104 



STUDY AND IDENTIFICATION OP BACTERIA 



palisades. Being a Gram-positive organism while the majority of the other patho- 

 genic bacilli are Gram-negative, it is of greatest importance to stain smears by this 

 method. It is not so strongly tenacious of the gentian violet as the cocci, so decolori- 

 zation should not be carried too far. 



The best medium for growing it is Loffler's blood-serum. 



An egg medium, made of the whole egg with glucose bouillon as described pre- 

 viously, is as suitable as Loffler's serum. Coagulated white of egg answers fairly 

 well, as will a hard-boiled egg the shell at one end being cracked and the white cut 

 with a sterile knife. This smooth side is then inoculated and the egg placed cut 

 side downward in a sherry glass. If an incubator is not at hand a tube may be 

 carried next the body in a pocket. The bacillus grows better on glycerine agar than 

 on plain agar. On such plates they appear as small, coarsely granular colonies with 

 a central dark area. In size the colonies resemble the streptococcus. On blood- 

 serum the' colonies are larger ^2 to ^ inch in diameter. 



FIG. 29. B. diphtheria stained by Neisser's method. (Mac Ned.} 



The diphtheria bacillus grows luxuriantly on blood agar and like the Streptococcus 

 pyogenes has a yellowish laked zone around the colony. The Hofmann and the 

 xerosis bacillus do not seem to have this haemolytic power. In bouillon it tends 

 to form a surface growth. It is at the surface that the toxin function is most marked, 

 hence in growing diphtheria for toxin formation we use Fernbach flasks which expose 

 a large surface to the air. It is a marked acid producer bouillon with a -f- 1 reac- 

 tion becoming +2.5 to +3 in thirty-six hours. The nitrate from a two or three 

 weeks' old broth culture is highly toxic, and is usually referred to as diphtheria toxin. 

 It is used in injecting horses to produce antitoxin. Ehrlich uses as a standard to 

 measure the toxicity of toxin the minimal lethal dose (M. L. D.). This is the amount 

 of toxin which will kill a 250-gram guinea-pig in just four days. Some toxins have 

 been produced whose M. L. D. was ^ O o c.c. or 0.002 so that i c.c. of such toxin would 

 kill 500 guinea-pigs. Theoretically, the measure of an antitoxin unit is the capacity 

 of neutralizing 200 units of a pure toxin. (On exposure to light, etc., toxin loses its 

 toxic power and is termed toxoid.) 



