Graphene nanomaterials and copper co-exposure in Populus nigra L.: ultrastructural, biochemical, and nutritional impact under in vitro conditions Simonetta Muccifora, Maria Adelaide Iannelli, Barbara Casentini, Lorenzo Camoni, Anna Fiorillo, Davide Gentile, Manuela Melucci, Valerio Giorgio Muzzini, Valentina Iori BMC Plant Biology, 2026 Abstract The wide application of graphene nanomaterials has led to their release into the environment, raising ecological risk concerns, especially when co-existing with other pollutants like copper (Cu), one of the most ubiquitous environmental metals. The impact of co-presence of these nanomaterials and Cu on woody plants remains unstudied and, in this regard, callus culture represents a reliable tool for toxicological studies. In this work, we investigated the effects of Cu in combination with two different graphene nanomaterials, graphene oxide (GO) and graphene nanoplatelets (GNP), on the cell ultrastructure, biochemical responses and nutrient uptake in callus culture of Populus nigra L., a pioneer tree species in the riparian ecosystem. GO and GNP alone showed an adverse impact on poplar cells, causing a significant reduction in dry weight and a notable increase in MDA levels, water and Ca uptake, and protein synthesis. Co-exposure to Cu and either GO or GNP increased dry weight while decreasing water content, MDA levels, antioxidant enzyme activities, and protein content. Furthermore, GO + Cu exposure promoted greater cellular metal uptake than the GNP + Cu treatment, indicating a greater effectiveness of GO as a Cu carrier, due to the higher presence of oxygen functional groups on its surface than GNP. Transmission Electron Microscopy (TEM) observations confirmed the cellular uptake of both GO and GNP, revealing distinct impacts on cell ultrastructure. These results provide useful information on the interaction between graphene nanomaterials and Cu for risk assessment and developing sustainable management strategies in agro-forestry.
Sex-Related Differences in Physiological and Biochemical Responses of Populus nigra to Bifunctionalized Silver Nanoparticles and Silver Ions Exposure In Vitro Valentina Iori, Davide Gentile, Barbara Casentini, Lorenzo Camoni, Anna Fiorillo, Elena Kuzminsky, Iole Venditti, Maria Adelaide Iannelli Plants, 2025 The aim of this research was to assess the sex-related responses to AgNPs stabilized with citrate (Cit) and glutathione (GSH), relative to silver ions supplied as AgNO3 in black poplar (Populus nigra L.), a dioecious, woody model species. The impact of the AgNPs-cit-GSH on male and female clones was evaluated by measuring key parameters of oxidative stress. The results showed that exposure to nanosilver resulted in lower Ag accumulation and reduced MDA levels in both genders compared to AgNO3. The female clone exhibited a dose-dependent response, characterized by an increase in dry weight (DW), along with a reduction in nutrient uptake, protein content, and ATPase activity, as well as an upregulation of glutathione-S-transferase (GST) activity compared to the control. The male clone displayed a specific treatment response. Exposure to AgNPs-cit-GSH caused a decrease in DW, water content, and nutrient uptake, accompanied by a rise in protein content as well as GST activity. In AgNO3-treated male cells, the increase in Ag content and MDA levels corresponded to a decrease in DW and a rise in protein, Cu, and Ca content. These findings offer valuable insights into sexual dimorphism in dioecious woody plants, a topic that has been largely understudied yet is critical for sustainable resource management strategies.
The Emerging Role of the Salt Tolerance-Related Protein in the Abiotic Stress Response of Arabidopsis thaliana Anna Fiorillo, Michela Manai, Elisa Falliti, Sabina Visconti, Lorenzo Camoni Plants, 2025 Abiotic stresses severely impair plant growth and productivity. To counteract stress, plants have evolved intricate strategies, including the induction of stress-responsive proteins. The Arabidopsis thaliana Salt Tolerance-Related Protein (STRP) has recently emerged as a key player in abiotic stress tolerance. STRP is a small, hydrophilic, intrinsically disordered protein that exhibits the potential to adopt distinct conformations depending on the cellular context. STRP is localized in the cytosol and nucleus and is associated with the plasma membrane. Stress induces the subcellular redistribution of STRP, accompanied by a significant increase (up to ten-fold) in its levels due to reduced degradation by the 26S proteasome. Reverse genetics studies have demonstrated that STRP can mitigate the detrimental effects of oxidative stress and participate in modulating stress-related gene expression. Although the exact mechanism of STRP remains unclear, its physicochemical properties suggest a dual role as a molecular shield, interacting with macromolecules without a fixed conformation, and as a binder of specific defense-related client proteins, adopting a defined tertiary structure. This review provides a comprehensive overview of STRP and its emerging role as a multifunctional player in abiotic stress responses, also highlighting its potential for strengthening crop resilience and maintaining agricultural productivity under global climate challenges.
