GROWTH OF GROUP A HEMOLYTIC STREPTOCOCCUS IN THE STEADY STATE

Abstract

The study in static cultures of the synthetic functions of bacteria and their growth is subject to serious limitations. These arise from the inability to specify and to insure constancy of various important cultural conditions such as growth rate, concentration of limiting metabolite and the concentrations of non-limiting metabolites when these are not present in large excess. In the case of acid-forming bacteria maintenance of a constant pH also constitutes a difficulty. This can be readily surmounted by the use of a pH-stat (Jacobsen and Leonis, 1951) but not without alteration of the ionic composition of the culture. The changing environment in static culture also limits the extent to which information derived in this way can be related to the metabolic activities of a microorganism maintained in the less variable environment of an infected host. The general solution to the limitations of static cultures is to be found in experimental arrangements which allow growth to proceed under steady-state conditions. A few years ago Monod (1950) and Novick and Szilard (1950) demonstrated the efficacy of continuous cultures (continuous dilution at constant volume) as a means of maintaining steady-state growth under controlled conditions. These authors described experimental devices for this purpose and considered some of the theoretical aspects of continuous cultures. They showed that it is possible to maintain a bacterial culture in a steady state in which the bacterial density and composition of medium remain constant as well as the rate of growth. Monod, particularly, emphasised the decisive advantages which steady-state kinetics provides for the investigation of various problems of bacterial physiology. Reference may be made to a recent review by Novick (1955) for a de1 This investigation was supported by a research grant (H-869) from the National Heart Institute of the National Institutes of Health, Public Health Service. scription of other devices used to obtain steadystate cultures. Since we are concerned with synthetic proceses associated with the growth of the hemolytic streptococcus, especially the formation of proteins, it appeared that our investigations would be greatly facilitated if they were carried out with steady-state cultures. We have therefore applied the continuous-culture technique to this organism and found it possible to achieve steadystate growth under a variety of conditions and over a wide range of rate of growth. In this paper we describe the experimental arrangement, the composition of the medium and the operating procedures which were developed for our purpose. Several of the factors which determine the steady-state conditions as well as the effects of their variation will be illustrated by our experimental results.