FIG 6 .

Growth within the pellicle confers protection to drug-intolerant M. tuberculosis strains. (A) H37Rv (upside down triangle), H37RvΔmmaA4 (diamond), and the complemented strain (star), grown planktonically in Middlebrook 7H9 medium containing RIF (2 µg/ml). Untreated controls of H37Rv (circle), H37RvΔmmaA4 (square), and the complemented strain (triangle) were included. CFUs were enumerated at the indicated time points by plating aliquots onto Middlebrook 7H10 agar plates. Results are representative of at least two independent experiments. (B) H37Rv (black bar) and H37RvΔmmaA4 (gray bar) strains were grown in Sauton’s medium under pellicle-promoting conditions for 3 weeks, followed by RIF (5 µg/ml) treatment for 1 week. CFU were determined by plating aliquots onto Middlebrook 7H10 agar plates. (C) H37Rv (black bar) and H37RvΔmmaA4 (Gray bar) strains were grown in coculture in Sauton’s medium under pellicle-promoting conditions for 3 weeks, followed by treatment with RIF (5 µg/ml) for 1 week. CFU proportions of H37Rv and H37RvΔmmaA4 strains in the mixed pellicle were determined by plating aliquots onto Middlebrook 7H10 agar plates with or without hygromycin (50 µg/ml), H37RvΔmmaA4 selectable marker. Experiments were performed in triplicate.