogenic agents by a barrier between the blood and the barrier, called the blood-brain barrier (BBB). Neuropathogenic Flaviviruses which can cross this BBB in order to reach their target cells in the CNS, are neuroinvasive. This study aims at understanding the cellular mechanisms by which YFV and JEV Flaviviruses cross the BBB and the effects of viral infection by WNV and ZIKV of the CNS cells during neocortex development.YFV is a hepatrotopic virus, which mostly infects the liver and the kidneys. The two live-attenuated vaccines against YFV, the FNV (for French Neurotropic Virus) vaccine and the 17D vaccine, were obtained empirically by several passages in suckling mouse brain of YFV virulent strains. These vaccines do not cause any disease targeting the liver or the kidneys, but can sometimes cause post-vaccine encephalitis. These encephalitis cases suggest that the vaccine strains have become neurovirulent and neuroinvasive. Due to high risks of post-vaccine encephalitis, the FNV vaccine use was discontinued in the 1980s.JEV is a neurotropic virus, causing acute encephalitis in South East Asia. To date, there is a live-attenuated vaccine against JEV, the JEV SA14-14-2 vaccine, which was obtained empirically by several passages in primary hamster kidney cells. This vaccine is less neurovirulent and less neuroinvasive than JEV virulent strains in mouse model, and it protects against JEV infections. However, some cases of post-vaccine encephalitis were reported. It thus seems that, in some cases, the vaccine strain JEV SA14-14-2 is able to cross the BBB and infect neuronal cells.The recent ZIKV epidemics in French Polynesia and South America were linked to an increase in the number of congenital malformations, rising questions regarding the capacity of a Flavivirus to induce CNS congenital malformations.In this study, we have identified cellular mechanisms involved in Flavivirus neuroinvasion and studied the effect of ZIKV and WNV infection of neuronal cells under development.To study CNS development, we have infected mouse embryos brain slices. We were able to show that ZIKV has a preferential tropism for neuronal progenitors, whereas WNV has a preferential tropism for neuronal cells. We also show that infection of neuronal progenitors by ZIKV impairs the cell life cycle, whereas no effect on the cell life cycle was observed for WNV-infected cells. Studies on apoptosis induction did not show any difference between both viruses at early time points of infection.To study Flavivirus neuroinvasion, we have used an in vitro model of BBB composed of human endothelial hCMEC/D3 cells that can form tight junctions. These cells were cultivated on Transwell inserts and placed above human neuronal cells. Using this system, we show that YFV FNV cross the BBB more efficiently than YFV 17D, suggesting that YFV FNV is more neuroinvasive than YFV 17D. This observation can explain the higher post-vaccine encephalitis risks associated with YFV FNV vaccine compared to YFV 17D vaccine. We also confirmed that JEV SA14-14-2 vaccine strain is less neuroinvasive than JEV RP9.We also examined how JEV crosses the BBB and the endothelial cell response following JEV treatment. We show that both JEV RP9 and SA14-14-2 are able to cross the BBB without infecting its endothelial cells and without disrupting the BBB. Preliminary results suggest that JEV RP9, but not JEV SA14-14-2, crosses the BBB by dynamin-dependant transcytosis. Transcriptomic analysis of endothelial cells treated by either virus show slight, but significant, differences in regulation of genes implicated in several pathways associated with CNS diseases
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Lequime, Sébastian. "Interactions flavivirus-moustiques : diversité et transmission." Thesis, Paris 6, 2016. http://www.theses.fr/2016PA066081/document.
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Les flavivirus sont des virus à ARN parmi lesquels certains sont des arbovirus transmis entre hôtes vertébrés par des vecteurs arthropodes, notamment des moustiques. L'interaction avec les moustiques est centrale dans la biologie des flavivirus par son influence sur leur diversité génétique et transmission, mais certains de ses aspects restent méconnus. Au cœur de cette thèse, des approches basées sur les « big data », générées par des technologies modernes ou par compilation de travaux plus anciens, ont éclairé d’un jour nouveau la complexité des relations moustique-flavivirus. En explorant des génomes de moustiques anophèles, nous avons identifié et caractérisé des éléments viraux endogènes d'origine flavivirale chez Anopheles sinensis et An. minimus, suggérant l'existence de flavivirus infectant les anophèles et révélant une facette insoupçonnée de leur diversité. Par ailleurs, nous avons exploré, par séquençage haut-débit, la fine interaction entre le génotype du moustique Aedes aegypti et la diversité intra-hôte du virus de la dengue-1. Nos résultats montrent un fort effet de la dérive génétique lors de l'infection initiale, diminuant l'importance relative de la sélection naturelle, et une modulation de la diversité génétique intra-hôte du virus par le génotype du moustique. Enfin, nous avons compilé la littérature sur la transmission verticale des arbovirus chez les moustiques, c'est-à-dire de la femelle infectée à sa descendance, afin d'identifier des facteurs techniques et biologiques sous-jacents. Nos résultats améliorent la compréhension de ce mode de transmission et des stratégies employées par les arbovirus pour persister dans l’environnementFlaviviruses are RNA virus among which some are arboviruses transmitted between vertebrate hosts and arthropod vectors, like mosquitoes. The interaction with mosquitoes is key in the biology of flaviviruses because it influences their genetic diversity and transmission. However, some aspects however are still poorly understood. At the heart of the work presented in this dissertation, strategies based on ‘big data’, both by taking advantage of modern technologies and by compiling older literature, highlighted new aspects of the complex relationships between flaviviruses and mosquitoes. While exploring Anopheles mosquito genomes, we identified and characterized endogenous viral elements of flaviviral origin in Anopheles sinensis and An. minimus, which supports the existence of flaviviruses infecting Anopheles mosquitoes and highlights new aspected of their diversity. Besides, we explored, by deep sequencing, the fine-tuned interaction between genotypes of the mosquito Aedes aegypti and the intra-host diversity of dengue virus 1. Our results showed a strong effect of genetic drift during initial infection, reducing the relative importance of natural selection, and a modulation of the intra-host viral genetic diversity by the mosquito genotype. Finally, we assembled the litterature on arbovirus vertical transmission in the mosquito vector, i.e. from an infected female to her offspring, in order to identify underlying technical and biological predictors. Our results increase our understanding of this transmission mode and the strategies employed by arboviruses to persist in their environment
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Lequime, Sébastian. "Interactions flavivirus-moustiques : diversité et transmission." Electronic Thesis or Diss., Paris 6, 2016. https://accesdistant.sorbonne-universite.fr/login?url=https://theses-intra.sorbonne-universite.fr/2016PA066081.pdf.
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Les flavivirus sont des virus à ARN parmi lesquels certains sont des arbovirus transmis entre hôtes vertébrés par des vecteurs arthropodes, notamment des moustiques. L'interaction avec les moustiques est centrale dans la biologie des flavivirus par son influence sur leur diversité génétique et transmission, mais certains de ses aspects restent méconnus. Au cœur de cette thèse, des approches basées sur les « big data », générées par des technologies modernes ou par compilation de travaux plus anciens, ont éclairé d’un jour nouveau la complexité des relations moustique-flavivirus. En explorant des génomes de moustiques anophèles, nous avons identifié et caractérisé des éléments viraux endogènes d'origine flavivirale chez Anopheles sinensis et An. minimus, suggérant l'existence de flavivirus infectant les anophèles et révélant une facette insoupçonnée de leur diversité. Par ailleurs, nous avons exploré, par séquençage haut-débit, la fine interaction entre le génotype du moustique Aedes aegypti et la diversité intra-hôte du virus de la dengue-1. Nos résultats montrent un fort effet de la dérive génétique lors de l'infection initiale, diminuant l'importance relative de la sélection naturelle, et une modulation de la diversité génétique intra-hôte du virus par le génotype du moustique. Enfin, nous avons compilé la littérature sur la transmission verticale des arbovirus chez les moustiques, c'est-à-dire de la femelle infectée à sa descendance, afin d'identifier des facteurs techniques et biologiques sous-jacents. Nos résultats améliorent la compréhension de ce mode de transmission et des stratégies employées par les arbovirus pour persister dans l’environnementFlaviviruses are RNA virus among which some are arboviruses transmitted between vertebrate hosts and arthropod vectors, like mosquitoes. The interaction with mosquitoes is key in the biology of flaviviruses because it influences their genetic diversity and transmission. However, some aspects however are still poorly understood. At the heart of the work presented in this dissertation, strategies based on ‘big data’, both by taking advantage of modern technologies and by compiling older literature, highlighted new aspects of the complex relationships between flaviviruses and mosquitoes. While exploring Anopheles mosquito genomes, we identified and characterized endogenous viral elements of flaviviral origin in Anopheles sinensis and An. minimus, which supports the existence of flaviviruses infecting Anopheles mosquitoes and highlights new aspected of their diversity. Besides, we explored, by deep sequencing, the fine-tuned interaction between genotypes of the mosquito Aedes aegypti and the intra-host diversity of dengue virus 1. Our results showed a strong effect of genetic drift during initial infection, reducing the relative importance of natural selection, and a modulation of the intra-host viral genetic diversity by the mosquito genotype. Finally, we assembled the litterature on arbovirus vertical transmission in the mosquito vector, i.e. from an infected female to her offspring, in order to identify underlying technical and biological predictors. Our results increase our understanding of this transmission mode and the strategies employed by arboviruses to persist in their environment
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Grard, Gilda. "Génomique et évolution des flavivirus transmis par les tiques et découverte d'un nouveau lignage du genre flavivirus." Aix-Marseille 2, 2006. http://www.theses.fr/2006AIX20679.
