Recombinant Bacillus subtilis spores expressing SARS-CoV-2 spike protein induced humoral, mucosal, and cellular immunity in mice

The COVID-19 pandemic greatly affected global health and economies. While vaccines have helped reduce mortality from the disease, the inability to prevent its transmission hinders full eradication. Mucosal vaccines capable of preventing virus transmission, hence, are urgently needed. A possible approach involves using Bacillus subtilis, a safe Gram-positive bacterium, as a bacterial spore vaccine delivery platform. Using this approach, the SARS-CoV-2 spike protein is engineered to be expressed in B. subtilis during bacterial sporulation. The recombinant bacterial spores, when used to immunize Balb/c mice, triggered significant immune responses. Two weeks after the second dose, mice showed elevated levels of SARS-CoV-2 spike-specific IgM, IgG, and sIgA. The immunization also boosted CD4⁺ and CD8⁺ T cell responses. Pro-inflammatory cytokines, such as IFN-γ, IL-2, and TNF-α, as well as anti-inflammatory cytokines like IL-10, were also upregulated upon restimulation with spike protein peptides. Additionally, serum from the immunized mice showed neutralizing activity against SARS-CoV-2. These findings suggest that B. subtilis spores expressing the SARS-CoV-2 spike protein can induce both humoral and cellular immunity, making them a promising platform for developing mucosal vaccines against SARS-CoV-2.