RESUMO
Plant biostimulants have received attention as sustainable alternatives to chemical fertilizers. Extracellular polymeric substances (EPSs), among the compounds secreted by plant growth-promoting rhizobacteria (PGPRs), are assumed to alleviate abiotic stress. This study aims to investigate the effect of purified EPSs on rice under abiotic stress and analyze their mechanisms. A pot experiment was conducted to elucidate the effects of inoculating EPSs purified from PGPRs that increase biofilm production in the presence of sugar on rice growth in heat-stress conditions. Since all EPSs showed improvement in SPAD after the stress, Enterobacter ludwigii, which was not characterized as showing higher PGP bioactivities such as phytohormone production, nitrogen fixation, and phosphorus solubilization, was selected for further analysis. RNA extracted from the embryos of germinating seeds at 24 h post-treatment with EPSs or water was used for transcriptome analysis. The RNA-seq analysis revealed 215 differentially expressed genes (DEGs) identified in rice seeds, including 139 up-regulated and 76 down-regulated genes. A gene ontology (GO) enrichment analysis showed that the enriched GO terms are mainly associated with the ROS scavenging processes, detoxification pathways, and response to oxidative stress. For example, the expression of the gene encoding OsAAO5, which is known to function in detoxifying oxidative stress, was two times increased by EPS treatment. Moreover, EPS application improved SPAD and dry weights of shoot and root by 90%, 14%, and 27%, respectively, under drought stress and increased SPAD by 59% under salt stress. It indicates that bacterial EPSs improved plant growth under abiotic stresses. Based on our results, we consider that EPSs purified from Enterobacter ludwigii can be used to develop biostimulants for rice.
RESUMO
Chitinases are enzymes that can break down chitin, a major component of the exoskeleton of insects and fungi. This feature makes them potential biopesticides in agriculture since they are considered a safe and environmentally friendly alternative to synthetic pesticides. In this work, we performed a comparative study between two different bacterial expression strains to produce a recombinant chitinase with improved stability. Escherichia coli strains Origami B and BL21 (DE3) were selected for their distinct cytosolic environment to express BhChitA chitinase of Bacillus halodurans C-125 and to investigate the role of disulfide bond formation and proper folding on its stability and activity. Expression of the recombinant BhChitA in bacterial strain containing oxidative cytosol (Origami B) improved its activity and stability. Although both expression systems have comparable biochemical properties (temperature range 20-80 °C and pH spectrum 3-10), BhChitA expressed in Origami strain seems more stable than expressed in BL21. Furthermore, the optimal expression conditions of the recombinant BhChitA has been carried out at 30 °C during 6 h for the Origami strain, against 20 °C during 2 h for BL21. On the other hand, no significant differences were detected between the two enzymes when the effect of metal ions was tested. These findings correlate with the analysis of the overall structure of BhChitA. The model structure permitted to localize disulfide bond, which form a stable connection between the substrate-binding residues and the hydrophobic core. This link is required for efficient binding of the chitin insertion domain to the substrate. BhChitA exhibited in vitro antifungal effect against phytopathogenic fungi and suppressed necrosis of Botrytis cinerea on detached tomato leaves. In vitro assays showed the influence of BhChitA on growth suppression of Botrytis cinerea (53%) Aspergillus niger (65%), Fusarium graminearum (25%), and Fusarium oxysporum (34%). Our results highlight the importance of the bacterial expression system with oxidative cytosol in producing promising biopesticides that can be applied for post-harvest processing and crop protection.