THE STUDY OF MICROORGANISMS IN THE SOIL 131 



Agar media containing CaCOa may be employed for isola- 

 tion purposes. 



Frequent transfer of the cultures and incubation in H2S 

 atmosphere will finally lead to isolation of pure cultures. 



Exercise 49 

 Oxidation of Sulphur and the Dissolving of Rock Phosphate 



Prepare a mixture of 60 parts good garden soil, 30 parts of 

 finely ground rock phosphate, and 10 parts of powdered sulphur. 

 Place in pots or tumblers. 



Add enough water to bring to 60 per cent saturation. 



Incubate at 25°C. 



At the end of every week, mix mixture well and add enough 

 water to keep at optimum. 



After 2, 7, 21, 42, 70, and 100 days, determine pH value, 

 amount of water soluble sulphate, and phosphate. Draw curves. 



After 2 and 60 days, determine also number of bacteria and 

 fungi by ordinary plate method. 



Exercise 50 

 Growth and Isolation of Thiobacillus thioparus 



Prepare 600 cubic centimeters of Medium 65. Distribute, 

 in 100 cubic centimeter portions, into 250 cubic centimeter 

 Erlenmeyer flasks. Do not sterilize. 



Inoculate two flasks with 1-gram portions of garden soil, two 

 with fresh manure. Incubate flasks. 



Test every 3 days for disappearance of thiosulphate, using a 

 dilute iodine solution. 



The flasks where the thiosulphate has first disappeared are used 

 for inoculation of two flasks with fresh medium. 



After the thiosulfate has disappeared in the second lot of flasks, 

 examine culture microscopically. 



Note bacteria and granules of precipitated sulphur. 



Culture is now plated upon thiosulphate agar. 



Incubate. 



Isolate from the sulphur-yellow colonies into flasks with sterile 

 liquid medium, in which the nitrogen salt and carbonate have 

 been sterilized separately and mixed aseptically. 



