Immunogenic insights and experimental validation of a multiepitope vaccine construct against Streptococcus pneumoniae

<i>Streptococcus pneumoniae</i> is a predominant cause of pneumonia, sinusitis, bacteremia, meningitis and otitis media especially in children and the elderly. The emergence of antibiotic resistance and shift towards non-vaccine serotypes has compromised the efficacy of the treatment regime and available vaccines. The available polysaccharide-based vaccines confer protection against limited serotypes. In this study, we have conducted the experimental validation of a promising multi epitope vaccine candidate. The vaccine construct was cloned into pET-28a vector and expressed in <i>Escherichia coli</i>. The expression of the protein was induced, and its purification was carried out by the Ni-NTA affinity chromatography. Further, purified multiepitope protein was subjected to the SDS-PAGE and western blot analysis. The secondary structure was determined by circular dichroism spectroscopy, and it was observed to closely align with the predicted structure by Alpha fold. The hemolysis assay confirmed its safety and tolerability. The BALB/c mice was administered with this multiepitope vaccine construct and immune response was evaluated. The antibody response was estimated and confirmed by western blot and ELISA analysis. The cytokine profiling assay revealed the involvement of interleukins, TNF-α, and GM-CSF. Together, this study provides the first experimental validation of a computationally designed multi-epitope vaccine against <i>S. pneumoniae</i>, demonstrating antibody titers, broad cytokine response, and absence of hemolytic toxicity.