Interaction between 2,4-Diacetylphloroglucinol- and Hydrogen Cyanide-Producing Pseudomonas brassicacearum LBUM300 and Clavibacter michiganensis subsp. michiganensis in the Tomato Rhizosphere

We have previously demonstrated that inoculation of tomato plants with 2,4-diacetylphloroglucinol (DAPG) and hydrogen cyanide (HCN) producing Pseudomonas brassicacearum LBUM300 could significantly reduce bacterial canker symptoms caused by Clavibacter michiganensis subsp. michiganensis ( Cmm ). In this study, in order to better characterize the population dynamics of LBUM300 in the rhizosphere of tomato plants, we characterized the role played by DAPG and HCN production by LBUM300 on rhizosphere colonization of healthy and Cmm -infected tomato plants. The impact of Cmm presence on the expression of DAPG and HCN biosynthetic genes in the rhizosphere was also examined. In planta assays were performed using combinations of Cmm and/or wild type LBUM300, DAPG- ( phlD - ) or HCN - ( hcnC - ) isogenic mutant stains. Populations of LBUM300 and phlD and hcnC gene expression were quantified in rhizosphere soil at several time points up to 264 h post-inoculation using culture-independent qPCR and RT-qPCR TaqMan assays, respectively. The presence of Cmm significantly increased rhizospheric populations of LBUM300. In Cmm -infected tomato rhizospheres, the populations of wild type LBUM300 and HCN- mutant, both producing DAPG, were significantly higher than the population of the DAPG- mutant. A significant up-regulation of phlD expression was observed in the presence of Cmm while hcnC expression was only slightly increased in the DAPG- mutant strain when Cmm was present. Additionally, biofilm production was found to be significantly reduced in the DAPG- mutant as compared to the wild type and the HCN- mutant. IMPORTANCE The results suggest that Cmm -infection of tomato plants contributes to increasing rhizospheric populations of LBUM300, a biocontrol agent, as well as overexpressing the DAPG biosynthetic operon in this bacterium. The increasing rhizospheric populations of LBUM300 represents one of the key factors in controlling Cmm in tomato plants, as DAPG-producing bacteria have shown the ability to decrease bacterial canker symptoms in tomato plants.