Improving photosynthetic efficiency and plant growth in controlled environments: the role of light quality, biostimulant application and ionising radiation

The possibility of growing higher plants, especially crops, in controlled environments allows reducing the variability of plant responses to the multiple stress occurring in the field and increasing the primary production. Among variables implicated in plant development and physiology, light represents a driver. Therefore, selecting specific wavelengths of visible light to obtain appropriate light quality regimes may significantly improve photosynthesis, biomass production, and secondary metabolites synthesis, resulting in an enrichment of food quality (in terms of nutraceutical compounds and antioxidants) and plant tolerance against abiotic stresses. The use of the light quality as 'natural fertiliser' alone or combined with other eco-friendly practices such as the employment of biostimulants could be a promising solution to enhance crop productivity preserving the overexploitation of soil and reducing the overuse of agrochemicals. The light spectrum modulation within growth chambers or vertical farms may find proper applications in the cultivation of crops in extreme environments on the Earth, such as hot or cold deserts or in the Space environment. This latter is highly unhospitable because it is characterised by many unfavourable ecological factors, including microgravity and cosmic radiations. In particular, ionising radiation is one of the significant constraints preventing plant growth and survival in Space. In the Bioregenerative Life Support Systems (BLSSs), conceived to sustain human life in Space, plants will have a crucial role in food production and air regeneration and CO2 removal. Thus, in the next future, the challenge to grow plants in Space must consider studying the effect of Space ionising radiation not only on humans and animals but also on plants. The PhD project is focused on the role of light quality in regulating the photosynthetic machinery of higher plants in controlled environments. In particular, it has been explored if and how specific light wavelengths during growth may modify plant physiological behaviour and phytochemical production in response to biostimulant application or exposure to ionising radiation. Among different variables affecting plant growth, biostimulants application was selected with the specific aim to improve the overall plant physiological performance in terms of primary and secondary metabolism in the context of sustainable agricultural practices. The ionising radiation was chosen as a space stress environmental factor in the view of experiments finalised to plant cultivation in Space. The experiments were carried out on widely consumed crops such as tomato, spinach, soybean and chard, considered important functional foods. Plants were grown in dedicated growth chambers under specific temperature, relative humidity, photoperiod, and light intensity condition, modulating the light spectrum to obtain specific light regimes promoting the photosynthetic performance. The outcomes of these experiments were utilised in the subsequent trials to test how light quality combined with ionising radiation or growth-promoting agents, i.e., biostimulants, may modify photosynthesis and antioxidant production. A downscaling investigative approach was adopted to analyse plant responses at different scale levels from cells and tissues to the whole organism. The outcomes of this research may have implications not only for developing sustainable protocols for indoor cultivation but also for plant growth in extreme environments on Earth and Space, such as the orbiting stations

Plant-Growth Promoting Microbes Change the Photosynthetic Response to Light Quality in Spinach

In this study, the combined effect of plant growth under different light quality and the application of plant-growth-promoting microbes (PGPM) was considered on spinach (Spinacia oleracea L.) to assess the influence of these factors on the photosynthetic performance. To pursue this goal, spinach plants were grown in a growth chamber at two different light quality regimes, full- spectrum white light (W) and red-blue light (RB), with (I) or without (NI) PGPM-based inoculants. Photosynthesis-light response curves (LRC) and photosynthesis-CO2 response curves (CRC) were performed for the four growth conditions (W-NI, RB-NI, W-I, and RB-I). At each step of LRC and CRC, net photosynthesis (PN), stomatal conductance (gs), Ci/Ca ratio, water use efficiency (WUEi), and fluorescence indexes were calculated. Moreover, parameters derived from the fitting of LRC, such as light-saturated net photosynthesis (PNmax), apparent light efficiency (Qpp), and dark respiration (Rd), as well as the Rubisco large subunit amount, were also determined. In not-inoculated plants, the growth under RB- regime improved PN compared to W-light because it increased stomatal conductance and favored the Rubisco synthesis. Furthermore, the RB regime also stimulates the processes of light conversion into chemical energy through chloroplasts, as indicated by the higher values of Qpp and PNmax in RB compared to W plants. On the contrary, in inoculated plants, the PN enhancement was significantly higher in W (30%) than in RB plants (17%), which showed the highest Rubisco content among all treatments. Our results indicate that the plant-growth-promoting microbes alter the photosynthetic response to light quality. This issue must be considered when PGPMs are used to improve plant growth performance in a controlled environment using artificial lighting

