WILT OF CUCURBITS. 221 



remote distances from the point of inoculation. Examined in drops of sterile water many of those 

 rods taken from near the tip were actively motile, while those taken lower down, i. e., from vessels 

 clogged solid with the bacteria, were not motile at all or only doubtfully so. The final condition 

 of the bacteria in the vessels of the plant appeared to be a zoogloeae stage. The organism was cul- 

 tivated from the interior of this plant at various levels and found to be Bacillus tracheipkUus. In 

 making these transfers the stems were shortened with a hot knife and the end dug into with a 

 flamed needle. On September 20, under these conditions, the following transfers into tubes of 

 sterile media were made from the interior of this plant : 



(1) Four inches from the tip. Beef broth; (2) Do., Cucumber broth; (3) Six inches from the tip, Potato broth; 

 (4) Do., Peptonized beef broth; (5) Eight inches from the tip, Beef broth; (6) Do., Potato broth; (7) Eleven 

 inches from the tip, Beef broth; (8) Do., Cucumber broth; (9) Fifteen inches from the tip Potato broth ; (10) Do., 

 Cucumber broth. 



One tube remained sterile (No. 2) ; 2 were contaminated (No. 8 with a pink organism and No. 10 

 with a white organism forming a pure white, wrinkled, fragile pellicle; 7 yielded moderately clouded 

 cultures exactly alike and presumably all of them pure cultures of Bacillus tracheiphtlus. The 

 behavior of the organism in 5 of these cultures (Nos. 1, 3, 5, 7, and 9 was tested further by transfers 

 to cylinders of sterile cooked potato. On this medium each one developed a thin, smooth, wet-shining 

 gray- white, sticky slime, scarcely distinguishable in color from the surface of the potato itself, and 

 perfectly characteristic as was afterwards learned, of the behavior of this organism on steamed 

 potato. 



(4.) The first sign appeared the fourth day after inoculation. The area in the vicinity of the 

 needle-pricks was the first part to become flabby and discolored. Within a period of 4 hours, on 

 the afternoon of September 5, the spot increased noticeably, being at least one-third larger at 4 o'clock 

 than at noon. By the sixth day the wilt had extended on one side to the extreme base of the leaf- 

 blade. The whole leaf drooped and two-thirds of it had changed color. The rest of the vine was 

 normal except the tip of one leaf which was bruised in repotting. By the eighth day the whole leaf- 

 blade had shriveled. On the eleventh day the blade of the leaf next above and of the leaf next below 

 the inoculated one began to droop slightly and on the twelfth day they were entirely shriveled, while 

 the next one above showed a tendency to droop. The thirteenth day the plant was photographed 

 (see plate 16, fig. 1 ). There were then 3 shriveled leaf-blades and 3 freshly-wilted leaves further up the 

 stem. At the top were three turgid leaves and at the bottom one. The fifteenth day the plant was 

 wilted throughout except the stem and the base of some of the petioles which looked normal. There 

 was no external indication of the cause of the disease. The stem was now cut open and examined 

 under the microscope: 6 inches below the inoculated leaf every vessel of every bundle contained 

 bacteria. Most of the vessels were gorged and large cavities had formed in the primary vessel- 

 parenchyma of three bundles. The bacteria here were not clearly motile. Six inches farther down 

 the vessels were still gorged but some of the bacteria were plainly motile. The tissue was less broken 

 down. The bacteria also occurred an inch from the tip of the stem, but less abundantly. The vessels 

 were full, however. The bacteria were also found in the petioles of the leaves where they were 

 abundant. The disorganization of the bundles in this plant had proceeded further than in No. 8, 

 examined the fourteenth day. Fourteen cultures were made from the interior of this plant as follows, 

 the stem being cut with a hot knife, and its interior dug into deeply with a flamed, steel-needle and 

 slime removed from the cavity : 



(1) 3 gelatin rolls; (2) 5 gelatin stabs; (3) 2 beef-broth tubes (1 peptonized); (4) 3 tubes of agar (stab cul- 

 tures). Two of the gelatin rolls remained sterile, one contained a mixed growth consisting of two small, white colonies, 

 two yellow ones, and a mold spore. Two of the gelatin stabs appeared to be pure cultures, the others were contami- 

 nated. The two beef-broths clouded typically, but on microscopic examination they were found to contain round 

 bodies as well as rods. The agar stabs all yielded typical, thin edged, gray-white, wet-shining surface growths. The 

 round bodies in the bouillon cultures may not have been contaminations as they afterwards appeared in tube 1, 

 September 17, made from vine 2. Also because a loop from one of these broths yielded a thin gray-white, wet- 

 shining, typical culture when transferred to potato.* 



(5 to 7.) One leaf-blade on each plant was inoculated with bacteria from a diseased squash-stem. 

 No result. The inoculated leaf did not wilt. Possibly the bacilli dried out in the pricks before they 

 got a start. 



(8.) One leaf-blade was inoculated with bacteria from a diseased squash-stem. The first signs 

 appeared the fourth day after inoculation. The area in the vicinity of the needle-punctures was at 



*In July, 1909, from a primary natural infection on a cucumber (petiole), a non-infectious white coccus was plated 

 out, B. tracheipkUus being dead. This coccus form is viscid and closely resembles B. tracheiphtlus on potato, but its 

 growth on agar, while smooth, is a denser chalkier white and it reddens litmus milk. It was stained by amyl Gram. 

 It did not grow in Fermi. 



