﻿CULTIVATION OF A BOVINE PIROPLASMA. 85 



As is already known, the parasite Piroplasma parvum is also char- 

 acterized by the fact that the infection can not be transferred by the 

 inoculation of the blood itself, and in this respect we have observed that 

 it differs greatly from its allied organisms, P. ligeminum, P. carus, etc. ; 

 however, on the other it shows a great similarity to "Haemoprotens" 

 which is a common parasite in the blood of birds. 



Observation 2. — Since the discovery of the important role taken by 

 ticks in infections with piroplasmata, many observers have in vain endea- 

 vored to prove the existence of the developmental forms in the bodies 

 of the ticks which live on infected animals. Very recently an actual 

 demonstration of a developmental change of the piroplasma was given 

 for the first time by E. Koch. 9 He described certain developmental 

 forms of Piroplasma oigeminum and P. parvum, the final stages of which 

 are as yet unknown. After his work was published, we endeavored to 

 repeat his experiments by feeding the cattle tick Rhipicephalus australis, 

 which is found abundantly on the native cattle of Japan and is regarded 

 as being a probable carrier of Piroplasma parvum. After several unsuc- 

 cessful trials, my efforts were directed toward the cultural method by 

 which Eogers and other observers were able successfully to demonstrate 

 the flagellate stage of the Leishman-Donovan bodies. 



The following different culture media were tested in our preliminary 

 experiments on the cultivation of the piroplasma; blood agar, sodium 

 citrate, both acidulated and nonacidulated ; beef extract, peptone water; 

 calf's serum; physiologic salt solution; common bouillon, etc. On the 

 4th of July, 1906, we first observed a few motile organisms in one test tube 

 which contained a small amount of infected blood mixed with acidulated 

 sodium citrate solution, prepared according to the method of Eogers. 

 The entire series of cultures was then carefully examined but no evidence 

 of development of any motile organism was obtained except in the 

 bouillon ones. This medium was apparently the most favorable, large 

 flagellates having abundantly developed therein after four days' incuba- 

 tion at a room temperature. Only a slight multiplication of the or- 

 ganisms took place in the first culture which we obtained in acidulated 

 sodium citrate. 



The length of the organism at its full-grown stage, as it is observed 

 in cultures, is about five times the diameter of an erythrocyte. It, at 

 this time, possesses a well-defined undulating membrane and a long 

 flagellum. The position of a nucleus and blepharoplast in the body of 

 the flagellate renders it impossible to distinguish it from a typical try- 

 panosoma developed in a culture. (PI. Ill, fig. 2.) 



The method which we have employed is a simple one, and is practically 



"Ibid (1905) 47, 1865, and Ztschr. f. Hyg. u. Infectionskrankh., Leipz. (1906) 

 54, 1. 



