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passive protection assay of monoclonal antibodies against dengue virus in suckling mice.dengue fever and dengue hemorrhagic fever/dengue shock syndrome are highly infectious diseases caused by dengue virus (dv). specific monoclonal antibodies (mabs) against dv are vital for diagnosis, pathological studies, and passive immune therapy. in this study, purified dv serotype 2 (dv2) was used as antigen and balb/c mice were immunized to induce specific antibodies. we established five hybridoma cell lines, called 78#, 1e7, 7f7, 8f12, and 8h1, respectively, and evaluated them by enzyme-link ...200919189182
application of the dengue virus ns1 antigen rapid test for on-site detection of imported dengue cases at airports.we used the dengue virus ns1 antigen (ag) rapid test for on-site detection of imported dengue cases at airports. among 22 positive cases of dengue identified from 850 patients with a fever suspected to have dengue, 17 were ns1 ag test positive. these findings demonstrate the usefulness of the ns1 ag rapid test in screening imported dengue cases at airports.200919193828
the cellular immune response plays an important role in protecting against dengue virus in the mouse encephalitis model.for several years, researchers have known that the generation of neutralizing antibodies is a prerequisite for attaining adequate protection against dengue virus. nevertheless, the cellular immune response is the principal arm of the adaptive immune system against non-cytopathic viruses such as dengue, as once the virus enters into the cell it is necessary to destroy it to eliminate the virus. to define the role of the cellular immune response in the protection against dengue, we selected the mo ...200919210225
dengue virus type 2 infections of aedes aegypti are modulated by the mosquito's rna interference pathway.a number of studies have shown that both innate and adaptive immune defense mechanisms greatly influence the course of human dengue virus (denv) infections, but little is known about the innate immune response of the mosquito vector aedes aegypti to arbovirus infection. we present evidence here that a major component of the mosquito innate immune response, rna interference (rnai), is an important modulator of mosquito infections. the rnai response is triggered by double-stranded rna (dsrna), whi ...200919214215
potential development of temephos resistance in aedes aegypti related to its mechanism and susceptibility to dengue virus.the addition of temephos to water containers as a larvicide against aedes aegypti was commonly used as a part of dhf control programs. the widespread, or long-term, application of insecticides can lead to the development of mosquito resistance to the insecticides through selection pressure. this presents a problem for disease control. therefore, this study was conducted in the laboratory to observe the potential development of resistance to temephos and the mechanism involved in ae. aegypti, and ...200319230585
evaluation of dengue nonstructural protein 1 antigen strip for the rapid diagnosis of patients with dengue infection.two hundred twenty samples obtained from 104 patients with dengue infection (n = 89) and other febrile illnesses (n = 15) were assayed for dengue nonstructural protein 1 (ns1) antigen by enzyme immunoassay and by an immunochromatography (lateral flow) test strip. the sensitivity and the specificity of dengue ns1 antigen strip were 98.9% and 90.6%, respectively.200919232857
evaluation of dengue ns1 test kits for the diagnosis of dengue fever.detection of the dengue ns1 antigen during the symptomatic phase of illness represents an important advance in the diagnosis of dengue fever. the sensitivity of 2 commercial kits was evaluated in a panel of 91 serum samples collected at defined intervals after onset of symptomatic dengue fever. the sensitivity of the bio-rad platelia (bio-rad laboratories, marnes-la-coquette, france) dengue ns1 assay was 73.6% (95% confidence interval [ci], 63.7-81.6). the panbio early elisa (panbio diagnostics, ...200919232865
detection of dengue nonstructural 1 (ns1) protein in vietnamese patients with fever.diagnosing dengue in febrile patients is challenging. of a total of 459 patients with acute undifferentiated fever, randomly selected from 12 primary health facilities and 1 clinic of the provincial malaria station in southern vietnam, dengue-specific antibody (ab) and ns1ag enzyme-linked immunosorbent assay (elisa) (platelia, bio-rad laboratories, hercules, ca 94547, us) were performed on acute (t0) and convalescent (t3 weeks) sera. polymerase chain reaction (pcr) was used for confirmation. bas ...200919232866
identification of novel target sites and an inhibitor of the dengue virus e protein.dengue and related flaviviruses represent a significant global health threat. the envelope glycoprotein e mediates virus attachment to a host cell and the subsequent fusion of viral and host cell membranes. the fusion process is driven by conformational changes in the e protein and is an essential step in the virus life cycle. in this study, we analyzed the pre-fusion and post-fusion structures of the dengue virus e protein to identify potential novel sites that could bind small molecules, which ...200919241120
rna interference mediated silencing of hsp60 gene in human monocytic myeloma cell line u937 revealed decreased dengue virus multiplication.heat shock proteins (hsps) or stress proteins are highly conserved molecules and expressed in all cell types under stressful conditions like heat, cold, hypoxia and infections. the objective of the present study was to determine the effect of dengue virus infection on relative expression of stress proteins and their role in the progression of the infection. as macrophages are the primary host for dengue, human promonocytic myeloblastoma u937 cells were infected with dengue virus type 2 new guine ...200919261350
in vitro assembly of nucleocapsid-like particles from purified recombinant capsid protein of dengue-2 virus.the capsid protein is one of the three structural proteins of flaviviruses and is the building block of the nucleocapsid. it has also a predominant role in the replication of dengue virus. to obtain nucleocapsid-like particles from recombinant dengue-2 capsid protein produced in e. coli, a purification process using cation exchange chromatography was established. the purified protein exhibited a molecular mass corresponding to a dimer; therefore, similar to that reported for alphaviruses, an in ...200919305942
virus-specific read-through codon preference affects infectivity of chimeric cucumber green mottle mosaic viruses displaying a dengue virus epitope.a cucumber green mottle mosaic virus (cgmmv) was used to present a truncated dengue virus type 2 envelope (e) protein binding region from amino acids 379 to 423 (eb4). the eb4 gene was inserted at the terminal end of the cgmmv coat protein (cp) open reading frame (orf). read-through sequences of tmv or cgmmv, caa-uag-caa-uua, or aaa-uag-caa-uua were, respectively, inserted in between the cp and the eb4 genes. the chimeric clones, prt, prg, and pcg+fsrtre, were transcribed into full-length capped ...200919325913
