Fig 1. Screening the MMV ‘Pathogen Box’ for inhibitors of O₂ consumption in T. gondii.

The O₂ consumption rate (OCR) of extracellular T. gondii parasites was measured in a 96-well plate using a Seahorse XFe96 extracellular flux analyzer. Compounds from the MMV ‘Pathogen Box’ were added to wells at a final concentration of 1 μM, and the change in OCR was monitored in real-time after each addition. Percent inhibition of OCR by each of the 400 compounds was calculated relative to complete inhibition observed after addition of the known OCR inhibitors atovaquone (1 μM) and antimycin A (10 μM), with each compound represented by a dot. A >30% inhibition cut off was applied (dotted line), with seven compounds inhibiting OCR by >30% at 1 μM (coloring of dots corresponds to coloring of labels of the chemical structures shown below). These hits included MMV689480/buparvaquone (burgundy), the endochin-like quinolone (ELQ) MMV671636 (green), MMV688754/trifloxystrobin (pink), MMV688978/auranofin (orange), MMV024397 (red), the aminopyrazole carboxamide MMV688853 (dark blue), and MMV021057/azoxystrobin (light blue). Data are from a single experiment, with the plate layouts and data points summarized in S1 Table.