scholarly journals Serological evaluation of an influenza A virus cold-adapted reassortant live vaccine, CR-37 (H1N1), in Japanese adult volunteers

1984 ◽  
Vol 92 (2) ◽  
pp. 231-242 ◽  
Author(s):  
N. Yamane ◽  
Y. Nakamura ◽  
M. Yuki ◽  
T. Odagiri ◽  
N. Ishida
Keyword(s):  
Influenza A Virus ◽  
Influenza A ◽  
Skin Tests ◽  
Wild Type Virus ◽  
Low Grade ◽  
Japanese Adult ◽  
Live Virus ◽  
Cold Adapted ◽  
Vaccine Preparation ◽  
Adult Volunteers

SUMMARYA cold-adapted influenza A virus, CR-37 (H1N1), derived from genetic reassortment between A/Ann Arbor/6/60 (H2N2) cold-adapted variant virus and A/California/10/78 (H1N1) wild-type virus, was tested in Japanese adult volunteers. The CR-37 live virus preparation induced only low-grade clinical reactionsin volunteers for the first 3–4 days after inoculation. Two vaccinees who did not show any antibody changes became febrile (over 38–0 °C). Skin tests using the vaccine preparation and uninfected allantoic fluid were performed, and indicated that one of these two vaccinees was positive for the CR-37 vaccine preparation. A high proportion of the vaccinees whose sera had a haemagglutination-inhibition (HI) antibody titre against thevaccine strain of ≤ 64 before inoculation, seroconverted in both HI and neuraminidase-inhibition (NA1) antibody titrations, and only a few seroconverted in the titration of antibody against type-specific internal antigens. The serological examinations against heterotypic H1N1 variants indicated that the cold-adapted live influenza virus vaccine could induce a broad spectrum of HI antibody reactivity and immunity of long duration.

scholarly journals Characterization of a Neuraminidase-Deficient Influenza A Virus as a Potential Gene Delivery Vector and a Live Vaccine

2004 ◽  
Vol 78 (6) ◽  
pp. 3083-3088 ◽  
Author(s):  
Kyoko Shinya ◽  
Yutaka Fujii ◽  
Hiroshi Ito ◽  
Toshihiro Ito ◽  
Yoshihiro Kawaoka
Keyword(s):  
Influenza Virus ◽  
Influenza A Virus ◽  
Influenza A ◽  
Fluorescent Protein ◽  
Cultured Cells ◽  
Mutant Cell ◽  
Wild Type Virus ◽  
Reading Frame ◽  
Foreign Genes ◽  
Green Fluorescent

ABSTRACT We recently identified a packaging signal in the neuraminidase (NA) viral RNA (vRNA) segment of an influenza A virus, allowing us to produce a mutant virus [GFP(NA)-Flu] that lacks most of the NA open reading frame but contains instead the gene encoding green fluorescent protein (GFP). To exploit the expanding knowledge of vRNA packaging signals to establish influenza virus vectors for the expression of foreign genes, we studied the replicative properties of this virus in cell culture and mice. Compared to wild-type virus, GFP(NA)-Flu was highly attenuated in normal cultured cells but was able to grow to a titer of >106 PFU/ml in a mutant cell line expressing reduced levels of sialic acid on the cell surface. GFP expression from this virus was stable even after five passages in the latter cells. In intranasally infected mice, GFP was detected in the epithelial cells of nasal mucosa, bronchioles, and alveoli for up to 4 days postinfection. We attribute the attenuated growth of GFP(NA)-Flu to virion aggregation at the surface of bronchiolar epithelia. In studies to test the potential of this mutant as a live attenuated influenza vaccine, all mice vaccinated with ≥105 PFU of GFP(NA)-Flu survived when challenged with lethal doses of the parent virus. These results suggest that influenza virus could be a useful vector for expressing foreign genes and that a sialidase-deficient virus may offer an alternative to the live influenza vaccines recently approved for human use.

scholarly journals The Influenza A Virus M2 Cytoplasmic Tail Is Required for Infectious Virus Production and Efficient Genome Packaging

