134 CONN, HARDING, KLIGLER, FROST, PRUCHA AND ATKINS 



mix with 10 cc. of alcoholic gentian violet prepared as above.) 

 The committee has not yet tested out this procedure. 



Sketches. Drawings of all the morphological characteristics 

 should be made on the blank spaces on the chart to the right of 

 the descriptions. Both typical and atypical forms should be 

 sketched, using care to designate which are typical. 



CULTURAL CHARACTERISTICS 



Cultures for the study of cultural characteristics should be 

 incubated at 37°C. in case of organisms of series I and III, and 

 at 25° in case of organisms of series II and IV, except that gelatin 

 cultures should be incubated at 20°. Room temperature may 

 be used in place of 25° at certain seasons of the year; but if a 

 minimum thermometer shows that the temperature falls below 

 22° during the course of the work, note should be made of the 

 fact. On the day when good growth first appears the proper 

 descriptive terms on the card should be underlined; after subse- 

 quent study, the changes should be noted in the space provided, 

 and sketches of the different stages should be made. 



PHYSIOLOGY 



Liquefaction of gelatin. Old method. The method in most 

 common use is to hold gelatin stab cultures six weeks at 20°C. 

 Plain gelatin should be used. 



Provisional method. It is recommended that the following 

 method proposed by Rothberg (1917) be put in provisional use 

 until experience shows its value. It is designed to distinguish 

 "true liquefiers" (organisms producing ecto-enzymes) from the 

 organisms that produce endo-enzymes of proteolytic action that 

 are released from the cell after death and cause liquefaction of 

 the gelatin if incubated for the long period mentioned above. 

 The method is to give the organism a preliminary cultivation for 

 eighteen to forty-eight hours (according to its rapidity of growth) 

 in a 1 per cent solution of gelatin at 25° or 37° according to its 

 temperature relations; then inoculate on surface of gelatin in test 

 tube and incubate fifteen days at 20°. 



