Plant biostimulants (PBs) represent a sustainable and efficient strategy to achieve stable crop yields under optimal or sub-optimal conditions, reducing chemical inputs. The new EU Regulation 2019/1009 has defined them as “substances and/or microorganisms that applied to the plant or rhizosphere stimulate natural processes, improve nutrient uptake and assimilation efficiency, tolerance to abiotic stresses, and product quality”. They include a large variety of biological products such as plant extracts, humic and fulvic acids, protein hydrolysates, phosphites, seaweed extracts, and living microorganisms, typically bacteria and fungi. They can promote plant growth, increase crop yields and quality, nutrient uptake, nitrogen (N) fixation, phosphorus (P), potassium (K) and micronutrient solubilization, biotic and abiotic stress tolerance. In this PhD thesis, the effect of Kocuria rhizophila and Streptomyces violaceoruber, two actinomycete strains, and Eranthis (Green Has Italia), a commercial non-microbial biostimulant, was evaluated to highlight their potential for improving growth performance of tomato (Solanum lycopersicum L., UC82, a commercial variety), used as plant model, grown under nutritional and drought stress. First, twenty-two bacterial strains were preliminary characterized for multiple PGPB traits. Among them, K. rhizophila and S. violaceoruber showed the best PGP activities, such as indole acetic acid production, phosphate solubilization, N_2 fixation, as well as drought and salt tolerance. The secondary metabolites and volatile organic compounds (VOCs) and the PGP activity of these actinomycete strains were evaluated on tomato. Their secreted and cellular metabolome revealed a rich arsenal of bioactive molecules, including antibiotics and siderophores, with S. violaceoruber, being the most effective PGPB strain. In particular, it was able to improve the seed germination index, hypocotyl and epicotyl growth and seedling growth. Furthermore, the produced VOCs showed antimicrobial activity, being able to modulate volatilome and exert control on the global DNA methylation of tomato seedlings. For these reasons, both K. rhizophila and S. violaceoruber were deeply studied for their PGPB activities. By a multi -omics approach, differential expressed genes as well as accumulated proteins and metabolites involved in the plant growth promotion in K. rhizophila-treated tomato plants compared to untreated control. Furthermore, eight gene modules based on their correlation with differential accumulated proteins and metabolites were identified by a weighted gene co-expression network analysis (WGCNA) approach. In particular, two modules showed the highest correlation with nine proteins, among which a nucleoside diphosphate kinase, and several metabolites, mainly belonging to amino acids and tricarboxylic acids (TCA). Our findings highlighted that sugars and amino acids, key nutrients for improving plant growth and yield, were strongly modulated by plant-K. rhizophila interaction. Furthermore, the ability of S. violaceoruber to mitigate N and/or drought stress condition was evaluated in different tomato growth systems: hydroponics, pots and plate-on-plate. In hydroponics, S. violaceoruber was able to increase morpho-physiological parameters such as fresh and dry root weight, root diameter and lateral root numbers, chlorophyll content, net photosynthesis rate, transpiration in tomato N stressed plants. Interestingly, it was able to restore these morpho-physiological parameters to the high N values. The undecylprodigiosin, mycelial red pigment produced by S. violaceoruber along tomato roots, confirmed root-bacteria interaction. In pots, the S. violaceoruber treatment increased the root fresh weight, shoot dry weight and SPAD index as well as net photosynthetic rate, transpiration rate and stomatal conductance of tomato treated plants. More interestingly, treated plants showed also an improved nitrogen use efficiency (NUE) and its component, nitrogen utilization efficiency (NUtE), useful traits to maintain high plant growth and yields. In plate-on-plate system, volatiles, produced by S. violaceoruber, applied by indirect inoculation, increased plant biomass, number and length of lateral roots, in control conditions, but they were not able to overcome drought stress effect. Finally, Eranthis, a commercial non-microbial biostimulant based on brown seaweed (Ascophylum nodosum and Laminaria digitata) and yeast extracts (Green Has Italia S.p.a (Canale, Italy), was evaluated for its ability to mitigate N stress. Under low N, Eranthis was able to increase fresh and dry weight, and lateral root number in tomato plants, grown in hydroponic system. This effect could be due to its composition rich in antioxidant molecules, such as flavonoids and flavanols, which contribute to reactive oxygen species (ROS) scavenging. In conclusion, both S. violaceoruber and K. rizophila and the no-microbial Eranthis could be considered as new candidates for developing novel biofertilizers with low environmental impact in a more sustainable cropping system. Further researches are needed to understand their mode of action to improve plant growth and stress tolerance, as well as their correct and efficient use in agriculture.
