Mutation spectra of chemical mutagens determined by Lac + reversion assay with Escherichia coli WP3101P–WP3106P tester strains

Abstract

We previously reported the development of mutation-specific Escherichia coli B tester strains WP3101 to WP3106 from strain WP2 uvrA. In this study we constructed their pKM101-containing derivatives WP3101P to WP3106P, and further isolated their rfa derivatives WP4101–WP4106 and WP4101P–WP4106P. The six kinds of F′ plasmids ( lacI −, lacZ − , proAB +), each of which carries a different lacZ allele, contained in the above strains were originally derived from E. coli K-12 strains CC101–CC106. All the tester strains show Lac − and Trp − phenotype. Assays for transitions and transversions are based upon Lac + reversion of a specific mutation located within the lacZ gene on an F′ plasmid. The trpE65(ochre) allele in the same strains enables them to be used for Trp + reversion assays as well. In the present paper, we evaluated the sensitivity, specificity, and usefulness of the newly developed tester strains. Strains WP3101P–WP3106P were highly sensitive to determine mutational profile of heterocyclic amines with S9 mix-mediated metabolic activation and most of the oxidative mutagens and free radical generators tested. Every type of base-pair substitutions induced by 2-amino-3,4-dimethylimidazo[4,5- f]quinoline (MeIQ) or 5-diazouracil were detected in strains WP3101P–WP3106P, while A:T→C:G and G:C→A:T mutations induced by MeIQ, and A:T→C:G, G:C→A:T, and G:C→C:G by 5-diazouracil were not detected in pKM101-free tester strains. In pKM101-carrying strains, cumene hydroperoxide induced all types of base substitutions, while formaldehyde preferentially induced G:C→T:A transversions. Phenazine methosulfate induced predominantly G:C→A:T transitions and G:C→T:A transversions, while H 2O 2 induced predominantly G:C→T:A and A:T→T:A transversions. Introduction of the rfa mutation considerably enhanced sensitivity to bulky mutagens such as polycyclic aromatic compounds. All six possible base substitutions induced by 9,10-dimethyl-1,2-benzanthracene (DMBA) were detected in tester strains WP4101P–WP4106P. In conclusion, our tester strains WP3101P–WP3106P and WP4101P–WP4106P permitted rapid and simple detection of specific mutations induced by variety of mutagens.