Isolation of ant mutants of Pseudomonas fluorescens strain Pf7-14 altered in antibiotic production, cloning of ant(+) DNA, and evaluation of the role of antibiotic production in the control of blast and sheath blight of rice

Pseudomonas fluorescens strain 7-14 (Pf7-14) isolated from the rice rhizosphere produces antibiotic(s) active against Pyricularia sp., Rhizoctonia solani, Pythium ultimum, and Gaeumannomyces graminis var. tritici. Mutagenesis of Pf714 with ethyl methane sulfonate (EMS) or transposon (mini-Tn5-Km) yielded antibiotic-defective mutants belonging to the following classes. Class I mutants, generated by EMS or mini-Tn5-Km, are totally deficient in antifungal antibiotics, i.e., are Ant-. Class II mutants were recovered following mutagenesis with mini-Tn5-Km; they produce reduced levels of antibiotics, i.e., are Ant(leaky). Class III mutants overproduce antibiotics; they were generated by EMS mutagenesis. Insertions of mini-Tn5-Km into at least four different chromosomal sites caused defects in antibiotic production. To further analyze ant genes affecting antibiotic production, we isolated Ant(+) plasmids from a genomic library of the strain Pf714. Of the several plasmids that appeared to carry ant loci, pAKC908 was unusual in that it restored antibiotic production in all of our Class I and Class II mutants. By mutagenesis of pAKC908 with Tn3HoHo1 we localized a DNA region required for antibiotic production. Southern hybridization data and the complementation pattern of the Ant(+) plasmids indicated clustering of the ant genes in Pf714. Field trials with an Ant-deficient mutant carrying a single-site mini-Tn5-Km insertion revealed that antibiotic production by Pf714 was required for the control of blast as well as sheath blight. The data also show that Pf714 was as effective as the fungicide tricyclazole in controlling these rice diseases. (C) 1996 Academic Press, Inc.