scholarly journals An Adenovirus Vector-Mediated Reverse Genetics System for Influenza A Virus Generation

2007 ◽  
Vol 81 (17) ◽  
pp. 9556-9559 ◽  
Author(s):  
Makoto Ozawa ◽  
Hideo Goto ◽  
Taisuke Horimoto ◽  
Yoshihiro Kawaoka
Keyword(s):  
Influenza Virus ◽  
Influenza A Virus ◽  
Influenza A ◽  
Reverse Genetics ◽  
Transfection Efficiency ◽  
Adenovirus Vector ◽  
Vero Cells ◽  
Efficient Gene ◽  
Cloned Cdna ◽  
Plasmid Transfection

ABSTRACT Plasmid-based reverse genetics systems allow the generation of influenza A virus entirely from cloned cDNA. However, since the efficiency of virus generation is dependent on the plasmid transfection efficiency of cells, virus generation is difficult in cells approved for vaccine production that have low transfection efficiencies (e.g., Vero cells). Here we established an alternative reverse genetics system for influenza virus generation by using an adenovirus vector (AdV) which achieves highly efficient gene transfer independent of cell transfection efficiency. This AdV-mediated reverse genetics system will be useful for generating vaccine seed strains and for basic influenza virus studies.

scholarly journals A One-Plasmid System To Generate Influenza Virus in Cultured Chicken Cells for Potential Use in Influenza Vaccine

2009 ◽  
Vol 83 (18) ◽  
pp. 9296-9303 ◽  
Author(s):  
Xiangmin Zhang ◽  
Wei Kong ◽  
Shamaila Ashraf ◽  
Roy Curtiss
Keyword(s):  
Influenza Virus ◽  
Reverse Genetics ◽  
Transfection Efficiency ◽  
Limiting Factor ◽  
Viral Rnas ◽  
Viral Yield ◽  
Plasmid Transfection ◽  
Dual Promoters ◽  
Plasmid Size ◽  
Potential Use

ABSTRACT Influenza virus has a set of ribonucleoproteins (RNPs) consisting of viral RNAs, influenza virus polymerase subunits, and nucleoprotein. Intracellular reconstitution of the whole set of RNPs via plasmid transfection results in the generation of influenza virus. By the use of reverse genetics and dual promoters, we constructed a 23.6-kb eight-unit plasmid that contains all the required constituents to generate influenza virus in chicken cells. Our “one-plasmid” system generated higher titers of influenza virus in chicken cells than the “eight-plasmid” system, enabling a simpler approach for generating vaccine seeds. Our study identified plasmid size as a potential limiting factor affecting transfection efficiency and hence the influenza viral yield from chicken cells.

scholarly journals Rescue of Influenza A Virus from Recombinant DNA

1999 ◽  
Vol 73 (11) ◽  
pp. 9679-9682 ◽  
Author(s):  
Ervin Fodor ◽  
Louise Devenish ◽  
Othmar G. Engelhardt ◽  
Peter Palese ◽  
George G. Brownlee ◽  
...  
Keyword(s):  
Influenza A Virus ◽  
Influenza A ◽  
Reverse Genetics ◽  
Recombinant Dna ◽  
Vero Cells ◽  
Influenza Viruses ◽  
Polymerase I ◽  
Negative Sense ◽  
Expression Plasmids ◽  
Delta Virus

ABSTRACT We have rescued influenza A virus by transfection of 12 plasmids into Vero cells. The eight individual negative-sense genomic viral RNAs were transcribed from plasmids containing human RNA polymerase I promoter and hepatitis delta virus ribozyme sequences. The three influenza virus polymerase proteins and the nucleoprotein were expressed from protein expression plasmids. This plasmid-based reverse genetics technique facilitates the generation of recombinant influenza viruses containing specific mutations in their genes.

scholarly journals A reverse-genetics system for Influenza A virus using T7 RNA polymerase

