Ultrastructural study of the reversion of protoplasts of Bacillus licheniformis to bacilli

The reversion of protoplasts of Bacillus licheniformis 6346 His- on a medium containing 2.5% agar has been studied in sectioned material after reaction with a ferritin-conjugated antibody specific to the peptidoglycan isolated from the walls of the bacilli. Freeze etching has also been used. Fibrils of material reacting with the antibody have been detected emerging from isolated areas of the protoplasts after 3 h of incubation. This material gradually covers the cell and can eventually (at 6 h) be seen in freeze-etched preparations as a fringe of up to 400 nm around the cells and covering the surfaces with particles that can be removed by lysozyme. At later stages the wall begins to take on a compact, well-defined appearance that can be seen in sections; however, the cells are still grossly deformed. A transitory emergence, beyond the wall of long fibers of 6 nm in diameter, takes place after about 12 h of incubation. These fibers react with the conjugated antibody and after freeze etching show a regular banded structure. They are probably indentical with the fibers isolated elsewhere (Elliott et al., 1975) and shown to contain all the wall constituents (i.e., peptidoglycan, teichoic acid, and teichuronic acid). These fibers are not detectable in the final stages of reversion.