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viral vaccine meeting held in barcelona, october 25-28, 2003. 200415098545
conserved rna secondary structures in flaviviridae genomes.presented here is a comprehensive computational survey of evolutionarily conserved secondary structure motifs in the genomic rnas of the family flaviviridae: this virus family consists of the three genera flavivirus, pestivirus and hepacivirus and the group of gb virus c/hepatitis g virus with a currently uncertain taxonomic classification. based on the control of replication and translation, two subgroups were considered separately: the genus flavivirus, with its type i cap structure at the 5' ...200415105528
conserved amino acids 193-324 of non-structural protein 3 are a dominant source of peptide determinants for cd4+ and cd8+ t cells in a healthy japanese encephalitis virus-endemic cohort.our earlier identification of the non-structural protein 3 (ns3) of japanese encephalitis virus (jev) as a dominant cd4+ as well as cd8+ t cell-eliciting antigen in a healthy jev-endemic cohort with a wide hla distribution implied the presence of several epitopes dispersed over the length of the protein. use of various truncated versions of ns3 in lymphocyte stimulation and interferon (ifn)-gamma secretion assays revealed that amino acids (aa) 193-324 of ns3 were comparable with, if not superior ...200415105530
accumulation of a 3'-terminal genome fragment in japanese encephalitis virus-infected mammalian and mosquito cells.japanese encephalitis virus (jev) contains a single positive-strand rna genome nearly 11 kb in length and is not formally thought to generate subgenomic rna molecules during replication. here, we report the abundant accumulation of a 3'-terminal 521- to 523-nucleotide (nt) genome fragment, representing a major portion of the 585-nt 3' untranslated region, in both mammalian (bhk-21) and mosquito (c6/36) cells infected with any of nine strains of jev. in bhk-21 cells, the viral genome was detected ...200415113895
purification and crystallization reveal two types of interactions of the fusion protein homotrimer of semliki forest virus.the fusion proteins of the alphaviruses and flaviviruses have a similar native structure and convert to a highly stable homotrimer conformation during the fusion of the viral and target membranes. the properties of the alpha- and flavivirus fusion proteins distinguish them from the class i viral fusion proteins, such as influenza virus hemagglutinin, and establish them as the first members of the class ii fusion proteins. understanding how this new class carries out membrane fusion will require ...200415016874
longitudinal studies of west nile virus infection in avians, yucatán state, méxico.following the introduction of west nile virus (wnv) into north america in 1999, surveillance for evidence of infection with this virus in migratory and resident birds was established in yucatán state, méxico in march 2000. overall, 8611 birds representing 182 species and 14 orders were captured and assayed for antibodies to wnv. of these, 5066 (59%) birds were residents and 3545 (41%) birds were migrants. twenty-one (0.24%) birds exhibited evidence of flavivirus infection. of these, 8 birds had ...200415018768
detection of australasian flavivirus encephalitic viruses using rapid fluorogenic taqman rt-pcr assays.the development of single, sensitive, fluorogenic reverse transcriptase-polymerase chain reaction (taqman) assays were required for the rapid and specific detection of three encephalitic viruses found in the australasian region, namely; japanese encephalitis virus (jev), murray valley encephalitis virus (mvev), and kunjin virus (kunv). primers and a fluorogenic probe were individually designed to be complementary to a nucleotide region encompassing the 3' terminus of the nonstructural (ns) 5 gen ...200415041213
phylogenetic analysis of bovine viral diarrhea viruses using five different genetic regions.phylogenetic analysis of the five different regions (5' non-coding region (5'ncr), n(pro), e2, ns3 and ns5b-3'ncr) of 48 japanese and reported bovine viral diarrhea virus (bvdv) genomes was performed. japanese bvdvs were segregated into bvdv1 subdivided into six subgroups and bvdv2. one isolate, so cp/75, isolated in 1975 and previously proposed as subgroup 1e according to its 5'ncr sequence, was quite unique and formed an independent lineage in the tree of any region. another isolate, 190cp, ob ...200414749175
fatal west nile virus encephalitis in a renal transplant recipient.west nile virus (wnv), a mosquito-transmitted single-stranded rna flavivirus, causes human disease of variable severity. we report clinical and pathologic findings of fatal encephalitis from the transmission of wnv from an organ donor to a kidney transplant recipient. the patient developed a febrile illness 18 days after transplantation, which progressed to encephalitis. postmortem examination demonstrated extensive viral encephalopathic changes. immunohistochemical studies highlighted wnv antig ...200414750237
detection of human anti-flavivirus antibodies with a west nile virus recombinant antigen microsphere immunoassay.we report a new, suspended-microsphere diagnostic test to detect antibodies to west nile (wn) virus in human serum and cerebrospinal fluid (csf). the microsphere immunofluorescence assay can be performed in less than 3 h on specimens of <or=30 microl. a recombinant wn virus envelope (e) protein antigen is covalently coupled to fluorescent polystyrene microspheres. after incubation with diluted serum or csf, antibodies bound to the e protein antigen are detected with fluorescently labeled anti-hu ...200414715733
mimicking live flavivirus immunization with a noninfectious rna vaccine.flaviviruses are human pathogens of world-wide medical importance. they have recently received much additional attention because of their spread to new regions (such as west nile virus to north america), highlighting their potential as newly emerging disease agents. using tick-borne encephalitis virus, we have developed and evaluated in mice a new genetic vaccine based on self-replicating but noninfectious rna. this rna contains all of the necessary genetic information for establishing its repli ...200414769933
safety and immunogenicity of a three dose regimen of two tetravalent live-attenuated dengue vaccines in five- to twelve-year-old thai children.the safety and immunogenicity of tetravalent live-attenuated dengue vaccines after a three dose vaccination series were evaluated in thai children.200414872173
alterations of pr-m cleavage and virus export in pr-m junction chimeric dengue viruses.during the export of flavivirus particles through the secretory pathway, a viral envelope glycoprotein, prm, is cleaved by the proprotein convertase furin; this cleavage is required for the subsequent rearrangement of receptor-binding e glycoprotein and for virus infectivity. similar to many furin substrates, prm in vector-borne flaviviruses contains basic residues at positions p1, p2, and p4 proximal to the cleavage site; in addition, a number of charged residues are found at position p3 and be ...200414963133
