Fig. 5
A time-shift between vector oogenesis and parasite lipid acquisition. a Lipid acquisition by mosquito ovaries precedes P. berghei oocyst development. The ratios of Nile Red mean intensities in the ovaries (orange line, N = 3) and oocysts growth (green line, N = 2) between control and Lp-depleted mosquitoes are shown. Mean oocyst sizes and Nile Red ovary intensities from pooled mosquitoes and experiments (N ≥ 2, 5–13 mosquitoes per time point per condition per experiment). b Lp-depleted mosquitoes (dsLp) and controls (Ctrl) were infected with P. berghei and offered (f–i) or not (b–e) a supplemental blood meal 7 days post infection (dpi) (3–63 mosquitoes per condition per experiment). b, f Salivary gland sporozoites were isolated and counted from control and Lp-deficient mosquitoes. Each dot represents the mean number of sporozoites per mosquito per experiment (N = 5; *p < 0.05; one-sided paired t-test, paired values for different dsRNA treatment within the same experiment). c, g Human hepatoma HepG2 cells were infected with 10,000 sporozoites and the number of extra-erythrocytic forms (EEFs) per field was gauged by microscopy 2 dpi. As a negative control, sporozoites from control mosquitoes were treated with cytochalasin D (Cyt D). Five microscopic fields were imaged per well per condition in triplicates, except for two cases for dsLp due to the low number of sporozoites. Mean number of EEFs of pooled experiments (N = 3) and SEM are shown (***p < 0.0001; Kruskal–Wallis test and Dunn’s multiple comparison test). d, h Representative fluorescent micrographs of GFP-expressing EEFs (green) developing in HepG2 cells. Cell nuclei are stained with DAPI (blue). Scale bar—10 µm. e, i Growth of EEFs produced by sporozoites from control and Lp-deficient mosquitoes. EEFs sizes of pooled experiments are shown (N = 3). The boxplots show the median (center line), the 25th to 75th percentiles (bounds of box) and the 10th and 90th percentiles (whiskers). The total numbers of EEFs are shown in brackets. e ***p < 0.0001; i p = 0.4223; one-sided Mann–Whitney test