concurrent chikungunya and dengue virus infections during simultaneous outbreaks, gabon, 2007.an outbreak of febrile illness occurred in gabon in 2007, with 20,000 suspected cases. chikungunya or dengue-2 virus infections were identified in 321 patients; 8 patients had documented co-infections. aedes albopictus was identified as the principal vector for the transmission of both viruses.200919331740
natural vertical transmission by stegomyia albopicta as dengue vector in brazil.the mosquito stegomyia albopicta is among the most important arbovirus vectors in the world, particularly for dengue viruses. their natural history suggests that biologically these viruses are highly adapted to their mosquito hosts and they were most likely mosquito viruses prior to becoming adapted to lower primates and humans. as well as being maintained by transmission among susceptible humans, dengue viruses may also be maintained by vertical transmission in mosquitoes during inter-epidemic ...200919347154
gene flow, subspecies composition, and dengue virus-2 susceptibility among aedes aegypti collections in senegal.aedes aegypti, the "yellow fever mosquito", is the primary vector to humans of the four serotypes of dengue viruses (denv1-4) and yellow fever virus (yfv) and is a known vector of chikungunya virus. there are two recognized subspecies of ae. aegypti sensu latu (s.l.): the presumed ancestral form, ae. aegypti formosus (aaf), a primarily sylvan mosquito in sub-saharan africa, and ae. aegypti aegypti (aaa), found globally in tropical and subtropical regions typically in association with humans. the ...200919365540
effects of a five-year citywide intervention program to control aedes aegypti and prevent dengue outbreaks in northern argentina.dengue has propagated widely through the americas. most countries have not been able to maintain permanent larval mosquito control programs, and the long-term effects of control actions have rarely been documented.200919399168
structure-based discovery of dengue virus protease inhibitors.dengue virus belongs to the family flaviviridae and is a major emerging pathogen for which the development of vaccines and antiviral therapy has seen little success. the ns3 viral protease is a potential target for antiviral drugs since it is required for virus replication. the goal of this study was to identify novel dengue virus (type 2; den2v) protease inhibitors for eventual development as effective anti-flaviviral drugs. the eudoc docking program was used to computationally screen a small-m ...200919428601
ns1 detection in addition to reverse transcriptase pcr and transcription-mediated amplification of dengue virus rna in acutely ill patients. 200919483079
antiviral effect of interferon lambda against west nile virus.type iii interferons (ifn), ifn-lambda or il-28/29, are new members of the ifn super-family. except for using distinct receptors, type i and type iii ifns share the same major post receptor signaling components to activate the transcription of a similar set of ifn-stimulated genes (isgs). to examine the antiviral effects of the new type ifns against west nile virus (wnv), we compared the antiviral effects of ifn-alpha and ifn-lambda on wnv virus-like particle (vlp) infection and replicon replica ...200919501257
the neovolcanic axis is a barrier to gene flow among aedes aegypti populations in mexico that differ in vector competence for dengue 2 virus.aedes aegypti is the main mosquito vector of the four serotypes of dengue virus (denv). previous population genetic and vector competence studies have demonstrated substantial genetic structure and major differences in the ability to transmit dengue viruses in ae. aegypti populations in mexico.200919564909
identification of amino acids in the dengue virus type 2 envelope glycoprotein critical to virus infectivity.the dengue virus envelope glycoprotein mediates virus attachment and entry and is the major viral antigen. the identification of 'critical' amino acids in the envelope glycoprotein that cannot be altered without loss of infectivity could have a major impact on the development of dengue virus vaccines and diagnostics. in this context, we determined whether six amino acids, previously predicted by computational analysis to play a critical role in the virus life cycle, were essential for virus viab ...200919587132
[construcion of a chimeric japanese encephalits virus/dengue virus-2].the prm/e gene of dv2 was cloned into the jev (sa14-14-2 strain) replicon vector which had been constructed previously, and the resulting recombinant plasmid was named ppartialdeltaprm/e. the constructed chimeric clone was linearized and then was transcripted into rna in vitro. the produced rna was transfected into bhk-21 cells. five to seven days later, cpe could be observed on the transfected bhk-21cells, and then the supernatant containing the chimeric virus was collected. the supernatant was ...200919634760
depletion of macrophages in mice results in higher dengue virus titers and highlights the role of macrophages for virus control.monocytes and macrophages are target cells for dengue infection. besides their potential role for virus replication, activated monocytes/macrophages produce cytokines that may be critical for dengue pathology. to study the in vivo role of monocytes and macrophages for virus replication, we depleted monocytes and macrophages in ifn-alphabetagammar knockout mice with clodronate liposomes before dengue infection. although less virus was first recovered in the draining ln in the absence of macrophag ...200919637226
essential role of platelet-activating factor receptor in the pathogenesis of dengue virus infection.severe dengue infection in humans causes a disease characterized by thrombocytopenia, increased levels of cytokines, increased vascular permeability, hemorrhage, and shock. treatment is supportive. activation of platelet-activating factor (paf) receptor (pafr) on endothelial cells and leukocytes induces increase in vascular permeability, hypotension, and production of cytokines. we hypothesized that activation of pafr could account for the major systemic manifestations of dengue infection. inocu ...200919666557
essential role of cyclophilin a for hepatitis c virus replication and virus production and possible link to polyprotein cleavage kinetics.viruses are obligate intracellular parasites and therefore their replication completely depends on host cell factors. in case of the hepatitis c virus (hcv), a positive-strand rna virus that in the majority of infections establishes persistence, cyclophilins are considered to play an important role in rna replication. subsequent to the observation that cyclosporines, known to sequester cyclophilins by direct binding, profoundly block hcv replication in cultured human hepatoma cells, conflicting ...200919680534
recombinant nucleocapsid-like particles from dengue-2 virus induce protective cd4+ and cd8+ cells against viral encephalitis in mice.virus-like particles are a highly effective type of subunit vaccine that mimics the overall structure of virus particles without containing infectious genetic material. in this work, a particulate form of the recombinant capsid protein from dengue-2 was evaluated in mice to determine the level of protection against viral challenge and to measure the antigen-induced cell-mediated immunity (cmi). the nucleocapsid-like particles (nlps) adjuvanted with alum did not induce antiviral antibodies. howev ...200919692540
