182 
to the present time no further experiments of importance with the living 
organism, either in a virulent or an attenuated condition, have been made 
on human beings, but a number of this nature have been performed on 
animals. 
The German Plague Commission ® (Gaffky, Pfeiffer, and Dieudonne) 
called attention to the fact that an ape (Macacus radiatus) which had 
been inoculated with the living pest organism and which after many days’ 
illness had recovered, was found to be fully immune about five weeks 
subsequent to the first inoculation, since, at that time, it resisted the 
injection of an entire oese of a virulent pest culture. For the purpose of 
attenuating the pest bacillus, agar cultures suspended in bouillon were 
exposed to a temperature of 51° C. during different periods of time. 
First experiment.—The culture was heated for two hours at 51° C. and after- 
wards proved to be sterile. A monkey, inoculated with one-fifth oese of this 
culture, acquired no immunity, succumbing to an injection of one oese of the 
virulent pest organism made twelve days later. 
Second experiment.—The culture was heated for one hour at the same temper- 
ature, but in this instance all of the bacilli were not killed. A monkey inoculated 
with one-fifth oese of this culture also showed no subsequent immunity. 
Third experiment.—The organism was heated for only one-half hour at 51° C. 
In this instance many of the bacilli were also not killed, and the monkey inoculated 
with one-fifth oese of this culture died of pest four days after this primary 
injection. 
In the foregoing observations, as was demonstrated by further experi- 
ments, the pest bacillus was not attenuated, but was either entirely killed 
or had retained its full virulence. In the second experiment, while the 
living organisms were still present in the vaccine, their number was 
obviously too small to produce the desired immunity or even to give rise 
to any decided reaction in the animal. This was conclusively shown to 
be the case by the following observation, in which a two-day agar culture 
was suspended in 5 cubic centimeters of bouillon and then further di- 
luted with saline solution, so that ape A received 1 cubic centimeter in a 
dilution of: 1 to 100,000; ape B, 1 cubic centimeter in a dilution of 1 to 
10,000, and ape C, 1 cubic centimeter in one of 1 to 1,000, subcutaneously. 
No apparent reaction occurred from these injections and only ape © 
showed any traces of immunity, this animal remaining alive for nine 
days following the injection of 1 oese of the virulent pest bacillus 
(although it succumbed after this period), while the other two animals 
died on the third day after such an inoculation. 
In another series of experiments, the commission attempted to obtain 
an attenuation of the organism by its exposure to the action of carbolic 
acid. However, no loss of virulence resulted by this method, the animals 
injected with the cultures which had been so treated all succumbing to 
pest infection. On account of the difficulties encountered in the attenua- 
tion of the plague bacillus or in obtaining cultures already attenuated, 
® Loc. cit., 303. 
