Background: Gluconacetobacter diazotrophicus Pal5 is an endophytic diazotrophic bacterium that lives in association with sugarcane plants. It has important biotechnological features such as nitrogen fixation, plant growth promotion, sugar metabolism pathways, secretion of organic acids, synthesis of auxin and the occurrence of bacteriocins. Results: Gluconacetobacter diazotrophicus Pal5 is the third diazotrophic endophytic bacterium to be completely sequenced. Its genome is composed of a 3.9 Mb chromosome and 2 plasmids of 16.6 and 38.8 kb, respectively. We annotated 3,938 coding sequences which reveal several characteristics related to the endophytic lifestyle such as nitrogen fixation, plant growth promotion, sugar metabolism, transport systems, synthesis of auxin and the occurrence of bacteriocins. Genomic analysis identified a core component of 894 genes shared with phylogenetically related bacteria. Gene clusters for gum-like polysaccharide biosynthesis, tad pilus, quorum sensing, for modulation of plant growth by indole acetic acid and mechanisms involved in tolerance to acidic conditions were identified and may be related to the sugarcane endophytic and plant-growth promoting traits of G. diazotrophicus. An accessory component of at least 851 genes distributed in genome islands was identified, and was most likely acquired by horizontal gene transfer. This portion of the genome has likely contributed to adaptation to the plant habitat. Conclusion: The genome data offer an important resource of information that can be used to manipulate plant/bacterium interactions with the aim of improving sugarcane crop production and other biotechnological applications.

The genome of the endophytic diazotrophic bacterial species Gluconacetobacter diazotrophicus PAL5 (PAL5) revealed the presence of a gum gene cluster. In this study, the gumD gene homologue, which is predicted to be responsible for the first step in exopolysaccharide (EPS) production, was insertionally inactivated and the resultant mutant (MGD) was functionally studied. The mutant MGD presented normal growth and nitrogen (N₂) fixation levels but did not produce EPS when grown on different carbon sources. MGD presented altered colony morphology on soft agar plates (0.3% agar) and was defective in biofilm formation on glass wool. Most interestingly, MGD was defective in rice root surface attachment and in root surface and endophytic colonization. Genetic complementation reverted all mutant phenotypes. Also, the addition of EPS purified from culture supernatants of the wild-type strain PAL5 to the mutant MGD was effective in partially restoring wild-type biofilm formation and plant colonization. These data provide strong evidence that the PAL5 gumD gene is involved in EPS biosynthesis and that EPS biosynthesis is required for biofilm formation and plant colonization. To our knowledge, this is the first report of a role of EPS in the endophytic colonization of graminaceous plants by a nitrogen-fixing bacterium.

Gluconacetobacter diazotrophicus is an aerobic diazotrophic plant-growth-promoting bacterium isolated from different gramineous plants. We showed that reactive oxygen species (ROS) were produced at early stages of rice root colonization, a typical plant defense response against pathogens. The transcription of the pathogen-related-10 gene of the jasmonic acid (JA) pathway but not of the PR-1 gene of the salicylic acid pathway was activated by the endophytic colonization of rice roots by G. diazotrophicus strain PAL5. Quantitative polymerase chain reaction analyses showed that, at early stages of colonization, the bacteria upregulated the transcript levels of ROS-detoxifying genes such as superoxide dismutase (SOD) and glutathione reductase (GR). To proof the role of ROS-scavenging enzymes in the colonization and interaction process, transposon insertion mutants of the SOD and GR genes of strain PAL5 were constructed. The SOD and GR mutants were unable to efficiently colonize the roots, indicated by the decrease of tightly root-associated bacterial cell counts and endophytic colonization and by fluorescence in situ hybridization analysis. Interestingly, the mutants did not induce the PR-10 of the JA-pathway, probably due to the inability of endophytic colonization. Thus, ROS-scavenging enzymes of G. diazotrophicus strain PAL5 play an important role in the endophytic colonization of rice plants.

Background: Inoculation with Gluconacetobacter diazotrophicus has shown to influence root development in red rice plants, and more recently, the induced systemic tolerance (IST) response to drought was also demonstrated. The goal of this study was to evaluate the inoculation effect of G. diazotrophicus strain Pal5 on the amelioration of drought stress and root development in red rice (Oryza sativa L.). Methods: The experimental treatments consist of red rice plants inoculated with and without strain Pal5 in presence and absence of water restriction. Physiological, biochemical, and molecular analyses of plant roots were carried out, along with measurements of growth and biochemical components. Results: The plants showed a positive response to the bacterial inoculation, with root growth promotion and induction of tolerance to drought. An increase in the root area and higher levels of osmoprotectant solutes were observed in roots. Bacterial inoculation increased the drought tolerance and positively regulated certain root development genes against the water deficit in plants. Conclusion: G. diazotrophicus Pal5 strain inoculation favored red rice plants by promoting various root growth and developmental mechanisms against drought stress, enabling root development and improving biochemical composition.