Publications

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the structural immunology of antibody protection against west nile virus.summary: recent investigations of the interaction between the west nile virus (wnv) envelope protein (e) and monoclonal antibodies (mabs) have elucidated fundamental insights into the molecular mechanisms of neutralization. structural studies have defined an epitope on the lateral ridge of domain iii (diii-lr) of the wnv e protein that is recognized by antibodies with the strongest neutralizing activity in vitro and in vivo. antibodies that bind this epitope are highly potent because they effici ...200818837784
substrate inhibition kinetic model for west nile virus ns2b-ns3 protease.west nile virus (wnv) has recently emerged in north america as a significant disease threat to humans and animals. unfortunately, no approved antiviral drugs exist to combat wnv or other members of the genus flavivirus in humans. the wnv ns2b-ns3 protease has been one of the primary targets for anti-wnv drug discovery and design since it is required for virus replication. as part of our efforts to develop effective wnv inhibitors, we reexamined the reaction kinetics of the ns2b-ns3 protease and ...200818855422
serological evidence of usutu virus occurrence in north-eastern italy.in the recent years, usutu virus (usuv), a flavivirus of the japanese encephalitis virus complex, has been reported in central europe. as part of a systematic surveillance programme to monitor possible entrance and/or circulation of vector-borne viruses, since 2001, sentinel-chicken flocks were placed throughout the italian territory nearby areas considered at risk of virus introduction. they have been periodically checked for the presence of antibodies against flaviviruses by indirect elisa, pl ...200818667029
spatial and temporal organization of tick-borne encephalitis flavivirus replicated rna in living cells.flaviviruses are positive rna viruses that assemble the replication complex in the cytoplasm of the infected cells. in order to get a dynamic view of the formation and distribution of flavivirus genomes in living cells we engineered a tick-borne encephalitis virus (tbev) replicon with an array of binding sites for the phage ms2 core protein. the modified tbev replicons were competent for rna replication and allowed the visualization of replicated genomic rna that accumulated in cytoplasmic struc ...200818667221
towards the design of antiviral inhibitors against flaviviruses: the case for the multifunctional ns3 protein from dengue virus as a target.new treatments are urgently needed to combat the increasing number of dengue fever cases in endemic countries as well as amongst a large number of travellers from non-endemic countries. of the 10 virus encoded proteins, ns3 (non-structural 3) and ns5 carry out all the enzymatic activities needed for polyprotein processing and genome replication, and are considered to be amenable to antiviral inhibition by analogy with successes for similar targets in human immunodeficiency virus and hepatitis c ...200818674567
one-step rt-pcr for detection of zika virus.zika virus (zikv) is an emerging mosquito-borne flavivirus circulating in asia and africa. human infection induces an influenza-like syndrome that is associated with retro-orbital pain, oedema, lymphadenopathy, or diarrhea. diagnosis of zika fever requires virus isolation and serology, which are time consuming or cross-reactive.200818674965
a glycosylated peptide in the west nile virus envelope protein is immunogenic during equine infection.using a monoclonal antibody directed to domain i of the west nile virus (wnv) envelope (e) protein, we identified a continuous (linear) epitope that was immunogenic during wnv infection of horses. using synthetic peptides, this epitope was mapped to a 19 aa sequence (wn19: e147-165) encompassing the wnv ny99 e protein glycosylation site at position 154. the inability of wnv-positive horse and mouse sera to bind the synthetic peptides indicated that glycosylation was required for recognition of p ...200819008394
insights into rna unwinding and atp hydrolysis by the flavivirus ns3 protein.together with the ns5 polymerase, the ns3 helicase has a pivotal function in flavivirus rna replication and constitutes an important drug target. we captured the dengue virus ns3 helicase at several stages along the catalytic pathway including bound to single-stranded (ss) rna, to an atp analogue, to a transition-state analogue and to atp hydrolysis products. rna recognition appears largely sequence independent in a way remarkably similar to eukaryotic dead box proteins vasa and eif4aiii. on ssr ...200819008861
[humoral immune response to structural and nonstructural proteins of tick-borne encephalitis virus in the natural history of the disease].a hundred and ninety-one patients who had fallen ill with tick-borne encephalitis (tbe) after bites by virus-carrying ticks were examined to reveal the characteristics of antibody production of the major classes of immunoglobulins (igm and igg) against structural protein e of the tbe virus and nonstructural protein ns1 of the tbe viral replicative complex in the natural history of an infectious process in different forms and variants of the disease. in the two-wave course of tbe, the infectious ...200818756813
discovery of a hepatitis c target and its pharmacological inhibitors by microfluidic affinity analysis.more effective therapies are urgently needed against hepatitis c virus (hcv), a major cause of viral hepatitis. we used in vitro protein expression and microfluidic affinity analysis to study rna binding by the hcv transmembrane protein ns4b, which plays an essential role in hcv rna replication. we show that hcv ns4b binds rna and that this binding is specific for the 3' terminus of the negative strand of the viral genome with a dissociation constant (kd) of approximately 3.4 nm. a high-throughp ...200818758449
cell-mediated immunity induced by chimeric tetravalent dengue vaccine in naive or flavivirus-primed subjects.three independent, phase 1 clinical trials were conducted in australia and in usa to assess the safety and immunogenicity of sanofi pasteur dengue vaccine candidates. in this context, dengue 1-4 and yellow fever 17d-204 (yf 17d)-specific cd4 and cd8 cellular responses induced by tetravalent chimeric dengue vaccines (cyd) were analyzed in flavivirus-naive or flavivirus-immune patients. tetravalent cyd vaccine did not trigger detectable changes in serum pro-inflammatory cytokines, whatever the vac ...200818762226
prevalence of west nile virus neutralizing antibodies in wild birds from the camargue area, southern france.the camargue area of southern france experienced the re-emergence of west nile virus (wnv) in the late summer of 2000 and 2004. immediately preceding the 2004 outbreak, samples were collected from 432 birds of 32 different species captured in mist nets and from 201 cattle egret (bubulcus ibis) nestlings sampled in their nests between 1 april and 12 june 2004. west nile virus neutralizing titers of >/=40 were detected in 4.8% (95% confidence limit, 2.9-7.5%) of the adult birds and in 1.6% (0.3-4. ...200818689669
