101 



transferred to 10 c. c. of srolatiii at .'!0° ('., shaken, and ponred into a sterile Petri 

 dish to form the clieck plate. 



(2) Tlie remainder of the cnlture (approximately 9 c. c.) was then plunged for 10 

 minutes into water at 4().r>0° to 46.60° C, and another 1 c. c. immediately pipetted 

 out into another tiihe of 10 c. c. of gelatin at 30° C, shaken, and poured into a 

 second Petri dish. 



(3) The culture was then allowed to cool to room temperatures after which 1 c. c. 

 was pipetted into another 10 c. c. of gelatin at 30° C, shaken and poured into a 

 third Petri dish. 



These 3 dishes were then put into the cool box, where they were kept at 12° to 16° 

 C, and examined and compared from time to time in the same way as the dishes of 

 agar. 



Results: (1) Colonies to the number of 2,000 to 3,000 per field (Zeiss 16 mm. and 

 12 comp. ocular) appeared in this dish. (2) More than 80 per cent of the germs 

 were destroyed by the heat, i. e., there were only 200 to 600 colonies per field in this 

 dish. (3) About 400 colonies per field appeared in this plate. 



The culture from which these 3 plates were inoculated was made with a single loop 

 from a broth culture 1 1 days old and this fact, together with its own age (28 hours at 

 24° to 28° C), precludes the idea that spores played any part in the results obtained. 



XI. February 3, 1898: Six tubes of alkaline beef broth, stock 286b (stock 286 con- 

 sisted of the broth from 1,000 grams of minced lean beef covered with 1,500 c. c. 

 distilled water and left in the ice box 24 hours. The fluid was finally made up to 

 2,000 c. c, titrated, and divided into four ecjual parts. Stock 286b received enough 



2N 



^ NaOH to render it exactly neutral to phenolphthalein, i. e., strongly alkaline to 



neutral litmus paper). These tubes were of Weber's resistant glass, 169 by 17 mm., 

 and very thin walled. Each received exactly 10 c. c. of the l)roth, in which, from 

 previous tests, the germ was known to grow readily, even when added in very small 

 'quantities. Each of these 6 tubes was inoculated with a drop of fluid from tube 1, 

 Jan. 29, a l)eef broth which was nicely clouded with good rolling clouds on shaking. 

 This broth had been clouded about 55 hours, but showed as yet very little precipitate 

 and no pellicle or zoogloea;. As much fluid was put into each tube as could be lifted 

 out on a medium sized (2 mm.) loop and 5 or 6 cm. of wire above it, i. e., an enor- 

 mous number of germs, as microscopic examination showed. About 15 or 20 

 minutes after inoculation 4 of the tubes were plunged into the hot water, while the 

 other 2 were held as checks. The exposed tubes were \mt well down into the bath 

 so that the surface of the broth was 5 to 8 cm. below the surface of the water, which 

 was in constant motion. The exposure was exactly 10 minutes. On removal, 2 of 

 the tul)es were cooled innnediately umler flowing water, while the other 2 were 

 allowed to cool gradually at room temperature (23° C. ). All were then screened 

 from the diffused light of the room and set away at room temjieratures which varied 

 from 15° to 25° ('. The temperature of the water l)ath at the beginning was 47.80° 

 C, falling slowly to 47.58° C. at the end. During the middle 8 minutes the range of 

 temperature was from 47.70° C. to 47.60° ('. These tubes were under observation 

 33 days. 



Results: (1) Checks. Both tubes clouded inside of 48 hours and passed through 

 a normal course of growth. (2) Cooled quickly. Both tubes remained perfectly 

 clear. (3) Cooled slowly. Both tubes remained perfectly clear. 



XII. February 3, 1898: This experiment was in all respects a duplicate of the 

 preceding, except that the water was a trifle cooler and that after iin)culation the 

 tubes were allowed to stand one-half hour before plunging. The temperature of the 

 water waa 47.45° C. at the beginning and 47.17° C. at the close. After one minute 

 the tem|)erature of the bath fell to 47.30° C. and during the next ^ minutes it gradu- 

 ally fell to 47.20°. During the remaining 3^ minutes the temperature fluctuated 

 between 47.16° C. and 47.18° C, being at the latter point most of the time. 



