248 Cultivation of Piroplasma 
L. Following the method of Miyajima (1907) (see p. 255) numerous 
cultures were made by mixing defibrinated blood and ordinary nutrient 
broth in proportions varying between 1—5 and 1—10. These were 
kept for various periods at temperatures rangiug between 18°C. and 
35° C. Though the red blood corpuscles were well preserved the 
parasites rapidly became rounded and lost their motility. No develop¬ 
mental forms like those described by Miyajima were seen and none 
with radiating processes. 
M. In the series of experiments about to be described certain very 
large irregular intracorpuscular forms with radiating spike-like processes 
were occasionally met with, apparently identical with the cultural forms 
described by Kleine, and resembling the free parasites observed by 
Koch in the early stage of infection in the tick. 
In the following description these are spoken of as forms with 
radiating processes and the processes as radii. 
In all the following experiments the cultures were made by adding 
a quantity (about 05 c.c.) of blood, defibrinated by shaking with glass 
beads for 20 minutes, to an equal quantity of 0‘6°/ 0 or 0'8°/ 0 salt solution 
or to a physiological saline solution (hereinafter referred to as “ P ” 
solution) with the following composition: 
Sodium chloride 
0*95% 
Potassium chloride 
0-025% 
Calcium chloride 
0-02% 
Sodium hyd. carbonate 
0-15% 
Dextrose 
0-1% 
Distilled water ... 
100 c.c. 
Dog I. The cultures were made in O'6°/ 0 salt solution. Some of 
these cultures were kept in test tubes plugged with cotton wool, and 
others in Petri dishes, slightly tilted, at various temperatures (14° C., 
16° C., 20 C. and 32° C.). Examinations of fresh and stained prepara¬ 
tions were made at various times. At 14° C. after 20 hours a few forms 
showed short blunt processes. At 16° C. no forms with radiating 
processes were seen. At 20° C. after 16 hours one form with short 
radii was found, but at 32° C. several parasites showing radii were 
observed after 24 hours. These experiments seemed to indicate that a 
temperature between 20° C. and 32° O. was the most suitable for the 
development of forms with radiating processes, and consequently in the 
subsequent experiments the cultures were kept at 24° C.—26° C. 
Dog II. Cultures were made in 0'6°/ 0 salt solution and were kept 
