J Clin Pathol: first published as 10.1136/jcp.11.6.483 on 1 November 1958. Downloaded from J. clin. Path. (1958), 11, 483. THE BACTERIAL CELL BY ROBERT CRUICKSHANK From the Bacteriology Department, University of Edinburgh Our approach to the bacterial cell depends on to these later. The nuclear or chromatin material our particular interests. The geneticist finds it a has the chemical and staining reactions of desoxy- most useful plastic tool for studying mutations and ribonucleic acid (D.N.A.), but cannot be regarded variations, transductions and transformations; the as a nucleus in the sense that we use the term for chemist is interested in it as a source of enzymes animal cells, since there is no nuclear membrane, of bewildering variety ; he also helps in identifying no centriole, and no unequivocal evidence of divi- the various components of the cell by fractiona- sion by mitosis. tion and detailed chemical analysis of bacterial cell masses; the cytologist is mainly concerned with Cell Wall the anatomy of the cell, using modern staining and Although a rigid lining membrane for the bac- microscopic methods, including phase-contrast and terial cell, similar to that for the fungus mycelium, electron microscopy; the bacteriologist is in- was postulated by Cohn in 1875, it is only since terested in the cell's interaction with its immediate the introduction of electron microscopy that cell environment, whether that be living tissue or dead walls have been clearly demonstrable. From bac- terial suspensions which have been mechanically and decaying matter. copyright. For clinical pathologists gathered together to do or sonically disrupted, the cell walls can be separ- honour to Virchow, it would seem most important ated from the electronically more opaque cyto- to discuss, if only in outline, our modern concepts plasm by high-speed centrifugation at 8,000 to of the anatomy and physiology of the bacterial 10,000 r.p.m., after first removing any intact cells cell. at 2,000 to 3,000 r.p.m. It is less easy to get pure cell walls with Gram-negative than with Gram- Definition positive cells, and mechanical disruption gives Bacteria belong to the kingdom of Protista, better results than sonic disintegration. The cell which also includes plant and animal forms and wall constitutes about 20% of the dry weight of http://jcp.bmj.com/ has many resemblances to the blue-green algae. Gram-positive cocci and as much as 45 % for According to majority opinion at the present time, C. diphtheriae. The thinner walls of Gram- the bacterium may be unicellular or multicellular, negative bacteria probably account for consider- with haploid nuclear or chromatin material occur- ably less than 20 % of the total weight. The thick- ring as a single body and dividing by transverse ness of the staphylococcal cell wall has been esti- fission. Cell division depends on constrictive mated at 15 to 20 mr,, while that of Gram-negative ingrowth of the cell wall and cytoplasmic mem- bacilli, like Bact. coli and salmonellae, is around on October 2, 2021 by guest. Protected brane. 10 to 15 m,u and of Mycobacterium tuberculosis Let us start with a simple diagrammatic repre- 23 mju. The selective rigidity of the outer casing sentation (Fig. 1) of the bacterial cell. It is en- is demonstrated by the finding that the cell walls closed in a cell wall from which there may be of rod-shaped organisms when specially prepared extruded flagella and fimbriae and may be sur- for electron microscopy retain their cylindrical rounded by a capsule. The main function of the form, whereas the protoplast assumes a spherical cell wall is to give form and rigidity and some form when the cell wall is dissolved by lysozyme. protection to the functional cell or protoplast. The Cell wall suspensions have a milky white opales- cytoplasm has a lining or membrane which acts as cent appearance, and in the case of chromogenic a selectively permeable osmotic barrier and con- bacteria there is no contained pigment which is tains within it many granules varying in size from associated with the small particles in the cyto- 10 to 20 mI,u composed mainly of ribonucleic acid plasm. (R.N.A.). Various inclusion bodies, such as There is considerable variation in the chemical volutin granules, lipid granules, etc., may be con- composition of the cell walls in different bacterial tained within the cytoplasm, and we shall refer species, the main constituents being peptide-poly- J Clin Pathol: first published as 10.1136/jcp.11.6.483 on 1 November 1958. Downloaded from 484 ROBERT CRUICKSHA NK MEMBRANE copyright. %1NCLUSION GRANULES 'CELL WALL: EXTRA- PROTOPLASTIC PROTOPLASt STRUCTURES FIG. 1.