Mutagenicity testing in Salmonella typhimurium strains possessing both the His reversion and Ara forward mutation systems and different levels of classical nitroreductase or O-acetyltransferase activities.

Abstract

The induction of forward mutations to L-arabinose resistance (AraR) and of reversions to histidine prototrophy (His+) can be quantitatively compared in Salmonella typhimurium BA strains. The BA bacteria carry the araD531 allele required for the Ara assay and a his auxotrophy (hisD3052 or hisG46) required for the His assay. In this study, 2 new sets of BA indicator strains have been constructed in order to combine the Ara forward and the His reverse mutation assays of S. typhimurium with deficiency, or over-production, in either classical nitroreductase or O-acetyltransferase for mutagenicity testing of nitro-containing chemicals. Nine mutagens with different chemical structures were tested to compare the specific mutagenic sensitivities of the new constructions with those of the parental and of the conventional TA indicator bacteria. The Ara test, which responded with high sensitivity to all chemicals tested, revealed important differences between the standard tester strains TA98 and TA100 with respect to the activation of mutagens considered to be dependent on classical nitroreductase activity. Total correspondence was found between the specific mutagenic sensitivities of the defective and the overproducing bacteria in the genetic background of TA98 but not in that of TA100. In the genetic background of TA100, chemicals such as nitrofurantoin and nitrofurazone displayed 10-fold reduced mutagenicity to the "classical nitroreductase" defective strain without increasing mutagenicity to the corresponding overproducing bacteria. This discrepancy might be attributed to the greater nitroreduction capability of strain TA100 (68.12 nmole/min/mg protein) as compared to TA98 (24.42 nmole/min/mg protein), by assuming that nitrofurantoin and nitrofurazone are such good substrates for classical nitroreductase that the additional enzyme activity produced from the corresponding overexpressing plasmid when present in TA100 no longer affected their metabolic activation. We propose that the Ara forward mutation test carried out in the set of over-producing bacteria constructed in the genetic background of TA98 might play a role for routine testing of large number of samples. The isogenic defective strains could be used in cases of uncertain results with the corresponding over-producing bacteria. Finally, the reversion of the his alleles accompanying the Ara assay in the BA strains could play a role in assessing the presence of mixtures of chemicals with different mutagenic specificity in samples of environmental relevance such as urban air, foods, and water.