ISO NITRIFICATION 



Di-potassium phosphate (K 2 HPO 4 ). . .5 gr. 



Ferrous sulphate . . . . .4 gr. 



Magnesium sulphate . . . . . 3 gr. 



Distilled water . . . . i litre. 



Take six or eight conical flasks with wide bases, 4 to 5 inches 

 in diameter : pour into each 50 c.c. of the solution, and add 

 .5 gram of basic magnesium carbonate. Inoculate one of the 

 flasks with about i gram of garden soil and keep at a tempera- 

 ture of 30 C. Test the solution for nitrites every two or three 

 days with zinc or cadmium iodide starch paste, and when this 

 reaction ceases test for nitrates with diphenylamine and sulphuric 

 acid (Ex. 74). Then add i c.c. of a 10 per cent, solution of 

 sodium nitrite and test again similarly until the added nitrite dis- 

 appears. Transfer a loopful from this flask into a second flask 

 of the solution, and when nitrate is formed in the second transfer 

 again to a third, and from the third to a fourth. 



The nitrate organisms may be grown on silica jelly impregnated 

 with the above nitrite solution and magnesium carbonate, in the 

 same manner as that described for the nitrite bacteria, Ex. 78. 



They grow, however, on the following agar medium, which is 

 more easily prepared : 



Sodium nitrite (puriss) . . . . 2 gr. 



carbonate (calcined) . . i gr. 



Di-potassium phosphate . . . .2 gr. 



Agar . ; . . .... 15 gr. 



Tap water . . . . . , i litre. 



From the last of several successive generations of cultivations 

 in the nitrite solution, already given above, inoculate 



(i) The melted agar medium in the usual manner with a 

 loopful of the solution: pour into a Petri dish, turn it upside 

 down in order to allow exuded water to run off the surface of the 

 medium. 



(ii) Make a streak culture on solidified nitrite agar in a Petri dish. 

 Incubate for three or four weeks at 30 C. The surface colonies 