2005 ◽  
Vol 79 (6) ◽  
pp. 3595-3605 ◽  
Author(s):  
Matthew F. McCown ◽  
Andrew Pekosz
Keyword(s):  
Influenza A Virus ◽  
Influenza A ◽  
Infectious Virus ◽  
Virus Production ◽  
Cytoplasmic Tail ◽  
Host Cells ◽  
Wild Type Virus ◽  
Type Virus ◽  
Wild Type ◽  
Virus Particles

ABSTRACT The M2 integral membrane protein encoded by influenza A virus possesses an ion channel activity that is required for efficient virus entry into host cells. The role of the M2 protein cytoplasmic tail in virus replication was examined by generating influenza A viruses encoding M2 proteins with truncated C termini. Deletion of 28 amino acids (M2Stop70) resulted in a virus that produced fourfold-fewer particles but >1,000-fold-fewer infectious particles than wild-type virus. Expression of the full-length M2 protein in trans restored the replication of the M2 truncated virus. Although the M2Stop70 virus particles were similar to wild-type virus in morphology, the M2Stop70 virions contained reduced amounts of viral nucleoprotein and genomic RNA, indicating a defect in vRNP packaging. The data presented indicate the M2 cytoplasmic tail plays a role in infectious virus production by coordinating the efficient packaging of genome segments into influenza virus particles.

scholarly journals Codon Deletions in the Influenza A Virus PA Gene Generate Temperature-Sensitive Viruses

2016 ◽  
Vol 90 (7) ◽  
pp. 3684-3693 ◽  
Author(s):  
Léa Meyer ◽  
Alix Sausset ◽  
Laura Sedano ◽  
Bruno Da Costa ◽  
Ronan Le Goffic ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Rna Polymerase ◽  
Influenza A Virus ◽  
Influenza A ◽  
Wild Type Virus ◽  
Deletion Mutants ◽  
Temperature Sensitive ◽  
Restrictive Temperature ◽  
Wild Type

ABSTRACTThe influenza virus RNA-dependent RNA polymerase, which is composed of three subunits, PB1, PB2, and PA, catalyzes genome replication and transcription within the cell nucleus. The PA linker (residues 197 to 256) can be altered by nucleotide substitutions to engineer temperature-sensitive (ts), attenuated mutants that display a defect in the transport of the PA–PB1 complex to the nucleus at a restrictive temperature. In this study, we investigated the ability of the PA linker to tolerate deletion mutations for furtherin vitroandin vivocharacterization. Four viable mutants with single-codon deletions were generated; all of them exhibited atsphenotype that was associated with the reduced efficiency of replication/transcription of a pseudoviral reporter RNA in a minireplicon assay. Using fluorescently tagged PB1, we observed that the deletion mutants did not efficiently recruit PB1 to reach the nucleus at a restrictive temperature (39.5°C). Mouse infections showed that the four mutants were attenuated and induced antibodies that were able to protect mice from challenge with a lethal homologous wild-type virus. Serialin vitropassages of two deletion mutants at 39.5°C and 37°C did not allow the restoration of a wild-type phenotype among virus progeny. Thus, our results identify codons that can be deleted in the PA gene to engineer genetically stabletsmutants that could be used to design novel attenuated vaccines.IMPORTANCEIn order to generate genetically stable live influenza A virus vaccines, we constructed viruses with single-codon deletions in a discrete domain of the RNA polymerase PA gene. The four rescued viruses exhibited a temperature-sensitive phenotype that we found was associated with a defect in the transport of the PA–PB1 dimer to the nucleus, where viral replication occurs. Thesetsdeletion mutants were shown to be attenuated and to be able to produce antibodies in mice and to protect them from a lethal challenge. Assays to select revertants that were able to grow efficiently at a restrictive temperature failed, showing that these deletion mutants are genetically more stable than conventional substitution mutants. These results are of interest for the design of genetically stable live influenza virus vaccines.

