474 BACTERIOLOGY. 



The tuberculin thus prepared should be tested upon 

 tuberculous guinea-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 grows 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 stage. By passing 1 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 made out of commercial meat extract does not 

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

 used for preparing* toxins. The commercial 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 T N NaOH in the 



