Transgenic Plasmodium berghei expressing P. vivax pre-erythrocytic antigens SSP3 and SPECT1 to elucidate their efficacy towards vaccine development

The development of an efficacious vaccine against Plasmodium vivax malaria remains a top global health priority. P. vivax forms hypnozoites, dormant forms of the parasite in the liver that can become reactivated after a few months or even years, causing relapse malaria and enhancing transmission. Therefore, targeting antigens expressed during the pre-erythrocytic stages offers the potential to prevent clinical malaria from both primary and relapse infections. The circumsporozoite protein (CSP) is the leading pre-erythrocytic vaccine candidate and forms the basis for the current P. falciparum malaria vaccines RTS, S and R21/Matrix M. Studies have shown that in P. vivax , CSP attributes limited protection, highlighting the need to identify additional pre-erythrocytic antigens to improve vaccine efficacy. Evaluation of new candidates often requires human volunteers, making initial screening complex and highlighting the need for robust preclinical models. In this study, we generated transgenic P. berghei parasite lines as research tools to investigate the potential efficacy of two P. vivax pre-erythrocytic vaccine candidates, SSP3 and SPECT1. The chimeric Pb-Pv SSP3 and Pb-Pv SPECT1 lines showed normal development throughout the parasite life cycle, in both mice and mosquitoes. Antisera raised against recombinant Pv SSP3 and Pv SPECT1 using TitreMax Gold adjuvant exhibited reduction in sporozoite invasion of hepatocytes in vitro , with mean inhibitions of 27% and 14%, respectively. In vivo sporozoite challenge studies showed sterile protection in 10% of mice immunized with r Pv SPECT1, whereas r Pv SSP3 provided no detectable protection. Although substantial protection was not observed for either antigen under the conditions tested, the transgenic P. berghei lines generated in this study serve as useful platforms for evaluating the efficacy of these antigens as vaccine candidates across different delivery systems and adjuvanted formulations.