ROUTINE TESTS FOR THE DESCRIPTIVE CHART V4,-13 



amount of solution No. 1. Replace the plug and push down until an inch or an inch and 

 a half above the surface of the culture. Place the tube upright in a boiling water-bath 

 and heat for 15 minutes without letting the culture solution come in contact with the 

 plug. The appearance of a red color on the plug indicates the presence of indole. 



The Kovacs (19£8) test is a simplification of that of Bohme, using only one solution; 

 it is now the method of choice in many laboratories: 



Para-dimethyl-amino-benzaldehyde 5 g. 



Amyl or butyl alcohol 75 ml. 



Hydrochloric acid, concentrated 25 ml. 



This reagent may be used as in the Bohme test, but no solution 2 is required. 



The Gnezda (1899) oxalic acid test is made as follows: dip a strip of filter paper in a 

 warm saturated solution of oxalic acid; on cooling, this is covered with crystals of the 

 acid. Dry the strip of paper thoroughly (sterilization by heat seems unnecessary), 

 and insert into the culture tube under aseptic conditions, bent at such an angle that it 

 presses against the side of the tube and remains near the mouth. Reinsert the plug 

 and incubate the culture. If indole is formed, the oxalic acid crystals take on a pink 

 color. 



It is recommended that the Gore or the Kovdcs test be used in a 

 routine way. In interj^reting the results obtained it must be re- 

 membered that when the reagents are added directly to the medium 

 they react with alpha-methyl-indole as well as with indole itself; but 

 as the former compound is non-volatile it cannot react to the Gore or 

 Gnezda tests. Hence the Ehrlich test unmodified is less specific for 

 indole than the Gore modification or the Gnezda test. 



Some samples of para-dimethyl-amino-benzaldehyde and of amyl 

 and butyl alcohol have been found unsatisfactory for the indole test. 

 It is well, therefore, to check new supplies of these chemicals against 

 samples known to be satisfactory. 



In early editions of this section of the ^lanual, the vanillin, or Steensma, test was 

 also described. It is now omitted, as it is regarded as unreliable; Fellers and Clough 

 (1925), for instance, have shown it to give too high a percentage of positive reactions 

 unconfirmable by any other test. 



The Production of Hydrogen Sulfide 



Hydrogen sulfide is generally detected in bacterial cultures by 

 observing the blackening which it produces in the presence of salts 

 of certain metals, such as lead, iron or bismuth, due to the dark color 

 of the sulfide of these metals. Two methods have been utilized for 

 employing these tests: one by incorporating the metallic salt in the 

 medium, and the other by using a test strip of filter paper impreg- 

 nated with the metallic salt in question. 



In early editions of this Manual four media containing either lead or 

 iron salts were given. The lead salt media, however, were discredited 