2007 ◽  
Vol 88 (4) ◽  
pp. 1281-1287 ◽  
Author(s):  
Emmie de Wit ◽  
Monique I. J. Spronken ◽  
Gaby Vervaet ◽  
Guus F. Rimmelzwaan ◽  
Albert D. M. E. Osterhaus ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Rna Polymerase ◽  
Influenza A Virus ◽  
Influenza A ◽  
Reverse Genetics ◽  
T7 Rna Polymerase ◽  
Influenza Virus A ◽  
Sense Orientation ◽  
Polymerase I ◽  
Rna Polymerase Promoter

The currently available reverse-genetics systems for Influenza A virus are all based on transcription of genomic RNA by RNA polymerase I, but the species specificity of this polymerase is a disadvantage. A reverse-genetics vector containing a T7 RNA polymerase promoter, hepatitis delta virus ribozyme sequence and T7 RNA polymerase terminator sequence has been developed. To achieve optimal expression in minigenome assays, it was determined that viral RNA should be inserted in this vector in the negative-sense orientation with two additional G residues downstream of the T7 RNA polymerase promoter. It was also shown that expression of the minigenome was more efficient when a T7 RNA polymerase with a nuclear-localization signal was used. By using this reverse-genetics system, recombinant influenza virus A/PR/8/34 was produced more efficiently than by using a similar polymerase I-based reverse-genetics system. Furthermore, influenza virus A/NL/219/03 could be rescued from 293T, MDCK and QT6 cells. Thus, a reverse-genetics system for the rescue of Influenza A virus has been developed, which will be useful for fundamental research and vaccine seed strain production in a variety of cell lines.

scholarly journals Reverse genetics approach towards understanding pathogenesis of H5N1 Hong Kong influenza A virus infection

2001 ◽  
Vol 356 (1416) ◽  
pp. 1841-1843 ◽  
Author(s):  
Masato Hatta ◽  
Gabriele Neumann ◽  
Yoshihiro Kawaoka
Keyword(s):  
Influenza Virus ◽  
Hong Kong ◽  
Influenza A ◽  
Reverse Genetics ◽  
Gene Segment ◽  
Influenza Viruses ◽  
H5n1 Influenza ◽  
Influenza A Virus Infection ◽  
Cloned Cdna ◽  
First Time

In 1990, Palese and colleagues established a method (reverse genetics) that allowed one to generate influenza virus containing a gene segment derived from cloned cDNA. Although this method contributed tremendously to our understanding of influenza pathogenesis, the requirement of helper viruses limited its use in many experimental settings. Recently, we and others established systems for the generation of influenza viruses entirely from cloned cDNAs. These systems require only DNA cloning and transfection techniques, and can therefore be easily implemented by laboratories working in the fields of molecular biology and virology. Thus, for the first time, a system is now available that allows highly efficient generation of influenza virus without technical limitations. Using this technology, we generated the same strain of H5N1 influenza viruses that caused an outbreak in Hong Kong in 1997, killing six people.

scholarly journals Influenza A Virus Hemagglutinin and Other Pathogen Glycoprotein Interactions with NK Cell Natural Cytotoxicity Receptors NKp46, NKp44, and NKp30

Viruses ◽  
2021 ◽  
Vol 13 (2) ◽  
pp. 156
Author(s):  
Jasmina M. Luczo ◽  
Sydney L. Ronzulli ◽  
Stephen M. Tompkins
Keyword(s):  
Influenza Virus ◽  
Nk Cells ◽  
Influenza A Virus ◽  
Influenza A ◽  
Nk Cell ◽  
Influenza Virus Infection ◽  
Target Cells ◽  
Natural Cytotoxicity ◽  
Activating Receptors ◽  
Natural Cytotoxicity Receptors