does variation in culex (diptera: culicidae) vector competence enable outbreaks of west nile virus in california?since the invasion of california by west nile virus (family flaviviridae, genus flavivirus, wnv) in 2003, we have annually monitored vector competence for the ny99 strain in culex tarsalis coquillett, culex pipiens quinquefasciatus say, culex p. pipiens l., and culex stigmatosoma dyar populations from four areas: deserts of coachella valley, densely urbanized maritime los angeles, southern san joaquin valley in kern county, and southern sacramento valley near davis in sacramento county. overall, ...200819058638
innate resistance to flavivirus infections and the functions of 2'-5' oligoadenylate synthetases.mouse susceptibility to experimental infections with flaviviruses is significantly influenced by a cluster of genes on chromosome 5 encoding a family of proteins with enzymatic properties, the 2'-5' oligoadenylate synthetases (oas). positional cloning of the locus in question has revealed that susceptibility of laboratory inbred strains to this class of virus is associated with a nonsense mutation in the gene encoding the oas1b isoform. analysis of the molecular structure of the cluster in diffe ...200818727488
extrinsic incubation periods for horizontal and vertical transmission of west nile virus by culex pipiens pipiens (diptera: culicidae).culex pipiens pipiens l. (diptera: culicidae), infected per os from a membrane feeder, transmitted west nile virus (family flaviviridae, genus flavivirus, wnv) at 26 degrees c horizontally during feeding to hamsters and suckling mice and vertically to f1 progeny during egg deposition. horizontal transmission rates increased with extrinsic incubation, with 75-100% of the females transmitting on days 16 through 25 postinfection (pi). no females deposited eggs infected with wnv after the first bloo ...200818533438
vector competence of aedes vexans (diptera: culicidae) for west nile virus and potential as an enzootic vector.vector competence of aedes vexans (meigen) and culex pipiens pipiens l. (diptera: culicidae) for west nile virus (family flaviviridae, genus flavivirus, wnv) was compared. infection rates of both species were similar 14 d after feeding on chickens, with wnv titers ranging from 10(4.2) to 10(8.7) plaque-forming units (pfu)/ml. median infectious doses and 95% confidence intervals (ci) were 10(6.0(5.8, 63)) and 10(5.7(5.4, 5.9)) pfu for ae. vexans and cx. p. pipiens, respectively. wnv transmission ...200818533439
using hydrologic conditions to forecast the risk of focal and epidemic arboviral transmission in peninsular florida.the accurate forecasting and tracking of arboviral transmission is becoming increasingly critical for the early recognition and management of arboviral epidemics. meteorological factors, especially rainfall and temperature, drive arboviral epidemics, but monitoring rainfall and temperature alone is not predictive of increased levels of vector-borne disease transmission. in florida, model simulations of water table depth (wtd) provide a measure of drought, and they have been shown to provide an a ...200818533440
arbovirus transmission by culex nigripalpus in florida, 2005.understanding the transmission patterns of west nile and st. louis encephalitis viruses (family flaviviridae, genus flavivirus, wnv and slev) could result in an increased ability to predict transmission risk to humans. to examine transmission patterns between vector and host, we trapped mosquitoes in three florida counties from june to november 2005 by using chicken-baited lard can mosquito traps. these traps were used to monitor for presence of wnv and slev in mosquitoes and subsequent transmis ...200818533444
persistent west nile virus transmission and the apparent displacement st. louis encephalitis virus in southeastern california, 2003-2006.west nile virus (family flaviviridae, genus flavivirus, wnv) invaded the colorado desert biome of southern california during summer 2003 and seemed to displace previously endemic st. louis encephalitis virus (family flaviviridae, genus flavivirus, slev, an antigenically similar flavivirus in the japanese encephalitis virus serocomplex). western equine encephalomyelitis virus (family togaviridae, genus alphavirus, weev), an antigenically distinct alphavirus, was detected during 2005 and 2006, ind ...200818533445
failure of japanese encephalitis vaccine and infection in inducing neutralizing antibodies against west nile virus, people's republic of china.we examined whether live attenuated japanese encephalitis (je) vaccine is effective in preventing west nile virus (wnv) infection in the people's republic of china. three groups were recruited into the study: patients with japanese encephalitis (je), healthy controls vaccinated with live attenuated 2 sa14-14-vaccine against je virus (jev), and unvaccinated healthy controls. serum samples were collected and screened for igg antibodies against jev by an indirect immunofluorescence assay. positive ...200818541783
effect of ns3 and ns5b proteins on classical swine fever virus internal ribosome entry site-mediated translation and its host cellular translation.a full-length ns3 (ns3f) and a truncated ns3 protein (ns3h) with an rna helicase domain possess rna helicase activity. using an in vitro system with a monocistronic reporter rna or dna, containing the csfv 5'-utr, we observed that both ns3f and ns3h enhanced internal ribosome entry site (ires)-mediated and cellular translation in a dose-dependent manner, but ns3 protease (ns3p) that lacks a helicase domain did not. ns3f was stronger than ns3h in promoting both translations. these results showed ...200818343841
efficient dengue virus (denv) infection of human muscle satellite cells upregulates type i interferon response genes and differentially modulates mhc i expression on bystander and denv-infected cells.dengue virus (denv) is a mosquito-borne flavivirus that causes an acute febrile disease in humans, characterized by musculoskeletal pain, headache, rash and leukopenia. the cause of myalgia during denv infection is still unknown. to determine whether denv can infect primary muscle cells, human muscle satellite cells were exposed to denv in vitro. the results demonstrated for the first time high-efficiency infection and replication of denv in human primary muscle satellite cells. changes in globa ...200818559930
use of synthetic peptides to represent surface-exposed epitopes defined by neutralizing dengue complex- and flavivirus group-reactive monoclonal antibodies on the native dengue type-2 virus envelope glycoprotein.the reactions of neutralizing monoclonal antibodies (mabs) that defined dengue virus (denv) complex, flavivirus subgroup or group neutralizing epitopes were tested against synthetic peptide sequences from domains i, ii and iii of the envelope (e) glycoproteins of different denv-2 genotypes/strains. the denv complex-reactive mab identified the surface-exposed 304-gkfkv/ivkeia-313 peptides and the denv complex-conserved 393-kkgssigq/km-401 peptides in domain iii, which were located adjacently in t ...200818559931
characterization of mosquito-adapted west nile virus.west nile virus (wnv), a mosquito-borne flavivirus, has significantly expanded its geographical and host range since its 1999 introduction into north america. the underlying mechanisms of evolution of wnv and other arboviruses are still poorly understood. studies evaluating virus adaptation and fitness in relevant in vivo systems are largely lacking. in order to evaluate the capacity for host-specific adaptation and the genetic correlates of adaptation in vivo, this study measured phenotypic and ...200818559933
