IMPORTANT PATHOGENIC BACTERIA 1 17 



EXERCISE 3. STUDY OF SUBGROXJP III 

 THE TYPHOID-DYSENTERY GROUP 



Use great care in handling members of this group. 



Inoculate agar slants from laboratory cultures of 

 B. typhosus, B. dysenteriae (Shiga), and B. fecalis 

 alkaligenes. 



1. Routine study. — Study the reaction on milk, 

 and test for indol. Preserve glucose agar cultures for 

 two weeks for the observation of involution forms. 



2. Special study. — ^Inoculate fermentation tubes in 

 the same manner as in the two preceding groups. Ob- 

 serve the absence of gas formation but note growth or 

 absence of growth in both arms. Compare, the results 

 with those of the colon and intermediate groups, 



3. Special study. — ^The staining of flagella. — To 

 demonstrate the presence of flagella on B. typhosus, 

 the following methpd will give good results (L6fl3er's 

 method) : 



a) The mordant: 



Tannic acid (20 per cent aqueous solution) . 10 parts 

 Ferrous sulphate (saturated aqueous solu- 

 tion) S parts 



Fuchsin (saturated alcoholic solution) i part 



Add one part i per cent NaOH solution for each 

 100 parts of stain. 



b) 'Prepare several cover sUps by flaming them, and 

 place them side by side on a piece of filter paper. 

 (This paper must be burned after using.) 



c) Place 4 or 5 loopfuls of water on a clean sUde. 



d) Make a light -suspension in this water of B. 

 typhosus from a 24-hour-old agar culture, taking care to 

 stir the suspension as little as possible. 



e) Place a loopful of water on each of the cover slips. 

 /) Carry over a loopful of the suspension on the 



