THE STUDY OF MICROORGANISMS IN THE SOIL 107 



If the nodule is small, crush, if the nodule is large, cut open 

 with a sterile knife and press out some of the inner contents into 

 a drop of sterile water. 



Make two or more loop transfers from the first drop of water in 

 a Petri dish to a second containing a few drops of water. 



Repeat these dilutions to a third and fourth Petri dish. Pour 

 Congo red mannitol agar plates (1 liter of Medium 79 + 10 cubic 

 centimeters of a solution of 1 gram of Congo red in 400 cubic 

 centimeters water), agitate until thoroughly mixed and incubate 

 at 28°C. 



Fig. 6.- 



-Bacteria from the nodules of pea ( X 1,500), stained with dilute carbol- 

 fuchsin. 



Instead of diluting in the melted agar, a suspension of the 

 bacteria may be spread over the surface of hardened agar. 



After 5, 10 and 20 days examine the plates. Fish off pure cul- 

 tures and transfer to slopes of Brom thymol blue mannitol agar. 

 Select about 5 pure cultures for each plant. The root nodule 

 bacteria usually form raised moist surface colonies, with round 

 edges, at first glistening, later changing to an opaque white. 



Prepare stained mounts of the root nodule bacteria. 



1. Fuchsin (carbol, according to Ziehl). Dilute with distilled 

 water, 1 part of stain and 9 parts of water. 



