FIG 3.

Succinate oxidation is the major contributor of electrons to the respiratory chain in M. tuberculosis. (A to C) Oxygen consumption rates (OCR) of the single and multiplexed sdhA1, sdhA2, and frdA knockdown strains when energized with OADC (A), succinate (B), or glycerol (C). Cultures were grown for 8 days in the presence of 100 ng/mL ATc to deplete cells of SDH/FRD enzymes in 7H9 medium containing the specified carbon sources before performing OCR measurements on cell suspensions. Data were normalized to the respective no-ATc control for each strain. 100% for OADC = ~30.0 pmol/(s × mL), for succinate = ~30.0 pmol/(s × mL), and for glycerol = ~35.0 pmol/(s × mL). Error bars represent the mean and standard deviation for two technical replicates, and data are representative of two independent experiments. Statistical significance was calculated using a one-way analysis of variance and a Dunnett test for multiple comparisons of each strain against the nontargeting control; ns, P > 0.05; *, P < 0.05; **, P < 0.01; ***, P < 0.001; and ****, P < 0.0001. (D) Chemical inhibition of both terminal oxidases with Q203 (400 nM) and ND-011992 (100 μM) demonstrates that succinate oxidation is the major contributor of electrons to the respiratory chain. Cultures were grown for 8 days in the presence of 100 ng/mL ATc to deplete cells of SDH/FRD enzymes in 7H9-OADC medium before performing OCR measurements on cell suspensions. Dashed horizonal lines in panel D represent the baseline OCR. Arrows represent the contribution of succinate oxidation to the OCR of M. tuberculosis and the residual OCR in the absence of succinate oxidation, respectively. Data are representative of two independent experiments.