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Gollins, S. W. "Mechanisms of flavivirus neutralization and cellular infection." Thesis, University of Oxford, 1985. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.355752.
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Dayaraj, Cecilia. "Molecular and immunological studies on flavivirus virulence." Thesis, University of Oxford, 1991. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.279888.
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Carney, Jennifer. "Viral Determinants of Flavivirus Neurotropism in Humans." Thesis, University of Liverpool, 2009. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.526956.
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Pacca, Carolina Colombelli. "Screening de novos antivirais inibidores de flavivirus." Faculdade de Medicina de São José do Rio Preto, 2013. http://bdtd.famerp.br/handle/tede/201.
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Introduction. Arboviruses, arthropod-borne viruses, are frequently associated with human outbreaks and represent a serious health problem. The genus Flavivirus, which includes both the Yellow Fever Virus (YFV) and Saint Louis Encephalitis Virus (SLEV), are important pathogens that result in high morbidity and mortality rates worldwide. In Brazil, YFV has a sylvatic cycle and occurs annually, despite the efficiency of the vaccine. Saint Louis Encephalitis is an infectious illness that can cause acute fever caused by SLEV, which is widely distributed in the Americas. The emergence of SLEV became a serious concern after the first related outbreak in Brazil in 2006, in the city of Sao Jose do Rio Preto. There is no specific antiviral drug for these viruses, only supporting treatment that can alleviate the symptoms and prevent complications. The need to develop effective and safe antiviral drugs is indispensable for the treatment of these infections. Objective. The aim of this work was to identify new possible antiviral drugs against the arboviruses that can cause acute fever and encephalitis (YFV and SLEV) and to evaluate the capacity of inhibition of these compounds in ABR mice. Material and Methods. Plaque reduction assay, flow citometry, immunofluorescence and cellular viability were used to test the compounds in vitro. ABR mice were inoculated with YFV, and the biological samples were tested for the presence of the virus through the use of plaque reduction assay and qPCR. Neutralization assay was also performed. Results. Treated cells showed efficient inhibition of viral replication at concentrations that presented minimal toxicity to the cells. The assays showed that ftalyl-tiazole and fenoxytiosemicarbazone were more effective, and that they reduced viral replication by 60% and 75% for YFV and SLEV, respectively. The analysis also revealed that the ABR mice inoculated with YFV had histopathological alterations in the liver; however, the samples did not present viral title. Neutralization assay showed a high concentration of antibodies in the serum. Conclusion. The inhibitions of viral replication were confirmed through the use of some assays in vitro, and the effectiveness of the selected compounds show that they are an option in the treatment of these viruses. More detailed studies are needed to determine the mechanism of action of these molecules. The mice were found to have histopathological alterations, which indicates viral infection; however, they also presented with high concentrations of antibodies. More studies about animal models are necessary to make in vivo experiments.
Introdução: Os arbovírus, vírus transmitidos por artrópodes, são freqüentemente associadas a surtos em seres humanos e representam um problema sério de saúde pública. Os vírus pertencentes ao gênero Flavivirus, tais como vírus da Febre Amarela (YFV) e vírus da Encefalite de Saint Louis (SLEV), são importantes patógenos que podem causar alta taxa de morbidade e mortalidade no mundo. No Brasil, YFV é mantido em ciclo silvestre notificados anualmente, a despeito da segurança e eficiência da vacina. A encefalite de Saint Louis é uma doença infecciosa febril aguda causada pelo SLEV amplamente distribuída nas Américas. A emergência do SLEV passou a ser um fato preocupante no Brasil a partir da constatação do primeiro surto no país em 2006, na cidade de São Jose do Rio Preto. Não existe tratamento específico para estas viroses, somente tratamento de suporte para ajudar a aliviar os sintomas e prevenir complicações. Desta forma, há uma grande necessidade de que sejam desenvolvidos antivirais efetivos e seguros para o tratamento destas infecções. Objetivos: O objetivo deste trabalho foi identificar potenciais compostos antivirais contra os arbovírus causadores de doença febril aguda e encefalites (YFV e SLEV) in vitro e avaliar a capacidade de inibição da replicação viral dos compostos in vivo em camundongos ABR. Materiais e Métodos: Para tanto, foram realizados ensaios de redução de placas, citometria de fluxo, imunofluorescencia, bem como testes de viabilidade celular para as analises in vitro. Além disto, camundongos ABR foram inoculados com YFV e seus materiais biológicos testados para a presença de partículas virais por ensaio de redução de placas e qPCR. Adicionalmente, foi realizado ensaio de neutralização do soro dos animais. Resultados: Celulas tratadas com os compostos mostraram eficiente inibição da replicação viral em concentrações que apresentam baixa citotoxicidade. Os ensaios mostraram que derivados de ftalyl-tiazole e fenoxytiosemicarbazone foram os mais eficazes na ação antiviral, apresentando redução de 60% e 75% para YFV e SLEV, respectivamente. Camundongos ABR inoculados com YFV apresentaram alterações histológicas no fígado, entretanto, não foi constatado título viral nas amostras testadas. O ensaio de neutralização mostra altas concentrações de anticorpos no soro dos animais. Conclusões: A inibição da replicação foi comprovada por vários ensaios in vitro evidenciando as moléculas como potentes alternativas para o tratamento dos vírus. Mais estudos são necessários para a determinação do mecanismo de ação destas moléculas. Os camundongos apresentaram alterações histopatológicas sendo um indicativo de infecção, entretanto, apresentam altas taxas de anticorpos. Mais estudos sobre modelo animal são necessários para a realização de ensaios in vivo.
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Singethan, Katrin. "Untersuchungen zur Inhibition Paramyxo- und Flavivirus-induzierter Membranfusion." kostenfrei, 2009. http://www.opus-bayern.de/uni-wuerzburg/volltexte/2009/3634/.
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Cunha, Mariana Sequetin. "Validação e uso de transcrição reversa seguida da reação em cadeia pela polimerase em tempo real (RT-qPCR) para a vigilância e diagnóstico de flavivírus transmitidos por mosquitos circulantes no Brasil." Universidade de São Paulo, 2018. http://www.teses.usp.br/teses/disponiveis/10/10133/tde-16102018-113026/.
