25 



Cytase. The greatest interest in this organism is its power of 

 destroying the cell walls of various plants. The quick spreading nature 

 of the rot shows that the cell-wall-destroying-enzyme must be elabor- 

 ated in considerable amounts. 



This enzyme was isolated in the following manner : — 



Sound potatoes were peeled and pieces cut out of the centre with 

 sterilized knives. These pieces were scorched over the naked flame 

 of a Bunsen burner and then dropped into wide mouth sterilized 

 flasks contain^rg 50-200 c. c. of sterilized distilled water. 



This operation, although carefully carried out in a chamber washed 

 with corrosive sublimate, was not always successful as a number of the 

 flasks became contaminated with foreign organism : however, some 

 flasks were obtained which contained nothing but B. oleraceae, and 

 these, after incubation at 25 degree C, for 10 days, were emptied into 

 a fine sterilized cloth and the juice pressed out. 



This juice was then filtered through absorbent cotton and treated 

 with four times its bulk of 94 per cent, alcohol, which gave a fine cloudy 

 precipitate. The mixture was frequently shaken and was left at room 

 temperature for 24 hours. After the final shaking the precipitate 

 was allowed to sediment for 12 hours when the supernatant liquid 

 was siphoned off, and the sediment collected on a hard filter paper, 

 w-ashed with alcohol, dried and then a hole was made in the filter and 

 the precipitate washed off into a sterile flask with sterilized distilled 

 water. This solution was then forced through a Pasteur-Cliamberland 

 filter, collected in a sterile flask and 5 c. c. portions of the liquid filled 

 into sterilized test tubes. The liquid was clear with a yellowish tinge 

 and was (juite sterile. (Incubation of tubes at 25" C.) 



Twenty test tubes were thus obtained and 8 of them were 

 treated as follows : — 



4 were heated to a temperature of 65 degrees C. for 10 minutes. 



4 were heated to a temperature of 212 degrees C. and then cooled. 



Small slices of potato and white turnip were then cut with sterile 

 knives and introduced into the tubes which were placed in a thermos- 

 tat at 20 degrees C. At the end of 24-36 hours the tubes were care- 

 fully examined and those that showed bacterial contamination were 

 put aside. The small pieces of tissue were fished out with a sterile 

 spatula and placed on a slide, a cover glass placed on top and the pre- 

 paration examined under microscope. The sections of turnips and 

 potato in the boiled and heated tubes were unchanged, they were 



