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t cell responses to an hla-b*07-restricted epitope on the dengue ns3 protein correlate with disease severity.dengue hemorrhagic fever (dhf), the severe manifestation of dengue virus (dv) infection characterized by plasma leakage, is more common in secondary dv infections in previously infected individuals and is associated with high levels of immune activation. to determine the ag specificity of this immune response, we studied the response to an hla-b*07-restricted t cell epitope, residues 221-232 of the dv ns3 protein, in 10 hla-b*07(+) thai children who were studied during and after acute dv infecti ...200212023403
complete genome sequence of montana myotis leukoencephalitis virus, phylogenetic analysis and comparative study of the 3' untranslated region of flaviviruses with no known vector.montana myotis leukoencephalitis virus (mmlv), a virus isolated from bats, causes an encephalitis in small rodents reminiscent of flavivirus encephalitis in humans. the complete mmlv genome is 10690 nucleotides long and encodes a putative polyprotein of 3374 amino acids. the virus contains the same conserved motifs in genes that are believed to be interesting antiviral targets (ntpase/helicase, serine protease and rna-dependent rna polymerase) as flaviviruses of clinical importance. phylogenetic ...200212124451
infection of scid mice with montana myotis leukoencephalitis virus as a model for flavivirus encephalitis.we have established a convenient animal model for flavivirus encephalitis using montana myotis leukoencephalitis virus (mmlv), a bat flavivirus. this virus has the same genomic organization, and contains the same conserved motifs in genes that encode potential antiviral targets, as flaviviruses that cause disease in man (n. charlier et al., accompanying paper), and has a similar particle size (approximately 40 nm). mmlv replicates well in vero cells and appears to be equally as sensitive as yell ...200212124452
an rna cap (nucleoside-2'-o-)-methyltransferase in the flavivirus rna polymerase ns5: crystal structure and functional characterization.viruses represent an attractive system with which to study the molecular basis of mrna capping and its relation to the rna transcription machinery. the rna-dependent rna polymerase ns5 of flaviviruses presents a characteristic motif of s-adenosyl-l-methionine-dependent methyltransferases at its n-terminus, and polymerase motifs at its c-terminus. the crystal structure of an n-terminal fragment of dengue virus type 2 ns5 is reported at 2.4 a resolution. we show that this ns5 domain includes a typ ...200212032088
etiology of encephalitis syndrome among hospitalized children and adults in takeo, cambodia, 1999-2000.whether or not japanese encephalitis virus (jev) is an important causative agent of acute encephalitis in cambodia remains unclear. this study was carried out to determine the cause of encephalitis syndrome among children and adults admitted to takeo provincial hospital from october 1999 to september 2000. ninety-nine cases were included in the study: 52 pediatric cases (12 were fatal) and 47 adult cases (10 were fatal). a causative agent such as human herpesvirus (hhv-3 or hhv-4), cryptococcus ...200212135294
catalytically active dengue virus ns3 protease forms aggregates that are separable by size exclusion chromatography.an active form of the dengue virus protease ns3 (cf40.gly.ns3pro) was expressed in escherichia coli. this construct consists of a critical 40 amino acid cofactor domain from ns2b fused to the n-terminal 184 amino acid protease domain of ns3 via a flexible, covalent linker (gly(4)sergly(4)). the recombinantly produced protein is soluble and has a hexa-histidine tag engineered at the n-terminus for ease of purification using metal affinity chromatography. however, the presence of lower molecular w ...200212135556
ns3 protease of langat tick-borne flavivirus cleaves serine protease substrates.langat (lgt) virus, initially isolated in 1956 from ticks in malaysia, is a naturally occurring nonpathogenic virus with a very close antigenicity to the highly pathogenic tick-borne encephalitis (tbe) western subtype virus and tbe far eastern subtype virus. ns3, the second largest viral protein of lgt virus, is highly conserved among flaviviruses and contains a characteristic protease moiety (ns3 pro). ns3 pro represents an attractive target for anti-protease molecules against tbe virus. we rep ...200212054734
st. louis encephalitis virus induced pathology in cultured cells.apoptosis is a highly regulated process of cellular self-destruction with diverse functions in multicellular organisms. it is known to be one of the mechanisms of viral pathogenesis. st. louis encephalitis virus (slev), an arthropod-borne flavivirus, causes encephalitis disease of varying severity mostly in north america and in some regions of south america. this virus induces cytopathic effects in vertebrate cell lines, however, the mechanism by which this occurs is yet to be elucidated. slev i ...200212111422
seasonal abundance of culex nigripalpus theobald and culex salinarius coquillett in north florida, usa.north florida is a transition zone between widespread culex nigripalpus populations to the south and focal culex salinarius populations to the north. culex nigripalpus is a vector of st. louis encephalitis (sle) and eastern equine encephalitis (eee) viruses in south florida, while cx. salinarius is a suspected new world vector of west nile (wn) virus. abundant vector populations are often a prerequisite for epidemic and epizootic transmission of arboviruses. extensive sle transmission has never ...200212125867
tick-borne encephalitis (tbe) in dogs.tick-borne encephalitis (tbe) is caused by a flavivirus and transmitted by ticks. it is known in dogs for nearly 30 years and the number of tbe cases is increasing. in addition to fever, cerebrocortical, thalamic, and brainstem symptoms occur simultaneously. not all tbe infections in dogs lead to clinical signs but peracute/lethal as well as subacute and chronic courses have been reported. tbe is a seasonal disease, depending on climate related tick activity. infected ticks are spreading the vir ...200212141763
modes of transmission of hemorrhagic fever. 200212150658
a nonsense mutation in the gene encoding 2'-5'-oligoadenylate synthetase/l1 isoform is associated with west nile virus susceptibility in laboratory mice.a mouse model has been established to investigate the genetic determinism of host susceptibility to west nile (wn) virus, a member of the genus flavivirus and family flaviviridae. whereas wn virus causes encephalitis and death in most laboratory inbred mouse strains after peripheral inoculation, most strains derived from recently trapped wild mice are completely resistant. the phenotype of resistance/susceptibility is determined by a major locus, wnv, mapping to chromosome 5 within the 0.4-cm-wi ...200212186974
safety and immunogenicity of tetravalent live-attenuated dengue vaccines in thai adult volunteers: role of serotype concentration, ratio, and multiple doses.dengue fever, caused by four serotypes of a mosquito-borne virus, is a growing problem in tropical countries. currently, there is no treatment or vaccine. we evaluated safety and immunogenicity of two doses, given six months apart, of seven formulations of dengue tetravalent live-attenuated vaccine (containing different concentrations of the component viruses) versus placebo in 59 flavivirus-seronegative thai adults. the first dose was the more reactogenic. most volunteers experienced clinically ...200212139219