cross-talk between rac1 and cdc42 gtpases regulates formation of filopodia required for dengue virus type-2 entry into hmec-1 cells.infection with dengue virus type-2 (denv-2) begins with virus adherence to cell surface receptors. in endothelial cells (hmec-1), a cell model for denv-2 infection, alpha 5 beta 3 integrin has been identified as a putative receptor for the virus. previous work had suggested that the actin cytoskeleton of hmec-1 cells plays an important role in virus entry and infection. in the present work, fixed and living hmec-1 cells expressing enhanced green fluorescent protein-actin were monitored for actin ...200919710257
viremia and antibody response in green monkeys (chlorocebus aethiops sabaeus) infected with dengue virus type 2: a potential model for vaccine testing.the increasingly limited availability and high cost of the hitherto most commonly used monkey species in dengue vaccine research has augmented the importance of identifying alternative suitable models for these studies. in this study we examined the capacity of green monkeys (chlorocebus aethiops sabaeus) to develop dengue viremia, and thus provide a potential model for dengue vaccine testing. monkeys were inoculated with two different doses of dengue virus type 2. all animals in both groups bec ...200919714858
induction of neutralization antibodies in mice by dengue-2 envelope dna vaccines.dengue (den) viruses have become a public health problem that affects approximately 100 million people worldwide each year. prevention measures rely on vector control programs, which are inefficient. therefore, a vaccine is urgently needed.200919715116
hepatic damage associated with dengue-2 virus replication in liver cells of balb/c mice.one difficulty in studying dengue virus (denv) is the lack of an experimental model that reproduces the human disease. in a previous work, we have shown that balb/c mice intraperitoneally inoculated with a denv-2 isolate presented viremia and mild focal areas of liver injuries. in this study, mice were inoculated by the intravenous route and presented extensive damage areas in the liver tissue, which were evaluated by histopathological and ultrastructural analysis. hepatic injury was noted mainl ...200919721415
viruses and autophagy.autophagy is an evolutionarily conserved intracellular process by which bulk cytoplasm is enveloped inside a double-membraned vesicle and shuttled to lysosomes for degradation. within the last 15 years, the genes necessary for the execution of autophagy have been identified and the number of tools for studying this process has grown. autophagy is essential for tissue homeostasis and development and defective autophagy is associated with a number of diseases. as intracellular parasites, during th ...019750559
stability and loss of a virus resistance phenotype over time in transgenic mosquitoes harbouring an antiviral effector gene.transgenic aedes aegypti were engineered to express a virus-derived, inverted repeat (ir) rna in the mosquito midgut to trigger rna interference (rnai) and generate resistance to dengue virus type 2 (denv2) in the vector. here we characterize genotypic and phenotypic stabilities of one line, carb77, between generations g(9) and g(17). the anti-denv2 transgene was integrated at a single site within a noncoding region of the mosquito genome. the virus resistance phenotype was strong until g(13) an ...200919754743
heterologous prime-boost strategy in non-human primates combining the infective dengue virus and a recombinant protein in a formulation suitable for human use.the aim of the present work was to test the concept of the heterologous prime-boost strategy combining an infective dengue virus with a recombinant chimeric protein carrying domain iii of the envelope protein.201019773190
viremia and the magnitude of the immune response upon infection of green monkeys with dengue virus type 2 are strain-dependent.testing in non-human primates is a generally accepted necessary step preceding the evaluation of dengue vaccine candidates in humans. a reduction of viremia in these animals after virus challenge is generally used as an indicator of vaccine efficacy. in this work, we compared the infectivity of three strains of dengue virus type 2 in a non-human primate model of dengue infection, with the aim of selecting a virus for vaccine protection studies. as a result, strain sb8553 produced the longest dur ...200919784699
[study on e protein epitopes and primary identification of main yellow virus].to analysis the e protein epitopes of dengue virus type 1-4, japanese encephalitis virus and yellow fever virus and to distinguish the shared or specific epitopes among them.200919799147
anti-dengue virus nonstructural protein 1 antibodies recognize protein disulfide isomerase on platelets and inhibit platelet aggregation.hemorrhagic syndrome is a hallmark of severe dengue diseases. we previously suggested a mechanism of molecular mimicry in which antibodies against dengue virus (dv) nonstructural protein 1 (ns1) cross-react with platelets. in the present study, we demonstrate that protein disulfide isomerase (pdi) on the platelet surface is recognized by anti-dv ns1 antibodies. anti-dv ns1 obtained from hyperimmunized mouse sera inhibited pdi activity and platelet aggregation, and both inhibitory effects were pr ...200919822367
phase 1 trial of the dengue virus type 4 vaccine candidate rden4{delta}30-4995 in healthy adult volunteers.rden4delta30-4995 is a live attenuated dengue virus type 4 (denv4) vaccine candidate specifically designed as a further attenuated derivative of the rden4delta30 parent virus. in a previous study, 5 of 20 vaccinees who received 10(5) plaque-forming units (pfu) of rden4delta30 developed a transient elevation of the serum alanine aminotransferase (alt) level and an asymptomatic maculopapular rash developed in 10 of 20. in the current study, 28 healthy adult volunteers were randomized to receive 10 ...200919861619
comprehensive analyses and characterization of haemophagocytic lymphohistiocytosis in vietnamese children.haemophagocytic lymphohistiocytosis (hlh) is a fatal haematological disorder with diverse aetiology. this prospective study was undertaken to characterize hlh cases in vietnamese children. clinical and laboratory data, genetic analyses and outcome of the hlh patients were analysed. a total of 33 patients were enrolled from march 2007 to december 2008, with a median age of 3 years. mutations of the sh2d1a (sap) and prf1 genes were detected in one patient, respectively. the virus association was h ...201019863536
induction of type i interferon by rna viruses: cellular receptors and their substrates.virus recognition and induction of interferon (ifn) are critical components of the innate immune system. the toll-like receptor (tlr) and rig-i-like receptor families have been characterized as key players in rna virus detection. signaling cascades initiated by these receptors are crucial for establishment of an ifn signaling mediated antiviral state in infected and neighboring cells and containment of virus replication as well as initiation of the adaptive immune response. in this review, we fo ...201019882216