west nile virus nonstructural protein 1 inhibits tlr3 signal transduction.the innate immune response is the first line of defense against foreign pathogens. the recognition of virus-associated molecular patterns, including double- and single-stranded rna, by pattern recognition receptors initiates a cascade of signaling reactions. these result in the transcriptional upregulation and secretion of proinflammatory cytokines that induce an antiviral state. many viruses have evolved mechanisms to antagonize these responses in order to help them establish a productive infec ...200818562533
changing the protease specificity for activation of a flavivirus, tick-borne encephalitis virus.the infectivity of flavivirus particles depends on a maturation process that is triggered by the proteolytic cleavage of the precursor of the m protein (prm). this activation cleavage is naturally performed by ubiquitous cellular proteases of the furin family, which typically recognize the multibasic sequence motif r-x-r/k-r. previously, we demonstrated that a tick-borne encephalitis virus (tbev) mutant with an altered cleavage motif, r-x-r, produced immature, noninfectious particles that could ...200818562534
comparative neuropathogenesis and neurovirulence of attenuated flaviviruses in nonhuman primates.based on previous preclinical evaluation in mice and monkeys, the chimeric tbev/den4delta30 virus, carrying the prm and e protein genes from a highly virulent far eastern strain of tick-borne encephalitis virus (tbev) on the backbone of a nonneuroinvasive dengue type 4 virus (den4), has been identified as a promising live attenuated virus vaccine candidate against disease caused by tbev. however, prior to use of this vaccine candidate in humans, its neurovirulence in nonhuman primates needed to ...200818353947
the flavivirus precursor membrane-envelope protein complex: structure and maturation.many viruses go through a maturation step in the final stages of assembly before being transmitted to another host. the maturation process of flaviviruses is directed by the proteolytic cleavage of the precursor membrane protein (prm), turning inert virus into infectious particles. we have determined the 2.2 angstrom resolution crystal structure of a recombinant protein in which the dengue virus prm is linked to the envelope glycoprotein e. the structure represents the prm-e heterodimer and fits ...200818369147
flavivirus methyltransferase: a novel antiviral target.many flaviviruses are significant human pathogens. no effective antiviral therapy is currently available for treatment of flavivirus infections. development of antiviral treatment against these viruses is urgently needed. the flavivirus methyltransferase (mtase) responsible for n-7 and 2'-o methylation of the viral rna cap has recently been mapped to the n-terminal region of nonstructural protein 5. structural and functional studies suggest that the mtase represents a novel antiviral target. her ...200818571739
characterization of asn130-to-ala mutant of dengue type 1 virus ns1 protein.the nonstructural protein 1 (ns1) of flavivirus has two n-glycosylation sites that are thought to be important for viral replication. effects of ns1 glycosylation site mutations on viral replication have been reported in several flaviviruses, but the results have differed. in this report, we examined the role of glycosylation site of ns1 on the replication of dengue type 1 virus (denv-1). denv-1 production was not detectable when full-length denv-1 rna, which has an n-glycosylation site asn130-t ...200818288598
prevalence of west nile virus neutralizing antibodies in spain is related to the behavior of migratory birds.west nile virus (wnv) is a bird flavivirus capable of infecting horses and humans that is transmitted by blood-sucking vectors. in europe and africa, sporadic infections and outbreaks causing human illness and deaths have occurred and have led to 2 mutually nonexclusive hypotheses regarding the circulation of wnv in europe: (1) the occurrence of endemic sylvatic cycles that occasionally result in human or equine infection, or (2) sporadic seeding of wnv by migratory birds from areas where the vi ...200818399777
[genotyping of japanese encephalitis viruses isolated in yunnan].explore the genotype of the epidemic japanese encephalitis virus (jev) in yunnan province from molecular level and to understand the molecular differences of the virus isolated from yunnan at different time, locality and host.200818574523
is quantum mechanics necessary for predicting binding free energy?to take into account polarization effects, the linear interaction energy model with continuum electrostatic solvation (liece) is supplemented by the linear-scaling semiempirical quantum mechanical calculation of the intermolecular electrostatic energy (qmliece). qmliece and liece are compared on three enzymes belonging to different classes: the west nile virus ns3 serine protease (wnv pr), the aspartic protease of the human immunodeficiency virus (hiv-1 pr), and the human cyclin-dependent kinase ...200818578469
closing the door on flaviviruses: entry as a target for antiviral drug design.with the emergence and rapid spread of west nile virus in the united states since 1999, and the 50-100 million infections per year caused by dengue virus globally, the threat of flaviviruses as re-emerging human pathogens has become a reality. to support the efforts that are currently being pursued to develop effective vaccines against these viruses, researchers are also actively pursuing the development of small molecule compounds that target various aspects of the virus life cycle. recent adva ...200818585795
a duplex real-time reverse transcriptase polymerase chain reaction assay for the detection of st. louis encephalitis and eastern equine encephalitis viruses.a duplex taqman real-time reverse transcriptase polymerase chain reaction (pcr) assay was developed for the detection of st. louis encephalitis virus (slev) and eastern equine encephalitis virus (eeev), for use in human and vector surveillance. the respective targets selected for the assay were the conserved ns5 and e1 genes of the 2 viruses. because of the insufficient number of ns5 sequences from slev strains in the genbank database, we determined the sequence of an approximately 1-kb region f ...200818715737
differential modulation of prm cleavage, extracellular particle distribution, and virus infectivity by conserved residues at nonfurin consensus positions of the dengue virus pr-m junction.in the generation of flavivirus particles, an internal cleavage of the envelope glycoprotein prm by furin is required for the acquisition of infectivity. unlike cleavage of the prm of other flaviviruses, cleavage of dengue virus prm is incomplete in many cell lines; the partial cleavage reflects the influence of residues at furin nonconsensus positions of the pr-m junction, as flaviviruses share basic residues at positions p1, p2, and p4, recognized by furin. in this study, viruses harboring the ...200818715923
japanese encephalitis in a finnish traveler on a two-week holiday in thailand.japanese encephalitis (je) virus is a mosquito-borne flavivirus, and one of the leading causes of epidemic encephalitis in southeast asia. reports of symptomatic jev encephalitis in tourists have been rare. we describe a case of symptomatic je transmitted in 2004 during a short two-week trip to common tourist attractions in thailand.200818556242
