Juan Manuel Ruiz’s research while affiliated with University of Granada and other places

Background and aims Asparagus Decline Syndrome (ADS) threatens the sustainability and productivity of asparagus (Asparagus officinalis L.) cultivation. This study aimed to characterize the physiological responses of asparagus plants to ADS, focusing on oxidative metabolism, hormonal regulation, and phenolic compounds profiles to understand the underlying mechanisms and inform management strategies. Methods A field trial was conducted in the south of Spain comparing asparagus plants grown in soil from a plot previously affected by the ADS with a control soil (not affected). The key parameters assessed included biomass and oxidative stress indicators, phytohormone and phenolic compounds profiles in the root and shoot, and the soil phenolic compounds. Results ADS-affected plants exhibited lower fresh and dry weight and volume, and elevated oxidative stress, as evidenced by increased malondialdehyde (MDA) and H2O2 levels, along with enhanced activities of superoxide dismutase (SOD) and ascorbate peroxidase (APX). Hormonal analysis revealed higher concentrations of abscisic acid (ABA) and jasmonic acid, alongside a concurrent reduction in indoleacetic, suggesting a stress-induced response likely contributing to growth inhibition. Furthermore, the depletion of caffeic acid in roots, alterations in flavonoid profiles in shoot tissues, and increased PPO activity were observed, potentially worsening oxidative stress and depleting antioxidant reserves. Finally, ferulic acid derivatives in the soil were identified as potential allelopathic compounds. Conclusion These findings highlight the complexity of ADS and underscore the importance of integrated management strategies, including soil health management, resistant varieties selection, and targeted modulation of plant physiological responses to mitigate the impacts of ADS on asparagus production.

Cadmium (Cd) stress considerably reduces crop yield. The use of biostimulants, like humic substances (HS), is a research strategy for increasing Cd tolerance. The present study evaluated the effectiveness of a HS-based product in improving Cd tolerance in lettuce. It was selected for its ability to enhance growth through improved photosynthesis and antioxidant capacity although its effectiveness under Cd stress is unclear. The study also investigated which mode of HS application (to roots or leaves) is more effective in enhancing Cd tolerance and reducing Cd accumulation. Plants were grown under Cd stress (100 µM CdCl 2 ), Cd stress + HS applied to the roots (R-HS) at 0.40 and 0.60 mL/L, and HS applied to the leaves (F-HS) at 7.50 and 10.00 mL/L, alongside a control (no CdCl 2 ). R-HS involved irrigating pots with HS diluted in a nutrient solution, whereas F-HS consisted of spraying leaves with HS diluted in distilled water. Cd reduced shoot and root dry weight (DW) by 35%. However, a lower reduction was observed under HS applications (16%, 22%, 19%, and 18% at R-HS 0.40 and 0.60 mL/L, and F-HS 7.50 and 10.00 mL/L, respectively). These results were attributed to increased antioxidant capacity, thiol biosynthesis, and photosynthesis activity. While the application mode did not affect the Cd tolerance, R-HS 0.60 mL/L increased shoot Cd (21%), whereas F-HS 10.00 mL/L reduced it (14%). Our study indicates that the HS-based product could enhance Cd tolerance in lettuce at the tested doses. In addition, R-HS 0.60 mL/L may be effective for Cd phytoextraction, whereas F-HS can help reduce Cd accumulation in leaves.

Water stress is one of the foremost global abiotic stressors limiting agricultural productivity. Biostimulants and bioactive compounds are emerging as promising tools to enhance crop stress tolerance. This study investigates the effects of Cytolan® Stress, a novel seaweed-derived biostimulant, on the water stress tolerance of lettuce plants. Three application strategies were evaluated: priming, where the biostimulant is applied before the onset of stress to prepare the plants for adverse conditions; buffering, involving application at the onset of stress to mitigate its immediate effects; and detoxifying, where the biostimulant is applied after stress to aid in plant recovery. Biomass, stress-related parameters, antioxidant activity, osmoprotectant levels, and photosynthesis-related metrics were analyzed to elucidate its potential mechanisms of action. The results demonstrated that Cytolan® Stress in priming and buffering applications significantly improved water stress tolerance, reducing biomass loss from 45% to only 25%. Moreover, the detoxifying treatment was the most effective, as plants showed biomass values similar to those of the control plants. The biostimulant reduced oxidative stress indicators while enhancing antioxidant defenses, including ascorbate (AsA)-glutathione (GSH) cycle, antioxidant compounds, and enzyme activities. In addition, Cytolan® Stress preserved photosynthesis performance under water stress conditions. These findings highlight the potential of Cytolan® Stress to mitigate drought stress effects in lettuce, offering broader implications for crop tolerance and resilience under water-limited conditions. Further studies are recommended to explore its efficacy across different crops and stress scenarios.

