Fig. 6: Functional analysis of potential resistant genes in parasite RSA and the effects of PfMYST knockdown on various antimalarial RSA.

a Ten potential resistant genes in response to DHA treatment are functional validated by episomal over-expression (OE), knockout (KO) or knockdown (KD). The overall results of RSA_0-3h assays of these transgenic lines are summarized in the last column. b Recrudescence Assay of PHISTb_KO, 2G4_KO, PHISTa_KO, DDX5_KO with the parent 3D7 line of transfection as control. Ring-stage parasites were exposed to three 6 h pulses at 700 nM DHA. Recrudescence curves were obtained based on parasitemia determined from Giemsa-stained smears. Data were presented as mean ± SEM from three independent experiments with technical duplicates. c,d RSA_0-3h for over-expression (OE), knockout (KO) or knockdown (KD) of “PfMYST-targeted genes”. Two OE, four KO and four KD lines were tested for their RSA_0-3h under 700 nM DHA treatment with WT 3D7 strain as a negative control. Data were presented as mean ± SEM from four independent experiments with technical duplicates. Statistical comparisons between vehicle control and GlcN-treated samples were conducted using a two-tailed student’s t test. Statistical comparisons between WT and mutant parasites were conducted using One-way ANOVA with Bonferroni correction, and the exact P-value was provided in Source Data. (*P < 0.05; ** P < 0.01; *** P < 0.001). e The impact of PfMYST knockdown on RSA_0-3h for seven antimalarial drugs, including AQ, PYR, CQ, PYA, LMF, MEF, and PPQ. RSA_0-3h was modified according to the IC₅₀ of each drug, with drug concentrations set at 10 x IC₅₀. Survival rates were calculated using the standard method. Data were presented as mean ± SEM from four independent experiments with technical duplicates. Statistical comparisons between vehicle control and GlcN-treated samples were conducted using a two-tailed student’s ttest. AQ: Amodiaquine, PYR: Pyrimethamine, CQ: Chloroquine, PYA: Pyronaridine, LMF: Lumefantrine, MEF: Mefloquine, PPQ: Piperaquine. f Relationship between the alerted resistance level and pharmacokinetic parameters, including drug half-life and T_max. Drugs with increased/decreased resistance were displayed in red/green circles, respectively. Source data are provided as a Source Data file.