NEUTRAL RED MEDIA 411 



Padlewsky's technique. This method also depends upon the use of a mixture 

 of bile and malachite green. Sow on plates prepared with the following medium : 



Distilled water, 

 Agar, - 

 Peptone, 

 Ox bile, - 

 Lactose, - 



100 c.c. 

 3 grams. 

 3 

 5 



1 gram. 



The medium should be slightly alkaline to litmus. After sterilization cool to 

 65 C. and add, firstly a mixture of 



1 per cent, aqueous solution of malachite green, - 0'5 c.c. 



Ox bile, - 0-5 



then, 



10 per cent, aqueous solution of sulphite of sodium. - 1 c.c. 



12. Method based upon the use of brilliant green. 



Conradi substitutes for malachite green a mixture of Brillantgrun-Kristall 



and picric acid. An agar containing these dyes favours the growth of the 



typhoid and paratyphoid bacilli while inhibiting that of most other organisms. 



The colon bacillus either does not grow at all or only in very small numbers. 



Prepare a slightly modified Hesse's agar : 



Water, 1 litre. 



Peptone, - 10 grams. 



Agar, - 30 



Liebig's extract, - 20 



Make alkaline, heat and filter. For every 1*5 litres of agar add 

 O'l per cent, aqueous solution of Brillantgrun-Kristall (extra 



pure, Hochst), - 10 c.c. 



1 per cent, aqueous solution of picric acid, - 10 ,, 



On this medium, the colonies of the typhoid bacillus are bright green and trans- 

 parent and thicker in the centre than at the margins : colonies of the paratyphoid 

 bacilli are larger, and yellowish-green in colour. 



13. Method based upon the use of neutral red. 



Savage has applied the property possessed by the colon bacillus of reducing 

 neutral red to the detection of that organism in water (p. 397). 



The method is only applicable to the detection of the colon bacillus and 

 does not indicate the presence of the typhoid bacillus. 



The reduction of neutral red, however, is not, as was formerly thought to 

 be the case, a specific property of the colon and paratyphoid bacilli : E. pyo- 

 cyaneus, B. fluorescens, B. enteritidis and some of the harmless saprophytic 

 organisms found in water give fluorescence in Savage's neutral-red broth ; 

 while, on the other hand, some strains of the colon bacillus exert hardly any 

 decolourizing action on neutral red (Sicre, Vincent). Savage's method is 

 therefore unreliable. 



Savage's technique. Prepare broth thus : 



Water, 1 litre. 



Beef, - - 250 grams. 



Boil, make up to 1 litre and add 



Peptone (Defresne), - 20 grams. 



Common salt, - 20 



Glucose, - 5 



Boil, cool, decant and add 10 c.c. of a 5 per cent, solution of neutral-red. Dis- 

 tribute in tubes and sterilize. The medium should be ruby-red in colour. 



Method of analysis. Sow a number of tubes of the medium with different quan- 

 tities (1 c.c. to 10 c.c.) of the suspected water. Incubate at 37 C. for 24 hours. 

 The presence of the colon bacillus is indicated by a beautiful green fluorescence or a 

 canary yellow tint according as to whether the water contains few or many colon bacilli. 



