466 CHOLERA 



to the fact that both indol and a nitrite are formed by the 

 spirillum in the medium, and hence, in applying the test for 

 indol, the addition of a nitrite is not necessary. It is essential 

 that the sulphuric acid should be pure, for if traces of nitrites 

 are present the reaction may be given by an organism which 

 has not the power of forming nitrites. 



Hcemolytic Test.— This method, introduced by Kraus, is performed by 

 means of agar plates (p. 45), a small quantity of sterile defibrinated blood 

 being added to the agar and thoroughly diffused; 1 if any organism has 

 hemolytic properties, a clear zone or areola forms around each colony by 

 the diffusion of haemoglobin. As a rule the cholera organism does not 

 produce hemolysis, but the result after twenty-four hours should be 

 taken, as later a clear zone may appear round a cholera colony (Greig). 

 It has, however, been found by several observers that the hemolytic test 

 is best carried out with a fluid culture. Greig, for example, adds varying 

 amounts, from 1 c.c. downwards, of a three days' culture in alkaline broth 

 to 1 c.c. of a 5 per cent, suspension of goats' corpuscles, the whole being 

 made up to 2 c.c, and thoroughly mixed. The tubes are placed in the 

 incubator for two hours at 37° C, and then in the ice-chest overnight, 

 the results being read next day. He found after testing more than 

 300 strains of true cholera spirilla that none of them produced 

 hemolysis, whereas this results with organisms of the El Tor group 

 (vide infra). 



Powers of Resistance. — In their resistance against heat, 

 cholera spirilla correspond with most spore-free organisms, and 

 are killed in an hour by a temperature of 55° C, and much 

 more rapidly at higher temperatures. They have comparatively 

 high powers of resistance against great cold, and have been 

 found alive after being exposed for several hours to the tempera- 

 ture of - 10° C. They are, however, killed by being kept in ice 

 for a few days. Against the ordinary antiseptics they have 

 comparatively low powers of resistance, and Pfuhl found that 

 the addition of lime, in the proportion of 1 per cent., to water 

 containing the cholera organisms was sufficient to kill them in 

 the course of an hour. 



As regards the powers of resistance in ordinary conditions, 

 the following facts may be stated ! In cholera stools kept at the 

 ordinary room temperature, the cholera organisms are rapidly 

 outgrown by putrefactive bacteria, but in exceptional cases they 

 have been found alive even after two or three months. In most 

 experiments, however, attempts to cultivate them even after a 

 much shorter time have failed. The general conclusion may be 

 drawn from the work of various observers, that the spirilla do 

 not multiply freely in ordinary sewage water, although they may 

 remain alive for a considerable period of time. On moist linen, 

 as Koch showed, they can nourish very rapidly. Though we 



