Nov. 1,1924 
Altemaria Leqfspot of Cauliflower 
435 
The above experiments indicate that 
a strong invertase; a weak amylase, if 
any; no cytase; and no pectinase were 
produced. To obtain further confirma¬ 
tion regarding the production of these 
enzyms by A. brassicae, fifty 200-cc. 
flasks, each containing 50 cc. of sweet- 
potato decoction, and a like number 
containing cabbage decoction, were pre¬ 
pared and inoculated. Several unin¬ 
oculated flasks were held as controls. 
The Czapek’s solution used in the ear¬ 
lier experiments did not appear to con¬ 
tain all the substances necessary for the 
best development of the fungus, hence 
these vegetable decoctions were used 
and proved to be very satisfactory; 500 
gm. of the vegetables per liter were used 
in preparing the decoctions. Part of 
the inoculated flasks were held at 31° 
in the dark and the remainder were 
placed at room temperature (25°) and 
in diffused light. After 12 days a 
thick mycelial felt was present on all 
of the inoculated flasks. The amount 
of growth on these media was very 
much better than that made on 
Czapek’s solution. The mycelial felt 
was very dark olive to almost black 
in color. Spores were present in 
great abundance. It might be added 
here that the fungus seems to sporulate 
even under the most adverse condi¬ 
tions, spores having been formed in 
considerable quantities even in Czapek’s 
solution without carbon. Further¬ 
more, it was found in the temperature 
experiments previously recorded that 
spores were formed at temperatures so 
high that they were extremely inju¬ 
rious to vegetative growth. In fact, 
sporulation seems to take place under 
any condition which will permit growth. 
The felts were removed from the 
sweet potato and cabbage decoctions, 
both those in the light and in the dark. 
Each set was kept separate through¬ 
out. These felts were washed in run¬ 
ning tap water for a few minutes to 
remove all traces of sugar and were 
then treated with acetone and ether in 
the manner described by Harter and 
Weimer (10) . The hyphae were then 
weighed out in 0.5 gm. samples, each 
being placed in a small Erlenmeyer 
flask. Twenty-five cc. of distilled 
water were added to two flasks of each 
set. To these flasks were added disks, 
1 cm. in diameter by 0.5 mm. thick, of 
sweet potato, carrot, and Irish potato 
and similar disks cut from the laminae 
of cauliflower leaves as well as small 
portions of the cauliflower curd. 
Twenty-five cc. of 0.75 per cent cane- 
sugar solution were added to each of two 
flasks containing hyphae grown on 
sweet-potato decoction in the light, to 
each of two flasks containing hyphae 
grown on sweet potato decoction in the 
dark, and to each of two flasks con¬ 
taining hyphae grown on cabbage de¬ 
coction in the dark. Twenty-five cc. 
of a 0.75 per cent starch paste and 25 
cc. of a cellulose suspension were added 
to each of two flasks containing hyphae 
from the same sources. Likewise, 
flasks containing the same amount of 
hyphae from the same sources with 25 
cc. of distilled water added were held 
as controls. Samples of the cane- 
sugar solution, starch paste, and cellu¬ 
lose suspension were also held as 
controls; 5 cc. of toluol were added to 
all of the flasks as an antiseptic, and 
they were then held at 45°. 
No maceration was evident in any of 
the flasks containing the hyphae and 
portion of vegetable in 96 hours, at 
which time the experiment was dis¬ 
continued. Here, as in the former test, 
the presence of pectinase could not be 
demonstrated. 
After 24 hours the flasks to which the 
cane sugar had been added were steam¬ 
ed and their contents tested for reduc¬ 
ing sugars by the Clark method (4). 
At the end of 30 hours the flasks to 
which the starch paste and cellulose had 
been added were similarly treated and 
tested. The control flasks containing 
samples of the cane sugar, starch paste, 
and cellulose were also tested. Like¬ 
wise, the control flasks containing 
hyphae and water were tested in the 
same way. The purpose of holding the 
hyphae in water was to determine the 
reducing sugar formed as a result of 
autolysis of the fungus itself. The 
average amount of reducing sugars 
present in the two flasks of each set 
minus that formed by autolysis and that 
in the original solutions are given in 
Table III. The data presented there 
show that a considerable quantity of 
both the cane sugar and the starch was 
hydrolyzed by the enzyms contained in 
the hyphae, denoting that invertase 
and amylase were produced by this 
fungus when growing on either sweet- 
potato or cabbage decoction. The 
number of tests made was not large 
enough to make the figures very 
accurate from a quantitative stand¬ 
point, but they are of value in that 
they show that considerable reducing 
sugar was produced in both the cane- 
sugar and starch solutions. Apparently, 
slightly more invertase was produced 
by the fungus when growing on cabbage 
than on sweet-potato decoction. How¬ 
ever, this difference in the results 
obtained may be within the range of 
experimental error. On the other hand 
the reverse seemed to be true of 
amylase. Light appeared to have a 
detrimental effect on both enzyms. 
