532 Messrs. Twort and Ingram. Isolating and [Nov. 7, 



A fifth batch of vaccine was made in a manner similar to vaccine No. 4, 

 except that the three cultures of Johne's bacillus were suspended in 6 c.c. of 

 0'8-per-cent.- sodium chloride in place of vaccine No. 3. 



As controls to the above vaccines we used diagnostic tuberculin prepared 

 t by the Pasteur Institute, diagnostic avian tuberculin prepared at the Koyal 

 Veterinary College, and a special timothy-grass bacillus vaccine prepared by 

 ourselves. This last was made in the same manner as ordinary diagnostic 

 tuberculin. The bacillus was grown for about three weeks in a glycerine 

 broth medium, which was then steamed and filtered through a Doulton 

 white porcelain filter. The sterile filtrate was placed in small sterile flasks 

 without previous concentration. The results of the tests conducted with 

 the above vaccines will be described under the head of experiments on 

 bovines. 



At the time that test-tube experiments were being carried out we 

 performed a number of inoculation experiments on animals. In the first 

 series we tested small laboratory animals such as mice, rats, guinea-pigs, 

 rabbits, hens, and pigeons. Several of each were inoculated either sub- 

 cutaneously or intraperitoneally, and some, such as rabbits and pigeons, 

 intravenously, while others were fed with food soaked in an emulsion of the 

 bacilli. Vigorous growing cultures of Johne's bacillus were made up into 

 thick emulsions with sterile normal saline ; the mice and rats were usually 

 inoculated with \ c.c, and the guinea-pigs, rabbits, hens, and pigeons with 

 \ to 1 c.c. of the emulsion. 



With the mice, rats, guinea-pigs, hens, and pigeons the results were 

 entirely negative. No lesions were found in the animals post mortem, even 

 when kept for nine months before killing. 



One mouse inoculated into the peritoneal cavity and killed after 14 days 

 showed a few acid-fast bacilli in one of the mesenteric glands, but they 

 could not be recovered in cultures, and it is probable that they were dead 

 and had been taken up by the glands. Sections of the mesenteric glands 

 showed no histological lesions. 



The only rabbit which showed any evidence of a lesion, probably produced 

 as a result of the inoculation (into the peritoneal cavity with 1 c.c. of a 

 thick emulsion of Johne's bacillus), was killed after four months. On 

 post-mortem examination a small whitish thickening was found in the wall 

 of the csecum, and films made from this area showed a few degenerated 

 acid-fast bacilli. Sections showed destruction and degeneration of the 

 tissues comprising the wall of the gut, with granular structureless material 

 containing a few degenerated acid-fast bacilli in the centre of the area. 

 Cultures made from this area on to various media, including the special 



