Delivery of chicken egg ovalbumin to dendritic cells by listeriolysin O-secreting vegetative Bacillus subtilis

Listeriolysin O (LLO), one of the most immunogenic proteins of Listeria monocytogenes and its main virulence factor, mediates bacterial escape from the phagosome of infected cell. Thus, its expression in non-pathogenic bacterial host may enable effective delivery of heterologous antigens to the host cell cytosol and lead to their processing predominantly through the cytosolic MHC class I presentation pathway. The aim of this project was to characterize a delivery of a model antigen, chicken egg ovalbumin (OVA) to the cytosol of dendritic cells by recombinant Bacillus subtilis vegetative cells expressing LLO. Our work indicates that LLO produced by non-sporulating, vegetative bacteria was able to support OVA epitope presentation by MHC I molecules on the surface of antigen presenting cells and consequently influenced OVA-specific cytotoxic T cells activation. Additionally, it was proven that genetic context of epitope sequence is of great importance, as only native, full-sequence OVA fused to N-terminal fragment of LLO was sufficient for effective epitope delivery and activation of CD8+ lymphocytes. These results demonstrate necessity for further verification of a fusion antigen potency of enhancing the MHC I presentation and they prove that LLO producing B. subtilis may represent a novel and attractive candidate for a vaccine vector.