62 PARASITIC AMOEBJS OF MAN. 



In 1897, Frosch demonstrated that it was pos- 

 sible to grow free-living amoebae upon culture media, 

 provided bacteria were present at the same time, and 

 that unless such bacteria were present it was impos- 

 sible to carry the organisms from one culture to 

 another. In the same year Casagrandi and Barba- 

 gallo confirmed these observations, but considered 

 that the association of the amoebae and bacteria was 

 simply an accidental one. Tsujitani, in 1898, cul- 

 tivated amoebae in association with cholera vibrios, and 

 by heating the cultures at 60 degrees C. he killed 

 the vibrios, thus obtaining a pure culture of the 

 amoebae. He found, however, that while the encysted 

 amoebae upon the cultures were capable of motility 

 and growth, multiplication did not occur when they 

 were transferred to sterile culture media. His ob- 

 servations, together with those of Zaubitzer, Mouton, 

 and Schardinger, definitely proved that it is impossible 

 to transplant pure cultures of amoebae free from all 

 bacteria. 



The most extensive work done upon the cultiva- 

 tion of the parasitic amoebae of man is that of Mus- 

 grave and Clegg. In 1904, these investigators pub- 

 lished a monograph upon this subject in which a 

 complete review was given of the literature, together 

 with their methods and results. They claim to have 

 been able to cultivate dysentery amoebae in symbiosis 



