Cost-effective production of vi polysaccharide through high-cell-density fermentation using animal component-free...

Salmonella enterica serovar Typhi remains a major public health concern that requires an effective strategy to produce vaccines. This study reports the development of a high cell density fermentation (HCD) process to increase the production of Vi- polysaccharide (ViPS). The strain has been confirmed by means of phenotypic, biochemical (VITEK 2), and serological methods. Media optimization identified soybean peptone as a cost-effective and non-animal component alternative that promotes robust growth. The optimized fed-batch fermentation process increased the cell density from OD₆₀₀ 19-20 under low-cell-density conditions to OD₆₀₀ 39-42.1 under high-cell-density conditions, resulting in an approximately 1.7-fold increase in Vi polysaccharide (ViPS) yield i.e., from 165-187.5 mg/L to 283.75-302.5 mg/L. A scaling to 30 L fermentation tanks yielded comparable yields (290-320 mg/L), demonstrating the robustness of the process. Thermal inactivation at 65 °C for 30 min achieved complete cell inactivation without compromising the integrity of the polysaccharide, as predicted by the predictive model (R² = 0.9947). NMR analysis confirms the structural integrity. The purified ViPS met WHO specifications, indicating suitability for cost-effective and scalable typhoid vaccine production.