258 GENERAL MICROBIOLOGY 



slanted nitrogen-free agar and note characteristic growth. 

 Why is nitrogen-free agar used for the cultivation of Ps. 

 radicicolaf 



18. Make permanent stains of pure culture and compare 

 with organisms on stained smear from nodule as to size, 

 shape, etc. Are involution forms present in either prepara- 

 tion? 



Do all species produce organisms of the same general 

 morphology in the respective nodules? In pure culture? 



19. Make a flagella stain from the pure culture as fol- 

 lows: 



a. Take a loopful of the mucilaginous growth from a 

 colony or an agar culture and spread it on a clean slide, 

 lashing it out in slender tongues. 



6. Let the film dry in air without killing or fixing. 



c. Flood the film a moment with saturated alcoholic 

 solution of gentian violet. 



d. Wash under the tap, dry and examine with the oil 

 immersion lens. 



20. The mucilage in which the cells lie will be found 

 to be deeply and evenly stained and the bacteria stained 

 scarcely at all, so that the preparation presents somewhat 

 the appearance of a photographic negative. 



The single polar flagellum may be demonstrated by 

 this stain, since it, like the protoplasm of the cells, refuses 

 the stain, and so it appears as a clear, uncolored streak in 

 the surrounding deeply stained mucilage. The flagella 

 are best seen at the margins and in thin places, inasmuch 

 as the mucilage in the denser areas masks the slender fla- 

 gella. 



21. Sometimes the roots of leguminous plants show, 

 instead of the normal nodules, lesions of crown-gall caused 

 by Bad. tumefaciens which somewhat resemble Ps. radici- 

 cola. 



22. For a rapid diagnosis, plate in the synthetic Congo 

 red medium which differentiates these two organisms; 



