AUTOTROPHIC BACTERIA 65 



the bacterium from the sediment was found to be regularly oval or 

 ellipsoidal in nature; it was also present in the medium. In order to 

 obtain pure cultures of these organisms Winogradsky 11 made use of the 

 two facts that the bacterium did not grow on the gelatin plate and in 

 nutrient bouillon but was found abundantly on the magnesium car- 

 bonate sediment. Some of that sediment was streaked out over a gela- 

 tin plate and those particles, which remained sterile on the plate, after 

 a few days incubation, were transferred into fresh flasks containing the 

 ammonium medium. The final steps in the study and isolation of the 

 bacteria concerned in the process of nitrification were the separation 



Fig. 8. Winogradsky flask for experiments on nitrification. Layer of MgCOj 

 on bottom of flask and mineral salt solution above it (after Omeliansky). 



of the nitrite and nitrate organisms and their cultivation upon specific 

 media. 



For the growth of the nitrite and nitrate forming bacteria, a 

 thorough aeration of the culture is essential. Winogradsky used 

 flasks of large diameter (12 cm) with flat bottoms, in which the 

 liquid formed only a shallow layer (less than 1 cm. in height). Boul- 

 anger and Massol 12 used scoria and allowed the liquid to reach only 

 half the height of the scoria layer, which was moistened at regular 



11 Winogradsky, S. Recherches sur les organismes de la nitrification. Ann. 

 Inst. Past. 4: 213-231, 257-274. 1890; 5: 92-100. 1891. 



12 Boulanger, E., and Massol, L. Etudes sur les microbes nitrificateurs. Ann. 

 Inst. Past. 17: 492-515. 1903; 18: 181-196. 1904. Compt. Rend. Acad. Sci. 

 687. 1905. 



