BACILLACE^: THE SPOROGENIC ANAEROBES 291 



spore-formers may be eliminated by successive transfers in 

 animals. 



The spores of CI. tetani resist the temperature of boiling water 

 for 5 to 30 minutes. Biological products to be introduced into 

 the human body need to be sterilized in the autoclave or else 

 carefully examined by anaerobic culture methods to insure their 

 freedom from tetanus spores. The danger of infection from this 

 source has been emphasized by Smith. ^ 



The colony in glucose gelatin or glucose agar consists of a 

 compact center with slender, radiating, straight or irregularly 

 curved threads about the periphery. Liquefaction of gelatin 

 becomes evident in stab-culture after about two weeks at 20° C. 

 Milk is sometimes but not always coagulated and the casein is 

 eventually digested. 



The cultures of the tetanus bacillus are extremely poisonous, 

 especially so when they are developed under very strict anaerobic 

 conditions. A nerve poison, tetanospasmin, and a hemolytic 

 poison, tetanolysin, are present. The former is the more impor- 

 tant constituent of the tetanus toxin. Neutral or slightly alka- 

 line plain nutrient broth, incubated in an atmosphere of hydrogen 

 for ten days after inoculation gives the most powerful toxin. 

 The bacteria-free fluid from such a culture has been found to kill 

 a mouse of lo-grams weight in a dose of 0.000 005 c.c. The toxin 

 is unstable in solution but very stable when dried. Dry material 

 of which 0.000 000 I gram is the fatal dose for a mouse is readily 

 obtained. The watery solution loses it toxicity when heated to 

 60° C. for 20 minutes, but when dry the toxin withstands heat- 

 ing at 120° C. for an hour. 



Tetanus presents essentially the same picture in inoculated 

 animals as in the natural disease, which is indeed, as a general 

 rule, merely an accidental inoculation. The presence of insoluble 

 material and of other bacteria mixed with them in a wound favors 

 the development of tetanus bacilli. The tetanus baciUi always 

 remain localized near the point of inoculation and may be hard 

 1 Journ. A. M. A., Mar. 21, igo8, Vol. L., pp. 929-934. 



