EXPERIMENT STATION BULLETINS. 485 



the classes of bacteria mentioned, a bile medium was prepared and tested 

 for trial by plating pure cultures of typhoid and lactic bacteria. 



Preparation of the Bile Medium. — Owing to the fact that no form of 

 bile salts, sodium taurocholate or glycocholate, were available at this 

 time, bile from a freshly slaughtered ox and sheep was substituted. The 

 agar was made in two lots, using the bile of the ox for one lot and that 

 of the sheep for the other. 



This medium was prepared as follows: 15 g. of agar was soaked in 

 500 cc. water over night. In the morning 20 g. peptone and 5 g. salt were 

 dissolved in 100 cc. hot water and added to the agar, which had been 

 digested previously. 20 g. lactose dissolved in 400 cc. hot water was 

 added next. This was divided into equal parts (500 cc. each) ; to one 

 was added the bile* of the sheep, to the other the ox bile* (about 30 cc. 

 each). 



Just before filtering, 1 per cent of a standard solution of Merck's puri- 

 fied litmus^ was added to each lot of agar. 



Broth cultures of the Bact, lactis acidi (lab.) and B. typliosus respec- 

 tively were plated on each lot of the litmus lactose bile agar to test its 

 efiiciency. The respective colonies could be distinguished readily from 

 one another. The presence of the bile in the medium seemed to inhibit 

 the growth of the lactic organism almost entirely. 



Two or three lactic colonies developed upon the ox-bile agar, none upon 

 the sheep-bile agar. The lactic colonies upon the ox-bile agar were very 

 small, round and transparent, resembling a minute drop of water. For 

 positive identification several of these colonies were transferred to broth, 

 incubated until signs of growth were shown and then transferred to 

 litmus milk tubes. The typical lactic reaction took place within 2-4 

 hours. 



The tyijhoid bacilli grew well upon both lots of this agar; both the 

 surface and sub-surface colonies were easy to difterentiate from those of 

 the lactic. 



This difi'erentiation being quite satisfactory, the same experiment was 

 continued, making duplicate plates in ox-bile agar and in "calcium-car- 

 bonate" agar. The counts for Bact. lactis acidi (lab.) were taken from 

 the calcium-carbonate agar and for B. typhosus from the bile agar. 



*The gall bladders were obtained from a freshly slaughtered carcass of a sheep and an 

 ox and were brought to the laboratory under sterile precautions, broken separately 

 into sterile deep culture dishes, and placed in cold storage until used. 