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Os flavivírus são considerados uma séria ameaça à saúde pública em diversas partes do mundo, pois muitos são agentes altamente patogênicos a seres humanos e animais, tais como os vírus da febre amarela, vírus do Oeste do Nilo, vírus da encefalite japonesa e vírus da dengue, capazes de causar encefalites ou febres hemorrágicas em seus hospedeiros. Muitos deles têm avançado a diferentes regiões geográficas onde sua circulação não havia sido detectada previamente, causando novos surtos. O diagnóstico clínico destas infecções é, muitas vezes, difícil, devido ao grande número de sintomas apresentados, que podem se confundir com outras enfermidades de diferentes causas etiológicas. Os principais métodos diretos utilizados atualmente no Brasil para detecção destes vírus são a inoculação intracerebral em camundongos neonatos, inoculação em culturas de células e RTPCR específica. O presente trabalho tem como objetivos avaliar a sensibilidade e validar a detecção dos vírus pertencentes ao gênero Flavivirus circulantes no Brasil por meio de uma reação single de RT-PCR em tempo real e implementá-la, tanto na rotina diagnóstica de casos com suspeita de arbovirose como na pesquisa de amostras de campo para monitoramento viral. Amostras dos flavivírus padrões da Febre Amarela, Bussuquara, Iguape, Ilheus, Encefalite de Saint Louis, Cacipacore e Zika foram quantificados por titulação em unidades formadoras de placa (UFP) ou TCID50 para se avaliar os limites de detecção para cada um deles por RT-qPCR que detecta o gênero Flavivirus. Os limites encontrados variaram de 0,01 UFP, para o vírus Ilheus, a 1 UFP, para os vírus da Febre Amarela e Iguape, e 1x101,6 TCID50/100µL para o vírus Bussuquara. Além disso, o presente trabalho foi capaz de identificar, após sequenciamento de cDNA gerado, os vírus Zika, isolado de um paciente febril, e os vírus Ilheus e Iguape, isolados a partir de diferentes espécies de Culicídeos, após uma única reação de RT-qPCR, e um possível novo flavivírus específico de insetos, isolado de mosquitos Aedes coletados em Guapiaçu, São Paulo. Não houve sinal de amplificação para os Alphavirus Mayaro e Chikungunya. O presente protocolo mostrou-se com alta sensibilidade e especificidade, podendo dessa forma ser utilizado para o diagnóstico diferencial dos diferentes flavivírus que ocorrem no Brasil, bem como para estudos de monitoramento viral em animais sentinelas e vetores, colaborando dessa forma com a saúde pública. Pode-se, ainda, detectar possíveis novos vírus específicos de artrópodesFlaviviruses are considered a serious threat to public health in many parts of the world, as many are highly pathogenic to humans and animals, such as Yellow Fever virus, West Nile virus, Japanese encephalitis virus and dengue virus, which are capable of causing encephalitis or hemorrhagic fever in their hosts. Many of them have spread to different geographic regions where their circulation had not been detected previously, causing new outbreaks. Diagnosis of these infections is often difficult, due to the large number of symptoms presented, which can be confused with other diseases of different etiological causes. The main direct methods currently used in Brazil for detecting these viruses are intracerebral inoculation in neonatal mice, inoculation in cell cultures and specific RT-PCR. The present work aims to evaluate the sensitivity and validate the detection of viruses belonging to the genus Flavivirus circulating in Brazil through a single real-time RT-PCR reaction and to implement it, both in the diagnostic routine of cases with arbovirus suspicions and in field samples for viral monitoring. Samples of the standard flaviviruses Yellow Fever, Bussuquara, Iguape, Ilheus, Saint Louis Encephalitis, Cacipacore and Zika were quantified by titration by plaque forming units (UFP) or TCID50 to evaluate the detection limits for each of them by RT- qPCR that detects genus Flavivirus. The limits found ranged from 0.01 PFU for Ilheus virus to 1 PFU for Yellow Fever and Iguape viruses and 1x101.6 TCID50 / 100L for the Bussuquara virus. In addition, the present work was able to identify, after cDNA sequencing Zika virus, isolated from a febrile patient, and both Ilheus and Iguape viruses, isolated from different species of Culicidae, and a possible new insect-specific flavivirus, isolated from Aedes mosquitoes collected in Guapiaçu, São Paulo. The Alphaviruses Mayaro and Chikungunya were not amplified. The present protocol shoed high sensitivity and specificity, and therefore it may may be used for the differential diagnosis of the different flaviviruses that occur in Brazil, as well as for viral monitoring studies in sentinel animals and vectors, thus collaborating with public health. It is also possible to detect new flavivirus that are arthopode-specific.
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Silva, Jaqueline Raymondi. "Pesquisa de infecções por Flavivírus da encefalite de Saint Louis, Rocio e Oeste do Nilo em cavalos, por inquérito sorológico e isolamento viral." Universidade de São Paulo, 2010. http://www.teses.usp.br/teses/disponiveis/17/17147/tde-23062010-150025/.
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Arboviroses são grave problema de saúde pública no Brasil e destas destacam-se aquelas causadas por Flavivírus, dos quais onze já foram descritos no Brasil. Destes, dois importantes em saúde pública, e que pertencem ao sorocomplexo da Encefalite Japonesa, são o vírus da encefalite de Saint Louis (SLEV) e o Rocio (ROCV). O vírus Oeste do Nilo (WNV), introduzido no continente americano em 1999, ainda não foi detectado no Brasil, contudo sua introdução é muito provável. Neste estudo, avaliou-se a circulação de SLEV, ROCV e WNV em cavalos, por tentativas de isolamento viral e inquérito soro-epidemiológico. As tentativas de isolamento viral, em 11 tecidos cerebrais de cavalos do estado da Paraíba, resultaram negativas. O inquérito sorológico, por IgG-ELISA tendo como antígeno peptídeos recombinantes do domínio III da proteína de envelope de SLEV, WNV e ROCV, foi utilizado em 753 soros de animais dos estados de São Paulo, Mato Grosso do Sul, Minas Gerais, Rio de Janeiro e Paraíba. Soros de 271 cavalos foram positivos para SLEV (35,98%), 254 para WNV (33,73%) e 144 para ROCV (19,12%). Portanto, o ELISA mostrou-se adequado, diagnosticando infecções prévias por estes Flavivírus. Também, observou-se intensa circulação destes vírus infectando cavalos nos locais de estudo. Ainda, obteve-se, pela primeira vez, evidencia de que WNV foi introduzido no Brasil e encontra-se a infectar cavalos nos estados pesquisados exceto Minas Gerais. Finalmente, o inquérito sorológico em cavalos mostrou-se uma abordagem adequada à vigilância das flaviviroses por SLEV, ROCV e WNV no Brasil.Arboviruses are a serious public health problem in Brazil and, from these, the most important are caused by Flavivirus. Eleven Flavivirus have been described in Brazil. Of these, Saint Louis Encephalitis Virus (SLEV) and Rocio Virus (ROCV) are major public health problems and belongs to the Japanese Encephalitis Serocomplex. West Nile Virus (WNV), introduced in the American continent in 1999, has not yet been detected in Brazil. In this study, it was evaluated the circulation of SLEV, WNV and ROCV in horses, by viral isolation attempts and a serosurvey. Viral isolation attempts were performed in 11 brain tissues of horses from Paraíba state with negative results. It was used for the serosurvey, an IgG-ELISA with recombinant peptides of domain III of SLEV, WNV and ROCV envelope protein as antigens. Sera from 753 animals from São Paulo, Mato Grosso do Sul, Minas Gerais, Rio de Janeiro and Paraíba states were tested, and 271 of them were positive for SLEV (35,98%), 254 for WNV (33,73%) and 144 for ROCV (19,12%). Therefore, this ELISA has been a suitable approach for diagnosis of ancient infections by these viruses. An intense circulation of flaviviruses infecting horses was observed in the study sites. Besides, it was found, for the first time, the presence of WNV in Brazil, infecting horses from all the studied states with the only exception of Minas Gerais. Finally, serosurvey in horses proved to be an appropriate approach for surveillance of Flavivirus infections by SLEV, WNV and ROCV.
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Araujo, Marina Reus Tassi de. "Expressão de proteínas recombinantes de vírus do gênero Flavivirus." reponame:Repositório Institucional da UFPR, 2010. http://hdl.handle.net/1884/22380.
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Gaunt, Michael W. "The epidemiology of louping ill virus and flavivirus evolution." Thesis, University of Oxford, 1997. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.363978.
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Lopes, Samuel Franco. "Identificação de Flavivirus em aves silvestres da Amazônia Central." Universidade Federal do Amazonas, 2011. http://tede.ufam.edu.br/handle/tede/4594.
