BACILLUS DIPHTHERIiE 515 



The cover-slip preparation, after having been fixed, is stained with so- 

 lution No. 1 for one to three seconds. It is then washed in water and 

 immersed for from three to five seconds in solution No. 2. With this 

 stain the bodies of the bacilli appear brown, the polar granules blue. 



Another method which has been extensively used is that of Roux. 

 The solutions required for this are : 



1. Dahlia violet 1 gram. 



Alcohol, 90 per cent 10 c.c. 



Aq. dest ad 100 " 



2. Methyl green 1 gram. 



Alcohol, 90 per cent 10 c.c. 



Aq. dest ad 100 " 



The two solutions are mixed, one part of 1 being added to three parts 

 of 2. Preparations are stained in this mixture for two minutes. The 

 polar bodies appear a dark violet. Other methods for the staining of 

 polar bodies have been recommended. There is very little advantage 

 in the use of these double stains and most bacteriologists employ for 

 routine work the simple stain with LoefHer's alkaline methylene blue. 



The significance of the polar bodies is not well understood. Their 

 discoverer, Ernst, regarded them as bodies analogous to the spores of 

 other organisms. The ease with which they are stained, however, and 

 the low temperatures to which the bacteria succumb make this appear 

 very unlikely. A more probable interpretation seems to be that of 

 Escherich ' who regards them as chromatic granules. 



Stained by Gram's method, the diphtheria bacilli retain the gentian- 

 violet. Care must be used in carrying out this method and strict timing 

 adhered to, since slight carelessness in this respect may lead to irregular 

 results. 



In stained smears from the throat or from cultures a characteristic 

 grouping of the baciUi has been observed. They lie usually in small 

 clusters, four or five together, parallel to each other, or at sharp angles. 

 Two organisms may often be seen attached to each other by their cor- 

 responding ends while their bodies diverge to form a "V" or "Y" shape. 



Biological Characteristics. — ^The diphtheria bacillus is a non-motile, 

 non-flagellated, non-spore-forming aerobe. Its preference for oxygen 

 is marked, but it will grow in anaerobic environment in the presence of 

 suitable carbohydrates. It does not liquefy gelatin. The bacillus grows 

 at temperatures varying between 19° C. and' 42° C, the most favorable 



^ Escherich, " Aetiologie, etc., d. Diphth.," Wien, 1894, 



