IV46-18 MANUAL OF METHODS FOR PURE CULTURE STUDY 



water non-spore-formers and with plant pathogens) because of slime 

 production, unusually fine flagella or flagella that are readily lost. 



Mordant: Solution A 



Tannic acid (10% aqu. solution) 18 ml. 



FeClsGHjO (6% aqu. solution) 6 ml. 



Solution B 



Solution A 3.5 ml. 



Basic fuchsin (0.5% in ethyl alcohol) 0.5 ml. 



HCl, concentrated 0.5 ml. 



Formalin 2.0 ml. 



Staining schedule: 



1. Prepare smears of young cultures, following carefully the 



procedure recommended on p. 15 under "Methods of handling 

 cultures". 



2. Filter the above Solution A onto the slide and allow it to remain 



3}/2 min. without heating. 



3. Pour off solution A, and without washing add solution B, also 



through a filter, and allow it to stand 7 min. without heating. 



4. Wash with distilled water. 



5. Before the slide dries, cover with Ziehl's carbol fuchsin (p. 5), 



allowing it to stand 1 min. on a hot plate heated just enough 

 for steam to be barely given off. 



6. Wash in tap water. 



7. Dry in the air and examine. 



Results: Similar to the preceding methods; but the background pre- 

 cipitate is usually finer and less conspicuous, thus interfering less with 

 the demonstration of unusually fine, delicate flagella. 



Staining flagella of anaerobes. O'Toole (19-12) calls attention to 

 certain difficulties in staining the flagella of anaerobes, and gives a 

 modification of the above Bailey stain which is intended to overcome 

 them. The method is not unlike that of Fisher and Conn who had 

 the O'Toole procedure in mind when working out their modifica- 

 tion. The O'Toole method does not seem to be as satisfactory as 

 the Fisher and Conn procedure for the above mentioned soil bacteria 

 and plant pathogens; but one must remember that it is particularly 

 recommended by its author for an entirely different type of organism. 



Capsule Stains— Recommended Procedures 



Bacterial capsules are more easily confused with artifacts than any 

 other structure pertaining to the organisms. Inasmuch as capsules 

 sometimes show merely as unstained areas around the cells, there is a 

 temptation to call any such surrounding area a capsule; very often, 

 however, they merely represent the tendency of a lightly stained sur- 

 rounding medium to retract from the cells on drying. For this 

 reason the best way to demonstrate capsules is actually to stain them 

 by some procedure which differentiates them from the cell itself. 

 Several of the flagella stains accomplish this, notably those of Bailey 

 and Leifson, given above. Much simpler is the procedure of Anthony 

 described below. The Anthony method can be recommended both 

 because of its simplicity and its dependability. Any of the other 



