The sporulation system of Bacillus subtilis as the basis of a multi-gene mutagen screening test

The sporulation system of B. subtilis provides the basis of a simple and unique test for the detection of forward mutations in any of several hundreds genes in 28--45 separate operons scattered throughout the chromosome. Non-sporulating or oligosporogenous mutant colonies are easily identified by their lack of a brown pigment normally present in spore-forming colonies. N-Methyl-N'-nitro-N-nitrosoguanidine (MNNG), ethyl methanesulfonate, acridine orange, acriflavin, nitrous acid, and UV irradiation are already known to produce sporulation mutants. This paper reports the dose dependence of sporulation mutant induction by 2-nitrosofluorene, ICR-191, nitrogen mustard, ethidium bromide and MNNG; mutagenesis is also demonstrated for aflatoxin B1 and 4-nitroquinoline-N-oxide. A mammalian liver enzyme metabolizing system was necessary for activation of aflatoxin B1. Auramine-O and 4-nitro-o-phenylenediamine failed to give a significant mutagenic effect under the conditions employed. The wide variety of mutagen classes detected indicates the general applicability of the test. This test, based on many genes throughout the chromosome, may prove less apt to exclude rare mutagenic "hot-spots" than systems based on the detection of mutations in a restricted region of the chromosome.