the flavivirus 3'-noncoding region: extensive size heterogeneity independent of evolutionary relationships among strains of tick-borne encephalitis virus.the sequences of the 3'-noncoding regions (ncr) of 12 strains of tick-borne encephalitis (tbe) virus were analyzed and found to vary in length from 350 to approximately 750 nucleotides. the size heterogeneity is restricted to a variable region following the stop codon, whereas the most 3'-terminal 350 nucleotides form a highly conserved core element containing several potentially important sequence motifs and secondary structure elements. a homoadenosine tract previously thought to form the 3'-t ...19957483260
complex formation between the ns3 serine-type proteinase of the hepatitis c virus and ns4a and its importance for polyprotein maturation.processing of the hepatitis c virus polyprotein is mediated by host cell signalases and at least two virally encoded proteinases. of these, the serine-type proteinase encompassing the amino-terminal one-third of ns3 is responsible for cleavage at the four sites carboxy terminal of ns3. the activity of this proteinase is modulated by ns4a, a 54-amino-acid polyprotein cleavage product essential for processing at the ns3/4a, ns4a/4b, and ns4b/5a sites and enhancing cleavage efficiency between ns5a ...19957494258
posttranslational signal peptidase cleavage at the flavivirus c-prm junction in vitro.we have investigated the cleavages at the flavivirus capsid-prm protein junction in vitro. when expressed in the absence of the flavivirus proteinase, capsid and prm, which are separated by an internal signal sequence, exist as a membrane-spanning precursor protein. here we show the induction of posttranslational signal peptidase cleavage of prm by trypsin cleavage of a cytoplasmic region of this precursor protein.19957494334
preparation of recombinant baculovirus by transfection of a ligated cdna fragment without prior plasmid amplification in e. coli. 19957495544
the langat model for tick-borne encephalitis virus. specific detection by rt-pcr.we have developed a reverse-transcriptase polymerase chain reaction assay for rapid detection of langat (lgt) virus, a flavivirus that is closely related to the highly pathogenic tick-borne encephalitis (tbe) viruses. unlike tbe viruses, lgt virus exhibits a significantly lower virulence for man. the assay serves as a safe alternative for the development and optimization of specific assays for the highly pathogenic subtypes of tbe viruses that are endemic throughout much of europe, the former so ...19937505284
epitope analysis of tick-borne encephalitis (tbe) complex viruses using monoclonal antibodies to envelope glycoprotein of tbe virus (persulcatus subtype).the arrangement of envelope protein epitopes of tick-borne encephalitis viruses (tbev) (persulcatus or eastern subtype, sofjin strain and ricinus or western subtype, minsk-256 strain) and kyasanur forest disease virus (kfdv) was investigated using competitive binding of monoclonal antibodies against the sofjin e protein. the e protein of tbev sofjin strain forms three antigenic domains: e1, e2 and e3, represented by 12, 9 and 2 epitopes respectively; two additional epitopes stand alone. domains ...19937505512
[structural proteins of hepatitis c virus]. 19937510088
an integrated target sequence and signal amplification assay, reverse transcriptase-pcr-enzyme-linked immunosorbent assay, to detect and characterize flaviviruses.we previously described a reverse transcriptase-pcr using flavivirus genus-conserved and virus species-specific amplimers (d. w. trent and g. j. chang, p. 355-371, in y. becker and c. darai; ed., frontiers of virology, vol. 1, 1992). target amplification was improved by redesigning the amplimers, and a sensitive enzyme-linked immunosorbent assay (elisa) technique has been developed to detect amplified digoxigenin (dig)-modified dna. a single biotin motif and multiple dig motifs were incorporated ...19947512096
tick-borne flavivirus ns1 gene: identification of conserved peptides and antigenic analysis of recombinant louping ill virus ns1 protein.the nucleotide sequence of the ns1 gene of louping ill (li) virus has been determined. the sequence shows a high degree of homology with other members of the tick-borne serocomplex of flaviviruses and a lower homology with the mosquito-borne flaviviruses. alignment of the deduced ns1 amino acid sequences with all tick-borne flavivirus ns1 sequences, identified four peptide regions which were conserved for all tick-borne flaviviruses, but were variable amongst mosquito-borne flaviviruses. a dendr ...19947513932
the flavivirus nonstructural protein ns3 is a dominant source of cytotoxic t cell peptide determinants.vaccinia virus recombinants encoding regions of the murray valley encephalitis virus (mve) genome, which together cover the entire viral coding region, were employed to identify the mve protein which is the dominant source of cd8+, cytotoxic, t cell antigenic determinant(s) presented by the mouse h-2kk major histocompatibility antigen. mve and west nile virus-immune, h-2k-restricted, effector cells recognized peptides derived from the mve nonstructural polyprotein segment, and in this region the ...19947516597
analysis of computer-predicted antibody inducing epitope on japanese encephalitis virus.theoretical methods to delineate antibody inducing epitopes have been employed to predict antigenic determinants on envelope glycoprotein (gpe) of japanese encephalitis (je), west nile (wn) and dengue (den) i-iv viruses. a predicted region on je virus gpe 74cpttgeahnekrad87 was synthesized, conjugated to klh (klh-peptide) and used in immunization of mice. a mouse monoclonal antibody (moab ivb4) reactive to the peptide was also found to react with native je virus gpe. characterization of the idio ...19937516621
identification of mosquito-borne flavivirus sequences using universal primers and reverse transcription/polymerase chain reaction.a reverse transcription/polymerase chain reaction (rt/pcr) protocol for the rapid detection and identification of flaviviruses was developed using a set of universal oligonucleotide primers. these primers correspond to sequences in the 3' non-coding region and in the ns5 gene which are highly conserved among the mosquito-borne flaviviruses. the sequences of the resulting amplified products were analysed for dengue 1, dengue 2, dengue 3, dengue 4, japanese encephalitis, west nile, yellow fever an ...19947520190
antibody capture immunoassay detection of japanese encephalitis virus immunoglobulin m and g antibodies in cerebrospinal fluid.immunoglobulin m (igm) and igg antibodies to japanese encephalitis virus (jev) were detected in acute-phase cerebrospinal fluid (csf) specimens from patients with acute encephalitis by using a solid-phase radioimmunoassay of the antibody capture type. of 12 patients with jev infections subsequently proven by hemagglutination inhibition serology, 11 had jev igm antibodies, as measured by antibody capture radioimmunoassay, in the first csf specimen (geometric mean titer, 1:2,500) compared with 0 o ...19827161371