contribution of disulfide bridging to epitope expression of the dengue type 2 virus envelope glycoprotein.the individual contributions of each of the six conserved disulfide (ss) bonds in the dengue 2 virus envelope (e) glycoprotein (strain 16681) to epitope expression was determined by measuring the reactivities of a panel of well-defined monoclonal antibodies (mabs) with llc-mk(2) cells that had been transiently transformed with plasmid vectors expressing e proteins that were mutant in their ss bonds. three domain i (di) epitopes (c1, c3, and c4) were affected by elimination of any ss bond and wer ...200414963174
structure of a flavivirus envelope glycoprotein in its low-ph-induced membrane fusion conformation.enveloped viruses enter cells via a membrane fusion reaction driven by conformational changes of specific viral envelope proteins. we report here the structure of the ectodomain of the tick-borne encephalitis virus envelope glycoprotein, e, a prototypical class ii fusion protein, in its trimeric low-ph-induced conformation. we show that, in the conformational transition, the three domains of the neutral-ph form are maintained but their relative orientation is altered. similar to the postfusion c ...200414963486
impaired t helper 1 function of nonstructural protein 3-specific t cells in japanese patients with encephalitis with neurological sequelae.japanese encephalitis virus (jev) is the principal cause of viral encephalitis. the predominance of children among patients with encephalitis in areas where jev is endemic that do not have immunization programs suggests that acquired immunity is critical in protecting adults against symptomatic jev encephalitis. we characterized and compared the t cell response to nonstructural (ns) protein 3 between healthy individuals naturally exposed to jev and patients in the convalescent phase of jev encep ...200414976606
recombinant kunjin virus replicon vaccines induce protective t-cell immunity against human papillomavirus 16 e7-expressing tumour.the persistence of the e7 oncoprotein in transformed cells in human papillomavirus (hpv)-associated cervical cancer provides a tumour-specific antigen to which immunotherapeutic strategies may be directed. self-replicating rna (replicon) vaccine vectors derived from the flavivirus kunjin (kun) have recently been reported to induce t-cell immunity. here, we report that inclusion of a ctl epitope of hpv16 e7 protein into a polyepitope encoded by a kun vector induced e7-directed t-cell responses an ...200414980484
determination of the disulfide bond arrangement of dengue virus ns1 protein.the 12 half-cystines of ns1 proteins are absolutely conserved among flaviviruses, suggesting their importance to the structure and function of these proteins. in the present study, peptides from recombinant dengue-2 virus ns1 were produced by tryptic digestion in 100% h(2)(16)o, peptic digestion in 50% h(2)(18)o, thermolytic digestion in 50% h(2)(18)o, or combinations of these digestion conditions. peptides were separated by size exclusion and/or reverse phase high performance liquid chromatogra ...200414981082
interim guidelines for the evaluation of infants born to mothers infected with west nile virus during pregnancy.west nile virus (wnv) is a single-stranded rna flavivirus with antigenic similarities to japanese encephalitis and st. louis encephalitis viruses. it is transmitted to humans primarily through the bites of infected mosquitoes. flavivirus infection during pregnancy has been associated rarely with both spontaneous abortion and neonatal illness but has not been known to cause birth defects in humans. during 2002, a total of 4,156 cases of wnv illness in humans, including 2,946 cases of neuroinvasiv ...200414985654
west nile virus: an overview of its spread in europe and the mediterranean basin in contrast to its spread in the americas.west nile (wn) virus is a mosquito-transmitted flavivirus. it is widely distributed in africa, the middle east, asia, and southern europe and was recently introduced to north america. birds are involved in the cycle of transmission as amplifying hosts. humans and horses are considered accidental dead-end hosts. wn fever was initially considered a minor arbovirosis, usually inducing a nonsymptomatic or a mild flu-like illness in humans, but some cases of encephalitis associated with fatalities we ...200414986160
characterization of a membrane-associated trimeric low-ph-induced form of the class ii viral fusion protein e from tick-borne encephalitis virus and its crystallization.the interaction of a dimeric membrane anchor-free form of the envelope protein e (se dimer) from tick-borne encephalitis virus with liposomes at acidic ph levels leads to its conversion into membrane-inserted se trimers. electron microscopy shows that these trimers have their long dimensions along the threefold molecular axis, which is oriented perpendicularly to the plane of the membrane, where the protein inserts via the internal fusion peptide. liposomes containing se at their surface display ...200414990739
virus spread, tissue inflammation and antiviral response in brains of flavivirus susceptible and resistant mice acutely infected with murray valley encephalitis virus.inborn resistance to flaviviruses, conferred by a single chromosome 5 locus flv, is a genetic trait operative in wild mice and a few strains of laboratory mice. in this study we have used in situ hybridisation to trace the spread of flavivirus genomic rna within the brains of flavivirus susceptible c3h/hejarc and congenic resistant c3h.pri- flv(r) mice following infection with murray valley encephalitis virus (mve) in parallel to studying a brain histopathology and induction of cellular genes in ...200414991436
solution structure of dengue virus capsid protein reveals another fold.dengue virus is responsible for approximately 50-100 million infections, resulting in nearly 24,000 deaths annually. the capsid (c) protein of dengue virus is essential for specific encapsidation of the rna genome, but little structural information on the c protein is available. we report the solution structure of the 200-residue homodimer of dengue 2 c protein. the structure provides, to our knowledge, the first 3d picture of a flavivirus c protein and identifies a fold that includes a large di ...200414993605
real-time reverse transcription loop-mediated isothermal amplification for rapid detection of west nile virus.a one-step, single tube, real-time accelerated reverse transcription loop-mediated isothermal amplification (rt-lamp) assay was developed for detecting the envelope gene of west nile (wn) virus. the rt-lamp assay is a novel method of gene amplification that amplifies nucleic acid with high specificity, efficiency, and rapidity under isothermal conditions with a set of six specially designed primers that recognize eight distinct sequences of the target. the whole procedure is very simple and rapi ...200414715762
conformational changes of the flavivirus e glycoprotein.dengue virus, a member of the flaviviridae family, has a surface composed of 180 copies each of the envelope (e) glycoprotein and the membrane (m) protein. the crystal structure of an n-terminal fragment of e has been determined and compared with a previously described structure. the primary difference between these structures is a 10 degrees rotation about a hinge relating the fusion domain dii to domains di and diii. these two rigid body components were used for independent fitting of e into t ...200415341726
