132 PRINCIPLES OF SOIL MICROBIOLOGY 



calcium or magnesium carbonates are added to the medium, or under anaerobic 

 conditions. Acid soils cause the production of highly vacuolated cells and 

 eventually kill the organism. These studies need further confirmation. 



Motility. In young agar slants, the organisms are found to be very 

 motile. Owing to the slime produced by the organism, the demonstra- 

 tion of flagella is very difficult; this was the reason for considerable 

 disagreement among the different investigators. It has come to be 

 recognized, 142 however, that the nodule bacteria possess two types of 

 flagellation: peritrichous and monotrichous. Differences, however, 

 have been reported even for a single strain. The soybean organism 

 was reported 143 by some as possessing peritrichic flagellation, but by 

 most other workers 144 as monotrichous. The differences thus obtained 

 were due either to the fact that cultures of various ages were employed 

 or different types of bacteria exist, even for the same plant (as Soja 

 max), in different parts of the world. 145 ' 146 



For staining of flagella, the following modification of the Loeffler's stain may be 

 used : 



Solution A 



parts 



Ferric chloride (1 : 20 aqueous solution) 1 



Saturated aqueous solution of tannic acid 3 



This solution improves with age; it should be at least a week or two old and 



should be filtered before using. 



Solution B 



parts 



Anilin oil 1 



95 per cent alcohol 4 



The bacterial suspension is allowed to air-dry on a clean cover glass. About 

 5 drops of solution A are then placed on the cover glass, followed immediately 

 by 1 to 2 drops of solution B. The combination is allowed to act at room tem- 

 perature for 2 minutes and is then washed in distilled water. The stain (30 parts 

 of saturated alcoholic solution of methylene blue, 13 parts of solution B as mord- 

 ant and 100 parts of 1 : 10,000 KOH solution) is applied for 2 minutes. 



142 Hansen, R. Note on the flagellation of the nodule organisms of the Legum- 

 inosae. Science. N. S. 50: 568-569. 1919. 



143 Wilson, J. K. Physiological studies of Bacillus radicicola of soybean {Soja 

 max Piper) and of factors influencing nodule production. Cornell Univ. Exp. 

 Sta. Bui. 386. 1917. 



144 Wright, 1925 (p. 127). 



145 Shunk, I. V. Notes on the flagellation of nodule bacteria of leguminosae. 

 Jour. Bact. 6: 239-246. 1921; Ibid. 5: 181-187. 1920. 



140 Fred and Davenport, 1918 (p. 582). 



