Denaturing gradient gel electrophoretic analysis of dominant 2,4-diacetylphloroglucinol biosynthetic phlD alleles in fluorescent Pseudomonas from soils suppressive or conducive to black root rot of tobacco

In Switzerland, similar types of rhizosphere pseudomonads producing the biocontrol compound 2,4-diacetylphloroglucinol (Phl) have been found in soils suppressive to Thielaviopsis basicola-mediated black root rot of tobacco as well as in conducive soils. However, most findings were based on the analysis of a limited number of Pseudomonas isolates, obtained from a single experiment and only from T. basicola-inoculated plants. Here, an approach based on denaturing gradient gel electrophoresis (DGGE) of dominant phlD alleles from tobacco rhizosphere provided different phlD migration patterns. Sequencing of phlD-DGGE bands revealed a novel phylogenetic cluster of phlD sequences found in both suppressive and conducive soils in addition to previously-documented phlD alleles. phlD-DGGE bands and alleles differed little from one plant to the next but more extensively from one sampling to the next during the three-year study. Three of the 13 bands and 12 of the 31 alleles were only found in suppressive soil, whereas five bands and 13 alleles were found exclusively in conducive soil. The population structure of phlD+ pseudomonads depended more on the individual soil considered and its suppressiveness status than on inoculation of tobacco with T. basicola. In conclusion, phlD-DGGE revealed additional phlD diversity compared with earlier analyses of individual Pseudomonas isolates, and showed differences in phlD+Pseudomonas population structure in relation to disease suppressiveness.