susceptibility of mosquito and tick cell lines to infection with various flaviviruses.the genus flavivirus consists of more than 70 virus species and subtypes, the majority of which are transmitted by mosquitoes or ticks, although some have no known vector (nkv). the ability of these viruses to infect cultured cells derived from mosquito or tick species offers a useful insight into the suitability of such vectors to harbour and replicate particular viruses. we undertook a comparative study of the susceptibility of mammalian vero cells, a clonal mosquito cell line (c6/36) and rece ...200415347394
[molecular analysis of hepatitis c virus in pathogenesis of chronic hepatitis c]. 200415359835
can we really learn from model pathogens?cytolytic t (tc) lymphocytes are the first order response of the adaptive immune system in the recovery from primary viral infections. these effector cells execute their function either by direct cytotoxicity through the fas or perforin pathway and/or by the release of cytokines that either directly or indirectly exert antiviral activity. mice respond to infection by closely related viruses with a vigorous tc response, which is characterized by extensive crossreactivity on target cells infected ...200415364054
use of recombinant e protein domain iii-based enzyme-linked immunosorbent assays for differentiation of tick-borne encephalitis serocomplex flaviviruses from mosquito-borne flaviviruses.the serological diagnosis of infection by flaviviruses is complicated by the presence of flavivirus cross-reactive antibodies that produce false-positive results for flavivirus infections, especially in regions where more than one virus is endemic. current diagnostic reagents for tick-borne flavivirus infection have been found to cross-react with yellow fever- or dengue virus-positive sera. this study utilized recombinant flavivirus e protein domain 3 (re-d3) as a diagnostic reagent to different ...200415364996
n- t-boc-amino acid esters of isomannide. potential inhibitors of serine proteases.hepatitis c, dengue and west nile virus are some of the most important flaviviruses, that share one important serine protease enzyme. serine proteases are the most studied class of proteolytic enzyme and, in these cases, a primary target for drug discovery. in this paper, we describe the synthesis and preliminary molecular modeling studies of a novel class of n- t-boc amino acid esters derived of isomannide as potential serine proteases inhibitors.200415365909
infectious entry of west nile virus occurs through a clathrin-mediated endocytic pathway.the pathway of west nile flavivirus early internalization events was mapped in detail in this study. overexpression of dominant-negative mutants of eps15 strongly inhibits west nile virus (wnv) internalization, and pharmacological drugs that blocks clathrin also caused a marked reduction in virus entry but not caveola-dependent endocytosis inhibitory agent, filipin. using immunocryoelectron microscopy, wnv particles were seen within clathrin-coated pits after 2 min postinfection. double-labeling ...200415367621
different chemokine expression in lethal and non-lethal murine west nile virus infection.west nile (wn) virus is a mosquito-borne flavivirus that can cause lethal encephalitis in humans and horses. the wn virus endemic in new york city (ny) in 1999 caused large-scale mortality of wild birds that was not evident in endemic areas in other parts of the world, and the pathogenesis of the wn virus strain isolated in ny (ny strain) appears to differ from that of previously isolated strains. however, the pathogenesis of ny strain infection remains unclear. this study examined cc (rantes/cc ...200415368509
cytopathological changes induced by selected brazilian flaviviruses in mouse macrophages.scanning and transmission electron microscopy were used to analyse the ultrastructure of peritoneal mouse macrophage cells infected with brazilian flavivirus (yellow fever, rocio, bussuquara and saint louis encephalitis viruses). macrophage cells collected 3 days after viral infection had a flattened shape, with an increased number of large spikes of cytoplasm prolongations, giving an appearance of hairy cells. cytopathological changes to the macrophage cells were similar regardless of the infec ...200415369478
in vivo monitoring of acute flavivirus (modoc) encephalitis with regional and whole-brain quantitative diffusion magnetic resonance imaging.in vivo imaging of structural changes in the brain of patients with encephalitis has become an important aid in diagnostic and therapeutic procedures. diffusion-weighted magnetic resonance imaging (dwi) was employed to quantitate regional and whole-brain diffusion-weighted mri changes in a hamster model for acute flavivirus encephalitis. the regional apparent diffusion coefficient (adc) was determined in hyperintense regions seen on t2-weighted images (i.e., the thalamic area and the temporal lo ...200415371156
experimental usutu virus infection of suckling mice causes neuronal and glial cell apoptosis and demyelination.usutu virus (usuv), a mosquito-borne flavivirus of the japanese encephalitis virus group has been responsible for avian mortality in austria since 2001. in the present study, the neuropathogenicity and neuroinvasiveness of usuv for 1-week-old suckling mice was investigated. after intraperitoneal inoculation, clinical signs like depression, disorientation, paraplegia, paralysis and coma were observed between 6 and 11 days post infection. histologically, there was widespread neuronal apoptosis esp ...200415372281
specific interaction between the classical swine fever virus ns5b protein and the viral genome.the ns5b protein of the classical swine fever virus (csfv) is the rna-dependent rna polymerase of the virus and is able to catalyze the viral genome replication. the 3' untranslated region is most likely involved in regulation of the pestivirus genome replication. however, little is known about the interaction between the csfv ns5b protein and the viral genome. we used different rna templates derived from the plus-strand viral genome, or the minus-strand viral genome and the csfv ns5b protein ob ...200415373834
transmission cycles, host range, evolution and emergence of arboviral disease.many pandemics have been attributed to the ability of some rna viruses to change their host range to include humans. here, we review the mechanisms of disease emergence that are related to the host-range specificity of selected mosquito-borne alphaviruses and flaviviruses. we discuss viruses of medical importance, including venezuelan equine and japanese encephalitis viruses, dengue viruses and west nile viruses.200415378043
dengue-specific cd8+ t cells have both protective and pathogenic roles in dengue virus infection.to analyze roles of memory t cells in the pathogenesis of dengue (den) virus infection, a den virus-specific cd8+ cell clone (2d42 cell) was employed to investigate its in vivo function after den virus infection using an animal model. hepg2 grafted severe combined immunodeficient (hepg2-grafted scid) mice were divided into three groups--group a: hepg2-grafted scid mice were inoculated intraperitoneally (ip) with 2d42 cells and then ip-infected with den virus type 2 (den-2); group b: hepg2-grafte ...200415388257
