BACTERIA DECOMPOSING CELLULOSES 197 



The abundant distribution in the soil and the rapid decomposition of 

 cellulose by this organism is thus established and can be demonstrated 

 without difficulty. In addition to the Sp. cytophaga, however, there are 

 other bacteria found in the soil which are capable of decomposing cellu- 

 lose under aerobic conditions. 



Extensive work has been carried out, in this connection, by Keller- 

 man and associates. 16 A special culture medium was developed, for 

 which the cellulose was prepared in the following manner: 



One liter of ammonium hydroxide, specific gravity 0.90, is poured into a glass- 

 stoppered bottle; 250 cc. of distilled water and 75 grams of pure copper carbonate 

 are added; the solution is shaken vigorously until all the copper is dissolved 

 (from 10 to 15 minutes is ordinarily required). Fifteen grams of high grade, 

 sheet filter paper is then added to the copper-ammonium solution and mixture 

 is shaken vigorously, at intervals of 10 minutes, for one-half hour. The solution 

 is examined carefully to see that the paper is completely dissolved. If any parti- 

 cles of paper remain in the solution, the shaking must be continued until the 

 solution is perfectly clear. 



The ammonium-copper-cellulose solution (250 cc.) is diluted to 10 liters with 

 cap water; a weak hydrochloric acid solution prepared by adding 500 cc. of con- 

 centrated acid to 10 liters of tap water is slowly added, with frequent shaking. 

 The addition of the acid is continued until the blue color disappears: a slight 

 excess of acid is added. The mixture is shaken vigorously and allowed to stand 

 a few minutes. The finely precipitated cellulose will rise to the top, due to the 

 large quantity of free hydrogen liberated in the precipitation process. The solu- 

 tion is shaken vigorously, at intervals of a few minutes, to dislodge the hydrogen. 

 As soon as the free hydrogen has escaped, the cellulose will settle rapidly. 



The cellulose is washed through repeated changes of water until free from 

 copper and chlorine. After the washing is complete, the cellulose in the solution 

 is brought up to 0.5 per cent, by allowing to settle a few days and siphoning off the 

 clear solution or by evaporation. The nutrient salts together with 1 per cent of 

 thoroughly washed agar are then added and heated in autoclave or boiled, until 

 the agar is dissolved. The medium is then tubed and sterilized in the usual way. 



Another method for the preparation of cellulose has been suggested 

 by Scales: 17 



Concentrated sulfuric acid (100 cc.) is diluted with 60 cc. of distilled water 

 in a 2-liter Erlenmeyer flask and cooled to 60° or 65°C. Five grams of filter paper, 



16 Kellerman, K. F,, and McBeth, I. G. The fermentation of cellulose. Cen- 

 trbl. Bakt. II, 34: 485-494. 1912; Kellerman, K. F., McBeth, I. G., Scales, F. M., 

 and Smith, N. R. Identification and classification of cellulose dissolving bac- 

 teria. Ibid. 39: 502-522. 1914. McBeth, I. G., and Scales, F. M. The destruc- 

 tion of cellulose by bacteria and filamentous fungi. U. S. Dept. Agr., Bur. PI. 

 Ind., Bui. 266, 1913. McBeth, I. G. Studies on the decomposition of cellulose 

 in soils. Soil Sci. 1: 437-487. 1916. 



17 Scales, F. M. A new method of precipitating cellulose for cellulose agar. 

 Centrbl. Bakt. II, 44: 661-663. 1915. 



