Optimization of Fermentation Conditions Using Design of Experiment (DoE) to Enhance the Production of Antigenic Capsular Polysaccharides From <i>Streptococcus pneumoniae</i> Serotype 18C for Pneumococcal Vaccine Development

Pneumococcal pneumonia, caused by Streptococcus pneumoniae, has emerged as a global health threat, especially in children below 5 years of age and elderly population. S. pneumoniae remains a major global health threat which necessitates scalable and high-yield vaccine production. This study presents an innovative fermentation platform for S. pneumoniae (serotype 18C) by optimizing an already refined media composition using data-driven Design of Experiment (DoE) approach. Unlike conventional applications, Plackett-Burman design (PBD) was strategically employed beyond screening, identifying statistically significant parameters and the interaction between key factors of dextrose, MgSO₄, pH, and aeration. Results reveal that pH level of 7.1 significantly enhances the antigenic capsular polysaccharide yield. Further, ANOVA analysis confirmed a strong synergistic effect between dextrose and MgSO₄. In addition, the present study also finds that fermentation at lower level of aeration (0.2 vvm) favors the bacterial growth aligning with S. pneumoniae's facultative anaerobic or microaerophilic nature. The downstream processing integrating centrifugation and diafiltration (100 kDa) improves the product recovery and purity reinforcing the process efficiency. The unconventional application of PBD combined with systematic optimization of refined process sets a new benchmark for pneumococcal fermentation strategies. The findings demonstrate that precise modulation of fermentation conditions can enhance the vaccine antigen yield addressing the scalability challenges in industrial production. Future work may focus on extending this DoE-based platform to other serotypes of S. pneumoniae and refining fermentation kinetics for next generation pneumococcal vaccine. This study not only challenges the traditional experimental set up, but paves the way for adaptive, data-driven bioprocess innovations in vaccine manufacturing.