single point mutation in tick-borne encephalitis virus prm protein induces a reduction of virus particle secretion.flaviviruses are assembled to bud into the lumen of the endoplasmic reticulum (er) and are secreted through the vesicle transport pathway. virus envelope proteins play important roles in this process. in this study, the effect of mutations in the envelope proteins of tick-borne encephalitis (tbe) virus on secretion of virus-like particles (vlps), using a recombinant plasmid expression system was analysed. it was found that a single point mutation at position 63 in prm induces a reduction in secr ...200415448368
structure of the dengue virus envelope protein after membrane fusion.dengue virus enters a host cell when the viral envelope glycoprotein, e, binds to a receptor and responds by conformational rearrangement to the reduced ph of an endosome. the conformational change induces fusion of viral and host-cell membranes. a three-dimensional structure of the soluble e ectodomain (se) in its trimeric, postfusion state reveals striking differences from the dimeric, prefusion form. the elongated trimer bears three 'fusion loops' at one end, to insert into the host-cell memb ...200414737159
high fidelity of yellow fever virus rna polymerase.three consecutive plaque purifications of four chimeric yellow fever virus-dengue virus (chimerivax-den) vaccine candidates against dengue virus types 1 to 4 were performed. the genome of each candidate was sequenced by the consensus approach after plaque purification and additional passages in cell culture. our data suggest that the nucleotide sequence error rate for sp6 rna polymerase used in the in vitro transcription step to initiate virus replication was as high as 1.34 x 10(-4) per copied ...200414694136
genetic vaccination of mice with plasmids encoding the ns1 non-structural protein from tick-borne encephalitis virus and dengue 2 virus.although there is a safe, inexpensive and efficacious vaccine against yellow fever, vaccination against other flavivirus diseases is less successful. there is no licensed vaccine against dengue fever and current vaccines against tick-borne encephalitis (tbe) and japanese encephalitis are expensive and require several injections. furthermore novel vaccines containing only virus envelope proteins may raise fears over antibody mediated enhancement (ade) of disease. here we report the successful use ...200414739654
tetracycline-inducible packaging cell line for production of flavivirus replicon particles.we have previously developed replicon vectors derived from the australian flavivirus kunjin that have a unique noncytopathic nature and have been shown to direct prolonged high-level expression of encoded heterologous genes in vitro and in vivo and to induce strong and long-lasting immune responses to encoded immunogens in mice. to facilitate further applications of these vectors in the form of virus-like particles (vlps), we have now generated a stable bhk packaging cell line, tetkuncprme, carr ...200414671135
nonsuppurative encephalitis in piglets after experimental inoculation of japanese encephalitis flavivirus isolated from pigs.nonsuppurative encephalitis was experimentally induced in 3-week-old piglets by a single intravenous inoculation of either of two strains (ib 2001 or as-6) of japanese encephalitis flavivirus (jev) isolated from field pigs. the lesions, which consisted of neuronal necrosis, neuronophagia, glial nodules, and perivas-cular cuffing, were distributed in the cerebrum, midbrain, pons, medulla oblongata, and cerebellum, particularly in the gray matter of the frontal and temporal lobes and thalamus. the ...200414715969
flavivirus encephalitis. 200415496637
analysis of adaptive mutations in kunjin virus replicon rna reveals a novel role for the flavivirus nonstructural protein ns2a in inhibition of beta interferon promoter-driven transcription.the establishment of persistent noncytopathic replication by replicon rnas of a number of positive-strand rna viruses usually leads to generation of adaptive mutations in nonstructural genes. some of these adaptive mutations (e.g., in hepatitis c virus) increase the ability of rna replication to resist the antiviral action of alpha/beta interferon (ifn-alpha/beta); others (e.g., in sindbis virus) may also lead to more efficient ifn production. using puromycin-selectable kunjin virus (kun) replic ...200415507609
chimerivax-west nile virus live-attenuated vaccine: preclinical evaluation of safety, immunogenicity, and efficacy.the availability of chimerivax vaccine technology for delivery of flavivirus protective antigens at the time west nile (wn) virus was first detected in north america in 1999 contributed to the rapid development of the vaccine candidate against wn virus described here. chimerivax-japanese encephalitis (je), the first live- attenuated vaccine developed with this technology has successfully undergone phase i and ii clinical trials. the chimerivax technology utilizes yellow fever virus (yf) 17d vacc ...200415507637
the transmembrane domains of the prm and e proteins of yellow fever virus are endoplasmic reticulum localization signals.the immature flavivirus particle contains two envelope proteins, prm and e, that are associated as a heterodimer. virion morphogenesis of the flaviviruses occurs in association with endoplasmic reticulum (er) membranes, suggesting that there should be accumulation of the virion components in this compartment. this also implies that er localization signals must be present in the flavivirus envelope proteins. in this work, we looked for potential subcellular localization signals in the yellow feve ...200415507646
the role of tumor necrosis factor in modulating responses of murine embryo fibroblasts by flavivirus, west nile.murine embryo fibroblasts (mef) transcribe tumor necrosis factor (tnf) mrna and secrete soluble tnf in response to infection by west nile virus (wnv) and tnf was demonstrated to be protective against wnv infection in vitro. tnf is not required for the wnv-induced upregulation of mhc-i expression on mef, as tnf deficiency did not affect the upregulation of major histocompatibility complex class i (mhc-i) by wnv. furthermore, nf-kappab was activated by wnv in tnf-deficient mef, demonstrating that ...200415518815
development and evaluation of an enzyme-linked immunosorbent assay for quantifying antibodies to japanese encephalitis virus nonstructural 1 protein to detect subclinical infections in vaccinated horses.antibodies to japanese encephalitis virus (jev) nonstructural 1 (ns1) protein constitute a marker of natural jev infection among populations vaccinated with inactivated je vaccine. in japan, with few recent human je cases, the natural infection rate is critical to evaluate the necessity of continuing je vaccination. a sensitive immunochemical staining method for detecting ns1 antibodies in individuals naturally and subclinically infected with jev was previously established. here, an enzyme-linke ...200415528700