classical swine fever virus ns5b-gfp fusion protein possesses an rna-dependent rna polymerase activity.rna-dependent rna polymerase (rdrp) is the replicase of positive-strand rna viruses. expression and characterization of the replicase are the first steps in the elucidation of the virus replication mechanism. we expressed nonstructural protein 5b (ns5b) of classical swine fever virus (csfv) as a fusion protein with green fluorescent protein (gfp) in porcine kidney cells (pk-15 cells), natural host cells of csfv. the expressed csfv ns5b-gfp fusion protein possessed rdrp activity. by fluorescence ...200212209316
expression, purification, and characterization of the rna 5'-triphosphatase activity of dengue virus type 2 nonstructural protein 3.dengue virus type 2 (den2), a member of the flaviviridae family of positive-strand rna viruses, contains a single rna genome having a type i cap structure at the 5' end. the viral rna is translated to produce a single polyprotein precursor that is processed to yield three virion proteins and at least seven nonstructural proteins (ns) in the infected host. ns3 is a multifunctional protein having a serine protease catalytic triad within the n-terminal 180 amino acid residues which requires ns2b as ...200212167347
flavivirus susceptibility in aedes aegypti.aedes aegypti is the primary vector of yellow fever (yf) and dengue fever (df) flaviviruses worldwide. in this review we focus on past and present research on genetic components and environmental factors in aedes aegypti that appear to control flavivirus transmission. we review genetic relationships among ae. aegypti populations throughout the world and discuss how variation in vector competence is correlated with overall genetic differences among populations. we describe current research into h ...200212234528
infection of polarized epithelial cells with flavivirus west nile: polarized entry and egress of virus occur through the apical surface.both polarized epithelial vero (c1008) and non-polarized vero (control) cells were grown on permeable cell culture inserts and infected either apically or basolaterally with west nile (wn) or kunjin (kun) virus. kun virus (closely related to wn virus) was used as a comparison. using indirect immunofluorescence and plaque assays of productive virus titres, entry of wn and kun viruses was confined to the apical surface of polarized epithelial cells. for the first time, these results provided evide ...200212237424
genome sequence analysis of tamana bat virus and its relationship with the genus flavivirus.tamana bat virus (tabv, isolated from the bat pteronotus parnellii) is currently classified as a tentative species in the genus flavivirus: we report here the determination and analysis of its complete coding sequence. low but significant similarity scores between tabv and member-viruses of the genus flavivirus were identified in the amino acid sequences of the structural, ns3 and ns5 genes. a series of cysteines located in the envelope protein and the most important enzymatic domains of the vir ...200212237426
the molecular biology of west nile virus: a new invader of the western hemisphere.west nile virus (wnv) is a mosquito-borne flavivirus that primarily infects birds but occasionally also infects humans and horses. in recent years, the frequency of wnv outbreaks in humans has increased, and these outbreaks have been associated with a higher incidence of severe disease. in 1999, the geographical distribution of wnv expanded to the western hemisphere. wnv has a positive strand rna genome of about 11 kb that encodes a single polyprotein. wnv replicates in the cytoplasm of infected ...200212142476
west nile virus.west nile (wn) virus is a mosquito-borne flavivirus and human, equine, and avian neuropathogen. the virus is indigenous to africa, asia, europe, and australia, and has recently caused large epidemics in romania, russia, and israel. birds are the natural reservoir (amplifying) hosts, and wn virus is maintained in nature in a mosquito-bird-mosquito transmission cycle primarily involving culex sp mosquitoes. wn virus was recently introduced to north america, where it was first detected in 1999 duri ...200212206968
in vitro determination of dengue virus type 2 ns2b-ns3 protease activity with fluorescent peptide substrates.the ns2b-ns3(pro) polyprotein segment from the dengue virus serotype 2 strain 16681 was purified from overexpressing e. coli by metal chelate affinity chromatography and gel filtration. enzymatic activity of the refolded ns2b-ns3(pro) protease complex was determined in vitro with dansyl-labeled peptide substrates, based upon native dengue virus type 2 cleavage sites. the 12mer substrate peptides and the cleavage products could be separated by reversed-phase hplc, and were identified by uv and fl ...200212297031
[new hepatitis viruses]serum obtained from a surgeon (with the initials gb) with acute icteric hepatitis, permited the passages of two virus to tamarins; furthermore was possible to detect anti-bodies against the putative epitope of this viruses, designed gbv-a and gbv-b in several human populations; many years after that two different researchers teams identified a new virus gbv-c or hvg, based on very similar aminoacid sequence of both.the role of gb-viruses and hepatitis g virus in human pathology is not completely ...199712221439
characterization and e protein expression of mutant strains during persistent infection of kn73 cells with japanese encephalitis virus.to study the character of mutants originating from japanese encephalitis viruses and the relationship between the characterization of mutant strains and e protein expression.200212411104
sentinel chicken surveillance programme in australia, 1 july 2001 to 30 june 2002.detection of flavivirus seroconversions in sentinel chicken flocks located throughout australia is used to provide an early warning of increased levels of murray valley encephalitis (mve) and kunjin virus activity in the region. during the 2001/2002 season low levels of flavivirus activity were detected in northern australia compared to previous years. mve and kunjin virus activity was detected in the kimberley and pilbara regions of western australia and the northern territory but not in north ...200212416705
solomon islands dengue seroprevalence study--previous circulation of dengue confirmed.since there were no confirmed outbreaks of dengue on record an extensive search of available records and a seroprevalence study were undertaken to determine if there was evidence of past dengue virus infections in the solomon islands. hospital and health department records revealed a suspected dengue outbreak in 1982 and deaths attributed to dengue in 1995 and 1996. serum samples from blood donors were analyzed for the presence of igg antibodies specific for flavivirus and dengue virus. a total ...200112418677
