THE B SUBUNIT OF Escherichia coli TOXIN LT FUSED WITH TRUNCATED rROP2 PROTEIN OF Neospora caninum INDUCES MIXED Th2/Th17 IMMUNE RESPONSE IN MICE

Abstract

Neospora caninum ROP2 protein is a promising vaccine antigen candidate for controlling neosporosis. In this study, we evaluated the immune response against a chimera formed by Escherichia coli heat labile B portion of the toxin (LTB) and its association with a truncated N. caninum ROP2 in a murine model. The recombinant proteins were expressed in E. coli, maintaining antigenic epitopes that were recognized by specific antibodies. Antigenicity was verified by western blot using serum from naturally infected cattle, and by ELISA using serum from mice infected experimentally with N. caninum. Immunogenicity was evaluated by total serum IgG, IgG1 and IgG2a isotypes in mice vaccinated with the experimental vaccines. Splenocyte transcription of the cytokines IL-10, IL-12, IL-17A and IFN-y were analyzed by qPCR. The chimeric construction rLTB/tROP2 modulated a significant increase (p<0.05) in total antibody titer against rROP2 by day 14 post inoculation, whereas the rROP2 induced higher (p<0.05) total IgG after the vaccine boost until the end of the experiment. Splenocytes from both groups stimulated with the rROP2 induced significant (p<0.05) higher IL-10 (18-27 folds), and IL-12 (7-12 folds) transcription, with a significant (p<0.0001) fold transcription increase for IL-17 (40-3000 folds). When stimulated with rLTB/tROP2, a significant (p<0.05) higher IL-10 (46-76 folds), IL-12 (4-25 folds) and IL-17 (40-400 folds) transcription were observed. Our results exhibit evidence that these recombinant proteins were able to modulate a mixed Th2/Th17 immune response, suggesting they may be a promising vaccine to be used for the control of N. caninum.