198 
Journal of Agricultural Research 
Vol. XXVI, No. 5 
was infested with Helminthosporium sativum, which develops on many of 
the wild grasses. This necessitated sterilizing the soil by the pressure- 
steam method for varying periods, depending on the pressure used. 
Four hours at i-pound pressure or less and one hour at 10 to 15 pounds 
gave satisfactory results. This soil after sterilization had a moisture¬ 
holding capacity of 67 per cent. Two varieties of wheat, Marquis (spring) 
and Harvest Queen (winter), and Hannchen and Hanna varieties of 
spring barley were used in these experiments. All seed was surface 
sterilized with a solution of mercuric chlorid and water (1:1,000) for 10 
minutes and thoroughly rinsed in sterile water before sowing. It was 
very difficult to obtain seed free from Helminthosporium infection and, 
as surface sterilization is not effective in controlling this infection, such 
seed had to be guarded against. One sample of Harvest Queen seed 
from the uplands of Madisun County, III., was for the most part free 
from infection, and this was used in much of the work. A small amount 
of Marquis seed, kindly supplied by G. H. Dungan, of the Illinois Agri¬ 
cultural Experiment Station, also proved to be practically free from 
infection, and the same was true of the seed of Hannchen and Hanna 
barley from the Aberdeen (Idaho) plots of Dr. H. V. Harlan, of the 
Office of Cereal Investigations, United States Department of Agriculture. 
The organisms used in the inoculations consisted of three single-spore 
strains of Helminthosporium sativum. The first, designated No. 51a, was 
isolated by the writer in May, 1920, from the crown of a Harvest Queen 
wheat plant, in the advanced stages of the rosette disease, growing near 
Granite City, Ill. The second, designated No. 350, was isolated by the 
writer in April, 1921, from an infected barley kernel obtained from a lot 
of seed grown in the vicinity of La Fayette, Ind. The third, designated 
No. 392, was isolated by Dr. R. W. Webb in the spring of 1921 from the 
same type of plant and from the same source as culture 51a. 
These strains were cultured on potato-glucose agar in Petri dishes. 
The spores were scraped from the surface of the medium and put into 
water. These spore suspensions were then used to inoculate the seed 
or the soil before sowing. 
In the case of seed inoculation, a given volume of spore suspension 
was placed in a test tube, such volume being just enough to moisten the 
number of seeds to be sown in a single pot. The suspension was care¬ 
fully measured by means of a pipette so as to insure uniformity of inocu¬ 
lation and then put into as many test tubes as there were pots to be 
inoculated. This measuring procedure was followed at the beginning of 
the inoculating operation. The seeds were previously counted out in 
definite numbers for each pot. At the time of sowing a particular pot, 
the seed was poured into the test tube of inoculum, well shaken, and 
emptied into a Petri dish, the small excess of suspension was drained off, 
and the seeds were quickly planted by means of forceps. Seeds were 
not introduced into the inoculum until just before planting. All seed 
was sown 1.5 inches deep. 
Owing to the fact that the spores of Helminthosporium sativum do not 
germinate to any extent in large quantities of water, no bad effects come 
from preparing all of the suspensions at the beginning of the sowing 
operations. The writer has had a spore suspension of this organism in 
the laboratory from April to November, 1921, with practically no germina¬ 
tion. Sowings of these spores were made on potato-glucose agar from 
time to time, and good germination took place until the latter part of 
the period, when the viability of the spores seemed to go down rapidly. 
