Escherichia coliK-12 Mutants Hyperproducing Chromosomal Beta-Lactamase by Gene Repetitions

Abstract

E. coli K-12 ampicillin-resistant mutants hyperproducing chromosomal .beta.-lactamase arose spontaneously from strains carrying ampA1 ampC+. Such mutants were found even in a recA background. Two Ampr-100 strains were analyzed genetically. The Ampr-100 resistance level of both strains could be transduced by direct selection for ampicillin resistance. Several classes of ampicillin-resistant transductants were found that differed from one another in the .beta.-lactamase activity and the ampicillin resistance mediated by an ampA1 ampC+-carrying strain. The data suggested that .beta.-lactamase hyperproduction was due to repetitions of the chromosomal amp genes. The size of the repeated region was calculated from cotransduction estimates, using the formula of Wu, and was about 1 min in 1 strain and 1.5 min in the other. Second-step Ampr-400 mutants were isolated from an Ampr-100 strain. The resistance of these mutants was apparently also due to repetitions, each mediating a resistance to about 10 .mu.g/ml. Mutants of wild-type strains that were moderately resistant to ampicillin also gave rise to intermediate-resistance classes, suggesting repetitions of the wild-type amp alleles. F'' factors hyperproducing chromosomal .beta.-lactamase by gene repetitions were constructed. They mediated levels of ampicillin resistance comparable to that of naturally occurring resistance plasmids. The expression of .beta.-lactamase hyperproduction was not affected by the presence of ampA and ampC alleles in trans and did not act in trans on the other alleles.