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Description
Conventional practices of field vegetable cultivation, like intense irrigation and fertilization, generate environmental problems - soil erosion, nitrate leaching, plastic material residues. Plant biostimulants represent an emerging class of agricultural input, which protect plants against abiotic stress, enhance / benefits nutrients uptake and improve yield quality. The goal of the project is to develop (bio)technologies for the production and the integrated utilization of next generation plant biostimulants, for field vegetables grown into high residue farming systems.
The ultimate outcome of the project is an integrated agro-bio-nano-technology, which determines phytonutrients bio-enhancement and selenium biofortification on vegetables and produces more with less in the areas with selenium deficit in soil. Both Romania and Norway are among countries with selenium deficit in soils
The project supports research cooperation between Romania and Norway and consolidates a strategic partnership, because Romanian and Norway Partners involved in the project have already a significant history of cooperation (including 2 ERA.NET common projects).
Summary of project results
Romania and Norway are among the countries with selenium soil deficits. Due to its narrow physiological window, selenium biofortification (through plant treatment) is a better approach than direct supplementation.
Physiological window of selenium. The U-shape relationship between selenium and disease status limits selenium supplementation to subjects with lower than optimal selenium status. The selenium (bio) fortification should present a lower risk of supplementation on subjects with Se optimal status or with supra-optimal status.
Although it is not recognized as a micronutrient essential for plants, selenium increase plant tolerance to abiotic stress. The STIM 4+ project addressed the challenges related to the development of an integrated agro-bio-nano-technology that targets optimal exploitation of the agrotechnical benefits of this selenium biofortification in combination with the high-residues vegetable farming. The STIM 4+ integrated agro-bio-nano-technology determines phytonutrients bio-enhancement and selenium biofortification on vegetables, produces more with less in the area with selenium deficit in soil and strengthens the benefits of high-residues vegetable farming while compensating its drawbacks.
The STIM 4+ activities focused on developing the next-generation plant biostimulants and their integrated utilization in the high-residues farming system. The STIM 4+ project delivered the following plant biostimulants: (i) inorganic plant biostimulants (zerovalent nanoselenium), (ii) organic plant biostimulants (strigolactone mimic, embedded in formulations based on stimuli-responsive glycodynameric chitosan and microalgae extract) and (iii) microbial plant biostimulant (multifaceted Trichoderma strain, responding to strigolactone).
One of the technical problems that limit the practical application of selenium nanoparticles is the tendency of hydrophobic selenium nanoparticles to aggregate in aqueous suspensions. The stable biogenic SeNPs suspensions were produced by a biotechnological process with a microalgae yeast consortium. The selected yeast strains reduce selenium salt toxicity to microalgae and contribute to forming a SeNPs amphiphilic biocorona, which stabilizes SeNPs in aqueous (spraying) solutions. The microalgae yeast consortia respond to strigolactone mimics by accelerating growth and enhanced flocculation. The strigolactone mimic application for more efficient biogenic SeNPs production is a side-ground result included in the patented process. The preparation of the stable biogenic SeNPs suspensions is important because it assures a precise application of selenium product with very narrow physiological window.
The strigolactone mimics developed during STIM 4+ project implementation are important due to their multifaceted action. Strigolactone mimic SL-20 improves nodulation of hairy vetch by 32% when combined with rhizobia and rhizobia helper (increasing the nitrogen fixed in the soil), enhances tolerance of hairy vetch to abiotic stress, and promotes the establishment of the hairy vetch crop. Strigolactone mimic SL-20 demonstrates synergic interaction with glycodynameric chitosan and selected Trichoderma strains. The integrated application of Trichoderma and formulated SL-20 increased the accumulation of lycopene and gamma-aminobutyrate (GABA) in tomato fruits. Strigolactone mimics SL-6 and SL-20 enhance mycorrhization and promote soil carbon sequestration through extractable glomalin-related soil protein.
Glycodynameric hydrogel based on chitosan and different aldehydes were used as two-in-one product, an organic plant biostimulant and biocompatible 3D porous formulation structure for strigolactone mimic, stimuli-responsive and with a hydrophilic – hydrophobic segregation feature.
