FIG 1.

Influence of supplemental oleate and palmitate on Toxoplasma replication and development. (A) Toxoplasma development in HFF assessed by plaque assays for 5 days with OA at the indicated concentrations. Data show histograms of the mean area of plaques ± SD from 4 independent experiments. *, P < 0.002, Student's t test. (B) Toxoplasma replication in HFF assessed by tritiated uracil incorporation in the presence of OA at the indicated concentrations at 24 h p.i. Data are percentages ± SD relative to the value for the control (to which no fatty acids were added, the value for which was set as 100%) for biological triplicates with experimental triplicates. *, P < 0.05, Student's t test. (C) Toxoplasma replication assessed by PV size at 24 h p.i. in the presence of OA, which was added to the medium at concentrations ranging from 0 to 0.5 mM. The numbers of parasites per PV in Toxoplasma-infected HFF are displayed as percentages for all PV. Data show the means ± SD for at least 1,000 randomly selected vacuoles from 3 independent experiments. P values were determined by Student's t test. *, P < 0.001; **, P < 0.0001. (D) Transmission EM of intracellular Toxoplasma incubated in the presence of 0.4 mM OA. PV containing 8, 4, 2, or 1 parasite are shown. In addition to LD, osmiophilic membranous structures (red arrowheads) in the residual body (RB), cytoplasm, or PV lumen were detected. hc, host cell; P, parasite. Bars, 250 nm. (E and F) Fluorescence microscopy of HFF infected with RFP-expressing Toxoplasma for 24 h in the presence of 0.5 mM OA, fixed, and stained with BODIPY 493/503. (E) Representative PV for at least 50 infected cells viewed at the same magnification. (F) Many large LD are detected in the parasite cytoplasm, and some are even bulging out from the parasite. (G) Transcriptional profile of TgDGAT upon addition of 0.5 mM OA to the medium for 24 h using real-time qPCR analysis. Data are means of means ± SD from 3 separate assays performed in technical triplicate. *, P < 0.003. (H) Toxoplasma replication in HFF assessed by measurement of tritiated uracil incorporation in the presence of PA at the indicated concentrations at 24 h p.i. Data are percentages ± SD relative to the value for the control (set as 100%) for biological triplicates with experimental triplicates. (I) Fluorescence microscopy of HFF infected with RFP-expressing Toxoplasma for 24 h in the presence of 0.2 or 0.4 mM PA, fixed, and stained with BODIPY 493/503. Representative PV from at least 30 infected cells are shown at the same magnification. Tg, T. gondii.