effect of triethylamine on the recovery of selected south american alphaviruses, flaviviruses, and bunyaviruses from mosquito (diptera: culicidae) pools.we evaluated the effect of triethylamine (tea) on the recovery of infectious virus from pools of mosquitoes for two south american alphaviruses (eastern equine encephalomyelitis and venezuelan equine encephalomyelitis subtypes iiic and id), one flavivirus (ilheus) and two bunyaviruses (mirim [guama group] and itaqui [group c]). mosquitoes were inoculated intrathoracically with virus, held for 7-10 d at 26 degrees c, and handled under one of four regimens before testing for the presence of virus ...200212349865
requirements for de novo initiation of rna synthesis by recombinant flaviviral rna-dependent rna polymerases.rna-dependent rna polymerases (rdrps) that initiate rna synthesis by a de novo mechanism should specifically recognize the template initiation nucleotide, t1, and the substrate initiation nucleotide, the ntpi. the rdrps from hepatitis c virus (hcv), bovine viral diarrhea virus (bvdv), and gb virus-b all can initiate rna synthesis by a de novo mechanism. we used rnas and gtp analogs, respectively, to examine the use of the t1 nucleotide and the initiation nucleotide (ntpi) during de novo initiati ...200212438578
complementation analysis of the flavivirus kunjin ns3 and ns5 proteins defines the minimal regions essential for formation of a replication complex and shows a requirement of ns3 in cis for virus assembly.we have previously reported successful trans-complementation of defective kunjin virus genomic rnas with a range of large lethal deletions in the nonstructural genes ns1, ns3, and ns5 (a. a. khromykh et al., j. virol. 74:3253-3263, 2000). in this study we have mapped further the minimal region in the ns5 gene essential for efficient trans-complementation of genome-length rnas in repbhk cells to the first 316 of the 905 codons. to allow amplification and easy detection of complemented defective r ...200212368319
ratios of subclinical to clinical japanese encephalitis (je) virus infections in vaccinated populations: evaluation of an inactivated je vaccine by comparing the ratios with those in unvaccinated populations.japanese encephalitis (je) virus is characterized as a virus that produces a large number of subclinical infections. in this report, we estimated a ratio of subclinical to clinical infections in vaccinated human populations who acquired natural infection with je virus, and evaluated protective capacity of the currently approved inactivated je vaccine by comparing the ratio with those reported for unvaccinated populations. we developed a sensitive immunostaining method for detecting nonstructural ...200212443667
prevalence of gb virus c/hepatitis g virus in hungary.in 1995 a new flavivirus, gb virus c/hepatitis g virus (gbv-c/hgv), was discovered. the aim of this study was to determine the prevalence of the virus in healthy persons and hepatitis patients in hungary. the sera of 408 healthy persons older than 60 years were tested for the presence of gbv-c/hgv antibodies, and 113 were positive (28%). eight of the 71 healthy persons younger than 60 years and twenty of the 51 sera (39%) taken from patients suffering from hepatitis of unknown origin proved to b ...200212443828
west nile virus vaccine.within the past 5 years, west nile encephalitis has emerged as an important disease of humans and horses in europe. in 1999, the disease appeared for the first time in the northeastern united states. west nile virus (a mosquito-borne flavivirus) has flourished in the north american ecosystem and is expected to expand its geographic range. in this review, the rationale for a human and veterinary vaccine is presented and a novel approach for rapid development of a molecularly-defined, live, attenu ...200112455232
epizootic activity of murray valley encephalitis virus in an aboriginal community in the southeast kimberley region of western australia: results of cross-sectional and longitudinal serologic studies.murray valley encephalitis (mve) virus is a mosquito-borne flavivirus causing severe encephalitis with a resultant high morbidity and mortality. in the period 1989-1993, we undertook a cross-sectional and longitudinal study by annually screening members of a small remote aboriginal community in northwestern australia for mve virus antibodies. of the estimated 250-300 people in the community, 249 were tested, and 52.6% had positive serology to mve. the proportion testing positive increased with i ...200212408675
prevalence of hepatitis g virus infection and homology of different viral strains in southern china.to investigate the prevalence of hepatitis g virus (hgv) infection and to analyse the homology of different hgv strains in southern china.200212439930
poly i:clc induction of the interferon system in mice: an initial study of four detection methods.induction of a large number of the components of the interferon (ifn) system (ifn genes, their mrnas, ifn proteins, ifn receptors, ifn signaling molecules, the ifn response genes, and their effector proteins) has been studied. less well studied is the comparative induction of these components in vivo. induction of ifn by double-stranded rna (dsrna) treatment mimics certain aspects of viral infection and induces the components of the ifn system. to determine the comparative sensitivity of detecti ...200212433283
the epidemiology of yellow fever in africa.yellow fever (yf) is still a major public heath problem, particularly in africa, despite the availability of a very efficacious vaccine. the world health organization estimates that there are 200,000 cases of yf annually, including 30,000 deaths, of which over 90% occur in africa. in the past 15 years, the number of yf cases has increased tremendously, with most of the yf activity in west africa. this increase in yf activity is in part due to a breakdown in yf vaccination and mosquito control pr ...200212475636
reactivity of serum samples from patients with a flavivirus infection measured by immunofluorescence assay and elisa.flavivirus infections are a significant public health problem, since several members of the flaviviridae family are highly pathogenic to humans. accurate diagnosis and differentiation of the infecting virus is important, especially in areas where many flaviviruses are circulating. in this study we evaluated a newly developed commercially available immunofluorescence assay (ifa) (indx, baltimore, md, usa) for the detection of igm and igg antibodies against dengue virus, yellow fever virus, japane ...200212467761
spontaneous mutations restore the viability of tick-borne encephalitis virus mutants with large deletions in protein c.the capsid protein, c, of tick-borne encephalitis virus has recently been found to tolerate deletions up to a length of 16 amino acid residues that partially removed the central hydrophobic domain, a sequence element conserved among flaviviruses which may be crucial for virion assembly. in this study, mutants with deletion lengths of 19, 21, 27, or 30 residues, removing more or all of this hydrophobic domain, were found to yield viable virus progeny, but this was without exception accompanied by ...200312477849
characterization of a siberian virus isolated from a patient with progressive chronic tick-borne encephalitis.a strain of tick-borne encephalitis virus designated zausaev (za) was isolated in siberia from a patient who died of a progressive (2-year) form of tick-borne encephalitis 10 years after being bitten by a tick. the complete genomic sequence of this virus was determined, and an attempt was made to correlate the sequence with the biological characteristics of the virus. phylogenetic analysis demonstrated that this virus belongs to the siberian subtype of tick-borne encephalitis virus. comparison o ...200312477807
