Drug-induced reversible and irreversible enzyme alterations underlying drug-resistance.

Abstract

p-Aminobenzoic acid-dependent M 48-34 strains ofEscherichia coligrown with and without sulphathiazole (ST) were investigated with respect to the enhanced capacities they acquire simultaneously for the synthesis of folic acid and resistance. The washed cells of the sensitive strain grown in drug-free media contain a small amount of intracellularly accumulated folic acid precursor(s). The precursors are converted by the washed cells into folic acid in the presence ofp-aminobenzoic acid (PABA) alone. The washed cells harvested from cultures containing a sub-inhibitory concentration of sulphathiazole contain a three-four times greater amount of intracellular folic acid precursor(s) which are converted into folic acid in the above manner. Following the utilization of the intracellular presursor(s) these cells on retesting manifest the capability of synthesizing folic acid from PABA and synthetic 2-amino- 4-hydroxy-6-carboxypteridine. This is a newly acquired character which is lacking in the cells grown in the absence of sulphathiazole. Higher intracellular folic acid-precursor accumula­tion and ability to synthesize folic acid from PABA and synthetic pteridine is possessed by the strains made resistant by successive transfers in the increasing concentrations of sulpha­thiazole. During these transfers, the amounts of sulphathiazole are so graded that the cells in the inocula are capable of surviving from 65 to 87·7% and therefore, the chances of selecting pre-existant resistant mutants, believed to occur at the rate of 10‒8to 10‒9, are reduced to a minimum or improbable level. The strain which is made resistant to 1·0μg ST by four successive transfers in graded amounts of sulphathiazole loses the acquired synthetic and resistance characters following three successive transfers in drug-free media, the strain made resistant to 15·0μg ST still possessed a fraction of the acquired abilities after twenty successive drug-free transfers, and the strain which was made resistant to 40μg ST did not show any decrease in the acquired characters after thirty successive drug-free transfers. It is to be noted that when the strain reversibly resistant to 1·0μg ST is grown in sulphathiazole and an antagonist concentration of PABA they fail to manifest the two characters. Similar concen­trations of PABA present during the growth of the resistant strains do not alter to any degree the acquired characters. Colonies of the parent sensitive cells appearing on agar plates con­taining various concentrations of sulphathiazole inoculated into drug-free media yield descendant cells which manifest the two characters, but lost them after two or three successive drug-free transfers, in a manner similar to 1·0μg ST-resistant cells discussed above.