Effects of medium composition on penicillin-induced hydrolysis and loss of RNA and culture turbidity in group A streptococci

Abstract

Exposure to penicillin G of exponentially growing cultures of group A streptococci growing in chemically defined medium (CDM) can lead to extensive loss of culture turbidity. Significant reductions in culture turbidity did not accompany comparable treatments of group A streptococci growing in Todd-Hewitt broth (THB). Studies with THB and a high-molecular-weight (>12,000) fraction of THB demonstrated that components in this complex medium inhibited the efflux of RNA hydrolysis products from otherwise intact cells. Hydrolysis products accumulated intracellularly and inhibited the extensive hydrolysis of RNA and consequently the loss of culture turbidity. Results of survival studies with cultures of group A streptococci exposed to penicillin G in THB demonstrated that this treatment protocol produces conditions of phenotypic tolerance relative to exposure in CDM. In combination, these findings provide further support for the hypothesis of RNA hydrolysis as the bactericidal mechanism of penicillin G action in this nonlytic death phenotype.Exposure of group A streptococci to antibiotics that inhibit cell wall synthesis leads to loss of viability without cellular lysis (nonlytic death) (3, 4, 10). The term nonlytic death originated with the observation that losses of viability were not accompanied by reductions in cell numbers or loss of culture turbidity. This phenomenon has been described for both gram-negative enteric bacilli (1) and group A streptococci (3, 4). However, in contrast to the previous studies with group A streptococci (3, 4), we observed significant reductions in culture turbidity and viability that were not associated with reductions in cell number (11). Results of radiolabeling studies demonstrated that these losses of culture turbidity were a direct reflection of penicillin G-induced destruction of RNA (11).In this report, we present results of experiments demonstrating that medium composition dramatically affects penicillin G-induced losses of turbidity and viability in cultures of group A streptococci. In addition, we show that this effect is associated with the presence of high-molecular-weight components in the medium, which inhibits the release of RNA hydrolysis products from cells and consequently inhibits the extent of penicillin G-induced hydrolysis of RNA.(A preliminary report of this work has been presented