ACTIVE IMMUNITY. 429 



virulence was not lost. Haffkine attenuated cultures of 

 the cholera spirillum by growing them in a current of air. 



(2) The virulence of an organism for a particular animal 

 may be lessened by passing the organism through the body 

 of another animal. Duguid and Burdon Sanderson found 

 that the virulence of the anthrax bacillus for bovine animals 

 was lessened by being passed through guinea-pigs, the dis- 

 ease produced in the ox by inoculation from the guinea-pig 

 being a non-fatal one. This discovery was confirmed by 

 Greenfield, who found that the bacilli cultivated from guinea- 

 pigs preserved their property in cultures, and could therefore 

 be used for protective inoculation of cattle. He also found 

 that the bacilli became attenuated when grown for successive 

 generations in aqueous humour. A similar principle was 

 applied in the case of swine plague by Pasteur, who found 

 that if the organism producing this disease was inoculated 

 from rabbit to rabbit, its virulence was increased for rabbits 

 but was diminished for pigs. Organisms which had been 

 passed through a series of rabbits produced in the pig illness, 

 but not death, and protection for at least a year resulted. 

 The method of vaccination against smallpox depends upon 

 the same principle. 



(3) Many organisms become diminished in virulence 

 when grown at an abnormally high temperature. The 

 method of Pasteur, already described (p. 293), for produc- 

 ing immunity in sheep against anthrax bacilli, depends 

 upon this fact. A virulent organism may also be attenuated 

 by being exposed to an elevated temperature which is 

 insufficient to kill it. Toussaint at an early date obtained 

 protective inoculation against anthrax by means of cultures 

 which had been exposed for a certain time to a temperature 

 of 55 C., though it is possible that in some cases the 

 bacilli were really killed, and immunity resulted from their 

 chemical products. 



(4) Still another method may be mentioned, namely, the 

 attenuation of the virulence by growing the organism in the 

 presence of weak antiseptics. Chamberland and Roux, for 

 example, succeeded in attenuating the anthrax bacillus by 