structure and dynamics of a predicted ferredoxin-like selenoprotein in japanese encephalitis virus.homologues of the selenoprotein glutathione peroxidase (gpx) have been previously identified in poxviruses and in rna viruses including hiv-1 and hepatitis c virus (hcv). sequence analysis of the ns4 region of japanese encephalitis virus (jev) suggests it may encode a structurally related but functionally distinct selenoprotein gene, more closely related to the iron-binding protein ferredoxin than to gpx, with three highly conserved uga codons that align with essential cys residues of ferredoxin ...200415530818
experimental transmission of karshi and langat (tick-borne encephalitis virus complex) viruses by ornithodoros ticks (acari: argasidae).selected species of mosquitoes and ornithodoros ticks were evaluated for their potential to transmit karshi and langat (tick-borne encephalitis virus complex) viruses in the laboratory. although there was no evidence of replication of karshi virus in either of the two mosquito species tested [ochlerotatus taeniorhynchus (wiedemann) or culex pipiens (l.)], karshi virus replicated in and was transmitted by all three species of ornithodoros ticks tested (ornithodoros parkeri cooley, ornithodoros so ...200415535630
evaluation of a diagnostic algorithm using immunoglobulin m enzyme-linked immunosorbent assay to differentiate human west nile virus and st. louis encephalitis virus infections during the 2002 west nile virus epidemic in the united states.a diagnostic algorithm was developed to differentiate between human infections of west nile virus (wnv) and st. louis encephalitis virus (slev) using positive-to-negative (p/n) ratios derived from the immunoglobulin m capture enzyme-linked immunosorbent assay (mac-elisa). to validate this algorithm, we tested 1,418 serum and cerebrospinal fluid (csf) samples from confirmed wnv and slev infections collected during the wnv epidemic of 2002 in the united states. wnv p/n-to-slev p/n ratios (w/s rati ...200415539517
chimpanzee fab fragments and a derived humanized immunoglobulin g1 antibody that efficiently cross-neutralize dengue type 1 and type 2 viruses.passive immunization with monoclonal antibodies from humans or nonhuman primates represents an attractive alternative to vaccines for prevention of illness caused by dengue viruses (denv) and other flaviviruses, including the west nile virus. in a previous study, repertoire cloning to recover fab fragments from bone marrow mrna of chimpanzees infected with all four denv serotypes (dengue virus serotype 1 [denv-1] to denv-4) was described. in that study, a humanized immunoglobulin g1 (igg1) antib ...200415542643
epitope determinants of a chimpanzee fab antibody that efficiently cross-neutralizes dengue type 1 and type 2 viruses map to inside and in close proximity to fusion loop of the dengue type 2 virus envelope glycoprotein.the epitope determinants of chimpanzee fab antibody 1a5, which have been shown to be broadly reactive to flaviviruses and efficient for cross-neutralization of dengue virus type 1 and type 2 (denv-1 and denv-2), were studied by analysis of denv-2 antigenic variants. sequence analysis showed that one antigenic variant contained a gly-to-val substitution at position 106 within the flavivirus-conserved fusion peptide loop of the envelope protein (e), and another variant contained a his-to-gln subst ...200415542644
histidine 39 in the dengue virus type 2 m protein has an important role in virus assembly.the mature flavivirus particle comprises a nucleocapsid core surrounded by a lipid bilayer containing the membrane (m) (derived from the precursor prm) and envelope (e) proteins. the formation of intracellular prm/e heterodimers occurs rapidly after translation and is believed to be important for the assembly and secretion of immature virus particles. in this study, the role of the his residue at position 39 in the m protein (m39) of dengue virus type 2 (denv-2) in the virus life cycle was inves ...200415557235
viral envelope protein glycosylation is a molecular determinant of the neuroinvasiveness of the new york strain of west nile virus.two new york (ny) strains of the west nile (wn) virus were plaque-purified and four variants that had different amino acid sequences at the n-linked glycosylation site in the envelope (e) protein sequence were isolated. the e protein was glycosylated in only two of these strain variants. to determine the relationship between e protein glycosylation and pathogenicity of the wn virus, 6-week-old mice were infected subcutaneously with these variants. mice infected with viruses that carried the glyc ...200415557236
toll-like receptor 3 mediates west nile virus entry into the brain causing lethal encephalitis.west nile virus (wnv), a mosquito-borne single-stranded (ss)rna flavivirus, causes human disease of variable severity. we investigated the involvement of toll-like receptor (tlr) 3, which recognizes viral double-stranded (ds)rna, on wnv infection. tlr3-deficient (tlr3(-/-)) mice were more resistant to lethal wnv infection and had impaired cytokine production and enhanced viral load in the periphery, whereas in the brain, viral load, inflammatory responses and neuropathology were reduced compared ...200415558055
rapid detection of west nile virus in birds using the vectest wnv antigen assay.to determine if the vectest west nile virus antigen assay (for testing mosquitoes) could be adapted to detect west nile virus (wnv) rapidly and accurately in birds for screening purposes.200415559727
a conserved histidine in the ij loop of the semliki forest virus e1 protein plays an important role in membrane fusion.the enveloped alphavirus semliki forest virus (sfv) infects cells via a low ph-triggered membrane fusion reaction mediated by the e1 protein. e1 is a class ii fusion protein that contains the hydrophobic fusion peptide loop and converts to a stable homotrimer during the fusion reaction. intriguingly, the fusion loop is closely associated with a loop connecting the i and j beta-strands. this ij loop plays a role in the cholesterol dependence of membrane fusion and is specifically susceptible to p ...200415564465
localization and characterization of flavivirus envelope glycoprotein cross-reactive epitopes.the flavivirus e glycoprotein, the primary antigen that induces protective immunity, is essential for membrane fusion and mediates binding to cellular receptors. human flavivirus infections stimulate virus species-specific as well as flavivirus cross-reactive immune responses. flavivirus cross-reactive antibodies in human sera create a serious problem for serodiagnosis, especially for secondary flavivirus infections, due to the difficulty of differentiating primary from secondary cross-reactive ...200415564505
complete genome analysis and molecular characterization of usutu virus that emerged in austria in 2001: comparison with the south african strain saar-1776 and other flaviviruses.here we describe the complete genome sequences of two strains of usutu virus (usuv), a mosquito-borne member of the genus flavivirus in the japanese encephalitis virus (jev) serogroup. usuv was detected in austria in 2001 causing a high mortality rate in blackbirds; the reference strain (saar-1776) was isolated in 1958 from mosquitoes in south africa and has never been associated with avian mortality. the austrian and south african isolates exhibited 97% nucleotide and 99% amino acid identity. p ...200415464850
