dPseudomonas protegens strain Pf-5 is a rhizosphere bacterium that suppresses soilborne plant diseases and produces at least seven different secondary metabolites with antifungal properties. We derived mutants of Pf-5 with single and multiple mutations in biosynthesis genes for seven antifungal metabolites: 2,4-diacetylphoroglucinol (DAPG), pyrrolnitrin, pyoluteorin, hydrogen cyanide, rhizoxin, orfamide A, and toxoflavin. These mutants were tested for inhibition of the pathogens Fusarium verticillioides and Fusarium oxysporum f. sp. pisi. Rhizoxin, pyrrolnitrin, and DAPG were found to be primarily responsible for fungal antagonism by Pf-5. Previously, other workers showed that the mycotoxin fusaric acid, which is produced by many Fusarium species, including F. verticillioides, inhibited the production of DAPG by Pseudomonas spp. In this study, amendment of culture media with fusaric acid decreased DAPG production, increased pyoluteorin production, and had no consistent influence on pyrrolnitrin or orfamide A production by Pf-5. Fusaric acid also altered the transcription of biosynthetic genes, indicating that the mycotoxin influenced antibiotic production by Pf-5 at the transcriptional level. Addition of fusaric acid to the culture medium reduced antibiosis of F. verticillioides by Pf-5 and derivative strains that produce DAPG but had no effect on antibiosis by Pf-5 derivatives that suppressed F. verticillioides due to pyrrolnitrin or rhizoxin production. Our results demonstrated the importance of three compounds, rhizoxin, pyrrolnitrin, and DAPG, in suppression of Fusarium spp. by Pf-5 and confirmed that an interspecies signaling system mediated by fusaric acid had parallel effects on antifungal metabolite production and antibiosis by the bacterial biological control organism.
Pseudomonas is a heterogeneous genus of Gammaproteobacteria composed of species with diverse ecological roles; certain strains are members of the plant microbiome contributing to plant growth and health. Our studies focus on the soil bacterium Pseudomonas protegens Pf-5, which is known for its capacity to suppress plant diseases and produce a large spectrum of metabolites with antibiotic activity (1, 2). The antibiotics produced by Pf-5 include pyrrolnitrin (3), pyoluteorin (4), analogs of rhizoxin (5, 6), hydrogen cyanide (7), 2,4-diacetylphloroglucinol (DAPG) (8), monoacetylphloroglucinol (MAPG) (9) (an intermediate in the DAPG biosynthetic pathway [10,11]), the lipopeptide orfamide A (12), and toxoflavin (13). The antibiotics produced by Pf-5 have distinct but overlapping activity spectra, and it is likely that multiple compounds contribute to the interactions of Pf-5 with target plant pathogens that result in disease suppression. Commonly, the role of an antibiotic in biological control is assessed by comparing the level of disease suppression provided by a wild-type strain versus an antibiotic-deficient mutant (14). For strain Pf-5, however, this approach has not been highly successful in providing evidence for the roles of antibioti...