serum antibodies to west nile virus in naturally exposed and vaccinated horses.a polyvalent elisa and plaque reduction neutralization tests (prnts) were used to measure serum antibodies to west nile virus (wnv) in horses naturally exposed to or vaccinated against this flavivirus in connecticut and new york state, usa. relying on a prnt as a 'gold standard', the main objective was to validate a modified elisa containing a recombinant wnv envelope protein antigen. it was also important to assess specificity by testing sera from horses that had other, undiagnosed illnesses. s ...200818719177
expression of the dengue virus type 2 ns5 protein in a cd4(+) t cell line inhibits hiv replication.human immunodeficiency virus (hiv) load is suppressed during dengue virus infection. the ns5a phosphoprotein of gb virus c (a related flavivirus) inhibits hiv replication in vitro. to determine whether the dengue virus ns5 protein inhibits hiv replication, cd4(+) t cell lines expressing this protein were generated. hiv replication in dengue virus ns5-expressing cells decreased by >90% compared with control cells (p < .01), and this was mediated in part by decreased hiv coreceptor (cxcr4) express ...200818721058
preliminary evaluation of exotic tick species and exotic pathogens imported on migratory birds into the british isles.field studies were carried out to determine whether ticks are being imported into the british isles on migratory birds. during spring and autumn migration 2004, ticks were collected from ringed birds at 11 bird observatories and 3 inland riparia riparia colonies. a total of 38 ticks of 4 species (ixodes ricinus, i. frontalis, i. lividus, i. arboricola) were collected from 12 species of bird. ticks were tested for viruses in the flavivirus and nairovirus genera, with no positives found. this data ...200818585865
potent cationic inhibitors of west nile virus ns2b/ns3 protease with serum stability, cell permeability and antiviral activity.west nile virus (wnv) has spread rapidly around the globe, efficiently crossing species from migrating birds into humans and other mammals. the viral protease ns2b-ns3 is important for wnv replication and recognizes dibasic substrate sequences common to other flaviviral proteases but different from most mammalian proteases. potent inhibitors of wnv protease with antiviral activity have been elusive to date. we report the smallest and most potent inhibitors known for this enzyme, cationic tripept ...200818729351
ligation-mediated amplification for effective rapid determination of viral rna sequences (rdv).emerging infectious diseases pose a significant risk to public health. methods for rapid detection of pathogens are needed to effectively treat these diseases. recently, we developed new methods for the rapid determination of viral rna sequences, rdv ver1.0 and ver2.0. we demonstrated that these methods were able to simultaneously detect cdna fragments of many different viruses without using sequence specific primers. however, some species of viruses, including the yokose virus (yokv), a flavivi ...200818595773
[serological monitoring of arbovirus infections in the estuary of the kuban river (the 2006-2007 data)].solid-phase enzyme immunoassay, neutralization test, and the hemagglutination-inhibition test were used to study the sera from human beings (152 samples), agricultural animals (n = 77), hares (n = 3), and wild birds (n = 69), collected in 2006-2007 in the kuban river estuary (temryuk district, krasnodar territory). there were specific antibodies against viruses of west nile (wh), tick-borne encephalitis (tbe) (flaviviridae, flavivirus), sindbis (togaviridae, alphavirus), the antigenic complex of ...200818756814
phosphorylation of yellow fever virus ns5 alters methyltransferase activity.serine/threonine phosphorylation of the nonstructural protein 5 (ns5) is conserved feature of flaviviruses, but the kinase(s) responsible and function(s) remain unknown. mass spectrometry was used to characterize phosphorylated residues of yellow fever virus (yfv) ns5 expressed in mammalian cells. multiple different phosphopeptides were detected. mutational and additional mass spectrometry data implicated serine 56 (s56), a conserved residue near the active site in the ns5 methyltransferase doma ...200818757072
[analysis of the efficiency of tick-borne encephalitis vaccination in the population in the natural foci of austria].tick-borne encephalitis (tbe), the preventive measure against which is vaccination, is a disease caused by flavivirus (tbe virus). the infection is common in many european countries and in much of central and eastern asia. formaline-killed whole-virion high-purified vaccine inactivated is widely used for immunization of the population in europe; however, the coverage of the population with vaccination greatly differs in the country where the natural foci of tbe are recorded. the exception is aus ...200818450105
viral membrane fusion.infection by viruses having lipid-bilayer envelopes proceeds through fusion of the viral membrane with a membrane of the target cell. viral 'fusion proteins' facilitate this process. they vary greatly in structure, but all seem to have a common mechanism of action, in which a ligand-triggered, large-scale conformational change in the fusion protein is coupled to apposition and merger of the two bilayers. we describe three examples--the influenza virus hemagglutinin, the flavivirus e protein and ...200818596815
present perspectives on flaviviral chemotherapy.flaviviruses are a group of arboviruses under the family of flaviviridae. flaviviruses are global pathogens, some of which cause neurotrophic disorders from mild febrile illness to lethal encephalitis while others cause severe hemorrhagic fever. these viruses are endemic to many parts of the world and periodic outbreaks take thousands of lives. with globalization of the world accompanied by gradual shift in global climate, the viruses are emerging in newer areas and we are in a point of exigency ...200818598919
mutation of mapped tia-1/tiar binding sites in the 3' terminal stem-loop of west nile virus minus-strand rna in an infectious clone negatively affects genomic rna amplification.previous data showed that the cellular proteins tia-1 and tiar bound specifically to the west nile virus 3' minus-strand stem-loop [wnv3'(-)sl] rna (37) and colocalized with flavivirus replication complexes in wnv- and dengue virus-infected cells (21). in the present study, the sites on the wnv3'(-)sl rna required for efficient in vitro t-cell intracellular antigen-related (tiar) and t-cell intracellular antigen-1 (tia-1) protein binding were mapped to short au sequences (uaauu) located in two i ...200818768985
synthesis and study of 9-deazaguanosine derivatives as potential inhibitors of rna virus replication.9-deazaguanosine and the alpha and beta anomers of its 2'-c-methyl counter part, have been synthesized and evaluated against a broad range of rna viruses, including hepatitis c virus.200818776524
flavivirus dna vaccine with a kick. 200818464782
early b-cell activation after west nile virus infection requires alpha/beta interferon but not antigen receptor signaling.the b-cell response against west nile virus (wnv), an encephalitic flavivirus of global concern, is critical to controlling central nervous system dissemination and neurological sequelae, including death. here, using a well-characterized mouse model of wnv infection, we examine the factors that govern early b-cell activation. subcutaneous inoculation with a low dose of replicating wnv results in extensive b-cell activation in the draining lymph node (ln) within days of infection as judged by upr ...200818786989
