Aflatoxin and anthraquinone biosynthesis by nitrosoguanidine-derived mutants of Aspergillus parasiticus

Abstract

The mutagen N-methyl-N′-nitro-N-nitrosoguanidine (NG) was shown to produce mutants of Aspergillus parasiticus NRRL 2999 that have an altered secondary biosynthesis capacity and nutritional requirements. Incubation of A. parasiticus (1 × 108) spores in 0.02% nitrosoguanidine for 60 min resulted in induced nutritional auxotrophs to a maximum frequency of 8% at 5% survival. Some 0.5% of the 5% survivors lacked the ability to synthesize aflatoxin; however, no other fluorescent metabolites were detected from these mutants. Another group of mutants was isolated that gave a red anthraquinone, subsequently identified as norsolorinic acid (NOR) and representing up to 3–4% of the mycelial dry weight mass. The wild-type high aflatoxin-producing strain yielded a small quantity of this pigment; however, all the NOR mutants synthesized limited amounts of aflatoxin B1 or none at all. Further mutagenesis of NOR mutants resulted in survivors still with reduced aflatoxin elaboration ability. These mutants also gave small quantities of the methoxylated pigment, solorinic acid. The wild-type A. parasiticus NRRL 2999 strain yields 50 times more aflatoxin B1 than NOR pigment, whereas the derived NOR mutants synthesized 100 to 200 times more NOR than aflatoxin B1. The synthesis of NOR in the induced NOR mutants paralleled growth, whereas aflatoxin B1 production occurs typically as growth begins to decline (biphasic synthesis). Such data indicate a possible metabolic block of secondary biosynthesis in NOR mutants since both NOR and aflatoxin B1 are acetate-derived polyketide metabolites.