Rabies virus production in high vero cell density cultures on macroporous microcarriers

2004 ◽  
Vol 85 (5) ◽  
pp. 506-515 ◽  
Author(s):  
A. Y. Yokomizo ◽  
M. M. Antoniazzi ◽  
P. L. Galdino ◽  
N. Azambuja ◽  
S. A. C. Jorge ◽  
...  
Keyword(s):  
Vero Cell ◽  
Cell Density ◽  
Rabies Virus ◽  
Virus Production ◽  
Macroporous Microcarriers

High Vero Cell Density and Rabies Virus Proliferation on Fibracel Disks Versus Cytodex-1 in Spinner Flask

2011 ◽  
Vol 14 (7) ◽  
pp. 441-448 ◽  
Author(s):  
S. Mehdi Hassanzade ◽  
Ali Zavareh ◽  
M. Ali Shokrgozar ◽  
Ali Ramezani ◽  
Ahmad Fayaz
Keyword(s):  
Vero Cell ◽  
Cell Density ◽  
Rabies Virus ◽  
Spinner Flask

Whole Genome Sequencing and Phylogenetic Analysis of the Rabies Virus Strain Moscow 3253 Adapted to a Vero Cell Line

2020 ◽  
Vol 35 (4) ◽  
pp. 237-242
Author(s):  
Ya. M. Krasnov ◽  
Zh. V. Alkhova ◽  
S. V. Generalov ◽  
I. V. Tuchkov ◽  
E. A. Naryshkina ◽  
...  
Keyword(s):  
Phylogenetic Analysis ◽  
Vero Cell ◽  
Whole Genome Sequencing ◽  
Cell Line ◽  
Rabies Virus ◽  
Genome Sequencing ◽  
Virus Strain ◽  
Whole Genome ◽  
Vero Cell Line ◽  
Rabies Virus Strain

scholarly journals PPEY Motif within the Rabies Virus (RV) Matrix Protein Is Essential for Efficient Virion Release and RV Pathogenicity

2008 ◽  
Vol 82 (19) ◽  
pp. 9730-9738 ◽  
Author(s):  
Christoph Wirblich ◽  
Gene S. Tan ◽  
Amy Papaneri ◽  
Peter J. Godlewski ◽  
Jan Marc Orenstein ◽  
...  
Keyword(s):  
Viral Replication ◽  
Rabies Virus ◽  
Rna Synthesis ◽  
Matrix Protein ◽  
Virus Production ◽  
Wild Type Virus ◽  
Wild Type ◽  
Conserved Sequence ◽  
Virion Release ◽  
The Matrix

ABSTRACT Late (L) domains containing the highly conserved sequence PPXY were first described for retroviruses, and later research confirmed their conservation and importance for efficient budding of several negative-stranded RNA viruses. Rabies virus (RV), a member of the Rhabdoviridae family, contains the sequence PPEY (amino acids 35 to 38) within the N terminus of the matrix (M) protein, but the functions of this potential L-domain in the viral life cycle, viral pathogenicity, and immunogenicity have not been established. Here we constructed a series of recombinant RVs containing mutations within the PPEY motif and analyzed their effects on viral replication and RV pathogenicity. Our results indicate that the first proline at position 35 is the most important for viral replication, whereas P36 and Y38 have a lesser but still noticeable impact. The reduction in viral replication was most likely due to inhibition of virion release, because initially no major impact on RV RNA synthesis was observed. In addition, results from electron microscopy demonstrated that the M4A mutant virus (PPEY→SAEA) displayed a more cell-associated phenotype than that of wild-type RV. Furthermore, all mutations within the PPEY motif resulted in reduced spread of the recombinant RVs as indicated by a reduction in focus size. Importantly, recombinant PPEY L-domain mutants were highly attenuated in mice yet still elicited potent antibody responses against RV G protein that were as high as those observed after infection with wild-type virus. Our data indicate that the RV PPEY motif has L-domain activity essential for efficient virus production and pathogenicity but is not essential for immunogenicity and thus can be targeted to increase the safety of rabies vaccine vectors.

scholarly journals Identification of a Role for Nucleolin in Rabies Virus Infection

2014 ◽  
Vol 89 (3) ◽  
pp. 1939-1943 ◽  
Author(s):  
S. Oksayan ◽  
J. Nikolic ◽  
C. T. David ◽  
D. Blondel ◽  
D. A. Jans ◽  
...  
Keyword(s):  
Rabies Virus ◽  
Infectious Virus ◽  
Virus Production ◽  
Host Cells ◽  
Nucleocytoplasmic Trafficking ◽  
P Protein ◽  
Rabies Virus Infection ◽  
Infected Cells ◽  
Protein Isoform ◽  
Nucleolin Expression

Rabies virus replicates in the cytoplasm of host cells, but rabies virus phosphoprotein (P-protein) undergoes active nucleocytoplasmic trafficking. Here we show that the largely nuclear P-protein isoform P3 can localize to nucleoli and forms specific interactions with nucleolin. Importantly, depletion of nucleolin expression inhibits viral protein expression and infectious virus production by infected cells. This provides the first evidence that lyssaviruses interact with nucleolin and that nucleolin is important to lyssavirus infection.

scholarly journals Optimization of Specifications for Scaled-Up Fixed Rabies Virus Cultivation (“Moscow 3253” Strain) in Vero Cell Culture

2014 ◽  
pp. 101-103
Author(s):  
S. V. Generalov ◽  
E. G. Abramova ◽  
Zh. V. Matveeva ◽  
I. M. Zhulidov ◽  
L. V. Savitskaya ◽  
...  
Keyword(s):  
Cell Culture ◽  
Vero Cell ◽  
Rabies Virus ◽  
Vero Cell Culture

scholarly journals Optimization of environment for high density Vero cell culture: effect of dissolved oxygen and nutrient supply on cell growth and changes in metabolites

