ISOLATION OF THE ORGANISM 429 



Any of the methods described for the isolation of the typhoid bacillus 

 are quite applicable to the isolation of the paratyphoid A bacillus. 



Agglutination reactions with the patients' serum supplemented by absorp- 

 tion tests should form a part of the diagnosis in every case. 



1. The most important step is to demonstrate the presence of the bacillus 

 in the blood stream. (The isolation of the paratyphoid A bacillus is com- 

 paratively easy during the first 4 or 5 days of the pyrexia : the chances of a 

 successful blood culture are greatly diminished by the 8th day, even when 

 the usual 5 c.c. of blood is withdrawn cf. enteric fever (Grattan and 

 Wood).) 



Grattan and Wood sow the blood (5 c.c.) on sterilized ox bile and after incubating 

 for 24 hours at 37 C. plate the growth on Conradi-Drigalski's original medium, 

 which they consider better than the more recent selective media l ; they incubate 

 again and then pick off colonies which resemble those of the typhoid bacillus, which 

 is indistinguishable on the Conradi-Drigalski medium from the paratyphoid A 

 bacillus. 



The organism isolated must be fully identified. Firstly, it must be shown 

 to belong to the paratyphoid group by a careful study of its morphological, 

 staining, cultural and fermentation reactions. (The characteristics of the para- 

 typhoid A bacillus may be conveniently summarized here : it is a short 

 stumpy gram-negative bacillus which does not liquefy gelatin, forms no 

 indol, does not clot milk but turns the medium permanently acid, ferments 

 glucose, mannite and dulcite with production of acid and gas, but has no 

 action on lactose and cane sugar.) Then the bacillus must be examined, as 

 regards its agglutination reactions, with known typhoid, paratyphoid A and 

 paratyphoid B serums. 



In the case of an organism isolated from the blood these tests are sufficient 

 for identification (Grattan and Wood). 



2. To isolate the organism from the urine or from the excreta some of the 

 material should be sown in dulcite broth or dulcite peptone water. Dulcite 

 is the enrichment medium par excellence for the paratyphoid group (Boycott). 

 After incubation 2 or 3 days sometimes elapse before the paratyphoid A 

 bacillus produces gas in dulcite media, but ultimately the amount formed is 

 considerable some of the culture may be plated on Conradi-Drigalski's or 

 M'Conkey's medium and the organisms isolated tested as above. But 

 according to Grattan and Wood when a bacillus resembling the paratyphoid 

 A bacillus is isolated from the stools it is necessary to carry the identification 

 a step further (by means of absorption tests) than in the case of a similar 

 organism isolated from the blood. 



A 24-hour growth of the suspected organism on agar is emulsified in 

 about 0'2 c.c. of paratyphoid A serum which is diluted according to its 

 titre, the object being to have an excess of bacilli for the amount of agglutinin 

 present : e.g. a serum having a titre of 1-300 may be diluted ten times. 



Incubate the emulsion for 2 hours at 37 C., centrifuge and put up the 

 clear supernatant liquid in a series of dilutions in sedimentation tubes and 

 test its agglutinating action with a known paratyphoid A bacillus. 



If the organism used for absorption be the paratyphoid A bacillus then 

 it will have completely removed the specific paratyphoid A agglutinins from 



1 None of these media will differentiate with any degree of certainty the typhoid bacillus 

 from the paratyphoid bacilli. That however is immaterial since it is unlikely that they 

 will be present together. The main use of these media is to differentiate at sight the 

 typhoid and paratyphoid bacilli from the colon bacillus which is invariably present and 

 usually in large numbers. 



