Jan. io, 1914 
Some Diseases of Pecans 
307 
Experimentation had already shown that the disease was readily con¬ 
trollable by Bordeaux mixture, and hence it was thought highly probable 
that it was of parasitic origin. Consequently, in the summer of 1912, 
materials for making cultures were taken directly into the field with ttie 
idea of locating the cause, if possible, by any of the ordinary methods of 
isolation. Leaves showing very recent infection were taken from the 
highest parts of the trees where there was little or no spattering from the 
soil. These leaves were placed in sterile Petri dishes and taken to the 
temporary laboratory, where the tiny spots were cut out at once with 
sterile scissors and transferred by the ordinary poured-plate method to 
Petri dishes of com meal and synthetic agar. 1 After 24 to 48 hours 
colonies became visible which had evidently originated from the diseased 
areas, and their appearance was quite uniform in all the cultures, except 
in a few cases where contaminations had entered. Transfers were then 
made to tube cultures. In this way strains of the fungus were obtained 
from Monticello, Fla., from Albany, Ga., and from San Antonio, Waco, 
and San Saba, Tex. 
Inoculations 
Circumstances connected with field travel prevented the making of 
any inoculation tests during the summer of 1912, but the following 
summer and winter trials were carried out upon pottted seedling trees in 
the greenhouse. The trees were sprinkled, inoculated from pure cul¬ 
tures, and covered after inoculation for several days with bell jars. Three 
strains of the fungus were used in this work: One from Texas, one 
from northern Florida, and a strain reisolated from an artificially infected 
leaf. 
Experiment No. i (Oct. 8, 1912).—The young leaves on four trees were inoculated 
from 1 ^-months-old, nonsporiferous synthetic-agar cultures (Florida strain 122), the 
slimy mycelial mass being smeared over portions of both leaf surfaces. These four 
trees and two moistened but uninoculated check trees were left under bell jars for 
five days. After a week small dark-brown specks were noted over the inoculated 
areas. In three weeks these spots were 1 to 2 mm. in diameter and in every way 
similar to natural infections. The check trees remained uninjured. 
Experiment No. 2 (Nov. 9, 1912).—The young leaves on three seedlings and the 
matured leaves on two others were inoculated as above only from 3-weeks-old, sporifer- 
ous com-meal-agar cultures (Florida strain 122). The five inoculated and five check 
trees were left under the bell jars for three days. Observation after two weeks showed 
the production of small, roundish, dark-brown specks, which at three weeks had 
become 1 to 3 mm. in diameter with small ashen-gray areas in the center. The lower 
1 Synthetic agar.—( i) 1,500 c. c. of distilled water and 36 grams of agar. Cook in double boiler for one 
hour at 15 pounds pressure. 
(2) 500 c. c. of distilled water, 200 grams of dextrose, 40 grams of peptone, 20 grams of ammonuim nitrate, 
5 grams of magnesium sulphate (crystals), 10 grams of potassium nitrate, 5 grams of potassium acid phos¬ 
phate (K2 HPCb), and 0.2 gram of sodium chlorid. 
Boil in double boiler for 30 minutes, add agar and cook for five minutes. Restore to volume, titrate, 
cool to 60 0 C., and add whites of two eggs. Cook to coagulate eggs, filter, tube, and sterilize. 
This formula is modified from that given by Francis Darwin and E. Hamilton Acton in their Practical 
Physiology of Plants, ed. 3, 1901, p. 68. 
I7°73°—!4- 3 
