Keene—Studies in Zygospore Formation. 1205 
rye) and rice. In every case zygospores have been secured 
when the medium was inoculated with spores from both strains, 
while at no time and on none of the above-mentioned media, 
have zygospores been obtained with the pure strains. 
No difficulty whatever was experienced in any inoculations 
in securing a plentiful production of zygospores when the two 
strains were inoculated into the same medium. The matter 
of humidity seems to be a very important secondary condition 
and the best results are invariably obtained if the cultures are 
kept in a cool, humid place. As soon as the medium begins 
to dry out, zygospore production ceases and there occurs a 
marked increase in vegetative development. Considerable dif¬ 
ficulty was experienced at first in bringing the zygospores to 
maturity, but later it was discovered that moisture conditions 
were responsible. If Petri dishes were used it was essential 
that a very heavy layer of agar be supplied, and the softer agars 
with high moisture content were more satisfactory. 
This substantiates the work done by Klebs (1898, 1902) in 
which he pointed out the relation of transpiration to spore 
formation in Sporodinia grandis, and corroborates the results 
obtained by Blakeslee (1904) on Rhizopus nigricans. There 
seems to be no evidence in Phycomyces, at least, that zygospore 
formation is affected by the concentration of the medium. 
Cytology of the Zygospore 
When the two branches from sexually different strains of 
Phycomyces nitens come together there occur a branching and 
lobing which tend to interlock the hyphae (PI. I, figs. 1 and 2). 
Previous to contact it is impossible to tell the gametophoric 
hyphae from any of the other parts of the mycelium. The 
nuclear conditions at this stage are the same as in the resting 
mycelium, as far as it has been possible to determine. There is 
no evidence of any excessive streaming or increase in the num¬ 
ber of nuclei. As the branches increase in length, they become 
erect, the terminal portions elongate, their inner surfaces lying 
parallel and appressed (fig. 3). The gametophores at this time 
are usually characterized by a yellow color which is due to the 
presence of oil as shown by tests. As the progametes continue 