Role of Poly(ADP-Ribose) Polymerase (PARP) Enzyme in the Systemic Acquired Acclimation Induced by Light Stress in Phaseolus vulgaris L. Plants

Plants are able to acclimate to environmental constraints through functional modifications that may also occur in tissues that are not directly exposed to stress. This process is termed “systemic acquired acclimation.” The present study aims to evaluate the involvement of PolyADP-ribose) polymerase (PARP) protein in the acclimation process to high light (HL) stress in Phaseolus vulgaris plants. For this purpose, some leaves located at the top of the plant, in the apical position, were directly exposed to HL (“inducing” leaves), while others on the same plant, distal from the top, continued to be exposed to growth light (“receiving” leaves) to verify the hypothesis that an “alert” message may be transferred from injured tissues to distal ones. Biochemical and eco-physiological analyses, namely PARP activity, H2O2 and water- and fat-soluble antioxidants (i.e., ascorbic acid, tocopherol, glutathione (GSH), phenols, carotenoids, etc.) content, and chlorophyll fluorescence measurements were performed on both “inducing” and “receiving” leaves. Even if no change in PARP expression was found, its activity increased in “receiving” unstressed leaves in response to the light stress duration experimented by “inducing” leaves, while antioxidant capacity declined. When the “receiving” leaves were exposed to HL, the PARP activity returned to the control value, while antioxidant capacity photosynthetic electron transport rate (Jf) decreased and increased, respectively, compared to Control. Our results seem to show an acclimation pathway triggered in remote tissues not yet subjected to stress, likely involving a reactive oxygen species wave activating the PARP enzyme in a mechanism still to be clarified. In addition, the increased tolerance of plants directly exposed to HL could implicate a boosted synthesis of soluble antioxidants accompanied by a reduction of PARP activity to reduce excessive consumption of NAD(P)

Aerated Buffalo Slurry Improves Spinach Plant Growth and Mitigates CO2 and N2O Emissions from Soil

Manure management is the main strategy for mitigating gas emissions from livestock farming. In this study, a laboratory-scale experiment was set up to identify suitable conditions to be applied in a farm-scale experiment. The liquid fraction (LF) of slurry was aerobically treated and greenhouse gas emissions from soil were evaluated. Furthermore, the value of treated LF as a fertilizer on spinach plants was also tested. The aeration of LF determined an increase in mean alkalinity due to ammonia loss. The mass fraction of heavy metals also decreased, likely due to the reduction in solubility. After being applied on soil, aerated LF determined lower CO2 and N2O emissions compared to untreated LF due to a reduced nitrogen load. Spinach plants fertilized with treated LF showed a lush growth and exhibited a lower heavy metal mass fraction as well as a higher content of antioxidants compared to plants fertilized with untreated slurry. Our results show that aeration might be an effective alternative for slurry management as it is able to produce an eco-friendly final product with a high fertilizing value

Light Spectral Composition Influences Structural and Eco-Physiological Traits of Solanum lycopersicum L. cv. ‘Microtom’ in Response to High-LET Ionizing Radiation

