The antibiotics phenazine-1-carboxylic acid (PCA) and 2,4-diacetylphloroglucinol (Phl) are major determinants of biological control of soilborne plant pathogens by various strains of fluorescent Pseudomonas spp. In this study, we described primers and probes that enable specific and efficient detection of a wide variety of fluorescent Pseudomonas strains that produce various phenazine antibiotics or Phl. PCR analysis and Southern hybridization demonstrated that specific genes within the biosynthetic loci for Phl and PCA are conserved among various Pseudomonas strains of worldwide origin. The frequency of Phl-and PCA-producing fluorescent pseudomonads was determined on roots of wheat grown in three soils suppressive to take-all disease of wheat and four soils conducive to take-all by colony hybridization followed by PCR. Phenazine-producing strains were not detected on roots from any of the soils. However, Phl-producing fluorescent pseudomonads were isolated from all three take-all-suppressive soils at densities ranging from approximately 5 ؋ 10 5 to 2 ؋ 10 6 CFU per g of root. In the complementary conducive soils, Phl-producing pseudomonads were not detected or were detected at densities at least 40-fold lower than those in the suppressive soils. We speculate that fluorescent Pseudomonas spp. that produce Phl play an important role in the natural suppressiveness of these soils to take-all disease of wheat.

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.

Microbiota of crop ancestors may offer a way to enhance sustainable food production

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