interferon antagonist function of japanese encephalitis virus ns4a and its interaction with dead-box rna helicase ddx42.the interferon (ifn) antagonists of japanese encephalitis virus (jev) proteins contribute to the je pathogenesis. most flavivirus non-structural (ns) proteins correlate with virus-induced inflammation and immune escape. ns4a proteins of west nile virus and dengue type 2 virus have been demonstrated to inhibit ifn signaling. in this study, jev ns4a without the c-terminal 2k domain has been demonstrated to partially block activation of an ifn-stimulated response element (isre)-based cis-reporter b ...200818588927
[isolation of influenza virus a (orthomyxoviridae, influenza a virus), dhori virus (orthomyxoviridae, thogotovirus), and newcastle's disease virus (paromyxoviridae, avulavirus) on the malyi zhemchuzhnyi island in the north-western area of the caspian sea].the paper presents the results of the 2003 and 2006 environmental virological monitoring surveys on the malyi zhemchuzhnyi island where a large breeding colony of sea gull (laridae) is located. in the past several years, expansion of cormorants (phalacrocorax carbo) has enhanced the intensity of populational interactions. the investigators isolated 13 strains of influenza a virus (orthomyxoviridae, influenza a virus) subtype h13n1 (from sea gulls (n = 4), cormorants (n = 9) 1 strain of dhori vir ...200818590134
association of hepatitis c virus replication complexes with microtubules and actin filaments is dependent on the interaction of ns3 and ns5a.the hepatitis c virus (hcv) rna replication complex (rc), which is composed of viral nonstructural (ns) proteins and host cellular proteins, replicates the viral rna genome in association with intracellular membranes. two viral ns proteins, ns3 and ns5a, are essential elements of the rc. here, by using immunoprecipitation and fluorescence resonance energy transfer assays, we demonstrated that ns3 and ns5a interact with tubulin and actin. furthermore, immunofluorescence microscopy and electron mi ...200818562541
dengue vaccines for travelers.dengue is an arthropod-borne infection caused by a flavivirus and spread by the aedes mosquitoes. many of the countries where dengue is endemic are popular tourist destinations and the disease is an increasingly important problem encountered by international travelers. personal protection against the day-feeding dengue vectors is problematic, indicating the urgent need for a dengue vaccine. this review discusses the challenges of vaccine development, current vaccine strategies and the prospects ...200818564012
clinical isolates of dengue virus with distinctive susceptibility to nitric oxide radical induce differential gene responses in thp-1 cells.in the present study, 10 clinical isolates of dengue virus were selected according to their susceptibility to the inhibitory effect of nitric oxide radical, no. five of them are nitric oxide-susceptible viruses while the other five are nitric oxide-resistant viruses. these isolates were investigated to identify genetic factors that are responsible for the different phenotypes. due to the evidence showing that no suppresses denv rna polymerase activity, we, therefore, hypothesized that the rdrp d ...200818455750
novel binding between pre-membrane protein and vacuolar atpase is required for efficient dengue virus secretion.flavivirus pre-membrane (prm) protein is important for proper folding and secretion of envelope (e) protein. however, other non-structural functions of prm protein in the context of virus life-cycle are poorly known. in this study, we aimed to elucidate if dengue virus (dv) prm protein interacts with host proteins and contributes to viral pathogenesis by screening human liver cdna yeast-two-hybrid library. our study identified vacuolar atpase (v-atpase) as a novel interacting partner of dv prm p ...200818573235
genetic study of the distribution of greek goat encephalitis virus in greece.tick-borne encephalitis virus (tbev) and louping ill virus (liv) are viruses in the mammalian tick-borne virus group/genus flavivirus, causing central nervous system disease. greek goat encephalitis virus (ggev), which was isolated from the brain of a newborn goat with neurological symptoms, is currently classified in the tbev group. the vector of ggev has not yet been specifically identified but is considered likely to be ixodes ricinus. a total of 3,144 ixodidae ticks collected during 2003-200 ...200818471057
identification and biochemical characterization of small-molecule inhibitors of west nile virus serine protease by a high-throughput screen.west nile virus and dengue virus are mosquito-borne flaviviruses that cause a large number of human infections each year. no vaccines or chemotherapeutics are currently available. these viruses encode a serine protease that is essential for polyprotein processing, a required step in the viral replication cycle. in this study, a high-throughput screening assay for the west nile virus protease was employed to screen approximately 32,000 small-molecule compounds for identification of inhibitors. le ...200818606844
key steps in the structure-based optimization of the hepatitis c virus ns3/4a protease inhibitor sch503034.the structures of both native and s139a holo-hcv ns3/4a protease domain were solved to high resolution. subsequently, structures were determined for a series of ketoamide inhibitors in complex with the protease. the changes in the inhibitor potency were correlated with changes in the buried surface area upon binding the inhibitor to the active site. the largest contributions to the binding energy arise from the hydrophobic interactions of the p1 and p2 groups as they bind to the s1 and s2 pocket ...200818421139
design, synthesis, and biological evaluation of antiviral agents targeting flavivirus envelope proteins.flavivirus envelope proteins (e proteins) have been shown to play a pivotal role in virus assembly, morphogenesis, and infection of host cells. inhibition of flavivirus infection of a host cell by means of a small molecule envelope protein antagonist is an attractive strategy for the development of antiviral agents. virtual screening of the nci chemical database using the dengue virus envelope protein structure revealed several hypothetical hit compounds. bioassay results identified a class of t ...200818610998
guidelines for plaque-reduction neutralization testing of human antibodies to dengue viruses.through the advisory committee on dengue and other flavivirus vaccines, the world health organization(who) has had a long-standing commitment to facilitate and to guide research and development of vaccines for medically important flaviviruses. recently, the paediatric dengue vaccine initiative (pdvi) was formed to accelerate the development, testing, and introduction of dengue (den)vaccines worldwide, partnering with who in this important public health effort. there are now a variety of den vacc ...200818476771
specific genetic markers for detecting subtypes of dengue virus serotype-2 in isolates from the states of oaxaca and veracruz, mexico.dengue (den) is an infectious disease caused by the den virus (denv), which belongs to the flavivirus genus in the family flaviviridae. it has a (+) sense rna genome and is mainly transmitted to humans by the vector mosquito aedes aegypti. dengue fever (df) and dengue hemorrhagic fever (dhf) are caused by one of four closely related virus serotypes (denv-1, denv-2, denv-3 and denv-4). epidemiological and evolutionary studies have indicated that host and viral factors are involved in determining ...200818625078