neuropathology of s. paulo south coast epidemic encephalitis (rocio flavivurus).the neuropathology of 8 cases of s. paulo south coast epidemic encephalitis (rocio flavivirus), a new arbo b virus encephalitis, is described. the topographic pattern of the lesions appears to be almost specific. the gray matter is predominantly affected. interstitial mononuclear infiltration, microglial proliferation and perivascular lymphocytic cuffing were seen. neuronophagia was seldom seen except during the acute phases of the disease. throughout the neuraxis, the gray matter was affected t ...19807359160
tick-borne encephalitis virus envelope protein e-specific monoclonal antibodies for the study of low ph-induced conformational changes and immature virions.a set of ten monoconal antibodies (mabs) specific for the tick-borne encephalitis (tbe) virus envelope protein e were prepared and characterized with respect to their functional activities, the location of their binding sites on protein e and the involvement of their epitopes in acid ph-induced conformational changes and interactions with the precursor to the membrane protein (prm) in immature virions. the majority of these mabs mapped to the previously defined antigenic domain a. all of the mab ...19957535997
lack of augmenting effect of interferon-gamma on dengue virus multiplication in human peripheral blood monocytes.the effect of interferon-gamma (ifn-gamma) on dengue virus multiplication in human peripheral blood monocytes was investigated. enriched monocytes were treated with ifn-gamma and then infected with dengue virus type 2 either directly or in the presence of optimal infection-enhancing levels of antibodies. pretreatment of monocytes from dengue-immune donors with 100 iu/ml of ifn-gamma caused 12- to 97-fold and 13- to 137-fold reduction of virus yields at 24 hr after infection in the absence and pr ...19957536230
oligomeric rearrangement of tick-borne encephalitis virus envelope proteins induced by an acidic ph.the flavivirus envelope protein e undergoes irreversible conformational changes at a mildly acidic ph which are believed to be necessary for membrane fusion in endosomes. in this study we used a combination of chemical cross-linking and sedimentation analysis to show that the envelope proteins of the flavivirus tick-borne encephalitis virus also change their oligomeric structure when exposed to a mildly acidic environment. under neutral or slightly alkaline conditions, protein e on the surface o ...19957529335
dengue virus-specific, hla-b35-restricted, human cd8+ cytotoxic t lymphocyte (ctl) clones. recognition of ns3 amino acids 500 to 508 by ctl clones of two different serotype specificities.dengue virus infections are a major cause of morbidity and mortality in tropical and subtropical areas of the world. we analyzed dengue virus-specific cd8+ cd4- ctl at the clonal level to further understand the role of cd8+ ctl in dengue virus infections. dengue virus-specific cd8+ ctl clones were established from lymphocytes of a dengue 4-immune adult. three patterns of dengue serotype specificities were identified: 1) specific for dengue 4, 2) cross-reactive for dengue 2 and dengue 4 (subcompl ...19957529799
flavivirus-cross-reactive, hla-dr15-restricted epitope on ns3 recognized by human cd4+ cd8- cytotoxic t lymphocyte clones.the role of flavivirus-cross-reactive t lymphocytes in recovery from and pathogenesis of flavivirus infections is not known. in the present paper, we have defined a flavivirus-cross-reactive epitope recognized by two cd4+ cd8- cytotoxic t lymphocyte (ctl) clones, jk4 and jk43. the t cell clones were established from the peripheral blood t lymphocytes of a dengue-4-immune donor, using a limiting-dilution method with dengue-4 antigen. these two t cell clones were cross-reactive for dengue virus ty ...19957561761
nucleotide sequence of the ns5 gene of banzi virus: comparison with other flaviviruses.banzi is a mosquito borne flavivirus which belongs to the uganda s serocomplex. no nucleotide sequence data have previously been reported from any virus of this serocomplex. we have determined the nucleotide sequence of the ns5 gene from banzi virus and the predicted amino acid sequence was elucidated. previously identified conserved rna polymerase, methyltransferase and flavivirus ns5 amino acid motifs were present in the banzi virus ns5 protein. these data add to the evidence for the functiona ...19957561771
low resolution mapping around the flavivirus resistance locus (flv) on mouse chromosome 5.although the phenomenon of innate resistance to flaviviruses in mice was recognized many years ago, it was only recently that the genetic locus (flv) controlling this resistance was mapped to mouse chromosome (chr) 5. here we report the fine mapping of the flv locus, using 12 microsatellite markers which have recently been developed for mouse chr 5. the new markers were genotyped in 325 backcross mice of both (c3h/hej x c3h/rv)f1 x c3h/hej and (balb/c x c3h/rv)f1 x balb/c backgrounds, relative t ...19957579886
[epidemiological analysis of cases of tbe in 1993 in bialystok district].111 cases of tbe was 20 times increased morbidity in comparison to 1992 in bialystok district in 1993. among hospitalized dominated male patients (52.35%). first cases was reported in may and last in november with a peak in july. the age of patients was from 6 to 78 years old, 63.06%, in age group from 31 to 59 years old. we established that 78.87% patients were citizens of bialystok, spending there recreation time in forest. despite occurrence increased cases of tbe in 1993 we conclude that amo ...19947597185
wesselsbron virus antibody in domestic animals in nigeria: retrospective and prospective studies.retrospective and prospective serological surveys to determine the prevalence of wesslsbron (wsl) virus infections in animal populations were carried out in different vegetational zones in nigeria. sera from 1,492 animals comprising 292 camels, 81 horses, 4 donkeys, 320 cattle, 235 sheep, 260 goats, 114 pigs, 101 dogs and 85 domestic fowls were assayed by haemagglutination-inhibition (hi) test for presence of antibodies to wsl virus and other flavivirus antigens: yellow fever (yf), potiskum (pot ...19957603342
molecular and serologic analysis in the transmission of the gb hepatitis agents.two flavivirus-like genomes have recently been cloned from infectious tamarin (saguinus labiatus) serum, derived from the human viral hepatitis gb strain, which is known to induce hepatitis in tamarins. in order to study the natural history of gb infections, further transmission studies were carried out in tamarins. reverse-transcription-polymerase chain reaction and enzyme-linked immunosorbant assays were developed for the detection of rna and antibodies associated with the two agents, gb virus ...19957623012
