Microenvironment of Mycobacterium smegmatis Culture to Induce Cholesterol Consumption Does Cell Wall Remodeling and Enables the Formation of Granuloma-Like Structures

Figure 3

The cell wall lipid profile is altered after culture in minimal medium. Lipid fractions of the mycobacterial cell wall were extracted from bacteria in stationary phase cultures, grown in Middlebrook 7H9 broth or minimal medium (MM) with or without glycerol and/or cholesterol supplementation (7H9+Gly, MM+Gly, MM+Gly+Chol, MM+Chol, and MM, respectively), by thin-layer chromatography (TLC) using solvent systems. The fractions were then stained for the following components: (a) glycolipids, using the orcinol staining reagent to identify PIMs; (b) phospholipids, using the molybdenum blue staining reagent to identify cardiolipin (CL), phosphatidylethanolamine (PE), and phosphatidylinositol (PI); (c) mycolic acids using chromic acid reagent; and (d) glycopeptidolipid (GPL) and dimycolate trehalose (TDM) using the orcinol staining reagent. The data are representative results of three independent experiments, showing only the relevant segments of the TLC plates.