Intracellular ATP and total adenylate concentrations are critical predictors of reovirus productivity from Vero cells.
The productivity of reovirus type-3 Dearing was studied in cultures of Vero cells in serum-free media. Viral productivity was dependent upon the metabolic state of the cells rather than the phase of growth at which the cells were infected. Cells at different energy states were established by 24-h incubation in nutrient-depleted media. This resulted in variable intracellular nucleotide concentrations but high cellular viability was maintained. Of the nucleotides analyzed at the time of infection only the intracellular [ATP] and total adenylate nucleotides were positively correlated with viral productivity. The correlated data followed a sigmoidal plot with an equation defined by polynomial regression analysis. Apparent threshold values of 3.2 fmol/cell and 3.3 fmol/cell were established for ATP and total adenylate, respectively, at which the viral production was 50% the maximal value. Cultures with lower ATP and total adenylate levels at the time of infection resulted in as much as a 95% reduction in overall viral titer compared to the control. The adenylate energy charge (AEC) showed a negative correlation with viral production with an AEC value >0.97 resulting in low virus productivity. Intracellular ATP or total adenylate concentration at the point of infection may be used as a predictor of viral yield in bioprocesses designed for virus/vaccine production.
Department of Microbiology, Buller Bldg., University of Manitoba, Winnipeg, Manitoba R3T 2N2, Canada.
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Burgener
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Coombs
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Butler
D000249:Adenosine Monophosphate / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D002455:Cell Division; D002522:Chlorocebus aethiops; D002851:Chromatography, High Pressure Liquid; D003470:Culture Media; D017930:Genes, Reporter; D005973:Glutamine; D007700:Kinetics; D008156:Luciferases / Q000235:genetics; D012087:Reoviridae / Q000254:growth & development* / Q000302:isolation & purification*; D014709:Vero Cells / Q000821:virology*
D003470:Culture Media; D005973:Glutamine; D000249:Adenosine Monophosphate; D000255:Adenosine Triphosphate; D008156:Luciferases