recombinant dengue virus-like particles from pichia pastoris: efficient production and immunological properties.the envelope glycoprotein (e) of flavivirus is the major structural protein on the surface of the mature virions. the complexes of premembrane (prm) and e play important roles in virus assembly and fusion modulation and in potential immunity-inducing vaccines. in the present study, the cdna encoding prm and e proteins of dengue virus type 2 (denv-2) was subcloned into the pgapzalphaa vector and further integrated into the genome of pichia pastoris under the control of the glyceraldehyde-3-phosph ...201019885726
bovine viral diarrhea virus ns4b protein is an integral membrane protein associated with golgi markers and rearranged host membranes.very little is known about bvdv ns4b, a protein of approximately 38 kda. however, a missense mutation in ns4b has been implicated in changing bvdv from a cytopathic to noncytopathic virus, suggesting that ns4b might play a role in bvdv pathogenesis. though this is one possible function, it is also likely that ns4b plays a role in bvdv genome replication. for example, bvdv ns4b interacts with ns3 and ns5a, implying that ns4b is part of a complex, which contains bvdv replicase proteins. other poss ...200919887001
the dengue virus type 2 envelope protein fusion peptide is essential for membrane fusion.the flaviviral envelope (e) protein directs virus-mediated membrane fusion. to investigate membrane fusion as a requirement for virus growth, we introduced 27 unique mutations into the fusion peptide of an infectious cdna clone of dengue 2 virus and recovered seven stable mutant viruses. the fusion efficiency of the mutants was impaired, demonstrating for the first time the requirement for specific fp aas in optimal fusion. mutant viruses exhibited different growth kinetics and/or genetic stabil ...201019913272
analysis of rickettsia typhi-infected and uninfected cat flea (ctenocephalides felis) midgut cdna libraries: deciphering molecular pathways involved in host response to r. typhi infection.murine typhus is a flea-borne febrile illness that is caused by the obligate intracellular bacterium, rickettsia typhi. the cat flea, ctenocephalides felis, acquires r. typhi by imbibing a bloodmeal from a rickettsemic vertebrate host. to explore which transcripts are expressed in the midgut in response to challenge with r. typhi, cdna libraries of r. typhi-infected and uninfected midguts of c. felis were constructed. in this study, we examined midgut transcript levels for select c. felis serine ...201020017753
in vitro antiviral activity of neem (azardirachta indica l.) bark extract against herpes simplex virus type-1 infection.herpes simplex virus type 1 (hsv-1) causes significant health problems from periodical skin and corneal lesions to encephalitis. we report here that an aqueous extract preparation from the barks of neem plant azardirachta indica acts as a potent entry inhibitor against hsv-1 infection into natural target cells. the neem bark extract (nbe) significantly blocked hsv-1 entry into cells at concentrations ranging from 50 to 100 microg/ml. the blocking activity of nbe was observed when the extract was ...201020041417
monoclonal antibody to dengue capsid protein: its application in dengue studies.dengue fever (df) and dengue hemorrhagic fever/dengue shock syndrome (dhf/dss) are considered the most important arthropod-borne viral diseases in terms of morbidity and mortality. the emergency and severity of dengue (den) infections increase the necessity of an early, quick and effective dengue laboratory diagnostic. viral isolation is considered a gold standard for diagnosis of dengue infection using monoclonal antibodies (mabs) as a tool for determining serotype specificity. alternatives hav ...200920061827
autophagy in viral replication and pathogenesis.autophagy is a catabolic process that is important for the removal of damaged organelles and long-lived proteins for the maintenance of cellular homeostasis. it can also serve as innate immunity to remove intracellular microbial pathogens. a growing list of viruses has been shown to affect this cellular pathway. some viruses suppress this pathway for their survival, while others enhance or exploit this pathway to benefit their replication. the effect of viruses on autophagy may also sensitize ce ...201020077024
the toll immune signaling pathway control conserved anti-dengue defenses across diverse ae. aegypti strains and against multiple dengue virus serotypes.dengue virus has become one of the most important arboviral pathogens affecting the world today. the virus is transmitted among humans by the mosquitoes aedes aegypti and ae. albopictus. like other vector-borne pathogens, this virus encounters innate immune defenses within the mosquito vector that limit infection. we have previously demonstrated the involvement of the toll pathway in the anti-dengue defense at 7 days after infection. in the present study, we have investigated the activity of thi ...201020079370
differential regulation of toll-like receptor-2, toll-like receptor-4, cd16 and human leucocyte antigen-dr on peripheral blood monocytes during mild and severe dengue fever.dengue fever (df), a public health problem in tropical countries, may present severe clinical manifestations as result of increased vascular permeability and coagulation disorders. dengue virus (denv), detected in peripheral monocytes during acute disease and in in vitro infection, leads to cytokine production, indicating that virus-target cell interactions are relevant to pathogenesis. here we investigated the in vitro and in vivo activation of human peripheral monocytes after denv infection. t ...201020113369
[identification and sequence analysis of e gene of dengue virus type 2 strain isolated from patient serum in shenzhen].to isolate and identify the pathogen of dengue fever from shenzhen city in 2005 - 2006, and to analyze the molecular characteristics of the isolated dengue virus strain as well as to explore its possible origin.200920137564
efficient, trans-complementing packaging systems for chimeric, pseudoinfectious dengue 2/yellow fever viruses.in our previous studies, we have stated to build a new strategy for developing defective, pseudoinfectious flaviviruses (pivs) and applying them as a new type of vaccine candidates. pivs combined the efficiency of live vaccines with the safety of inactivated or subunit vaccines. the results of the present work demonstrate further development of chimeric pivs encoding dengue virus 2 (den2v) glycoproteins and yellow fever virus (yfv)-derived replicative machinery as potential vaccine candidates. t ...201020137799
innate antiviral immunity in drosophila.the study of drosophila, and other genetically tractable insects, has expanded our understanding of innate immunity and more recently antiviral innate mechanisms. the drosophila antiviral program includes inflammatory signaling cascades as well as antiviral rna silencing and autophagy. this review will highlight the recent discoveries in antiviral immunity in insects and will reveal some of the lessons learned.020137906