I biostimolanti rappresentano una strategia sostenibile ed efficiente per ottenere rese colturali stabili in condizioni ottimali o sub-ottimali, riducendo gli input chimici. Il nuovo Regolamento UE 2019/1009 li definisce come "sostanze e/o microrganismi che applicati alla pianta o alla rizosfera stimolano i processi naturali, migliorano l'efficienza di assorbimento e assimilazione dei nutrienti, la tolleranza agli stress abiotici e la qualità del prodotto". Essi comprendono un'ampia varietà di prodotti biologici, come estratti vegetali, acidi umici e fulvici, idrolizzati proteici, fosfiti, estratti di alghe marine, e microrganismi vivi, tipicamente batteri e funghi. I biostimolanti possono promuovere la crescita delle piante, aumentare la resa e la qualità delle colture, l'assorbimento dei nutrienti, la fissazione dell'azoto (N), la solubilizzazione di fosforo (P), potassio (K) e micronutrienti, la tolleranza agli stress biotici e abiotici. In questo progetto di tesi, è stato valutato l'effetto di Kocuria rhizophila e Streptomyces violaceoruber, due ceppi di actinomiceti, e di Eranthis (Green Has Italia), un biostimolante commerciale non microbico, nel migliorare le performance di crescita di pomodoro (Solanum lycopersicum L., UC82, varietà commerciale), usato come pianta modello, in diverse condizioni di stress nutrizionale ed idrico. In primo luogo, 22 ceppi batterici sono stati caratterizzati per le molteplici caratteristiche di batteri promotori di crescita delle piante (PGPB). Tra questi, K. rhizophila e S. violaceoruber, sono risultati i ceppi migliori per le loro attività PGPB, quali la produzione di acido indoloacetico, la solubilizzazione dei fosfati, la fissazione di N2 e la tolleranza alla siccità e al sale. Sono stati valutati i loro metaboliti secondari, i composti organici volatili (VOCs) e la loro attività PGP sul pomodoro. Il loro metaboloma secreto e cellulare ha rivelato un ricco arsenale di molecole bioattive, tra cui antibiotici e siderofori, e tra questi, S. violaceoruber si è dimostrato il ceppo più efficace. In particolare, esso era in grado di migliorare l'indice di germinazione, la crescita dell'ipocotile e dell'epicotile e la crescita delle piantine. Inoltre, i VOCs prodotti mostravano attività antimicrobica, essendo in grado di modulare il volatiloma e di esercitare un controllo sulla metilazione del DNA delle piantine di pomodoro. Alla luce di questi risultati, sia K. rhizophila che S. violaceoruber sono stati studiati più a fondo per le loro attività PGPB. Mediante un approccio multi-omico, sono stati identificati geni, proteine e metaboliti, coinvolti nella promozione della crescita delle piante, attivati da K. rhizophila in piante di pomodoro. Inoltre, con un approccio WGCNA, sono stati identificati otto moduli genici correlati con proteine e metaboliti accumulati in modo differenziale. In particolare, due moduli hanno mostrato la più alta correlazione con nove proteine, tra cui una nucleoside difosfato chinasi, ed