CONRADI AND DBIGALSKI'S METHOD 407 



very rapidly through the walls of a porous bougie, and the various modifications of 

 the method which have been introduced seem to have little to recommend them. 



5. Chantemesse's methods. 



Chantemesse has introduced two methods of isolating the typhoid bacillus, 

 both of which depend upon obtaining surface colonies on carbolic agar. For 

 isolating the organism from stools and water the second is not only more 

 rapid but is simpler. 



First method. Filter 5 or 6 litres of the suspected water through a Chamberland 

 bougie. Wash the surface of the filter in 200 c.c. of a 3 per cent, peptone solution. 

 Incubate the latter at 37 C. and arrange the culture so that air can be bubbled through 

 it while incubating. Add more peptone solution at the end of 12 hours and again 

 at the end of 24 hours. Then centrifuge the culture. The typhoid bacilli being 

 motile and isolated (i.e. not grouped in clumps) remain in suspension while non- 

 motile organisms and those massed together in zooglea masses go to the bottom of 

 the vessel. With the supernatant liquid sow a number of Esmarch's roll tubes by 

 the dilution method using carbolic-agar as the medium. 



Carbolic-agar. Dissolve 30 grams of peptone and 20 grams of agar in a litre of 

 water, and make feebly alkaline (p. 31), tube in quantities of 10 c.c. and sterilize. 

 Immediately before use melt the agar and add four drops of a 5 per cent, solution of 

 carbolic acid to each tube ( =0'1 per cent, of carbolic acid). 



Incubate the cultures at 37 C. Growth appears in about 16-20 hours. Sow 

 all colonies at all resembling the typhoid bacillus on the various media used for 

 differentiating the organism. 



Second method (recommended). Sow the suspected material directly on 

 litmus-lactose-carbolic-agar. 



Litmus-lactose-carbolic-agar. Prepare agar as above and add 2 per cent, lactose. 

 Tube in quantities of 10 c.c. and sterilize. Just before use melt a number of tubes 

 of lactose-agar and to each add 1 c.c. of sterile neutral litmus solution and 4 drops 

 of a 5 per cent, solution of carbolic acid. Mix thoroughly, pour into Petri dishes 

 in thin layers (1-2 mm. deep) and allow to set. 



Dip a fine sterile badger-hair brush in a tube of sterile water to which a 

 trace of the suspected stool has been added, and without recharging it sow 

 in succession six surface plates of litmus-lactose-carbolic-agar. 



If water is to be examined, filter it through a Chamberland bougie and sow 

 litmus-lactose-carbolic-agar plates with the deposit left on the bougie. For 

 spreading the plates use a glass rod bent at a right angle (Drigalski's spatula). 



Incubate the plates at 37 C. and after 12-15 hours numerous colonies will 

 be found on the plates some red (the colon bacillus) and some blue (the typhoid 

 bacillus). Test the blue colonies by the agglutination reaction. 



6. Conradi-Drigalski's method. 



This method, in principle the same as that of Chantemesse, is in very general 

 use in Germany. 



The suspected material is sown on the surface of agar containing lactose 

 and litmus. Crystal- violet is used in place of the carbolic acid in Chante- 

 messe's medium, and is found to be just as effective in restraining the majority 

 of organisms while allowing the growth of the colon and typhoid bacilli. 



Conradi and Drigalski's medium. 



(a) Preparation. Macerate 1500 grams of minced beef in 2 litres of water for 

 24 hours : boil the mixture for an hour : filter : make up to 2 litres with water : 

 add 



Peptone (Witte), .... 20 grams. 



Nutrose, 20 



Salt, - 10 



boil, add 60 grams of agar ; heat until the agar is dissolved : make feebly alkaline 



