RESEARCHES ON CERTAIN PROBLEMS OF PLAGUE IMMUNITY 7 



virulence and habit of the culture. To obtain powerful toxins, the 

 bacillus must have been recently isolated from the animal body. After 

 a period of cultivation on artificial media, and especially if high tempera- 

 tures have been employed, it loses its toxigenic property, which may, 

 however, be recovered in part at least by fresh passage through the 

 animal body. 



Kolle (1903), in proceeding to study the production and nature of 

 plague toxin, selected for his experiments such cultures as, from their 

 behaviour in the animal body, might be expected to prove toxigenic on 

 artificial media. He observed that, after a large number of passages 

 through the rat, plague cultures showed a tendency to lose their property 

 of multiplication in the blood, tissues, and organs of the animal body. 

 The death of the animals resulted almost entirely from the toxins 

 produced at the site of inoculation and in the neighbouring buboes. In 

 other words, the cultures, before their frequent passage, tended to produce 

 a plague septicaemia, after their passage a plague toxaemia. The 

 toxaemic symptoms came on at an early period, when the multiplication 

 of the bacilli in the rats was occurring solely at the site of inoculation. 

 Kolle employed an alkaline bouillon. 



Whatever the nature of the flask employed for the cultivation of the 

 bacillus provision was made for an abundant supply of oxygen by having 

 only a moderately thin layer of medium at the bottom of the vessel. 

 Occasionally a little blood from the rat or rabbit was added to the 

 bouillon, in which case the culture was observed to be very abundant. 

 The cultures were grown some at 37° C, others at 35° C, others at 22° 

 C, and still others in the ice-safe at from 6° - 10° C. 



A number of methods were employed to obtain the toxin, apart 

 from the bacilli. By the addition to the cultures of phenol and toluol, 

 and subsequent centrifugalisation, a sterile supernatant fluid was obtained 

 which was tested for toxicity. The Chamberland filter was also 

 employed, the filtrate being tested for sterility, as in certain cases the 

 plague bacilli have been found to pass the filter. 



Kolle employed for his experiments rats of 180-200 grammes weight. 

 In his experience the guinea-pig and rabbit are too resistant, while the 

 mouse gives irregular results or proves too susceptible. Intraperitoneal 

 injection was in every case employed. 



In young cultures^ of from 3-4 days, no toxin could be demon- 



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