tumor necrosis factor alpha protects against lethal west nile virus infection by promoting trafficking of mononuclear leukocytes into the central nervous system.west nile virus (wnv) is a neurotropic flavivirus that has emerged globally as a significant cause of viral encephalitis in humans, especially in immunocompromised individuals. previous studies have shown essential protective roles for antiviral cytokines (e.g., alpha interferon [ifn-alpha] and ifn-gamma) against wnv in mice. however, studies using cell culture offer conflicting answers regarding whether tumor necrosis factor alpha (tnf-alpha) has an anti-wnv function. to test the biological sig ...200818632856
differential cholesterol binding by class ii fusion proteins determines membrane fusion properties.the class ii fusion proteins of the alphaviruses and flaviviruses mediate virus infection by driving the fusion of the virus membrane with that of the cell. these fusion proteins are triggered by low ph, and their structures are strikingly similar in both the prefusion dimer and the postfusion homotrimer conformations. here we have compared cholesterol interactions during membrane fusion by these two groups of viruses. using cholesterol-depleted insect cells, we showed that fusion and infection ...200818632857
construction and evaluation of a chimeric pseudoinfectious virus vaccine to prevent japanese encephalitis.multiple vaccines exist to control japanese encephalitis (je), but all suffer from problems. we have developed a new type of flavivirus vaccine, a pseudoinfectious virus (replivax wn) that prevents west nile virus (wnv)-induced disease. here, we describe production of a chimeric replivax (replivax je) that expresses the je virus (jev) prm and e proteins. our prototype replivax je replicated poorly in cells, but blind passage produced a better-growing derivative, and analyses of this derivative a ...200818433947
dengue in travellers is still underestimated.dengue fever (df), one of the main emerging virus diseases, also increases in german travellers, due to the increasing number of travellers to tropical regions.200818477850
improvement of the purification of saint louis encephalitis virus ns2b-ns3 recombinant protease expressed in escherichia coli.the ns2b-ns3 serine protease of saint louis encephalitis virus (slev), a potential target for antiviral drug design, has been over-expressed as a recombinant his-tag protein in escherichia coli for future structural determination. the production process resulted in a soluble protease with co-purification of dnak, a bacterial molecular chaperone already described in e. coli protein expression. two approaches were tested to remove this specific contaminant. the fusion protein bound to the purifica ...200818485841
molecular characterization of two rocio flavivirus strains isolated during the encephalitis epidemic in são paulo state, brazil and the development of a one-step rt-pcr assay for diagnosis.rocio virus (rocv) was responsible for an explosive encephalitis epidemic in the 1970s affecting about 1,000 residents of 20 coastland counties in são paulo state, brazil. rocv was first isolated in 1975 from the cerebellum of a fatal human case of encephalitis. clinical manifestations of the illness are similar to those described for st. louis encephalitis. rocv shows intense antigenic cross-reactivity with japanese encephalitis complex (jec) viruses, particularly with ilheus (ilhv), st. louis ...200818488087
a comparative biochemical analysis of the ns2b(h)-ns3pro protease complex from four dengue virus serotypes.the two-component protease ns2b-ns3 of dengue virus mediates proteolytic processing of the polyprotein precursor and therefore represents a target for the development of antiviral drugs. the amino acid sequences of the ns3 serine protease and the ns2b cofactor exhibit relatively low degrees of conservation among the 4 serotypes thus suggesting that differences in enzyme activity exist which could modulate their susceptibility to future protease inhibitors. in this study we have addressed the que ...200818439431
a complex rna motif defined by three discontinuous 5-nucleotide-long strands is essential for flavivirus rna replication.tertiary or higher-order rna motifs that regulate replication of positive-strand rna viruses are as yet poorly understood. using japanese encephalitis virus (jev), we now show that a key element in jev rna replication is a complex rna motif that includes a string of three discontinuous complementary sequences (tdcs). the tdcs consists of three 5-nt-long strands, the left (l) strand upstream of the translation initiator aug adjacent to the 5'-end of the genome, and the middle (m) and right (r) st ...200818669441
standardized quantitative rt-pcr assays for quantitation of yellow fever and chimeric yellow fever-dengue vaccines.yellow fever-dengue chimeras (cyds) are being developed currently as live tetravalent dengue vaccine candidates. specific quantitative assays are needed to evaluate the viral load of each serotype in vaccine batches and biological samples. a quantitative real-time rt-pcr (qrt-pcr) system was developed comprising five one-step qrt-pcrs targeting the e/ns1 junction of each chimera, or the ns5 gene in the yellow fever backbone. each assay was standardized using in vitro transcribed rna qualified ac ...200818501437
a survey on cases of tick-borne encephalitis in european countries.the european network for diagnostics of "imported" viral diseases (enivd) is finalising a project to improve the diagnostic and monitoring of encephalitis viruses in europe. part of this study was to analyse the present surveillance situation for tick-borne encephalitis (tbe), which is the most important flavivirus infection of the central nervous system in the european union (eu) and russia. a questionnaire was mailed to contact points in all member states of the eu and three non-eu countries ( ...200818445446
limited replication of yellow fever 17dd and 17d-dengue recombinant viruses in rhesus monkeys.for the development of safe live attenuated flavivirus vaccines one of the main properties to be established is viral replication. we have used real-time reverse transcriptase-polymerase chain reaction and virus titration by plaque assay to determine the replication of yellow fever 17dd virus (yfv 17dd) and recombinant yellow fever 17d viruses expressing envelope proteins of dengue virus serotypes 2 and 4 (17d-denv-2 and 17d-denv-4). serum samples from rhesus monkeys inoculated with yfv 17dd and ...200818506257
phylogenetic and evolutionary analyses of st. louis encephalitis virus genomes.st. louis encephalitis virus belongs to the japanese encephalitis virus serocomplex of the genus flavivirus, family flaviviridae. since the first known epidemic in 1933, the virus has been isolated from a variety of geographical, temporal, and host origins. we have sequenced 10,236 nucleotides of the open reading frame (93.6% of the full-length genome) of 23 of these strains, and have used the sequences to conduct phylogenetic analyses, in order to investigate the forces shaping the evolution of ...200818374605