establishment and characterization of dengue virus type 2 nonstructural protein 1 specific t cell lines.immunity against dengue viruses (denv) infection may include cellular immune responses which involve in the immunopathology of denv infection hosts. this study was to establish short-term dengue virus type 2 (denv2) nonstructural protein 1 (ns1) specific t cells from splenocytes from balb/c mice immunized with denv2 ns1 in vitro, which may be used to identify immunopathologic mechanism of dengue. nine denv2 ns1 specific t cell lines were successfully established by using limiting dilution method ...201020138364
pt(ii) and ag(i) complexes with acesulfame: crystal structure and a study of their antitumoral, antimicrobial and antiviral activities.two new complexes of platinum(ii) and silver(i) with acesulfame were synthesized. acesulfame is in the anionic form acesulfamate (ace). the structures of both complexes were determined by x-ray crystallography. for k(2)[ptcl(2)(ace)(2)] the platinum atom is coordinated to two cl(-) and two n-acesulfamate atoms forming a trans-square planar geometry. each k(+) ion interacts with two oxygen atoms of the s(o)(2) group of each acesulfamate. for the polymeric complex [ag(ace)](n) the water molecule b ...201020149461
isolation and characterization of selective and potent human fab inhibitors directed to the active-site region of the two-component ns2b-ns3 proteinase of west nile virus.there is a need to develop inhibitors of mosquito-borne flaviviruses, including wnv (west nile virus). in the present paper, we describe a novel and efficient recombinant-antibody technology that led us to the isolation of inhibitory high-affinity human antibodies to the active-site region of a viral proteinase. as a proof-of-principal, we have successfully used this technology and the synthetic naive human combinatorial antibody library hucal gold(r) to isolate selective and potent function-blo ...201020156198
six rna viruses and forty-one hosts: viral small rnas and modulation of small rna repertoires in vertebrate and invertebrate systems.we have used multiplexed high-throughput sequencing to characterize changes in small rna populations that occur during viral infection in animal cells. small rna-based mechanisms such as rna interference (rnai) have been shown in plant and invertebrate systems to play a key role in host responses to viral infection. although homologs of the key rnai effector pathways are present in mammalian cells, and can launch an rnai-mediated degradation of experimentally targeted mrnas, any role for such re ...201020169186
the length of and nonhydrophobic residues in the transmembrane domain of dengue virus envelope protein are critical for its retention and assembly in the endoplasmic reticulum.the morphogenesis of many enveloped viruses, in which viral nucleocapsid complex interacts with envelope (e) protein, is known to take place at various sites along the secretory pathway. how viral e protein retains in a particular intracellular organelle for assembly remains incompletely understood. in this study, we investigated determinants in the e protein of dengue virus (denv) for its retention and assembly in the endoplasmic reticulum (er). a chimeric experiment between cd4 and denv precur ...201020181718
two lineages of dengue virus type 2, brazil. 201020202456
pichia pastoris-expressed dengue virus type 2 envelope domain iii elicits virus-neutralizing antibodies.a tetravalent dengue vaccine that can protect against all four serotypes of dengue viruses is a global priority. the host-receptor binding, multiple neutralizing epitope-containing carboxy-terminal region of the dengue envelope protein, known as domain iii (ediii), has emerged as a promising subunit vaccine antigen. one strategy to develop a tetravalent dengue subunit vaccine envisages mixing recombinant ediiis, corresponding to the four dengue virus serotypes. towards this objective, a recombin ...201020211204
selection and identification of b-cell epitope on ns1 protein of dengue virus type 2.ns1 of dengue virus (denv) is an important non-structural protein, which plays an important role in denv replication and dengue infection. in this study, using the phage-displayed peptide library screening method and purified anti-denv2-ns1 polyclonal antibody immunoglobulin g (igg) as target, which was generated from the purified recombinant expressed denv2-ns1 protein immunization on rabbit, seven b-cell epitopes of denv2-ns1 protein were screened. considering the results of comprehensive bioi ...201020211209
study on the determinants of suckling mice neurovirulence of dengue 2 virus.pdvws501 was a genomic-length cdna clone of dengue 2 virus, through which infectious virus (mon501) could be rescued. mon501 was neurovirulent in mice, whose e residues 62 and 203 were lys and asn, respectively. two genomic-length cdna clones (tb62 and tb203) were constructed by pointed mutation of pdvws501 with ol-pcr, e62 of tb62 and e203 of tb203 were converted to glu and asp, respectively. rna transcripts of pdvws501, tb62 and tb203 were produced in vitro and electroporated into bhk-21 cells ...200320213366
exploring different strategies to express dengue virus envelope protein in a plant system.dengue virus envelope glycoprotein (e-protein) is the main protein associated with immunity induction. to produce a candidate for subunit vaccines and to provide an antigen for diagnostic kits, it was expressed in a novel plant system using deconstructed viral modules. a truncated version of the e-protein was designed to be expressed alone and co-expressed with dengue virus structural proteins. as well, the critical domain iii of e-protein was fused to hepatitis b core antigen (hbcore). the reco ...201020213522
replication of not-known-vector flaviviruses in mosquito cells is restricted by intracellular host factors rather than by the viral envelope proteins.chimeric yellow fever virus 17d (yfv-17d) and dengue virus type 2 (denv2) carrying the surface proteins of modoc virus (modv), a not-known-vector (nkv) flavivirus, replicated efficiently in mammalian (vero-b) and mosquito (c6/36) cells, whereas modv failed to replicate in mosquito cells. transfection of c6/36 cells with modv rna did not result in virus replication; however, transfection of these mosquito cells with yfv-17d or denv2 rna did. the inability of nkv viruses (such as modv) to infect a ...201020219898
both e protein glycans adversely affect dengue virus infectivity but are beneficial for virion release.the e protein of most flaviviruses is modified by asn-linked glycosylation at residue 153/154 and in the case of the four dengue virus (denv) serotypes by a second glycan at residue 67. however, the absence of e protein glycosylation among numerous natural isolates of different flaviviruses suggests that the glycan, per se, is not critically important in the virus life cycle. consistent with this notion, we show that ablation of both glycans from the denv-2 e protein reduces but does not prevent ...201020219924
