Le virus West Nile (VWN) est un virus neurotrope principalement transmis par piqûre de moustique et dont le réservoir est constitué par la faune aviaire sauvage. Les souches circulant en Europe appartiennent à 4 lignages génétiques différents à l’origine de nombreuses épidémies d’ampleur modérée à faible et limitées géographiquement, contrairement à ce qui a été observé en Amérique du Nord. En 1998 en Israël, une importante épidémie a a été associée pour la première fois à une forte mortalité de la faune aviaire sauvage. Le virus (souche IS-98-ST1, lignage 1a) a été isolé du cerveau d’une cigogne moribonde. L’objet de cette thèse a été de construire un clone infectieux de la souche IS-98-ST1 afin d’en explorer les propriétés de neuroinvasion et de pouvoir mettre en évidence les déterminants moléculaires de sa virulence.Le virus obtenu à partir de la construction clone infectieux s’est révélé posséder les mêmes propriétés biologiques que le virus parental, que ce soit in vitro sur cellules Vero ou in vivo sur souris sensibles ou résistantes ou encore sur l’embryon de poulet. L’embryon de poulet est présenté ici comme un nouveau modèle d’évaluation de la virulence du VWN. Un modèle cellulaire neuronale (lignée de neuroblastomes humains, SK-N-SH) est aussi évalué dans ce manuscrit. En conclusion, un nouvel outil de génétique inverse a été obtenu pour le VWN. Cet outil permettra de travailler sur l’impact de mutations ponctuelles, ou de modifications plus importantes touchant un ou plusieurs gènes viraux sur la virulence du VWN, spécifiquement dans le contexte européen. / West Nile virus (WNV) is a neurotropic virus mainly transmitted through mosquito bites. Wild birds represent the main reservoir hosts. Strains circulating in Europe belong to four lineages and have caused numerous but limited epidemics over the last few years. In 1998, an important outbreak associated to huge bird fatalities caused by a highly neuroinvasive strain (IS-98-ST1) took place in Israel. We aimed at producing a new infectious clone, based on the lineage 1a IS-98-ST1 WNV strain, for the characterization of its neuroinvasion properties as well as the molecular determinants of European WNV virulence. The growth kinetics of recombinant and parental WNV were similar in Vero cells. Moreover, the phenotypes of recombinant and parental WNV were indistinguishable in terms of viremia, viral load in the brain and mortality in susceptible and resistant mice. Finally, the pathobiology of the infectious clone was examined in embryonated chicken eggs, proposed as a new model for the evaluation of WNV virulence. The potential of human neuroblastoma cells (SK-N-SH) to discriminate between highly and mildly virulent WNV strains was assayed. In conclusion: a new molecular tool that is useful for the study of molecular determinants of WNV virulence has been generated. We take advantage of the high genetic stability of our one-piece infectious WNV cDNA clone to produce mutant viruses through the insertion of point mutations or the exchange of genetic fragments between WNV strains into the backbone of the IS-98-ST1 infectious clone.

Source: Masters Abstracts International, Volume: 47-06, page: 3501. Adviser: Kristy O. Murray. Includes bibliographical references.

Thesis (M.S.) -- University of Tennessee, Knoxville, 2005. / Title from title page screen (viewed on July 13, 2005). Thesis advisor: Reid R. Gerhardt. Document formatted into pages (xii, 77 p. : ill. (some col.), maps (some col.)). Vita. Includes bibliographical references (p. 67-76).

