256 THE DIPHTHERIA BACILLUS 



organism in broth. On the other hand, when the organism is grown at 39 C. 

 on glycerin-agar or on broth with a current of air passing over it, it loses its 

 virulence rapidly, so that on inoculation into a guinea-pig nothing more than 

 a local oedema results (Roux and Yersin). 



The same fact may be observed by drying a false membrane from a case 

 of diphtheria and exposing it to the air : the organism remains alive for a 

 long time, but if fragments of the membrane be sown from day to day it 

 will be found that the number of non-virulent colonies increases. The 

 bacilli thus artificially attenuated have all the characteristics of the pseudo- 

 diphtheria Hofmann's bacillus (see p. 273). [It would seem that the 

 explanation of this result is to be found in the supposition that both the 

 diphtheria bacillus and Hofmann's bacillus were present in the original 

 membrane and that the former died out. All attempts to convert a diph- 

 theria bacillus into an Hofmann and vice versa have invariably failed (p. 274). ] 



(b) Restoration of virulence. -It is impossible to restore the virulence of 

 an organism which has become so attenuated as to have entirely lost its viru- 

 lence for the guinea-pig (Roux and Yersin). 



On the other hand, Roux and Yersin succeeded in restoring the virulence 

 of an organism which produced nothing more than a slight oedema in the 

 guinea-pig ; they inoculated the bacillus, and with it a virulent culture of a 

 streptococcus, into a guinea-pig : the animal succumbed with symptoms of 

 diphtheria and the bacillus recovered from the fluid of the local oedema was 

 found to have very distinctly increased in virulence. According to Blasi 

 and Russo-Travalli association with the colon bacillus has a similar effect 

 in restoring the virulence of the diphtheria bacillus. Trumpp, by inoculating 

 a mixture of small doses of diphtheria toxin and an almost non-virulent 

 bacillus, was also able to restore the virulence of the organism. 



(c) Exaltation of virulence. Roux and Yersin failed to raise the virulence 

 of the diphtheria bacillus by passage through guinea-pigs or rabbits. Bar- 

 dach after passing the bacillus through twenty-five dogs noted a distinct 

 increase in virulence for the dog but only a slight increase in virulence for 

 the guinea-pig. 



Martin grew the bacillus in collodion sacs in the peritoneal cavities of a 

 series of rabbits and succeeded in raising the virulence for that animal, but 

 found that the virulence for the guinea-pig was unaltered. 



[Thus the virulence of the diphtheria bacillus, while easily lowered, is 

 difficult to increase by laboratory methods.] 



[2. Bio-chemical reactions.] 



[(a) Action on carbohydrates. " All strains of the diphtheria bacillus 

 produce acid from glucose, galactose, laevulose, and maltose. Most form 

 acid out of dextrine and glycerine. On lactose the action is very variable, 

 and only a few strains act on saccharose. All tests on mannite yielded 

 negative results " (Graham-Smith). No gas is formed in any of the media. 



[In testing the action of diphtheria bacilli upon carbohydrates the most 

 suitable medium to which to add the carbohydrate is Hiss's serum-water 

 medium (Chap. XL.). To this solution, 1 per cent, of the carbohydrate is 

 added. 



[If sugar-free broth be used the results may not be so uniform, because as 

 has already been pointed out some strains of the diphtheria bacillus do not 

 grow readily on such broth when first isolated from the body. 



[(b) Indol. The diphtheria bacillus does not appear to form indol (Theobald 

 Smith and others), but some observers are said to have obtained an indol 

 reaction after prolonged cultivation.] 