Natural killer (NK) cells are part of the innate immunity repertoire, and function in the recognition and destruction of tumorigenic and pathogen-infected cells. Engagement of NK cell activating receptors can lead to functional activation of NK cells, resulting in lysis of target cells. NK cell activating receptors specific for non-major histocompatibility complex ligands are NKp46, NKp44, NKp30, NKG2D, and CD16 (also known as FcγRIII). The natural cytotoxicity receptors (NCRs), NKp46, NKp44, and NKp30, have been implicated in functional activation of NK cells following influenza virus infection via binding with influenza virus hemagglutinin (HA). In this review we describe NK cell and influenza A virus biology, and the interactions of influenza A virus HA and other pathogen lectins with NK cell natural cytotoxicity receptors (NCRs). We review concepts which intersect viral immunology, traditional virology and glycobiology to provide insights into the interactions between influenza virus HA and the NCRs. Furthermore, we provide expert opinion on future directions that would provide insights into currently unanswered questions.

scholarly journals Thapsigargin Is a Broad-Spectrum Inhibitor of Major Human Respiratory Viruses: Coronavirus, Respiratory Syncytial Virus and Influenza A Virus

Viruses ◽  
2021 ◽  
Vol 13 (2) ◽  
pp. 234
Author(s):  
Sarah Al-Beltagi ◽  
Cristian Alexandru Preda ◽  
Leah V. Goulding ◽  
Joe James ◽  
Juan Pu ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Respiratory Syncytial Virus ◽  
Influenza A Virus ◽  
Broad Spectrum ◽  
Influenza A ◽  
Respiratory Viruses ◽  
Influenza Viruses ◽  
Virus Challenge ◽  
2009 H1n1 ◽  
Syncytial Virus

The long-term control strategy of SARS-CoV-2 and other major respiratory viruses needs to include antivirals to treat acute infections, in addition to the judicious use of effective vaccines. Whilst COVID-19 vaccines are being rolled out for mass vaccination, the modest number of antivirals in use or development for any disease bears testament to the challenges of antiviral development. We recently showed that non-cytotoxic levels of thapsigargin (TG), an inhibitor of the sarcoplasmic/endoplasmic reticulum (ER) Ca2+ ATPase pump, induces a potent host innate immune antiviral response that blocks influenza A virus replication. Here we show that TG is also highly effective in blocking the replication of respiratory syncytial virus (RSV), common cold coronavirus OC43, SARS-CoV-2 and influenza A virus in immortalized or primary human cells. TG’s antiviral performance was significantly better than remdesivir and ribavirin in their respective inhibition of OC43 and RSV. Notably, TG was just as inhibitory to coronaviruses (OC43 and SARS-CoV-2) and influenza viruses (USSR H1N1 and pdm 2009 H1N1) in separate infections as in co-infections. Post-infection oral gavage of acid-stable TG protected mice against a lethal influenza virus challenge. Together with its ability to inhibit the different viruses before or during active infection, and with an antiviral duration of at least 48 h post-TG exposure, we propose that TG (or its derivatives) is a promising broad-spectrum inhibitor against SARS-CoV-2, OC43, RSV and influenza virus.

scholarly journals STUDIES ON INFLUENZA VIRUS

1941 ◽  
Vol 73 (5) ◽  
pp. 581-599 ◽  
Author(s):  
Edwin H. Lennette ◽  
Frank L. Horsfall
Keyword(s):  
Influenza Virus ◽  
Chick Embryo ◽  
Influenza A Virus ◽  
Influenza A ◽  
Complement Fixation ◽  
Soluble Antigen ◽  
Swine Virus ◽  
Specific Fixation ◽  
Mouse Lungs

Influenza complement fixation tests designed for use with ferret serum are described. Complement-fixing antigens derived from influenza ferret lungs were unsatisfactory due to their low content of soluble antigen; those prepared from mouse lungs or developing chick embryo membranes proved to be better antigenically and were reliable when the various reagents in the test were properly adjusted to eliminate non-specific fixation of complement. The results of cross complement fixation tests indicated that the soluble antigens of the PR8 and W.S. strains of influenza A virus were closely similar, if not identical. They indicated also that the soluble antigen of swine virus possessed components present in the antigens of the human strains of virus.

scholarly journals Dicer is involved in protection against influenza A virus infection

