192 Nitrogen Fixation— Nonsymbiotic 



ago, but it is only within the last decade or two that the true bio- 

 logical nature of the process has been established. Numerous other 

 claims concerning the ability of various organisms to fix atmospheric 

 nitrogen have been questioned. An organism is considered unable 

 to fix nitrogen if no increase in combined nitrogen can be demon- 

 strated by chemical analysis. Even then, if such an increase can be 

 demonstrated, the importance of the reaction in the soil itself may 

 still be open to question. 



Berthelot claimed in 1885 to have demonstrated that, when a soil 

 is exposed to the air, its nitrogen content gradually increases and the 

 fixation of nitrogen is biological in nature. This claim was not sub- 

 stantiated. It was not until six years later that the capacity for 

 nitrogen fixation by nonsymbiotic organisms was established by Wino- 

 gradsky. Clostridium pasteiiriamim, an anaerobic organism belong- 

 ing to the group of butyric acid bacteria, was the first organism to 

 be found capable of bringing about an increase in the amount of 

 combined nitrogen in the medium. An available source of energy 

 was required for this purpose. A definite ratio was found to exist 

 between the carbohydrate consumed and the amount of the nitrogen 

 fixed. 



Following Winogradsky's work, Beijerinck demonstrated in 1901 

 that nonsymbiotic nitrogen fixation can be carried out by aerobic 

 bacteria belonging to the genus Azotohacter. Other organisms, desig- 

 nated as Graniilobacter, were also found capable of fixing atmos- 

 pheric nitrogen. In addition to these, numerous other bacteria 

 in the soil were found capable of fixing small amounts of nitrogen 

 on artificial culture media, especially when freshly isolated from 

 the soil. 



It has been claimed that various other organisms, in addition to 

 the bacteria, have been found capable of fixing varying amounts of 

 atmospheric nitrogen. These organisms ranged from different groups 

 of algae and fungi to a variety of animal forms. Most of these claims 

 have remained unsubstantiated. However, some of the blue algae 

 and some of the purple ( nonsulf ur ) bacteria have been found capable 

 of fixing molecular nitrogen. 



Classification of Nitrogen-Fixing Organisms 



The nitrogen-fixing bacteria recjuire sources of energy that they 

 are able to obtain from certain organic compounds of carbon, which 

 are also used for cell svnthesis. These organisms can be classified 