formation of polymeric glycoprotein complexes from a flavivirus: tick-borne encephalitis virus.treatment of tick-borne encephalitis (tbe) virus with triton x-100 (tx-100), octylglucoside (og) or cetyltrimethylammonium bromide (ctab) caused dissociation of the virus envelope into dimers or monomers of the glycoprotein v3. by centrifugation into detergent-free sucrose density gradients, these subunits were found to reassociate and to form haemagglutinating homogeneous glycoprotein complexes sedimenting at 15 to 16, 16 to 18 and 11 to 23s after tx-100, og and ctab treatment, respectively. gl ...19807420059
up-regulation of mhc class i by flavivirus-induced peptide translocation into the endoplasmic reticulum.flavivirus infection of mammalian cells increases the cell surface expression of major histocompatibility complex (mhc) class i molecules, the recognition elements for cytotoxic t cells. here, we show that the mechanism for flavivirus-induced up-regulation of class i mhc involves an increase in peptide supply to the endoplasmic reticulum. flavivirus-mediated peptide supply for mhc class i assembly is independent of the peptide transporters for class i antigen presentation, since infection of cla ...19957544229
association between ns3 and ns5 proteins of dengue virus type 2 in the putative rna replicase is linked to differential phosphorylation of ns5.dengue virus type 2, a member of the family flaviviridae, encodes a single polyprotein precursor consisting of 3391 amino acids residues that is processed to at least 10 mature proteins by host and viral proteases. the ns3 protein contains a domain commonly found in cellular serine proteinases that in cooperation with ns2b is involved in polyprotein processing. in addition, ns3 and ns5 proteins contain conserved motifs found in several rna helicases and rna-dependent rna polymerases, respectivel ...19957642575
a novel complex formed between the flavivirus e and ns1 proteins: analysis of its structure and function.we examined the structural features and functional significance of a novel complex which forms between the envelope (e) protein and nonstructural protein ns1 of murray valley encephalitis (mve) virus. western blot analysis of virus-infected c6/36 cell lysates revealed that the undenatured form of this e-ns1 complex was a heat-sensitive e-(ns1 dimer) complex. furthermore, the e-ns1 complex was observed in cells infected with kunjin, japanese encephalitis, west-nile and kokobera viruses which indi ...19957646339
epitope mapping of dengue 1 virus e glycoprotein using monoclonal antibodies.ten monoclonal antibodies (mabs) were raised against dengue 1 (den 1, hawaii) virus e glycoprotein. specificity of the mabs was tested by elisa and immunofluorescence. eight were den 1 type-specific, one was den group-reactive (dgr) and one was flavivirus cross-reactive (fcr). two of these type specific mabs exhibited haemagglutination-inhibition (hi) and neutralized (n) den 1 virus in vivo (hs). these two mabs showed 100% protection against a challenge of 100 ld50 of den 1 virus in adult swiss ...19957646356
viral infection of macrophages profoundly alters requirements for induction of nitric oxide synthesis.activated mouse macrophages can produce high levels of nitric oxide, an antimicrobial effector molecule recently also implicated in antiviral defense. as viral infection may alter macrophage functions, nitric oxide production was investigated in murine macrophages infected with a flavivirus, tick-borne encephalitis virus. infected macrophages produced high levels of nitric oxide upon stimulation with lipopolysaccharide without priming, while in control macrophages induction of nitric oxide produ ...19957676626
expression and identification of hepatitis c virus polyprotein cleavage products.hepatitis c virus (hcv) is the major cause of transfusion-acquired non-a, non-b hepatitis. hcv is an enveloped positive-sense rna virus which has been classified as a new genus in the flavivirus family. like the other two genera in this family, the flaviviruses and the pestiviruses, hcv polypeptides appear to be produced by translation of a long open reading frame and subsequent proteolytic processing of this polyprotein. in this study, a cdna clone encompassing the long open reading frame of th ...19937679746
definition of an hla-dpw2-restricted epitope on ns3, recognized by a dengue virus serotype-cross-reactive human cd4+ cd8- cytotoxic t-cell clone.we previously reported that the clone jk34 was cross-reactive for dengue virus types 1, 2, 3, and 4 and recognized ns3 (i. kurane, m. a. brinton, a. l. samson, and f. a. ennis, j. virol. 65:1823-1828, 1991). in the present experiments, we defined the epitope at the amino acid level, with 93 15-mer overlapping peptides which cover the entire ns3. a peptide 4 which contains amino acids 251 to 265 of ns3 sensitized the autologous b lymphoblastoid cell line (lcl) to the lysis by jk34. the smallest p ...19937690424
comparative analysis of ns3 sequences of temporally separated dengue 3 virus strains isolated from southeast a combination of pcr and direct-cycle sequencing using consensus primers, we analyzed approximately 400-bp fragments within the ns3 genes of twenty-one dengue virus type 3 strains isolated from five neighboring southeast asian countries at different time intervals from 1956 to 1992. the majority of base disparities were silent mutations, with few predicted amino acid substitutions, thus emphasizing the strict conservation of the ns3 gene. phylogenetic trees constructed on the basis of these n ...19947698880
the virus causing encephalomyelitis in sheep in spain: a new member of the tick-borne encephalitis group.the nucleotide and deduced primary amino acid sequence of the envelope gene of two virus isolates from the brains of spanish sheep with encephalomyelitis, were determined and compared with those of other flaviviruses. the amino acid alignments showed that the spanish viruses shared 95 to 96 per cent homology with the envelope protein of louping ill virus and western european tick-borne encephalitis virus. in comparison, the maximum variation in amino acid identities among strains of louping ill ...19957709053
[meningoencephalo-myeloradiculitis due to flavivirus: bi-brachial paralysis and respiratory insufficiency].3 patients developed rapid onset of fever and nuchal stiffness. paresis of brachial muscles occurred within 4 days and all patients had respiratory failure that needed mechanical ventilation. at the peak of the disease there were bilateral asymmetrical severe atrophy of brachial, shoulder and neck muscles, cranial nerve pareses and absent or weak deep reflexes in the upper extremities. csf analyses showed sterile lymphocytic pleocytosis. in 2 cases the patients suffered a tick bite in switzerlan ...19957709179
