CHESTNUT BLIGHT FUNGUS 
sterilized. By this means the bark was transported to the laboratory 
where the tests were made on the following day. 
In the laboratory loo cc. of sterile tap water was poured into a 
shallow sterile dish. The piece of bark to be tested was trimmed down 
to 2 X 3 cm., stood on end in the dish of water, held by the corners 
and the outer surface scraped thoroughly with a cutting needle. 
The scalpels and needles were flamed before using, the hands and 
arms washed in mercuric chloride, and the entire operation carried on 
in a culture room. The dish was shaken at intervals for several hours , 
after which time i cc. of its contents was transferred to a flask con- 
taining ICQ cc. of sterile tap water. This flask was also permitted to 
stand for at least an hour, and was shaken at intervals. From this 
flask I cc. and fractions were transferred to sterile Petri dishes; and 
it was also found advantageous in many instances to pour several 
plates using fractions of i cc. from the wash water. Chestnut bark 
agar was used in all the cultures. 
Incubation took place at laboratory temperatures and the plates 
were carefully watched for at least ten days. This was necessary on 
account of the fact *that many of the spores of the blight fungus, 
although viable, would not begin to germinate at once, and colonies of 
Endothia parasitica frequently made their appearance 3 to 5 days 
after they were due under normal conditions. 
Discussion of Results 
Six series of tests, representing 36 pieces of healthy chestnut bark, 
were made between December 22, 1913, and June 4, 1914. Of these 
36 pieces only five failed to yield positive results. The remaining 
31 showed the presence of viable pycnospores of the chestnut blight 
fungus in numbers varying from 33 to 172,222 per square centimeter 
of bark surface (Table I). None of the colonies appeared in cultures 
at the time at which ascospore colonies usually show (3), but it should 
be stated that it was impossible to tell from the time of appearance 
of the colonies that all originated from pycnospores, on account of 
the frequency of delayed germination. It is a known fact, however, 
that the ascospores are forcibly ejected into the air, and that none 
are washed down the tree trunks as is the case with pycnospores (4, 5). 
We therefore feel certain that the colonies appearing in the cultures 
originated from pycnospores. 
Positive results were obtained almost uniformly during December 
