Gene amplification contributes to sulfonamide resistance in Escherichia coli

Abstract

A sulfathiazole-resistant strain of Escherichia coli was isolated and shown to contain a fourfold tandemly amplified segment of DNA 18 kilobase pairs in length in addition to a mutationally altered dihydropteroate synthase, the target enzyme for sulfonamide inhibition. The amplified DNA contained a gene designated sur that contributed to sulfathiazole resistance when present in greater amounts than those in the wild type. Sulfathiazole resistance was markedly decreased upon loss of the amplified DNA after nonselective growth. Plasmids that contained sur also conferred only weak sulfathiazole resistance on wild-type strains. Comparison of the restriction maps of the amplified DNA, wild-type DNA, and sur-containing plasmids showed that a DNA rearrangement occurred before or concomitant with the DNA amplification event. The DNA rearrangement resulted from an IS5 insertion, which, in conjunction with an IS5 element residing near sur in the wild-type strain, resulted in an -IS5-sur-IS5- configuration. Homologous recombination could account for duplication and subsequent amplification of the sur region. High-copy-number plasmids containing the sur locus did not express a sulfathiazole-resistant dihydropteroate synthase, nor did they overexpress wild-type dihydropteroate synthase. These data suggest that the high level of sulfathiazole resistance in this strain results from a synergistic effect of two different mutations.