UsingPlasmodium knowlesias a model for screeningPlasmodium vivaxblood-stage malaria vaccine targets reveals new candidates

Plasmodium vivaxis responsible for the majority of malaria cases outside Africa. UnlikeP. falciparum, theP. vivaxlife-cycle includes a dormant liver stage, the hypnozoite, which can cause infection in the absence of mosquito transmission. An effective vaccine againstP. vivaxblood stages would limit symptoms and pathology from such recurrent infections, and therefore could play a critical role in the control of this species. Vaccine development inP. vivax, however, lags considerably behindP. falciparum, which has many identified targets with several having transitioned to Phase II testing. By contrast only oneP. vivaxblood-stage vaccine candidate based on the Duffy Binding Protein (PvDBP), has reached Phase Ia, in large part because the lack of a continuousin vitroculture system forP. vivaxlimits systematic screening of new candidates. We used the close phylogenetic relationship betweenP. vivaxandP. knowlesi, for which anin vitroculture system in human erythrocytes exists, to test the scalability of systematic reverse vaccinology to identify and prioritiseP. vivaxblood-stage targets. A panel ofP. vivaxproteins predicted to function in erythrocyte invasion were expressed as full-length recombinant ectodomains in a mammalian expression system. Eight of these antigens were used to generate polyclonal antibodies, which were screened for their ability to recognize orthologous proteins inP. knowlesi. These antibodies were then tested for inhibition of growth and invasion of both wild typeP. knowlesiand chimericP. knowlesilines modified using CRISPR/Cas9 to exchangeP. knowlesigenes with theirP. vivaxorthologues. Candidates that induced antibodies that inhibited invasion to a similar level as PvDBP were identified, confirming the utility ofP. knowlesias a model forP. vivaxvaccine development and prioritizing antigens for further follow up.AUTHOR SUMMARYMalaria parasites cause disease after invading human red blood cells, implying that a vaccine that interrupts this process could play a significant role in malaria control. MultiplePlasmodiumparasite species can cause malaria in humans, and most malaria outside Africa is caused byPlasmodium vivax. There is currently no effective vaccine against the blood stage of any malaria parasite, and progress inP. vivaxvaccine development has been particularly hampered because this parasite species cannot be cultured for prolonged periods of time in the lab. We explored whether a related species,P. knowlesi, which can be propagated in human red blood cellsin vitro, can be used to screen for potentialP. vivaxvaccine targets. We raised antibodies against selectedP. vivaxproteins and testedtheir ability to recognize and preventP. knowlesiparasites from invading human red blood cells, thereby identifying multiple novel vaccine candidates.