194 



BACTERIA IN RELATION TO PLANT DISEASES. 



CAPSULE STAINS. 



Ribbert's Method of Staining Capsules. 



Water 100 



Alcohol 50 



Glacial acetic acid 12.5 



Warm and add dahlia to saturation. The 

 covers are barely touched to this stain, and are 

 then washed in water. The cover may then be 

 mounted in glycerin or balsam. The stain 

 keeps well. If the cover is left on the stain too 

 long the capsule becomes deep blue and can not 

 be distinguished from the body of the organism. 



Friedlaender's Capsule Stain. 

 (See '85 Friedlaender, Bibliog., XIII.) 

 Richard Muir"s Capsule Stain. 



1 i ) Mordant : 



Mercuric chloride (sat. aq. sol.) 2 



Tannin (20 per cent in water) 2 



Potash alum (sat. aq. sol.) 5 



The dried films are mordanted 2 minutes. 



They are then washed in water, in alcohol, and 



again in water. 



(2) Stain : Carbol-fuchsin 2 to 3 minutes, with 

 gentle heat. 



Wash with water; re-mordant 2 to 3 minutes; 

 wash again with water. 



(3) Counter-stain: Methylene blue (sat. aq. 

 sol.) 2 minutes. Bleach in methyl alcohol, clear 

 in xylol. 



Welch's Capsule Stain. 



Fix in glacial acetic acid. After a few sec- 

 onds pour off the acid and flood with anilin- 

 water gentian violet; repeat this operation until 

 all acid is removed ; wash and examine in salt 

 solution (0.85 to 2 per cent). 



Kaufmann's Method. 



Stain 2 hours at 35 C. in Loeffler's methylene 

 blue; wash in water containing caustic potash 

 or soda (i : 1500), dry; expose 2 minutes in Vi 

 per cent silver nitrate; wash again in the alka- 

 line water ; counterstain 30 seconds in basic 

 fuchsin water (i cc. sat. ale. sol. in 20 cc. aq. 

 dest.) ; expose again to the alkaline water (sec- 

 onds) ; dry and mount. Best adapted to demon- 

 stration of capsules in fresh tissues. The bac- 

 terial body is blue and the capsule red. 



Moore's Night-blue Capsule Stain. 



(See '99 Moore, Bibliog., XIII.) 

 Boni's Method. 



Mix white of I egg, glycerin 50 cc., formalin 

 2 drops; shake well and filter. The bacteria 

 are placed in a drop of this fluid, spread, and 

 heated until the glycerin has entirely evaporated. 

 .Stain 20 to 30 seconds in carbol-fuchsin, wash 

 in water, dry and counterstain 4 to 6 minutes 

 in Loeffler's methylene blue, wash in water, dry, 

 and mount in Canada balsam. The background 

 is red, body of organism blue, and periphery 

 colorless. (See 'oo Boni, Bibliog., XIII.) 



SPORE STAINS. 



Mauser's Spore Stain. 



Pass cover-glass quickly three times through 

 flame. Drop on carbol-fuchsin and heat for 5 

 minutes over flame, renewing the stain as it boils 

 away. Nearly decolorize in dilute sulphuric or 

 acetic acid (5 per cent). Wash very thoroughly. 

 Counterstain with a dilute watery solution of 

 methylene blue or with Loeffler's methylene blue 

 (Festschrift fur Zenker). 



Mailer's Spore Stain. 

 (See '91 Moeller, Bibliog., XI.) 

 Neisser's Spore Stain. 



The cover is floated on hot anilin-fuchsin for 

 an hour. The temperature should be near the 

 boiling point. The cover-glass is then washed 

 in water and decolorized in acid alcohol (i part 

 hydrochloric acid, 3 parts alcohol). Care must 

 be taken not to expose to the acid alcohol too 



long; otherwise the color will be removed from 

 the spores also. The cover is now stained for 

 contrast in a saturated aqueous solution of 

 methylene blue. 



Fiocca's Spore Stain. 



The prepared cover is placed in a watch-glass 

 or test-tube containing 20 cc. of 10 per cent am- 

 monia (water solution) and 10 to 20 drops of 

 alkalin methylene blue or other alkalin solu- 

 tion of anilin color. Then the fluid is heated 

 to the giving off of steam and left for 3 to 15 

 minutes. It is now passed for a moment ( ?) 

 through 20 per cent nitric or sulphuric acid, then 

 thoroughly washed in water and stained for con- 

 trast, if desired, in an aqueous solution of vesu- 

 vin, malachite green, or safranin (saturated?). 

 (See '93, Fiocca, Bibliog., XL) 



(For other methods, e. g., Foth's, Klein's, 

 Aujeszky's, see Bibliog., XI.) 



