458 HEMOGLOBINOPHILIC BACILLI 
Isolation and Culture. — Unlike the PfeiflFer bacillus, B. pertussis 
can be made to grow in media which do not contain hemoglobin. For 
initial growths outside of the human body, however, Bordet and 
Gengou have recommended a potato-glycerin-blood agar medium 
which is claimed to be far more efficient than blood-agar.^ The Avery 
oleate-rabbit-blood agar is even a better medium for this purpose (see 
p. 2.3.S). The Bordet-Gengou bacillus is more readily isolated from 
the bronchial secretion during the first paroxysms- than later in the 
disease. Gultures are obtained from bronchial mucus which has been 
washed several times in sterile water, then spread on the surface of the 
potato medium and incubated at 37° C. After twenty-four to forty- 
eight hours' incubation colonies appear as very minute, transparent 
growths which resemble dew drops; colonies of B. influenzae frequently 
develop at the same time, but the colonies of the latter are somewhat 
larger than those of B. pertussis. Secondary transplantations of B. 
pertussis upon fresh potato-glycerin-blood agar grow more luxuriantly 
than of B. influenzae, how^ever, and after repeated transfers the Bordet- 
Gengou bacillus will grow^ upon ascitic agar. The influenza bacillus 
will not grow in media free from hemoglobin. Ordinary media, unless 
ascitic fluid or blood serum is added, are wholly unsuited for the 
growth of B. pertussis. 
B. pertussis, like the Pfeift'er bacillus, is an aerobic organism. Anaer- 
obic development has not been obtained. The optimum temperature 
of growth is 37° C; Wollstein^ states that slight development takes 
place even at 5° to 10° C. An exposure of thirty minutes at 57° to 
60° C, prevents further development in artificial media. The organisms 
may remain viable upon the potato-glycerin-agar medium for two 
months. 
Products of Growth.— According to Wollstein,'* no acid is produced 
in glucose, lactose, saccharose or mannitol serum broth. No enzymes 
have been demonstrated in cultures of the organism, and it produces 
no visible changes in hemoglobin. Extracelhilar toxins have never 
been demonstrated, but autolyzed cultures introduced intravenously 
into rabbits frequently kill them within tw^enty-four to forty-eight 
hours. Subcutaneous injections of autolysates may cause local 
necrosis, but generalized symptoms fail to appear. Similar results 
have been obtained with endotoxin obtained by grinding the bacilli 
to an impalpable powder and injecting a saline suspension of them.^ 
1 It is prepared as follows: 100 grams finely chopped potatoes are boiled in 200 cc. 
of 4 per cent glycerin for a short time, then cooled. To every 100 cc. of the potato- 
glycerin extract there is added 300 cc. of 7.5 per cent agar containing 0.5 per cent NaCl. 
The glycerin-potato extract replaces the usual peptone-meat-juice nutrients of nutrient 
agar. The mixture is heated to boiling, filtered and sterilized in test-tubes, about 3 cc. 
of medium per tube. (Old potatoes which are slightly alkaline in reaction are much 
better than new potatoes which are usually acid in reaction, in preparing the glycerin 
extract.) To each tube of the sterilized glycerin-potato agar medium is added an equal 
volume of sterile, defibrinated human or rabbit's blood, while the medium is still warm, 
45° to 50° C. Then the mixture is cooled in an inclined position. 
2 Wollstein: Jour. Exp. Med., 1909, 11, 41. ' Loc. cit. 
4 Loc. cit. 6 Bordet and Gengou: Centralbl. f. Bakteriol., ref., 1909, 43, 273. 