noninfectious recombinant antigen for detection of st. louis encephalitis virus-specific antibodies in serum by enzyme-linked immunosorbent assay.proper surveillance of virus activity and a timely response to viral outbreaks depend upon the rapid diagnosis of viral infections. the immunoglobulin m (igm) antibody-capture enzyme-linked immunosorbent assay (mac-elisa) is a fast, sensitive test routinely used for the diagnosis of the medically important west nile and st. louis encephalitis flaviviruses. however, the suckling mouse brain-derived (smb) antigen used in this assay is tedious to prepare and has a risk of exposing personnel to live ...200415472331
interaction of west nile virus with alpha v beta 3 integrin mediates virus entry into cells.the functional receptor for the flavivirus west nile (wnv) infection has been characterized in this study with a combination of biochemical and molecular approaches. a 105-kda protease-sensitive glycoprotein that binds wnv was isolated from the plasma membrane of cells permissive to wnv infection. the protein was subjected to peptide sequencing, and this glycoprotein was identified as a member of the integrin superfamily. infection of wnv was shown to be markedly inhibited in vero cells pretreat ...200415475343
the ns3 protein of hepatitis c virus induces caspase-8-mediated apoptosis independent of its protease or helicase activities.apoptosis has been implicated in the pathogenesis of hepatitis c virus (hcv)-related disease. here, we show that expression of hcv ns3, or the ns2/ns3 precursor protein, in mammalian cells results in induction of apoptosis and activation of caspases. hcv ns3-induced apoptosis was blocked by a caspase-8, but not a caspase-9-specific inhibitor. hcv ns3 coimmunoprecipitated with caspase-8, but not with other caspases or with fadd. coexpression of hcv ns3 and caspase-8 resulted in aggregation of the ...200415476874
enhancement of dengue virus translation: role of the 3' untranslated region and the terminal 3' stem-loop domain.an essential step for a productive infection by the dengue flavivirus (den) is translation of the m(7)g-capped, nonpolyadenylated positive-sense rna genome. we have recently identified sequences within the den 3' untranslated region (utr) that modulate viral translation. here, we show that the den type 2 (den2) 3'utr stimulated translation of m(7)g-capped den2 5'utr-containing reporter mrnas in baby hamster kidney (bhk) cells compared to a 3' vector sequence. analogous to the 3' poly(a) tail, th ...200415476880
proinflammatory cytokines and chemokines in humans with japanese encephalitis.japanese encephalitis virus (jev), the mosquito-borne flavivirus, annually causes an estimated 35,000-50,000 encephalitis cases and 10,000-15,000 deaths in asia, and there is no antiviral treatment. the role played by the immune response in determining the outcome of human infection with jev is poorly understood, although, in animal models of flavivirus encephalitis, unregulated proinflammatory cytokine responses can be detrimental.200415478067
characterization of tick-borne encephalitis virus from estonia.tick-borne encephalitis virus (tbev) is a severe problem in estonia. in the present article the first genetic analysis of estonian tbev strains is described. in total, seven tbev strains were isolated from ticks (ixodes ricinus and i. persulcaus), rodents (apodemus agrarius and cletrionomys glareolus), and serum from a tick-borne encephalitis (tbe) patient. the nucleic acid sequences of the viral genome encoding almost the complete e protein (nt 41-1250) and the 3'-ncr-termini of the estonian tb ...200415484275
[immunity to flavivirus in amerindian population of the sierra de perijá, zulia state, venezuela].little information is available about flavivirus infection in amerindian populations in western venezuela. on this account the activity and seroprevalence of these viruses were determined and the hypothesis concerning the existence of a sylvatic cycle, conditioning the infection transmission of these viruses in indigenous populations, was studied. for this, blood samples from yukpas (n=144) and barí (n=110) communities were collected, 35 (yukpas=25 and barí=10) of which were processed for viral ...200415602900
bovine viral diarrhoea virus is internalized by clathrin-dependent receptor-mediated endocytosis.bovine viral diarrhoea virus (bvdv) is a pestivirus within the family flaviviridae. in contrast to the members of the genus flavivirus, nothing is known about the viral entry route for pestiviruses. in this study, the process of bvdv infection following attachment to the cell surface was examined. bvdv clearly co-localizes with clathrin, with early endosome antigen-1 (eea-1), an early endosome marker, and also with lysosomal-associated membrane protein-2 (lamp-2), a lysosomal marker. bvdv intern ...200415606865
[west nile virus, an emerging arbovirus].the west nile virus (family flaviviridae, genus flavivirus), first isolated in 1937 in africa, is an arthropod-borne virus transmitted through culex mosquitoes bite. its natural cycle includes a vector (culex mosquitoes), a reservoir (birds of various species) and humans and large mammals are epidemiological dead-ends. progressive dissemination: this virus recently drew attention after several epidemics and epizooties were identified in france, russia, romania, and israel. however, the most stri ...200415614177
isolation of bovine viral diarrhea virus 1, a pestivirus from autopsied lamb specimen from tamil nadu, india.an epizootic of febrile illness among the madras red breed of sheep had occurred in 1994 in verrapuram, chennai, india. the epizootic was suspected as rift valley fever (rvf)-like sickness based on clinical features. however, its etiological agent could neither be isolated nor implicated conclusively. during the post-epizootic period a male lamb died of similar clinical features and the spleen was immediately collected. inoculation of spleen suspension in infant mouse brain yielded a virus that ...200415745045
infection of human dendritic cells by dengue virus activates and primes t cells towards th0-like phenotype producing both th1 and th2 cytokines.dengue viruses (dv) infection is an important public health issue all over the world. although the pathogenesis remains unclear, the overwhelmingly triggered immune responses have been consistently observed. recently, we and other researchers demonstrated that the natural hosts for dv are dendritic cells (dc), the primary sentinels of immune system. in light of the significance of t cells in dengue virus pathogenesis, here, we examine the possible consequences of dc-t cell interaction that is su ...200415624700
[tick borne flavivirus encephalitis]. 200415624474
