DIRECT INOCULATION OF CONIFEROUS STEMS WITH 
DAMPING-OFF FUNGI 1 
By Annie Rathbun-Gravatt 
Junior Pathologist , Office of Forest Pathology , United States Department of 
Agriculture 
INTRODUCTION 
Hartley’s (2, 3 ) 2 and Spaulding’s ( 8) 
summaries of the important literature 
on the damping-off of conifers have 
brought out the fact that aerial stem 
infections are less frequent than sub¬ 
terranean infections. Most of these 
published reports have been based upon 
field observations or upon experiments 
in which inoculum was added to the 
soil at the time the seeds were sown, 
instead of being applied directly to the 
seedlings. These methods, while giv¬ 
ing the combined loss from the various 
types of damping-off, (germination 
loss, root damping-off, stem damping- 
off, etc.), are not entirely satisfactory 
for showing just what part of this loss 
each type causes. Results secured 
from the direct inoculation of coniferous 
taproots have been published pre¬ 
viously ( 6 ). This paper will present 
the results from the direct inoculation 
of coniferous stems. 
METHODS OF INOCULATION 
The seedlings for most of the experi¬ 
ments were grown in autoclaved sand 
in 2-inch pots in the greenhouse of the 
United States Department of Agri¬ 
culture at Washington, D. C. They 
were watered only with boiled water. 
The pots were kept from the time of 
seeding until the time of inoculation in 
glass-walled cupboards previously dis¬ 
infected with mercuric chloride. At 
the time of inoculation the pots were 
taken from the cupboard and all but 
10 seedlings were removed from each 
pot. The seedlings were inoculated 
while the seed-coats were still clinging 
to the cotyledons of most of them. 
The seedlings grown in this way, 
though normally green, were taller 
than seedlings of the same age usually 
are, the light intensity in the glass 
cupboards being apparently insuffi¬ 
cient for normal growth. 
The original plan was to inoculate 
each seedling at the soil surface be¬ 
cause usually in seedbeds aerial infec¬ 
tions occur at the soil surface rather 
than above. Almost immediately, 
however, this method was discarded 
in favor of inoculation platforms 
(fig. 1), since it was found very im¬ 
practical to test the fungi by inocu¬ 
lation at the soil surface without the 
fungi’s growing from one seedling to 
another through the soil. The method 
adopted is believed to give a fair in¬ 
dication of the results which would 
be obtained at the soil surface. The 
platforms consisted of equilateral tri¬ 
angles with sides three-eighths of an 
inch long cut from library cards. A 
toothpick was inserted through a hole 
in the center of each triangle. An inocu¬ 
lation platform was placed beside each 
seedling, and upon it in contact with 
the stem was placed the inoculum. 
Each toothpick was pushed into the 
soil until the triangular platform was 
about three-eighths of an inch above 
the soil surface. 
Inoculum for each seedling consisted 
of either a 4 by 4 by 2 mm. cube of 
corn-meal agar, or rice mush about 
the size of a swollen rice grain, plus 
the mycelium and any spores growing 
upon it. In case both kinds of inocu¬ 
lum were used in the same experiments 
one kind was used on 5 seedlings, and 
the other kind on the 5 seedlings on 
the other side of the same pot. Each 
control pot was divided in the same 
manner, each half being treated with 
one kind of sterile medium. In each 
experiment there were usually 150 
control seedlings, 75 for each medium 
when two kinds were used. At least 
10 seedlings were inoculated with each 
fungous line grown on each medium. 
Individual damp chambers were 
made for each pot, consisting of trun¬ 
cated sheet celluloid cones 5 inches 
high and 2^ inches in diameter at the 
base. Moisture was furnished by 
i Received for publication, June 5, 1924—issued, April, 1925. 
s Reference is made by number (italic) to “Literature cited,” p. 339. 
Journal of Agricultural Research, 
Washington, D. C. 
( 327 ) 
Vol. XXX, No. 4 
Feb. 15,1925 
Key No. G-407 
