TOLC-DERIVED PAN-RICKETTSIAL PEPTIDE VACCINES ELICIT B- AND T- CELL IMMUNE RESPONSES

Abstract

Intro: The putative TolC-derived antigenic epitope may trigger an effective immune response and prove to be a novel vaccine that protects against all rickettsial infections. Methods: A total of 421 healthy volunteers were recruited and tested for the Typhus group-, Spotted Fever group- and Scrub Typhus- specific IgG ELISA. Epitopes that are highly immunogenic and conserved across groups of rickettsial species were identified through in silico approaches. An in-house ELISA was developed using identified B cell peptide as antigen and the serum was tested for peptide-specific pre-existing antibodies. The MHC I-presented peptides are screened for their T-cell stimulation using human PBMCs isolated from the buffy coat using Ficoll Paque. After seeding, the cells were stimulated in RPMI media with TolC-peptides and phytohemagglutinin as control. Following incubation at 37°C with 5% CO2, the supernatant was measured for an antigen-specific relative increase in IFN-γ using real-time RT-PCR and ELISA. Findings: Among the peptide epitopes screened, B-cell epitope IYPEGGAQYSRIRSAKNQTRNSA/VVQ and T-cell epitope KLYEAKITR were conserved across three rickettsial groups. Of the 415 samples tested, 266 were positive for any of the three rickettsial IgG. In-house peptide-specific ELISA indicated the presence of pre-existing antibodies in 167 samples against identified TolC B-cell peptide epitopes. In TolC T-cell peptide epitope-treated PBMC cultures, a significant relative increase in IFN-γ was observed by real-time RT-PCR and ELISA compared to mock-treated cultures. The ELISA method yielded a high number of readouts, however, the two methods showed a significant correlation. Discussion: Rickettsial pathogens need a combination of B- cell and T-cell immunity for considerable protection against them. The peptide epitopes induced antibody response and IFN-γ by human PBMCs. Conclusion: The B- and T-cell epitopes identified from the TolC protein were highly conserved and immunogenic. This could serve as a potential vaccine for protection against both the spotted fever group and typhus group of rickettsial pathogens.