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surface expression of influenza virus neuraminidase, an amino-terminally anchored viral membrane glycoprotein, in polarized epithelial cells.we have investigated the site of surface expression of the neuraminidase (na) glycoprotein of influenza a virus, which, in contrast to the hemagglutinin, is bound to membranes by hydrophobic residues near the nh2-terminus. madin-darby canine kidney or primary african green monkey kidney cells infected with influenza a/wsn/33 virus and subsequently labeled with monoclonal antibody to the na and then with a colloidal gold- or ferritin-conjugated second antibody exhibited specific labeling of apica ...19853016520
the influenza virus nucleoprotein synthesized from cloned dna in a simian virus 40 vector is detected in the nucleus.we obtained dna sequences coding for the nucleoprotein (np) of an influenza a virus by reverse transcription of virion rna with synthetic oligonucleotide primers. terminal sequence analysis showed that the cloned gene contained a full-length copy of the virion rna segment. the np-specific dna was inserted into the late region of a simian virus 40 vector, and the dna recombinant was propagated in the presence of an early simian virus 40 temperature-sensitive mutant helper. infection of african gr ...19836296449
impairment of multicycle influenza virus growth in vero (who) cells by loss of trypsin activity.we demonstrated that influenza virus replication in vero (who) cells, a subline of african green monkey kidney cells, is impaired by rapid inactivation of trypsin in the culture fluids. trypsin inactivation was caused by a factor secreted by vero cells into the media. repeated addition of trypsin to the culture medium of influenza virus-infected vero cells restores the multicycle growth pattern of influenza a virus strains, allowing high yields to be obtained at a low multiplicity of infection. ...19957884927
african green monkey kidney (vero) cells provide an alternative host cell system for influenza a and b viruses.the preparation of live, attenuated human influenza virus vaccines and of large quantities of inactivated vaccines after the emergence or reemergence of a pandemic influenza virus will require an alternative host cell system, because embryonated chicken eggs will likely be insufficient and suboptimal. preliminary studies indicated that an african green monkey kidney cell line (vero) is a suitable system for the primary isolation and cultivation of influenza a viruses (e. a. govorkova, n. v. kave ...19968764064
a host restriction-based selection system for influenza haemagglutinin transfectant viruses.during the 1996 influenza epidemic in vienna we obtained influenza a virus specimens (vienna/47/96, vienna/81/96) which grow efficiently in african green monkey kidney (vero) cells but not in embryonated chicken eggs. amplification of the specimens in vero cells resulted in progeny that agglutinated human but not chicken erythrocytes. reassortment analysis suggested that the haemagglutinin (ha) might be responsible for the host restriction. vero cells were infected with the vienna/47/96 virus an ...19989634082
generation of high-yielding influenza a viruses in african green monkey kidney (vero) cells by reverse genetics.influenza a viruses are the cause of annual epidemics of human disease with occasional outbreaks of pandemic proportions. the zoonotic nature of the disease and the vast viral reservoirs in the aquatic birds of the world mean that influenza will not easily be eradicated and that vaccines will continue to be needed. recent technological advances in reverse genetics methods and limitations of the conventional production of vaccines by using eggs have led to a push to develop cell-based strategies ...200414747549
live cold-adapted influenza a vaccine produced in vero cell line.the african green monkey kidney (vero) cell line was used as a substrate for the development of a live cold-adapted (ca) reassortant influenza vaccine. for that purpose, a new master strain was generated by an adaptation of the wild type (wt) a/singapore/1/57 virus to growth at 25 degrees c in a vero cell line. the resulting cold-adapted (ca) muster strain a/singapore/1/57ca showed temperature sensitive (ts) phenotype and was attenuated in animal models and protective in the challenge experiment ...200415163508
immunization of primates with a newcastle disease virus-vectored vaccine via the respiratory tract induces a high titer of serum neutralizing antibodies against highly pathogenic avian influenza virus.the ongoing outbreak of highly pathogenic avian influenza virus (hpaiv) in birds, the incidence of transmission to humans with a resulting high mortality rate, and the possibility of a human pandemic warrant the development of effective human vaccines against hpaiv. we developed an experimental live-attenuated vaccine for direct inoculation of the respiratory tract based on recombinant avian newcastle disease virus (ndv) expressing the hemagglutinin (ha) glycoprotein of h5n1 hpaiv (ndv-ha). expr ...200717715243
establishment of canine rna polymerase i-driven reverse genetics for influenza a virus: its application for h5n1 vaccine production.in the event of a new influenza pandemic, vaccines whose antigenicities match those of circulating strains must be rapidly produced. here, we established an alternative reverse genetics system for influenza virus using the canine polymerase i (poli) promoter sequence that works efficiently in the madin-darby canine kidney cell line, a cell line approved for human vaccine production. using this system, we were able to generate h5n1 vaccine seed viruses more efficiently than can be achieved with t ...200818045936
