NATURE OF THE " ESSENTIAL SUBSTANCE " 93 



occurred, and, as we have already remarked, in 

 point of fact some growth took place on most of the 

 media. 



The portion that was insoluble in chloroform, but 

 soluble in water, always gave a good growth, and 

 we were able to reduce the amount of essential sub- 

 stance in the chloroform soluble portion by extract- 

 ing with water, especially by dissolving it in a little 

 chloroform and shaking with water in a separating 

 funnel. 



In the case of the bovine tubercle bacillus grown on 

 Dorset's egg medium, we obtained apparently the 

 same extracts as with the B, phlei, yet Johne's 

 bacillus failed to grow on media containing the ex- 

 tracts of the bovine tubercle bacillus. 



To investigate the nature of the "essential sub- 

 stance," we now carried out experiments to ascertain, 

 if possible, the general nature of the substance or 

 substances from which it was formed, and from these 

 experiments we soon obtained some interesting results. 

 We experimented with B. phlei grown on various 

 media, and found that when grown on glycerine-agar, 

 or glycerine-broth, the bacilli formed a good medium ; 

 but, on the other hand, when grown on Dorset's egg 

 medium, the results were not nearly so good, and 

 when grown on glycerine-liver broth for four weeks 

 at 37° C, the bacillus was useless for making media 

 for Johne's bacillus. From B. phlei grown on liver- 

 broth we obtained apparently the same extracts as 

 before, but the extracts were unsuitable for making 

 into media. The same results were obtained, as a rule, 

 with the other acid-fast bacilli. The human and avian 

 tubercle bacilli made the best media for Johne's bacillus 

 when they had been grown on glycerine-beef broth, or 

 glycerine-beef broth-agar, although the avian type, 



