78 Messrs. J. G. Thomson and D. Thomson. 



So far, only the asexual generation of the malaria parasite has been grown 

 in vitro. The next step is to cultivate the sexual generation as it occurs in 

 the human host and in the mosquito. "We have attempted both, but, so far, 

 without definite success. We have no doubt, however, that this will also 

 be accomplished sooner or later. In 1912, Thomson, J. G., and Sinton 

 successfully cultivated the human trypanosome, and the development they 

 obtained was apparently that which takes place in the stomach of the tsetse 

 fly. Joukoff (1913) states that he has cultivated the mosquito cycle of the 

 malarial parasite, though his results have not yet been confirmed. There 

 is no reason to doubt that the phases of development of protozoa in insects 

 may be produced in the culture tube. 



Cultivation Technique, 



Our method of cultivation is practically the same as that of Bass and 

 Johns, except that it is less complicated ; 10 c.c. of blood is drawn from a 

 vein and transferred to a sterile test-tube containing a thick wire leading 

 to the bottom of the tube from the cotton wool plug. 1/10 c.c. of a 

 50-per-cent. solution of glucose is added to this tube, preferably before 

 adding the blood. The blood is defibrinated by gently stirring with the 

 thick wire. Defibrination should be complete in about 5 minutes. The 

 wire with the clot is then removed and the blood is poured into several 

 smaller sterile tubes (about 1-inch column of blood in each). A rubber cap 

 is placed over the cotton wool plugs to prevent evaporation and the tubes are 

 then transferred (standing upright) to an incubator at a temperature of 37° 

 to 41° C. The corpuscles settle in a short time, leaving about half an inch of 

 clear serum at the top. It is apparently unnecessary to remove the 

 leucocytes by centrifugalisation. 



Further Observations on Cultivation. 



We have grown four complete generations of parasites in one tube by the 

 above method, and we do not see why their growth should not continue 

 indefinitely, provided fresh serum and corpuscles be added. It is not the 

 presence of leucocytes which prevents further development. This is due to 

 degenerative changes taking place in the corpuscles and serum. If the serum 

 and corpuscles be kept in a sterile condition in an ice chest they remain 

 unchanged for a long time, but at the temperature required for the growth of 

 the parasites visible changes take place in a few days. The corpuscles 

 become fragmented and form a brownish debris and the serum becomes dark 

 brown in colour. When this occurs the parasites are unable to continue 

 their developmental cycle. Bass and Johns, in their original paper (1912), 