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The Amazon region has the highest biodiversity on the planet, as well as the largest number of arboviruses isolated, mainly due to the great diversity of species of wild vertebrates and hematophagous arthropods. Among the arboviruses, Flavivirus stand out both for producing the highest number of infections and human diseases, such as the severity of these diseases. Moreover, it is known that birds act as reservoir of some poorly studied Flavivirus, such as: Ilheus, Saint Louis Encephalits, Rocio, Cacipacoré and Bussuquara. Despite intensive studies in the brazilian amazon, especially in certain areas of the state of Para, few epidemiological information about most of these viruses were obtained. In this study, a specie of Flavivirus has been identified in whole blood samples from wild birds, captured in-situ (Alter do Chão/PA) and ex-situ (Manaus) by polymerase chain reaction preceded by reverse transcription (RT- PCR) followed by Multiplex-Nested PCR (MN-PCR) tests for species-specific identification. Among the 189 samples, 7(4,23%) were suggestive of Ilheus virus. The diagnostic technique used was effective in identifying the genus Flavivirus in samples of wild birds presenting itself as practical, fast and secure for the identification of brazilian arboviruses. The circulation of enzootic viruses both in captive and free-living birds, increase the role of birds as host in the cycles of transmission of zoonoses.
A Amazônia apresenta os maiores índices de biodiversidade do planeta, assim como o maior número de arbovírus isolados, principalmente em função da grande diversidade de espécies de artrópodes hematófagos e vertebrados silvestres. Entre os arbovírus, destacam-se os Flavivírus tanto por produzirem o maior número de infecções e doenças humanas, como pela gravidade destas doenças. Além disso, sabe-se que as aves atuam como reservatório de alguns Flavivirus pouco estudados como: Ilhéus, Saint Louis, Rocio, Cacipacoré e Bussuquara. Apesar de estudos intensivos realizados na Amazônia brasileira, sobretudo em certas áreas do Estado do Pará, poucas informações epidemiológicas a respeito da maioria desses vírus foram obtidas. Nesse estudo, foi identificada uma espécie de Flavivirus em amostras de sangue total de aves silvestres, capturadas in-situ (Alter do chão/PA) e ex-situ (Manaus/AM) através da reação em cadeia de polimerase conjugada a transcrição reversa (RT-PCR) seguida por multiplex nested-PCR (MN-PCR) para testes de identificação espécie-específicos. Entre as 189 amostras analisadas, foram encontrados 7 (4,23%) amplicons sugestivos do vírus Ilhéus entre as amostras positivas. A técnica de diagnóstico utilizada mostrou-se eficaz para a identificação do gênero Flavivírus em amostras de aves silvestres apresentando-se como uma alternativa prática, rápida e segura para a identificação de arboviroses brasileiras. A circulação enzoótica dos vírus tanto em aves de cativeiro como de vida livre reforçam o papel das aves como hospedeiro nos ciclos de transmissão destas zoonoses.
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Moureau, Grégory. "Génomique des Flavivirus : Contribution à l'analyse taxonomique et phylogénique." Thesis, Aix-Marseille, 2012. http://www.theses.fr/2012AIXM5002.
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Les virus à ARN - à l'exception des rétrovirus - représentent plus de 200 pathogènes humains et/ou vétérinaires majeurs. Ils sont pour la plupart considérés comme émergents, durant les dernières années ils ont été retrouvés au-delà de leur territoire d'origine, dans de nouvelles régions du monde. Les plus connus de ces virus sont le virus du West Nile, le virus chikungunya, la grippe, le coronavirus SRAS, l'entérovirus 71 (agent responsable de la fièvre aphteuse), les virus de la dengue, l'hépatite C virus, le virus de la fièvre hémorragique de Crimée Congo, le virus de la vallée du Rift, l'encéphalite japonaise et plusieurs entérovirus humains. En terme de mortalité et morbidité ils sont à l'origine de plus 100 millions de cas par an et la menace a tendance à augmenter. Ces statistiques sont démoralisantes quand on considère les immenses progrès de la médecine et de la science durant ces dernières dizaines d'années. Les recherches présentées dans ma thèse portent sur le genre Flavivirus, au sein duquel on retrouve le virus de la fièvre jaune, un pathogène humain responsable d'épidémies majeures en Afrique et en Amérique latine durant les 300 à 400 dernières années. Ce virus est toujours responsable d'épidémies majeures en Afrique malgré un vaccin très bien toléré et des plus efficaces. On assiste au même scénario avec le virus de l'encéphalite japonaise en IndeWithout including the retroviruses, there are in excess of 200 RNA viruses that are recognised human and/or animal pathogens, many of which are considered to be emerging viruses because during recent years they have dispersed beyond their original territories causing epidemics in new regions of the World. The more well known of these emerging viruses include, West Nile virus, chikungunya virus, influenza virus, SARS coronavirus, EV71 virus (the aetiological agent of hand foot and mouth disease), dengue virus, hepatitis C virus, Crimean Congo haemorrhagic fever virus, Rift Valley fever virus, Japanese encephalitis virus and many human enteroviruses from a variety of genera. The combined global morbidity and mortality figures for these viruses add up to 100s of millions per year and the current trend appears to be upwards. This is a depressing statistic when one considers the amazing medical and scientific achievements that we have witnessed during the past decades. The studies described in my thesis were focused on the genus Flavivirus the type species of which is yellow fever virus, another terrifyingly virulent human pathogen that has caused so much suffering in Africa and Latin America during the past 300 to 400 years. This virus continues to cause major epidemics in Africa despite the availability of one of the safest and most effective vaccines with which to control infections due to yellow fever virus. Indeed similar comments can be made in the context of the flavivirus Japanese encephalitis virus in India
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Bos, Sandra. "Undestanding the viral molecular factors involved in Zika virus pathogenicity in humans." Thesis, La Réunion, 2019. http://www.theses.fr/2019LARE0005.
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Le virus Zika (ZIKV) est un phénomène épidémiologique sans précédent qui surprit le monde entier. Pendant de nombreuses années, il fut considéré comme un virus anodin responsable d’une poignée d’infections humaines, auto-limitées et bénignes, en Afrique et en Asie du Sud-est. Mais, après des décennies de propagation silencieuse, une première épidémie éclata en Micronésie en 2007 - tel un signal d'alarme. Quelques années plus tard, une soudaine épidémie de ZIKV de plus grande ampleur se déclara dans les îles du Pacifique avant d'atteindre le Brésil en 2015. Au cours de cette période, Zika fut associé à de graves complications neurologiques, mettant en évidence son fort potentiel pathogène pour l'homme. Depuis son émergence, plus de 80 pays et territoires ont été touchés par la pandémie de ZIKV, désormais reconnu comme un virus neurotrope et tératogène. L'association des souches contemporaines de ZIKV à des formes graves de maladie chez l'homme, qui n'ont jamais été signalées auparavant, a soulevé l'hypothèse d'une pathogénicité nouvellement acquise. Ainsi, mes travaux de doctorat visaient à déterminer si l'ampleur de l'épidémie actuelle pouvait en partie avoir été facilitée par des facteurs viraux qui auraient renforcé la fitness du ZIKV. À cette fin, mon projet de recherche s'est concentré sur l'identification des facteurs moléculaires viraux impliqués dans la pathogénicité du virus Zika chez l’homme à partir du développement de clones moléculairesZika virus (ZIKV) is an unprecedented epidemiological phenomenon which surprised the world. For many years, it was considered a trivial virus responsible for only a handful of human infections, self-limited and benign, in Africa and Southeast Asia. But then, after decades of silent spread, a first epidemic broke out in Micronesia in 2007 – like a warning signal. A few years later, a sudden Zika outbreak of larger scale occurred in the Pacific islands before reaching Brazil in 2015. During this period, Zika was associated with severe neurological complications, highlighting its serious pathogenic potential for humans. Since its emergence, more than 80 countries and territories have been affected by the ZIKV pandemic, which is now recognized as a neurotropic and teratogenic virus. The association of contemporary ZIKV strains with severe forms of disease in humans, that have never been reported before, has raised the hypothesis of newly acquired pathogenicity. In this regard, my doctoral research aimed to determine whether the scope of the current epidemic was partly facilitated by viral factors that improved ZIKV fitness. To this end, my research project focused on the identification of the viral molecular factors involved in Zika virus pathogenicity in humans based on the development of molecular clones
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Fulop, Lynda Dorothy. "Molecular analysis of flavivirus genome sequences : implications for virus classification." Thesis, University of Surrey, 1995. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.308496.
