500 NITROGEN-FIXING BACTERIA 



the pore-spaces in the roots of certain plants. Though suspecting it of being 

 capable of fixing atmospheric nitrogen, he was unable to demonstrate this con- 

 clusively. Numerous strains of Rhizobium have since been described, but as it 

 is doubtful whether they are separate species, we shall confine ourselves to a 

 description of Rhiz. leguminosarum. 



In pure cultures on nutrient agar this organism occurs in the form of rods, 

 1-5-3 ji X 0-5 [JL, but by varying the constituents of the medium it can be made 

 to pass through a cycle of changes, described by Bewley and Hutchinson (1920) 

 as follows : (1) Pre-swarmer stage, non-motile. When a pure culture is inoculated 

 into a neutral soil solution, the organisms assume this form in 4 to 5 days. 

 Diameter about 0-4 ^. (2) Pre-swarmer stage ; larger, non-motile, cocci, 0-8 [x 

 in diameter. Appear in mannitol agar. (3) Swarmer stage ; very actively motile. 

 Cells are ellipsoidal ; 0-9 // X 0-18 p. (Beijerinck). They are so small as to be 



^ 



) 



i 







ff ■' 



1 



Fig. 97. — Rhizobium leguminosarum. 

 1. Pre-swarmer, first stage. 2. Pre-swarmer, second stage. 3. Swarmer. 4. Motile rod. 



5. Highly vacuolated rods. 

 (After Bewley and Hutchinson.) 



able to pass through a Chamberland filter. Appear in carbohydrate media. (4) 

 Rod stage ; motile. Appearance favoured by carbohydrates ; as long as these 

 are abundant, the organism remains in this stage. Dimensions 3-4 // X 1 /^. (5) 

 Stage of high vacuolation. In a neutral soil extract, or in a medium in which 

 the carbohydrate has been exhausted, the organisms become highly vacuolated ; 

 the chromatin divides into a number of bands. Later these bands become rounded 

 off and escape from the rod as the coccoid pre-swarmers (Fig. 97). The forma- 

 tion of the pre-swarmers may also be induced by the addition of calcium and 

 magnesium carbonate to the medium, or by incubating the culture anaerobically. 

 As Lewis (1938) points out, there is no reason to regard this succession of morpho- 

 logical changes as essentially different from that seen in other groups of bacteria, 

 or as indicating the occurrence of a specialized form of reproduction ; the vacuolated 

 state appears to depend on the presence of fat globules. 



Calcium phosphate causes a change from pre-swarmers to rods. Acid soils 

 favour the production of highly vacuolated cells, and eventually kill the organisms. 



