196 Journal of Agricultural Research voi. xix. No. s 



climatic periods in the Philippines. Two of the series of inoculations with 

 orchard soil infusions were carried on in the middle of the rainy season 

 when the soil was kept constantly wet by the rains. The third series of 

 inoculations was carried on at the beginning of the dry season when the 

 soil dried out and became dusty to a considerable extent. The attempt 

 was made to secure the soil for each day's infusion at different depths. 

 Soil was frequently taken from the surface and just as frequently from 

 a depth of 10 inches. It is thought that the inoculations shown here 

 were made from soil infusions which are entirely representative of the 

 different conditions in the Lamao soils. 



Inasmuch as the question of the existence or nonexistence of P. citri 

 in the soil is an important point in canker control work, the following 

 test was undertaken to corroborate further the preceding experiments. 



EXPERIMENT III 



INOCULATED SOIL IN BOXES 



Orchard soil was autoclaved twice, one hour each time at 45 pounds 

 pressure. The soil was placed in thin la5^ers on plates, so that the steam 

 would penetrate easily. The autoclaved soil was placed in a seed-house 

 flat which measured 18 by 24 by 5 inches. The soil was air-dried and was 

 inoculated with 1,500 cc. of an infusion of P. citri in sterile water. This 

 flat was then placed at a level with the soil and covered with cheesecloth 

 to prevent animals from disturbing it. The flat received the full play of 

 sun, wind, and rain and was exposed to the same conditions as exist 

 beneath a tree in the orchard. 



Another flat of the same size containing unsterilized soil was inocu- 

 lated with an equal amount of an identical infusion. This flat was placed 

 under identical conditions with the flat of sterilized soil but at several 

 yards' distance to prevent distribution of the canker organism too easily; 

 it was also covered with cheesecloth. 



On the first day after inoculation, a small portion of the inoculated soil 

 from the autoclaved flat was removed with a spatula to a clean dry- 

 sterilized culture tube. To this about 10 cc. of tap water were added; 

 the tube was shaken vigorously for several minutes; and the resulting 

 infusion was spread upon the upper and lower surfaces of five actively 

 growing pummelo leaves. Each leaf was then punctured 40 times with 

 a new needle, and a new coating of the infusion was spread over the 

 leaves and over the punctures. For this spreading of the infusion small 

 cotton swabs such as are used for collecting diphtheria cocci from sus- 

 pected cases were used. A new swab was used for each tube of infusion. 



Five infusions were made each successive day from the flat of inocu- 

 lated autoclaved soil. On each successive day five infusions were made 

 in the same way from the unsterilized inoculated soil, and each of these 

 was spread upon five actively growing leaves, each leaf being subse- 

 quently punctured 40 times. 



