398 A MANUAL OF BACTERIOLOGY 



in the sulphuric or nitric acid solution, washed, dried, and 

 mounted. They can also be stained by Gram's method, which 

 usually brings out the beaded appearance very markedly, or by 

 any of the other methods mentioned under Sputum. Differentiation 

 from the leprosy bacillus will be found at pp. 405, 407, and from 

 the smegma bacillus and other acid-fast organisms at p. 408. 



3. Urine. The tubercle bacillus is often very difficult to 

 demonstrate in urine. The urine must be allowed to stand in a 

 conical glass for twenty-four hours or centrifuged, and film 

 specimens are prepared with the sediment and treated by one of 

 the methods for sputum given above. Several specimens should 

 be made and must be very carefully examined. The sediment 

 may also be treated by the antiformin method. It is important 

 to exclude the smegma bacillus, and the urine is preferably drawn 

 off by a. catheter. Staining may be carried out by HouselFs 

 method, by which the smegma bacillus is decolorised, viz. after 

 staining in warm carbol-fuchsin the specimen is washed and 

 dried. It is then immersed in acid alcohol (alcohol + 3 per cent, 

 hydrochloric) for ten minutes, washed in water, counter-stained 

 for a few seconds in a saturated alcoholic solution of methylene 

 blue, washed, dried, and mounted (see also p. 408). An electro- 

 lytic method for the concentration of the tubercle bacilli has been 

 devised by Kuss. 1 



If a diagnosis is of importance inoculation should be resorted 

 to. Two guinea-pigs are inoculated subcutaneously in the thigh 

 or abdomen with 0-5 to 1 c.c. of the deposit from the sedimented 

 or centrifuged urine, or one may be inoculated subcutaneously, 

 the other intra-peritoneally. If tubercle bacilli are present the 

 animals may show signs of tuberculosis as early as two to three 

 weeks after inoculation. Sometimes, of course, the animals may 

 die from some intercurrent infection before the tuberculous 

 infection has had time to develop. Delepine recommended the 

 inoculations to be made on the inner aspect of the leg about the 

 level of the knee. The order of infection after inoculation is as 

 follows : the popliteal, superficial and deep inguinal, and sub- 

 iumbar glands, the retrohepatic, mediastinal and bronchial, 

 deep cervical, and subscapular glands, the spleen, liver, and 

 lungs. The inoculated animals arc killed in two to three weeks, 



1 Proc. Roy. Soc. Lond., B. 1909. 



