scholarly journals Cell culture–based production of defective interfering influenza A virus particles in perfusion mode using an alternating tangential flow filtration system

2021 ◽  
Author(s):  
Marc D. Hein ◽  
Anshika Chawla ◽  
Maurizio Cattaneo ◽  
Sascha Y. Kupke ◽  
Yvonne Genzel ◽  
...  
Keyword(s):  
Influenza A Virus ◽  
Hollow Fiber ◽  
Influenza A ◽  
Tubular Cell ◽  
Cell Retention ◽  
Virus Particles ◽  
Tangential Flow ◽  
Perfusion Process ◽  
Flow Filtration ◽  
Retention Device

AbstractRespiratory diseases including influenza A virus (IAV) infections represent a major threat to human health. While the development of a vaccine requires a lot of time, a fast countermeasure could be the use of defective interfering particles (DIPs) for antiviral therapy. IAV DIPs are usually characterized by a large internal deletion in one viral RNA segment. Consequentially, DIPs can only propagate in presence of infectious standard viruses (STVs), compensating the missing gene function. Here, they interfere with and suppress the STV replication and might act “universally” against many IAV subtypes. We recently reported a production system for purely clonal DIPs utilizing genetically modified cells. In the present study, we established an automated perfusion process for production of a DIP, called DI244, using an alternating tangential flow filtration (ATF) system for cell retention. Viable cell concentrations and DIP titers more than 10 times higher than for a previously reported batch cultivation were observed. Furthermore, we investigated a novel tubular cell retention device for its potential for continuous virus harvesting into the permeate. Very comparable performances to typically used hollow fiber membranes were found during the cell growth phase. During the virus replication phase, the tubular membrane, in contrast to the hollow fiber membrane, allowed 100% of the produced virus particles to pass through. To our knowledge, this is the first time a continuous virus harvest was shown for a membrane-based perfusion process. Overall, the process established offers interesting possibilities for advanced process integration strategies for next-generation virus particle and virus vector manufacturing.Key points• An automated perfusion process for production of IAV DIPs was established.• DIP titers of 7.40E + 9 plaque forming units per mL were reached.• A novel tubular cell retention device enabled continuous virus harvesting.

scholarly journals Cell culture-based production of defective interfering influenza A virus particles in perfusion mode using an alternating tangential flow filtration system

2021 ◽  
Author(s):  
Marc Dominique Hein ◽  
Anshika Chawla ◽  
Maurizio Cattaneo ◽  
Sascha Young Kupke ◽  
Yvonne Genzel ◽  
...  
Keyword(s):  
Influenza A Virus ◽  
Hollow Fiber ◽  
Influenza A ◽  
Hollow Fiber Membrane ◽  
Cell Retention ◽  
Tangential Flow Filtration ◽  
Virus Particles ◽  
Tangential Flow ◽  
Perfusion Process ◽  
Flow Filtration

Respiratory diseases including influenza A virus (IAV) infections represent a major threat to human health. While the development of a vaccine requires a lot of time, a fast countermeasure could be the use of defective interfering particles (DIPs) for antiviral therapy. IAV DIPs are usually characterized by a large internal deletion in one viral RNA segment. Consequentially, DIPs can only propagate in presence of infectious standard viruses (STVs), compensating the missing gene function. Here, they interfere with and suppress the STV replication and might act "universally" against many IAV subtypes. We recently reported a production system for purely clonal DIPs utilizing genetically modified cells. In the present study, we established an automated perfusion process for production of a DIP, called DI244, using an alternating tangential flow filtration (ATF) system for cell retention. Viable cell concentrations and DIP titers more than 10-times higher than for a previously reported batch cultivation were observed. Further, we investigated a novel tubular cell retention device for its potential for continuous virus harvesting into the permeate. Very comparable performances to typically used hollow fiber membranes were found during the cell growth phase. During the virus replication phase the tubular membrane, in contrast to the hollow fiber membrane, allowed 100% of the produced virus particles to pass through. To our knowledge, this is the first time a continuous virus harvest was shown for a membrane-based perfusion process. Overall, the process established offers interesting possibilities for advanced process integration strategies for next-generation virus particle and virus vector manufacturing.

Evidence for specific packaging of the influenza A virus genome from conditionally defective virus particles lacking a polymerase gene

Vaccine ◽  
2006 ◽  
Vol 24 (44-46) ◽  
pp. 6647-6650 ◽  
Author(s):  
Emmie de Wit ◽  
Monique I.J. Spronken ◽  
Guus F. Rimmelzwaan ◽  
Albert D.M.E. Osterhaus ◽  
Ron A.M. Fouchier
Keyword(s):  
Influenza A Virus ◽  
Influenza A ◽  
Virus Genome ◽  
Polymerase Gene ◽  
Virus Particles ◽  
Defective Virus

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 THE PRODUCTION OF FEVER BY INFLUENZAL VIRUSES

1949 ◽  
Vol 90 (4) ◽  
pp. 321-334 ◽  
Author(s):  
Robert R. Wagner ◽  
Ivan L. Bennett ◽  
Virgil S. LeQuire
Keyword(s):  
Influenza A Virus ◽  
Low Temperatures ◽  
Influenza A ◽  
Immune Serum ◽  
Influenza B ◽  
Chick Embryos ◽  
Febrile Response ◽  
Virus Particles ◽  
Chicken Erythrocytes

