404 ISOLATION OF THE TYPHOID BACILLUS 



B. Grimbert's method. 



Grimbert attributes the failure of Eisner's method partly to the want 

 of uniformity of the medium due to variations in the chemical composition 

 of potatoes, and partly to the fact that Eisner did not test the reaction of 

 his medium. According to Grimbert the addition of iodide of potassium is 

 not essential : ordinary gelatin can be used if the reaction be such that 

 10 c.c. are neutralized by 5 c.c. of lime water, though it is better to have a 

 medium of constant chemical composition. Grimbert's medium is used in 

 the same way as Eisner's, but the colonies are more slow in developing and 

 the earliest do not appear before the third day. The method, as a matter of 

 fact, has hardly any advantage over Eisner's original method. 

 Technique. To 1,000 c.c. of water add : 



Maltose, - 1 gram. 



Soluble starch, 2 grams. 



Asparagin, - 2 



Neutral phosphate of potassium. 2 ,, 



Potassium sulphate, - 2 



Magnesium sulphate, 2 ,, 



Ammonium bimalate, - 2 ,. 



Magnesium carbonate, - 1 gram. 



Dissolve 15 per cent, of gelatin in the mixture, clear with white of egg, heat to 

 115, filter, and test the reaction thus : dilute 10 c.c. of the gelatin with 50 c.c. of 

 warm distilled water, add a few drops of an alcoholic solution of phenol-phthalein, 

 then run in lime water until a permanent rose pink colour is obtained. If more 

 than 3 c.c. of lime water are required to neutralize the gelatin reduce the acidity by 

 the addition of a small quantity of normal soda solution until 10 c.c. of the gelatin 

 are neutralized with 5 c.c. of lime water. 



Immediately before use 1 per cent, of iodide or bromide of potassium may be 

 added. 



C. Remy's method. 



Kemy suggests the use of a medium which is more nutritive and less acid 

 than Grimbert's. By means of his " differential gelatin " he has been able 

 to isolate the typhoid bacillus from stools in all the cases of enteric fever 

 which he has investigated. 



This " differential gelatin " has no greater selective property than Eisner's 

 medium and the majority of micro-organisms grow in it. Still, liquefying 

 species are to some extent checked and the inhibiting influence of the colon 

 bacillus on the typhoid bacillus is not apparent on this medium. 



Technique. Preparation of the " differential gelatin." To a litre of water in a 

 flask add : 



Asparagin, - 6 grams. 



Oxalic acid, - - 0'5 gram. 



Lactic acid, - - 0'15 



Citric acid, - - .... Q*15 



Di- sodium phosphate, - - 5 grams. 



Potassium sulphate, - - - - 1*25 ,, 



Sodium chloride, - - - - - - - - 2 



Witte's peptone, - .... 30 



Heat to 110 C. for 15 minutes, and on taking the flask out of the autoclave pour 

 the boiling liquid into another flask containing 120-150 grams of best quality gelatin. 

 Shake the flask until the gelatin is dissolved, add soda solution until the mixture is 

 slightly alkaline, heat in the autoclave again to 110 C. for 15 minutes, then add 

 sufficient half-normal sulphuric acid x to render the medium acid to such an extent 

 that 10 c.c. require the addition of 0'2 c.c. of half-normal solution of soda to neutralize ; 2 

 mix by shaking well, then heat in the steamer at 100 C. for 10 minutes and filter. 



1 A normal solution contains 98 grams H. 2 S0 4 per litre. 



2 This acidity is equivalent to 0'5 gram of H. 2 S0 4 per litre. 