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Dejarnac, Ophélie. "Molecular and cellular basis of phosphatidylserine receptors mediated flavivirus infection." Thesis, Sorbonne Paris Cité, 2017. http://www.theses.fr/2017USPCC297/document.
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Le virus de la dengue (DENV) et le virus Zika (ZIKV) sont deux virus transmis par le moustique et responsables de maladies importantes chez l’Homme. En absence de vaccin et de traitements antiviraux efficaces, ces pathogènes représentent des problèmes de santé publique majeurs. Les bases moléculaires des interactions qu’établissent le DENV et ZIKV et la cellule hôte lors de l’entrée virale sont peu connues. Notre laboratoire a récemment identifié, les protéines TIM (TIM-1 et TIM-4) et TAM (Tyro3 et Axl), deux familles de récepteurs à la phosphatidylsérine (PtdSer) impliqués dans la reconnaissance et l’élimination des cellules apoptotiques par phagocytose, comme de nouveaux facteurs d’entrée du DENV. Les récepteurs TIM et TAM permettent l’infection par le DENV en interagissant avec la PtdSer associée aux virions selon un mécanisme similaire à la reconnaissance des cellules apoptotiques (mimétisme apoptotique). L’objectif général de mon travail de thèse a été d’explorer les mécanismes moléculaires et cellulaires par lesquels TIM-1 et Axl médient l’entrée des flavivirus. A l’aide de techniques d’imagerie en temps réel nous avons montré que TIM-1 et DENV sont co-internalisés et que TIM-1 joue un rôle actif dans l’entrée du DENV. Notamment, nous avons montré que deux résidus lysine présentes dans le domaine cytoplasmique de TIM-1 sont importantes pour l’ubiquitination du récepteur et pour l’endocytose du virus. La recherche de partenaires de TIM-1 par des études de spectrométrie de masse a permis d’identifier STAM, un membre du complexe ESCRT-0 impliqué dans le trafic des récepteurs ubiquitinés, comme facteur important pour l’infection. Collectivement, nos résultats suggèrent très fortement que TIM-1 est le premier récepteur bona fide caractérisé pour le DENV.Identifier les facteurs d’entrée du ZIKV représente un enjeu majeur dans la compréhension du tropisme et de la pathogénèse associée à ce virus. Nous avons montré que le récepteur Axl est essentiel pour l’entrée du ZIKV dans les cellules microgliales, les astrocytes du cerveau humain en développement ainsi que dans les fibroblastes de la peau. Nos études ont démontré un double rôle du récepteur Axl dans l’infection par ZIKV. Axl lie et permet l’internalisation des particules virales, mais aussi, contribue à l’établissement d’un environnement favorable à la réplication virale en inhibant la réponse immunitaire innée. En conclusion, ce travail a contribué à améliorer notre compréhension des mécanismes d’entrée des virus DENV et ZIKV. Nos résultats indiquent que ces deux virus exploitent plusieurs récepteurs aux phospholipides pour initier leur cycle infectieux, ce qui pourrait contribuer à l’élargissement de leur tropismeDengue virus (DENV) and ZIKA virus (ZIKV) are two mosquito-borne viruses responsible for important diseases in humans. Since there is currently no vaccine neither antiviral treatment available against these human pathogens, they are two major health concerns. The molecular basis of DENV and ZIKV host cell interactions leading to virus entry are poorly understood, hampering the discovery of new targets for antiviral intervention. Our laboratory recently discovered that TIM (TIM-1 and TIM-4) and TAM (Tyro3 and Axl) proteins, two receptor families that contribute to the phosphatidylserine (PtdSer)-dependent phagocytic removal of apoptotic cells, are DENV entry factors. TIM and TAM receptors mediate DENV infection by interacting with virion-associated PtdSer through a mechanism similar to the recognition and engulfment of apoptotic cells by phagocytes (viral apoptotic mimicry). The general objective of my PhD was to establish a detailed understanding of the molecular mechanisms by which TIM-1 and Axl mediated infection. Using live imaging, we demonstrated that TIM-1 and DENV are co-internalised and TIM-1 play an active role during DENV endocytosis. We showed that TIM-1 cytoplasmic domain is essential for DENV internalization, especially, we identified two lysine residues that are essential for TIM-1 ubiquitination and DENV endocytosis. Proteomic analysis of TIM-1 interacting partners identified STAM, a member of the ESCRT-0 complex involved in intracellular sorting of ubiquitinated cargos, as an essential host factor for DENV infection. Collectively our results establish TIM-1 as the first identified DENV bona fide receptor.Identifying ZIKV entry factors represents a major challenge in the understanding of ZIKV tropism and pathogenesis. We showed that Axl is responsible for ZIKV infection of microglial cells and astrocytes in the human developing brain and primary fibroblasts in human skin, suggesting an important role of this receptor during ZIKV life cycle. We also highlighted the dual role of the Axl receptor in ZIKV infection, which simultaneously promotes viral entry and dampens the innate immune response to facilitate a post entry step of the ZIKV life cycle. In conclusion, this work provided new insights in our understanding of the DENV and ZIKV entry program. Both viruses engage phospholipid receptors for their infectious entry, providing a rational to ascertain therapeutic strategies targeting virion-associated phospholipids
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Defrocourt, Christophe. "L'encéphalite japonaise en 2003 : épidémiologie et nouvelles perspectives de vaccin." Rouen, 2004. http://www.theses.fr/2004ROUEP008.
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L'encéphalite japonaise est une maladie croissante en Asie du Sud-Est et dans les pays du Pacifique occidental puisqu'elle s'étend aujourd'hui à l'Australie et à la Russie. Elle est actuellement la première cause d'encéphalite virale en Asie et se rencontre surtout en zone rurale. Cette maladie est provoquée par un virus de la famille des Flavivirus. Elle est véhiculée par une piqûre de moustique de la famille des Culex. Les porcs et les oiseaux aquatiques ont un rôle de réservoirs dans la transmission de la maladie et seule leur augmentation explique la transmission à l'homme, puisqu'e celui-ci est un hôte accidentel. L'encéphalite japonaise est une maladie souvent bénigne mais peut cependant être fatale et laisser d'importantes séquelles chez les survivants. Les personnes les plus touchées sont les enfants et les personnes âgées ou immunodéficientes. Actuellement, aucun médicament efficace n'existe, mais la vaccination et une protection mécanique permettenet d'empêcher la maladie. De nouveaux vaccins sont en cours de recherche et la terminologie du "Chimerivax" va apporter dans peu de temps une vaccination à immunisation rapide et aussi efficace que celle utilisée contre le virus de la fièvre jaune. De plus, un accord entre professionnels pharmaceutiques et instituts de recherches donne l'espoir de la fabrication d'un traitement curatif.
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Scaramozzino, Natale. "Flavivirus : étude d'une cible diagnostique, la région NS codant pour la polymérase et d'une cible thérapeutique, la protéase NS3 du virus Langat." Université Joseph Fourier (Grenoble), 2002. http://www.theses.fr/2002GRE19004.
