3.1 Batch fermentation
Batch cultivation was conducted at 10 L bioreactor as previously described for the production of cGMP lots at 60 L (Gonçalves et al., 2014). Cells were harvested at the same OD ~ 6.0, to evaluate process reproducibility, downscale methods and have a sample product as a standard. Furthermore, batch fermentation was conducted to the highest OD to verify if harvesting at a higher cell density would interfere with vaccine quality. Figure 2 shows that the highest OD was 9.0-10.0; at this time-point (5 h) all glucose had been consumed. Dry cell weight was 4.15 ± 0.33 g/L at the highest OD, higher than the 1.26 g/L yield obtained in the culture of encapsulated strain 23F for polysaccharide production (Gonçalves et al., 2002) and the 1.13 g/L yield of strain 14 (Leal et al., 2011). The disruption of thelytA gene (which encodes for one of the autolysins of pneumococcus) in the strain used in this study has contributed to the ability to achieve higher biomass production. At the highest OD, lactate production was 19.0 ± 0.7 g/L and acetate production was 10.3 ± 1.1 g/L, values that were similar to other pneumococcal strains (Gonçalves et al., 2002; Gogola-Kolling et al., 2014). At 3.5 h, when cultures reached OD 6.0, the concentration of lactate and acetate were about 10.0 g/L and 4.5 g/L, respectively, similar to cGMP conditions at 60 L (Gonçalves et al., 2014).