role of the transmembrane domains of prm and e proteins in the formation of yellow fever virus envelope.flavivirus envelope proteins have been shown to play a major role in virus assembly. these proteins are anchored into cellular and viral membranes by their c-terminal domain. these domains are composed of two hydrophobic stretches separated by a short hydrophilic segment containing at least one charged residue. we investigated the role of the transmembrane domains of prm and e in the envelope formation of the flavivirus yellow fever virus (yfv). alanine scanning insertion mutagenesis has been us ...200312502797
novelity in gb virus c/hepatitis g virus and its controversy.several novel human rna viruses were identified in 1995-96 and were partially characterized that apparently can cause acute and chronic hepatitis both in monkeys and humans. these new viruses are related to the flavivirus hepatitis c. it is known to be distinct from other human hepatitis viruses (a, b, c, d, e). three viruses, identified by investigators at abbott labs, have been termed gb-a, gb-b and gb-c. gb-a and gb-b are likely tamarin viruses whereas gb-c infects humans only. all these thre ...200212476863
[cloning and sequencing of cdna in ns3 region of chinese hepatitis g virus isolated from henan province].to study the genetic characteristics of non-structural region 3 of hgv in china, a hgv-rna positive sample was collected from henan. hgv rna was extracted and amplified by rt-nested-pcr. we got four overlap fragments and they were cloned into plasmid vector pcdna ii. the cdna was sequenced by the sanger's method. nucleotide homology between henan strain and other reported hgv isolates (pnf2161, r10291, gbv-c) was 85.7% to 93.8% over the 2137 nucleotides examined. comparison of amino acid sequenc ...199812515161
[detection of hepatitis g virus infection among clinical patients with hepatitis/liver diseases in china by reverse transcription-nested polymerase chain reaction].according to two highly conserved genome sequences within the helicase(ns3) region and 5'-uncoding(5' utr) region, we designed two sets of primer pairs to detect hgv rna by rt-nested pcr in order to study hgv infection in chinese population. three hundred and fifty-four serum specimens of various liver diseases were collected from beijing, qin huangdao and henan areas. seventy-nine out of 354(22.3%) specimens were hgv rna positive. among 254 known clinical hepatitis/liver disease samples, 50(19. ...199812515201
[identification of hepatitis c or/and g virus rna in one tube by reverse transcription nested polymerase chain reaction].hepatitis c and g viruses belong to the flaviviridae family. they have similar modes of transmission. the dual infection of hgv and hcv were reported. a simultaneous detection for hcv or/and hgv was established by reverse transcription nested polymerase chain reaction(rt-nested pcr). the primers were derived from the 5' non-coding region of hcv and ns3 region of hgv, and the length of the pcr product was between 170 bp and 300 bp. rt-nested pcr can be finished in one test tube and hgv and/or hcv ...199812515202
[sequencing and analysis of the ns5 region of hgv/gbv-c gene].to study the genomic organization of non structural region 5 of hgv in china, 186 nucleotides of ns5 region of hgv were amplified, cloned and sequenced. the data were analyzed with computer programs. the results indicated that the gene sequence homology between 2 hgv isolates in this study was 95.2%. the comparison of these 2 isolates and 3 foreign isolates reported previously showed that the sequence homology was in the range of 84.4% to 92.4%, and was obvious by divergent from the foreign isol ...199812526350
strategies for the identification of inhibitors of west nile virus and other flaviviruses.the recent outbreaks of west nile virus (wnv) and other emerging and re-emerging flaviviruses, have made prevention and treatment of flavivirus infections a global public health priority. no inhibitor is currently available for use as an antiflavivirus therapy. in this communication, i review (i) stages of flavivirus replication representing potential targets for antiviral drug discovery; (ii) strategies for identification of flavivirus inhibitors; and (iii) existing antiviral inhibitors reporte ...200212476954
laboratory-acquired west nile virus infections--united states, 2002.west nile virus (wnv), a mosquito-borne flavivirus introduced recently to north america, is a human, equine, and avian neuropathogen. the majority of human infections with wnv are mosquito-borne; however, laboratory-acquired infections with wnv and other arboviruses also occur. this report summarizes two recent cases of wnv infection in laboratory workers without other known risk factors who acquired infection through percutaneous inoculation. laboratory workers handling fluids or tissues known ...200212537288
mycophenolic acid inhibits dengue virus infection by preventing replication of viral rna.dengue fever is a mosquito-borne viral disease of global importance with no available antiviral therapy. we assessed the ability of mycophenolic acid (mpa), a drug currently used as an immunosuppressive agent, to inhibit dengue virus (dv) antigen expression, rna replication, and virus production. pharmacological concentrations of mpa effectively blocked dv infection, decreasing the percentage of infected cells by 99% and the levels of secreted virus by up to a millionfold. results were reproduce ...200212504563
interferons, interferon inducers, and interferon-ribavirin in treatment of flavivirus-induced encephalitis in mice.we evaluated the prophylactic and therapeutic efficacy of interferon alpha-2b, pegylated interferon alpha-2b, poly(i. c), and ampligen against modoc virus encephalitis in an animal model for flavivirus infections. all compounds significantly delayed virus-induced morbidity (paralysis) and mortality (due to progressive encephalitis). viral load (as measured on day 7 postinfection) was significantly reduced by 80 to 100% in the serum, brain, and spleen in mice that had been treated with either int ...200312543691
sequence diversity of the capsid gene and the nonstructural gene ns2b of dengue-3 virus in vivo.previously, we studied the envelope (e) gene of dengue virus and reported that dengue-3 virus is present as a quasispecies. to investigate the extent of intrahost sequence variation of other dengue viral genes, we examined in this study the capsid (c) gene and the nonstructural gene, ns2b, derived directly from plasma dengue viruses from 18 confirmed dengue-3 patients. using reverse transcription-pcr, multiple clones of a 360-nucleotide region covering the c gene and of a 404-nucleotide region c ...200212482670
the extent of homologous recombination in members of the genus flavivirus.the family flaviviridae includes important human pathogens, such as dengue (den) virus, yellow fever (yf) virus and hepatitis c virus, many of which have emerged or re-emerged in recent years. until recently, flavivirus evolution was thought to proceed in a clonal manner, with diversity generated mainly through the accumulation of mutational changes. however, this assumption has now been shown to be invalid, with homologous recombination demonstrated in all three genera of the flaviviridae: sinc ...200312560576