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Les flavivirus sont des virus responsables d'une considérable morbidité et mortalité dans le monde en causant des encéphalites sévères, des fièvres hémorragiques, ainsi que des symptômes fébriles chez l'homme. L'absence de symptômes cliniques spécifiques pour une infection liée à un virus donné, et la présence simultanée de différents arbovirus dans une même région impose la nécessité de disposer d'un diagnostic de genre par PCR. Parmi les amorces proposées dans la littérature, une seule paire a permis la détection des différents flavivirus testés avec une sensibilité limite de 105 doses infectieuses. ML-1. La PCR semi-nichée développée au laboratoire utilise 2 nouvelles amorces et permet la détection des différents flavivirus avec une sensibilité de moins de 200 doses infectieuses. ML-1. Après le séquençage des produits amplifiés, la construction d'un dendrogramme permet d'orienter le diagnostic vers les principales espèces de flavivirus. Actuellement, il n'existe pas de traitement spécifique des infections à flavivirus. La protéase virale est une cible privilégiée pour ce type de thérapie. La protéase NS3 du virus Langat, virus utilisé comme modèle du virus de l'encéphalite à tiques, a été exprimée, purifiée et son activité enzymatique, sous sa forme recombinante, a été caractérisée. L'activité protéasique a été réalisée par hydrolyse de différents substrats peptidiques chromogéniques. Cette protéase virale clive in vitro les substrats comportant en position P1 un acide aminé basique tel l'arginine ou la lysine. Ces observations sont confirmées par l'étude comparative des sites de clivage naturels au sein de la polyprotéine des flavivirus. Cette protéase recombinante devrait pouvoir être utilisée pour le criblage de molécules antiprotéases utilisables dans le traitement des infections à flavivirusFlaviviruses are responsible for considerable morbidity and mortality and may cause severe encephalitis, hemorrhagic fever, hepatitis, and febrile symptoms in vertebrates, including humans (. . . ). Different p-nitroanilide substrates, defined on canonic sequences for their susceptibility to Ser-protease, were applied to the proteolytic assays of the protein. The highest values were obtained from substrates containing an Arg or Lys (amino acid) residue at the P1 position. This purification method will facilitate the future development of reliable testing procedures for anti-proteases directed to NS3 proteins
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Figueiredo, Mario Luis Garcia de. "Identificação de flavivirus infectando culicídeos de 1999 a 2007 no Brasil." Universidade de São Paulo, 2010. http://www.teses.usp.br/teses/disponiveis/42/42132/tde-19052010-153321/.
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Introdução: Arbovírus são vírus transmitidos por artrópodos, pertencendo, principalmente, aos gêneros Flavivirus (Flaviviridae), Alphavirus (Togaviridae) e Orthobunyavirus (Bunyavirus). Os Flavivirus, em sua maioria, são associados a zoonoses, causando doenças humanas febrís, febres hemorrágicas e encefalites. Inclusive, causam epidemias que são sério problema de saúde pública. Este estudo mostra uma pesquisa de Flavivirus em culicídeos, de diferentes regiões do país, utilizando uma técnica para identificação viral por RT-PCR com primers Flavivirusespecíficos e uma Multiplex-nested-PCR com primers espécie-específicos. Métodos: Culicídeos foram capturados, quantificados, identificados, agrupados em lotes por espécie e congelados. No laboratório, os animais foram macerados e tiveram o RNA extraído. Estes extratos foram submetidos a RT-PCR gênero-específica e à Multiplex-nested-PCR, para detecção e identificação dos vírus a nível de espécie. Resultado: De 3317 culicídios adultos e 571 larvas coletados em 4 diferentes regiões do Brasil, Sul, Sudeste, Norte e Nordeste, fez-se 246 lotes de mosquitos e desses foi possível obter amplicon sugestivo de Flavivirus em 16 (6,5%). Em 3 lotes contendo larvas de Aedes albopictus obteve-se amplicon sugestivo de vírus do dengue tipo 3. Também, em 13 lotes contendo Haemagogus leucocelaenus, Aedes aegypti e Aedes albopictus foi possível obter amplicons sugestivos de vírus do dengue tipos 1 e 2. Dos amplicons obtidos, 4 tiveram nucleotídios seqüenciados o que permitiu confirmar a presença dos vírus do dengue tipo 3 e Cacipacoré. Conclusão: O trabalho permitiu concluir que: a metodologia de RT-PCR para Flavivirus seguida de Multiplex-nested-PCR espécie-específica foi adequada para detecção e identificação destes vírus em culicídios; amplificaram-se genomas de Flavivirus em 6,5% dos lotes de culicídios estudados; vírus do dengue tipo 1 e tipo 2 foram encontrados infectando Aedes aegypti de Santos em 1999, Manaus em 2005-2006 e Foz do Iguaçu; vírus do dengue tipos 2 e 3 foram encontrados em Aedes albopictus de Santos em 1999 e Manaus em 2005-2006, sugerindo que este mosquito participe na transmissão de dengue; vírus do dengue tipo 3 foi encontrado em larvas de Aedes albopictus mostrando transmissão vertical do vírus; vírus do dengue tipo 1 foi encontrado infectando Haemagogus sp. sugerindo existência de ciclo silvático deste vírus; Aedes aegypti do Amazonas estavam infectados com o vírus Cacipacoré.Introduction: Arbovirus are rodent-borne viruses mostly from Flavivirus (Flaviviridae), Alphavirus (Togaviridae) e Orthobunyavirus (Bunyavirus) genus. Flavivirus, are commonly zoonotic and can cause febrile illness, haemorrhagic fever and encephalitis. Flavivirus outbreaks occur in Brazil and are a major public health problem. We show here a research looking for Flavivirus infections in Culicidae by a RT-PCR using Flavivirus-especific primers and a Multiplex-nested-PCR using specie-specific primers for virus identification. Methods: Culicidae were captured, quantified, identified, pooled based on the specie and frozzen. In the laboratory, the animals were crushed and had the RNA extracted. These extracts were tested by a Flavivirus genus-specific RT-PCR followed by a specie-specific Multiplex-nested-PCR. Results: From 3317 captured adult Culicidae and 571 collected larvae in 4 different regions of Brazil, 246 pools were obtained and from these, Flavivirus indicative amplicons were obtained in 16 (6.5%). Amplicons of dengue type 3 were obtained from 3 pools of Aedes albopictus larvae. It was also possible to obtain indicative amplicons of dengue types 1 and 2 in 13 pools of Haemagogus leucocelaenus, Aedes aegypti and Aedes albopictus. Besides, 4 amplicons had the nucleotides sequenced, confirming the mosquito infection by dengue type 3 and Cacipacoré viruses. Conclusion: The technique combining a Flavivirus genus-specific RT-PCR followed by a specie-specific Multiplex-nested-PCR was suitable for detection of these viruses in the mosquitoes; Flavivirus infecting Culicidae were detected in 6.5% of the analyzed mosquito pools; dengue virus type 1 and type 2 were found infecting Aedes aegypti from Santos (1999), Manaus (2005-2006) and Foz do Iguaçu cities; dengue type 2 virus was found in Aedes albopictus from Santos city (1999) and Manaus city (2005-2006), suggesting that this mosquito could be participating on dengue transmition; dengue type 3 virus was found in Aedes albopictus larvae showing the vertical transmission of this virus; dengue type 1 virus was found infecting Haemagogus sp. what suggests on the existence of a sylvatic maintenance cycle of this virus; Aedes aegypti from Amazonas state were found infeted by Cacipacoré virus.
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Nguyen, Jennifer B. "Molecular Mechanisms of Host-Pathogen Interactions in Flavivirus and Hookworm Infection." Thesis, Yale University, 2014. http://pqdtopen.proquest.com/#viewpdf?dispub=3580786.
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Microbial pathogens and their hosts have evolved complex adaptations to ensure their individual survival, resulting in a so-called "molecular arms race." While hosts may have acquired diverse mechanisms to protect themselves from the microbial invader, pathogens have developed elaborate strategies to evade and subvert these defenses. Viruses and hookworms are important pathogens which have evolved to successfully invade and infect their human hosts. Although structural biology has provided significant mechanistic insight into these processes of invasion, many specific host-pathogen interactions and their dynamics have not been well studied or characterized.
The work presented in this dissertation clarifies the mechanisms of cellular entry of one particular family of viruses, the flaviviruses, and discusses strategies for viral clearance by host cells. Additional insight into the role of a cytoplasmic DNA sensor, LRRFIP1, in mediating an innate immune response to non-flavivirus microbial infection is presented. Finally, strategies for the development of small-molecule or peptide inhibitors of virus entry and hookworm infection are proposed.
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Caleiro, Giovana Santos. "Investigação da presença do retrovírus da Reticuloendoteliose aviária (REV) e do anticorpo IgG do vírus Oeste do Nilo (WNV) em aves." Universidade de São Paulo, 2018. http://www.teses.usp.br/teses/disponiveis/99/99131/tde-04092018-090320/.
