Nitric oxide synthase expression in Pseudomonas koreensis MME3 improves plant growth promotion traits.

The development of novel biotechnologies that promote a better use of N to optimize crop yield is a central goal for sustainable agriculture. Phytostimulation, biofertilization, and bioprotection through the use of bio-inputs are promising technologies for this purpose. In this study, the plant growth-promoting rhizobacteria Pseudomonas koreensis MME3 was genetically modified to express a nitric oxide synthase of Synechococcus SyNOS, an atypical enzyme with a globin domain that converts nitric oxide to nitrate. A cassette for constitutive expression of synos was introduced as a single insertion into the genome of P. koreensis MME3 using a miniTn7 system. The resulting recombinant strain MME3:SyNOS showed improved growth, motility, and biofilm formation. The impact of MME3:SyNOS inoculation on Brachypodium distachyon growth and N uptake and use efficiencies under different N availability situations was analyzed, in comparison to the control strain MME3:c. After 35 days of inoculation, plants treated with MME3:SyNOS had a higher root dry weight, both under semi-hydroponic and greenhouse conditions. At harvest, both MME3:SyNOS and MME3:c increased N uptake and use efficiency of plants grown under low N soil. Our results indicate that synos expression is a valid strategy to boost the phytostimulatory capacity of plant-associated bacteria and improve the adaptability of plants to N deficiency. KEY POINTS: • synos expression improves P. koreensis MME3 traits important for rhizospheric colonization • B. distachyon inoculated with MME3:SyNOS shows improved root growth • MME3 inoculation improves plant N uptake and use efficiencies in N-deficient soil.

Signs of a phyllospheric lifestyle in the genome of the stress-tolerant strain Azospirillum brasilense Az19.

Azospirillum brasilense Az19 is a plant-beneficial bacterium capable of protecting plants from the negative effects of drought. The objective of this study was to determine and analyze the genomic sequence of strain Az19 as a means of identifying putative stress-adaptation mechanisms. A high-quality draft genome of ca. 7 Mb with a predicted coding potential of 6710 genes was obtained. Phylogenomic analyses confirmed that Az19 belongs to the brasilense clade and is closely related to strains Az39 and REC3. Functional genomics revealed that the denitrification pathway of Az19 is incomplete, which was in agreement with a reduced growth on nitrate under low O2 concentrations. Putative genes of the general stress response and oxidative stress-tolerance, as well as synthesis of exopolysaccharides, carotenoids, polyamines and several osmolytes, were detected. An additional poly-beta-hydroxybutyrate (PHB) synthase coding gene was found in Az19 genome, but the accumulation of PHB did not increase under salinity. The detection of exclusive genes related to DNA repair led to discover that strain Az19 also has improved UV-tolerance, both in vitro and in planta. Finally, the analysis revealed the presence of multiple kaiC-like genes, which could be involved in stress-tolerance and, possibly, light responsiveness. Although A. brasilense has been a model for the study of beneficial plant-associated rhizobacteria, the evidence collected in this current study suggests, for the first time in this bacterial group, an unexpected possibility of adaptation to the phyllosphere.