Chemical Detoxification of Flexner Dysentery Antigen: II. Studies of Accelerated Growth Rates.

A major problem which arises in work on the chemistry of bacteria is that of obtaining adequate amounts of organisms with the facilities of the ordinary research laboratory. For this purpose, in growing bacteria for chemical studies on Flexner dysentery organisms (Barnes, Dewey, Henry, and Lupfer, in press), a method was developed for obtaining large amounts of this organism rapidly, and in high concentration, using simple constituents and standard laboratory apparatus. The method is based on the observations that the organism flourishes in the presence of aeration (Topley and Wilson, 1936; Hoberman and Dubos, unpublished observations), and that growth is enhanced if the effect of the hydrogen ions produced during growth is buffered by suitable salts (Hoberman and Dubos). Conditions for culturing organisms in which a large surface is exposed to a gas phase have been described by several investigators (Mironova, 1941; Clifton, 1943; Johnson, Bruce, and Dutcher, 1943; Hoberman and Dubos). By this method the virulent type III (Z) strain with which we were concerned was grown to concentrations 8 to 10 times those obtained over 18 to 24 hours by usual culture methods (table 1, figure 1). Also, the rate of growth was increased so greatly that each liter of culture medium produced this concentration in 21 to 3 hours. In this period, therefore, the final result was a net increase of 60to 80-fold over the usual 20-hour growth rates. If, at the end of the 3-hour period, the bacteria were kept in the logarithmic phase of growth through the prompt addition of a fresh liter of medium, the effect could be reproduced as often as desired. The logarithms of the values for turbidity and live count are plotted against time in figure 2. It may be seen from this figure that the rapid regeneration rate of the organisms in still media during the first hour was continued after shaking began. During the logarithmic phase. the number of live organisms doubled every 40 minutes, but after about 2' hours of shaking the regeneration rate decreased sharply. The relative increase in mass during the shaking period was only 3-fold, whereas that in numbers was 30-fold (table 1, figure 1). It is apparent that there is an extensive mean loss in mass for the individual organism during this period of rapid growth. However, the yield of antigen, as judged