Humic substances (HS) are commonly employed as plant biostimulants to enhance crop yields. However, the HS mechanisms of action, as well as the differences between radicular and foliar modes of application, remain unclear. Here, we explored the changes in phytohormonal balance as possible mechanism of HS to enhance lettuce (Lactuca sativa L.) growth, and the difference between both modes of application. For this purpose, BLACKJAK®, a HS‐based product was applied as radicular (R) and foliar (F) at the concentrations (mL/L): 0.20 (R1), 0.40 (R2), 0.60 (R3), 0.80 (R4), 5.00 (F1), 7.50 (F2), 10.00 (F3), and 12.50 (F4). The experiment was performed in pots filled with vermiculite:perlite (3:1) and HS were applied three times with a periodicity of 10 days. Shoot and root growth parameters were measured. In addition, the phythormones indole‐3‐acetic acid (IAA), gibberellins (GAs), trans‐zeatine (tZ), isopentenyl adenine (iP), 1‐aminocyclopropane‐1‐carboxylic acid (ACC), abscisic acid (ABA), jasmonic acid (JA), and salicylic acid (SA), were analysed by U‐HPLC‐MS. BLACKJAK® application resulted in higher shoot growth at doses R1, R2, R3, F2, and F3, whereas root biomass was increased at R2, R3, F2, F3, and F4, showing radicular better plant growth than foliar applications. Furthermore, HS changed phytohormonal balance in shoots and roots. However, it was with radicular applications, especially at R2, where phytohormonal profile was best associated with plant growth due to the increases observed in IAA, GAs, JA, SA, tZ, and decreased ABA. However, further research is needed to clarify the involvement of hormones in the growth‐promoting action of HS.

Increasing crop yield with low-N supplies has become one of the main aims of current agriculture to reduce the excessive use of chemical fertilizers. A sustainable strategy to improve crop productivity, N assimilation, and N Use Efficiency (NUE) under limit-N growth conditions is the application of biostimulants, such as humic substances (HS). Here, we evaluated the effectiveness of an HS-based biostimulant, BLACKJAK®, in improving lettuce growth and NUE under N-deficit conditions. Thus, BLACKJAK® was applied radicularly (R) and foliarly (F) at the following doses: R-HS 0.40 mL/L, R-HS 0.60 mL/L, F-HS 7.50 mL/L, and F-HS 10.00 mL/L. Three N levels were applied: optimal (7 mM) and N-deficit (3 mM and 1 mM). The results showed that shoot dry weight (DW) was reduced at 3 mM N (−32%) and 1 mM N (−42%). However, R and F BLACKJAK® enhanced plant growth at all three N levels, especially with F-HS at 10.00 mL/L, which showed an increase of 43% in shoot DW at 3 and 1 mM N, compared to plants not treated with HS. BLAKCJAK® also improved photosynthesis, NO3− and organic N accumulation, the activity of N assimilation enzymes, and the concentration of amino acids and proteins, regardless of the N level. In addition, HS enhanced NUE parameters under all N conditions, except for R-HS 0.60 mL/L at 1 mM N. Hence, our study suggests that the HS-based product BLACKJAK® could be a good candidate for reducing chemical fertilizer use and improving lettuce growth and NUE under low N conditions, although further research is required.

Humic substances (HS) have been defined as a potential plant biostimulant to improve crop yield in a sustainable and environmentally friendly way. Leonardite-suspension concentrate (SC) is a type of HS extracted from lignite that is currently employed to enhance various physiological aspects of plants. However, the different effects between both modes of SC application (root and foliar) are poorly understood, especially on photosynthesis performance. Therefore, this study aimed to investigate the influence of a leonardite-SC-based product (BLACKJAK®), on lettuce growth and photosynthesis efficiency, while comparing both methods of application. For this purpose, four root (R): R1 (0.20 mL/L), R2 (0.40 mL/L), R3 (0.60 mL/L), and R4 (0.80 mL/L), and four foliar: F1 (5.00 mL/L), F2 (7.50 mL/L), F3 (10.00 mL/L), and F4 (12.50 mL/L) BLACKJAK® doses were applied to lettuce plants. Related shoot and root growth parameters, photosynthetic efficiency, and sugar and starch content were assessed in lettuce plants. The results showed that BLACKJAK® improved shoot and root biomass, foliar area, and root length, especially at intermediate doses (R2, R3, F2, and F3), with R3 demonstrating the greatest growth increases. Similarly, the main photosynthetic parameters analyzed (net photosynthetic rate and Rubisco carboxylation efficiency), and the soluble sugars and starch content were improved by the same doses, with R3 showing the best photosynthetic performance. Hence, our study suggests that BLACKJAK® improves lettuce yield and photosynthetic efficiency, particularly with radicular application at R3.

Salinity stress constitutes one of the main abiotic stresses that considerably reduces crop yield. An approach to enhance plant growth under salt stress involves the addition of humic substances (HS) to roots or leaves. Here, we evaluated the potential use of BLACKJAK®, an HS-based product, to enhance salt tolerance in lettuce (Lactuca sativa L.). For this aim, plants were exposed to salinity (100 mM NaCl), salinity + HS: radicular (R)-HS (0.40 mL/L and 0.60 mL/L) and foliar (F)-HS (7.50 mL/L and 10.00 mL/L), along with a control (without NaCl). Parameters related to plant growth, Na⁺ and K⁺ accumulation, photosynthetic activity, oxidative stress, enzymatic and non-enzymatic antioxidants, as well as proline levels were evaluated. Results showed that R and F-HS considerably enhanced salinity tolerance. In addition, F-HS offered a greater improvement of plant growth in relation to shoot fresh weight, shoot relative growth rate, and foliar area, being 10.00 mL/L the best dose. This tolerance could be associated with reduced Na⁺ translocation to the shoot and enhanced shoot K⁺ accumulation, decreasing Na⁺/K⁺ ratio. Furthermore, HS improved the net photosynthetic rate, Rubisco carboxylation efficiency, and photosystem II performance, and reduced ROS levels and lipid peroxidation. Hence, our data show the potential use of BLACKJAK® to improve lettuce tolerance to salinity, with foliar application slightly better than radicular to achieve this aim, especially at 10.00 mL/L dose.