Les flavivirus ayant un impact en médecine vétérinaire sont largement distribués dans le monde (à l’exemple de la fièvre West Nile (WNF) présente sur les cinq continents ou de l’Encéphalite japonaise (EJ) en Asie du Sud-Est) et sont responsables de maladies à dominante neurologique chez l’homme et/ou le cheval.La virémie étant généralement brève lors de ces infections virales, les méthodes de diagnostic utilisées sont essentiellement sérologiques. Or le chevauchement fréquent des aires de répartition des flavivirus complique le diagnostic sérologique. En effet, des réactions sérologiques croisées entre flavivirus sont observées lors de l’utilisation de méthodes de diagnostic usuelles telles que l’ELISA et l’immunofluorescence (IF). Les résultats sérologiques doivent donc être confirmés par la méthode fastidieuse de séroneutralisation (SNT) virale avec les différents flavivirus existants dans la région. De plus, le risque d’émergence sur un territoire donné de nouveaux flavivirus comme le virus Zika au Brésil ou en Amérique du Nord ne peut être exclu.Nous avons donc développé dans la première partie de ce travail une nouvelle stratégie de diagnostic sérologique multiplexe à l’aide de la méthode d’immuno-essai (MIA) sur billes. Sachant que la glycoprotéine d’enveloppe soluble (sE) des flavivirus est divisée en 3 domaines (D) structuraux, DI, DII et DIII et que les épitopes du DIII sont spécifiques de chaque flavivirus, des protéines EDIII recombinantes de différents flavivirus d’intérêt ont été synthétisées en système d’expression Drosophila S2 et leur antigénicité a été testée sur sérums équins et ovins. Nous avons obtenu des résultats très encourageants en démontrant que l’utilisation des EDIII associée avec la capacité de multiplexage de la méthode MIA apparaît comme une réponse adaptée au défi du diagnostic sérologique des infections à flavivirus.Nous avons en outre utilisé la même méthode de multiplexage mais avec des antigènes NS1 du WNV pour mettre en place un test DIVA (Differentiating Infected from Vaccinated Animals) permettant de différencier les chevaux infectés par le WNV des chevaux vaccinés avec un vaccin recombinant WNV.Un autre écueil en médecine vétérinaire est le traitement des affections à flavivirus. En effet l’arsenal thérapeutique est limité et le traitement est avant tout symptomatique. Nous avons dans la seconde partie de ce travail testé in vitro une molécule antivirale à large spectre, le sr1057 sur nos virus d’intérêt (WNV et JEV). Cette molécule qui provient d’une stratégie de criblage développée par l’Institut Pasteur a probablement pour cible la cellule de l’hôte car elle est capable d’inhiber des virus très différents, à génome à ARN de polarité positive ou négative et ADN.Les résultats que nous avons obtenus avec ce composé sont en demi-teinte pour les flavivirus testés avec une efficacité démontrée pour le JEV mais plus modeste pour le WNV. Ils n’excluent cependant pas une possible utilisation de cet antiviral en médecine vétérinaire équine car une activité inhibitrice in vitro sur les virus de l’Herpes équin de type I et de l’artérite virale a été confirmée par d’autres collaborateurs. / Flaviviruses with a major impact in veterinary medicine are widely distributed (e.g. West Nile fever (WNF) has spread across the five continents and Japanese Encephalitis (JE) is reported in South-East Asia) and are mainly responsible for neurological diseases in humans and/or horses.After flavivirus infection, viremia in mammal hosts is generally short and consequently indirect methods are mostly used to diagnose flavivirus infections. However, frequent spatial overlapping in their circulation areas renders the interpretation of serological assays difficult. Indeed, cross-reactions between flaviviruses are observed in rapid serological tests such as in ELISA and immunofluorescence assays (IFA). Serological assay results should thus be confirmed by the tedious comparative virus neutralization test (VNT) using a panel of viruses known to circulate in the area. Moreover, the risk of emergence of new flaviviruses such as reported recently with the Zika virus in Brazil or in North America should be considered when studying flavivirus epidemiology.In the first section, a new strategy aiming at improving the serological diagnosis of flavivirus infections was developed using the multiplexing capacity of microsphere immunoassays (MIA). The flavivirus soluble envelope (sE) glycoprotein ectodomain is composed of three domains (D), e.g. DI, DII and DIII, with EDIII containing virus-specific epitopes. Recombinant EDIIIs of different flaviviruses were synthesized in the Drosophila S2 expression system. The microspheres coupled with rEDIIIs were assayed with equine and ovine sera from natural and experimental flavivirus infections or non-immune samples. Very encouraging results have been obtained and this innovative multiplex immunoassay based on flavivirus rEDIIIs appears to be a powerful alternative to ELISAs and VNTs for veterinary diagnosis of flavivirus-related diseases.MIA with WNV nonstructural 1 protein were also implemented to differentiate Infected from Vaccinated Animals (DIVA). Such a DIVA approach was only successful when horses had been immunized with a recombinant canarypox vaccine, while horses receiving inactivated WNV vaccine developed immune responses close to the ones induced after natural infection.Another pitfall in veterinary medicine is the lack of therapeutics for viral diseases and specifically for flaviviruses. The therapeutic arsenal is indeed rather limited and animals are generally administered supportive treatments only. In the second part, the results of the in vitro testing of a broad spectrum antiviral named sr1057 on WNV and JEV replication are presented. This chemical, identified from a unique screening strategy developed by Pasteur Institute, is probably targeting the host cell and was found to inhibit the replication of varied RNA and DNA viruses belonging to different virus families. The sr1057 compound was not as efficient at inhibiting the replication of flaviviruses as for other RNA+ viruses, with a modest antiviral effect demonstrated for WNV and a higher efficacy on JEV. This antiviral presents however potentials for applications in equine veterinary medicine because it efficiently inhibited equine herpes virus-1 and equine arteritis virus in vitro, as clearly shown by other collaborators.