Evaluation of Influenza A/Hong Kong/123/77 (H1N1) ts -1A2 and Cold-Adapted Recombinant Viruses in Seronegative Adult Volunteers

1980 ◽  
Vol 29 (2) ◽  
pp. 348-355 ◽  
Author(s):  
Brian R. Murphy ◽  
Margret B. Rennels ◽  
R. Gordon Douglas ◽  
Robert F. Betts ◽  
Robert B. Couch ◽  
...  
Keyword(s):  
Hemagglutination Inhibition ◽  
Influenza A ◽  
Wild Type Virus ◽  
Enzyme Linked Immunosorbent Assay ◽  
Serological Response ◽  
Type Virus ◽  
Wild Type ◽  
Cold Adapted ◽  
Recombinant Viruses ◽  
Inhibition Response

Two attenuated influenza A donor viruses, the A/Udorn/72 ts -1A2 and the A/Ann Arbor/6/60 cold-adapted ( ca ) viruses, are being evaluated for their ability to reproducibly attenuate each new variant of influenza A virus to a specific and desired level by the transfer of one or more attenuating genes. Each of these donor viruses has been able to attenuate influenza A viruses belonging to the H3N2 subtype by the transfer of one or more attenuating genes. To determine whether these two donor viruses could attenuate a wild-type virus that belonged to a different influenza A subtype, ts -1A2 and ca recombinants of a wild-type virus representative of the A/USSR/77 (H1N1) Russian influenza strain were prepared and evaluated in adult doubly seronegative volunteers at several doses. The recombinants derived from both donor viruses were attenuated for the doubly seronegative adults. Less than 5% of infected vaccinees developed a febrile or systemic reaction, whereas five of six recipients of wild-type virus developed such a response. The 50% human infectious dose (HID 50 ) for each recombinant was approximately 10 5.0 50% tissue culture infective doses. The virus shed by the ts -1A2 and ca vaccinees retained the ts or ca phenotype, or both. This occurred despite replication of the recombinant viruses for up to 9 days. No evidence for transmission of the ca or ts -1A2 recombinant virus to controls was observed. A serum hemagglutination inhibition response was detected in less than 50% of the infected vaccinees. However, with the more sensitive enzyme-linked immunosorbent assay, a serological response was detected in 100% of the ca vaccinees given 300 HID 50 and approximately 70% of ca or ts vaccinees who received 10 to 32 HID 50 of virus. These results indicate that the recombinants derived from both donor viruses were satisfactorily attenuated and were stable genetically after replication in doubly seronegative adults although they induced a lower serum hemagglutination inhibition response than that found previously for H3N2 ts and ca recombinants.

scholarly journals Influenza A Virus NS1 Protein Prevents Activation of NF-κB and Induction of Alpha/Beta Interferon

2000 ◽  
Vol 74 (24) ◽  
pp. 11566-11573 ◽  
Author(s):  
Xiuyan Wang ◽  
Ming Li ◽  
Hongyong Zheng ◽  
Thomas Muster ◽  
Peter Palese ◽  
...  
Keyword(s):  
Virus Infection ◽  
Influenza A Virus ◽  
Influenza A ◽  
Influenza Virus Infection ◽  
Wild Type Virus ◽  
Ns1 Protein ◽  
Wild Type ◽  
Functional Link ◽  
Beta Interferon ◽  
Alpha Beta

ABSTRACT The alpha/beta interferon (IFN-α/β) system represents one of the first lines of defense against virus infections. As a result, most viruses encode IFN antagonistic factors which enhance viral replication in their hosts. We have previously shown that a recombinant influenza A virus lacking the NS1 gene (delNS1) only replicates efficiently in IFN-α/β-deficient systems. Consistent with this observation, we found that infection of tissue culture cells with delNS1 virus, but not with wild-type influenza A virus, induced high levels of mRNA synthesis from IFN-α/β genes, including IFN-β. It is known that transactivation of the IFN-β promoter depends on NF-κB and several other transcription factors. Interestingly, cells infected with delNS1 virus showed high levels of NF-κB activation compared with those infected with wild-type virus. Expression of dominant-negative inhibitors of the NF-κB pathway during delNS1 virus infection prevented the transactivation of the IFN-β promoter, demonstrating a functional link between NF-κB activation and IFN-α/β synthesis in delNS1 virus-infected cells. Moreover, expression of the NS1 protein prevented virus- and/or double-stranded RNA (dsRNA)-mediated activation of the NF-κB pathway and of IFN-β synthesis. This inhibitory property of the NS1 protein of influenza A virus was dependent on its ability to bind dsRNA, supporting a model in which binding of NS1 to dsRNA generated during influenza virus infection prevents the activation of the IFN system. NS1-mediated inhibition of the NF-κB pathway may thus play a key role in the pathogenesis of influenza A virus.

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