FIGURE 3.

LOS content of the various M. marinum mutants. Following extraction, polar glycolipids from the wild type and the wbbL2, MMAR_2331, and MMAR_2321 deletion mutants as well as their corresponding complemented strains (wbbL2_C, 2331_C, and 2321_C) were separated by two-dimensional TLC using the solvent system chloroform/methanol/water (60:30:6, v/v/v) in the first direction and chloroform/acetic acid/methanol/water (40:25:3:6, v/v/v/v) in the second direction. Glycolipids were detected with orcinol/sulfuric acid staining and charring. In parallel, TLC plates were stained with iodine vapor, and the glycolipid intermediates accumulating in the different mutants (labeled 1 and 2 for wbbL2, 3 for MMAR_2331, and 4 for MMAR_2321) were individually extracted for further structural identification. PIM, phosphatidylinositol mannosides.