Since 2014, an Asian lineage of Zika virus has caused outbreaks, and it
has been associated with neurological disorders in adults and congenital
defects in newborns. The resulting threat of the Zika virus to human
health has prompted the development of new vaccines, which have yet to
be approved for human use. Vaccines based on the attenuated or
chemically inactivated virus will require large-scale production of the
intact virus to meet potential global demands. Intact viruses are
produced by infecting cultures of susceptible cells, a dynamic process
that spans from hours to days and has yet to be optimized. Here, we
infected Vero cells adhesively cultured in well-plates with two Zika
virus strains: a recently isolated strain from the Asian lineage, and a
cell-culture-adapted strain from the African lineage. At different time
points post-infection, virus particles in the supernatant were
quantified; further, microscopy images were used to quantify cell
density and the proportion of cells expressing viral protein. These
measurements were performed across multiple replicate samples of
one-step infections every four hours over 60 hours and for multi-step
infections every four to 24 hours over 144 hours, generating a rich
dataset. For each set of data, mathematical models were developed to
estimate parameters associated with cell infection and virus production.
The African-lineage strain was found to produce a 14-fold higher yield
than the Asian-lineage strain in one-step growth and a 7-fold higher
titer in multi-step growth, suggesting a benefit of cell-culture
adaptation for developing a vaccine strain. We found that image-based
measurements were critical for discriminating among different models,
and different parameters for the two strains could account for the
experimentally observed differences. An exponential-distributed delay
model performed best in accounting for multi-step infection of the Asian
strain, and it highlighted the significant sensitivity of virus titer to
the rate of viral degradation, with implications for optimization of
vaccine production. More broadly, this work highlights how image-based
measurements can contribute to discrimination of virus-culture models
for the optimal production of inactivated and attenuated whole-virus
vaccines.
89. Adeno-Associated Virus Production Using a Disposable Fixed-Bed Bioreactor From Bench-Scale To Industrial Scale