118 BACTERIAL DISEASES OF PLANTS 



bacteria have held the stain less than some substance between 

 them. Where only tissue differences are to be brought out, 

 methyl green followed by acid fuchsin (see Part III, No. XIV) 

 leaves little to be desired, at least in certain plants, e.g., crown 

 gall on the Paris daisy, sunflower or house geranium, where the 

 parenchymatic tissues become red or pink, and the lignified 

 tissues blue. 



Staining Bacteria from Cultures. — A variety of stains are 

 useful for staining smears and streaks from dilutions of pure 

 cultures e.g., methylene blue. Gentian violet, basic fuchsin, 

 carbol fuchsiUj, amyl Gram. It is important to start with clean 

 slides or covet's, to dilute the culture so that the bacteria shall 

 not be crowded, and so to stain and wash that the bacteria 

 shall be sharply and deeply colored on a clean clear background. 

 A momentary bath in alcohol following the stain is sometimes 

 serviceable, or in acid alcohol if the organism is an acid fast 

 organism, or in iodine followed by ethyl alcohol if it stains by 

 Gram's method. 



Flagella Staining. — This is more troublesome than either of 

 the preceding and will tax to the uttermost the ingenuity and 

 capacity of the average student. Some never learn to do it, 

 most are able to accomplish it with perseverance. A very few 

 become experts and obtain beautiful preparations with com- 

 parative ease. To succeed, the culture must be suitable. One 

 seldom obtains good preparations from cultures that are not 

 actively motile. Some species are much more obdurate than 

 others. Attempts should be made as a rule only from very young 

 agar-streak cultures (6-24 hours) and not then unless an inspec- 

 tion of the organism on the margin of a hanging drop shows it to 

 be actively motile. Sometimes better success may be had by 

 transfers from bouillon to agar than from agar to agar, or by 

 flooding a young agar culture with 15-20 cc. of distilled water 

 and then taking motile organisms from the top part of the fluid. 



The sHdes or covers used must be absolutely clean and of 

 good quality. The mordanting fluid should be fresh. The 

 bacterial film should be evenly spread and the individual bacteria 

 widely separated but not too widely. The covers maybe lightly 

 flamed before staining but this is not always necessary. Very 