[biological properties and antigenic interconnection between tiuleniy and karshi flaviviruses].biological properties of flaviviruses tyuleny and karshi, new for the science, were studied. the viruses are highly pathogenic for laboratory animals; they replicate fairly well in primary chick embryo cells and ps cells. bhk-21 cells were found susceptible to replication of karshi virus. tyuleny and karshi viruses were found capable of initiating and maintaining a persistent infection in primary brain cell cultures from suckling syrian hamsters for at least 3 months. both the viruses showed cyt ...19947716902
studies on serological cross-reaction in sequential flavivirus infections.acute- and convalescent-phase sera from patients with dengue (den) hemorrhagic fever (dhf) and japanese encephalitis (je) that contained pre-existing flavivirus antibodies were tested for cross-reacting antibodies to den, je and yellow fever (yf) viruses by a neutralization (n) test. a fourfold or greater rise in n antibody titer in the convalescent-phase was considered significant. of 39 dhf cases, obtained at chiang mai university hospital, thailand, 15 (38.5%) showed a rise in den antibody ti ...19947723688
identification of two flavivirus-like genomes in the gb hepatitis agent.a subtractive pcr methodology known as representational difference analysis was used to clone specific nucleotide sequences present in the infectious plasma from a tamarin infected with the gb hepatitis agent. eleven unique clones were identified, seven of which were examined extensively. all seven clones appeared to be derived from sequences exogenous to the genomes of humans, tamarins, saccharomyces cerevisiae, and escherichia coli. in addition, sequences from these clones were not detected in ...19957724574
japanese encephalitis virus nonstructural protein ns3 has rna binding and atpase activities.sequence data suggest that japanese encephalitis virus (jev) protein ns3 is a multifunctional protein with sequence motifs characteristic of a protease and a helicase. to examine the functions of jev-ns3, a fusion protein of ns3 in escherichia coli was generated. analysis by western blot using monospecific rabbit antisera generated against the fusion protein (anti-mbjen3) showed that ns3 was localized in the membrane fraction of jev-infected cells and the particulate fraction of bacteria extract ...19957732656
rapid, single-step rt-pcr typing of dengue viruses using five ns3 gene order to detect and type dengue viruses in serum specimens, four type-specific downstream primers were designed for use with a consensus upstream primer in a reverse transcription and polymerase chain reaction (rt-pcr) assay. rt-pcr using these five primers amplified ns3 gene fragments of diagnostic sizes of 169, 362, 265 and 426 base pairs for dengue virus types 1, 2, 3 and 4, respectively, but not for japanese encephalitis, kunjin and yellow fever viruses. the conventional two-step rt-pcr p ...19957738139
immunodominant epitopes on the ns1 protein of mve and kun viruses serve as targets for a blocking elisa to detect virus-specific antibodies in sentinel animal serum.two mosquito-borne flaviviruses, murray valley encephalitis (mve) and kunjin (kun), are the aetiological agents of australian encephalitis. mve causes a severe and potentially fatal form of the disease while kun is responsible for only a few relatively mild cases. therefore it is important that serological tests used in flavivirus surveillance differentiate between infections with these two viruses. however, this has been hampered in the past by the close antigenic relationships between flavivir ...19957738140
isolation of wesselsbron virus from ostriches. 19957740726
a community-based study of subclinical flavivirus infections in children in an area of tamil nadu, india, where japanese encephalitis is endemic.a characteristic feature of the epidemiology of japanese encephalitis (je) is the occurrence of a large number of subclinical infections. the reporting of only overt cases underestimates the total level of virus transmission, a knowledge of which is essential for the evolution of control strategies. we carried out a 3-year prospective serological study between 1989 and 1991 in a primary health centre in tamil nadu where je is endemic. each year paired specimens, taken before and after the transm ...19957743596
the filovirus enigma. 19957746050
[stegomyian indices and epidemiological status of yellow fever in a rural area of the ivory coast].the m'bahiakro region, located in the center of ivory coast and inhabited by the n'gain, has been the scene of a yellow fever epidemic since 1982. this region reunites all the conditions for cyclic emergence of the flavivirus given the current epidemiologic pattern of yellow fever in africa. in view of this situation orstom, in collaboration with the pasteur institute of ivory coast, has created a pilot zone for epidemiologic surveillance of yellow fever in this region. the n'gain region which h ...19947746124
early induction of interferon-independent virus-specific icam-1 (cd54) expression by flavivirus in quiescent but not proliferating fibroblasts--implications for virus-host interactions.west nile virus (wnv) infection of human embryonic fibroblasts can induce expression of icam-1 by two distinct mechanisms. an early and direct mechanism occurs within 2 hr of virus infection which is cytokine independent, and an indirect mechanism occurs within 24 hr of virus infection and is regulated by the release of ifn-type 1. virus-inactivated, conditioned supernatants removed from wnv-infected fibroblast cultures at 4 hr did not alter icam-1 expression on fresh, untreated fibroblasts, whe ...19957747416
adherence status regulates the primary cellular activation responses to the flavivirus west nile.increases in cell-surface intercellular adhesion molecule-1 (icam-1; cd54) and major histocompatibility complex antigen class i (mhc-i) and class ii (mhc-ii) expression during flavivirus infection of murine macrophages was strongly dependent on adherence status. cd54 and mhc expression was significantly increased during infection with the flavivirus west nile (wnv) on adherent, but not on non-adherent, macrophages. in contrast, increased cd54 and mhc-i expression was induced by interferon-gamma ...19957751002
the envelope glycoprotein from tick-borne encephalitis virus at 2 a resolution.the crystallographically determined structure of a soluble fragment from the major envelope protein of a flavivirus reveals an unusual architecture. the flat, elongated dimer extends in a direction that would be parallel to the viral membrane. residues that influence binding of monoclonal antibodies lie on the outward-facing surface of the protein. the clustering of mutations that affect virulence in various flaviviruses indicates a possible receptor binding site and, together with other mutatio ...19957753193