nss encoded by groundnut bud necrosis virus is a bifunctional enzyme.groundnut bud necrosis virus (gbnv), a member of genus tospovirus in the family bunyaviridae, infects a large number of leguminosae and solanaceae plants in india. with a view to elucidate the function of nonstructural protein, nss encoded by the small rna genome (s rna), the nss protein of gbnv- tomato (karnataka) was over-expressed in e. coli and purified by ni-nta chromatography. the purified rnss protein exhibited an rna stimulated ntpase activity. further, this activity was metal ion depend ...201020305786
dengue virus replicons: production of an interserotypic chimera and cell lines from different species, and establishment of a cell-based fluorescent assay to screen inhibitors, validated by the evaluation of ribavirin's activity.the prevention and treatment of flavivirus infections are public health priorities. dengue fever is the most prevalent mosquito-borne viral disease of humans, affecting more than 50 million people annually. despite the urgent need to control dengue infections, neither specific antiviral therapies nor licensed vaccines exist and the molecular basis of dengue pathogenesis is not well understood. in this study we produced a novel dengue virus type 2 (dv2) subgenomic replicon that expresses a fusion ...201020307577
antagonism of the complement component c4 by flavivirus nonstructural protein ns1.the complement system plays an essential protective role in the initial defense against many microorganisms. flavivirus ns1 is a secreted nonstructural glycoprotein that accumulates in blood, is displayed on the surface of infected cells, and has been hypothesized to have immune evasion functions. herein, we demonstrate that dengue virus (denv), west nile virus (wnv), and yellow fever virus (yfv) ns1 attenuate classical and lectin pathway activation by directly interacting with c4. binding of ns ...201020308361
characterization of recombinant dengue-2 virus derived from a single nucleotide substitution in the 5' noncoding region.variants of wild-type dengue serotype 2 (den-2) virus containing nucleotide substitutions at positions 14, 15, or 57 in the 5' ncr were constructed by pcr-mediated site-directed mutagenesis. all three viruses containing a single point substitution demonstrated attenuation phenotype as evidenced by decreases replication and plaque size in cell culture assay. all three variants were less neurovirulent in newborn mice compared to the wild type. the mutants were immunogenic in adult mice immunogenic ...201020339476
reemergence of dengue in mauritius.dengue reemerged in mauritius in 2009 after an absence of >30 years, and >200 cases were confirmed serologically. molecular studies showed that the outbreak was caused by dengue virus type 2. phylogenetic analysis of the envelope gene identified 2 clades of the virus. no case of hemorrhagic fever was recorded.201020350397
a plasmid encoding parts of the dengue virus e and ns1 proteins induces an immune response in a mouse model.a denv-2 plasmid named peii*eiii/ns1*,containing sequences encoding portions of the envelope protein that are potentially involved in the induction of neutralizing antibodies and a portion of the ns1 sequence that is involved in protection, is reported in this work. the synthesized subunit protein was recognized by human sera from infected patients and had the predicted size. the immunogenicity of this construct was evaluated using a mouse model in a prime-boost vaccination approach. the priming ...201020390312
refolded dengue virus type 2 ns1 protein expressed in escherichia coli preserves structural and immunological properties of the native protein.the dengue virus ns1 protein has been shown to be a protective antigen under different experimental conditions but the recombinant protein produced in bacterial expression systems is usually not soluble and loses structural and immunological features of the native viral protein. in the present study, experimental conditions leading to purification and refolding of the recombinant dengue virus type 2 (denv-2) ns1 protein expressed in escherichia coli are described. the refolded recombinant protei ...201020399232
dengue ns1 antigen detection: a useful tool in early diagnosis of dengue virus infection.this study was undertaken to evaluate the efficacy of ns1 antigen (ag) assay as an early marker for dengue virus (dv) infection.201020404453
a functional abca1 gene variant is associated with low hdl-cholesterol levels and shows evidence of positive selection in native americans.it has been suggested that the higher susceptibility of hispanics to metabolic disease is related to their native american heritage. a frequent cholesterol transporter abca1 (atp-binding cassette transporter a1) gene variant (r230c, rs9282541) apparently exclusive to native american individuals was associated with low high-density lipoprotein cholesterol (hdl-c) levels, obesity and type 2 diabetes in mexican mestizos. we performed a more extensive analysis of this variant in 4405 native american ...201020418488
wss45, a sulfated alpha-d-glucan, strongly interferes with dengue 2 virus infection in vitro.to investigate the mode of action of wss45, one sulfated derivative of an alpha-d-glucan from the gastrodia elata bl, on the multiplication cycle of dengue virus serotype 2 (dv2), including initial infection and intracellular replication.201020418898
high-throughput procedure for tick surveys of tick-borne encephalitis virus and its application in a national surveillance study in switzerland.tick-borne encephalitis (tbe), a viral infection of the central nervous system, is endemic in many eurasian countries. in switzerland, tbe risk areas have been characterized by geographic mapping of clinical cases. since mass vaccination should significantly decrease the number of tbe cases, alternative methods for exposure risk assessment are required. we established a new pcr-based test for the detection of tbe virus (tbev) in ticks. the protocol involves an automated, high-throughput nucleic ...201020453126
the envelope glycoprotein domain iii of dengue virus type 2 induced the expression of anticoagulant molecules in endothelial cells.dengue virus (dv) causes a non-specific febrile illness known as dengue fever (df), and a severe life-threatening illness, dengue hemorrhagic fever/dengue shock syndrome (dhf/dss). hemostatic changes induced by this virus involve three main factors: thrombocytopenia, endothelial cell damage, and significant abnormalities of the coagulation and fibrinolysis systems. the pathogenesis of bleeding in dv infections remains unknown. in this article, we focused on the dv activating endothelial cells an ...201020526654
optimization of conditions for secretion of dengue virus type 2 envelope domain iii using pichia pastoris.we have developed a recombinant clone of the methylotrophic yeast pichia pastoris capable of secreting dengue virus type 2 envelope domain iii (sediii-2). we explored various induction parameters including media composition, temperature, ph, and methanol concentration, to optimize conditions for sediii-2 expression in shake flask culture. induction at 20°c in the presence of 2% (v/v) methanol in a medium buffered to ph 5.8 resulted in highest secretion of sediii-2. this yield could be further en ...201020538539
