BACILLUS TETANI 243 



Cultivation. At 20 to 24 C. growth occurs slowly and spores 

 are produced in six to ten days ; at 37 C. development is much 

 more rapid and spore formation begins within twenty-four to thirty 

 hours. Under ordinary conditions the tetanus bacillus is a strict 

 anaerobe. If, however, aerobic bacilli are growing with it in culture 

 medium it will develop even when air is admitted. Presumably 

 the air is used up by the associated aerobes. Growth occurs 

 abundantly on gelatin or agar containing 1 to 2 per cent glucose ; 

 the colonies have a fleecy thread-like margin radiating from a 

 heavier central portion. In gelatin stabs growth appears along 

 the needle track and from it outgrowths extend, giving the appear- 

 ance of an inverted fir tree ; liquefaction takes place slowly, gen- 

 erally with the production of a gas of characteristic and disagree- 

 able odor. 



As the tetanus bacillus is almost always associated with other 

 organisms the most successful method of obtaining a pure culture 

 is by taking advantage of the resistance of its spores to heat. 

 The material containing the organism may be inoculated into a 

 tube of glucose broth and incubated at 37 C. for two days. By 

 that time sporulation will have occurred and the culture may then 

 be heated to 80 C. for three quarters of an hour in order to destroy 

 the associated vegetative forms. From the heated culture agar 

 anaerobic plates are made, and if the tetanus bacilli are the only 

 spore-bearing anaerobic organisms present a pure culture may be 

 readily obtained. 



Resistance. In the vegetative form the tetanus bacillus is 

 destroyed by the same agencies that kill spore-free bacteria. On 

 the other hand, few forms of life are more resistant than tetanus 

 spores; in a dried condition they may retain their vitality for 

 years. In 5 per cent carbolic acid they are killed in ten hours. 

 The addition of 0.5 per cent hydrochloric acid hastens the germi- 

 cidal effect and destroys them in two hours. Bichloride of mercury 

 1 to 1000 kills them in three hours and in thirty minutes if 0.5 

 per cent hydrochloric acid be added to the solution. The spores 

 are completely destroyed when exposed to dry heat at 160 C. for 

 one hour or to steam at 120 C. for twenty minutes. 



