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of Kolle and an old Manila culture which had been grown continuously 
upon artificial media for three years and whose virulence has been still 
further reduced by artificial means. The attenuation of this last culture 
was further brought about by growing the bacillus at a temperature of 
from 41° to 43° C. in flasks of alcoholic bouillon for three weeks at a 
time, as recommended by Otto. Cultures from these flasks were then inoc- 
ulated on agar for many generations, a fresh generation being made every 
day for several weeks, and the organisms always cultivated at the same 
high temperature. Beginning with 0.05 cubic centimeter of absolute 
alcohol and 50 cubic centimeters of bouillon, the amount was gradually 
increased in successive cultures up to 5 cubic centimeters in 50 of bouillon. 
Before making inoculations in man, the action of the attenuated culture 
was of course carefully tested in animals. 
In the present paper it is merely my desire to call attention to the 
fact that vaccination in man can with safety be performed with attenu- 
ated cultures of the living plague organism, and therefore only the human 
inoculations undertaken with one strain of this bacillus will be referred 
to..* The organism in question (J/a. V) possesses so little virulence 
that in a series of twelve guinea pigs and thirty monkeys inoculated with 
from one to two entire agar slant cultures, not one succumbed from the 
effects of the inoculation.’ It was with this culture that the first 
experiments were performed in human beings. Since I believed that the 
guinea pig is an equally if not even a more susceptible organism than 
man to the pathogenic action of the plague bacillus, it was presumed that 
if this animal could invariably withstand the action of such large amounts 
as two whole agar slant cultures of the organism, much smaller quanti- 
ties could be inoculated into human beings with safety, and indeed, before 
performing the experiments on man, | felt thoroughly convinced of this 
fact; nevertheless, the human inoculations were performed as carefully 
and with as much deliberation as possible, 
The first injections were carried on upon prisoners under sentence of 
death; in the first case one-hundredth oese of the attenuated culture 
was inoculated subcutaneously without any noticeable. effect. After ten 
days, ten other individuals were inoculated with the same dose, in order 
to demonstrate that no special natural immunity against the plague 
organism had been existent in the first instance. In this manner the 
amount of living organisms given was gradually increased, a single person 
being first inoculated with the larger dose and then, after it had been 
observed that no unfavorable effects occurred, from five to ten other 
“ The results of these experiments were communicated in a paper read before the 
Manila Medical Society at its meeting on November 6, 1905. 
* But one of the animals of the series perished. Monkey No. 1299 died about 
twelve hours after inoculation with the avirulent pest organism, of a staphylo- 
coccus and streptococcus pyemia which had existed prior to the inoculation. A 
large suppurating wound existed over the abdomen. 