1986 ◽  
Vol 81 (1) ◽  
pp. 65-103
Author(s):  
A.T. Nahapetian ◽  
J.N. Thomas ◽  
W.G. Thilly
Keyword(s):  
Cell Growth ◽  
Dissolved Oxygen ◽  
Vero Cell ◽  
Cell Density ◽  
Lactate Production ◽  
Nutrient Supply ◽  
High Density ◽  
Limiting Factors ◽  
Utilization Rate ◽  
Maximum Cell Density

This study was initiated for optimization of the environment of a technologically useful mammalian cell line for high density production. Cultures of Vero cells on microcarriers were perfused with 100%, 50%, 25% and 12.5% modified L15 media (galactose was replaced with 10 mM-fructose, with 4 mM-glutamine and 5% foetal bovine serum) in phosphate-buffered saline at either 4 or 8 vol. day-1. Cell growth, pH, dissolved oxygen, and changes in the metabolites, lactate to pyruvate and lactate to ammonia indices, demonstrated that under the conditions used in the present study, perfusion of cultures with 50% L15 medium in PBS at 8 vol. day-1 provided the optimum microenvironment for Vero cell growth. The highest cell density in the perfused cultures was 3 X 10(7) cells ml-1, which at these conditions was ten times higher than the maximum cell density (3 X 10(6) cells ml-1) obtained in a batch culture. Nutrient supply and conditioning factors were the most probable growth-limiting factors in cultures that were perfused with 12.5% and 25% L15 media, while multilayering, limitation of available oxygen, and accumulation of metabolic end products in the cellular microenvironment were the most probable causes of a density-dependent inhibition of cell growth observed under the optimized and overfed (supply of 100% L15 medium at the rate of 8 vol. day-1) culture conditions. Under the optimized environmental condition, the major source of energy was probably glutamine during the first week. However, significant utilization of fructose became evident at higher cell densities during the second week, when lactate production dramatically declined and reached an almost undetectable level, while respiration progressively assumed the predominant role in energy production. It is postulated that ‘available’ oxygen in the multicell-layered microenvironment of the optimized cultures was higher than in the overfed culture due to the greater utilization rate of oxygen for oxidation of excess nutrients in the overfed culture.

High cell density cultivations by alternating tangential flow (ATF) perfusion for influenza A virus production using suspension cells

Vaccine ◽  
2014 ◽  
Vol 32 (24) ◽  
pp. 2770-2781 ◽  
Author(s):  
Yvonne Genzel ◽  
Thomas Vogel ◽  
Johannes Buck ◽  
Ilona Behrendt ◽  
Daniel Vazquez Ramirez ◽  
...  
Keyword(s):  
Cell Density ◽  
Influenza A Virus ◽  
Influenza A ◽  
High Cell Density ◽  
Virus Production ◽  
High Cell ◽  
Suspension Cells ◽  
Tangential Flow

scholarly journals Dominance of a Nonpathogenic Glycoprotein Gene over a Pathogenic Glycoprotein Gene in Rabies Virus

2007 ◽  
Vol 81 (13) ◽  
pp. 7041-7047 ◽  
Author(s):  
Milosz Faber ◽  
Marie-Luise Faber ◽  
Jianwei Li ◽  
Mirjam A. R. Preuss ◽  
Matthias J. Schnell ◽  
...  
Keyword(s):  
Brain Tissue ◽  
Rabies Virus ◽  
Lower Risk ◽  
Rna Synthesis ◽  
Vaccine Candidate ◽  
Neuroblastoma Cells ◽  
Virus Production ◽  
Viral Rna ◽  
Glycoprotein Gene

ABSTRACT The nonpathogenic phenotype of the live rabies virus (RV) vaccine SPBNGAN is determined by an Arg→Glu exchange at position 333 in the glycoprotein, designated GAN. We recently showed that after several passages of SPBNGAN in mice, an Asn→Lys mutation arose at position 194 of GAN, resulting in GAK, which was associated with a reversion to the pathogenic phenotype. Because an RV vaccine candidate containing two GAN genes (SPBNGAN-GAN) exhibits increased immunogenicity in vivo compared to the single-GAN construct, we tested whether the presence of two GAN genes might also enhance the probability of reversion to pathogenicity. Comparison of SPBNGAN-GAN with RVs constructed to contain either both GAN and GAK genes (SPBNGAN-GAK and SPBNGAK-GAN) or two GAK genes (SPBNGAK-GAK) showed that while SPBNGAK-GAK was pathogenic, SPBNGAN-GAN and SPBNGAN-GAK were completely nonpathogenic and SPBNGAK-GAN showed strongly reduced pathogenicity. Analysis of genomic RV RNA in mouse brain tissue revealed significantly lower virus loads in SPBNGAN-GAK- and SPBNGAK-GAN-infected brains than those detected in SPBNGAK-GAK-infected brains, indicating the dominance of the nonpathogenic phenotype determined by GAN over the GAK-associated pathogenic phenotype. Virus production and viral RNA synthesis were markedly higher in SPBNGAN-, SPBNGAK-GAN-, and SPBNGAN-GAK-infected neuroblastoma cells than in the SPBNGAK- and SPBNGAK-GAK-infected counterparts, suggesting control of GAN dominance at the level of viral RNA synthesis. These data point to the lower risk of reversion to pathogenicity of a recombinant RV carrying two identical GAN genes compared to that of an RV carrying only a single GAN gene.

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