AUTOTROPHIC BACTERIA 81 



According to Winogradsky, the sulfuric acid formed is neutralized 

 by the calcium carbonate or bicarbonate present in the water, since the 

 reaction of the water cultures of these bacteria was not found to become 

 acid. In reference to the physiology of these organisms, the results of 

 Winogradsky can be summarized as follows: (1) The sulfur bacteria 

 oxidize hydrogen sulfide and accumulate sulfur in the form of small 

 spheres, consisting of soft amorphous sulfur which never crystallizes 

 in the living cells. (2) They oxidize the sulfur to sulfuric acid, which 

 is at once neutralized, by the carbonates present, into sulfates. (3) 

 Without sulfur, the organisms soon die off. (4) They can live and 

 multiply in liquid containing only traces of organic substances. 



This last point was refuted by Keil, 44 who demonstrated that the 

 organisms are autotrophic and do not need organic substances for their 

 growth. Keil claims to have isolated pure cultures of Beggiatoa and 

 Thiothrix, and found that these organisms are capable of living in 

 media free from any traces of organic matter, although the presence of 

 small quantities of organic substances is not detrimental to them. 

 The raw cultures were obtained by Keil by placing a layer of black mud 

 containing these bacteria on the bottom of a glass container, 3 to 4 cm. 

 high, covering it with 2 to 3 cm. of river water and placing in the dark, 

 at room temperature. The Beggiatoa formed a white layer over the 

 mud. The Thiothrix could be easily distinguished by the fact that they 

 were fastened at one end. By adding water from a sulfur spring to 

 Petri dishes, then placing these under a bell-jar, the amount of gas 

 necessary for the growth could readily be ascertained. Ammonium 

 salts were found to be used as sources of nitrogen and only carbonic 

 acid as a source of carbon. Carbon dioxide pressure may vary within 

 the limits of 0.5 and 350 mm. (25 mm. is the optimum) ; oxygen may 

 vary within 10 to 20 mm., and H 2 S within 0.6 to 1.7 mm. The presence 

 of carbonates is important for the neutralization of the acids. 



The pure cultures were obtained from the enriched culture by the 

 mere mechanical process of washing out all other organisms first with 

 ordinary water and then with sterile water. This was followed by 

 growth under the bell-jar, at definite gas pressures and frequent changes 

 of medium. Further information on this group of organisms is found 



44 Keil, F. Beitriige zur Physiologie der farblosen Schwefelbakterien. Beitr. 

 Biol. Pflanz, II: 335-372. 1912. 