new inferences from tree shape: numbers of missing taxa and population growth rates.the relative positions of branching events in a phylogeny contain information about evolutionary and population dynamic processes. we provide new summary statistics of branching event times and describe how these statistics can be used to infer rates of species diversification from interspecies trees or rates of population growth from intraspecies trees. we also introduce a phylogenetic method for estimating the level of taxon sampling in a clade. different evolutionary models and different samp ...200212554454
cleavage of protein prm is necessary for infection of bhk-21 cells by tick-borne encephalitis virus.flavivirus particles are synthesized in an immature form containing heterodimers of the proteins prm and e. shortly before release from the cell, prm is cleaved by the host protease furin to yield mature virions. in this study, the furin-mediated cleavage of the tick-borne encephalitis (tbe) virus protein prm was prevented by specific mutagenesis of the cleavage site. this resulted in the production of immature tbe virions, which were shown to be completely non-infectious in bhk-21 cells. this f ...200312533715
a rapid and convenient variant of fusion-pcr to construct chimeric flaviviruses.so far, full-length cdnas of chimeric flaviviruses have been constructed by restriction-enzyme cleavage of the gene(s) to be exchanged or by fusion-pcr of two amplified pcr fragments. the construction of a chimeric flavivirus by a faster and more convenient variant of the standard fusion-pcr is reported. a modoc/yellow fever chimeric virus was engineered in which the structural prm and e genes of yellow fever virus 17d were replaced by the homologous genes of modoc virus. in two pcr steps, a fus ...200312565155
[expression of non-structural region 3 gene of the chinese hgy and analysis of the antigenicity of the recombinant proteins].this study is to analyze the antigenicity of the ns3 proteins of chinese hgv and their potential use in the serological diagnosis.199912569774
[a study of hgv infection in liver tissue of hepatocellular carcinoma].to investigate the prevalence of hepatitis g virus (hgv) in hepatocellular carcinoma (hcc).199912569791
intrauterine west nile virus infection--new york, 2002.west nile virus (wnv), a mosquito-borne flavivirus and human neuropathogen, is epidemic in the united states. in 2002, newly recognized mechanisms of person-to-person wnv transmission were described, including possible transmission from mother to infant through breast milk. wnv has not been previously associated with intrauterine infection or adverse birth outcomes. this report describes a case of transplacental wnv transmission. pregnant women should take precautions to reduce their risk for wn ...200212537289
wicking assays for the rapid detection of west nile and st. louis encephalitis viral antigens in mosquitoes (diptera: culicidae).the recent outbreaks of west nile (wn) encephalitis and st. louis encephalitis (sle) in the united states have highlighted the need for rapid and specific methods of detecting arboviral antigens in mosquitoes. we evaluated rapid, field-usable assays for detecting and differentiating wn and sle viruses in mosquito pools, based on a patent-pending, immunochromatographic technology (vectest) formatted on a dipstick. the device provides results in less than 20 min and can be used in laboratories wit ...200312597660
epidemiology of hepatitis c virus infection in australia.since the discovery in 1989 of hepatitis c virus (hcv) as the infectious agent responsible for the vast majority of post-transfusion non-a non-b hepatitis the patterns of transmission and clinical consequences of this highly prevalent flavivirus have been widely studied.200312600649
polystyrene derivatives substituted with arginine interact with babanki (togaviridae) and kedougou (flaviviridae) viruses.outbreaks of new or old diseases appear primarily in tropical zones such as africa, south and central america, or asia. among these diseases, those induced by arboviruses (the best known of which are being yellow fever, dengue, ebola, and sindbis) are under intensive observation by the world health organization. rapid isolation and identification of the viral species is the first step in the diagnosis, study, and control of epidemics. one major problem with the isolation of viruses is capturing ...200312601758
impact of direct virus-induced neuronal dysfunction and immunological damage on the progression of flavivirus (modoc) encephalitis in a murine model.flavivirus encephalitis is believed to be the result of two main mechanisms: (i) direct damage to and dysfunction of neurons as a result of viral replication and (ii) destruction of the brain tissue by an inflammatory response. the differential impact of both mechanisms on the progression of flavivirus encephalitis has not been clearly determined. we have now studied the encephalitis caused by modoc virus (modv) infection in (i) severe combined immunodeficiency (scid) mice, (ii) immunocompetent ...200312587070
enzyme-linked immunosorbent assay using recombinant antigens expressed in mammalian cells for serodiagnosis of tick-borne encephalitis.a recombinant plasmid that expresses the tick-borne encephalitis (tbe) virus premembrane (prm) and envelope (e) proteins in mammalian cells was constructed. recombinant proteins retained antigenic and conformational structures similar to those of native virus proteins, and transfected cells released virus-like particles (vlps), which were 1.13-1.14 g/ml in density and 20-30 nm in diameter, into the culture medium. recombinant e proteins were used for the development of an enzyme-linked immunosor ...200312609684
[serological diagnosis of dengue and yellow fever infections in suspected cases from pará state, brazil, 1999].from june to december 1999, 785 serum samples were obtained from patients clinically suspected of having dengue or yellow fever. the patients were referred by public health centers distributed within the six mesoregions of par state, brazil. serum samples were tested for flavivirus antibodies by hemagglutination inhibition test and for dengue and yellow fever viruses by enzyme-linked immunosorbent assay for igm detection. of the sera collected, 563 (71.7%) were positive by hi test and out of th ...200312612738
hepatitis g virus (hgv): current perspectives.five viruses are usually associated with hepatitis in humans: a-e. in addition to these viruses as aetiological agents of hepatitis, there remain a number of patients with hepatitis in whom no virus could be identified. it was therefore postulated that there may be other agents which may be causing hepatitis. recently, two viruses have been associated with hepatitis: hepatitis g virus (hgv), and transfusion transmissible virus (ttv). hepatitis g virus (hgv) is a single stranded rna virus which r ...200212593580
spinal cord disease in west nile virus infection. 200312575663
