478 BACTERIOLOGY. 



2. PLATE CULTURES. From another slimy flake 

 prepare a set of gelatin plates. Expose them to a tem- 

 perature of from 20 to 22 C., and after sixteen, twenty- 

 two, and thirty-six hours observe the appearance of the 

 colonies. Usually after about twenty-two hours the 

 colonies of this organism can easily be identified by one 

 familiar with them. 



3. PEPTONE CULTURES. With another slimy flake 

 start a culture in a tube of peptone solution either the 

 solution of Dunham or, as Koch proposes, a solution of 

 double the strength of that of Dunham (Witte's peptone 

 is to be used, as it gives the best and most constant 

 results). Keep this at from 37 to 38 C., and at the 

 end of from six to eight hours prepare cover-slips from 

 the upper layers (without shaking) and examine them 

 microscopically. If comma bacilli were present in the 

 original material, and are capable of multiplication, they 

 will be found in this locality in almost pure culture. 

 After the microscopic examination prepare a second 

 peptone culture from the upper layers of the one just 

 examined, also a set of gelatin plates, and with what 

 remains make the test for indol by the addition of 10 

 drops of concentrated sulphuric acid for each 10 c.c. of 

 fluid contained in the tube. If comma bacilli are grow- 

 ing in the tube, the rose color characteristic of the 

 presence of indol should appear. 



By following this plan "a bacteriologist who is 

 familiar with the morphological and biological peculi- 

 arities of this organism should make a more than probable 

 diagnosis at once by microscopic examination alone, and a 

 positive diagnosis in from twenty to, at most, twenty- 

 four hours after beginning the examination." (Koch.) 