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As aves podem carregar um grande número de patógenos. As aves migratórias, por viajarem longas distâncias, são as principais responsáveis pela disseminação de agentes infecciosos. Entre os agentes, destacam-se os vírus, como por exemplo o retrovírus da Reticuloendoteliose aviária (REV), amplamente distribuído; e o vírus da febre do Oeste do Nilo (WNV), uma virose reemergente, com caráter zoonótico. Os principais sintomas da Reticuloendoteliose aviária incluem anemia, doença de Runting e síndrome não neoplásica aguda. Já o agente etiológico da Febre do Nilo Ocidental, é o Flavivirus West Nile (WNV).. As aves são seus hospedeiros definitivos e os humanos são hospedeiros acidentais, podendo manifestar quadro febril, e em menor porcentagem, meningite e encefalite. Mosquitos dos gêneros Culex e Aedes spp são os principais transmissores do vírus. Ao contrário do REV que não dispõe de evidências de sua circulação no Brasil, há evidências do WNV em aves e equinos e mais recentemente, em humanos. O objetivo desse trabalho foi investigar a presença do REV e do WNV em aves silvestres e de cativeiro da cidade de São Paulo e do Norte do estado do Pará. Sangue, soro e swab de cloaca foram coletados, totalizando mais de 1000 amostras. Através de técnicas moleculares foi possível detectar a presença do REV em 74 amostras (16%), todas do estado do Pará. O sequenciamento parcial dessas amostras e sua filogenia sugeriu que a migração de aves EUA-Brasil possa ter sido a rota utilizada. Através de ELISA anti-IgG de WNV, 4 amostras de São Paulo foram positivas. Apresentamos a primeira evidência do REV no país e sugerimos a presença do WNV no estado de São Paulo.Birds can carry a large number of pathogens. The migratory birds are most responsible for the spread of infectious agents due to long distance travels. Among these pathogens, the most notable are viruses, such as the avian Reticuloendotheliosis retrovirus (REV), widely distributed; and the West Nile virus (WNV), a reemerging zoonotic disease. The main symptoms of avian reticuloendotheliosis include anemia, Runting\'s disease and acute nonneoplastic syndrome. The etiological agent of West Nile fever is Flavivirus West Nile (WNV). Birds are their definitive hosts and humans are accidental hosts, which generaly present febrile symptoms, but at less proportion,, meningitis and encephalitis. Mosquitoes of the genus Culex and Aedes spp are the main vectors of the virus. Differently from the REV that has no evidence of its circulation in Brazil, there is evidence of WNV in birds and horses and more recently in humans. The objective of this work was to investigate the presence of REV and WNV in wild birds and captive birds from the city of São Paulo and Northern from Pará State. Blood, serum and cloacal swab were collected, resulting in more than 1000 samples. Through molecular techniques it was possible to detect the presence of REV in 74 samples (16%), all from the State of Pará. The partial sequencing of these samples and their phylogeny suggested that the migration of US-Brazil may have been the route for the virus entry. Through anti-WNV IgG ELISA, 4 samples from São Paulo were positive. We present the first evidence of REV in the country and suggest the presence of WNV in the state of São Paulo.
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Cruz, Ana Cecilia Ribeiro. "Caracterização Molecular e Biológica do vírus dengue circulante no Brasil." reponame:Repositório Institucional da FIOCRUZ, 2005. https://www.arca.fiocruz.br/handle/icict/4111.
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Fundação Oswaldo Cruz.Instituto Oswaldo Cruz. Rio de janeiro, RJ, Brasil
A febre do dengue está amplamente distribuída por todas as áreas tropicais do mundo. Em certas áreas endêmicas, as formas graves da doença, febre hemorrágica do dengue (FHD) e síndrome de choque do dengue (SCD), ocorrem com freqüência. Outras manifestações clínicas, como, por exemplo, encefalopatias, encefalites, mielite transversa, também tem sido observadas. O nosso objetivo foi a caracterização genética e biológica de amostras de vírus VDEN2 e 3, com ênfase no caráter de neurovirulência do vírus VDEN2. As amostras de VDEN 2 e 3 utilizadas foram isoladas na Serviço de Arbovírus do Instituto Evandro Chagas a partir de soro de pacientes com quadro clínico de dengue (FD e FHD) e manifestações neurológicas. Os vírus padrão foram o VDEN2 Nova Guiné C e 44-2, VDEN3 H87. As 11 seqüências do vírus VDEN2 quando comparados entre si e com 16 outras seqüências do VDEN2 apresentaram percentual médio de identidade nucleotídica e aminoácido de 89,5 e 99,8% respectivamente. Os estudos filogenéticos realizados mostram os isolados brasileiros pertencem ao genótipo III de origem Asiática. As 10 seqüências do vírus VDEN3 quando comparados entre si e com 16 outras seqüências do VDEN3 apresentaram percentual médio identidade nucleotídica e aminoácido de 89,6 e 100% respectivamente.Os estudos filogenéticos realizados mostram que estão agrupados ao genótipo III também de origem Asiática. O virus VDEN2 isolado a partir de caso mielite transversa (Bel61082) foi caracterizado pelo sequenciamento nucleotidico da região estrutural assim como em testes de neurovirulência em camundongos recém-nascidos. A susceptibilidade de culturas primárias de neurônios e astrócitos, células de glioma humano, também foi estudada para fins de caracterização do caráter neurovirulento do vírus.O estudo de neurovirulência em infecção de camundongo mostrou sinais aparentes de doença apenas após a inoculação da cepa NGC, mas a replicação do vírus Bel 61082 pode ser detectada por RT-PCR em suspensão de cérebro de camundongo. Obtivemos evidências experimentais de apoptose nas células neuronais infectadas com VDEN2 NGC e Bel61082. No sequenciamento nucleotídico, a principal alteração observada está localizada na proteína M 28, região esta conhecida como pró-apoptótica.
The dengue fever is widely distributed in all tropical areas of the world. In certain endemic areas, severe disease, dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS), occur frequently. Other clinical manifestations such as, for example, encephalopathies, encephalitis, transverse myelitis, has also been observed. Our objective was to characterize genetic and biological samples of viruses VDEN2 and 3, with emphasis on the neurovirulence of the virus VDEN2. The samples of DENV 2 and 3 used were isolated in the Service of the Instituto Evandro Chagas Arboviruses from the serum of patients with clinical dengue (DF and DHF) and neurological manifestations. Viruses were standard VDEN2 New Guinea C and 44-2, VDEN3 H87. The 11 sequences of the virus VDEN2 compared among themselves and with 16 other sequences showed VDEN2 average percentage of nucleotide and amino acid identity of 89.5 and 99.8% respectively. Phylogenetic studies performed show the Brazilian isolates belonged to genotype III of Asian origin. The 10 sequences of the virus VDEN3 compared among themselves and with 16 other sequences showed VDEN3 average percentage nucleotide and amino acid identity of 89.6 and 100% respectively.The phylogenetic studies carried out show that they are grouped to genotype III also of Asian origin. The virus isolated from VDEN2 case transverse myelitis (Bel61082) was characterized by nucleotide sequencing of the framework region as well as tests neurovirulence in mice newborn. The susceptibility of primary cultures of neurons and astrocytes, human glioma cells was also investigated for the characterization of the character neurovirulent vírus.O study of infection in mouse neurovirulence showed no apparent signs of disease after inoculation only NGC strain, but virus replication 61,082 Bel can be detected by RT-PCR on mouse brain suspension. We obtained experimental evidence of apoptosis in neuronal cells infected with NGC and VDEN2 Bel61082. In nucleotide sequencing, the main change observed is located at 28 M protein, an area known as pro-apoptotic.
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COLE, E. R. "Estudo Fitoquímico do Óleo Essencial dos Frutos da Aroeira (Schinus terebinthifolius RADDI) e Sua Eficácia no Combate ao Dengue." Universidade Federal do Espírito Santo, 2008. http://repositorio.ufes.br/handle/10/4634.
