10 BULLETIN" 924, U. S. DEPARTMENT OF AGRICULTURE. 
which had been isolated from an injured grapefruit leaf. The 
spores were washed off the twigs and mixed with those from the 
culture tubes. This wash water, which was clouded with fresh viable 
spores, was used in saturating wads of absorbent cotton, which in 
turn were placed on the fruit. The inoculated fruit was covered 
with two or three layers of waxed paper for 48 hours. At the 
expiration of this time the paper and wet cotton were removed and 
the fruit left unprotected. To serve the purpose of a control, other 
fruits were similarly treated, except that the cotton was wetted 
with sterile water. These inoculations gave negative results. 
Similar inoculation tests were made during the fall of 1919, using 
as inoculum a mixture of a number of strains of ColleiotricJium gloec- 
sporioides isolated from typical tear stains on grapefruit and from 
dead grapefruit twigs, as follows: 
On October 20, 1919, on almost fully grown grapefruit; on October 27, on grapefruit 
showing faint yellowing ; on November 7 , on grapefruit approximately one-half colored ; 
on November 15, on grapefruit almost fully colored; and on December 3, on mature 
grapefruit. This test was repeated during the late spring, summer, and fall of 1920, 
using mixed inoculum from the same strains of Colletotrichum gloeosporioides. Inocu- 
lations were made on May 15, on grapefruit averaging 1 inch in diameter; on May CI, 
on grapefruit averaging 1J inches in diameter; on June 15, on grapefruit averaging 
If inches in diameter; on June 30, on grapefruit averaging 2^ inches in diameter; 
on July 15, on grapefruit averaging 2 J inches in diameter: on July 30, on grapefruit 
averaging 3 inches in diameter; and on November 1 and 6. on grapefruit just be- 
ginning to color. 
The results of all these tests were negative. Not the slightest 
symptom of tear-stain in any of the fruits inoculated during 1919 
could be detected as late as February, 1920, when the crop was har- 
vested. The fruit inoculated during 1920 was free from tear-stain 
when final observations were made in November. 
Observations were made to determine the frequency of association 
of tear-stain with dead twigs that might harbor Colletotrichum or 
other fungi. Unsprayed groves with more than the average propor- 
tion of dead wood present were examined carefully during the past 
four years. Among the properties inspected in Florida 4 are in Lee 
County, 10 in Polk County, 2 in Hillsboro County, 3 in Pinellas 
County, 3 in Osceola County, 10 in Orange County, 2 in Volusia 
County, 4 in Brevard County, 3 in St. Lucie County, and 4 in Dade 
County. The data obtained indicate that dead twigs, spurs, etc.. are 
found immediately over not more than 10 per cent of the tear-stained 
fruit, and that in damp, densely shaded, low-hammock properties, 
where trees have an unusually large number of dead twigs and where 
environmental factors would appear to be especially favorable for the 
development of fungi, tear-stained fruits are very seldom found. On 
the other hand, tear-stain is most abundant in higher and drier lo- 
cations where light and moisture favor the greatest rust-mite 
development. 
