288 SPECIFIC MICRO-ORGANISMS 



may occur as a granular sediment, as a diffuse cloudiness or as a 

 pellicle on the surface, depending upon the reaction and pepton 

 content of the medium and the vigor of growth of the culture. 

 The growth on the surface produces the best yield of toxin. 

 Acid is produced in dextrose broth. The organism is killed 

 when moist by heating to 60 C. for 20 minutes. It is fairly 

 resistant to drying and has been found alive in bits of dry diph- 

 theritic membrane after four months. 



Roux and Yersin in 1888 filtered broth cultures of the diph- 

 theria bacillus through porcelain filters and found the filtrate 



FIG. 120. Colonies of B. diphtheria on agar. X2oo. (After Park and Williams.) 



extremely poisonous. By injecting it into animals they were 

 able to produce the signs of local and general intoxication which 

 are observed in the natural disease. A favorable medium for 

 toxin production is a veal broth containing 2 per cent pepton 

 and having a titre of 9 c.c. 1 of normal sodium hydroxide above 

 the neutral point to litmus. It should be placed in flasks in a 

 thin layer to allow abundant air supply. Incubation for from 

 5 to 10 days gives the maximum toxicity. The filtrate from such 

 a culture may kill a 250 gram guinea-pig in a dose of 0.002 c.c. 

 Less powerful toxin is frequently obtained, so that sometimes 

 even 0.5 c.c. or more may be required to kill a guinea-pig, and 



1 Per 1000 c.c. of the medium. 



