FIG 7 .

Cholesterol-altering FDA-approved drugs kill C. burnetii. C. burnetii-infected HeLa cells (3 days postinfection) were treated for 3 h with the indicated drug (20 µM) with or without the vATPase inhibitor bafilomycin A1 (100 nM). (A) Blocking acidification with bafilomycin at least partially rescues bacterial killing by amiodarone, clemastine, haloperidol, and spiperone. Values were normalized to the value with DMSO and without bafilomycin, and the means plus SEM from three experiments are shown. ****, P < 0.00001 compared to the value without bafilomycin, as determined by two-way ANOVA with Sidek’s multiple-comparison test. (B) Filipin labeling of mCherry-C. burnetii-infected HeLa cells suggests that amiodarone and clemastine alter cholesterol trafficking. In amiodarone-treated cells, more filipin labeling in PVs is observed, while clemastine-treated cells show an overall increase in filipin labeling around the PV. Coverslips were fixed following drug treatment for 3 h, and the cells were stained with filipin (cholesterol) and CD63. Filipin images were taken under identical capture settings and processed identically in ImageJ. The filipin fluorescence intensity is shown using a lookup table, with blue showing the lowest fluorescence intensity and yellow showing the highest fluorescence intensity. The white arrows point to PVs, and representative PVs are shown in the insets. Bars = 10 µm.