Identification of Immune Responses to Japanese Encephalitis Virus Specific T Cell Epitopes

Abstract

Background: Due to the similarity between the dengue (DENV) and the Japanese encephalitis virus (JEV) there is potential for immune cross-reaction. We sought to identify T cell epitopes that are specific to JEV and do not cross react with DENV.Methodology: 20mer peptides were synthesized from regions which showed >90% conservation. Using IFNγ cultured ELISpot assays, we investigated JEV-specific T cell responses in DENV − and JEV − non-immune individuals (DENV − JEV − = 21), JEV seronegative and had not received the JE vaccine, but who were DENV seropositive (DENV + JEV − = 22), JEV + (seropositive for JEV and had received the JE vaccine), but seronegative for DENV (DENV − JEV + = 23). We further assessed the responses to these peptides by undertaking ex vivo IFNγ assays and flow cytometry.Results: None of DENV − JEV − individuals responded to any of the 20 JEV-specific peptides. High frequency of responses was seen to 6/20 peptides by individuals who were JEV + but DENV − , where over 75% of the individuals responded to at least one peptide. P34 was the most immunogenic peptide, recognized by 20/23 (86.9%) individuals who were DENV − JEV + , followed by peptide 3 and peptide 7 recognized by 19/23 (82.6%). Peptide 34 from the NS2a region, showed <25% homology with any flaviviruses, and <20% homology with any DENV serotype. Peptide 20 and 32, which were also from the non-structural protein regions, showed <25% homology with DENV. Ex vivo responses to these peptides were less frequent, with only 40% of individuals responding to peptide 34 and 16-28% to other peptides, probably as 5/6 peptides were recognized by CD4+ T cells.Discussion: We identified six highly conserved, T cell epitopes which are highly specific for JEV, in the Sri Lankan population. Since both JEV and DENV co-circulate in the same regions and since both JE and dengue vaccines are likely to be co-administered in the same geographical regions in future, these JEV-specific T cell epitopes would be useful to study JEV-specific T cell responses, in order to further understand how DENV and JEV-specific cellular immune responses influence each other.

Development and validation of an assay for detection of Japanese encephalitis virus specific antibody responses

“…The specificity of these JEV peptides was determined by using Clustal Omega of European Bioinformatics Institute (EBI) ( www.ebi.ac.uk ) to confirm that they did not significantly cross-react with the DENV ( S1 Table ). Out of these 36 peptides, only 22 peptides were successful in the synthesis with 90% purity (Genscript USA) and were used for further analysis as previously described [ 17 ].…”