high titer growth of human and avian influenza viruses in an immortalized chick embryo cell line without the need for exogenous proteases.the current method of growing influenza virus for vaccine production is through the use of embryonated chicken eggs. this manufacturing system yields a low concentration of virus per egg, requires significant downstream production for purification, and demands a considerable amount of time for production. we have demonstrated an immortalized chick embryo cell line, termed pbs-1, is capable of growing unmodified recent isolates of human and avian influenza a and b viruses to extremely high titers ...200818524432
cell-based influenza vaccines: progress to date.human vaccines against influenza have been available for almost 60 years and, until recently, were prepared almost entirely from viruses grown in the allantoic cavity of 9- to 11-day-old embryonated chicken eggs. manufacture involving eggs is not sufficiently flexible to allow vaccine supplies to be rapidly expanded, especially in the face of an impending pandemic. other problems may arise from the infections of progenitor flocks that adversely affect egg supplies, and from the manufacturing pro ...200818627206
newcastle disease virus-vectored vaccines expressing the hemagglutinin or neuraminidase protein of h5n1 highly pathogenic avian influenza virus protect against virus challenge in monkeys.h5n1 highly pathogenic avian influenza virus (hpaiv) causes periodic outbreaks in humans, resulting in severe infections with a high (60%) incidence of mortality. the circulating strains have low human-to-human transmissibility; however, widespread concerns exist that enhanced transmission due to mutations could lead to a global pandemic. we previously engineered newcastle disease virus (ndv), an avian paramyxovirus, as a vector to express the hpaiv hemagglutinin (ha) protein, and we showed that ...201019923177
distinct severe acute respiratory syndrome coronavirus-induced acute lung injury pathways in two different nonhuman primate species.acute lung injury (ali) and acute respiratory distress syndrome (ards), caused by influenza a virus h5n1 and severe acute respiratory syndrome coronavirus (sars-cov), supposedly depend on activation of the oxidative-stress machinery that is coupled with innate immunity, resulting in a strong proinflammatory host response. inflammatory cytokines, such as interleukin 1ß (il-1ß), il-8, and il-6, play a major role in mediating and amplifying ali/ards by stimulating chemotaxis and activation of neutr ...201121325418
impact of host cell line adaptation on quasispecies composition and glycosylation of influenza a virus hemagglutinin.the genome of influenza a viruses is constantly changing (genetic drift) resulting in small, gradual changes in viral proteins. alterations within antibody recognition sites of the viral membrane glycoproteins hemagglutinin (ha) and neuraminidase (na) result in an antigenetic drift, which requires the seasonal update of human influenza virus vaccines. generally, virus adaptation is necessary to obtain sufficiently high virus yields in cell culture-derived vaccine manufacturing. in this study det ...201122163276
the multi-basic cleavage site of the hemagglutinin of the highly pathogenic a/vietnam/1203/2004 (h5n1) avian influenza virus acts as a virulence factor in a host-specific manner in mammals.highly pathogenic avian influenza (hpai) viruses of the h5 and h7 subtypes typically possess multiple basic amino acids around the cleavage site (mbs) of their hemagglutinin (ha) protein, a recognized virulence motif in poultry. to determine the importance of the h5 ha mbs as a virulence factor in mammals, recombinant wild-type hpai a/vietnam/1203/2004 (h5n1) viruses that possessed (h5n1) or lacked (δh5n1) the h5 ha mbs were generated and evaluated for their virulence in balb/c mice, ferrets, an ...201122205751
evaluation of replication, immunogenicity and protective efficacy of a live attenuated cold-adapted pandemic h1n1 influenza virus vaccine in non-human primates.we studied the replication of influenza a/california/07/09 (h1n1) wild type (ca09wt) virus in two non-human primate species and used one of these models to evaluate the immunogenicity and protective efficacy of a live attenuated cold-adapted vaccine, which contains the hemagglutinin and neuraminidase from the h1n1 wild type (wt) virus and six internal protein gene segments of the a/ann arbor/6/60 cold-adapted (ca) master donor virus. we infected african green monkeys (agms) and rhesus macaques w ...201222789506
replication of live attenuated cold-adapted h2n2 influenza virus vaccine candidates in non human primates.the development of an h2n2 vaccine is a priority in pandemic preparedness planning. we previously showed that a single dose of a cold-adapted (ca) h2n2 live attenuated influenza vaccine (laiv) based on the influenza a/ann arbor/6/60 (aa ca) virus was immunogenic and efficacious in mice and ferrets. however, in a phase i clinical trial, viral replication was restricted and immunogenicity was poor. in this study, we compared the replication of four h2n2 laiv candidate viruses, aa ca, a/tecumseh/3/ ...201525444799
