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The fungus Ustilago maydis is a biotrophic pathogen of maize (Zea mays). In its genome we have identified an ortholog of YAP1 (for Yeast AP-1-like) from Saccharomyces cerevisae that regulates the oxidative stress response in this organism. yap1 mutants of U. maydis displayed higher sensitivity to H 2 O 2 than wild-type cells, and their virulence was significantly reduced. U. maydis yap1 could partially complement the H 2 O 2 sensitivity of a yap1 deletion mutant of S. cerevisiae, and a Yap1-green fluorescent protein fusion protein showed nuclear localization after H 2 O 2 treatment, suggesting that Yap1 in U. maydis functions as a redox sensor. Mutations in two Cys residues prevented accumulation in the nucleus, and the respective mutant strains showed the same virulence phenotype as Dyap1 mutants. Diamino benzidine staining revealed an accumulation of H 2 O 2 around yap1 mutant hyphae, which was absent in the wild type. Inhibition of the plant NADPH oxidase prevented this accumulation and restored virulence. During the infection, Yap1 showed nuclear localization after penetration up to 2 to 3 d after infection. Through array analysis, a large set of Yap1-regulated genes were identified and these included two peroxidase genes. Deletion mutants of these genes were attenuated in virulence. These results suggest that U. maydis is using its Yap1-controlled H 2 O 2 detoxification system for coping with early plant defense responses.

The plasmid pME6863, carrying the aiiA gene from the soil bacterium Bacillus sp. A24 that encodes a lactonase enzyme able to degrade N-acyl-homoserine lactones (AHLs), was introduced into the rhizosphere isolate Pseudomonas fluorescens P3. This strain is not an effective biological control agent against plant pathogens. The transformant P. fluorescens P3/pME6863 acquired the ability to degrade AHLs. In planta, P. fluorescens P3/pME6863 significantly reduced potato soft rot caused by Erwinia carotovora and crown gall of tomato caused by Agrobacterium tumefaciens to a similar level as Bacillus sp. A24. Little or no disease reduction was observed for the wild-type strain P3 carrying the vector plasmid without aiiA. Suppression of potato soft rot was observed even when the AHL-degrading P. fluorescens P3/pME6863 was applied to tubers 2 days after the pathogen, indicating that biocontrol was not only preventive but also curative. When antagonists were applied individually with the bacterial plant pathogens, biocontrol activity of the AHL degraders was greater than that observed with several Pseudomonas 2,4-diacetylphloroglucinol-producing strains and with Pseudomonas chlororaphis PCL1391, which relies on production of phenazine antibiotic for disease suppression. Phenazine production by this well characterized biological control strain P. chlororaphis PCL1391 is regulated by AHL-mediated quorum sensing. When P. chlororaphis PCL1391 was co-inoculated with P. fluorescens P3/pME6863 in a strain mixture, the AHL degrader interfered with the normally excellent ability of the antibiotic producer to suppress tomato vascular wilt caused by Fusarium oxysporum f. sp. lycopersici. Our results demonstrate AHL degradation as a novel biocontrol mechanism, but also demonstrate the potential for non-target interactions that can interfere with the biocontrol efficacy of other strains.

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Pseudomonas putida KT2442 is a root-colonizing strain which can use proline, one of the major components in root exudates, as its sole carbon and nitrogen source. A P. putida mutant unable to grow with proline as the sole carbon and nitrogen source was isolated after random mini-Tn5-Km mutagenesis. The mini-Tn5 insertion was located at the putA gene, which is adjacent to and divergent from the putP gene. The putA gene codes for a protein of 1,315 amino acid residues which is homologous to the PutA protein of Escherichia coli, Salmonella enterica serovar Typhimurium, Rhodobacter capsulatus, and several Rhizobium strains. The central part of P. putida PutA showed homology to the proline dehydrogenase of Saccharomyces cerevisiae and Drosophila melanogaster, whereas the C-terminal end was homologous to the pyrroline-5-carboxylate dehydrogenase of S. cerevisiae and a number of aldehyde dehydrogenases. This suggests that in P. putida, both enzymatic steps for proline conversion to glutamic acid are catalyzed by a single polypeptide. The putP gene was homologous to the putP genes of several prokaryotic microorganisms, and its gene product is an integral inner-membrane protein involved in the uptake of proline. The expression of both genes was induced by proline added in the culture medium and was regulated by PutA. In a P. putida putA-deficient background, expression of both putA and putP genes was maximal and proline independent. Corn root exudates collected during 7 days also strongly induced the P. putida put genes, as determined by using fusions of the put promoters to lacZ. The induction ratio for the putA promoter (about 20-fold) was 6-fold higher than the induction ratio for the putP promoter.

Pseudomonas putida BIRD-1 is a plant growth-promoting rhizobacterium whose genome size is 5.7 Mbp. It adheres to plant roots and colonizes the rhizosphere to high cell densities even in soils with low moisture. This property is linked to its ability to synthesize trehalose, since a mutant deficient in the synthesis of trehalose exhibited less tolerance to desiccation than the parental strain. The genome of BIRD-1 encodes a wide range of proteins that help it to deal with reactive oxygen stress generated in the plant rhizosphere. BIRD-1 plant growth-promoting rhizobacteria properties derive from its ability to enhance phosphorous and iron solubilization and to produce phytohormones. BIRD-1 is capable of solubilizing insoluble inorganic phosphate forms through acid production. The genome of BIRD-1 encodes at least five phosphatases related to phosphorous solubilization, one of them being a phytase that facilitates the utilization of phytic acid, the main storage form of phosphorous in plants. Pyoverdine is the siderophore produced by this strain, a mutant that in the FvpD siderophore synthase failed to grow on medium without supplementary iron, but the mutant was as competitive as the parental strain in soils because it captures the siderophores produced by other microbes. BIRD-1 overproduces indole-3-acetic acid through convergent pathways.

Pseudomonas putida are strict aerobes that proliferate in a range of temperate niches and are of interest for environmental applications due to their capacity to degrade pollutants and ability to promote plant growth. Furthermore solvent-tolerant strains are useful for biosynthesis of added-value chemicals. We present a comprehensive comparative analysis of nine strains and the first characterization of the Pseudomonas putida pangenome. The core genome of P. putida comprises approximately 3386 genes. The most abundant genes within the core genome are those that encode nutrient transporters. Other conserved genes include those for central carbon metabolism through the Entner-Doudoroff pathway, the pentose phosphate cycle, arginine and proline metabolism, and pathways for degradation of aromatic chemicals. Genes that encode transporters, enzymes and regulators for amino acid metabolism (synthesis and degradation) are all part of the core genome, as well as various electron transporters, which enable aerobic metabolism under different oxygen regimes. Within the core genome are 30 genes for flagella biosynthesis and 12 key genes for biofilm formation. Pseudomonas putida strains share 85% of the coding regions with Pseudomonas aeruginosa; however, in P. putida, virulence factors such as exotoxins and type III secretion systems are absent.