complete nucleotide sequence of yellow fever virus vaccine strains 17dd and 17d-213.the complete nucleotide sequence of the genome from two yellow fever (yf) virus vaccine strains, 17dd and 17d-213, has been determined. comparison of these sequences with those of other yf viruses including the parental virulent asibi strain allowed the identification of 48 nucleotide sequence differences which are common to all 17d substrains. this is a significant reduction from the 67 nucleotide changes originally reported as being 17d-specific and potentially related to viral attenuation. th ...19957754673
immunisation against japanese encephalitis. 19957760642
alphavirus and flavivirus glycoproteins: structures and functions. 19957774004
dengue-3 (16562) pgmk 33 vaccine: neurovirulence, viremia and immune responses in macaca fascicularis.investigation of monkey neurovirulence of dengue-3 viruses (den-3, 16562) was undertaken to provide an evaluation of the relative safety of virus strain attenuated for potential use of live virus vaccine. ten flavivirus-negative, cynomolgus monkeys (macacafascicularis) were used in the test. the animals were inoculated intrathalamically, intraspinally and intramuscularly with den-3 pgmk 33 attenuated live virus vaccine (6 monkeys): parent virus (2) and control cell culture fluid (2). blood sampl ...19947777925
analysis of flavivirus envelope proteins reveals variable domains that reflect their antigenicity and may determine their pathogenesis.studies on the molecular basis of flavivirus neutralisation, attenuation and tropism indicate that amino acid substitutions, in different parts of the envelope gene, may be responsible for the altered phenotypes. however, the association of particular substitutions with individual characteristics has proven difficult. comparative analysis of all known tick-borne flavivirus envelope proteins through sequence alignment and a sliding window, reveals clusters of amino acid variation distributed thro ...19957785318
an arbovirus cline across the northern hemisphere.the mode and tempo of arbovirus evolution and dispersal can help to explain the dynamics of pandemics, viral outbreaks, and emerging viruses. by comparing nucleotide and deduced amino acid sequences of their envelope proteins, we describe the continuous distribution of the tick-borne encephalitis (tbe) complex viruses, the most important flaviviruses in europe, across major geographical areas and the conditions under which mutations occur. the analyses reveal a correlation between the geographic ...19957793067
susceptibility of the sf9 insect cell line to infection with adventitious viruses.sf9, the insect cell line commonly used for gene expression by recombinant baculovirus (bv), can be infected by st. louis encephalitis (sle) virus, a flavivirus, resulting in a persistent, productive, and cytopathic infection, while retaining the ability to be infected with a recombinant baculovirus (rbv). we now demonstrate using double immunofluorescence that single cells are dually infected with sle virus and rbv. fourteen additional viruses including additional flaviviruses, other arbovirus ...19947811453
isolations of west nile and bagaza viruses from mosquitoes (diptera: culicidae) in central senegal (ferlo).during october-november 1990, 31,497 mosquitoes consisting of 25 different species were collected in barkedji, ferlo area (senegal), and tested for virus infection. viruse were isolated from 55 of 407 pools. eighteen pools were found positive for both bagaza virus (bga) and west nile virus (wn). one alphavirus (babanki [bbk] and 72 flaviviruses (19 bga, 53 wn) were isolated from culex poicilipes theobald (29 wn, 8 bga), c. neavei theobald (3 wn, 1 bga), mimomyia hispida theobald (8 wn, 6 bga, an ...19947815413
intracellular interference of tick-borne flavivirus infection by using a single-chain antibody fragment delivered by recombinant sindbis virus.a single-chain antibody fragment that identifies a neutralizing epitope on the envelope protein of louping ill and some other tick-borne flaviviruses was previously expressed in soluble form from bacteria and shown to be functionally active in vitro. to see whether or not the single-chain antibody could bind and inactivate infectious virus in vivo, we have used recombinant sindbis virus as a delivery vehicle for intracellular expression of the antibody fragment. the variable genes and interchain ...19957815482
reverted virulence of attenuated tick-borne encephalitis virus mutant is not accompanied with the changes in deduced viral envelope protein amino acid sequence.serial passages of tick-borne encephalitis (tbe) virus strain 4387 isolated from the liver and lungs of the bank vole through the salivary glands of ixodes ricinus ticks led to a reduction of its virulence for laboratory mice infected via peripheral route. when attenuated mutants were passaged through mouse brains, virulent phenotypes have appeared in the 3rd mouse passage. after 5 consecutive passages the virus was more pathogenic for mice after peripheral inoculation than the parental 4387 str ...19947817894
a single nine-amino acid peptide induces virus-specific, cd8+ human cytotoxic t lymphocyte clones of heterogeneous serotype is generally accepted that virus-specific cd8+ cytotoxic t lymphocytes (ctls) recognize nine-amino acid peptides in conjunction with hla class i molecules. we recently reported that dengue virus-specific cd8+ ctls of two different serotype specificities, which were established by stimulation with dengue virus, recognize a single nine-amino acid peptide of the nonstructural protein ns3 of dengue virus type 4 (d4v) in an hla-b35-restricted fashion. to further analyze the relationships between t ...19957544398
bhk cell proteins that bind to the 3' stem-loop structure of the west nile virus genome rna.the first 83 3' nucleotides of the genome rna of the flavivirus west nile encephalitis virus (wnv) form a stable stem-loop (sl) structure which is followed in the genome by a smaller sl. these 3' structures are highly conserved among divergent flaviviruses, suggesting that they may function as cis-acting signals for rna replication and as such might specifically bind to cellular or viral proteins. cellular proteins from uninfected and wnv-infected bhk-21 s100 cytoplasmic extracts formed three di ...19957637011
synthesis and secretion of recombinant tick-borne encephalitis virus protein e in soluble and particulate form.a quantitative study was performed to investigate the requirements for secretion of recombinant soluble and particulate forms of the envelope glycoprotein e of tick-borne encephalitis (tbe) virus. full-length e and a carboxy terminally truncated anchor-free form were expressed in cos cells in the presence and absence of prm, the precursor of the viral membrane protein m. formation of a heteromeric complex with prm was found to be necessary for efficient secretion of both forms of e, whereas only ...19957637027