the capsid-coding region hairpin element (chp) is a critical determinant of dengue virus and west nile virus rna synthesis.dengue virus (denv) and west nile virus (wnv) are members of the flavivirus genus of positive-strand rna viruses. rna sequences and structures, primarily in the untranslated regions, have been shown to modulate flaviviral gene expression and genome replication. previously, we demonstrated that a structure in the denv coding region (chp) enhances translation start codon selection and is required for viral replication. here we further characterize the role of the chp in the denv life cycle. we dem ...200818676000
genetic and serologic properties of zika virus associated with an epidemic, yap state, micronesia, 2007.zika virus (zikv) is a mosquito-borne flavivirus first isolated in uganda from a sentinel monkey in 1947. mosquito and sentinel animal surveillance studies have demonstrated that zikv is endemic to africa and southeast asia, yet reported human cases are rare, with <10 cases reported in the literature. in june 2007, an epidemic of fever and rash associated with zikv was detected in yap state, federated states of micronesia. we report the genetic and serologic properties of the zikv associated wit ...200818680646
tick-borne encephalitis.we review the epidemiological and clinical characteristics of tick-borne encephalitis, and summarise biological and virological aspects that are important for understanding the life-cycle and transmission of the virus. tick-borne encephalitis virus is a flavivirus that is transmitted by ixodes spp ticks in a vast area from western europe to the eastern coast of japan. tick-borne encephalitis causes acute meningoencephalitis with or without myelitis. morbidity is age dependent, and is highest in ...200818514730
communicable diseases network australia national arbovirus and malaria advisory committee annual report, 2006-07.this report describes the epidemiology of mosquito-borne disease in australia for the mosquito-borne disease season 1 july 2006 to 30 june 2007, which was moderately low compared to previous seasons. ross river virus (rrv) infections (55%), barmah forest virus (bfv) infections (29%) and overseas acquired malaria (11%) were the most common mosquito-borne diseases reported in 2006-07. the number, proportion and rate of national bfv notifications were the second highest on record since 1998-99. the ...200818522303
analysis of the effects of alterations in the tick-borne encephalitis virus 3'-noncoding region on translation and rna replication using reporter replicons.the 3'-noncoding region (3'-ncr) of the flavivirus genome includes a variable region that tolerates the insertion of heterologous genetic information. natural isolates of tick-borne encephalitis virus (tbev) have particularly long variable regions, which, for some strains, include an internal poly(a) tract. we constructed luciferase reporter replicons of tbev to analyze the impact of various manipulations of the 3'-ncr on viral rna translation and replication. the choice of the reporter gene, it ...200818533218
impact of aerial spraying of pyrethrin insecticide on culex pipiens and culex tarsalis (diptera: culicidae) abundance and west nile virus infection rates in an urban/suburban area of sacramento county, california.in response to an epidemic amplification of west nile virus (family flaviviridae, genus flavivirus, wnv), the sacramento and yolo mosquito and vector control district (symvcd) sprayed ultralow-volume (ulv) formulations of pyrethrin insecticide (evergreen ec 60-6: 6% pyrethrin insecticide, 60% piperonylbutoxide; mgk, minneapolis, mn, applied as 0.003 kg/ha [0.0025 lb/acre] ) over 218 km2 in north sacramento and 243.5 km2 in south sacramento on three consecutive evenings in august 2005. we evaluat ...200818714879
preservation of a critical epitope core region is associated with the high degree of flaviviral cross-reactivity exhibited by a dengue-specific cd4+ t cell clone.dengue is a member of the flaviviridae, a large group of related viruses some of which co-circulate in certain regions (e.g. dengue and yellow fever in south america). immune responses cross-reactive between different dengue serotypes are important in the pathogenesis of dengue disease but it is not known whether previous infection with one flavivirus might affect the clinical course of subsequent infections with other members of the family. cd4+ t cells have been shown to be important in the pr ...200818383038
effective inhibition of japanese encephalitis virus replication by small interfering rnas targeting the ns5 gene.japanese encephalitis virus (jev), a mosquito-borne flavivirus, causes an acute infection of the central nervous system resulting in encephalitis of humans and many kinds of animals. ns5, the largest and most conserved flavivirus protein, is homologous to methyltransferase and rna-dependent rna polymerase. rna interference is an effective anti-viral strategy to inhibit viral replication in vitro. in this study, four short hairpin rna (shrna) expression vectors (ps4.1-ns5-201, ps4.1-ns5-455, ps4. ...200818190994
host-seeking heights, host-seeking activity patterns, and west nile virus infection rates for members of the culex pipiens complex at different habitat types within the hybrid zone, shelby county, tn, 2002 (diptera: culicidae).host-seeking heights, host-seeking activity patterns, and west nile virus (family flaviviridae, genus flavivirus, wnv) infection rates were assessed for members of the culex pipiens complex from july to december 2002, by using chicken-baited can traps (ct) at four ecologically different sites in shelby county, tn. host-seeking height was assessed by ct placed at elevations of 3.1, 4.6, and 7.6 m during one 24-h period per month. host-seeking activity was assessed by paired ct placed at an elevat ...200818402144
crystal structure of the catalytic domain of japanese encephalitis virus ns3 helicase/nucleoside triphosphatase at a resolution of 1.8 a.the ns3 protein of japanese encephalitis virus (jev) is a large multifunctional protein possessing protease, helicase, and nucleoside 5'-triphosphatase (ntpase) activities, and plays important roles in the processing of a viral polyprotein and replication. to clarify the enzymatic properties of ns3 protein from a structural point of view, an enzymatically active fragment of the jev ntpase/helicase catalytic domain was expressed in bacteria and the crystal structure was determined at 1.8 a resolu ...200818201743
construction and characterization of a second-generation pseudoinfectious west nile virus vaccine propagated using a new cultivation system.safer vaccines are needed to prevent flavivirus diseases. to help develop these products we have produced a pseudoinfectious west nile virus (wnv) lacking a functional c gene which we have named replivax wn. here we demonstrate that replivax wn can be safely propagated at high titer in bhk cells and vaccine-certified vero cells engineered to stably express the c protein needed to trans-complement replivax wn growth. using these bhk cells we selected a better growing mutant replivax wn population ...200818423946
single-round infectious particles enhance immunogenicity of a dna vaccine against west nile virus.dna vaccines encoding replication-defective viruses are safer than inactivated or live attenuated viruses but may fail to stimulate an immune response sufficient for effective vaccination. we augment the protective capacity of a capsid-deleted flavivirus dna vaccine by co-expressing the capsid protein from a separate promoter. in transfected cells, the capsid-deleted rna transcript is replicated and translated to produce secreted virus-like particles lacking the nucleocapsid. this rna is also pa ...200818425125