-Diagrammatic representation of a bacterial cell (extracellular structures omitted at bottom of figure). saccharide complexes. Certain major differences a cell wall precursor, since it contained muramic http://jcp.bmj.com/ occur between the Gram-positive and Gram- acid and the amino-acids in the same ratio (Park negative organisms, the former having a limited and Strominger, 1957). These findings may be linked range of amino-acids, whereas the Gram-negative to the early suggestion by Duguid (1946), follow- bacteria have the same full range of amino-acids ing observed morphological changes in penicillin- as have most proteins. The lipid content of the inhibited bacteria, that penicillin interferes speci- cell wall in Gram-negative bacteria is usually much fically with the formation of the cell wall while greater than that of Gram-positive organisms allowing growth to proceed until the organism on October 2, 2021 by guest. Protected (around 20% compared with 2-4%), and this high bursts its defective envelope and so undergoes content of lipid may be related to the lipo-poly- lysis, and to the recent observations by Cooper saccharide of the 0 antigen or endotoxin. Poly- (1954, 1955) that penicillin is specifically bound to saccharides and hexosamine are present in about a bacterial lipid fraction which could be the cyto- equal amounts in both Gram-positive and Gram- plasmic membrane. At last we seem to be learning negative bacteria. something about the mode and site of action of In the past few years more detailed analyses penicillin and perhaps of other antibiotics. have been made of the chemical content of the cell Another application of our knowledge of the wall and its possible precursors (see Park, 1958). chemistry of the cell wall concerns the action of Thus, analysis of staphylococcal cell walls has lysozyme, which acts on a mucopolysaccharide shown that muramic acid, D-glutamic acid, lysine with the release of N-acetyl hexosamine. When and DL-alanine are present in a ratio of about resistant variants of the susceptible M. lysodeik- I: 1: 1: 3, while examination of penicillin-inhibited ticus are obtained, chemical analysis shows no staphylococci revealed an accumulation of a change in their sugar or amino-acid content but uridine-5'-pyrophosphate complex which could be a 100-fold increase in the 0-acetyl group. These J Clin Pathol: first published as 10.1136/jcp.11.6.483 on 1 November 1958. Downloaded from THE BACTERIAL CELL 485 findings suggest that lysozyme resistance may be correct term than capsular swelling reaction) to due to an ability of the organism to acetylate study the structural elements in the capsule and certain cell wall hydroxyl groups associated with also as an indicator of the localization of surface the substrate, which in the sensitive strains are free antigens. When specific antiserum is added to and receptive (Brumfitt, Wardlaw, and Park, 1958). capsulated bacteria, a precipitation reaction quickly Other lytic enzymes, such as that derived from occurs which renders the capsule visible without Streptomyces albus, act on cell walls and can be changing its size or shape. This reaction has been used for preparing bacterial solutions for precipita- used in the identification of such bacterial species tion reaction, such as Lancefield grouping. as pneumococcus, streptococcus, Klebsiella, Haemophilus, Neisseria, Pasteurella, and Bacillus, Capsules and has helped to show that the capsular substance Much controversy and confusion have arisen of the anthrax bacillus is not a polysaccharide but over the definition of bacterial capsules, and vari- a glutamic acid polypeptide, while that of ous alternative or additional names have been used, Pasteurella pestis is also a protein or a protein- such as envelope, slimy layer, ectoplasm, and outer polysaccharide complex. coat. We may define a capsule as a microscopi- cally demonstrable slimy or gelatinous layer cover- Flagella ing the cell wall and having a definite external sur- face. The term "envelope" is used by some Flagella are unusual in that they lie outside the bacteriologists if the external boundary is not cell wall and may be removed without killing the sharp or the slime has a tendency to dissolution; cell, yet have a characteristic movement which in- others have described a " capsule membrane," but dicates that they are part of the living cell. Despite there seems no sound support for either of these the ingenious artistry and arguments of Pijper, we may accept the flagella as locomotor organelles two terms. A good deal of the confusion has copyright. arisen from difficulties in the demonstration of the with their origin in a basal granule inside the cell capsule. The two most reliable methods are the wall. Structurally, they are non-tapering filaments wet-film India ink technique, used as a negative in the shape of a cylindrical helix, which may have stain, and the specific antigen-antibody reaction either one of two different wavelengths. The fila- erroneously called the Neufeld Quellung or cap- ments vary in length up to 12 ,u and are approxi- sular swelling reaction.