DopGE : METHODS OF CULTURE OF ASCOBOLACEAE 159 
water. In this case no surface film of moisture was visible on 
the agar nor on the cover of the dish. Twelve plates were pre- 
pared and the spores heated in the usual manner. No spores 
germinated. Small pieces (about 5 mm. square) of another agar 
medium containing a good supply of moisture were inserted in 
_ the plates of hard agar. Spores were then stabbed into the pieces 
of soft agar and heated. Practically all of the spores germinated. 
The mycelium grew out on the hard agar and produced a large 
number of apothecia in about ten days. This demonstrates the 
possibility of growth and reproduction under conditions not favor- 
able for spore germination. 
ASCOBOLUS CARBONARIUS Karst. 
In my first experiments on the effects of heat on the germina- 
tion of the spores of this species, small pieces of the apothecia 
were placed in Petri dishes on goose dung agar prepared as above 
described and placed in the sterilizing oven, which was then slowly 
heated to a temperature of 65° C., the time required being 30 
minutes. They were then removed from the oven and left at 
room temperature. The following morning, in the cultures that 
had been heated, hundreds of spores had germinated; not only 
ripe spores, but half grown, hyaline spores had germinated, sending 
out fully as long and vigorous germ tubes as the others. In the 
unheated control plates there was no evidence of germination. A 
medium was then made up with an extract of heated soil as a 
nutrient. Twenty plates of this agar medium were poured and 
imoculated with spores taken from the glass slides. Ten of these 
plates were heated to 65°—-75° C., the other ten were left at room 
temperatures. About 30 c.c. of the heated soil decoction was 
Poured into each of twenty more Petri dishes and ascospores sowed 
imthese. Ten of the dishes were heated to 65° C., and ten reserved 
unheated as controls. About 12 hours after, it was found that 
fully 90 per cent of the spores in both the liquid and the solid media 
that had been heated had germinated, while there was no germina- 
tion in the unheated controls. Many small hyaline spores had 
also germinated. Comparison showed that these were half grown 
“pores of A scobolus carbonarius which had failed to reach maturity 
before being expelled from the asci. Further attempts to induce 
