STUDIES ON CARBON DIOXIDE IV. THE INFLUENCE OF GASEOUS ENVIRONMENT ON GROWTH AND TOXIN PRODUCTION OF C. DIPHTHERIAE

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Abstract
In this study of the influence of different atmospheres on the diphtheria bacillus and its extracellular toxin, the subcutaneous and intracutaneous methods of toxicity determinations were compared. The skin test was found to be a reliable and economical means of determining the approximate toxicity of Corynebacterium diphtheriae culture-filtrates. Test atmospheres containing different concentrations of CO2 and O were supplied to the cultures by an aeration system with which fairly constant gaseous environments could be maintained. Aeration with CO2-free atmospheres containing 10-50% O resulted in marked irregularity in growth and in the toxin content of C. diphtheriae cultures, and permitted an increased rate of destruction of toxin once it was formed, a rate directly proportional to the concentration of O in the atmospheres passed over the cultures. The presence of from 3-10% CO2 in the atmospheres passed over the cultures resulted in increased growth and toxin production. Uniform maximum toxin formation with a given medium was obtained in cultures grown under an atmosphere containing 10% CO2 and 10-20% O. No decrease in tox-icity occurred in any of these cultures on prolonged incubation. An increased CO2 tension also prevented the usual rapid destruction of the bacterial cells after the period of maximum growth. The reaction of cultures grown in meat-infusion broth containing 2 % Difco-pro-teose peptone was maintained at about pH 8.0, pH 7.8 and ph 7.5 by aeration with atmospheres containing 5, 10 and 20% CO2 respectively. The general conclusion is drawn that CO2 plays an important role in the growth and toxin production of C. diphtheriae, mainly by (a) acting either as a catalyst or food in stimulating growth and toxin formation; (b) by controlling the reaction of the culture medium during growth; and (c) by preventing the destruction of the toxin once it is formed.

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