Figure 5

Induction of GFP in M. smegmatis MSOHG4 strain by intracellular organic hydroperoxides of macrophages. (A) GFP expression assessed by microscopy. BMDM cells grown on glass coverslips were infected with M. smegmatis MSOHG4 bearing ohrR-pohr-gfpuv and phsp60-rfp fusions for 2, 4, 12 and 24 h at 37 °C. After washing, the coverslips were examined under Nikon TiE Inverted Fluorescence Microscope using 60X objective. DIC, Cy3 and FITC indicate images obtained using these filters. ‘Merge’ indicates the merged images of DIC, CY3 and FITC. I, Images of uninfected BMDM, and II–V, images of BMDM infected with MSOHG4 for 2, 4, 12 and 24 h of infection, respectively. (B) GFP expression assessed by flow cytometry. BMDM cells grown on culture dishes were infected with M. smegmatis wild type (I), MSOHG2 (II) and MSOHG3 (III) for 4 h at 37 °C. After washing, cells were processed for flow cytometry. Flow analysis was performed in BD FACSAriaII using excitation and emission wavelengths of 395 nm and 509 nm, respectively.