﻿STUDIES IN PLAGUE IMMUNITY. 301 



As mentioned above, Otto showed that it was not possible to increase 

 the pathogenicity of strains of pest bacilli of moderate virulence by 

 repeated passages through animals. The strains with which he per- 

 formed his experiments Avere all capable of killing guinea pigs in 

 moderate doses at the time of the commencement of the work. 



I have attempted to increase the virulence of a more attenuated cul- 

 ture of the pest bacillus (one which in large doses was not capable of 

 killing guinea pigs) by experiments performed on monkeys. The strain 

 "A virulent Manila" was employed for the inoculations. This organism 

 was chosen because it had at an earlier time possessed a much greater 

 virulence and because its pathogenicity had been reduced artificially. 

 (See p. 310.) It was therefore interesting to see if the virulence could 

 be reclaimed. 



Experiments of this nature were first undertaken with the idea of 

 ascertaining how long the attenuated organisms would remain alive in 

 the tissues after subcutaneous inoculation. The technique of the experi- 

 ments was as follows : 



The abdomen of a monkey was first shaved and a suspension of the organism 

 inoculated subcutaneously. The skin was then carefully massaged until the 

 fluid was apparently completely absorbed. After varying periods of time, the skin 

 of the abdomen was scrubbed several times with ether and alcohol and a small 

 incision made with a sterile knife through the dermis. Cultures were then taken 

 from the drops of blood or from the pus which escaped from the incised wound. 



Usually, when the injection was made beneath the skin of the ab- 

 domen, a few hours later about the point of inoculation an oadematous 

 swelling appeared which did not entirely disappear for forty-eight 

 hours. In an earlier series of experiments made with a slightly more 

 attenuated pest culture, it was found that the organisms were still very 

 numerous in the tissues six to eight hours after the inoculation, after 

 which time they gradually disappeared so that cultures made twenty-four 

 hours subsequent to the injection generally, though not always, remained 

 sterile. It seemed probable that the more resistant organisms were those 

 which remained alive in the tissues for the longest time and that there 

 was a true "survival of the fittest," An attempt was therefore made 

 to ascertain if it would be possible to increase the pathogenicity of a 

 slightly more virulent culture by this same procedure. As soon as the 

 cultures made on agar from the incision in the animal developed, they 

 were inoculated subcutaneously into a second monkey and so on through- 

 out the series. The strain used for this series of experiments ("Avirulent 

 Manila") was so attenuated at the time of their commencement that 

 one 24-hour agar slant culture usually, but not invariably, caused death 

 from subacute plague infection in a guinea pig of about 250 grams 

 weight. This culture was passed through twenty-five monkeys according 

 to the method above described. At the end of this time its virulence 