una serie di metaboliti appartenenti ad amminoacidi e TCA. I risultati hanno evidenziato che zuccheri ed aminoacidi, nutrienti chiave per la crescita e resa delle piante, sono stati fortemente modulati dall'interazione pianta-K. rhizophila. Successivamente, è stata valutata la capacità di S. violaceoruber di mitigare le condizioni di stress da N e/o idrico in pomodoro uitlizzando diversi sistemi di crescita: idroponica, vaso e plate-on-plate. In idroponica, S. violaceoruber era in grado di aumentare i parametri morfo-fisiologici come il peso delle radici fresche e secche, il diametro, il numero di radici laterali, il contenuto di clorofilla, il tasso di fotosintesi netta e la traspirazione delle piante di pomodoro sottoposte ad N stress. Inoltre, il trattamento era in grado di ripristinare questi parametri morfo-fisiologici alla condizione di alto N. L’undecilprodigiosina, pigmento rosso miceliare prodotto da S. violaceoruber lungo le radici del pomodoro, conferma l'interazione radice-batterio. In vaso, il trattamento con S. violaceoruber aumentava il peso fresco delle radici, il peso secco dei germogli, l'indice SPAD, il tasso fotosintetico netto, il tasso di traspirazione e la conduttanza stomatica delle piante di pomodoro. Il trattamento ha anche influenzato lo sviluppo della pianta migliorando l'efficienza d'uso dell'azoto (NUE) e l'utilizzo dell'azoto (NUtE), utile per mantenere alte le rese. Nel sistema plate-on-plate, i composti volatili prodotti da S. violaceoruber, applicato tramite inoculazione indiretta, aumentavano la biomassa della pianta, il numero e la lunghezza delle radici laterali, in condizioni di controllo, ma non sono stati in grado di mitigare l'effetto dello stress idrico. Infine, Eranthis, un biostimolante commerciale di natura non microbica, costituito da estratti di alghe brune (Ascophylum nodosum and Laminaria digitata) e lieviti (Green Has Italia S.p.a (Canale, Italy), è stato valutato per la sua capacità di mitigare l’N stress. Il trattamento con Eranthis ha influenzato i parametri morfologici delle radici, aumentando il peso fresco e secco e il numero di radici laterali nelle piante di pomodoro, coltivate in un sistema idroponico, in condizioni di limitazione di N. Questo effetto potrebbe essere dovuto alla sua composizione ricca di molecole antiossidanti, come flavonoidi e flavanoli, che sono in grado di contribuire alla rimozione dei ROS. In conclusione, S. violaceoruber e K. rizophila potrebbero essere considerati come nuovi candidati per lo sviluppo di nuovi biofertilizzanti a basso impatto ambientale per un sistema colturale più sostenibile. Sono necessarie ulteriori ricerche per meglio comprendere le modalità d'azione dei biostimolanti, microbici e non, nel migliorare la crescita delle piante e la tolleranza agli stress, per un loro più efficace utilizzo in un'agricoltura sostenibile.
Evaluation of Biostimulants activity in mitigating the N and drought stress in tomato / Caldiero, Ciro. - (2024 Mar 20).