eared dove (zenaida auriculata, columbidae) as host for st. louis encephalitis virus (flaviviridae, flavivirus).st. louis encephalitis virus (slev) is an emerging flavivirus in south american countries. its ecology and biological transmission cycles are scarcely known. eared doves (zenaida auriculata) have frequently been found infected by slev, and therefore, could be suspected as slev hosts. thirty post-hatch-year eared doves were subcutaneously inoculated with the genotype v slev 78v-6507 viral strain and subsequently bled. no deaths or clinical signs of illness were observed in the inoculated doves. t ...200818429694
serologic survey of potential vertebrate hosts for west nile virus in poland.a survey for antibodies to west nile virus (wnv; genus ,flavivirus) was carried out by plaque-re-duction neutralization microtesting in 78 horses, 20 domestic chickens, and 97 wild birds belonging to 10 species from different areas in poland. specific antibodies were detected in five juvenile (hatching-year) birds collected in 2006: three white storks (ciconia ciconia) in a wildlife rehabilitation center (5.4% of all examined storks; the antibody titers in each bird were 1:320, 1:160, and 1:20), ...200818433332
human outbreak of st. louis encephalitis detected in argentina, 2005.an outbreak of flavivirus encephalitis occurred in 2005 in córdoba province, argentina.200818249032
pathogenic flaviviruses.haemorrhagic disease, encephalitis, biphasic fever, flaccid paralysis, and jaundice are typical manifestations of diseases in human beings after infections by mosquito-borne or tick-borne flaviviruses such as yellow fever, dengue, west nile, st louis encephalitis, japanese encephalitis, tick-borne encephalitis, kyasanur forest disease, and omsk haemorrhagic fever. although the characteristics of these viruses are well defined, they are still unpredictable with increases in disease severity, unus ...200818262042
the two-component ns2b-ns3 proteinase represses dna unwinding activity of the west nile virus ns3 helicase.similar to many flavivirus types including dengue and yellow fever viruses, the nonstructural ns3 multifunctional protein of west nile virus (wnv) with an n-terminal serine proteinase domain and an rna triphosphatase, an ntpase domain, and an rna helicase in the c-terminal domain is implicated in both polyprotein processing and rna replication and is therefore a promising drug target. to exhibit its proteolytic activity, ns3 proteinase requires the presence of the cofactor encoded by the upstrea ...200818442976
viral haemorrhagic fever and vascular alterations.pathogenesis of viral haemorrhagic fever (vhf) is closely associated with alterations of the vascular system. among the virus families causing vhf, filoviruses (marburg and ebola) are the most fatal, and will be focused on here. after entering the body, ebola primarily targets monocytes/macrophages and dendritic cells. infected dendritic cells are largely impaired in their activation potency, likely contributing to the immune suppression that occurs during filovirus infection. monocytes/macropha ...200818278167
insights into the oligomerization state-helicase activity relationship of west nile virus ns3 ntpase/helicase.west nile virus (wnv) is a member of the flaviviridae family of positive-strand rna viruses. its viral rna is translated to produce a polyprotein precursor that is further processed into three structural and seven non-structural proteins. the non-structural protein 3 (ns3) possess both protease and helicase activities. the c-terminal portion of the ns3 contains the atpase/helicase domain presumably involved in viral replication. this domain has been expressed in escherichia coli, purified in sol ...200818448185
genetic interactions among the west nile virus methyltransferase, the rna-dependent rna polymerase, and the 5' stem-loop of genomic rna.flavivirus methyltransferase catalyzes both guanine n7 and ribose 2'-oh methylations of the viral rna cap (gpppa-rna-->m(7)gpppam-rna). the methyltransferase is physically linked to an rna-dependent rna polymerase (rdrp) in the flaviviral ns5 protein. here, we report genetic interactions of west nile virus (wnv) methyltransferase with the rdrp and the 5'-terminal stem-loop of viral genomic rna. genome-length rnas, containing amino acid substitutions of d146 (a residue essential for both cap meth ...200818448528
cholesterol effectively blocks entry of flavivirus.japanese encephalitis virus (jev) and dengue virus serotype 2 (den-2) are enveloped flaviviruses that enter cells through receptor-mediated endocytosis and low ph-triggered membrane fusion and then replicate in intracellular membrane structures. lipid rafts, cholesterol-enriched lipid-ordered membrane domains, are platforms for a variety of cellular functions. in this study, we found that disruption of lipid raft formation by cholesterol depletion with methyl-beta-cyclodextrin or cholesterol che ...200818448543
long-term balancing selection at the west nile virus resistance gene, oas1b, maintains transspecific polymorphisms in the house mouse.oligoadenylate synthetases (oass) are interferon-inducible enzymes that participate in the first line of defense against a wide range of viral infection. recent studies have determined that oas1b, a member of the oas gene family in the house mouse (mus musculus), provides specific protection against flavivirus infection (e.g., west nile virus, dengue fever virus, and yellow fever virus). we characterized the nucleotide sequence variation in coding and noncoding regions of the oas1b gene for a la ...200818460447
kunjin replicon-based simian immunodeficiency virus gag vaccines.an rna-based, non-cytopathic replicon vector system, based on the flavivirus kunjin, has shown considerable promise as a new vaccine delivery system. here we describe the testing in mice of four different sivmac239 gag vaccines delivered by kunjin replicon virus-like-particles. the four vaccines encoded the wild type gag gene, an rna-optimised gag gene, a codon-optimised gag gene and a modified gag-pol gene construct. the vaccines behaved quite differently for induction of effector memory and ce ...200818462846
[study of the diagnostic potential of various regions of tick-borne encephalitis virus glycoprotien e].bioinformation techniques were used to analyze envelope glycoprotein of tick-borne encephalitis virus in order to determine the potential diagnostically significant antigenic clusters. five selected regions of the amino acid sequence of protein e were retranslated to nucleic sequences, by employing the optimum code for e. coli. the resultant dna fragments were synthesized by polymerase chain reaction from synthetic oligonucleotides and expressed in e. coli cells. the recombinant antigen replicat ...200818318135
persistence of antibodies to west nile virus nonstructural protein 5.because igm antibody against west nile virus (wnv) pre-membrane/envelope (prem/e) recombinant protein may persist for more than 1 year, an assay distinguishing recent from past wnv infection would be useful. published findings for a single patient suggest that the presence of antibody against wnv nonstructural protein 5 (ns5) indicates recent infection.200818467165
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