Microbial Structures and Staining Reactions 111 



dismiss the hundreds of gram negative organisms described in 

 Bergeys Manual of Determinative Bacteriology. Identification of 

 organisms is an ehmination procedure, and the gram stain represents 

 a very important early step in this process. 



Without discussing complex chemical and physical theories 

 proposed to explain this staining difference of bacteria, a few broad 

 concepts will be mentioned. Some persons believe that there is 

 a difference in the permeability ( intactness ) of the cell walls, with 

 gram negative cells being more permeable than the gram positive 

 bacteria, both for the entrance and for the egress of the crystal 

 violet dye. The chemical composition of the surface of the cells 

 is another theory put forth to explain the difference in gram stain- 

 ing reactions between cells. Gram positive bacteria contain a 

 chemical compound called magnesium ribonucleate at or near the 

 cell surface. When these organisms are stripped of this chemical, 

 they become negative in their staining reactions. If the magnesium 

 ribonucleate is "replated" on these stripped cells, they revert to their 

 gram positive status. Attempts to convert true gram negative 

 cells by this plating technic had met with failure until recent 

 studies indicated that if a viscous ribonucleate is employed, it ;s 

 possible to change gram negative bacteria into gram positive 

 organisms. That the gram stain depends upon surface phenomena 

 is fairly well agreed, but the exact mechanism of the reaction is 

 still in doubt. 



CAPSULE STAIN 



It is sometimes difficult to demonstrate capsules on bacteria, 

 and some failures undoubtedly are due to the ease with which some 

 slime layers can slip off the cells during the staining process. In 

 fact, slime can be demonstrated in some liquid cultures, but the 

 bacteria in that culture may fail to show capsules around the 

 cell wall. 



A generally accepted technic for staining capsules employs 

 India ink, nigrosine, or Congo red (so-called negative stains) as 

 background material against which the unstained organisms stand 

 out. By counter-staining with a basic dye like crystal violet, the 

 bacterial cell will take up the dye while the capsule stains only 



