Snapshots of mycobacterial time-lapse movie. Images of phase-contrast, green fluorescence, and red fluorescence channels taken at indicated times from the time-lapse movie following the reporter strain of M. smegmatis. GFP signal, representing induction of the icl gene, increases upon addition of acetate (Ac), whereas the constitutively-expressed DsRed2 signal is present throughout the experiment. Main challenges of automated analysis of such a movie are noted: lack of visible edges between adjacent cells, lack of clear division site, cell shape variations (arrows), and non-cell objects present in the images (arrowhead). Scale bar, 5 μm.