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The Cultivation of Trypanosoma rhodesiense, Stephens and 

 Fantham. 



By H. Bayon, M.D., Genoa and Wurzburg, Beit Memorial Besearch Fellow, 

 Lister Institute. 



(Communicated by Sir Konald Boss, K.C.B., F.B.S. Beceived June 20, 1912.) 



Employing a strain of Trypanosoma rhodesiense kindly supplied to me by 

 Sir Bonald Boss, K.C.B., F.B.S., I have carried out some cultural experiments. 

 The following media were found to be successful. (1) Clegg's Amoeba agar 

 to which is added twice its amount of rabbits' blood which has been frozen 

 and thawed rapidly so as to cause the haemoglobin to diffuse into the serum ; 

 and (2) the following formula : — agar 15 grm., glucose 10 grm., water 1000 grm., 

 and twice the volume of rabbits' blood added. 



The trypanosomes were taken from sub-inoculated rats on the third day of 

 infection. Five drops of blood were placed on the agar, which is a diffluent 

 mass resting at the bottom of the culture tube. These were incubated at 22° 

 to 25° C. The tubes were examined after five days and then every two days, 

 usually, up to the 30th day. It was found that if the culture was successful 

 a multiplication of the trypanosomes occurred, which could be seen actually 

 taking place under the microscope. The trypanosomes were easily recognised 

 as T. rhodesiense because of the size and position of the kinetonucleus, the 

 length and breadth of the posterior end, and the position of the trophonucleus. 

 This trypanosome was differentiated from other easily culturable trypano- 

 somes, as for example T. lewisi, by inoculation into rats. 0"5 c.c. of the 

 diffluent culture injected intra-peritoneally caused T. rhodesiense to appear in 

 the blood 10 days after inoculation and the rat died in a further five days. 

 From these rats the trypanosomes were recovered and again cultivated. 



The forms of trypanosomes as commonly seen in these cultures are long and 

 slender with a definite flagellum. They move sinuously, with the flagellar end 

 first. The nucleus is longer and narrower than that seen in the blood, and 

 stains with G-iemsa's method a light purple. Volutin-granules are frequently 

 present. The kinetonucleus can be seen separated from the blepharoplast and 

 its usual position is somewhat to the side of the trypanosome. These forms 

 remain actively motile up to the 21st day of cultivation. They are virulent 

 up to the eighth day of culture in the quantity inoculated as mentioned 

 above. In certain cases rosette forms are imitated by the trypanosomes 

 clumping together ; however, a little pressure on the cover-glass causes them 

 to separate and to swim away individually. True brood-forms have also been 



