474 BACTERIOLOGY. 



The tuberculin thus prepared should be tested upon 

 tuberculous guinqa-pigs. The fatal dose should be ascer- 

 tained and the effect on the temperature of the animals 

 should be observed. 



Diphtheria Toxin. 



The diphtheria bacillus when it ^rows in bouillon pro- 

 duces a powerful toxin. The ordinary bouillon which is al- 

 kaline at the time of inoculation becomes acid in about 24 

 hours and then gradually, in about 4 or 5 days, it returns to 

 an alkaline condition. The toxin is produced especially 

 during the alkaline sta^e. By passing a current of air 

 over the liquid, the acid stage can be shortened and, hence, 

 the toxicity of the culture is increased. If, however, the 

 aeration is continued for more than 5 days it will cause an 

 oxidation of the toxin, and, as a result, the liquid decreases 

 in toxicity. 



Bouillon mad>e out of commercial meat extract does not 

 give rise to acid products, and hence such media have been 

 used for preparing toxins. The cbmmercial peptons vary a 

 great deal in their composition, and while some give a 

 bouillon which does not change in reaction, others will give 

 rise to acid products. The production of a temporary acid 

 reaction in bouillon made out of meat is undoubtedly due to 

 the presence of sugar. This can be removed by allowing 

 the meat to ferment. 



The bouillon best adapted for the cultivation of the 

 diphtheria bacillus is prepared as follows : 



1. — 500 g. of chopped beef are added to 1000 c.c. of water; the 

 mass is thoroughly mixed and set aside for 20 hours at 35°. The diges- 

 tion at this temperature serves to destroy the sugar that may be pres- 

 ent. The liquid is then strained through well washed muslin. 5 g. of 

 common salt and 20 g. of "Witte's pepton (2 per cent.) are added to 1 

 liter of the filtrate. The liquid is then titrated with ^ NaOH in the 