This study evaluated if specific light quality (LQ) regimes (white fluorescent, FL; full- spectrum, FS; red-blue, RB) during plant growth modified morphological and photosynthetic traits of Solanum lycopersicum L. ‘Microtom’ plants irradiated at the dry seed stage with 25 Gy 48Ca ions (IR). The irradiation reduced plant size while it increased leaf dry matter content (LDMC) and relative water content (RWC) compared to the control. FS and RB light regimes determined a decrease of plant height and a rise of RWC compared to FL plants. The irradiation under FS and RB regimes favoured the development of dwarf plants and improved the leaf water status. Under the FL regime, irradiated plants showed reduced photosynthesis and stomatal conductance. The opposite behavior was observed in RB irradiated plants in which gas exchanges were significantly stimulated. RB regime enhanced Rubisco expression in irradiated plants also inducing anatomical and functional adjustments (i.e., increase of leaf thickness and incidence of intercellular spaces). Finally, 48Ca ions did not prevent fruit ripening and the achievement of the ‘seed-to seed’ cycle, irrespective of the LQ regime. Overall, the present study evidenced that RB light regime was the most effective in optimising growth and photosynthetic efficiency of ‘Microtom’ irradiated plants. These outcomes may help to develop proper cultivation protocols for the growth of dwarf tomato in Controlled Ecological Life Support Systems (CELSS)

Photosynthetic, Molecular and Ultrastructural Characterization of Toxic Effects of Zinc in Caulerpa racemosa Indicate Promising Bioremediation Potentiality

Caulerpaceae are unconventional green algae composed of multinucleated, single siphonous cells. The species of Caulerpa are acquiring major scientific interest for both their invasion in the Mediterranean ecological niche and for the production of valuable natural metabolites. Furthermore, the abilities of Caulerpa spp. in the biorecovery of polluted waters were recently investigated. Among heavy metal contaminants in marine systems, zinc (Zn) is considered a critical pollutant, progressively accumulating from plastic leachates. In this study, the responses of Caulerpa racemosa to different levels (5-10 mg L-1) of Zn were studied for 14 days under laboratory-controlled conditions. Effects of Zn were monitored by measuring the growth rate, photosynthetic efficiency and gene expression. Moreover, the ability of Caulerpa to remove Zn from seawater was monitored. Zn induced detrimental effects by decreasing the relative growth rate (RGR) and maximal PSII photochemical efficiency (Fv/Fm). Moreover, C. racemosa, grown in contaminated seawater, reduced the levels of Zn to a final concentration of 1.026 and 1.932 mg L-1 after 14 days, thus demonstrating efficient uptake. Therefore, our results characterized the effects of zinc on C. racemosa and the possible role of this alga as being effective in the bioremediation of marine seawater

Modulation of Antioxidant Compounds in Fruits of Citrus reticulata Blanco Using Postharvest LED Irradiation

Phlegrean mandarin fruits are already known for health-promoting properties due to the high concentration of phytochemicals in peel, pulp, and seed. Biotic and abiotic factors, including light, may modulate their biosynthesis, metabolism, and accumulation. In this context, light-emitting diodes (LED) have recently been applied to control nutritional traits, ripening process, senescence, fruit shelf-life, and pathogenic microbial spoilage of fruits. This study investigated the effect of the seven-day exposure of Phlegrean mandarin fruits to two LED regimes, white (W) and red–blue (RB), to test the possibility that the storage under specific light wavelengths may be used as green preservation technology that enhances fruit phytochemical properties. To pursue this aim, the antioxidant activity and polyphenolic profile of the pulp and peel of mandarins under W and RB light regimes were evaluated and compared with Control fruits not exposed to LED treatment. Our results indicated that storage under W and RB treatments modulates the antioxidant content in pulp and peel differently. Compared to W, the RB regime increases the ascorbic acid, flavonoid, anthocyanin, and carotenoid concentrations, while the polyphenol profile analysis reveals that the number of important phytochemicals, i.e., quercetin rutinoside, chlorogenic acid, sinensetin, and rutin, are higher under W. The overall data demonstrated that postharvest LED irradiation is a valid tool for modifying fruit phytochemical properties, which also boosts specific bioactive compounds

Antioxidant Properties of Pulp, Peel and Seeds of Phlegrean Mandarin (Citrus reticulata Blanco) at Different Stages of Fruit Ripening