monoclonal antibodies directed against tick-borne encephalitis virus with neutralizing activity in vivo.monoclonal antibodies (moabs) were raised against the tick-borne encephalitis (tbe) virus, strain k23. the reactivities of 14 selected moabs were characterized by elisa, western blot analysis, haemagglutination inhibition, immunoprecipitation, in vivo protection and in vitro neutralization tests. all moabs reacted only with the glycoprotein e. the binding epitope of one moab could be delimited by a synthetic peptide to amino acids 306-339 representing one immunodominant loop structure of the gly ...19947817895
[immuno-allergic meningeal reaction following anti-flavivirus vaccination]. 19947824472
hepatitis c virus-encoded nonstructural protein ns4a has versatile functions in viral protein processing.a transient protein expression system in cos-1 cells was used to study the role of hepatitis c virus (hcv)-encoded ns4a protein on hcv nonstructural polyprotein processing. by analyzing the protein expression and processing of a deletion mutant polypeptide, ns delta 4a, which encodes the entire putative hcv nonstructural polyprotein except the region encoding ns4a, the versatile functions of ns4a were revealed. most of the ns3 processed from ns delta 4a was localized in the cytosol fraction and ...19957853491
mutagenesis of the yellow fever virus ns2b/3 cleavage site: determinants of cleavage site specificity and effects on polyprotein processing and viral replication.the determinants of cleavage site specificity of the yellow fever virus (yf) ns3 proteinase for its 2b/3 cleavage site have been studied by using site-directed mutagenesis. mutations at residues within the garr decreases s sequence were tested for effects on cis cleavage of an ns2b-3(181) polyprotein during cell-free translation. at the p1 position, only the conservative substitution r-->k exhibited significant levels of cleavage. conservative and nonconservative substitutions were tolerated at ...19957853494
an amino-terminal domain of the hepatitis c virus ns3 protease is essential for interaction with ns4a.hepatitis c virus (hcv) genomic rna is translated into a large polyprotein that is processed into structural and nonstructural proteins. processing at the n termini of several nonstructural proteins requires sequences contained in both ns3 and ns4a. ns3 contains a serine protease, whereas the function of ns4a in proteolysis is yet to be determined. by using the vaccinia virus-t7 hybrid expression system to transiently express hcv polypeptides in hela cells, we studied the effect of several n-ter ...19957853516
phylogeny of tyu, sre, and cfa virus: different evolutionary rates in the genus flavivirus.the nucleotide and deduced amino acid sequence of the envelope (e) gene of tyuleniy (tyu) and saumarez reef (sre) virus have been determined and the data used to classify these viruses in relation to the other tick- and mosquito-borne viruses in the genus flavivirus. the phylogenetic trees obtained by maximum parsimony and distance methods for 22 flavivirus e genes showed that tyu and sre virus are a sister group of the tbe virus complex. the trees were consistent with the flavivirus serological ...19957856087
cryosubstitution technique reveals new morphology of flavivirus-induced structures.cryotechniques in combination with electron microscopy were used in an attempt to obtain more precise morphological details of flavivirus-induced structures. from conventional chemical fixation procedure, proliferation of endoplasmic reticulum, formation of microtubule paracrystals and clusters of smooth membrane vesicles (with 'thread-like' enclosures) were observed. these induced changes are typical for flavivirus infections. the images obtained from cryosections were disappointing as the stru ...19947868648
dengue fever virus and japanese encephalitis virus synthetic peptides, with motifs to fit hla class i haplotypes prevalent in human populations in endemic regions, can be used for application to skin langerhans cells to prime antiviral cd8+ cytotoxic t cells (ctls)--a novel approach to the protection of humans.flaviviruses were reported to induce cd8+ cytotoxic t cells in infected individuals, indicating that nonapeptides, proteolytic cleavage products of the viral precursor protein, enter the endoplasmic reticulum in infected cells and interact with hla class i molecules. the assembled hla class i molecules are transported to the plasma membrane and prime cd8+ t cells. current knowledge of the interaction of viral peptides with hla molecules is reviewed. based on this review, an idea is presented to ...19947871759
formation of the flavivirus envelope: role of the viral ns2b-ns3 of the late processing events in the flavivirus replication cycle involves cleavage of the intracellular form of the flavivirus capsid protein (cint) to the mature virion form (cvir) lacking the carboxy-terminal stretch of hydrophobic amino acids which serves as a signal peptide for the downstream prm protein. this cleavage event was hypothesized to be effected by a viral protease and to be associated with virion formation. we have proposed a model of flavivirus virion formation in which pro ...19957884844
[genome sequence and antigen structure of the powassan virus: analysis of genetic elements of tick-transmitted flaviviruses]. 19957886964
processing of japanese encephalitis virus non-structural proteins: ns2b-ns3 complex and heterologous proteases.processing of japanese encephalitis (je) virus non-structural (ns) proteins expressed by recombinant vaccinia viruses was analysed to characterize the responsible viral protease. analysis of the processing of polyprotein ns2a-2b-3' containing the n-terminal 322 amino acids of ns3 revealed products consistent with cleavages at the predicted intergenic junctions as well as at one or possibly two sites within ns2a. cleavage at the alternate site(s) containing the cleavage sequence motif within ns2a ...19957897348
[differences in susceptibility to peripheral infection with japanese encephalitis virus among inbred strains of mouse].we compared susceptibility of inbred mouse strains against intracerebral as well as peripheral challenge of a flavivirus, japanese encephalitis (je) virus, and the results were summarized as follows: (1) seven inbred mouse strains (c3h/he, c57bl/6, balb/c, akr/n, nc, nzb, dba/2) could be classified into following 3 groups by their mortality and infection rate when they were subcutaneously challenged with a wild je virus strain isolated from field-caught mosquitoes at a single passage in mouse (b ...19947900284
el niño-southern oscillation and vector-borne disease. 19937901589
