106 LABORATORY MANUAL OF MICROBIOLOGY 



just one-millionth of an acre. The tubes are usually 6 to 9 

 inches long. 



By means of this tube, several samples are obtained from 

 different parts of the field. The soil of the various samples is 

 then well mixed and sifted. 



An aliquot portion of soil is then suspended in 10 to 20 vol- 

 umes of water and well shaken. 



The heavy particles of soil are allowed to settle for 5 seconds 

 and the supernatant liquid immediately poured into another 

 vessel. 



The residue is washed two or three times with clean water and 

 the washings added to original liquid. 



The liquid is allowed to run through a series of superimposed 

 sieves, ranging from 16- to 200-mesh per inch and the residual 

 material is washed with a little water. When the liquid flowing 

 through the finest sieve shows no animals, it is discarded. 



The animals are washed away from each sieve with a small 

 amount of water and washings with animals placed in a series of 

 tubes. 



The animals settle more readily to the bottom than the clay. 

 The suspended liquid can be removed after 30 minutes' settling 

 and replaced by clean water. The mixture of inorganic particles 

 and organisms is then examined under the microscope. 



NITROGEN-FIXING BACTERIA 

 Exercise 18 



The Isolation of Bacteria from the Root Nodules of Various 

 Leguminous Plants 



Thoroughly wash the roots and nodules of several leguminous 

 plants {e.g., red clover, alfalfa or sweet clover, pea or vetch, 

 and soybean) under the tap. 



Compare the number, size, color and position of the nodules on 

 the roots of these different leguminous plants. 



Select a large and firm nodule, cut off, and immerse for 3 to 5 

 minutes in mercuric chloride solution (1:1,000), or in 70 per 

 cent alcohol. Remove alcohol or mercuric chloride by washing in 

 sterilized water and place the nodule on a sterile surface (flamed 

 slide or Petri dish). 



