108 CLASSIFICATION AND DESCRIPTION OF BACTERIA 
One of the faetors which has inhibited a more extended use of the Chart 
has been the lack of standard methods to use with it. Several of the 
tests need more study. The Committee on the Chart for the Identi- 
fication of Bacterial Species of the Society of American Bacteriologists 
his made a preliminary report on methods to be used with the Chart 
(Conn, H. J., 1918). Since their report represents the attempt on the 
part of an organized society to standardize technique, parts of their 
report will be reproduced here. 
Media. The importance of using carefully prepared media does 
not need emphasis. Greatest care should be used in adjusting the 
reaction to true neutrality by means of brom thymol blue. Ail media 
should be prepared after the Standard Methods of the American Public 
Health Association. 
Invigoration of Cultures. The Committee* has made the following 
recommendation with regard to this important factor: 
Provided a medium can be found upon which the organism to be studied 
grows vigorously, it should be invigorated before study, even though freshly 
isolated from its natural habitat. The procedure to be employed is as follows: 
Prepare duplicate subcultures in standard gluclose broth, and on standard agar 
slopes, placing cultures of each at 37° and 25°. On the basis of resulting growth 
the organism falls into one of the following series: 
Seres I. Organisms which produce good growth (surface growth, dis- 
tinct turbidity, or heavy precipitate) in twenty-four hours at 37° in glucose 
broth. 
Senes II. Organisms which do not produce good growth in twenty-four 
hours as above, but do in forty-eight hours at 25° in glucose broth. 
Series III. Organisms which do not grow well in glucose broth but do 
produce good growth on the surface of agar in twenty-four hours at 37°. 
Series IV. Organisms excluded from the above groups but which produce 
good growth on the surface of agar in forty-eight hours at 25° C. 
Record the series number on the chart at the proper place and proceed 
with the invigoration by inoculating into another tube of glucose broth for 
organisms of Series III and IV. Incubate this tube at the temperature, and 
for the time, called for in the series which it belongs; then transfer from this 
tube to a third tube and incubate as before. From this third culture make a 
gelatin or agar plate and incubate at the temperature previously used until 
colonies of sufficient size for isolation are obtained. 
Transfer from a typical colony to one or more agar slants and incubate 
one day at 37° or for two days at 25° according to the temperature relation of 
the organism studied. 
* The term ‘‘ Committee ’’ when used in this chapter will refer to the Committee 
on the Chart for Identification of Bacterial Species of the Society of American Bac- 
teriologists. 
