Monitoring of the relation between 2,4-diacetylphloroglucinol-producing Pseudomonas and Thielaviopsis basicola populations by real-time PCR in tobacco black root-rot suppressive and conducive soils

Abstract

Natural suppressiveness of Swiss soils to Thielaviopsis basicola-mediated tobacco black root rot is thought to depend mainly on fluorescent pseudomonads producing the antimicrobial compound 2,4-diacetylphloroglucinol. However, the relation between these phl+Pseudomonas populations and both the T. basicola population and disease suppressiveness in these soils is unknown, and real-time PCR tools were used to address this issue. Significant rhizosphere levels of phl+ pseudomonads had been evidenced before in suppressive as well as conducive soils, but this was done using culture-based approaches only. Here, a phlD-based real-time PCR method targeting all phlD+ genotypes, unlike the strain-specific real-time PCR methods available so far, was developed and validated (detection limit around 4 log cells g−1 soil and amplification efficiency >80%). When implemented on Swiss soils suppressive or conducive to black root rot, it clarified the hypothesis that suppressiveness does not require higher levels of phlD+ pseudomonads. The parallel assessment of T. basicola population by real-time PCR (method of Huang and Kang, 2010) suggested that suppressiveness was not due to the inability of the pathogen to colonize the rhizosphere and tobacco roots in suppressive soils, but rather that phl+ pseudomonads might act by limiting root penetration by the pathogen in suppressive soils. In conclusion, an effective real-time PCR method was achieved for phlD+ pseudomonads and can be used to monitor this key functional group in various environmental conditions, including here to better understand the ecology of suppressive soils.