epitope-blocking enzyme-linked immunosorbent assays for the detection of serum antibodies to west nile virus in multiple avian species.we report the development of epitope-blocking enzyme-linked immunosorbent assays (elisas) for the rapid detection of serum antibodies to west nile virus (wnv) in taxonomically diverse north american avian species. a panel of flavivirus-specific monoclonal antibodies (mabs) was tested in blocking assays with serum samples from wnv-infected chickens and crows. selected mabs were further tested against serum samples from birds that represented 16 species and 10 families. serum samples were collecte ...200312624027
tick-borne encephalitis.tick-borne encephalitis (tbe) is one of the most dangerous human infections occurring in europe and many parts of asia. the etiological agent tick-borne encephalitis virus (tbev), is a member of the virus genus flavivirus, of the family flaviviridae. tbev is believed to cause at least 11,000 human cases of encephalitis in russia and about 3000 cases in the rest of europe annually. related viruses within the same group, louping ill virus (liv), langat virus (lgtv) and powassan virus (powv), also ...200312615309
diagnosis of tick-borne encephalitis.the actual diagnosis of a tick-borne encephalitis (tbe) must be established in the laboratory because of the non-specific clinical features it presents. the method of choice is the demonstration of specific igm- and igg-serum antibodies by enzyme-linked immuno-sorbent assay (elisa), since these antibodies are detectable in practically every case at the time of hospitalization. early after onset of disease in the cerebrospinal fluid specific antibodies can only be found in 50% of the patients, bu ...200312628812
history of tbe vaccines.tick borne encephalitis was a frequent viral cns disease in europe in the last century, with several hundred hospitalisation cases per year. the causative agent is tick-borne encephalitis virus (tbev), a flavivirus that is transmitted by ticks. the first tissue culture derived vaccine was developed in the early 1970s and this was commercialised as a collaboration between the institute of virology, vienna, austria and immuno ag vienna (now baxter healthcare). this vaccine was highly successful du ...200312628813
enhancing biosynthesis and secretion of premembrane and envelope proteins by the chimeric plasmid of dengue virus type 2 and japanese encephalitis virus.we have constructed a series of plasmids encoding premembrane (prm) and envelope (e) protein genes of dengue virus type 2 (den-2). these plasmids included an authentic den-2 prm-e construct (pcbd2-14-6), and two chimeric constructs, 90% den-2 e-10% japanese encephalitis (je) virus e (pcb9d2-1j-4-3) and 80% den-2 e-20% je e (pcb8d2-2j-2-9-1). monoclonal antibody (mab) reactivity indicated that all three plasmids expressed authentic den-2 virus e protein epitopes representative of flavivirus domai ...200312620809
role of type i and type ii interferon responses in recovery from infection with an encephalitic flavivirus.we have investigated the contribution of the interferon (ifn)-alpha/beta system, ifn-gamma and nitric oxide to recovery from infection with murray valley encephalitis virus, using a mouse model for flaviviral encephalitis where a small dose of virus was administered to 6-week-old wild-type and gene knockout animals by the intravenous route. we show that a defect in the ifn-alpha/beta responses results in uncontrolled extraneural virus growth, rapid virus entry into the brain and 100 % mortality. ...200312604807
inoculation of plasmids encoding japanese encephalitis virus prm-e proteins with colloidal gold elicits a protective immune response in balb/c mice.we established a simple and effective method for dna immunization against japanese encephalitis virus (jev) infection with plasmids encoding the viral prm and e proteins and colloidal gold. inoculation of plasmids mixed with colloidal gold induced the production of specific anti-jev antibodies and a protective response against jev challenge in balb/c mice. when we compared the efficacy of different inoculation routes, the intravenous and intradermal inoculation routes were found to elicit strong ...200312634382
intracellular assembly and secretion of recombinant subviral particles from tick-borne encephalitis virus.it is believed that flavivirus assembly occurs by intracellular budding of the nucleocapsid into the lumen of the endoplasmic reticulum (er). recombinant expression of tick-borne encephalitis (tbe) virus envelope proteins prm and e in mammalian cells leads to their incorporation into enveloped recombinant subviral particles (rsps), which have been used as a model system for studying assembly and entry processes and are also promising vaccine candidates. in this study, we analyzed the formation a ...200312634393
steady-state cleavage kinetics for dengue virus type 2 ns2b-ns3(pro) serine protease with synthetic peptides.the n-terminal part of the ns3 protein from dengue virus contains a trypsin-like serine protease responsible for processing the nonstructural region of the viral polyprotein. enzymatic activity of the ns2b-ns3(pro) precursor incorporating a full-length ns2b cofactor of dengue virus type 2 was examined by using synthetic dodecamer peptide substrates encompassing native cleavage sequences of the ns2a/ns2b, ns2b/ns3, ns3/ns4a and ns4b/ns5 polyprotein junctions. cleavage of the dansylated substrates ...200312625822
cross-protective and infection-enhancing immunity in mice vaccinated against flaviviruses belonging to the japanese encephalitis virus serocomplex.the japanese encephalitis virus serocomplex is a group of mosquito-borne flaviviruses that cause severe encephalitic disease in humans. the recent emergence of several members of this serocomplex in geographic regions where other closely related flaviviruses are endemic has raised urgent human health issues. thus, the impact of vaccination against one of these neurotropic virus on the outcome of infection with a second, serologically related virus is unknown. we show here that immunity against m ...200312639478
antibody and interleukin-12 treatment in murine models of encephalitogenic flavivirus (st. louis encephalitis, tick-borne encephalitis) and alphavirus (venezuelan equine encephalitis) infection.early and sustained treatment with interleukin-12 (il-12) ameliorated disease in a mouse model of infection with the encephalitogenic flavivirus, st. louis encephalitis virus (slev, japanese encephalitis serogroup). however, this effect was not reproduced in murine infections with either the flavivirus tick-bore encephalitis virus (tbev) or the alphavirus venezuelan equine encephalitis virus (veev). il-12 exacerbated tbev disease when used in conjunction with monoclonal antibody (mab), suggestin ...200312639298
a stable full-length yellow fever virus cdna clone and the role of conserved rna elements in flavivirus replication.yellow fever virus (yf) is the prototype member of the flavivirus genus. here, we report the successful construction of a full-length infectious cdna clone of the vaccine strain yf-17d. yf cdna was cloned into a low-copy-number plasmid backbone and stably maintained in several e. coli strains. transcribed rnas had a specific infectivity of 10(5)-10(6) p.f.u. ( micro g rna)(-1), and the resulting virus exhibited growth kinetics, plaque morphology and proteolytic processing similar to the parental ...200312692292
