DIPHTHERIA BACILLUS. 345 
temperatures only alter it very gradually. Kept from 
light and air and in cold storage it keeps almost un- 
altered for years. 
The Production of Toxin in Culture Media. The arti- 
ficial production of toxin in cultures of the diphtheria 
bacillus has been found to depend upon definite condi- 
tions, which are of practical importance in obtaining 
toxin for the inoculation of horses, and also of theoret- 
ical interest in explaining why cases of apparently equal 
local severity have such different degrees of toxic ab- 
sorption. The researches of Roux and Yersin laid the 
foundation of our knowledge. Their investigations have 
been continued by Theobald Smith, Spronck, ourselves, 
and others. After an extensive series of investigations 
we (Park and Williams) came to the following conclu- 
sions: Toxin is produced by fully virulent diphtheria 
bacilli at all times during their life when the conditions 
are favorable. Under less favorable conditions some 
bacilli are able to produce toxin while others are not; 
or it may be that some conditions favor some bacilli 
while they are deleterious to others. Diphtheria ba- 
cilli may find conditions suitable for luxuriant growth, 
but unsuitable for the production of toxin. The re- 
quisite conditions for a good development of toxin, as 
judged by the behavior of a number of cultures, are a 
temperature from about 35° to 87.5° C., a suitable cul- 
ture medium, such asa 2 per cent. peptone nutrient 
bouillon of an alkalinity which should be about 8 c.e. 
of normal soda solution per litre above the neutral 
point to litmus, and prepared from a suitable peptone 
and meat. The culture fluid should be in compara- 
tively thin layers and in large-necked Erlenmeyer 
flasks, so as to allow of a free access of air. The 
