52 MEDICAL BACTERIOLOGY 



to 4oC., the bacteria are put in it and the tube shaken to distribute them, it is 

 then solidified. 



Stab cultures made in solid media, usually gelatin or agar, are made by 

 thrusting a needle or straight platinum wire coated with bacteria through the 

 long axis of a tube of media nearly to the bottom. 



Gas formation by bacteria is best observed by cultivating in liquid media in 

 fermentation tubes, tubes so constructed that displacement of fluid by generated 

 gas is easily discernible. 



The best form of fermentation tube consists of an upright cylindrical tube, 

 about 5 inches long and ^g inch in diameter, closed at the top and connected 

 at the bottom by a narrower short U-tube to a spherical chamber, of about 10 

 cc. capacity, which has an opening at the top. These fermentation tubes are 

 plugged with cotton and sterilized in a hot-air oven. The media to be used in 

 them is best handled if tubed, sterilized and stored in test-tubes; each test-tube 

 containing sufficient medium to completely fill the upright arm and one-half 

 of the spherical portion of a fermentation tube. 



When a test is to be made the suspected material is first placed in the fer- 

 mentation tube and the culture medium then poured into the bulb. By tipping 

 the fermentation tube the suspected substance and culture medium mix and 

 flow into the upright arm. Every bubble of air must be displaced from the 

 upright portion of the fermentation tube, so that the formation of gas in the 

 tube may be recognized, as it accumulates at the top of the upright arm. 



Indol. Some bacteria produce indol (C 8 H 7 N) when cultured on appropriate 

 media, and this fact is taken advantage of in the identification and differentia- 

 tion of certain organisms. It must be remembered that an organism, like the 

 colon bacillus, which ordinarily produces indol, may at times temporarily lose 

 its ability to do so. 



Dunham's solution has been largely used as a culture medium favorable to 

 indol production, but the Trypsinized peptone-water of Rivas seems superior. 



The organism to be tested is planted in a test-tube containing Dunham's 

 or Rivas' medium and incubated at 37C. If only one tube is planted it is 

 tested for indol after 48 hours of incubation. Sometimes sufficient indol is 

 produced in 6, 12 or 24 hours to be detected and hence when the determination 

 is desired as early as possible, it is expedient to plant several tubes and examine 

 one 6 hours, another 12 hours, another 24 hours, and, finally, one 48 hours after 

 incubation. 



The presence of indol is disclosed by adding 5 to 20 drops of a 0.2 per cent, 

 solution of potassium nitrite, shake the tube, then add slowly an equal quantity 

 of a 25 per cent, solution of pure sulphuric or hydrochloric acid. If indol is 

 present the culture medium turns red, immediately after the addition of acid, 

 or several minutes later. 



For an elaborate discussion of indol production by bacteria, see "Studies 

 on Indol," Rivas, D., Centralblatt fur Bakteriologie, Parasitekunde und 

 Infektionskrankheiten, Orig., 1912, Ixiii Bd. 



Plating. The cultivation of bacteria in broad, shallow, circular, covered 



