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Root inoculation of Arabidopsis thaliana ecotype Columbia with Pseudomonas fluorescens CHA0r partially protected leaves from the oomycete Peronospora parasitica. The molecular determinants of Pseudomonas fluorescens CHA0r for this induced systemic resistance (ISR) were investigated, using mutants derived from strain CHA0: CHA400 (pyoverdine deficient), CHA805 (exoprotease deficient), CHA77 (HCN deficient), CHA660 (pyoluteorin deficient), CHA631 (2,4-diacetylphloroglucinol [DAPG] deficient), and CHA89 (HCN, DAPG- and pyoluteorin deficient). Only mutations interfering with DAPG production led to a significant decrease in ISR to Peronospora parasitica. Thus, DAPG production in Pseudomonas fluorescens is required for the induction of ISR to Peronospora parasitica. DAPG is known for its antibiotic activity; however, our data indicate that one action of DAPG could be due to an effect on the physiology of the plant. DAPG at 10 to 100 microM applied to roots of Arabidopsis mimicked the ISR effect. CHA0r-mediated ISR was also tested in various Arabidopsis mutants and transgenic plants: NahG (transgenic line degrading salicylic acid [SA]), sid2-1 (nonproducing SA), npr1-1 (non-expressing NPR1 protein), jar1-1 (insensitive to jasmonic acid and methyl jasmonic acid), ein2-1 (insensitive to ethylene), etr1-1 (insensitive to ethylene), eir1-1 (insensitive to ethylene in roots), and pad2-1 (phytoalexin deficient). Only jar1-1, eir1-1, and npr1-1 mutants were unable to undergo ISR. Sensitivity to jasmonic acid and functional NPR1 and EIR1 proteins were required for full expression of CHA0r-mediated ISR. The requirements for ISR observed in this study in Peronospora parasitica induced by Pseudomonas fluorescens CHA0r only partially overlap with those published so far for Peronospora parasitica, indicating a great degree of flexibility in the molecular processes leading to ISR.