46 ELEMENTARY BACTERIOLOGY LABORATORY EXERCISES 
SECTION 17: BACTERIOLOGY OF WATER 
EXERCISE 66 
STUDY OF THE COLI-AEROGENES GROUP OF BACTERIA 
Make a detailed study of cultures representing different types of the coli- 
aerogenes group. Determine: (1) morphology, (2) staining reactions, (3) spore 
formation, (4) cultural characteristics on agar slopes, (5) gelatin lique- 
faction, (6) indol production, (7) milk reactions, (8) type of colony on brom 
cresol purple lactose agar, (9) fermentation reactions in glucose, lactose, and 
sucrose broths, (10) methyl red test, (11) Voges-Proskauer reaction. 
The Methyl Red Test 
Grow the cultures in a medium containing: 
0.5% glucose 
0.5% peptone : 
0.5% dibasic potassium phosphate 
Incubate for 5 days at 30° C. (37° C. for 2 days may be used). 
Determine the resulting hydrogen ion concentration using methyl red as the 
indicator. Cultures which are acid to this indicator are designated positive to 
the methyl red test. 
The Voges-Proskauer Reaction 
This test is based upon the ability of certain organisms to produce acetyl- 
methyl-carbinol from glucose. For this purpose part of the culture which is used 
for the methyl red test may be utilized. (Ordinary glucose broth could be used 
for the Vosges-Proskauer reaction, but not for the methyl] red test.) 
After incubation, test as follows: to 5 c.c. of the glucose broth culture, add 
5 cc. of a 10% solution of KOH. Allow to stand over night. A positive test’for 
acetyl-methyl-carbinol is indicated by the presence of an eosin pink color. 
Another test for the presence of acetyl-methyl-carbinol, which gives quicker 
results, may be used. Add 5 c.c. of a 10% solution of KOH to 1 to 4 c.c. of a 48- 
hour culture in methyl red media (or glucose broth). Boil the culture for a few 
minutes by holding the tube in boiling water. The reaction may be hastened by 
shaking the tube. 
EXERCISE 67 
EXAMINATION OF WHOLESOME WATER 
Obtain some water by allowing the faucet to run at least ten minutes before 
collecting the sample in a sterile flask. 
