DESCRIPTION OF JOHNE'S BACILLUS 65 



method of staining, Johne's bacillus is coloured a 

 bright magenta and the rest of the material blue. 



When staining films made from pure cultures, 

 counter-staining with methylene blue is not necessary. 



Johne's bacillus may also be stained by Gram's 

 method, and is, therefore, Gram- positive. In this 

 method films of bacilli are treated with hot aniline- 

 gentian violet for three or four minutes ; the stain is 

 then poured off, and Lugol's iodine solution floated 

 over the surface of the film, and allowed to remain for 

 about a minute ; the films are then treated with abso- 

 lute alcohol until no more colour can be extracted. If 

 examined under the microscope, the bacilli are found 

 to be stained a blackish-violet colour. 



From what has been said it is clear that in morph- 

 ology and staining reactions Johne's bacillus agrees 

 with the various varieties of tubercle bacilli. It shows 

 no character that cannot be met with under certain 

 conditions in the tubercle bacilli. 



Cultivation of the Bacillus. — Johne and Frothingham, 

 after making their original discovery of the presence 

 of acid-fast bacilli in the intestinal mucous membrane 

 obtained from cases of pseudo-tuberculous enteritis, 

 attempted to cultivate the bacilli outside the animal 

 body. As a medium they used glycerine-agar, expect- 

 ing to obtain cultures of some variety of the tubercle 

 bacillus. Their cultures, however, remained sterile, 

 and on microscopic examination they found no evi- 

 dence of multiplication of the bacilli. In 1904, Stuur- 

 man obtained, from a case of Johne's disease at Leyden, 

 a pure culture of an acid-fast bacillus, and again, in 

 1908, from the inguinal gland of a guinea-pig inoculated 

 with a small piece of the intestine of an animal affected 

 with Johne's disease he obtained another culture, 

 which showed characters similar to the first. These 