interferon alfa-2a in japanese encephalitis: a randomised double-blind placebo-controlled trial.japanese encephalitis virus (jev), although confined to asia, causes about 35000-50000 cases and 10000 deaths every year, and is the most important cause of encephalitis worldwide. there is no known antiviral treatment for any flavivirus. results from in-vitro studies and work in animals have shown inteferon alfa has antiviral activity on japanese encephalitis and other flaviviruses; therefore, we aimed to assess the efficacy of inteferon alfa-2a in japanese encephalitis.200312642049
architecture of the flaviviral replication complex. protease, nuclease, and detergents reveal encasement within double-layered membrane compartments.flavivirus infection causes extensive proliferation and reorganization of host cell membranes to form specialized structures called convoluted membranes/paracrystalline arrays and vesicle packets (vp), the latter of which is believed to harbor flaviviral replication complexes. using detergents and trypsin and micrococcal nuclease, we provide for the first time biochemical evidence for a double membrane compartment that encloses the replicative form (rf) rna of the three pathogenic flaviviruses w ...200312700232
the importance of being outer: consequences of the distinction between the outer and inner surfaces of flavivirus glycoprotein e.glycoprotein e of west nile, dengue and other flaviviruses is the principal stimulus for the development of neutralizing antibodies and contains a fusion peptide responsible for inserting the virus into the host cell membrane. this glycoprotein lies flat on the surface of the virion and therefore only epitopes on the outer or lateral surface are important immunogens. changes in antigen recognition after exposure of the virus to low ph have yielded clues to the fusion process.200312648938
characterization of rna synthesis, replication mechanism, and in vitro rna-dependent rna polymerase activity of japanese encephalitis virus.in vitro rna-dependent rna polymerase assays revealed that the jev replication complex (rc) synthesized viral rna utilizing a semiconservative and asymmetric mechanism. peak viral replicase activity and levels of viral rna observed 15-18 h postinfection (h p.i.) preceded maximum viral titers in the culture medium seen 21 h p.i. among divalent cations, mg(2+) was essential and exhibited cooperative binding for its two replicase-binding sites. mn(2+), despite sixfold higher affinity for the replic ...200312667804
a new cell line from the embryonic tissues of culex tritaeniorhynchus and its susceptibility to certain flaviviruses.a new cell line from the embryonic tissue of culex tritaeniorhynchus mosquito was established. morphological studies carried out at the 45th passage level (p-45) showed four different cell types viz. epithelial-like cells, fibroblast-like cells, giant cells and vacuolated cells. karyological studies indicated diploid (2n = 6) chromosomes in majority of cells irrespective of passage level. a twelve-fold increase of cell number was observed in 10 days at p-49. the cells could be preserved in liqui ...200212693860
recent advances in japanese encephalitis.japanese encephalitis (je), the most important cause of epidemic encephalitis worldwide, is confined to asia, but its geographical area is spreading. west nile virus, and other closely related flaviviruses, cause similar disease elsewhere. recent cryoelectron microscopic studies have characterized the flavivirus envelope protein as a new class of viral fusion protein (class ii), and examined its arrangement on the virion surface. changes in the envelope protein's hinge region, or its putative re ...200312707858
interferon, ribavirin, 6-azauridine and glycyrrhizin: antiviral compounds active against pathogenic flaviviruses.ribavirin, interferon-alpha (ifn-alpha), 6-azauridine and glycyrrhizin were tested in vitro for their antiviral activities against 11 pathogenic flaviviruses belonging to principal antigenic complexes or individual serogroups of medical importance: dengue, japanese encephalitis, mammalian tick-borne and yellow fever virus (yfv) groups. antiviral activity was estimated by the reduction of the cytopathic effect of each flavivirus in vero cells and by the reduction in virus titer. cytotoxicity was ...200312719009
temperature sensitive mutations in the genes encoding the ns1, ns2a, ns3, and ns5 nonstructural proteins of dengue virus type 4 restrict replication in the brains of mice.using reverse genetics, it is possible to readily add well-defined attenuating mutations to the genome of wild type or incompletely attenuated dengue (den) viruses to generate vaccine candidates that exhibit the desired balance between attenuation and immunogenicity. here, we describe the identification of eight temperature sensitive missense mutations distributed in four non-structural protein genes that specify a 60- to 10,000-fold range of restricted replication in the suckling mouse brain co ...200312721805
viral internal ribosome entry site structures segregate into two distinct morphologies.an increasing number of viruses have been shown to initiate protein synthesis by a cap-independent mechanism involving internal ribosome entry sites (iress). predictions of the folding patterns of these rna motifs have been based primarily on sequence and biochemical analyses. biophysical confirmation of the models has been achieved only for the ires of hepatitis c virus (hcv), which adopts an open structure consisting of two major stems. we have conducted an extensive comparison of flavivirus a ...200312743317
evasion of innate and adaptive immunity by flaviviruses.after a virus infects an animal, antiviral responses are generated that attempt to prevent dissemination. interferons, antibody, complement, t and natural killer cells all contribute to the control and eradication of viral infections. most flaviviruses, with the exception of some of the encephalitic viruses, cause acute disease and do not establish persistent infection. the outcome of flavivirus infection in an animal is determined by a balance between the speed of viral replication and spread, ...200312752684
interaction of flaviviruses with cells of the vertebrate host and decoy of the immune response.flaviviruses cause endemic and epidemic disease with significant morbidity and mortality throughout the world. in contrast to viruses that avoid the host immune response by down-regulating cell surface major histocompatibility complex expression, infection by members of the neurotropic japanese encephalitis serogroup induce virus-directed functional increases in expression of class i and ii major histocompatibility complex and various adhesion molecules, resulting in increased susceptibility to ...200312752685
mhc class i up-regulation by flaviviruses: immune interaction with unknown advantage to host or pathogen.in contrast to many other viruses that escape from cytotoxic t cell recognition by down-regulating major histocompatibility complex class i-restricted antigen presentation, flavivirus infection of mammalian cells up-regulates cell surface expression of major histocompatibility complex class i molecules. two putative mechanisms for flavivirus-induced major histocompatibility complex class i up-regulation, one via activation of the transcription factor nf-kappab, the second by augmentation of pept ...200312752686
