54 ELEMENTARY BACTERIOLOGY LABORATORY EXERCISES 
EXERCISE 79 
STUDY OF BACTERIUM COLI AND BACTERIUM AEROGENES 
Review the general morphological, cultural, and physiological characteristics 
of organisms of the coli-aerogenes group as determined in Exercise 66. 
What action do the members of this group of organisms have upon lactose? 
Do they liquefy gelatin? From these characteristics, what action would you ex- 
pect them to have on milk? 
Inoculate tubes of plain milk, litmus milk, and brom cresol purple milk with 
Bacterium coli and Bacterium aerogenes. Also inoculate Smith fermentation 
tubes containing lactose broth with these organisms. Incubate all the cultures at 
37° C. for two days. 
Note carefully the type of fermentation produced in milk by these organisms: 
whether acid is produced, the milk curdled, type of curd produced, evidences of 
gas formation in the curd and in the fermentation tube, and whether there is any 
evidence of peptonization or casein digestion. Titrate a sample from the plain 
milk culture and calculate its acidity as lactic acid. 
EXERCISE 80 
STUDY OF LACTOBACILLUS BULGARICUS 
From the milk culture of Lactobacillus bulgaricus furnished make stained 
preparations using methylene blue and Gram’s stain. Study the morphology of 
the organism and make drawings. Note whether Gram positive or negative. 
Inoculate: (1) nutrient agar slope, (2) plain nutrient broth, (3) litmus 
milk, (4) brom cresol purple milk, and (5) three tubes of plain milk. Incubate 
at 37° C. 
Observe the agar slope and plain bouillon cultures. Is there much evidence 
of growth on these media? Observe the milk cultures for: acid production, coag- 
ulation of milk, type of curd produced, evidences of gas formation, and casein 
digestion. Titrate the plain milk cultures and calculate the percentage of lactic 
acid after 2, 7, and 14 days’ incubation at 37° C. 
Although members of, the Lactobacillus group do not grow rapidly enough 
at low temperatures to take part in the ordinary souring of milk, they are always 
present in ordinary milk and will develop upon prolonged incubation. Fill a 
small bottle with skimmilk. The bottle should be filled to the neck and then 
tightly stoppered so as to prevent mold growth. Incubate at 37° C. Make micro- 
scopic preparations using methylene blue and Gram’s stain after 2, 7, and 14 
days’ incubation. Make a drawing of a typical field at each of these periods. 
Determine the percentage of lactic acid produced after 2, 7, and 14 days’ incu- 
bation at 37° C. 
