68 



STUDY AND IDENTIFICATION OF BACTERIA 



In Tarozzi's method, pieces of fresh sterile organs are added to 

 bouillon. Pieces of kidney, liver, or spleen are best suited. After 

 adding the tissue the media may be heated to 80 C. for a few minutes 

 without interfering with the anaerobic condition producing properties 

 of the fresh tissues. This method is practically the same as that 

 recommended by Smith (see Tetanus). This is also a feature of 

 Noguchi's method of culturing Treponema pallidum, 



The Method of Liborius. 



In this it is necessary to have a test-tube containing about 4 inches of a i% 

 glucose agar. Glucose acts as a reducing agent and furnishes energy. It is con- 

 venient to add about i/io of i% of sulphindigo- 

 tate of soda; the loss of the blue color at the site 

 of the colony enabling us to pick them out. The 

 tube of agar should be boiled just before using to 

 expel remaining oxygen from the tube. Now 

 rapidly bring down the temperature to about 42 

 C., by placing the tube in cold water, and inocu- 

 late the material to be examined. A second or 

 third tube may be inoculated from the first, just 

 as in ordinary diluting methods for plate cultures. 

 Having inoculated the tubes, solidify them as 

 quickly as possible, using tap water or ice-water. 

 The anaerobic growth develops in the depths of 

 the medium. Some pour a little sterile vaseline 

 or paraffin or additional agar on the top of the 

 medium in the tube as a seal 'from the air. 

 Others have recommended the inoculation of 

 some aerobe, as B. prodigiosus, on the surface. 

 This latter method is not advisable. A deep stab 

 culture is often sufficient. 



The Method of Buchner. 



In this method one gram each of pyrogallic 

 acid and caustic potash or soda for every 100 c.c. 

 of space in the vessel containing the culture is 

 used to absorb the oxygen. It is convenient to 

 drop in the pyrogallic acid; then put in place the 

 inoculated tubes or plates; then quickly pouring 

 in the amount of caustic soda, in a 10% aqueous 

 solution, to immediately close the containing 

 vessel. A large test-tube in which a smaller one containing the inoculated medium 

 is placed, and which may be closed by a rubber stopper, is very convenient. A 

 good rubber-band fruit jar is satisfactory. A desiccator may be used for plates. 



FIG. 20. Arrangement of 

 tubes for cultivation of anae- 

 robes by Buchner's method. 



(Williams.) 



