THE STUDY OF OBLIGATELY ANAEROBIC BACTERIA 127 



sterilization and handling of the Spray dish. This is eliminated in the Bray^ dish, 

 which is Pyrex and essentially the same design as the Spray dish. In the Bray dish, 

 however, the need for the lip is eliminated, since the top of the bottom section is 

 slightly smaller in diameter than the remainder of the bottom section. This allows 

 the top to fit down over the rim of the bottom section. 



Method, Pour anaerobic medium in the top half of the dish, and after 

 soHdification, streak from sample or culture, or pour seeded plate. After 

 inverting dish, place 10 ml of 20 per cent aqueous NaOH solution in one 

 section of the bottom dish and 4 ml of 40 per cent aqueous pyrogallic acid 

 in the other. Seal dish with plasticene or tape. Tilt dish to mix solu- 

 tions and place in incubator. 



Advantages. Anaerobiosis is attained quickly. It is a useful method for single 

 plate culture. Since each plate is a single unit, observations may be made at any 

 time and any particular plate of a series may be opened when visual inspection reveals 

 growth to be at the desired stage. Recommended for clinical laboratory technicians 

 seeking a quick method of purification of possible pathogenic types. Disadvantages. 

 Preparation of individual dishes is time-consuming, and in work on any scale, anaerobic 

 jars may be preferred. Loss of anaerobiosis due to leaks in seal is not uncommon. 

 The excess of alkali present results in absorption of CO 2, which may be disadvantage- 

 ous with some strains. In addition, a small amount of CO is given off during oxida- 

 tion of the pyrogallate. Some strains are inhibited by this substance. Preliminary 

 replacement of air by an inert gas before introduction of the pyrogallate will minimize 

 this effect. 



Anaerobic Jar with Alkaline Pyrogallol for Plate Culture 



For each 100 ml of jar capacity 1 g of pyrogallic acid and 10 ml oi 

 2.5N NaOH are used. Plates are stacked in the jar, together with 

 anaerobic indicator tube, and pyrogallic acid is added to alkali in a large- 

 diameter test tube. The jar is quickly sealed, using plasticine or silicone 

 grease. 



Advantages. General availability of materials and suitability for a variety of jars, 

 which need be only sealable. Disadvantages. As noted above, absence of CO2 and 

 presence of CO may inhibit growth of some strains. 



Tube Culture 



Materials for method of Griffin (1932) : (1) Two test tubes with approxi- 

 mately 5^ in. diameter (one empty and the other containing a liquid or 

 slant culture of the anaerobe), (2) two one-holed rubber stoppers to fit 

 tubes, (3) short piece of small-diameter rubber tubing, (4) two short 

 pieces of glass tubing of diameter to fit tightly in holes of rubber stoppers, 

 (5) small glass vial, (6) dry pyrogallic acid, (7) strong aqueous NaOH. 



1 Bray anaerobic dish. Corning Glass Works, Corning, New York, Pyrex No. 3155, 

 or dealer. 



