Rhizoxin analogs, orfamide A and chitinase production contribute to the toxicity of Pseudomonas protegens strain Pf‐5 to Drosophila melanogaster

Abstract

Pseudomonas protegens strain Pf-5 is a soil bacterium that was first described for its capacity to suppress plant diseases and has since been shown to be lethal to certain insects. Among these is the common fruit fly Drosophila melanogaster, a well-established model organism for studies evaluating the molecular and cellular basis of the immune response to bacterial challenge. Pf-5 produces the insect toxin FitD, but a ΔfitD mutant of Pf-5 retained full toxicity against D. melanogaster in a noninvasive feeding assay, indicating that FitD is not a major determinant of Pf-5's oral toxicity against this insect. Pf-5 also produces a broad spectrum of exoenzymes and natural products with antibiotic activity, whereas a mutant with a deletion in the global regulatory gene gacA produces none of these exoproducts and also lacks toxicity to D. melanogaster. In this study, we made use of a panel of Pf-5 mutants having single or multiple mutations in the biosynthetic gene clusters for seven natural products and two exoenzymes that are produced by the bacterium under the control of gacA. Our results demonstrate that the production of rhizoxin analogs, orfamide A, and chitinase are required for full oral toxicity of Pf-5 against D. melanogaster, with rhizoxins being the primary determinant.

Characterization of Toxin Complex Gene Clusters and Insect Toxicity of Bacteria Representing Four Subgroups of Pseudomonas fluorescens

“…Furthermore, evaluation of fitD mutants in this and a companion study [40] show that even a specific bacterial strain, such as P . protegens Pf-5, employs different lethality mechanisms on different insect hosts.…”

Novel mechanism of metabolic co-regulation coordinates the biosynthesis of secondary metabolites in Pseudomonas protegens

“…P. protegens can occupy different environments, including the soil, plant surfaces, and insects, and interact with different organisms including bacteria, fungi, protozoa, and insects (Brodhagen et al, 2004; Jousset et al, 2009; Kidarsa et al, 2013; Henkels et al, 2014; Loper et al, 2016). Both DAPG and pyoluteorin contribute to the antagonism of P. protegens against a range of bacteria, fungi, and oomycetes (Ohmori et al, 1978; Keel et al, 1992) and both facilitate the bacterium’s establishment in fungal tissues, such as basidiocarps of the button mushroom Agaricus spp.…”

Secondary Metabolism and Interspecific Competition Affect Accumulation of Spontaneous Mutants in the GacS-GacA Regulatory System in Pseudomonas protegens

“…We hypothesized that secondary metabolism contributes to the accumulation of spontaneous Gac Ϫ mutants. To test this hypothesis, we evaluated the accumulation of Gac Ϫ mutants of a Pf-5 derivative strain that contains mutations in hcnB, ofaA, phlA, pltA, prnC, and rzxB (referred to as the 6-fold mutant hereafter) (35). This 6-fold mutant does not produce any of the six secondary metabolites: hydrogen cyanide, orfamide A, 2,4-diacetylphloroglucinol, pyoluteorin, pyrrolnitrin, or rhizoxin (35).…”

“…To test this hypothesis, we evaluated the accumulation of Gac Ϫ mutants of a Pf-5 derivative strain that contains mutations in hcnB, ofaA, phlA, pltA, prnC, and rzxB (referred to as the 6-fold mutant hereafter) (35). This 6-fold mutant does not produce any of the six secondary metabolites: hydrogen cyanide, orfamide A, 2,4-diacetylphloroglucinol, pyoluteorin, pyrrolnitrin, or rhizoxin (35). After 6 days of growth, the proportion of Gac Ϫ mutants was significantly lower in cultures of this 6-fold mutant relative to wild-type Pf-5 (Fig.…”