african green monkeys recapitulate the clinical experience with replication of live attenuated pandemic influenza virus vaccine candidates.live attenuated cold-adapted (ca) h5n1, h7n3, h6n1, and h9n2 influenza vaccine viruses replicated in the respiratory tract of mice and ferrets, and 2 doses of vaccines were immunogenic and protected these animals from challenge infection with homologous and heterologous wild-type (wt) viruses of the corresponding subtypes. however, when these vaccine candidates were evaluated in phase i clinical trials, there were inconsistencies between the observations in animal models and in humans. the vacci ...201424807726
treatment with anti-c5a antibody improves the outcome of h7n9 virus infection in african green monkeys.patients infected with influenza a(h7n9) virus present with acute lung injury (ali) that is due to severe pneumonia and systemic inflammation. it is often fatal because there are few effective treatment options. complement activation has been implicated in the pathogenesis of virus-induced lung injury; therefore, we investigated the effect of targeted complement inhibition on ali induced by h7n9 virus infection.201525433014
sp-303, an antiviral oligomeric proanthocyanidin from the latex of croton lechleri (sangre de drago).sp-303, a large proanthocyanidin oligomer isolated from the latex of the plant species croton lechleri (eupborbiaceae) has demonstrated broad activity against a variety of dna and rna viruses. in cell culture, sp-303 exhibits potent activity against isolates and laboratory strains of respiratory syncytial virus (rsv), influenza a virus (flu-a) and parainfluenza virus (piv). parallel assays of sp-303 and ribavirin showed comparable activity against these viruses. sp-303 also exhibits significant ...199423195881
primate lentiviruses are differentially inhibited by interferon-induced transmembrane proteins.interferon-induced transmembrane (ifitm) proteins inhibit the entry of a large number of viruses. not surprisingly, many viruses are refractory to this inhibition. in this study, we report that different strains of hiv and siv are inhibited by human ifitm proteins to various degrees, with siv of african green monkeys (siv(agm)) being mostly restricted by human ifitm2. interestingly, siv(agm) is as much inhibited by human ifitm2 as by ifitm3 of its own host african green monkeys. our data further ...201525463599
evaluation of different embryonating bird eggs and cell cultures for isolation efficiency of avian influenza a virus and avian paramyxovirus serotype 1 from real-time reverse transcription polymerase chain reaction-positive wild bird surveillance samples.virus isolation rates for influenza a virus (fluav) and avian paramyxovirus serotype 1 (apmv-1) from wild bird surveillance samples are lower than molecular detection rates for the specific viral genomes. the current study was conducted to examine the possibility of increased virus isolation rates from real-time reverse transcription polymerase chain reaction (real-time rt-pcr) using alternative virus isolation substrates such as embryonating duck eggs (edes), embryonating turkey eggs (etes), ma ...201222529126
cathelicidin-derived antimicrobial peptides inhibit zika virus through direct inactivation and interferon pathway.zika virus (zikv) is a neurotrophic flavivirus that is able to infect pregnant women and cause fetal brain abnormalities. although there is a significant effort in identifying anti-zikv strategies, currently no vaccines or specific therapies are available to treat zikv infection. antimicrobial peptides, which are potent host defense molecules in nearly all forms of life, have been found to be effective against several types of viruses such as hiv-1 and influenza a. however, they have not been te ...201829706959
immune profiling of influenza-specific b- and t-cell responses in macaques using flow cytometry-based assays.influenza remains a significant global public health burden, despite substantial annual vaccination efforts against circulating virus strains. as a result, novel vaccine approaches are needed to generate long-lasting and universal broadly cross-reactive immunity against distinct influenza virus strains and subtypes. several new vaccine candidates are currently under development and/or in clinical trials. the successful development of new vaccines requires testing in animal models, other than mic ...202032741011
influenza-infected newborn and adult monkeys exhibit a strong primary antibody response to hemagglutinin stem.the specificity of antibodies (abs) generated against influenza a virus (iav) infection can significantly alter protection and viral clearance. at present, the impact of age upon this process is relatively unexplored. here, we evaluated the ab response in newborn and adult african green monkeys following infection with iav using a strain that enables us to determine the immunodominance (id) hierarchy of the ab response to hemagglutinin (ha), the principal target of protective abs. this revealed ...202032078584
development of high-yield influenza a virus vaccine viruses.vaccination is one of the most cost-effective ways to prevent infection. influenza vaccines propagated in cultured cells are approved for use in humans, but their yields are often suboptimal. here, we screened a/puerto rico/8/34 (pr8) virus mutant libraries to develop vaccine backbones (defined here as the six viral rna segments not encoding haemagglutinin and neuraminidase) that support high yield in cell culture. we also tested mutations in the coding and regulatory regions of the virus, and c ...201526334134
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