Evaluation of Biostimulants activity in mitigating the N and drought stress in tomato
Caldiero, Ciro
2024-03-20
Abstract
Plant biostimulants (PBs) represent a sustainable and efficient strategy to achieve stable crop yields under optimal or sub-optimal conditions, reducing chemical inputs. The new EU Regulation 2019/1009 has defined them as “substances and/or microorganisms that applied to the plant or rhizosphere stimulate natural processes, improve nutrient uptake and assimilation efficiency, tolerance to abiotic stresses, and product quality”. They include a large variety of biological products such as plant extracts, humic and fulvic acids, protein hydrolysates, phosphites, seaweed extracts, and living microorganisms, typically bacteria and fungi. They can promote plant growth, increase crop yields and quality, nutrient uptake, nitrogen (N) fixation, phosphorus (P), potassium (K) and micronutrient solubilization, biotic and abiotic stress tolerance. In this PhD thesis, the effect of Kocuria rhizophila and Streptomyces violaceoruber, two actinomycete strains, and Eranthis (Green Has Italia), a commercial non-microbial biostimulant, was evaluated to highlight their potential for improving growth performance of tomato (Solanum lycopersicum L., UC82, a commercial variety), used as plant model, grown under nutritional and drought stress. First, twenty-two bacterial strains were preliminary characterized for multiple PGPB traits. Among them, K. rhizophila and S. violaceoruber showed the best PGP activities, such as indole acetic acid production, phosphate solubilization, N_2 fixation, as well as drought and salt tolerance. The secondary metabolites and volatile organic compounds (VOCs) and the PGP activity of these actinomycete strains were evaluated on tomato. Their secreted and cellular metabolome revealed a rich arsenal of bioactive molecules, including antibiotics and siderophores, with S. violaceoruber, being the most effective PGPB strain. In particular, it was able to improve the seed germination index, hypocotyl and epicotyl growth and seedling growth. Furthermore, the produced VOCs showed antimicrobial activity, being able to modulate volatilome and exert control on the global DNA methylation of tomato seedlings. For these reasons, both K. rhizophila and S. violaceoruber were deeply studied for their PGPB activities. By a multi -omics approach, differential expressed genes as well as accumulated proteins and metabolites involved in the plant growth promotion in K. rhizophila-treated tomato plants compared to untreated control. Furthermore, eight gene modules based on their correlation with differential accumulated proteins and metabolites were identified by a weighted gene co-expression network analysis (WGCNA) approach. In particular, two modules showed the highest correlation with nine proteins, among which a nucleoside diphosphate kinase, and several metabolites, mainly belonging to amino acids and tricarboxylic acids (TCA). Our findings highlighted that sugars and amino acids, key nutrients for improving plant growth and yield, were strongly modulated by plant-K. rhizophila interaction. Furthermore, the ability of S. violaceoruber to mitigate N and/or drought stress condition was evaluated in different tomato growth systems: hydroponics, pots and plate-on-plate. In hydroponics, S. violaceoruber was able to increase morpho-physiological parameters such as fresh and dry root weight, root diameter and lateral root numbers, chlorophyll content, net photosynthesis rate, transpiration in tomato N stressed plants. Interestingly, it was able to restore these morpho-physiological parameters to the high N values. The undecylprodigiosin, mycelial red pigment produced by S. violaceoruber along tomato roots, confirmed root-bacteria interaction. In pots, the S. violaceoruber treatment increased the root fresh weight, shoot dry weight and SPAD index as well as net photosynthetic rate, transpiration rate and stomatal conductance of tomato treated plants. More interestingly, treated plants showed also an improved nitrogen use efficiency (NUE) and its component, nitrogen utilization efficiency (NUtE), useful traits to maintain high plant growth and yields. In plate-on-plate system, volatiles, produced by S. violaceoruber, applied by indirect inoculation, increased plant biomass, number and length of lateral roots, in control conditions, but they were not able to overcome drought stress effect. Finally, Eranthis, a commercial non-microbial biostimulant based on brown seaweed (Ascophylum nodosum and Laminaria digitata) and yeast extracts (Green Has Italia S.p.a (Canale, Italy), was evaluated for its ability to mitigate N stress. Under low N, Eranthis was able to increase fresh and dry weight, and lateral root number in tomato plants, grown in hydroponic system. This effect could be due to its composition rich in antioxidant molecules, such as flavonoids and flavanols, which contribute to reactive oxygen species (ROS) scavenging. In conclusion, both S. violaceoruber and K. rizophila and the no-microbial Eranthis could be considered as new candidates for developing novel biofertilizers with low environmental impact in a more sustainable cropping system. Further researches are needed to understand their mode of action to improve plant growth and stress tolerance, as well as their correct and efficient use in agriculture.File | Dimensione | Formato | |
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