36 A MANUAL OF BACTERIOLOGY 



eliminated. This method does not lead to a pure cultivation, for 

 several forms besides the nitrifying organisms persistently main- 

 tain themselves in these mineral solutions. 



So recourse was had to gelatin plate cultivations. Although 

 several organisms were isolated in this manner, none of them 

 possessed the slightest nitrifying power. 



Frankland, and later Warington (1890), succeeded in isolating 

 nitrous organisms by the dilution method. Nitrifying solutions 

 were diluted, and traces inoculated into ammoniacal solutions ; in 

 some of these nitrification occurred, although no growth could be 

 obtained on gelatin, and they were found to contain the nitrous 

 organism only. A little later Winogradsky isolated nitrous 

 organisms, first by modified gelatin plates, and afterwards by the 

 silica jelly method. 



This is carried out as follows : Sodium carbonate is fused in the 

 blowpipe, and fine white sand is added so long as effervescence is 

 produced. The mass is allowed to cool, and is then dissolved in 

 water. The solution is poured into an excess of very dilute 

 hydrochloric acid (silicic acid and sodium chloride being formed). 

 The solution is dialysed and sterilised. For use, some of this is 

 placed in a sterile dish and is mixed with the following solution and 

 inoculated : 



Ammonium sulphate . . . 0-4 grm. 

 Magnesium sulphate . . 0-5 



Di-potassium hydrogen phosphate . 0-1 

 Calcium chloride .... trace 

 Sodium carbonate .... 0-6-0-9 grm. 

 Water 100 c.c. 



This mixture sets to a jelly in five to fifteen minutes. 



Winogradsky has also made use of agar for plates, but this 

 medium is not so suitable as the silica jelly. A 2 per cent, aque- 

 ous agar is prepared and poured into Petri dishes ; the film is then 

 sown with Proteus, and allowed to grow for seven to ten days. It 

 is then thoroughly washed, collected, boiled, and mixed with the 

 salts mentioned above. The object of growing the Proteus upon 

 it as a preliminary is to eliminate the organic matter admixed 

 with the agar. 



Nitrification in the soil is thus brought about by two 

 groups of organisms. The first oxidises ammonia into 