West Nile virus (WNv) is an arbovirus and is transmitted by infected mosquitoes after feeding on the blood of birds carrying the virus. The Canadian WNv national surveillance system has just completed its tenth year of operation. The thesis is to evaluate the surveillance system and analyze multi-year human data. The evaluation includes the use of multiple lines of complementary methods such as the US CDC surveillance guidelines, Canadian Evaluation Framework, document review and a survey. Logistic and Poisson regressions were used for data analyses. WNv has become endemic in most parts of Canada since the virus occurred in 2001. The virus activity is peak around August. High numbers of human cases with WNv neurological syndrome identified pose a significant health concern due to the long term sequelae among affected patients. WNv national surveillance met its main objectives and there is a continual need for the surveillance.

Introduction: Vector-borne infectious diseases represent a major public health problem in both developing and developed nations. In particular, West Nile Virus (WNV), a mosquito-borne disease that can lead to severe disease and death in humans, caused over 2,100 reported cases in the United States last year (CDC, 2016). In Maricopa County, Arizona WNV has caused 474 reported cases during the last five years, with a case-fatality rate at 7.8%. Aim: To examine the association between weather patterns and incidence of WNV in Maricopa County, AZ from 2007 to 2013. Methods: We analyzed weekly data on climatological variables and WNV incidence from Maricopa County, AZ. The specific independent variables of interest were precipitation, minimum temperatures, mean temperatures, and maximum temperatures. A full model was generated using multiple linear regression, and a stepwise selection procedure yielded a minimal model. Results: The full multiple linear regression model explains 45.30% of the observed variance in WNV incidence. The variable showing a significant impact on WNV incidence in this model was rainfall (p <0.0001). Stepwise selection results explained 45.16% of the variance observed in the data. This model included two significant predictors: precipitation and maximum temperature. Conclusion: Climatic variables, particularly the amount of rainfall and maximum temperatures, significantly influence WNV dynamics in Maricopa County, Arizona. These findings are in line with prior studies and could be useful to guide mosquito control programs in the state of Arizona.

West Nile virus (WNv) is an arbovirus and is transmitted by infected mosquitoes after feeding on the blood of birds carrying the virus. The Canadian WNv national surveillance system has just completed its tenth year of operation. The thesis is to evaluate the surveillance system and analyze multi-year human data. The evaluation includes the use of multiple lines of complementary methods such as the US CDC surveillance guidelines, Canadian Evaluation Framework, document review and a survey. Logistic and Poisson regressions were used for data analyses. WNv has become endemic in most parts of Canada since the virus occurred in 2001. The virus activity is peak around August. High numbers of human cases with WNv neurological syndrome identified pose a significant health concern due to the long term sequelae among affected patients. WNv national surveillance met its main objectives and there is a continual need for the surveillance.