The multifaceted Trichoderma strain, Trichoderma pseudokoningii Tpk20, selected during project implementation and deposited under number DSM 34838 at Leibniz Institute DSMZ · German Collection of Microorganisms and Cell Cultures, is important because it increases the water use efficiency of vegetable plants. Integrated with the other plant biostimulants, it optimizes vegetable response to deficit irrigation, reducing the costs of vegetable cultivation in the field.
Healthy and vigorous vegetables transplants were produced by using an extract of the microalgae biostimulated by beneficial yeasts.
Plant biostimulant effect of extract produced from yeast-stimulated micro-algae on cucumber. The field experiments were organized at Valea-Stânei, with the support of Vegetable Cooperative Luica-Călărăși. The vegetables growers proposed deficit irrigation as a practical solution to manage the difficulties of field experiments. This practical solution emphasizes the complementary interactions between the components of the integrated agro-bio-nano-technology in increasing water use efficiency and saving costs.
Field experiments of integrated agro-nano-biotechnology. a) Aerial aspect of the field experiment in deficit conditions, demonstrating the benefits of the proposed integrated technology. The HV treated plots present a luxurious growth of tomatoes and higher yield and the bare control plant have limited growth and lower yield. b) Equipment used for determination in field of the plant photosynthetic parameters. c) aspects of the experimental plots. d) green tomatoes fruits from the experiments. e) suction lysimeter, used to sample soil solutions from experimental plots. f) portable porometer, used to determine stomatal conductance. g) portable fluorimeter, used to determine photosynthetic yield by pulse modulated chlorophyll fluorescence.
In a joint activity, the first toxicity assessment of the multifaceted strigolactone mimic (SL-6) to aquatic organisms was done. The main outcome of the risk assessment in terms of both the aquatic ecotoxicity and genotoxicity. The activity involved producing zygotes (equivalent to terrestrial plant seeds), harvesting zygotes and growing thallus (equivalent to stem and leaves of the terrestrial plants) in controlled condition, under different SL-6 concentrations. On the end, the effect of the strigolactone mimic was evaluated by stereomicroscopic assay.
Determination of SL-6 effects on F. vesiculosus brown seaweed. producing zygotes from female and male receptacles, harvesting and washing zygotes, cultivation of F. vesiculosus on controlled conditions (14 hours light, 10 hours dark, 20°C) and different concentrations of SL-6 for 14 days, examination of the thallus on stereomicroscope at different time of growing, images of the effects of SL-6 on F. vesiculosus thalus.
The ecotoxicology assay demonstrate that SL-6 is without negative effects at a dose 100 times higher than the doses applied to treat vegetables cultivated in high-residues mulch.
The integrated agro-bio-nano-technology produce more with less in areas with a deficit in selenium and challenging climatic conditions. The quality of the vegetables produced with the integrated agro-bio-nano-technology is higher. The better yield of the field grown vegetables with lower water consumption is important for vegetable growers, that increase the profitability of their farming. The quality of the field grown vegetable is important for consumers.
The taste of vegetable fruits produced with the integrated technology is better and the shelf life of is longer due to phytonutrients accumulation, including polyamine. The safety of the fresh vegetables produced by integrated technology was tested by comet assay and by determination of effects on proliferation of Caco-2 (DSMZ ACC 169), a cell line derived from a colorectal adenocarcinoma. The extracts from PB- treated tomatoes effectively prevented DNA damage. Most probably this is a result of the higher phytonutrients accumulation. There were no differences between effects on Caco-2 of extracts from fruits of vegetables cultivated in bare soil and those grown in hair-vetch mulch treated with biostimulants, except SeNPs treated experimental variants (due to the apoptotic effect of selenium on Caco-2 cancer cell).
Summary of bilateral results
The project results on the development of the biostimulants based on new strigolactone mimics, new Trichoderma strains and new chitosan-based glycodynamers illustrate the contribution of the project STIM 4+ to increased capacities of the Romanian partners. Three Ph.D. (female) students participated in developing and studying these new biostimulants. A young (female) Ph.D. student worked in Norway on the ecotoxicological and genotoxicity studies done for the first time worldwide for strigolactone mimics.