A. “Vignal tubes” of the style shown ([Fig. 125]) are made from glass tubes of about 6 to 8 mm. outside diameter, sealed at the small end, plugged with cotton above the constriction and sterilized. The medium, agar or gelatin, which has been previously inoculated with the anaërobic culture, is then drawn up into the tube, after breaking off the tip, as far as the constriction. The tube is then sealed in the flame at the small end and also at the constriction. Since it is full of the medium and sealed, access of air is prevented. This forms an excellent means for “isolation” ([Chapter XVIII]); the tube needs merely to be cut with a file at the point where colonies appear, then these may be readily transferred.
Fig. 125.—Vignal tubes. × ⅓ 1, the sterile tube ready for inoculation; 2, fourth dilution tube showing a few isolated colonies, one near the figure; 3, third dilution showing colonies isolated but numerous; 4, second dilution tube showing colonies still more numerous; 5, first dilution tube showing colonies so numerous and small as to give a cloudy appearance to the tube. In use tube 2 would be filed in two at the colony and inoculations made from it.
B. “Fermentation tubes” form a simple means for growing liquid cultures of anaërobes, the growth occurring in the closed arm only, while with facultative anaërobes, growth occurs both in the closed arm and in the open bulb. A little “paraffin oil” (a clear, heavy petroleum derivative) may be poured on the fluid in the open bulb as a very efficient seal, though it is not usually necessary.
C. “Deep culture tubes.”—The medium, agar, gelatin or a liquid is poured into tubes until they are approximately one-half full, a little paraffin oil is poured on the surface (not essential always), then the tubes are plugged and sterilized. Inoculation is made to the bottom and anaërobes grow well ([Fig. 126]).
Fig. 126.—Deep tubes showing anaërobic
growth. 1, shows a few small gas bubbles; 2, shows the medium broken up by the excessive development of gas.
D. For slope or plate, or any type of surface cultures the Novy jar ([Fig. 127]) is the most practical device. It is good practice to combine the vacuum method, the hydrogen replacement method and the oxygen absorption method in using these jars. In operation a solution of 20 per cent. NaOH is poured on the bottom of the jar to a depth of 1 or 2 cm., the cultures are placed on glass supports above the alkali and a short wide tube of strong pyrogallol is set in on the bottom in such a way that it may be easily upset and mixed with the alkali when it is desired to do so. The cover is now clamped in position with all joints well vaselined. Then the outlet tube is connected with a suction pump and the air drawn out. Meanwhile the inlet tube has been connected with a hydrogen generator, and after the jar is exhausted hydrogen is allowed to flow in, and this process is repeated until one is satisfied that the air is replaced. The suction exhausts the air from the tubes or plates so that much less time is required to replace the air with hydrogen. Finally the stop-cock is closed, and the pyrogallol solution is gently shaken down and mixed with the alkali so that any remaining oxygen will be absorbed.
