Abstract. The simian parasite Plasmodium knowlesi is now the commonest cause of malaria in Malaysia and can rapidly cause severe and fatal malaria. However, microscopic misdiagnosis of Plasmodium species is common, rapid antigen detection tests remain insufficiently sensitive and confirmation of P. knowlesi requires polymerase chain reaction (PCR). Thus available point-of-care diagnostic tests are inadequate. This study reports the development of a simple, sensitive, colorimetric, high-throughput loop-mediated isothermal amplification assay (HtLAMP) diagnostic test using novel primers for the detection of P. knowlesi. This assay is able to detect 0.2 parasites/μL, and compared with PCR has a sensitivity of 96% for the detection of P. knowlesi, making it a potentially field-applicable point-of-care diagnostic tool.Human infection with the simian malarial parasite, Plasmodium knowlesi results from transmission from long-tailed and pigtailed macaques, via the Anopheles leucosphyrus mosquito vector, 1 and has been documented in all countries of southeast Asia, except Laos and Timor Leste.1 In Sabah, Malaysia, while the total number of reported malaria cases has decreased since 1992, the absolute number and proportion due to P. knowlesi have increased significantly such that in 2013, 62% of malaria notifications were due to P. knowlesi, making it the most common cause of malaria in this region.
2Plasmodium knowlesi malaria is difficult to diagnose by conventional microscopy. Ring and trophozoite stages of P. knowlesi appear morphologically very similar to both Plasmodium falciparum and Plasmodium malariae.