422 



SPORE CYCLE OF BACILLUS SUBTILIS 



We have found no references in the literature to the particular prob- 

 lem involved, but it may be interesting to present in tabular form the 

 limiting reactions of the medium for some bacteria. 



Table I indicates that these organisms survive in a certain limited 

 range of hydrogen ion concentrations, and that slight changes in the 

 concentration of the hydrogen ion influence the organism to a great 

 extent. 



Method of Procedure. 



Preparation of Media. — The media used in this investigation had 

 the same composition and were prepared in the same way as those de- 

 scribed in a previous article.- The pH was determined electrometri- 

 cally and was found to be as shown in Table II. 



TABLE II. 



Preparation of Media of Different pH Values. 



Throughout the following account, we have used the approximate 

 figures in referring to the various pH values; i.e., desired pH is used 

 to denote the hydrogen ion concentration of the broth. 



Method Used in Obtaining Free Spores. — The spores of the organ- 

 ism were obtained as follows: A portion of a young agar culture of 

 Bacillus subtilis was emulsified in sterile 0.85 per cent salt solution. 

 The emulsion was then transferred to a Roux flask containing stand- 

 ard agar, and the organisms were distributed over the surface of the 

 medium. The flask was incubated at 30°C. for 15 days and was then 

 kept at 25°C. for 3 weeks. At the time of the experiment the culture 

 contained practically nothing but free spores. 



Preparation of Moist Chambers. — Two moist chartibers were made 

 for each pH value for each temperature. The chambers were pre- 

 pared in the usual way, using paraffin of high melting point to seal 

 the ring to the slide. By means of a platinum loop 2 mm. in diameter, 



^ Itano, A., Massachusetts Agric. Exp. Sta., Butt. 167, 1916, 164. 



