18 



youngest unfolded leaves and also into the healthy portions of the 

 older ones. Besides brushing- in the cultures, al^rasion inoculations and 

 also h3"podermic injections with an ordinary hypodermic syringe were 

 made. 



Ten plants were thus treated with B. dianthi during the season of 



1897, the three methods of infection being followed in the case of dif- 

 ferent branches of each plant, but the disease was not produced in any 

 case. An occasional spot developed on some of the plants after treat- 

 ment, but such spots were always close to a spot which had been previ- 

 ously marked with India ink, and so could not be attributed to infec- 

 tion b}^ the germs applied. Negative results also were obtained with 

 the white germ mentioned. 



Further infection experiments with B. dianthi were carried out in 



1898, and these will be described later on in this bulletin. The results 

 of the writer's work so far showed no evidence that the disease was 

 caused by bacteria, and a search was therefore instituted for other 

 possible external causes. 



FIRST COLONIZATION EXPERIMENTS AVITH APHIDES. 



Arthur and Bolley regarded aphides as "such efficient bearers of 

 the contagion that every leaf on a plant ma}^ be inoculated at hundreds 

 of points," and indeed the writer observed that the spots were 

 extremely common on many plants which had been attacked by the 

 aphides. A careful study revealed the fact that carnations are seldom, 

 if ever, entirely free from aphides — they may often be found between 

 the 3^oung appressed leaves of plants apparently free from them. 

 Ten 3"oung plants were taken into the laboratory and entirely freed 

 from the insects by first picking off all that could be found, and then 

 fumigating the plants lightly with hydrocyanic acid gas. The plants 

 were kept unaer moist bell jars, and as soon as growth free from spot 

 had developed aphides were colonized on various leaves. In the case 

 of the youngest, most rapidly growing leaves spots could be detected 

 with a hand lens three days after the tissues had been punctured by 

 the aphides; in leaves somewhat older a longer time elapsed before the 

 spots became visible; while in leaves half size, especially if the plant 

 was making slow growth, the spots could not be seen for about two 

 weeks after the puncture was made. 



The spot appeared first as a minute translucent dot, accompanied by 

 a .slight .swelling of the tissues. On some of the very young leaves the 

 spots were unusually large, being from 2 to 3 mm. in diameter h\ the 

 time the leaf had reached full size. Sections were cut from the spots 

 so produced and were stained as before described, but no parasitic 

 organisms could be found in the tissues. The cells had enlarged and 

 the chloroplasts had lost their colox and had failed to develop or had 

 shrunk in the manner characteristic of the disease. The peculiar 