In this work, we assess the potential of waste products of Phlegrean mandarin (Citrus reticulata Blanco), namely seeds and peel, to be reutilized as a source of bioactive compounds beneficial for the human diet. Starting from the evidence that the by-products of this specific cultivar are the most powerful sources of antioxidants compared to pulp, we have investigated if and how the bioactive compounds in peel and seeds may be affected by fruit ripening. Three stages of fruit ripening have been considered in our study: unripe fruits = UF, semi-ripe fruits = SRF, ripe fruits = RF. The overall results indicated that RF showed the highest concentration of antioxidants. Among fruit components, peel was the richest in total antioxidant capacity, total polyphenol content, total flavonoids, total chlorophylls and carotenoids, while seeds exhibited the highest concentration of total condensed tannins and ascorbic acid. The liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay indicates the occurrence, in peel extracts, of 28 phenolic compounds, mainly flavonoids (FLs); in seeds, 34 derivatives were present in the first stage (UF), which diminish to 24 during the ripening process. Our data indicated that the content of phytochemicals in citrus strongly varies among the fruit components and depends on the ripening stage. The higher antioxidant activity of peel and seeds, especially in RF, encourage a potential use of by-products of this specific citrus cultivar for industrial or pharmacological applications. However, to maximize the occurrence of desired bioactive compounds, it is important also to consider the ripening stage at which fruits must be collected

Impact of Different Fertilizer Sources under Supplemental Irrigation and Rainfed Conditions on Eco-Physiological Responses and Yield Characteristics of Dragon’s Head (Lallemantia iberica)

The effects of the irrigation regime and different fertilizer sources on the eco-physiological responses and yield characteristics of dragon’s head were explored in a factorial experiment based on a randomized complete block design with 12 treatments and 3 replications in the 2019 growing season. The treatments included six different fertilizer sources (animal manure, vermicompost, poultry manure, biofertilizer, chemical fertilizer, and control) and two irrigation regimes (rainfed and supplemental irrigation). The results indicated the positive effects of supplementary irrigation and the application of vermicompost, poultry manure, and animal manure by increasing the absorption of nutrients (phosphorus and potassium) and improving relative water contents, chlorophyll and carotenoid contents, and the fixed oil percentage of dragon’s head. The activities of catalase, ascorbate peroxidase, and superoxide dismutase decreased in the rainfed plants, whereas organic fertilizer application increased the antioxidant enzyme activity. The highest grain yield (721 kg ha−1), biological yield (5858 kg ha−1), total flavonoids (1.47 mg g−1 DW), total phenol (27.90 mg g−1 DW), fixed oil yield (200.17 kg ha−1), and essential oil yield (1.18 kg ha−1) were noted in plants that were treated with vermicompost under supplemental irrigation. Therefore, it is recommended that organic fertilizers such as vermicompost and poultry manure be used to substitute chemical fertilizers. These practices can help popularize organic crops using rainfed and supplementary irrigation

Counteracting the negative effects of copper limitations through the biostimulatory action of a tropical plant extract in grapevine under pedo-climatic constraints

In southern Mediterranean areas, vineyards are facing the combination of increasing air temperature, drought and frequency of extreme events (e.g., heat waves) due to climate change. Since most of the berry growth and ripening phases occur during the aridity period, such environmental constraints are responsible for limitations in yield and berry quality. Within this scenario, to achieve vineyard sustainability, renewed approaches in vineyard management have been proposed and the use of plant biostimulants seems a prominent and environmental friendly practice. The aim of this study was to test four combinations of a tropical plant extract and conventional chemicals for disease control on morpho-anatomical, physiological, biochemical and berry quality in Vitis vinifera L. subsp. vinifera “Aglianico.” In particular, we aimed to evaluate the possibility to counteract the negative effects of the reductions in copper distribution, by applying the tropical plant extract enriched with: micronutrients, enzymes involved in the activation of natural defense, aminoacids, and vitamins. The halved dose of Cu in combination with the tropical plant extract allowed maintaining a reduced vegetative vigor. In the second year of treatment, the addition of the plant extract significantly improved leaf gas exchanges and photochemistry as well as the synthesis of photosynthetic pigments. At berry level, the plant extract induced an increase in phenolics accompanied by a decrease in soluble sugars. The overall results showed that the expected differences in growth performance and productivity in vines are linked to different eco-physiological and structural properties induced by the various treatments. The tropical plant extract also primed plant defenses at the leaf and fruit levels, mainly due to modifications of some structural and biochemical traits, respectively