brefeldin a affects west nile virus replication in vero cells but not c6/36 cells.a fungal metabolite brefeldin a (bfa) was used to study virus-host interaction in glycoprotein processing in west nile virus-infected vero and c6/36 cells. the results indicated that as little as 1 microgram/ml of bfa resulted in complete breakdown in the golgi organelle in infected vero cells. this led to modifications of the glycoproteins which could not be efficiently used in infectious virion formation. in contrast, as much as 10 micrograms/ml of bfa in culture medium did not affect either g ...19937903673
the interactions of the flavivirus envelope proteins: implications for virus entry and release.viral membrane proteins play an important role in the assembly and disassembly of enveloped viruses. oligomerization and proteolytic cleavage events are involved in controlling the functions of these proteins during virus entry and release. using tick-borne encephalitis virus as a model we have studied the role of the flavivirus envelope proteins e and prm/m in these processes. experiments with acidotropic agents provide evidence that the virus is taken up by receptor-mediated endocytosis and th ...19947913359
detection of transfusion-associated hepatitis caused by non-a, non-b, non-c flavivirus.sera of patients suffering from acute hepatitis, and different forms of chronic hepatitis were found to be reactive to reagents prepared from the yellow fever virus (yf) vaccine strain. serum samples of 1974 patients were tested, and 133 of them were positive. hepatitis c virus specific antibodies were absent from the majority of them. the frequency of antibodies to other flaviviruses (tick-borne encephalitis, west nile) and hepatitis b virus markers was similar to that measured among the popula ...19947921854
specificity of the hepatitis c virus ns3 serine protease: effects of substitutions at the 3/4a, 4a/4b, 4b/5a, and 5a/5b cleavage sites on polyprotein processing.cleavage at four sites (3/4a, 4a/4b, 4b/5a, and 5a/5b) in the hepatitis c virus polyprotein requires a viral serine protease activity residing in the n-terminal one-third of the ns3 protein. sequence comparison of the residues flanking these cleavage sites reveals conserved features including an acidic residue (asp or glu) at the p6 position, a cys or thr residue at the p1 position, and a ser or ala residue at the p1' position. in this study, we used site-directed mutagenesis to assess the impor ...19947933136
cytokine concentrations in cerebrospinal fluid in flavivirus infections. 19937939960
processing of flavivirus structural glycoproteins: stable membrane insertion of premembrane requires the envelope signal peptide.the flavivirus structural proteins capsid (c), premembrane (prm), and envelope (e) are cleaved in that order from the n-terminus of the polyprotein by the er intralumenal enzyme signal peptidase. the prm-e and e-ns1 junctions contain hydrophobic domains with both transmembrane and signal function. these domains reside at the c-termini of prm and e, respectively, after cleavage. we studied the functions of the 37-amino-acid c-terminus of the dengue virus type 4 (den4) prm (amino acids 243-279 of ...19947941319
flavivirus west nile (sarafend) egress at the plasma membrane.west nile (sarafend) virus was distinctly observed to bud from the plasma membrane rather than mature intracellularly. this has been observed with transmission electron microscopy. using conventional scanning electron microscopy, budding at the plasma membrane especially at the filopodia was clearly illustrated. immunogold labelling against the virus envelope protein was also performed to confirm this mode of exit. the gold particles were observed to be located at the sites where virus budding w ...19947944952
generation of long flavivirus expression cassettes by in vivo recombination and transient dominant selection.assembly of expression cassettes coding for large segments of viral polyproteins is often complicated or impossible due to the instability of the resulting recombinant (re-) plasmids during propagation in escherichia coli. using the transient dominant selection approach described for the construction of vaccinia virus recombinants (re-vv), we have constructed several intermediate vectors and developed a procedure which enables direct assembly of long expression cassettes in the vv genome by in v ...19947958993
kinetics and cross-reactivity of the virus-specific antibody-forming cells in mice during primary and secondary infection with japanese encephalitis virus and related flaviviruses.a modified antibody-forming cell assay was used to enumerate splenocytes secreting antibodies to japanese encephalitis (je) virus and four other flaviviruses in infected cells as the target antigens. the optimal viral antigen expression for the assay was standardized in the infected cells for each virus and the incubation time varied depending of the cell line and the virus. the kinetics of response of je virus-infected mice readily showed igg isotype switching. antibody-forming splenocytes of m ...19947962258
expression of cloned envelope protein genes from the flavivirus tick-borne encephalitis virus in mammalian cells and random mutagenesis by pcr.the structural membrane proteins prm and e of the flavivirus tick-borne encephalitis (tbe) virus were expressed in mammalian cells for the purpose of probing the structure and molecular interactions of these proteins. advantage was taken of the natural error frequency of the taq polymerase used in the pcr amplification to generate a randomly mutated population of genes that were then cloned directly into plasmid expression vectors under the control of an sv40 promoter. analysis of the mutation f ...19947975266
towards a new generation of flavivirus vaccines.flavivirus diseases have caused great public health concern for over three centuries, with diseases like yellow fever, dengue, japanese encephalitis and tick-borne encephalitis causing thousands of deaths. although yellow fever epidemics can be brought under control by the use of vaccine or mosquito-control measures, there have been many examples of its re-emergence as an epidemic disease. similarly, the use of vaccines or arthropod-control measures have failed to prevent the spread of other fla ...19947975848
elisa for detecting dengue and japanese encephalitis viral antigen in indirect elisa has been developed for detecting two viruses in triturated, experimentally infected mosquitoes: dengue (den) in aedes aegypti (l.) and ae. albopictus (skuse) and japanese encephalitis (je) viral antigen in culex tritaeniorhynchus giles. den antigen from four strains of the virus, representing each of the four serotypes, and je antigen were captured with a polyclonal anti-flavivirus igg and detected with a monoclonal antibody (4g2) that reacts with all flaviviruses. minimum vira ...19947979627
