Co-expression of Japanese encephalitis virus prM–E–NS1 antigen with granulocyte-macrophage colony-stimulating factor enhances humoral and anti-virus immunity after DNA vaccination

Abstract Japanese encephalitis virus (JEV) is an agent of Japanese encephalitis, and granulocyte-macrophage colony-stimulating factor (GM-CSF) is an attractive DNA vaccine adjuvant for its antigen presentation. In the present study, we have constructed DNA vaccines that carried JEV prM–E–NS1 genes with or without the GM-CSF gene. Immunization with the bicistronic plasmid pCAG-JEGM that co-expresses GM-CSF and viral prM–E–NS1, resulted in the highest IgG response and sufficient protection against virus-challenged BALB/c mice. However, much to our surprise, co-inoculation of the GM-CSF plasmid with the pCAG-JE plasmid expressing viral prM–E–NS1 lead to a low antibody titer and a relatively low survival rate. Moreover, anamnestic antibody-mediated protection played a dominant role in the mice JEV challenge model, according to the enhancement of post-challenge neutralizing antibody titers and further adoptive transfer experiments. Taken together, this study should encourage further development of JEV DNA vaccine strategies and caution against the use of cytokines as an adjuvant.