2007 ◽  
Vol 88 (10) ◽  
pp. 2627-2635 ◽  
Author(s):  
Alexey A. Matskevich ◽  
Karin Moelling
Keyword(s):  
Virus Infection ◽  
Influenza A Virus ◽  
Mammalian Cells ◽  
Influenza A ◽  
Virus Production ◽  
Vero Cells ◽  
Antiviral Response ◽  
Protein Levels ◽  
Infected Cells ◽  
Influenza A Virus Infection

In mammals the interferon (IFN) system is a central innate antiviral defence mechanism, while the involvement of RNA interference (RNAi) in antiviral response against RNA viruses is uncertain. Here, we tested whether RNAi is involved in the antiviral response in mammalian cells. To investigate the role of RNAi in influenza A virus-infected cells in the absence of IFN, we used Vero cells that lack IFN-α and IFN-β genes. Our results demonstrate that knockdown of a key RNAi component, Dicer, led to a modest increase of virus production and accelerated apoptosis of influenza A virus-infected cells. These effects were much weaker in the presence of IFN. The results also show that in both Vero cells and the IFN-producing alveolar epithelial A549 cell line influenza A virus targets Dicer at mRNA and protein levels. Thus, RNAi is involved in antiviral response, and Dicer is important for protection against influenza A virus infection.

scholarly journals Development of Neuraminidase Subtype-Specific Reference Antisera by Recombinant Protein Expressed in Baculovirus

2012 ◽  
Vol 20 (2) ◽  
pp. 140-145 ◽  
Author(s):  
Kyu-Jun Lee ◽  
Jun-Gu Choi ◽  
Hyun-Mi Kang ◽  
Kwang-Il Kim ◽  
Choi-Kyu Park ◽  
...  
Keyword(s):  
Influenza Virus ◽  
Avian Influenza ◽  
Influenza A Virus ◽  
Diagnostic Tests ◽  
Influenza A ◽  
Recombinant Baculovirus ◽  
Cross Reactivity ◽  
Specific Reference ◽  
Simple Method ◽  
Avian Influenza A Virus

ABSTRACTOutbreaks of avian influenza A virus infection, particularly the H5N1 strains that have affected birds and some humans for the past 15 years, have highlighted the need for increased surveillance and disease control. Such measures require diagnostic tests to detect and characterize the different subtypes of influenza virus. In the current study, a simple method for producing reference avian influenza virus antisera to be used in diagnostic tests was developed. Antisera of nine avian influenza A virus neuraminidases (NA) used for NA subtyping were produced using a recombinant baculovirus. The recombinant NA (rNA) proteins were expressed in Sf9 insect cells and inoculated intramuscularly into specific-pathogen-free chickens with the ISA70 adjuvant. The NA inhibition antibody titers of the rNA antiserum were in the ranges of 5 to 8 and 6 to 9 log2units after the primary and boost immunizations, respectively. The antisera were subtype specific, showing low cross-reactivity against every other NA subtype using the conventional thiobarbituric acid NA inhibition assay. These results suggest that this simple method for producing reference NA antisera without purification may be useful for the diagnosis and surveillance of influenza virus.

scholarly journals REACTIONS BETWEEN INFLUENZA VIRUS AND A COMPONENT OF ALLANTOIC FLUID

1948 ◽  
Vol 88 (4) ◽  
pp. 463-484 ◽  
Author(s):  
Paul H. Hardy ◽  
Frank L. Horsfall
Keyword(s):  
Influenza Virus ◽  
Influenza A Virus ◽  
Influenza A ◽  
Allantoic Fluid ◽  
Partial Dissociation

Evidence is presented which shows that there is a component present in normal allantoic fluid, probably mucoprotein in nature, capable of combining with influenza A virus (PR8), and that following combination between this component and the virus only partial dissociation of the complex occurs. Evidence is also presented which strongly suggests that the component is present in virus-infected allantoic fluid in which it is in part combined with the virus and in part free although altered by viral action. The probability that the component is present as well in highly purified preparations of influenza virus, and its effect upon various reactions obtained with this agent are discussed.

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