WILT OF CUCURBITS. 233 



tissues of the stem were considerably broken down. The organism was cultivated out using the same 

 method as in No. 29 and found to be Bacillus tracheiphilus. Samples were also saved in alcohol for 

 paraffin sections. The following day the second leaf below was flabby. The twenty-ninth day this 

 vine, together with the others, was dug up to make room for new plants. A portion of it was saved 

 dry. Search was made in it for spores. Potato-culture No. 9, November 1 2 (from No. 9, November 

 8, which was inoculated direct from the interior of this vine) when examined November 28 was 

 covered with a smooth, wet-shining thin gray-white slime, exactly typical of B. tracheiphilus. This 

 was the most viscid culture seen. It strung up readily 30 to 40 cm., once 50 and once 53 cm. Exam- 

 ined in a hanging drop the rods were of variable length, but many short. All were about the same 

 breadth, and none were motile. A cover-glass preparation was stained. 



Remarks. Signs appeared on four of the plants in 5! days after inoculation. Two 

 failed to take the disease, and signs appeared on the other four as follows : On two at the 

 end of the sixth day, on one at the end of the eighth day or the beginning of the ninth and 

 on one (that inoculated in the fruit) the tenth day. On October 31, the day on which six 

 plants first showed the disease the temperature of the hothouse was 72 F. at 10 a. m. and 

 80 a few hours later. 



The history of the culture used for making the foregoing inoculations is as follows: 



(1) Beef broth No. i, Sept. 17, inoculated direct from vine No. 2, which was inoculated direct from a cucumber 

 plant diseased naturally. 



(2) Streak from i on slant agar (Red x September 27). 



(3) Potato cylinder (No. 8, October 17) inoculated with one colony from margin of the preceding. 



(4) Potato broth (tube 8, October 23) made from 3. 



(5) Vine 25, etc., inoculated from tube 8, October 23. 



INOCULATIONS OF NOVEMBER 2, 1894. 



An attempt was made to discover whether squash-bugs and cucumber-beetles were the 

 means of introducing this organism into the tissues of the plant. Cucumber plants (Cucumis 

 sativus) were used for the experiment. Some of them were sprayed with a pure culture of 

 Bacillus tracheiphilus and placed under bell-jars along with the insects. Other plants were 

 not thus sprayed but were placed in insect cages into which the bugs or beetles were intro- 

 duced after infecting them with the bacteria. Several attempts were made to infect 

 some of the plants. The infectious material first used was obtained from a potato-broth- 

 culture (tube 8, October 28) which was from slant potato-culture No. 8, October 17. The 

 latter was the first sub-culture from a colony, the original source of which was the interior 

 of vine No. 2. The potato-broth was faintly cloudy with rolling clouds when shaken and 

 was full of an actively motile bacillus, as determined by examination of a hanging drop. 

 The greater part of this tube was poured into about three times as much sterile water. For 

 the cage experiments this diluted culture was sprayed on three pieces of cut squash-fruit 

 each about 2X2 inches so that the whole cut surface was wet. The sprayed squash was in 

 three covered dishes. Into one dish I put the spotted beetle (Diabrotica 12-punctata). Into 

 another I put the striped beetle (Diabrotica vittata) and into a third, a dozen squash-bugs 

 (Coreus tristis). These insects were collected from a squash-field the preceding afternoon 

 and were quite lively. After allowing them to feed upon and crawl over the infected squash- 

 flesh for an hour, they were transferred to cucumber-vines inside of three insect-cages. This 

 work was completed about i p. m. For the bell- jar experiments I used the same liquid, 

 but the plants themselves were sprayed with it. These latter experiments (Nos. 35, 36, 

 39, 40) were begun about 3 p. m. 



(34 a to e.) Five well-grown vines in three pots were placed in an insect-cage. The vines were 6 to 

 10 inches high and had 18 good leaves in addition to the cotyledons which hadnotyetwithered. About 

 10 squash-bugs were taken from the sprayed squash-fruit already described and were put into the 

 cage. Inside of the first half hour they began to stick their beaks into the vines and they were very 

 particular to put them into the veins. The next morning they were not active. Only 3 were on the 

 vines and these were not sucking. At 1 1 a.m. these bugs were taken out and put on pieces of squash 

 freshly sprayed with the same fluid as on the preceding day. This fluid had remained in the atomizer 

 and a microscopic examination showed that the bacilli were still motile. At noon the bugs were once 



