Intact Cells, Protoplasts and ""Ghosts" of a Bacillus 



Ibl 



The cytoplasmic areas and the nuclear equivalents 

 are seen. Sometimes the outer border of the cyto- 

 plasm appears slightly darker than the rest of it, 

 suggesting the presence of a cytoplasmic membrane. 

 These findings are in accordance with the fact that 

 the protoplasts very much show the same biochemi- 

 cal and physiological capabilities as the intact cells. 

 The protoplasts thus respire at the same rate as the 

 cells (13), they synthesize protein and nucleic acid 

 from low molecular weight compounds (4) and they 

 are able to produce adaptive enzymes (3, 5, 16). 

 Moreover, bacteriophage particles and spores de- 

 velop within them (I, 7, 9, 10). Under certain 

 conditions protoplasts even exhibit growth and divi- 

 sion phenomena (6). 



The absence of the cell wall structure in proto- 

 plasts has earlier been indicated by analytical data 

 (13), by light microscopical investigations (12, 14) 

 and by the fact that they do not absorb phage 

 particles (13). 



The "ghosts** consists of shells enclosing some 

 protoplasmic material more or less in the form of 

 granules but otherwise empty, as may be apparent 

 from figs. 3 and 4. Thus earlier observations by means 

 of light microscopy (12, 15), strongly suggesting that 

 the "ghosts*" represent cytoplasmic membranes, are 

 confirmed. On the other hand it should be empha- 

 sized that some protoplasmic material evidently re- 

 mains enclosed within this membrane even after 

 the osmotic lysis of the protoplasts. 



Staining experiments performed on whole cells 

 and "ghosts** using Sudan Black seem to indicate 

 that the granules of Bacillus M. are not of lipid 

 nature. In all events they should not be identified 

 with the granules of polymerized /j-hydroxy-butyric 



Fig. 1. Section of intact cells of Bacillus M. Magnification 

 >: 28,000. 



Fig. 2. Section of protoplasts of Bacillus M. Magnification 

 X 38,000. 



Fig. 3. Section of "ghosts" of Bacillus M. Magnification 

 25,000. 



Fig. 4. Electron micrograph of drop preparation of "ghost" 

 of Bacillus M. 



acid found in genuine B. nwi^atcrium strains. They 

 seem instead to have more in common with the 

 non-lipid granules that have recently been isolated 

 by Georgi, Militzer and Decker (2) from the ther- 

 mophilic bacterium Bacillus stcarothcrniophilus. 

 These workers report that the granules contain 

 desoxyribonucleic acid among other things and, from 

 the enzymological point of view, cytochrome c and 

 phosphatases. 



It should, however, be borne in mind that the 

 granules found in the bacterial "ghosts" may be 

 artefacts. The osmotic lysis must be regarded as a 

 rather violent disintegration process, readily causing 

 rearrangements between the protoplasmic constitu- 

 ents of the bacterial cell. Such an interpretation is 

 strengthened by the fact that no granules of the size 

 and the electron density found in the "ghosts" of 

 Bacillus M, are seen in sections of intact cells or 

 protoplasts. 



References 



Brenner, S. and Stent, G., Bioc/iim. Bioplivs. Acta 17, 



473 (1955). 

 Georgi, C. E., Militzer, W. E., and Decker, T. S., 



J. Bactoriol. 70, 716 (1955). 

 Landman, O. E. and Spiegelman, S., Pioc. Natl. Acad. 



Sci. Wash. 41, 698 (1955). 

 McQuillen, K., Biochim. Biophys. Acta 17, 382 (1955). 

 ^ /. Gen. Microbiol. 13, iv (1955). 

 ^ Biochim. Biophys. Acta 18, 458 (1955). 

 Mutsaars, W., Ann. Inst. Pasteur 89, 166 (1955). 

 Sai.ion, M. R. J., Nature 170, 746 (I952J. 



— /. Gen. Microbiol. 13, iv (1955). 



Salton, M. R. J. and McQuillen, K., Biochim. Biophvs. 



Acta 17, 465 (1955). 

 ToMCSiK, J., Symp. Soc. Gen. Microbiol. 6, 53 (1955). 

 ToMcsiK, J. and Guex-Holzer, S., Schwei:. Z. allgcm. 



Pathol, u. Bakteriol. 15, 517 (1952). 

 Weibull, C, J. Bacterial. 66, 688 (1953). 



— E.xptl. Cell Research 9, 139 (1955). 



— ibid. 10, 214 (1956). 



WiAME, J. M., SroRCK, R., and Vanderwinkel, E., 

 Biochim. Biophys. Ada 18, 353 (1955). 



12. 



13. 

 14. 

 15. 

 16. 



