THE STUDY OF MICROORGANISMS IN THE SOIL 105 

 Exercise 16 



Winogradsky's Method of Microscopic Analysis of Soil 



Take several soil samples, mix carefully, and powder. Add 1 

 gram of soil (on dry basis) to 4 cubic centimeters of distilled water 

 and shake vigorously for 5 minutes. 



Allow to settle for 30 seconds, then pour off suspension into 

 small tube of hand centrifuge. 



Add twice 3-cubic centimeter portions of water to residue, 

 shake 1 minute, allow to settle for 30 seconds and pour into same 

 tube of centrifuge. These manipulations take about 10 minutes. 

 The combined extract (10 cubic centimeters) has in the meanwhile 

 formed another sediment. 



Pipette off 5 cubic centimeters of the supernatant liquid 

 into another centrifuge tube and again centrifuge a few minutes; 

 a third sediment is formed. 



Stained preparations are then made of the three sediments, 

 of the centrifuged and non-centrifuged suspensions. 



A drop is placed upon a slide, to cover 1 square centimeter. 



Dry in oven and cover with a small portion of 1 per cent warm 

 agar solution or 0.1 per cent cold agar solution. 



When the agar has dried, a few drops of absolute alcohol are 

 added to fix the preparations. 



This is followed by staining with a 1 per cent solution of 

 Erythrosin in 5 per cent phenol solution, allowing the dye to act 

 for 5 to 15 minutes in the cold or slight warming, then washing 

 a few seconds in water. Attempt to differentiate between short 

 rods and cocci, bacilli, Azotobacter cells, spores and filaments of 

 fungi, actinomyces filaments, and protozoan cysts. 



Exercise 17 



Determination of Numbers of Nematodes and Other Worms and 



Insects in Soil^ 



Obtain several soil sampling tubes consisting of open cylinders 

 (made of tin or galvanized iron), 72.1 millimeters of internal 

 diameter, with the rim of one end sharpened and the other rein- 

 forced. The area of the internal cross-section of the tube is 



1 Cobb, N. A., Bur. PI. Ind., U. S. Dept. Agr., Agr. Tech. Circ, 1, 1918. 



