The emergence of artemisinin resistance in Southeast Asia imperils efforts to reduce the global malaria burden. We genetically modified the Plasmodium falciparum K13 locus using zinc-finger nucleases and measured ring-stage survival rates after drug exposure in vitro; these rates correlate with parasite clearance half-lives in artemisinin-treated patients. With isolates from Cambodia, where resistance first emerged, survival rates decreased from 13 to 49% to 0.3 to 2.4% after the removal of K13 mutations. Conversely, survival rates in wild-type parasites increased from ≤0.6% to 2 to 29% after the insertion of K13 mutations. These mutations conferred elevated resistance to recent Cambodian isolates compared with that of reference lines, suggesting a contemporary contribution of additional genetic factors. Our data provide a conclusive rationale for worldwide K13-propeller sequencing to identify and eliminate artemisinin-resistant parasites.

The level of human group IIA secreted phospholipase A2 (hGIIA sPLA2) is increased in the plasma of malaria patients, but its role is unknown. In parasite culture with normal plasma, hGIIA is inactive against Plasmodium falciparum, contrasting with hGIIF, hGV, and hGX sPLA2s, which readily hydrolyze plasma lipoproteins, release nonesterified fatty acids (NEFAs), and inhibit parasite growth. Here, we revisited the anti-Plasmodium activity of hGIIA under conditions closer to those of malaria physiopathology where lipoproteins are oxidized. In parasite culture containing oxidized lipoproteins, hGIIA sPLA2 was inhibitory, with a 50% inhibitory concentration value of 150.0 ± 40.8 nM, in accordance with its capacity to release NEFAs from oxidized particles. With oxidized lipoproteins, hGIIF, hGV, and hGX sPLA2s were also more potent, by 4.6-, 2.1-, and 1.9-fold, respectively. Using specific immunoassays, we found that hGIIA sPLA2 is increased in plasma from 41 patients with malaria over levels for healthy donors (median [interquartile range], 1.6 [0.7 to 3.4] nM versus 0.0 [0.0 to 0.1] nM, respectively; P < 0.0001). Other sPLA2s were not detected. Malaria plasma, but not normal plasma, contains oxidized lipoproteins and was inhibitory to P. falciparum when spiked with hGIIA sPLA2. Injection of recombinant hGIIA into mice infected with P. chabaudi reduced the peak of parasitemia, and this was effective only when the level of plasma peroxidation was increased during infection. In conclusion, we propose that malaria-induced oxidation of lipoproteins converts these into a preferential substrate for hGIIA sPLA2, promoting its parasite-killing effect. This mechanism may contribute to host defense against P. falciparum in malaria where high levels of hGIIA are observed.

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