IMPORTANT PATHOGENIC BACTERIA 117 



EXERCISE 3. STUDY OF SUBGROUP 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 (Loffler's 

 method) : 



a) The mordant: 



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

 Ferrous sulphate (saturated aqueous solu- 

 tion) 5 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 slips 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 slide. 



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 



