284 BACTERIOLOGY 



growth at 35 C. on an agar surface 10 cm. long) of cholera 

 bacillus having been drawn into a small pipette, the point 

 of the latter is bored through the abdominal wall of a guinea- 

 pig, at a spot previously sterilised by cauterisation after clip- 

 ping the hair short, and the contents are blown out into the 

 peritoneal cavity. Immediately after death (which takes place 

 in twenty-four hours) the abdomen is opened with the strictest 

 antiseptic precautions, and the fluid found in the iliac fossae 

 is transferred by means of a sterile pipette to a sterilised 

 test-tube, in which it is left in the incubator for 8 or 10 

 hours at 35 C., in such a position that the largest possible 

 surface of fluid is exposed to the air. The process of intra- 

 peritoneal injection, &c., is then repeated, and this is done 

 twenty to thirty times in succession, until the highest pos- 

 sible degree of virulence is attained, which may be over a 

 hundred times as great as that of the original growth. 

 Cultures of the exalted virus can then be made, but do not 

 retain their full virulence for many generations. 



The vaccine actually injected may contain the living 

 bacilli (since these perish in the tissues) , or may be ' car- 

 bolised,' so as to kill them. Carbolised vaccines are less dan- 

 gerous and liable to contamination, and may be kept in- 

 definitely in sealed tubes, but are not so powerful. The 

 vaccine is thus obtained : A standard culture is made by 

 inoculating the whole surface of an agar tube and keeping 

 it at 35 C. for 24 hours, after which an emulsion is pre- 

 pared by mixing the growth with 2 or 3 c.cm. of bouillon, 

 and then diluting up to 8 c.cm. for living vaccines ; or with 

 6 c.cm. of sterile \ per cent, carbolic acid solution for car- 

 bolised vaccines. The amount of either to be injected is 

 1 c.cm., and the injection should be made under the skin of 

 the flank. ^ 



Results of vaccination. Injection of such a vaccine 

 causes a rise of temperature accompanied by malaise and 



