(A) For pharmacological inhibition of acid ceramidase (Ac), C57BL/6 mice were either treated with 750 µg carmofur or with vehicle as control by daily intraperitoneal (i.p.) injection starting 1 day prior to P. yoelii infection. (B) Parasitemia of infected carmofur- and vehicle-treated mice was determined at indicated time points by microscopy of Giemsa-stained blood films (n = 10–11). (C) Spleen weight of P. yoelii-infected carmofur- and vehicle-treated mice 7 days post infection (p.i.) (n = 9). (D) Ceramide, sphingosine, S1P, and sphingomyelin levels in bone marrow of infected carmofur- and vehicle-treated mice 7 days p.i. were determined by high-performance liquid chromatography mass spectrometry (HPLC-MS/MS) (n = 6–9). (E) Frequencies of Ter119+CD71+ reticulocytes in blood of P. yoelii-infected carmofur- and vehicle-treated mice were analyzed by flow cytometry on days 0, 3, 7, and 10 (n = 12–18). (F) Percentages of erythroblasts (Eb, CD71+FSChigh), reticulocytes (Ret, CD71+FSClow), and mature erythrocytes (Ery, CD71-FSClow) in bone marrow of P. yoelii-infected carmofur- and vehicle-treated mice on day 7 (n = 9). (G) P. falciparum growth inhibition assay to test potential effects of carmofur on intraerythrocytic development in vitro. Micrographs of infected erythrocytes that were either nontreated or exposed 48 hr to increasing doses of carmofur (top rows) or chloroquine (bottom row). Shown are representative images of standard growth assays (left panel). Bars: 2 µm. Dose–response assay (48 hr) of carmofur or DMSO as control against asexual P. falciparum growth (starting at ring stages, n = 3 in triplicates). 50% inhibitory concentration (IC50) is indicated. Results from 2–3 independent experiments are presented as mean ± SEM. Statistical analyses were performed using two-way ANOVA, followed by Sidak’s post-test for (A), unpaired Student’s t-test for (C) and (F), and Mann–Whitney U-test for (D) and (E) (*p<0.05, **p<0.01, ****p<0.0001).