is flavivirus resistance interferon type i-independent?interferon type i comprises a group of major virus-inducible host antiviral factors that control infection with a great number of human and animal viruses. they are ubiquitously expressed cytokines that interfere with virus replication within different cell types by activating a number of host genes and several parallel antiviral pathways. two major intracellular actors of ifn-i-induced antiviral states are ribonucleic acid-dependent protein kinase and 2'-5'-oligoadenylate synthetases/rnase l, b ...200312752687
[hepatitis c virus infection mimicking hemorrhagic fever: a case report]acute c hepatitis, compared with hepatitis a or b, may sometimes be associated with extrahepatic manifestations of a haematologic nature. the first case of hepatitis associated with aplastic anemia was reported by lorenz in 1955. we observed a patient who developed acute hepatitis, associated with febrile purpura: pancytopenia was present, but not aplasia. this association is frequent in flavivirus-related infections, like dengue and yellow fever, and principally affects children and young peopl ...200012714845
isolation of a new flavivirus related to cell fusing agent virus (cfav) from field-collected flood-water aedes mosquitoes sampled from a dambo in central kenya.cell fusing agent virus (cfav) is an rna insect virus that was isolated from a line of aedes aegypti mosquito cells and has been assigned to the family flaviviridae, genus flavivirus. we report here the first isolation of a cfa-like virus from field-collected mosquitoes. mosquito larvae and pupae were sampled from flooded dambos in central province, kenya during the short rain season of 1999. specimens were reared to adults, identified and pooled by species and were tested for the presence of vi ...200312756616
genetic and phenotypic characterization of the newly described insect flavivirus, kamiti river virus.we have described in the accompanying paper by sang, et al., ([57], arch virol 2003, in press) the isolation and identification of a new flavivirus, kamiti river virus (krv), from ae. macintoshi mosquitoes that were collected as larvae and pupae from flooded dambos in central province, kenya. among known flaviviruses, krv was shown to be most similar to, but genetically and phenotypically distinct from, cell fusing agent virus (cfav). krv was provisionally identified as an insect-only flavivirus ...200312756617
[west nile fever may even be spread to sweden].west nile virus (wnv) is a flavivirus, which was first isolated in uganda 1937. this virus has attracted attention in the past years. wnv is one of the worlds most widespread flaviviruses and has caused recent outbreaks among humans and animals. west nile fever (wnf) is a mosquito borne zoonotic disease. the last decade the epidemiological pattern of the virus and the severity of the outbreaks have changed. this article reviews the epidemiological changes and current situation of wnf, presents a ...200312756695
lipophilic derivatization of synthetic peptides belonging to ns3 and e2 proteins of gb virus-c (hepatitis g virus) and its effect on the interaction with model lipid membranes.the synthesis by solid-phase methodologies of peptides belonging to structural and non-structural proteins of gb virus c as well as its n-alpha-acylation with myristate and palmitate fatty acids is described. to explore the peptide-lipid interactions we have used liposomes composed of dipalmitoylphosphatidylcholine as model membranes and complementary spectroscopic and calorimetric techniques. our results show that structural and more clearly the structural lipophilic peptide sequences incorpora ...200312753379
flavivirus capsid is a dimeric alpha-helical protein.the capsid proteins of two flaviviruses, yellow fever virus and dengue virus, were expressed in escherichia coli and purified to near homogeneity suitable for biochemical characterization and structure determination by nuclear magnetic resonance. the oligomeric properties of the capsid protein in solution were investigated. in the absence of nucleic acid, both proteins were predominantly dimeric in solution. further analysis of both proteins with far-uv circular dichroism spectroscopy indicated ...200312768036
stable rna structures can repress rna synthesis in vitro by the brome mosaic virus replicase.a 15-nucleotide (nt) unstructured rna with an initiation site but lacking a promoter could direct the initiation of rna synthesis by the brome mosaic virus (bmv) replicase in vitro. however, bmv rna with a functional initiation site but a mutated promoter could not initiate rna synthesis either in vitro or in vivo. to explain these two observations, we hypothesize that rna structures that cannot function as promoters could prevent rna synthesis by the bmv rna replicase. we documented that four d ...200312702814
crystallization and preliminary x-ray diffraction analysis of langat virus envelope protein domain iii.the putative receptor-binding domain (domain iii) of the flavivirus langat envelope glycoprotein has been crystallized using the hanging-drop vapor-diffusion method at 277 k. two distinct crystal morphologies were observed to grow under the same conditions. the crystal forms both belong to a trigonal space group, p3(1)21 or p3(2)21, with unit-cell parameters a = 80.93, c = 132.1 a and a = 104.8, c = 219.5 a for forms i and ii, respectively. complete data sets to 2.9 and 3.35 a, respectively, hav ...200312777770
molecular detection of west nile virus rna.west nile virus is a mosquito-borne flavivirus that is primarily maintained in nature in a mosquito-bird-mosquito transmission cycle. mammals, including humans and horses, are incidentally infected through biting by mosquitoes infected with west nile virus. since 1994, west nile virus outbreaks have occurred with a high incidence of severe disease in humans and horses. in the usa, west nile virus was first detected in 1999 in new york city and has since spread to 39 states in humans. the virus h ...200312779009
traditional and novel approaches to flavivirus vaccines.yellow fever, dengue, japanese encephalitis and tick-borne encephalitis viruses are the medically most important members of the flavivirus genus composed primarily of arboviruses. in this paper, we review the commercially available traditional flavivirus vaccines against yellow fever, japanese encephalitis, and tick-borne encephalitis, as well as modern approaches to flavivirus vaccines. formalin inactivation technology has been employed to produce killed vaccines. flaviviruses have been attenua ...200312782056
role of heparan sulfate for attachment and entry of tick-borne encephalitis virus.attachment of the flavivirus tick-borne encephalitis (tbe) virus to different permissive cell lines was investigated by a newly established quantitative assay using fluorescence-labeled virus. previous work had shown that bhk-21 cell-adapted mutants of tbe virus had acquired potential heparan sulfate (hs) binding sites on the outer surface of protein e. quantitative analysis of one of these mutants indicated that it attached to hs-expressing cell lines with a 10- to 13-fold higher affinity than ...200312706093
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