30UTINE TESTS FOR THE IDENTIFICATION OF BACTERIA 157 



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. 



THE PRODUCTION OF HYDROGEN SULFIDE 



Hydrogen sulfide is generally detected in bacterial cultures by observ- 

 ing the blackening which it produces in the presence of salts of certain 

 metals, such as lead, iron, or bismuth, owing 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 impregnated with the metallic 

 salt in question. 



In early editions of the manual four media containing either lead or 

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

 some time ago because of the toxic properties of these salts, and Hunter 

 and Crecelius (1938) show the superiority of bismuth media over iron 

 media. Zobell and Feltham (1934), moreover, have shown distinct 

 advantages from the use of lead acetate test strips, without any of these 

 metallic salts in the media. The advantage of the test strip technic is 

 that it is more sensitive and does not introduce the possibility of inhibit- 

 ing the bacterial growth if the concentration of metallic salt in the medium 

 is too great. It is important, as emphasized by Hunter and Crecelius, 

 that the indicator and method employed be stated when results are 

 given. Untermohlen and Georgi (1940) suggest use of nickel or cobalt 

 salts but specially emphasize the variations in results w^ith different media 

 and indicators. 



When using the test-strip technic the bacteria may be grown in ordinary broth, 

 peptone solution alone, or a peptone agar suitable to the organism in question. One 

 must be certain that the peptone contains available sulfur compounds. This can be 

 determined by running a check tube inoculated with a slow hydrogen sulfide producer. 

 For this procedure the test strip should be prepared by cutting white filter paper 

 mto strips approximately 5 by 50 mm, soaking them in a saturated solution of lead 

 acetate, sterilizing them in plugged test tubes, and drying in an oven at 120°C. One 

 of these strips should be replaced in the. mouth of the culture tube before incubation in 

 such a position that one-quarter to one-half of the strip projects below the cotton plug. 

 These tubes should be incubated at about the optimum temperature of the organism 

 under investigation and examined daily to notice whether or not blackening of the test 

 strip has occurred. 



Because of the inconvenience of the test-strip technic, media in which 

 iron salts are incorporated are now generally preferred. A dehydrated 

 medium of such composition is available and has been found quite 

 satisfactory. 



