270 TUBERCULOSIS 



Staining Reactions. — The tubercle bacillus takes up the 

 ordinary stains very slowly and faintly, and for successful stain- 

 ing one of the most powerful solutions ought to be employed, e.g., 

 gentian-violet or fuchsin, along with aniline-oil water or solution 

 of carbolic acid. Further, such staining solutions require to be 

 applied for a long time, or the staining must be accelerated by 

 heat,, the solution being warmed till steam arises and the 

 specimen allowed to remain in the hot stain for two or three 

 minutes. One of the best and most convenient methods is the 

 Ziehl-Neelsen method (see p. 105). The bacilli present this 

 further peculiarity, however, that after staining has taken place 

 they resist decolorising by solutions which readily remove the 

 colour from the tissues and from other organisms which may be 

 present. Such decolorising agents are sulphuric or nitric acid 

 in 20 per cent, solution. Preparations can thus be obtained in 

 which the tubercle bacilli alone are coloured by the stain first 

 used, and the tissues can then be coloured by a contrast stain. 

 Within recent years certain other bacilli have been discovered 

 which present the same staining reactions as tubercle bacilli ; 

 they are therefore called " acid-fast " (vide infra). The spores 

 of many bacilli become decolorised more readily than tubercle 

 bacilli, though some retain the colour with equal tenacity. 



Muck's Method. — Much maintains that in addition to the ordinary 

 acid-fast bacillus, the organism exists in the form of a bacillus which is 

 not acid-fast and also in the form of free granules. These two forms are 

 demonstrable by certain modifications of Gram's method, of which the 

 following is specially suitable : — 



Methyl-violet B.N., 10 c.c. of a saturated alcoholic solution in 100 c.c. of 

 a 2 per cent, watery solution of carbolic acid ; stain by boiiing over the 

 flame for a few minutes or at 37° C. for 24-48 hours, then treat with Gram's 

 iodine for 1-5 minutes, 5 per cent, nitric acid for one minute, 3 per cent, 

 hydrochloric acid for 10 seconds, and complete the decolorisation with a 

 mixture of acetone and alcohol in equal parts. 



There seems to be no doubt that in certain conditions more tubercle 

 bacilli can be demonstrated in the tissues by Much's method than by the 

 Ziehl-Neelsen method. 



Chemical Composition.— Bulloch and Macleod, by treating tubercle 

 bacilli with hot alcohol and ether, extracted a wax whiijh gave the 

 characteristic staining reactions of the bacilli themselves. The remains 

 of the bacilli, further, when extracted with caustic potash, yielded a body 

 which was probably a chitin, and which was acid-fast when stained for 

 twenty-four hours with carbol-fuchsin. Benians considers that a waxy 

 material in some way encloses the protoplasm and fatty constituent, and 

 confers on the organism the property of resisting the penetration of acid 

 and alcohol. 



Cultivation. — The medium first used by Koch was inspissated 

 blood serum (vide p. 40). If inoculations are made on this 



