THE PITCHER LIQUOR OF NEPENTHES. 121 



cleavage of the dipeptide had not occurred after digestion for nine 

 days in the incubator. In a second experiment, after digestion for 

 twenty-one days in the incubator, followed by seven days in the 

 room, a distinctly positive reaction for free tryptophane was ob- 

 tained by the bromine and acetic acid test. Hence, liquor from 

 stimulated pitchers apparently hydrolyzed glycyltryptophane, pro- 

 vided the period of incubation was sufficiently long. 



Bacteria of the Pitcher Liquor. 



The bacteriological study was made in collaboration with E. 

 Quintard St. John. Unopened and opened pitchers were studied 

 separately. 



Unopened Pitchers. — Sterile scissors were used in cutting the 

 plant tissues. The prolonged midrib or tendril, which carries the 

 pitcher, was severed ; the top portion of the pitcher was rapidly 

 passed through the flame and was immediately cut off. The cut 

 edge of the pitcher was then flamed rapidly ; and the liquor was 

 withdrawn at once with a sterile pipette, and plated on plain nu- 

 trient agar. After incubation for four days at 2)7° C., the plates 

 were examined for bacterial growth. 



Twelve pitchers were studied in this manner, the liquor from 

 each pitcher being plated separately. Colonies invariably failed to 

 develop on the plates ; hence the liquor in unopened pitchers was 

 sterile. 



Opened Pitchers. — Partly opened pitchers, which had not been 

 invaded by insects, were used in two experiments. The liquor 

 from each of these pitchers contained a goodly number of bacteria 

 which grew on plain nutrient agar at 37° C. 



All the other experiments were conducted on liquor from open, 

 active pitchers in which insect remains were present. 



Total Count. — With each of five pitchers, several successive di- 

 lutions of the liquor were sown on plain nutrient agar, and the 

 plates were incubated at 37° C. for four days ; the colonies were 

 then counted. The number of bacteria per c.c. of pitcher liquor 

 was as low as 48,000 in one pitcher, and as high as 8,000,000 in 

 another pitcher; the other pitchers gave values: 450,000, 1,200,000, 

 and 1,900,000, respectively. 