tick-borne virus diseases of human interest in europe.several human diseases in europe are caused by viruses transmitted by tick bite. these viruses belong to the genus flavivirus, and include tick-borne encephalitis virus, omsk haemorrhagic fever virus, louping ill virus, powassan virus, nairovirus (crimean-congo haemorrhagic fever virus) and coltivirus (eyach virus). all of these viruses cause more or less severe neurological diseases, and some are also responsible for haemorrhagic fever. the epidemiology, clinical picture and methods for diagnos ...200415606630
live measles vaccine expressing the secreted form of the west nile virus envelope glycoprotein protects against west nile virus encephalitis.the schwarz strain of measles virus (mv), a live attenuated rna virus, is one of the safest and most effective human vaccines available. immunization with mv vaccine expressing heterologous antigen is an attractive strategy to prevent emerging viral diseases. west nile virus (wnv), which recently emerged in north america, is an important mosquito-borne flavivirus that causes numerous cases of human encephalitis, thus urging the development of a vaccine. to evaluate the efficacy of recombinant mv ...200415609230
[comparative analysis of virulence of the siberian and far-east subtypes of the tick-born encephalitis virus].the siberian subtype of the tick-borne encephalitis virus (tev) is different from the far-east subtype by a moderate virulence observed in siberian hamsters and by a low infection development rate (100 strains were compared). no differences were found in neuro-invasiveness. clinical findings and experiments with monkeys denote the ability of the siberian subtype to provoke severe forms of tick-borne encephalitis (tbe). the inflammation-and-degenerative changes were localized in the brain cortex, ...200415597957
identification of residues in the dengue virus type 2 ns2b cofactor that are critical for ns3 protease activation.proteolytic processing of the dengue virus polyprotein is mediated by host cell proteases and the virus-encoded ns2b-ns3 two-component protease. the ns3 protease represents an attractive target for the development of antiviral inhibitors. the three-dimensional structure of the ns3 protease domain has been determined, but the structural determinants necessary for activation of the enzyme by the ns2b cofactor have been characterized only to a limited extent. to test a possible functional role of t ...200415564480
performance of immunoglobulin g (igg) and igm enzyme-linked immunosorbent assays using a west nile virus recombinant antigen (prem/e) for detection of west nile virus- and other flavivirus-specific antibodies.focus technologies developed an indirect immunoglobulin g (igg) enzyme-linked immunosorbent assay (elisa) and a mu-capture igm elisa for the detection of west nile virus (wnv)-specific antibodies based on a wnv prem/e protein recombinant antigen. normal and disease state serum panels were used to assess the performance characteristics of the two wnv elisa kits. totals of 807 and 1,423 sera were used to assess the igg elisa and igm elisa kits, respectively. the focus technologies igg elisa had a ...200415472323
flavivirus isolations from mosquitoes collected from saibai island in the torres strait, australia, during an incursion of japanese encephalitis virus.adult mosquitoes (diptera: culicidae) were collected in january and february 2000 from saibai island in the torres strait of northern australia, and processed for arbovirus isolation during a period of japanese encephalitis (je) virus activity on nearby badu island. a total of 84 210 mosquitoes were processed for virus isolation, yielding six flavivirus isolates. viruses obtained were single isolates of je and kokobera (kok) and four of kunjin (kun). all virus isolates were from members of the c ...200415347396
emerging flaviviruses: the spread and resurgence of japanese encephalitis, west nile and dengue viruses.mosquito-borne flaviviruses provide some of the most important examples of emerging and resurging diseases of global significance. here, we describe three of them: the resurgence of dengue in tropical and subtropical areas of the world, and the spread and establishment of japanese encephalitis and west nile viruses in new habitats and environments. these three examples also illustrate the complexity of the various factors that contribute to their emergence, resurgence and spread. whereas some of ...200415577938
[the prevalence of hepatitis g virus in patients with hepatitis b and c virus infections].gb virus c/hepatitis g virus (gbv-c/hgv) is a recently identified flavivirus, which has been frequently detected in patients with chronic hepatitis b virus (hbv) and hepatitis c virus (hcv) infections. in this study, the prevalence of hgv-rna and antibodies to e2 envelope antigen (anti-e2) which is a marker of past infection, have been investigated in the samples of patients with hcv and hbv infections, and the prevalence rates were compared with the control group. the study group consisted of 5 ...200415700668
[residual neurovirulence of the chimera of langat and denge-4 flaviviruses in intracerebral infection of monkeys].the residual neurovirulence of the previously obtained chimeric virus tp21/den4, containing the rna region encoding the pre m and e structural enzymes, strain tp21, virus langat, and the remaining part of the genome from the denge 4 virus, was studied in experiments with monkeys cercopithecus aethiops involving the intracerebral administration of the virus. the tick-borne encephalitis virus, strain absettarov, was used as positive control. a comparative analysis of the experimental and published ...200515747868
an infectious west nile virus that expresses a gfp reporter gene.west nile virus is a mosquito-borne, neurotropic flavivirus that causes encephalitis in humans and animals. since being introduced into the western hemisphere in 1999, wnv has spread rapidly across north america, identifying this virus as an important emerging pathogen. in this study, we developed a dna-launched infectious molecular clone of wnv that encodes a gfp reporter gene. transfection of cells with the plasmid encoding this recombinant virus (pwnii-gfp) resulted in the production of infec ...200515749120
novel flavivirus or new lineage of west nile virus, central europe.a flavivirus (strain 97-103) was isolated from culex pipens mosquitoes in 1997 following floods in south moravia, czech republic. the strain exhibited close antigenic relationship to west nile virus (wnv) prototype strain eg-101 in a cross-neutralization test. in this study, mouse pathogenicity characteristics and the complete nucleotide and putative amino acid sequences of isolate 97-103, named rabensburg virus (rabv) after a nearby austrian city, were determined. rabv shares only 75%-77% nucle ...200515752439
japanese encephalitis in india. 200515970050
development of immunoglobulin m capture enzyme-linked immunosorbent assay to differentiate human flavivirus infections occurring in australia.we report the development of a flavivirus immunoglobulin m (igm) capture enzyme-linked immunosorbent assay (mac-elisa) which improves the determination of an infecting flavivirus serotype over that by current serological methods. a panel of 165 igm-positive sera from flavivirus patients with specific diagnostic results was tested by the flavivirus mac-elisa using a panel of 10 antigens. for 134 of these sera (81.2%), the highest reactivity was demonstrated against the infecting virus, which was ...200515753249
