Production of reovirus type-1 and type-3 from Vero cells grown on solid and macroporous microcarriers.
Two strains of reovirus were propagated in Vero cells grown in stationary or microcarriers cultures. Vero cells grown as monolayers on T-flasks or in spinner cultures of Cytodex-1 or Cultispher-G microcarriers could be infected with reovirus serotype 1, strain Lang (T1L), and serotype 3, strain Dearing (T3D). A regime of intermittent low speed stirring at reduced culture volume was critical to ensure viral infection of cells in microcarrier cultures. The virus titre increased by 3 to 4 orders of magnitude over a culture period of 150 h. Titres of the T3D reovirus strain were higher (43%) compared to those of the T1L strain in all cultures. Titres were significantly higher in T-flask and Cytodex-1 microcarrier cultures compared to Cultispher-G cultures with respect to either reovirus type. The viral productivity in the microcarrier cultures was dependent upon the multiplicity of infection (MOI) and the cell/bead ratio at the point of infection. A combination of high MOI (5 pfu/cell) and high cell/bead loading (>400 for Cytodex-1 and >1,000 for Cultispher-G) resulted in a low virus productivity per cell. However, at low MOI (0.5 pfu/cell) the virus productivity per cell was significantly higher at high cell/bead loading in cultures of either microcarrier type. The maximum virus titre (8.5 x 10(9) pfu/mL) was obtained in Cytodex-1 cultures with a low MOI (0.5 pfu/cell) and a cell/bead loading of 1,000. The virus productivity per cell in these cultures was 4,000 pfu/cell. The lower viral yield in the Cultispher-G microcarrier cultures is attributed to a decreased accessibility of the entrapped cells to viral infection. The high viral productivity from the Vero cells in Cytodex-1 cultures suggests that this is a suitable system for the development of a vaccine production system for the Reoviridae viruses.
Berry
J M
JM
Department of Microbiology and Medical Microbiology, University of Manitoba, Winnipeg, Manitoba, Canada.
Barnabé
N
N
Coombs
K M
KM
Butler
M
M
D000818:Animals; D001709:Biotechnology; D002522:Chlorocebus aethiops; D003470:Culture Media; D005069:Evaluation Studies as Topic; D012089:Mammalian orthoreovirus 3 / Q000502:physiology*; D008863:Microspheres; D018165:Orthoreovirus / Q000502:physiology*; D014709:Vero Cells; D010948:Viral Plaque Assay; D014765:Viral Vaccines / Q000302:isolation & purification; D014776:Virus Cultivation / Q000379:methods*; D014779:Virus Replication
D003470:Culture Media; D014765:Viral Vaccines