West Nile Virus (WNV) first appeared in the western hemisphere in 1999, and has since spread across the United States and into Mexico and the Caribbean. It has been hypothesized that WNV has spread rapidly via migratory birds, and that various avian species may facilitate viral amplification during winter months. The goals of this research were to determine the role of American Tree Sparrows (Spizella americana) in the spread of WNV during their igrations and to determine the role of the Northern Cardinal (Cardinalis cardinalis) in winter survivorship and subsequent spring amplification of WNV. Additional wintering avian species were sampled to provide a general survey of the prevalence of WNV in winter in south-central Kansas. Blood samples were taken from the brachial vein of migratory and wintering birds captured using mist nets at four wintering feeding stations at the Wichita State University Field Station. Some samples were taken from retrapped birds within a single winter to determine if winter transmission occurs. Some birds were resampled in consecutive winters to monitor seroconversion rates. Analysis of serum samples were performed, in triplicate, using an epitope-blocking ELISA. The current study was conducted during the consecutive winters of 2003-04 and 2004-05. It was concluded that resident species had an increased incidence of WNV exposure when compared to that of migratory species. This difference suggests that migratory species may not have as important a role in the dissemination of WNV as first hypothesized. Also, minimal, if any, winter transmission occurs on communal feeding grounds. Viral amplification during the winter was not demonstrated, although one individual seroconverted during a single winter. / Thesis (M.S.)--Wichita State University, Dept. of Biological Sciences / Includes bibliographical references (leaves 35-39)

The 1,2,5-Thiadiazolidin-3-one 1,1-dioxide scaffold was used in the design and synthesis of inhibitors of Dengue Virus and West Nile Virus proteases and human tryptase. The scaffold was successfully used in the synthesis of potential inhibitors of Dengue Virus and West Nile Virus proteases. Inhibitors of Human tryptase synthesized based on the 1,2,5-Thiadiazolidin-3-one 1,1- dioxide scaffold were shown to be effective mechanism-based inhibitors of the enzyme. / Thesis (M.S.)--Wichita State University, College of Liberal Arts and Sciences, Dept. of Chemistry. / "July 2006." / Includes bibliographic references (leaves 64-66).

RNase footprinting and nitrocellulose filter-binding assays were previously used to map one major and two minor binding sites for the cell protein eEF1A on the 3’(+) stem loop (SL) RNA of West Nile virus (WNV) (2). Base substitutions in the major eEF1A binding site or adjacent areas of the 3’(+) SL were engineered into a WNV infectious clone. Mutations that decreased, as well as ones that increased, eEF1A binding in in vitro assays had a negative affect on viral growth. None of these mutations affected the efficiency of translation of the viral polyprotein from the genomic RNA, but all of the mutations that decreased in vitro eEF1A binding to the 3’ SL RNA also decreased viral minus-strand RNA synthesis in transfected cells. Also, mutations that increased the efficiency of eEF1A binding to the 3’ SL RNA increased minus-strand RNA synthesis in transfected cells, which resulted in decreased synthesis of genomic RNA. These results strongly suggest that the interaction between eEF1A and the WNV 3’ SL facilitates viral minus-strand initiation. eEF1A colocalized with viral replication complexes (RC) in infected cells and antibody to eEF1A coimmunoprecipitated viral RC proteins, suggesting that eEF1A facilitates an interaction between the 3’ end of the genome and the RC. eEF1A bound with similar efficiency to the 3’ terminal SL RNAs of four divergent flaviviruses, including a tick-borne flavivirus, and colocalized with dengue RC in infected cells. These results suggest that eEF1A plays a similar role in the RNA replication of all flaviviruses.