Comparative production of recombinant capripoxvirus in Vero cells using microcarrier- and macrocarrier-based dynamic cell culture systems under serum-free conditions

Brucellosis remains one of the most significant zoonotic diseases worldwide, and the absence of licensed human vaccines highlights the need for novel vaccine platforms and reproducible laboratory-scale manufacturing strategies. In the present study, the replication efficiency of a recombinant capripoxvirus expressing a Brucella antigen was evaluated in Vero cells cultured under serum-free conditions using two 3D carrier-based cultivation systems. Vero cells were cultivated in serum-free medium either on Cytodex 1 microcarriers in a 500-mL reusable spinner flask operated in repeated-batch mode or on BioNOC II® macrocarriers in single-use 500-mL BelloCell™ 500AP vessels integrated into the BelloStage™-3000 system and operated according to the Tide Motion principle with medium recirculation. Cells were infected with recombinant SPPV(TKΔ)-OMP16 at a multiplicity of infection of 0.1, and total cell yield, metabolic parameters, and virus production were subsequently monitored throughout cultivation. The microcarrier and macrocarrier systems achieved maximum total cell yields of 7.0 × 10⁸ and 3.1 × 10⁹ cells, respectively. Peak virus titers reached 5.75 log₁₀ TCID₅₀/mL at 120 h post-infection in the microcarrier system and 7.25 log₁₀ TCID₅₀/mL at 168 h post-infection in the macrocarrier system, corresponding to a 1.5 log₁₀ increase in virus titer (p < 0.01). After normalization to the working culture volume, the macrocarrier system exhibited 4.4-fold higher volumetric cell productivity. In addition, virus productivity normalized to the total number of cells present at the time of infection was approximately sevenfold higher than that observed in the microcarrier system. These findings demonstrate that the macrocarrier-based dynamic cultivation system enables superior total cell yield and enhanced recombinant virus production under serum-free laboratory-scale conditions. KEY POINTS: • BioNOC II® macrocarriers increased volumetric cell productivity by 4.4-fold compared with Cytodex 1. • BioNOC II® macrocarriers increased peak virus titers by 1.5 log₁₀ TCID₅₀/mL. • Cell-specific viral productivity was approximately sevenfold higher in the macrocarrier-based bioreactor system.