Development of an experimental human blood-stage model for studying Plasmodium knowlesi

BackgroundThe zoonotic parasite Plasmodium knowlesi is an increasing cause of human malaria in Southeast Asia. We aimed to develop a P. knowlesi induced blood-stage malaria (IBSM) model to enable further study of parasite biology, host responses to infection, and activity of antimalarial treatments. MethodsWe manufactured and characterised a P. knowlesi parasite bank using the P. knowlesi YH1 strain adapted to grow in human serum. This P. knowlesi bank was evaluated in an IBSM study involving four healthy adults intravenously inoculated with P. knowlesi-infected erythrocytes using a dose escalation strategy (10-fold increases from ~2,800 viable parasites). Parasitemia was monitored by qPCR. All participants received curative treatment with artemether-lumefantrine. Endpoints included safety and infectivity of the P. knowlesi parasite bank. FindingsFour participants were inoculated. Two participants became infected. Participant two (administered ~28,000 viable parasites) had detectable parasites (~6 parasites/mL) immediately prior to protocol-defined treatment on day 21. Participant three (administered ~280,000 viable parasites) developed detectable parasites on day 17, peaking at 202 parasites/mL just after protocol-defined treatment on day 24. The parasite multiplication rate over 32 hours (PMR32) was 2{middle dot}33 (95% CI 1{middle dot}66 - 3{middle dot}25). Adverse events were mostly mild and unrelated to the challenge agent. InterpretationA P. knowlesi YH1-HS parasite bank suitable for use in human volunteers was successfully manufactured; however, infectivity was suboptimal. Further optimisation of the P. knowlesi parasite bank and/or study design will be required to improve infectivity. FundingNational Health and Medical Research Council of Australia and the Royal Brisbane and Womens Hospital Foundation. Research in contextO_ST_ABSEvidence before this studyC_ST_ABSSeveral studies conducted during the 1930s through to the 1960s report the results of P. knowlesi human infection studies. This includes the first experimental infection of three individuals in 1931, followed by a number of studies reporting the use of P. knowlesi as malariotherapy for neurosyphilis. Several additional studies involving the experimental infection of humans with P. knowlesi were conducted in the 1960s, following the first reported case of naturally acquired P. knowlesi malaria. No P. knowlesi human infection studies have been conducted since the 1960s. Added value of this studyIn this study we developed a Good Manufacturing Practice (GMP)-compliant P. knowlesi parasite bank suitable for use in human volunteers. This involved adapting the P. knowlesi YH1 strain to growth in human serum, resulting in the development of a P. knowlesi line (YH1-HS) which can be readily cultured under the same conditions routinely used for P. falciparum. We next tested the P. knowlesi YH1-HS parasite bank in four healthy volunteers using a dose escalation study. Two participants became infected. Implications of all the available evidenceThe development of a reliable and reproducible P. knowlesi volunteer infection study would be an important advance for P. knowlesi research. While this study resulted in the development of the first GMP-compliant P. knowlesi parasite bank, infectivity was suboptimal, suggesting that further optimisation of the P. knowlesi parasite bank and/or study design will be required, as well as further investigation into host and/or parasite factors that may impact infectivity.