AUTOTROPHIC BACTERIA 95 



Lieske 89 working with Spirophyllum confirmed the original observa- 

 tions of Winogradsky. The oxidation of FeC0 3 takes place according 

 to the following equation: 



2FeC0 3 + H 2 = Fe 2 (OH) 6 + 2C0 2 + 29 calories 

 Lieske used a medium having the following composition: 



(NH 4 ) 2 S0 4 1.5 grams K 2 HP0 4 0.05 gram 



KC1 0.05 gram Ca(N0 3 ) 2 0.01 gram 



MgS0 4 0.05 gram Distilled water 1000 cc. 



The medium was placed in Erlenmeyer flasks (100 cc.) to a height of 2 cm., 

 sterilized and allowed to stand two days. Coarse iron filings, which had been 

 dry sterilized for one hour at 160°C. were then added, 0.05 gram to each flask. 

 The flasks were inoculated with a small amount of culture of iron bacteria, placed 

 under a bell-jar in a cool place, and C0 2 added up to 1 per cent. This resulted in 

 the formation of about 0.01 per cent of FeC0 3 , which remained constant as long 

 as there was metallic iron present. 



Spirophyllum developed in four days in the cultures to which iron was 

 added. Pure cultures were obtained by repeated transfers to sterile 

 flasks. Similar results were obtained in 1888 by Winogradsky for 

 Leptothrix ochracea, although he did not work with pure cultures. 

 Organic matter in concentrations of over 0.01 per cent (peptone, aspara- 

 gine, sugar) produced an injurious effect upon the growth of Spiro- 

 phyllum. This organism can oxidize no other iron salt, except the 

 bicarbonate, and also not MnC0 3 , showing it to be strictly auto- 

 trophic and highly specialized. Leptothrix ochracea was found by 

 Lieske to be able to utilize manganese carbonate as well as iron car- 

 bonate as a source of energy and to be facultative autotrophic, capable 

 of existing also in organic media. 



The two media used by Lieske for pure culture study have the following com- 

 position: 



I II 



Distilled water 1000 cc. Manganese car- 



Agar 10.0 grams bonate saturated 



Manganese acetate. . . 0.1 gram solution 1:10 



NaHCOs 0.001 per cent 



(NH 4 ) 2 S0 4 0.001 per cent 



K 2 HP0 4 and 

 MgS0 4 Traces 



One may well agree with Harder 79 that certain iron-depositing organ- 

 isms, such as Spirophyllum, require ferrous bicarbonate in solution and 



89 Lieske, R. Beitrage zur Kenntnis der Physiologie von Spirophyllum ferru- 

 gineum Ellis, einem typischen Eisenbakterium. Jahrb. wiss. Bot. 49: 91-127. 

 1911; Zur Ernahrungsphysiologie der Eisenbakterien. Centrbl. Bakt. II, 49: 

 413-425. 1919. 



