Polymorphisms within the prnD and pltC genes from pyrrolnitrin and pyoluteorin-producing Pseudomonas and Burkholderia spp.

Abstract

Pyrrolnitrin (PRN) and pyoluteorin (PLT) are broad-spectrum antibiotics produced by several strains of Pseudomonas and Burkholderia species. Both antibiotics play an important role in the suppression of multiple plant pathogenic fungi. Primers were developed from conserved sequences and amplified prnD and pltC fragments from 18 Pseudomonas and four Burkholderia spp. of worldwide origin that produce either PRN or PLT or both. Subsequent RFLP (restriction fragment length polymorphisms) analysis of the 438-bp pltC fragment showed no polymorphisms among PLT-producing Pseudomonas strains. Polymorphisms within the 786-bp prnD fragment, however, allowed the assessment of the diversity among PRN-producing Pseudomonas and Burkholderia spp. to a level similar to that obtained by three 10-mer primers in random amplified polymorphic DNA analysis. Phylogenetic analysis of 16S rDNA sequences of strains representative of PRN-producing Pseudomonas and Burkholderia species correlated well with their taxonomic status. Phylogenetic relationships inferred from each of the four prn genes and from the complete sequence of the prn biosynthetic locus were similar to 16S rDNA-based phylogeny for most strains, except for Burkholderia pyrrocinia DSM 10685. Both RFLP analysis and comparison of the prn gene sequences showed that B. pyrrocinia DSM 10685 was more closely related to PRN-producing Pseudomonas strains, suggesting that lateral gene transfer may have occurred. Colony hybridization and PCR with PRN- and PLT-specific probes and primers showed that Pseudomonas and Burkholderia spp. harboring the prnD and pltC gene were not present at detectable levels on roots of wheat grown in five agricultural soils collected in The Netherlands, two of them being naturally suppressive to Gaeumannomyces graminis var. tritici. These results suggest that PRN- and PLT-producing Pseudomonas and Burkholderia sp. do not contribute to the natural suppressiveness found in these Dutch take-all decline soils.

Effect of Organic Management of Soils on Suppressiveness to Gaeumannomyces graminis var. tritici and its Antagonist, Pseudomonas fluorescens

“…Pseudomonas fluorescens (Pf32-gfp) Pseudomonas fluorescens strain 32 was isolated from roots of wheat growing in a Dutch take-all suppressive soil where wheat had been grown in monoculture for 27 years (Souza et al, 2003). This strain effectively controls Ggt and produces 2,4-DAPG.…”

The Production, Separation and Stability of Pyoluteorin: A Biological Pesticide

“…The limit of detection (LOD) of this method was 0.107 g/mL, by setting the signal-to-noise ratio at 3:1, and the limit of quantification (LOQ) for Plt was 0.36 g/mL by setting the signal-to-noise ratio at 10:1. The LOD of our method was much lower than that of HPLC (10 g/mL) (de Souza & Raaijmakers, 2003) and CEZ (0.66 g/mL) . This result suggests that this method can meet with the requirement of pesticide residue analysis.…”

“…As a potential biological pesticide, understanding the stability of Plt under different environmental conditions as well as its residue in soil after incorporation into the environment was great important for its commercial use. Some analytical methods of Plt from the fermentation broth based on HPLC have been reported (de Souza & Raaijmakers, 2003;La Fuente et al, 2004). Kim et al described a method for quantification analysis of Plt in fermentation broth with liquid chromatography-mass spectrometric (LC-MS) (Kim et al, 2003).…”

Screening of Rhizobacteria for Their Plant Growth Promotion Ability and Antagonism Against Damping off and Root Rot Diseases of Broad Bean (Vicia faba L.)

“…Genomic DNA of each of the fifteen promising isolates along with the five reference strains (from MTCC, IMTECH, Chandigarh) were isolated and subjected to RAPD analysis by PCR-RAPD methods [23], using seven oligonucleotide primers (OPA-1, 2, 4, 18, 20, OPD-2 and 5), separately. The reactions were carried out in final volume of 25 ll containing 19 PCR buffer, 1.5 mM MgCl2, 800 lM dNTPs and 1.0-2.0 units Taq DNA polymerase.…”