Pseudomonas fluorescens MFE01 delivers a putative type VI secretion amidase that confers biocontrol against the soft-rot pathogen Pectobacterium atrosepticum.

The type VI secretion system (T6SS) is a contractile nanomachine widespread in Gram-negative bacteria. The T6SS injects effectors into target cells including eukaryotic hosts and competitor microbial cells and thus participates in pathogenesis and intermicrobial competition. Pseudomonas fluorescens MFE01 possesses a single T6SS gene cluster that confers biocontrol properties by protecting potato tubers against the phytopathogen Pectobacterium atrosepticum (Pca). Here, we demonstrate that a functional T6SS is essential to protect potato tuber by reducing the pectobacteria population. Fluorescence microscopy experiments showed that MFE01 displays an aggressive behaviour with an offensive T6SS characterized by continuous and intense T6SS firing activity. Interestingly, we observed that T6SS firing is correlated with rounding of Pectobacterium cells, suggesting delivery of a potent cell wall targeting effector. Mutagenesis coupled with functional assays then revealed that a putative T6SS secreted amidase, Tae3Pf , is mainly responsible for MFE01 toxicity towards Pca. Further studies finally demonstrated that Tae3Pf is toxic when produced in the periplasm, and that its toxicity is counteracted by the Tai3Pf inner membrane immunity protein.

Pseudomonas fluorescens MFE01 uses 1-undecene as aerial communication molecule.

Bacterial communication is a fundamental process used to synchronize gene expression and collective behavior among the bacterial population. The most studied bacterial communication system is quorum sensing, a cell density system, in which the concentration of inductors increases to a threshold level allowing detection by specific receptors. As a result, bacteria can change their behavior in a coordinated way. While in Pseudomonas quorum sensing based on the synthesis of N-acyl homoserine lactone molecules is well studied, volatile organic compounds, although considered to be communication signals in the rhizosphere, are understudied. The Pseudomonas fluorescens MFE01 strain has a very active type six secretion system that can kill some competitive bacteria. Furthermore, MFE01 emits numerous volatile organic compounds, including 1-undecene, which contributes to the aerial inhibition of Legionella pneumophila growth. Finally, MFE01 appears to be deprived of N-acyl homoserine lactone synthase. The main objective of this study was to explore the role of 1-undecene in the communication of MFE01. We constructed a mutant affected in undA gene encoding the enzyme responsible for 1-undecene synthesis to provide further insight into the role of 1-undecene in MFE01. First, we studied the impacts of this mutation both on volatile organic compounds emission, using headspace solid-phase microextraction combined with gas chromatography-mass spectrometry and on L. pneumophila long-range inhibition. Then, we analyzed influence of 1-undecene on MFE01 coordinated phenotypes, including type six secretion system activity and biofilm formation. Next, to test the ability of MFE01 to synthesize N-acyl homoserine lactones in our conditions, we investigated in silico the presence of corresponding genes across the MFE01 genome and we exposed its biofilms to an N-acyl homoserine lactone-degrading enzyme. Finally, we examined the effects of 1-undecene emission on MFE01 biofilm maturation and aerial communication using an original experimental set-up. This study demonstrated that the ΔundA mutant is impaired in biofilm maturation. An exposure of the ΔundA mutant to the volatile compounds emitted by MFE01 during the biofilm development restored the biofilm maturation process. These findings indicate that P. fluorescens MFE01 uses 1-undecene emission for aerial communication, reporting for the first time this volatile organic compound as bacterial intraspecific communication signal.