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Previous issue date: 2008-03-07O dengue é considerado a mais importante das arboviroses humanas, constituindo atualmente um dos principais problemas de Saúde Pública no mundo. A doença é causada por um vírus do gênero Flavivirus, com quatro diferentes sorotipos, transmitidos por mosquitos do gênero Aedes, sendo o Aedes aegypti seu principal vetor. O controle do vetor por meio de inseticidas sintéticos figura como a principal medida adotada no combate a doença, uma vez que não existem vacinas efetivas contra os diferentes sorotipos do vírus. Os inseticidas, apesar de eficazes, apresentam sérios problemas relacionados ao seu uso, dentre os quais podem ser destacados os danos ambientais, a toxicidade para uso humano e o risco de desenvolvimento de larvas e adultos resistentes.
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Ottendorfer, Christy L. "Impact of West Nile virus on the natural history of St. Louis encephalitis virus in Florida." [Tampa, Fla] : University of South Florida, 2008. http://purl.fcla.edu/usf/dc/et/SFE0002452.
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Machado, Daiane Cristina. "Sequenciamento do genoma completo do vírus Culex flavivirus (Flaviviridae) isolado no Brasil /." São José do Rio Preto, 2016. http://hdl.handle.net/11449/154714.
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Orientador: Maurício Lacerda NogueiraBanca: Marilia de Freitas Calmon
Banca: Aripuanã Sakurada Aranha Watanabe
Resumo: No Brasil, a presença de ecossistemas com uma grande riqueza e diversidade de animais e cidades grandes e densamente povoadas oferece condições para a ocorrência de diversas arboviroses. Vários arbovírus transmitidos por mosquitos do gênero Culex pertencem ao gênero Flavivirus. Vírus desse gênero são conhecidos por causar doenças em humanos e animais, no entanto, flavivírus que não infectam ou causam doença em vertebrados, replicando somente em artrópodes têm sido descritos. O vírus Culex flavivirus é um exemplo. Ele foi isolado de espécies de mosquitos do gênero Culex na América do Norte, América Central, América do Sul, África e Ásia. Apesar dos estudos realizados em outras partes do mundo, no Brasil há ainda pouca informação sobre esse vírus. Nesse trabalho, realizamos o seqüenciamento do genoma completo do vírus Culex flavivirus (CxFV) isolado de mosquitos Culex de São José do Rio Preto (SP), por meio das técnicas de RT-PCR e seqüenciamento nucleotídico. O genoma viral do isolado CxFV_RPBR07_2007, o qual foi obtido no presente estudo apresentou 10.706 nucleotídeos com uma ORF de 10.092 nucleotídeos (3.364 aminoácidos) flanqueada por uma 5'UTR de 32 nucleotídeos e uma 3'UTR de 582 nucleotídeos. Análises filogenéticas revelaram que o CxFV_RPBR07_2007 agrupa-se com outros flavivírus de insetos, como o Cell fusing agent virus (CFAV) e o Kamiti River virus (KRV), e relaciona-se intimamente com isolados de CxFV do México (América Latina) e de Uganda (África); mantendo relação também com isolados dos Estados Unidos (América do Norte) e do Japão (Ásia). Esses agrupamentos foram também observados em outros estudos filogenéticos
Abstract: In Brazil, the presence of ecosystems with a wealth and diversity of animals, and cities large and densely populated offers conditions for the occurrence of many arboviruses. Several arboviruses transmitted by mosquitoes of the genus Culex belong to the genus Flavivirus. Viruses of this genus are known to cause disease in humans and animals, however, flaviviruses do not infect or cause disease in vertebrates, replicating only in arthropods have been described. Culex flavivirus virus is an example. He was isolated from species of mosquitoes of the genus Culex in North America, Central America, South America, Africa and Asia. Although studies conducted in other parts of the world, in Brazil there is still little information about this virus. In this work we performed the sequencing of the complete genome of Culex flavivirus virus (CxFV) isolated from Culex mosquitoes in São José do Rio Preto (SP) using the techniques of RT-PCR and nucleotide sequencing. The viral genome of the isolated (CxFV_RPBR07_2007) obtained in the present study presented 10.706 nucleotides with an ORF of 10.092 nucleotides (3.364 amino acids) flanked by a 5'UTR of 32 nucleotides and a 3'UTR of 582 nucleotides. Phylogenetic analyzes revealed that the CxFV_RPBR07_2007 groups with other insect flaviviruses such as the Cell fusing agent virus (CFAV) and Kamiti River virus (KRV), and is closely related to CxFV isolates from Mexico (Latin America) and Uganda (Africa); also maintaining relationship with isolates from the United States of America (North America) and Japan (Asia). These groupings were also observed in other phylogenetic studies
Mestre
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Machado, Daiane Cristina [UNESP]. "Sequenciamento do genoma completo do vírus Culex flavivirus (Flaviviridae) isolado no Brasil." Universidade Estadual Paulista (UNESP), 2014. http://hdl.handle.net/11449/154714.
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000869213.pdf: 1151866 bytes, checksum: 95a9e9bbe8d1c2b6b9784eae9f7abb27 (MD5)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)
Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)
Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)
No Brasil, a presença de ecossistemas com uma grande riqueza e diversidade de animais e cidades grandes e densamente povoadas oferece condições para a ocorrência de diversas arboviroses. Vários arbovírus transmitidos por mosquitos do gênero Culex pertencem ao gênero Flavivirus. Vírus desse gênero são conhecidos por causar doenças em humanos e animais, no entanto, flavivírus que não infectam ou causam doença em vertebrados, replicando somente em artrópodes têm sido descritos. O vírus Culex flavivirus é um exemplo. Ele foi isolado de espécies de mosquitos do gênero Culex na América do Norte, América Central, América do Sul, África e Ásia. Apesar dos estudos realizados em outras partes do mundo, no Brasil há ainda pouca informação sobre esse vírus. Nesse trabalho, realizamos o seqüenciamento do genoma completo do vírus Culex flavivirus (CxFV) isolado de mosquitos Culex de São José do Rio Preto (SP), por meio das técnicas de RT-PCR e seqüenciamento nucleotídico. O genoma viral do isolado CxFV_RPBR07_2007, o qual foi obtido no presente estudo apresentou 10.706 nucleotídeos com uma ORF de 10.092 nucleotídeos (3.364 aminoácidos) flanqueada por uma 5'UTR de 32 nucleotídeos e uma 3'UTR de 582 nucleotídeos. Análises filogenéticas revelaram que o CxFV_RPBR07_2007 agrupa-se com outros flavivírus de insetos, como o Cell fusing agent virus (CFAV) e o Kamiti River virus (KRV), e relaciona-se intimamente com isolados de CxFV do México (América Latina) e de Uganda (África); mantendo relação também com isolados dos Estados Unidos (América do Norte) e do Japão (Ásia). Esses agrupamentos foram também observados em outros estudos filogenéticos
In Brazil, the presence of ecosystems with a wealth and diversity of animals, and cities large and densely populated offers conditions for the occurrence of many arboviruses. Several arboviruses transmitted by mosquitoes of the genus Culex belong to the genus Flavivirus. Viruses of this genus are known to cause disease in humans and animals, however, flaviviruses do not infect or cause disease in vertebrates, replicating only in arthropods have been described. Culex flavivirus virus is an example. He was isolated from species of mosquitoes of the genus Culex in North America, Central America, South America, Africa and Asia. Although studies conducted in other parts of the world, in Brazil there is still little information about this virus. In this work we performed the sequencing of the complete genome of Culex flavivirus virus (CxFV) isolated from Culex mosquitoes in São José do Rio Preto (SP) using the techniques of RT-PCR and nucleotide sequencing. The viral genome of the isolated (CxFV_RPBR07_2007) obtained in the present study presented 10.706 nucleotides with an ORF of 10.092 nucleotides (3.364 amino acids) flanked by a 5'UTR of 32 nucleotides and a 3'UTR of 582 nucleotides. Phylogenetic analyzes revealed that the CxFV_RPBR07_2007 groups with other insect flaviviruses such as the Cell fusing agent virus (CFAV) and Kamiti River virus (KRV), and is closely related to CxFV isolates from Mexico (Latin America) and Uganda (Africa); also maintaining relationship with isolates from the United States of America (North America) and Japan (Asia). These groupings were also observed in other phylogenetic studies