Fig. 8

FNT-1 is dispensable in wild-type T. gondii but becomes highly fitness-conferring in ΔBCKDH parasites. a Table highlighting the conservation of formate/nitrite transporters (FNTs) across apicomplexans and their fitness score from a recent screen fitness screen of metabolic genes. b Plaque assay, testing the fitness during the lytic cycle of RH parasites and mutant parasites lacking BCKDH, FNT-1 or both. c Quantification of plaque area size comparing different strains: RH, ΔBCKDH, ΔFNT-1 and ΔBCKDH/ΔFNT-1 double-KO parasites. Error bars represent the standard deviation between 3 independent infections. Per infection, the areas of > 20 plaques were quantified, and statistically significant differences were determined by a t-test comparing the mutants as indicated (n.s., non-significant; ***p < 0.001). d Intracellular growth assay of RH and mutant parasites over 36 h. Error bars represent the standard deviation between 3 independent infections. Per infection, > 100 vacuoles were counted. e, f Metabolomic analysis of T. gondii RH and mutant parasites (ΔBCKDH, ΔFNT-1 and ΔBCKDH/ΔFNT-1). Levels of intracellular metabolites relative to levels in RH (dashed line) (e) and of metabolites secreted into the medium by purified parasites normalised to valine, an essential amino acid present in the culture medium at 0.8 mM (f). Error bars in e and f represent the standard deviation between replicates (n = 4). Statistically significant differences between each mutant and RH were assessed using a t-test and are indicated (*p < 0.05; **p < 0.005; ***p < 0.001). BCKDH, branched-chain α-keto acid dehydrogenase-complex; Glc, glucose; Hex-P, hexose-phosphate; Lac, lactate; Cit, citrate; Suc, succinate; Mal, malate; Glu, glutamate; Asp, aspartate; Ala, alanine; Val, valine; Leu, leucine; Gly, glycine; Ser, serine; Thr, threonine; MyIno, myo-inositol; MyIno-P, myo-inositol-phosphate; SDHP, sedoheptulose-7-phosphate; Pyr, pyrimidine; Ino, inosine