The intravenous injection of the PR8 strain of influenza A virus, the Lee strain of influenza B, and the "B" strain of Newcastle disease virus produces fever in rabbits. This phenomenon has been studied in relation to certain in vitro properties of these viruses. Saline suspensions of virus prepared by centrifugation or elution from chicken erythrocytes produced fever. Fluids from which most of the virus particles had been removed were non-pyrogenic. Exposure to temperatures which destroyed the infectivity of the virus for chick embryos did not prevent fever. However, heating sufficient to destroy the hemagglutinin also rendered virus non-pyrogenic. The injection of erythrocytes onto which virus had been adsorbed produced fever. Heated virus adsorbed onto erythrocytes, which failed to elute, produced no elevation of temperature, although heated virus alone was pyrogenic. Neutralization of virus with specific immune serum prevented fever. Antipyrine was capable of abolishing the febrile response to virus. Certain differences between the febrile response in rabbits to the injection of viruses and that following bacterial pyrogens were noted. The period between injection and beginning of temperature rise is longer with virus than with bacterial pyrogens. Relatively low temperatures inactivate the fever-producing capacity of viruses, whereas bacterial pyrogens withstand prolonged autoclaving, and the neutralization of viral fever by specific immune serum contrasts sharply with the failure of antibody to affect the response to bacterial pyrogens. Certain previous observations on the lymphopenia produced in rabbits by the injection of influenzal viruses were confirmed. The capacity of virus preparations to induce fever in rabbits closely parallels their capacity to induce lymphopenia. It was concluded that the fever-producing property of influenzal viruses is closely associated with the capacity to agglutinate erythrocytes.

scholarly journals Distinct Domains of the Influenza A Virus M2 Protein Cytoplasmic Tail Mediate Binding to the M1 Protein and Facilitate Infectious Virus Production

2006 ◽  
Vol 80 (16) ◽  
pp. 8178-8189 ◽  
Author(s):  
Matthew F. McCown ◽  
Andrew Pekosz
Keyword(s):  
Influenza A Virus ◽  
Influenza A ◽  
Reverse Genetics ◽  
Infectious Virus ◽  
Virus Assembly ◽  
Virus Production ◽  
Cytoplasmic Tail ◽  
M2 Protein ◽  
Virus Particles ◽  
M1 Protein

ABSTRACT The cytoplasmic tail of the influenza A virus M2 protein is highly conserved among influenza A virus isolates. The cytoplasmic tail appears to be dispensable with respect to the ion channel activity associated with the protein but important for virus morphology and the production of infectious virus particles. Using reverse genetics and transcomplementation assays, we demonstrate that the M2 protein cytoplasmic tail is a crucial mediator of infectious virus production. Truncations of the M2 cytoplasmic tail result in a drastic decrease in infectious virus titers, a reduction in the amount of packaged viral RNA, a decrease in budding events, and a reduction in budding efficiency. The M1 protein binds to the M2 cytoplasmic tail, but the M1 binding site is distinct from the sequences that affect infectious virus particle formation. Influenza A virus strains A/Udorn/72 and A/WSN/33 differ in their requirements for M2 cytoplasmic tail sequences, and this requirement maps to the M1 protein. We conclude that the M2 protein is required for the formation of infectious virus particles, implicating the protein as important for influenza A virus assembly in addition to its well-documented role during virus entry and uncoating.

scholarly journals Interferon-inducedSus scrofaMx1 blocks endocytic traffic of incoming influenza A virus particles

2010 ◽  
Vol 41 (3) ◽  
pp. 29 ◽  
Author(s):  
Mélanie Palm ◽  
Mutien-Marie Garigliany ◽  
François Cornet ◽  
Daniel Desmecht
Keyword(s):  
Influenza A Virus ◽  
Influenza A ◽  
Virus Particles ◽  
Endocytic Traffic

Influenza A Virus Subpopulations and Their Implication in Pathogenesis and Vaccine Development

2020 ◽  
Vol 8 (1) ◽  
pp. 247-267 ◽  
Author(s):  
Amir Ghorbani ◽  
John M. Ngunjiri ◽  
Chang-Won Lee
Keyword(s):  
Influenza A Virus ◽  
Influenza A ◽  
Vaccine Development ◽  
Current Knowledge ◽  
Genomic Diversity ◽  
Biologically Active ◽  
Virus Particles ◽  
Live Attenuated Influenza Vaccine ◽  
Minor Portion ◽  
A Minor

The concept of influenza A virus (IAV) subpopulations emerged approximately 75 years ago, when Preben von Magnus described “incomplete” virus particles that interfere with the replication of infectious virus. It is now widely accepted that infectious particles constitute only a minor portion of biologically active IAV subpopulations. The IAV quasispecies is an extremely diverse swarm of biologically and genetically heterogeneous particle subpopulations that collectively influence the evolutionary fitness of the virus. This review summarizes the current knowledge of IAV subpopulations, focusing on their biologic and genomic diversity. It also discusses the potential roles IAV subpopulations play in virus pathogenesis and live attenuated influenza vaccine development.

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