50 
Tetanus spores may live a long time in vaccine virus. We have 
found them alive and virulent on diy points after two hundred and 
ninety-five days and in giycerinated virus sealed in capillary tubes 
three hundred and fifty-five days. 
Tetanus ma}^ become a contaminating element of vaccine before it 
leaves the heifer. During the period of three to five days which 
elapses between the vaccination of the heifer and the removal of the 
virus there is opportunity for tetanus to find a lodgment in the erup- 
tion on the heifer’s body surface, provided tetanus is present in the 
stall or stable surroundings of the animal. 
If tetanus reaches the heifer’s vaccinated area it may contaminate 
both the ‘"dry points,” which are made directly from the “lymph,” 
and also the vaccine pulp, which the manufacturer subjects for a longer 
or shorter time to the germicidal action of glycerin before he markets 
it as “giycerinated virus.” 
Tetanus added to gl}merinated vaccine virus does not germinate 
when kept hermetically sealed and under anaerobic conditions in small 
capillary tubes. It gradually loses its virulence both in the tubes and 
on the ivory points. Although the virulence is lost, the tetanus spores 
are not necessaril}" dead; for, while they will not produce sj^mptoms 
when inoculated directly into mice, they ma}’ be revived into active 
virulent cultures b}" growing* in fresh bouillon under favorable con- 
ditions. That is to say, the vegetabilit}^ of the spore remains active 
long after it has lost its power to produce the disease when inoculated 
directly into mice. 
Therefore, in looking for tetanus in vaccine virus it is best to make 
cultures first and study the growth for end spore-bearing rods and 
then test the elfects of the culture in animals. 
The spores of tetanus lose their virulence and die much more quickly 
in the vaccine l}^mph on diy points than in the giycerinated tubes. 
The vitality of tetanus in glj^cerinated virus depends largely upon 
the number of spores contaminating the virus. Large quantities, 
namel^y forty times the minimal lethal dose (lOxMLD), remain alive 
and active over a }*ear. Smaller amounts may disappear in four to 
seven months. Yerj small amounts (1 X MLD) have in one of our 
experiments lost virulence in one month and failed to grow in bouillon 
in two months. 
On the other hand, these verv small amounts of tetanus mav remain 
active for months in giycerinated virus in capillar}^ tubes. In one 
instance as small a quantity as 0.000055 c. c., which was about two and 
a half times the minimal fatal dose, remained active seven months. 
In vaccinal “lymph” on dry points the spores may begin to lose 
their virulence in two weeks and be dead in two months. Usually 
they live longer. We found them alive and virulent in points that 
had been contaminated with 50 X MLD about ten months (two hundred 
and ninety-five days). 
O 