antiviral actions of flavanoid-derived compounds on dengue virus type-2.dengue viruses, mosquito-borne members of the flaviviridae family, are the causative agents of dengue fever and its associated complications, dengue haemorrhagic fever and dengue shock syndrome. to date, more than 2.5 billion people in over 100 countries are at risk of infection, and approximately 20 million infections were reported annually. there is currently no treatment or vaccine available for dengue infection. this study employed a whole-cell organism model or in vitro methods to study the ...201020567498
dengue-2-virus-interacting polypeptides involved in mosquito cell infection.for the design of effective antiviral strategies, understanding the fundamental steps of the virus life cycle, including virus-host interactions, is essential. we performed a virus overlay protein binding assay followed by proteomics for identification of proteins from membrane fractions of a7 (aedes aegypti) cells, c6/36 (aedes albopictus) cells and the midgut brush border membrane fraction of ae. aegypti mosquito that bind to dengue-2 virus. actin, atp synthase β subunit, hsc 70, orisis, prohi ...201020571839
differential gene expression from midguts of refractory and susceptible lines of the mosquito, aedes aegypti, infected with dengue-2 virus.suppressive subtractive hybridization was used to evaluate the differential expression of midgut genes of feral populations of aedes aegypti (diptera: culicidae) from colombia that are naturally refractory or susceptible to dengue-2 virus infection. a total of 165 differentially expressed sequence tags (ests) were identified in the subtracted libraries. the analysis showed a higher number of differentially expressed genes in the susceptible ae. aegypti individuals than the refractory mosquitoes. ...201020572793
tnf-alpha-dependent regulation of cxcr3 expression modulates neuronal survival during west nile virus encephalitis.the chemokine cxcl10 exerts antiviral effects within the central nervous system (cns) through the recruitment of virus-specific t cells. however, elevated levels of cxcl10 may induce neuronal apoptosis given its receptor, cxcr3, is expressed by neurons. using a murine model of west nile virus (wnv) encephalitis, we determined that wnv-infected neurons express tnf-alpha, which down-regulates neuronal cxcr3 expression via signaling through tnfr1. down-regulation of neuronal cxcr3 decreased cxcl10- ...201020579746
structural optimization and de novo design of dengue virus entry inhibitory peptides.viral fusogenic envelope proteins are important targets for the development of inhibitors of viral entry. we report an approach for the computational design of peptide inhibitors of the dengue 2 virus (denv-2) envelope (e) protein using high-resolution structural data from a pre-entry dimeric form of the protein. by using predictive strategies together with computational optimization of binding "pseudoenergies", we were able to design multiple peptide sequences that showed low micromolar viral e ...201020582308
structure and function analysis of therapeutic monoclonal antibodies against dengue virus type 2.dengue virus (denv) is the most prevalent insect-transmitted viral disease in humans globally, and currently no specific therapy or vaccine is available. protection against denv and other related flaviviruses is associated with the development of antibodies against the viral envelope (e) protein. although prior studies have characterized the neutralizing activity of monoclonal antibodies (mabs) against denv type 2 (denv-2), none have compared simultaneously the inhibitory activity against a gene ...201020592088
sequence-function analysis of the sendai virus l protein domain vi.the large (about 2200 amino acids) l polymerase protein of nonsegmented negative-strand rna viruses (order mononegavirales) has six conserved sequence regions ("domains") postulated to constitute the specific enzymatic activities involved in viral mrna synthesis, 5'-end capping, cap methylation, 3' polyadenylation, and genomic rna replication. previous studies with vesicular stomatitis virus identified amino acid residues within the l protein domain vi required for mrna cap methylation. in our r ...201020609457
cytoplasmic viral replication complexes.many viruses that replicate in the cytoplasm compartmentalize their genome replication and transcription in organelle-like structures that enhance replication efficiency and protection from host defenses. in particular, recent studies with diverse positive-strand rna viruses have further elucidated the ultrastructure of membrane-bound rna replication complexes and how these complexes function in close coordination with virion assembly and budding. the structure, function, and assembly of some po ...020638644
nucleocapsid-like particles of dengue-2 virus enhance the immune response against a recombinant protein of dengue-4 virus.in this study, we evaluate in mice a novel formulation containing nucleocapsid-like particles of dengue-2 virus (recnlp) co-immunized with a chimeric protein composed of the dengue-4 envelope domain iii fused twice within the meningococcal p64k protein of neisseria meningitidis (pd24). the animals receiving the pd24-recnlp mixture showed the highest levels of antiviral antibodies. similar results were obtained for ifnγ secretion levels, indicating a functional th1 cellular response. consistently ...201020640909
vipr: a probabilistic algorithm for analysis of microbial detection microarrays.all infectious disease oriented clinical diagnostic assays in use today focus on detecting the presence of a single, well defined target agent or a set of agents. in recent years, microarray-based diagnostics have been developed that greatly facilitate the highly parallel detection of multiple microbes that may be present in a given clinical specimen. while several algorithms have been described for interpretation of diagnostic microarrays, none of the existing approaches is capable of incorpora ...201020646301
natural attenuation of dengue virus type-2 after a series of island outbreaks: a retrospective phylogenetic study of events in the south pacific three decades ago.dengue is an expanding arboviral disease of variable severity characterized by the emergence of virus strains with greater fitness, epidemic potential and possibly virulence. to investigate the role of dengue virus (denv) strain variation on epidemic activity we studied denv-2 viruses from a series of south pacific islands experiencing outbreaks of varying intensity and clinical severity. initially appearing in 1971 in tahiti and fiji, the virus was responsible for subsequent epidemics in americ ...201020663532
evaluation of extracellular subviral particles of dengue virus type 2 and japanese encephalitis virus produced by spodoptera frugiperda cells for use as vaccine and diagnostic antigens.new or improved vaccines against dengue virus types 1 to 4 (denv1 to denv4) and japanese encephalitis virus (jev), the causative agents of dengue fever and japanese encephalitis (je), respectively, are urgently required. the use of noninfectious subviral extracellular particles (eps) is an inexpensive and safe strategy for the production of protein-based flavivirus vaccines. although coexpression of premembrane (prm) and envelope (e) proteins has been demonstrated to produce eps in mammalian cel ...201020668137
