Figure 5.

Mutations FASII, Lipoate Synthesis, and Biotin Metabolic Pathways Affect Liver-Stage Development

(A) Pathway maps for FAS II, lipoate metabolism, and biotin metabolism in the Plasmodium apicoplast. See Table S4 for gene IDs, enzyme functions, and reactions. Pep, phosphoenolpyruvate; Pyr, pyruvate; Ac-CoA, acetyl-CoA; Mal-CoA, malonyl-CoA; Mal-ACP, malonyl-[acp]; Acetoac-ACP, acetoacetyl-[acp]; Octanoyl-ACP, octanoyl-[acp]; Octanoyl-E2, protein N6-(octanoyl)lysine; Lipoyl-E2, protein N6-(lipoyl)lysine.

(B) Schematic representation of phenotypes of single knockout (KO) mutants. Green, phenotype not significantly different from wild type (WT) parasites. Red, phenotype significantly different (>2-day delay in pre-patent period).

(C) Size of 250 cultured EEFs (48 hpi) per mutant; median and interquartile ranges are shown in red. ^∗ = p < 0.05 by Kruskal-Wallis test.

(D) Relative maturation of EEFs measured as conversion of infected host cells to detached cells at 48 hpi. Error bars show standard deviations from 8 biological replicates (for PDH-E2) or 3 biological replicates (all other mutants). The results were statistically evaluated by a one-way analysis of variance (ANOVA) test with Dunnet’s multiple comparisons (^∗∗p ≤ 0.01; ^∗∗∗p ≤ 0.001).

(E) The number of mice that developed blood-stage infections after injection of 5,000 mutant sporozoites and the mean delay (range) in pre-patency compared to mice infected with WT sporozoites. ^∗, gene KO mutants with a significantly “slow fitness” blood stage phenotype (Bushell et al., 2017). See Figure S4B for plots showing the course of blood stage infections after sporozoite injection.