DopGE : METHODS OF CULTURE OF ASCOBOLACEAE 155 
to transfer these germinated spores to the agar plates. When the 
spores were removed from the slide the germ tubes were either 
broken off or injured in some manner, but if the slides themselves 
were placed on the medium the mycelium would continue to grow 
and finally produce ascocarps. These cultures invariably became 
contaminated with foreign fungi. 
Small pieces of dung or filter paper upon which young ascocarps 
were growing, were next placed in the agar medium. Although 
many attempts were made to obtain cultures in this manner, not 
the slightest further growth of the mycelium was obtained. 
Several plates prepared in this manner were placed in a drying 
oven and heated slowly for 40 minutes. The temperature of 
the oven was thus gradually raised to 80° C. The plates were 
then withdrawn and allowed to stand in the laboratory at 
foom temperatures. After 24 hours it was found that the 
spores thrown out upon the medium from the mature ascocarps 
before the cultures had been heated, had germinated and the 
mycelium was growing vigorously. This experiment was re- 
peated with a number of plates, and unheated controls were main- 
tained at room temperatures. In all the plates that had been 
heated the spores germinated, while none of the spores in the 
controls did so. 
A series of experiments, some of which are tabulated below, 
Were made to test more fully the effect of heat on germination 
and to determine the approximate minimum, optimum, and 
maximum temperatures for spore germination. Controls at room 
temperatures were maintained in ninety cases. It will be seen from 
the table that about 80 per cent of the spores heated to 60°-70° C. 
germinated. In no case were the spores in the controls even 
swollen. The method as finally worked out may be described as 
follows: Spores for inoculation were obtained by laying glass slides 
°n corks over pieces of dung bearing ripe ascocarps. With the 
use of a Zeiss binocular, the spores were removed with a sterilized 
Platinum needle and stabbed into the medium about 1.5 cm. from 
the edge. If more than one plate was to be inoculated it was 
found necessary to moisten the spores by blowing the breath on 
the slide, since they dry out rapidly and adhere to the slide so 
firmly that their removal is impossible without destroying them. 