a balb/c mouse model shows that liver involvement in dengue disease is immune-mediated.in the present study, balb/c mice were used to develop a model for the hepatic injury associated to dengue infection. histological analysis after subcutaneous inoculation with a low viral dose of dengue-2 virus showed kupffer cell hyperplasia and an increased inflammatory cellular infiltrate next to the bile ducts on days 5, 7 and 14 post-inoculation, mainly characterized by the presence of mononuclear cells. the liver mrna transcription level of il-1β was highest on the 5th day post-infection ( ...201020673760
characterization of dengue virus type 2: new insights on the 2010 brazilian epidemic.dengue viruses (denv) serotypes 1, 2, and 3 have been causing yearly outbreaks in brazil. in this study, we report the re-introduction of denv2 in the coast of são paulo state. partial envelope viral genes were sequenced from eighteen patients with dengue fever during the 2010 epidemic. phylogenetic analysis showed this strain belongs to the american/asian genotype and was closely related to the virus that circulated in rio de janeiro in 2007 and 2008. the phylogeny also showed no clustering by ...201020676363
morphological changes in vero cells postinfection with dengue virus type-2.although no specific antiviral tablets or injections that can kill the dengue virus are currently available, adequate care and treatment could control its morbidity. interaction of dengue virus to target cells could be an important feature for virus propagation. ultrastructural analysis of this interaction was studied with vero cells. vero cells were treated with dengue virus type-2 at different time intervals at multiplicity of infection (m.o.i) < 10, m.o.i > 10, and m.o.i = 100. it was found t ...201020687131
domain-iii fg loop of the dengue virus type 2 envelope protein is important for infection of mammalian cells and aedes aegypti mosquitoes.the fg extended loop in domain iii of the dengue virus type 2 (denv2) envelope protein is postulated to be a molecular determinant for host cell infectivity. to determine the contribution of the fg loop to virus infectivity, an infectious cdna clone of denv2 was manipulated by deleting amino acids in the loop (vepgδ) to mimic tick-borne flaviviruses or by substituting these aas with rgd or rgdk/s to mimic motifs present in other mosquito-borne flaviviruses. we found the fg loop to be dispensable ...201020708768
transgene-mediated suppression of dengue viruses in the salivary glands of the yellow fever mosquito, aedes aegypti.controlled sex-, stage- and tissue-specific expression of antipathogen effector molecules is important for genetic engineering strategies to control mosquito-borne diseases. adult female salivary glands are involved in pathogen transmission to human hosts and are target sites for expression of antipathogen effector molecules. the aedes aegypti 30k a and 30k b genes are expressed exclusively in adult female salivary glands and are transcribed divergently from start sites separated by 263 nucleoti ...201020738425
index cluster study of dengue virus infection in nicaragua.traditional study designs do not identify acute asymptomatic or pre-symptomatic dengue virus (denv) infections, thus limiting our understanding of immunologic and viral factors that modulate infection outcome. in the 2006 and 2007 dengue seasons, we conducted a pilot index cluster study in managua, nicaragua, in which 442 persons living within 50 meters of 22 index cases identified through an ongoing pediatric cohort study were evaluated for denv infection. post-enrollment and pre-enrollment den ...201020810839
quantitative analysis of replication and tropisms of dengue virus type 2 in aedes albopictus.dengue virus serotype 2 (denv-2) rna replication profiles and tropisms were studied by using quantitative rt-pcr (q-rtpcr) in intrathoracically infected aedes albopictus. the virus rna replication profiles were diverse in mosquito organs. in fat body, brain, salivary gland, and malpighian tubes, it peaked at 8, 23, 23, and 27 days post-infection, respectively, and then, all declined. in midgut, it increased all the time and had no trend of decline. in ovary, it had no apparent increase. subseque ...201020810842
upregulation of a novel eukaryotic translation initiation factor 5a (eif5a) in dengue 2 virus-infected mosquito cells.dengue virus, a mosquito-borne flavivirus, is the etiological agent of dengue fever, dengue hemorrhagic fever, and dengue shock syndrome. it generally induces apoptosis in mammalian cells, but frequently results in persistent infection in mosquito cells. that mechanism remains to be explored. in turn, a genomic survey through subtractive hybridization (pcr-select cdna subtraction) was conducted in order to find gene(s) that may play a role in interactions between the virus and its host cells.201020819232
roles of small gtpase rac1 in the regulation of actin cytoskeleton during dengue virus infection.increased vascular permeability is a hallmark feature in severe dengue virus (dv) infection, and dysfunction of endothelial cells has been speculated to contribute in the pathogenesis of dengue hemorrhagic fever/dengue shock syndrome (dhf/dss). rho-family gtpase rac1 is a significant element of endothelial barrier function regulation and has been implicated in the regulation of actin remodeling and intercellular junction formation. yet there is little evidence linking rac1 gtpase to alteration i ...201020824170
the human immune response to dengue virus is dominated by highly cross-reactive antibodies endowed with neutralizing and enhancing activity.antibodies protect against homologous dengue virus (denv) infection but can precipitate severe dengue by promoting heterotypic virus entry via fcγ receptors (fcγr). we immortalized memory b cells from individuals after primary or secondary infection and analyzed anti-denv monoclonal antibodies (mabs) thus generated. mabs to envelope (e) protein domain iii (diii) were either serotype specific or cross-reactive and potently neutralized denv infection. di/dii- or viral membrane protein prm-reactive ...201020833378
virucidal activity and chemical composition of essential oils from aromatic plants of central west argentina.the essential oils of seven aromatic plants from central west argentina were isolated by steam distillation and analyzed by a gas chromatography/mass spectrometry technique. the oils were screened for cytotoxicity and in vitro inhibitory activity against herpes simplex virus type 1 (hsv-1), dengue virus type 2 (denv-2) and junin virus (junv). the oils showed a variable virucidal action according to the virus. junv was the least susceptible virus in comparison with hsv-1 and denv-2. the better re ...201020839642
molecular mechanisms involved in the early steps of flavivirus cell entry.flaviviruses enter their host cells by receptor-mediated endocytosis, a well-orchestrated process of receptor recognition, penetration and uncoating. recent findings on these early steps in the life cycle of flaviviruses are the focus of this review.201020869460
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