substrate determinants for cleavage in cis and in trans by the hepatitis c virus ns3 proteinase.processing of the hepatitis c virus polyprotein is accomplished by a series of cotranslational and posttranslational cleavages mediated by host cell signalases and two virally encoded proteinases. of these the ns3 proteinase is essential for processing at the ns3/4a, ns4a/4b, ns4b/5a, and ns5a/5b junctions. processing between ns3 and ns4a occurs in cis, implying an intramolecular reaction mechanism, whereas cleavage at the other sites can also be mediated in trans. sequence analysis of the amino ...19957983710
[viral encephalitis after tick bite].tbe is caused by a flavivirus and is endemic in certain areas of central and eastern europe, such as austria, the south of germany, hungary, czechoslovakia, poland, the former yugoslavia and sng and certain areas of sweden and finland. recent developments in the eastern european countries have made them popular with danish travellers, who are thus exposed to infection. until now tbe has been a very rare disease in denmark. the clinical course is biphasic being preceded by an incubation period of ...19947985272
use of a flavivirus rna-dependent rna polymerase assay to investigate the antiviral activity of selected compounds.we have developed an assay using flavivirus rna-dependent rna polymerase to test the inhibitory activity of potential antiviral agents. the effects of antiviral agents on rna synthesis were examined in this assay using extracts of vero cells infected with dengue virus type 2 or kunjin virus. several different classes of known polymerase inhibitors were tested. the synthesis of double-stranded replicative form rna was inhibited in a dose-dependent fashion in the presence of the polyoxometalate hp ...19947993077
dengue haemorrhagic fever (dhf) outbreak in outbreak of dengue haemorrhagic fever (dhf) occurred in calcutta between september and december, 1990. children and young adults were the major victims. haemorrhagic manifestations and shocks were the main features in most of the hospitalised cases. five mouse pathogenic agents were isolated from 105 acute cases and all were identified as den-3. hi and cf test with 55 paired sera revealed evidence of dengue infection in 33 (60 per cent) and flavivirus group reaction including dengue in 17 (30 ...19938014433
[flavivirus encephalitis in an urban area]. 19948029200
arboviruses causing human disease in the australasian zoogeographic region.over 65 arboviruses have been reported from countries in the australasian zoogeographic region, but only a few have been implicated in human disease. these include the flaviviruses murray valley encephalitis (mve), kunjin (kun), kokobera (kok), and dengue, particularly types 1 and 2; the alphaviruses ross river (rr), barmah forest (bf), and sindbis (sin); and the bunyaviruses, gan gan and trubanaman. in this paper recent epidemiological and clinical results pertaining to these viruses are review ...19948031248
processing of dengue type 4 and other flavivirus nonstructural proteins.dengue type 4 (den4) and other flaviviruses employ host and viral proteases for polyprotein processing. most proteolytic cleavages in the den4 nonstructural protein (ns) region are mediated by the viral ns2b-ns3 protease. the n-terminal third of ns3, containing sequences homologous to serine protease active sites, is the protease domain. to determine required sequences in ns2b, deletions were introduced into den4 ns2b-30% ns3 cdna and the expressed polyproteins assayed for self-cleavage. a 40 am ...19948032267
a case of japanese encephalitis.we report a case of japanese encephalitis that occurred in a woman who had spent only a few days in an area where she could have been exposed to the virus. the risks and protective efficacy of vaccination against japanese encephalitis virus for travellers who visit endemic areas for only a short period are discussed.19948034997
kinetic and structural analyses of hepatitis c virus polyprotein processing.recombinant vaccinia viruses were used to study the processing of hepatitis c virus (hcv) nonstructural polyprotein precursor. hcv-specific proteins and cleavage products were identified by size and by immunoprecipitation with region-specific antisera. a polyprotein beginning with 20 amino acids derived from the carboxy terminus of ns2 and ending with the ns5b stop codon (amino acids 1007 to 3011) was cleaved at the ns3/4a, ns4a/4b, ns4b/5a, and ns5a/5b sites, whereas a polyprotein in which the ...19948035505
processing of the intracellular form of the west nile virus capsid protein by the viral ns2b-ns3 protease: an in vitro study.according to the existing model of flavivirus polyprotein processing, one of the cleavages in the amino-terminal part of the flavivirus polyprotein by host cell signalases results in formation of prm (precursor to one of the structural proteins, m) and the membrane-bound intracellular form of the viral capsid protein (cint) retaining the prm signal sequence at its carboxy terminus. this hydrophobic anchor is subsequently removed by the viral protease, resulting in formation of the mature viral c ...19948057458
[a serological survey regarding flaviviridae infections on the island of réunion (1971-1989)].serological prevalence of flaviviridae was studied on reunion island by testing 2,507 human sera from a randomised sample. each serum was tested against 5 viruses (yellow fever, dengue type 1 and 2, west nile and wesselsbron) using haemagglutination inhibition test: 42.68% of human sera were found positive. the multivalent reactions represent practically three fourths of the positive ones. a severe dengue type 2 outbreak on the island in 1977-1978 and the possible circulation of a flavivirus may ...19948061530
postvaccinal plexus neuropathy following vaccination against tick-borne encephalitis and tetanus in a competitive athlete. 19948086776
rapid identification of flavivirus using the polymerase chain reaction.a rapid and accurate detection and identification system was developed for flaviviruses that makes use of reverse transcription-polymerase chain reaction (rt-pcr). a primer pair (yf-1 and yf-3), which corresponds to the highly conserved sequence at 3' noncoding region among flaviviruses, was useful for identification of mosquito-borne flaviviruses. nine sets of species specific primer pairs were also selected to identify and distinguish species, i.e., yellow fever, west nile, murray valley encep ...19938097200
complete genomic sequence of powassan virus: evaluation of genetic elements in tick-borne versus mosquito-borne flaviviruses.the complete nucleotide sequence of the positive-stranded rna genome of the tick-borne flavivirus powassan (10,839 nucleotides) was elucidated and the amino acid sequence of all viral proteins was derived. based on this sequence as well as serological data, powassan virus represents the most divergent member of the tick-borne serocomplex within the genus flaviviruses, family flaviviridae. the primary nucleotide sequence and potential rna secondary structures of the powassan virus genome as well ...19938097605
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