287 
Cytology of Pythium tiltimum, n. sp. 
remained fresh for days. It must be remembered that if 
boiled cress-seedlings are used the material is attacked at 
once, and vigorous growth is the result. The species is 
clearly a true saprophyte. 
Observations on Serial Sections. 
Preparation of the material. At first Petri-dish cultures, 
three, four, or more days old, were taken and examined in situ 
under the microscope. If in suitable condition they were 
fixed in Hermann’s fluid, while still in the dish and under 
microscopic control. They were then passed into paraffin 
through the medium of xylol, and the material was finally sec- 
tioned thick and stained with gentian violet. A complete 
series of preparations was found, illustrating all stages in the 
cytology, and then it became evident that the method was 
not an absolutely satisfactory one. The nuclei could not 
easily be followed at the stage immediately succeeding fertili- 
zation, as had been the case indeed in previous years with 
Saprolegnia and Achlya , and in addition there was a difficulty 
in following the details in the maturation of the oospores. 
The preparations used to furnish the illustrations were with 
four exceptions made as follows : — 
Absolutely pure cultures were grown in Petri dishes and 
fixed under microscopic control by immersion in Flemming’s 
stronger solution for periods of five hours or more. The 
blackening was removed by hydrogen peroxide and the de- 
hydration carried out with great caution. No collapse took 
place. Here the microscopic control came to an end. The 
material was transferred to paraffin through the medium of 
xylol. The alcohol was gradually displaced by xylol and the 
xylol by paraffin. Notwithstanding these precautions, shrink- 
ing and some amount of collapse took place at this stage, 
although curiously enough chromacetic acid material passed 
through the imbedding oven at the same time under precisely 
similar conditions showed no shrinking of any kind. Sections 
were cut of 5 \i thickness and stained with gentian violet. 
