170 PRINCIPLES OF SOIL MICROBIOLOGY 



placed in the upper part of a Petri dish, then covered with the lower part, placed 

 into the upper part. Solid medium may also be placed in deep layers in ordinary 

 containers, then inoculated with a long platinum loop reaching to the bottom 

 of the container (Liborius). The agar can be taken out from the deep tube, by 

 stabbing to the bottom a sterile glass or metallic tube, 2 mm. in diameter, so as to 

 admit air. 40 



2. Cultivation of anaerobes in vacuo. This method was also proposed by 

 Pasteur and consists in placing the medium in a tube with a capillary end, inocu- 

 lating, pumping out the air, then sealing the end. Petri dishes can also be placed 

 in an ordinary desiccator, from which the atmosphere is then pumped out. The 

 method described by Meyer 41 can be used for the cultivation of bacteria at differ- 

 ent partial oxygen tensions. 



3. Absorption of oxygen from the atmosphere. The most common method of 

 absorption of oxygen from the atmosphere is carried out by means of alkaline 

 pyrogallate solution introduced by Buchner. 42 A mixture of equal portions of 

 10 per cent solutions of pyrogallol and KOH are often used, or 5 per cent solution 

 of the first and 12.5 per cent of the second. Buchner used 1 gram of pyrogallic 

 acid and 10 cc. of 10 per cent solution of KOH for every 100 cc. of air space. 42 The 

 method of Buchner was modified for liquid media : 44 the sterile cotton plug is pushed 

 into the tube; 1 cc. of 20 per cent pyrogallic acid and 1 cc. of 20 per cent KOH are 

 placed upon it, the tube is then closed with a rubber stopper. An alkaline pyro- 

 catechin FeS04 solution to be used as a sensitive reagent for determining traces 

 of oxygen has been suggested. 45 The following method is very convenient: 45 

 About 15 to 20 cc. of agar medium is placed in a large tube, about 1 inch in 

 diameter, the tube is plugged with cotton and sterilized, then slanted. Imme- 

 diately after inoculation, the cotton plug is pressed deeply into the tube, about 

 1 to 2 inches above the tip of slant. One cubic centimeter of a 20 per cent solu- 

 tion of pyrogallic acid (or a tabloid containing 0.13 gram of the acid) and 0.25 cc. 

 of a 40 per cent solution of KOH are poured upon the plug, the tube closed 

 with a rubber stopper, turned upside down and placed in the incubator. 



In the case of Petri dishes, Omeliansky used a combination of evacuation and 

 absorption of oxygen. 43 Ten per cent solution of KOH is poured upon the bottom 

 of a desiccator and an open Petri dish containing dry pyrogallol is placed in it. 

 The dishes containing fresh medium and inoculated are then placed into the 



40 Burri, R., Staub, W., and Hohl, J. Si'issgriinf utter und Buttersaure- 

 bazillen. Schweiz. Milchztg. 45: nos. 78, 83. 1919. 

 "Meyer, 1905 (p. 162). 



42 Buchner, H. Eine neue Methode zur Kultur anaerober Mikroorganismen. 

 Centrbl. Bakt. 4: 149. 1888. 



43 Omeliansky, W. L. Ein einfacher Apparat zur Kultur von Anaeroben im 

 Reagenzglase. Centrbl. Bakt. II, 8: 711-714. 1902. 



44 Wright, J. H. A method for the cultivation of anaerobic bacteria. Centrbl. 

 Bakt. I, 29: 61. 1901. 



46 Binder, K., and Weinland, R. F. liber eine neue scharfe Reaktion auf ele- 

 mentaren Sauerstoff. Ber. deut. chem. Gesell. 46: 255-259. 1913. 



46 Buchanan, R. M. An inset absorption appliance for the test-tube culture of 

 Anaerobes. Centrbl. Bakt. I, Orig., 74: 526-527. 1914. 