peptide inhibitors of dengue virus and west nile virus infectivity.viral fusion proteins mediate cell entry by undergoing a series of conformational changes that result in virion-target cell membrane fusion. class i viral fusion proteins, such as those encoded by influenza virus and human immunodeficiency virus (hiv), contain two prominent alpha helices. peptides that mimic portions of these alpha helices inhibit structural rearrangements of the fusion proteins and prevent viral infection. the envelope glycoprotein (e) of flaviviruses, such as west nile virus ( ...200515927084
comparative analysis of immunoglobulin m (igm) capture enzyme-linked immunosorbent assay using virus-like particles or virus-infected mouse brain antigens to detect igm antibody in sera from patients with evident flaviviral infections.the use of immunoglobulin m (igm) antibody-capture enzyme-linked immunosorbent assay (mac-elisa) serves as a valuable tool for the diagnosis of acute flaviviral infections, since igm antibody titers are detectable early, peak at about 2 weeks postinfection, and subsequently decline to lower levels over the next few months. traditionally, virus-infected tissue culture or suckling mouse brain (smb) has been the source of viral antigens used in the assay. in an effort to provide a reliable source o ...200516000440
[the occurrence rate of hgv/gbv-c rna and risk factors in patients of narcological dispensary in novosibirsk].the occurrence rate of hgv/gbv-c rna, genotypic variety of isolates and various risk factors of infection with hgv/gbv-c were evaluated in 500 patients of the narcological dispensary of novosibirsk. the occurrence rate of hgv/gbv-c rna among all examined blood sera was 33.6%. at the same time in blood sera with hcv markers the occurrence rate of hgv/ gbv-c was 42.9% and in sera with negative results for markers hcv--25%. for gene typing of obtained isolates the direct sequencing of the amplifica ...200515881934
characterization of west nile viral replication and maturation in peripheral neurons in culture.the north american west nile virus (wnv), new york 1999 strain, appears to be highly neurotropic, and its neuroinvasiveness is an important aspect of human disease. the authors have developed an in vitro model to study wnv replication and protein processing in neurons. they compared wnv infection of the dorsal root ganglion (drg) neurons (sensory neurons) and pc-12 cells (sympathetic neurons) to wnv infection of the mosquito cell line, c6/36, and vero cells. wnv infection of both neuronal cell t ...200515804955
the identification of immunodominant linear epitopes of dengue type 2 virus capsid and ns4a proteins using pin-bound peptides.we have used multi-pin peptide synthesis strategy to identify b-cell epitopes on two small dengue virus proteins, capsid and ns4a. we have identified several linear, immunodominant epitopes on both these proteins. almost all these epitopes mapped to regions predicted to be hydrophilic based on kyte and doolittle profiles. of the capsid epitopes identified in this study, the most immunogenic ones mapped to the c-terminal alpha4 helix, which lies on the solvent-exposed surface of the capsid dimer. ...200516022901
recombination and flavivirus vaccines: a commentary. 200515811640
alkhumra haemorrhagic fever: case report and infection control details. 200515816213
competitive inhibition of the dengue virus ns3 serine protease by synthetic peptides representing polyprotein cleavage sites.the ns3 serine protease of dengue virus is required for the maturation of the viral polyprotein and consequently represents a promising target for the development of antiviral inhibitors. however, the substrate specificity of this enzyme has been characterized only to a limited extent. in this study, we have investigated product inhibition of the ns3 protease by synthetic peptides derived from the p6-p1 and the p1'-p5' regions of the natural polyprotein substrate. n-terminal cleavage site peptid ...200515823576
first record of aedes (aedimorphus) vexans vexans (meigen) in australia.aedes (aedimorphus) vexans vexans has become widely distributed in the australasian and oceanic islands zoogeographic regions, and in this paper, we describe the 1st confirmed report of ae. vexans vexans in australia. a total of 45 adult individuals were collected around the town of kununurra in the northeast kimberley region of western australia during the late wet seasons between 1996 and 2003. the majority (84%) was collected at trap sites in or near the ord stage i irrigation area. aedes vex ...200516033126
the suitability of yellow fever and japanese encephalitis vaccines for immunization against west nile virus.seven volunteers involved in flavivirus studies have been immunized with commercial japanese encephalitis and yellow fever vaccines je-vax and yf-vax. strong homologous and cross-reactive with west nile virus (wnv) antibody responses with titers 1:1600 to 1:51200 were found in all donors. all donors developed high levels of yellow fever virus (yfv) and japanese encephalitis virus (jev) neutralizing antibodies with titers 1:50 to 1:1600 and 1:20 to 1:640, respectively, and wnv neutralizing antibo ...200515939510
alkhumra virus infection, a new viral hemorrhagic fever in saudi arabia.four patients with typical acute viral hemorrhagic fever were identified in the holy city of makkah, saudi arabia, between 8 and 23 february 2001, the hajj (pilgrimage) period of that year. tests for rift valley fever (rvf), crimean-congo hemorrhagic fever (cchf), and dengue were negative. blood specimens were sent to the centres for disease control and prevention (cdc), atlanta for viral culture and testing for other hemorrhagic fever viruses. a new flavivirus closely related to the tick-borne ...200516038757
stereophysicochemical variability plots highlight conserved antigenic areas in flaviviruses.flaviviruses, which include dengue (dv) and west nile (wn), mutate in response to immune system pressure. identifying escape mutants, variant progeny that replicate in the presence of neutralizing antibodies, is a common way to identify functionally important residues of viral proteins. however, the mutations typically occur at variable positions on the viral surface that are not essential for viral replication. methods are needed to determine the true targets of the neutralizing antibodies.200515845145
dendritic cell-specific intercellular adhesion molecule 3-grabbing non-integrin (dc-sign)-mediated enhancement of dengue virus infection is independent of dc-sign internalization signals.dengue virus (dv) is a mosquito-borne flavivirus that causes hemorrhagic fever in humans. in the natural infection, dv is introduced into human skin by an infected mosquito vector where it is believed to target immature dendritic cells (dcs) and langerhans cells (lcs). we found that dv productively infects dcs but not lcs. we show here that the interactions between dv e protein, the sole mannosylated glycoprotein present on dv particles, and the c-type lectin dendritic cell-specific intercellula ...200515855154
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