BACILLUS DIPHTHERIA 163 



described a double stain (see Technique, p. 23) which can be used as a 

 means of differential diagnosis. In sections the bacilli can be stained by 

 Loffler's method, also by the Gram and Cladius methods. 



Biological Characters. Aerobic and non-liquefying, growing most 

 freely in the presence of oxygen, but is also a facultative anaerobe. The 

 growth takes place only between 20 and 42 C. on all slightly alkaline 

 media ; the optimum temperature is about 35 C. 



On Gelatine Plates. Small, round, white colonies develop, which under 

 a low power appear yellowish-brown in colour, and granular, with 

 irregular borders, rarely reaching a diameter of over 1-5 mm.; the 

 development is very slow, nothing very characteristic being observed in 

 less than seventy-two hours. 



In Gelatine Stab Cultures. The same above mentioned ; small, white, 

 round colonies, not exceeding the above size, develop along the inocula- 

 tion track. At 24 C. surface growth and indications of a nail-shaped 

 growth appear. The gelatine is not liquefied. 



On Agar. Best on glycerine agar plates ; in twenty-four to forty-eight 

 hours small, greyish-white, shining colonies develop, which macroscopic- 

 ally often exhibit a stratified appearance ; and under a low power 

 appear granular, with irregular borders. 



On Agar Stroke Cultures. In twenty-four hours small, transparent, 

 slightly elevated colonies appear ; the further growth is very scanty, and 

 does not extend far from the inoculation track. 



In Agar Stab Cultures. Colonies develop along the inoculation track 

 with a minimal amount of surface growth. 



In a mixture of glycerine agar and human blood serum the growth is 

 much more pronounced and extensive (see Photograph of Culture 

 growing in this medium, Fig. 59). 



On Lqffler's Serum (for preparation of same see Technique, p. 56). In 

 twenty-four hours somewhat large, whitish opaque colonies of firm 

 consistence develop, which only increase slightly in size during the 

 next few days. This medium is the best for diphtheria bacilli, and is 

 always used for differential diagnosis. 



Bouillon. In this medium the bacillus grows in fine clumps, which 

 fall to the bottom of the tube, or are deposited on the sides without 

 causing any clouding of the medium. The bouillon may appear diffusely 

 clouded to the naked eye, but when examined microscopically in a 

 hanging-drop the clumpy arrangement is easily observed. In bouillon 

 kept at 35 C. for some time a whitish film often forms over part of the 

 surface. The reaction of the bouillon is subject to changes frequently 

 at first it is acid, and subsequently again alkaline. These changes can be 

 observed in the medium when a little rosolic acid is added. These 

 reactions are attributed to the primary fermentation of muscle sugar 

 often present in the bouillon. 



