470 CHOLERA 



sac and cause toxic symptoms. By greatly increasing the 

 virulence of the organism, then growing it in bouillon and 

 filtering the cultures on the third and fourth day, they obtained 

 a fluid which was highly toxic to guinea-pigs (the fatal dose 

 usually being ^ c.c. per 100 grm. weight). The symptoms 

 closely resemble those obtained by Pfeiffer. They found that 

 the toxicity of the nitrate was not altered by boiling, appar- 

 ently this toxic substance is different from Pfeiffer's endotoxin. 

 In a recent research Huntemuller has obtained from various 

 strains an acutely acting extracellular toxin which is very labile 

 and which he believes to be identical with the haemolysin. He 

 has obtained an antitoxin to this toxin. The diversity in the 

 results obtained by various workers seems only explicable on the 

 view that different strains vary greatly as regards production of 

 extracellular toxin. It may be stated that, as a rule, the greater 

 part of the toxic substance is closely bound up with the bacterial 

 protoplasm, and is only set free on its disintegration. 



Immunity. As this subject is discussed later, only a few 

 facts will be here stated, chiefly for the purpose of making clear 

 what follows with regard to the means of distinguishing the 

 cholera spirillum from other organisms. The guinea-pig or any 

 other animal may be easily immunised against the cholera 

 organism by repeated injections (conveniently made into the 

 peritoneum) of non-fatal doses of dead spirilla ; later the living 

 organisms may be used. In this way a high degree of immunity 

 against the organism is developed ; and further, the blood serum 

 of an animal thus immunised (anti-cholera serum) has markedly 

 protective power when injected, even in a small quantity, into a 

 guinea-pig along with five or ten times the fatal dose of the 

 living organism. Under these circumstances the spirilla undergo 

 a granular transformation and, ultimately, solution ; this pheno- 

 menon is generally known as Pfeiffer's reaction, and was applied 

 by him to distinguish the cholera spirillum from organisms 

 resembling it. The following are the details : 



Pfeiffer's Reaction. A loopful (2 mgrm.) of a recent agar culture of the 

 organism to be tested is added to 1 c.c. of ordinary bouillon containing 

 001 c.c. of anti-cholera serum. The mixture is then injected into the 

 peritoneal cavity of a young guinea-pig (about 200 grm. in weight), and 

 the peritoneal fluid of this animal (conveniently obtained by means of 

 capillary glass tubes inserted into the peritoneum) is examined micro- 

 scopically after a few minutes. If the spirilla injected have been cholera 

 spirilla, it will be found that they become motionless, swell up into 

 globules, and ultimately break down and disappear positive result. If 

 they are found active and motile, then the possibility of their being 

 true cholera spirilla may be excluded negative result. In the former 



