FIG 2.

Intracellular activity of RFB on M. abscessus-infected THP-1 cells. (A and B) Macrophages were infected with (A) M. abscessus S-morphotype and (B) R-morphotype expressing TdTomato (MOI of 2:1) for 3 h prior to treatment with RIF (50 μg/ml), AMK (50 μg/ml), RFB (12.5 μg/ml), or DMSO. CFU were determined at 0, 1, and 3 dpi. Data are mean values ± SD for three independent experiments. Data were analyzed using a one-way analysis of variance (ANOVA) Kruskal-Wallis test. (C and D) Percentage of infected THP-1 macrophages at 0, 1, and 3 days after infection with (C) M. abscessus S or (D) M. abscessus R. Data are mean values ± SD for three independent experiments. Data were analyzed using a one-way ANOVA Kruskal-Wallis test. (E) Percentage of S-infected macrophage categories and (F) percentage of R-infected macrophage categories infected with different numbers of bacilli (<5 bacilli, 5 to 10 bacilli, and >10 bacilli). The categories were counted at 0 or at 1 and 3 days postinfection in the absence of antibiotics or in the presence of RIF or AMK at 50 μg/ml or RFB at 12.5 μg/ml. Values are means ± SD from three independent experiments performed in triplicate. (G) Four immunofluorescent fields were taken at 1 day postinfection showing macrophages infected with M. abscessus expressing Tdtomato (red). The surface and the endolysosomal system of the macrophages were detected using anti-CD63 antibodies (green). The nuclei were stained with DAPI (blue). White arrows indicate individual or aggregate mycobacteria. Scale bar, 20 μm. **, P ≤ 0.01; ***; P ≤ 0.001.