Search

The mean volumes of stationary-phase cells of wild-type and asporogenous mutants of Bacillus subtilis have been measured. Mutants blocked at stage 0 of sporulation either produced cells that had the same volume as the developing sporangium or they divided to produce cells of one-half this volume. The order of expression of the genes affected by the mutations in these strains was determined by biochemical characterization and by construction of double sporulation mutants. Mutants that produced small cells were blocked at an earlier stage of sporulation than those that produced large cells. It is suggested that the following dependent sequence must occur before the formation of the prespore spetum: (i) the initiation of sporulation, (ii) a signal to block the final central division site, and (iii) a signal to activate a polar septum site.

An autoradiographic study revealed two different patterns of cell wall constructioni associated with two types of morphogenesis in Mucor rouxii. In hyphae, the cell wall was preferentially synthesized in the apical region; these cylindrical walls seemed to be generated by a sharply descending gradient of wall synthesis radiating from the apex. In spherical cells (germinating spores, yeast cells), wall formation occurred largely, if not entirely, in uniformly dispersed fashion over the entire cell periphery.

The growth of the cell wall of Bacillus megaterium KM was examined by labeling the cell wall uniformly with (3)H-alpha, epsilon-diaminopimelic acid and examining by radioautography the distribution of labeled cell wall material in daughter cells. The results indicate that the label is distributed uniformly to all daughter cells and rule out a single growth point, either central or apical, for these cells. The results strongly suggest that cell wall growth occurs by uniform deposition of new wall over the whole cell surface.

2-Deoxy-d-glucose (2DG) acted as a competitive inhibitor of the synthesis of cell wall components in Saccharomyces cerevisiae protoplasts. The synthesis of fibrillar glucan cell wall component was inhibited at a glucose to 2DG ratio of 4:1 in the cultivating medium. The completion of the formation of cell wall by the synthesis of the amorphous mannan-protein cell wall component was inhibited at a glucose to 2DG ratio of about 20:1. The inhibition could be reversed by increasing the glucose to 2DG ratio in the nutrient medium. No incorporation of 2DG into fibrillar glucan cell wall component was observed.

Hancock, R. (Harvard Medical School, Boston, Mass.), and P. C. Fitz-James. Some differences in the action of penicillin, bacitracin, and vancomycin on Bacillus megaterium. J. Bacteriol. 87:1044-1050. 1964.-Penicillin and cycloserine do not inhibit the growth of protoplasts of Bacillus megaterium, indicating that inhibition of cell-wall synthesis is the only significant process by which they inhibit growth of bacteria. In contrast, bacitracin and vancomycin inhibit growth of protoplasts and bacteria at similar concentrations, indicating that they have important sites of action other than their known inhibition of cell-wall synthesis. At concentrations which inhibit mucopeptide synthesis, penicillin, bacitracin, and vancomycin each cause an increased rate of efflux of K ions from growing bacteria. This effect of penicillin is prevented by chloramphenicol or hypertonic sucrose, whereas the effects of bacitracin and vancomycin are unchanged under these conditions. It is concluded that bacitracin and vancomycin have direct effects on the cytoplasmic membrane, and it is proposed that their inhibition of cell-wall synthesis could be a consequence of these effects. Bacitracin and vancomycin do not compete with penicillin for binding to cells of B. megaterium, a further indication that they have a different primary site of action.

A fine-structure study of synchronously dividing Bacillus megaterium revealed the sequence of events involved in the division of the cell. First, a mesosome develops as a concentric fold of the plasma membrane at the site of septum formation. The mesosome contains membrane-bound vesicular structures, 300 to 500 A in diameter, plus a large membrane-bound structure, 2,000 A in diameter. These larger vesicles are peculiar to mesosomes in this stage of division and are not observed in the mesosomes involved in spore septum formation. The transverse septum originates within the mesosome and remains enclosed during its subsequent growth across the cell. An intimate association is observed between mesosome vesicles, mesosome membrane, and the growing edge of the transverse septum. Prior to completion of the septum, the membranes bounding the mesosome fuse, and further wall thickening occurs within the structure formed by this fusion. At this time, the septum only equals the parent cell wall in thickness. The doubling in thickness of the septum, which is required for the production of two normal daughter cell walls, occurs during a second phase of wall thickening, which is characterized by the appearance of a constriction at the base of the septum. As the constriction widens, the wall in this region thickens, forming the typical rounded poles of the daughter cells. Capsular synthesis at the poles occurs during this second phase of wall thickening. Throughout the division process, the nuclear material appears to be associated at one end with a mesosome at or near the pole of the cell and at the other end to the mesosome involved in septum formation. This association frequently takes the form of a stalklike extension of the mesosome penetrating into the chromatin fibrils.

1. When Bacillus subtilis was grown in a medium in which sporulation occurred well-defined morphological changes were seen in thin sections of the cells. 2. Over a period of 7.5hr. beginning 2hr. after the initiation of sporulation the following major stages were observed: axial nuclear-filament formation, spore-septum formation, release of the fore-spore within the cell, development of the cortex around the fore-spore, the laying down of the spore coat and the completion of the corrugated spore coat before release of the spore from the mother cell. 3. The appearance of refractile bodies and 2,6-dipicolinic acid and the development of heat-resistance began between 5 and 6.5hr. after initiation of sporulation. 4. The appearance of 2,6-dipicolinic acid and the onset of refractility appeared to coincide with a diminution of electron density in the spore core and cortex. 5. Heat-resistance was associated with the terminal stage, the completion of the spore coat. 6. The spore coat was composed of an inner and an outer layer, each of which consisted of three or four electron-dense laminae. 7. Serial sections through cells at an early stage of sporulation showed that the membranes of each spore septum were always continuous with the membranes of a mesosome, which was itself in close contact with the bacterial or spore nucleoid. 8. These changes were correlated with biochemical events occurring during sporulation.

Hyphae of Aspergillus nidulans continued to synthesize all the major polysaccharide components of the cell wall when cycloheximide was added to cultures. In the presence of cycloheximide, hyphae did not elongate, but electron microscopy showed that the walls became thicker around the cell. The conclusion that cycloheximide changed wall synthesis from extension at the apex to subapical thickening was supported by grain distributions on radioautograms of mutant hyphae labeled with galactose. These findings are discussed in relation to the control of hyphal wall synthesis and are compared with the effects of protein synthesis inhibitors on wall formation in gram-positive bacteria.

Insertion of new wall during the growth of Bacillus licheniformis occurred discretely and probably at the sites of incipient cross wall formation.

This paper describes the process by which the cell wall of Proteus mirabilis, as measured by the presence of the O antigen, develops during the differentiation of swarmers from short cells on an agar surface. The sequence was followed by fluorescent-antibody staining, with both the direct and reverse methods. When the organisms were labeled with fluorescent antibody by the direct method, they showed a progressive diminution of the marker along the cell surface and some increase in the length of the bacteria. However, the label had become completely diluted out before typical swarmers developed. When the bacteria were exposed initially to unlabeled antibody by the reverse technique, and then incubated with fluorescent antibody, they showed a progressive increase both in the intensity of the label along their entire periphery and in cellular length, culminating in the formation of swarmers. It is concluded that in P. mirabilis, as in the few other gram-negative bacteria examined so far, cell wall synthesis takes place diffusely, i.e., by intercalation of new with old components along the length of the cell wall.