Apr. 15,1914 
Experiments with Apple Leaf-Spot Fungi 
59 
to seven days either by inclosure in paper bags or by beginning the experi¬ 
ments during an extended rainy period. It soon became evident that 
zonate enlargements were developing from some of the check spots as 
well as from some of the inoculated ones, and in nearly every case the 
species of Alternaria appeared in culture from these enlargements. It 
was suspected, however, that the Alternaria was only a saprophyte and 
that water standing on the leaf for some time and soaking into the dead 
spot had thence reached the intercellular spaces of the adjacent living 
portion of the leaf and caused death by preventing gaseous exchange. 
Accordingly it was thought that if a branch of a susceptible variety with 
spots burned on its leaves were placed under very moist conditions the 
zoned areas could be induced quickly and abundantly. 
On July 8 a branch of the York Imperial variety having sound leaves was selected 
and the cut end placed in a flask of distilled water. Without previous washing or 
sterilization the leaves were scorched with the heated end of the steel rod. The 
branch was then drenched with distilled water and placed under a bell jar partly lined 
with wet filter paper. Twice during the experiment the branch was taken out, and 
after the water on the leaves had been allowed to evaporate it was again drenched and 
placed under the bell jar. Under these conditions the leaves of the York Imperial 
apple held up as well as if they had been on the tree, there being no noticeable etiola¬ 
tion. In nine days 86 spots out of in, or 77 per cent, had enlarged zones from 1 to 4 
mm. in diameter, there being two or three zones to each spot. Such enlargements 
were then cut out from the leaves, dipped for a moment in 95 per cent alcohol, in mer- 
curic-chlorid solution (1 to 1,000) for two minutes, in sterile water for five minutes, and 
then placed upon culture media. Of these cultures 17 out of 21 developed the Alter¬ 
naria and 4 were sterile. 
On July 19 this experiment was repeated, with the addition of two others, as follows: 
Experiment i.— Healthy leaves on branches of York Imperial apples were scorched 
with the heated end of a steel rod, sprinkled with distilled water, and placed under a 
bell jar lined with wet filter paper. 
Experiment 2.—Treatment was the same as in No. 1, except that the branches 
were first immersed in 95 per cent alcohol for a moment, then in mercuric-chlorid solu¬ 
tion (1 to 1,000) for two minutes, and washed in sterile water for five minutes. 
Experiment 3 . —One branch was treated as in No. 1 and one as in No. 2 , but neither 
was placed under a bell jar or kept moist in any way. 
The results after nine days are summarized in Table I. 
TabeE I.— Results of inoculation experiments of July IQ on branches of York Imperial 
apples. 
Experiment No. 
Number of 
burned 
spots. 
Number of 
enlarged 
spots. 
Average 
width of 
enlarge¬ 
ments. 
Percentage 
of enlarged 
spots. 
Mm. 
71 
7 
72 
44 
3 
3 
7 
3. 
39 
0 
0 
O 
Cultures were made as in the previous experiment, except that pieces of material were 
left in the mercuric-chlorid solution for three minutes. Of these 9 out of 17 developed 
the Alternaria, 7 contained bacteria, and 1 was sterile. In these experiments it will 
be seen that sterilization of the leaf surfaces practically prevented the disease. 
On July 28 a series of four experiments was begun. Experiments 1,2, and 3 were 
performed in the same manner as Nos. 1,2, and 3 of July 19. Experiment 4 was per¬ 
formed in the same manner as experiment 2, except that after the washing in sterile 
water the leaves were sprayed with water containing Alternaria spores from cultures 
made from the spot enlargements of the experiment of July 8. 