A Biostimulant Based on Ecklonia maxima and Yeast Extract Increases the Resistance of Tomato Plants Toward Pseudomonas syringae pv. tomato DC3000 Anna Fiorillo, Michela Manai, Mauro Marra, Lorenzo Camoni Physiologia Plantarum, 2025 Plant biostimulants represent a promising option to improve agricultural production and stress resistance while reducing the use of fertilizers and pesticides. Despite various evidence demonstrating the beneficial role of biostimulants in preventing the negative effects of abiotic stress on plants, the ability of biostimulants to bolster defense mechanisms has been brought to light only recently. In this work, the impact of a biostimulant based on Ecklonia maxima and yeast extracts (S/Y) on the response of tomato infected with Pseudomonas syringae pv. tomato DC3000 (Pst DC3000) was assessed. S/Y was selected after a screening to identify biostimulants capable of conferring resistance to Pst DC3000. S/Y boosts the early events of the plant's innate immunity. Indeed, biostimulation increased the Pst DC3000‐induced oxidative burst by upregulating the NADPH oxidase/respiratory burst oxidase homolog and apoplastic class III peroxidases expression. Moreover, the deposition of callose was also promoted. Due to improved activation of early defense responses by S/Y, disease symptoms and bacterial spread 72 h after the infection were significantly reduced. Finally, levels of salicylic acid, a key hormone in plant innate immunity, were increased by S/Y, whilst those of jasmonic acid and auxin, which are negative regulators in defense responses to Pst DC3000, were hampered. Overall, these findings show that S/Y mitigates infection symptoms by acting on different defense mechanisms, thus providing evidence of the potential of the biostimulant to improve plants' response to biotic stresses.
Phenotypical and biochemical characterization of tomato plants treated with triacontanol Michela Manai, Anna Fiorillo, Monica Matuozzo, Mei Li, Chiara D’Ambrosio, Loris Franco, Andrea Scaloni, Vincenzo Fogliano, Lorenzo Camoni, Mauro Marra Scientific Reports, 2024 Biostimulants are heterogeneous products designed to support plant development and to improve the yield and quality of crops. Here, we focused on the effects of triacontanol, a promising biostimulant found in cuticle waxes, on tomato growth and productivity. We examined various phenological traits related to vegetative growth, flowering and fruit yield, the metabolic profile of fruits, and the response of triacontanol-treated plants to salt stress. Additionally, a proteomic analysis was conducted to clarify the molecular mechanisms underlying triacontanol action. Triacontanol application induced advanced and increased blooming without affecting plant growth. Biochemical analyses of fruits showed minimal changes in nutritional properties. The treatment also increased the germination rate of seeds by altering hormone homeostasis and reduced salt stress-induced damage. Proteomics analysis of leaves revealed that triacontanol increased the abundance of proteins related to development and abiotic stress, while down-regulating proteins involved in biotic stress resistance. The proteome of the fruits was not significantly affected by triacontanol, confirming that biostimulation did not alter the nutritional properties of fruits. Overall, our findings provide evidence of the effects of triacontanol on growth, development, and stress tolerance, shedding light on its mechanism of action and providing new insights into its potential in agricultural practices.
Effect of Polyethylene Glycol-Simulated Drought Stress on Stomatal Opening in “Modern” and “Ancient” Wheat Varieties Ilva Licaj, Anna Fiorillo, Maria Chiara Di Meo, Ettore Varricchio, Mariapina Rocco Plants, 2024 Climate change is leading to an increase in the intensity, duration, and frequency of severe droughts, especially in southern and southeastern Europe, thus aggravating water scarcity problems. Water deficit stress harms the growth, physiology, and yield of crops like durum wheat. Hence, studying ancient wheat varieties’ stress responses could help identify genetic traits to enhance crop tolerance to environmental stresses. In this background, this study aimed to investigate the effects of PEG 6000-stimulated drought stress in the ancient wheat variety Saragolla and the modern one Svevo by analyzing various biochemical and molecular parameters that can especially condition the stomatal movement. Our data revealed that drought stress caused a significant increase in the levels of total soluble sugars, ABA, and IAA in both selected cultivars to a greater extent in the Saragolla than in the Svevo. We demonstrated that, under water deficit stress, calcium dynamics as well as the expression of ERF109, MAPK3/6, MYB60, and TaTPC1, involved in the activation of drought-related calcium-sensitive pathways, display significant differences between the two varieties. Therefore, our study provided further evidence regarding the ability of the ancient wheat variety Saragolla to better cope with drought stress compared to the modern variety Svevo.
14-3-3 Proteins and the Plasma Membrane H+-ATPase Are Involved in Maize (Zea mays) Magnetic Induction Anna Fiorillo, Ambra S. Parmagnani, Sabina Visconti, Giuseppe Mannino, Lorenzo Camoni, Massimo E. Maffei Plants, 2023 The geomagnetic field (GMF) is a natural component of the biosphere, and, during evolution, all organisms experienced its presence while some evolved the ability to perceive magnetic fields (MF). We studied the response of 14-3-3 proteins and the plasma membrane (PM) proton pump H+-ATPase to reduced GMF values by lowering the GMF intensity to a near-null magnetic field (NNMF). Seedling morphology, H+-ATPase activity and content, 14-3-3 protein content, binding to PM and phosphorylation, gene expression, and ROS quantification were assessed in maize (Zea mays) dark-grown seedlings. Phytohormone and melatonin quantification were also assessed by LG-MS/MS. Our results suggest that the GMF regulates the PM H+-ATPase, and that NNMF conditions alter the proton pump activity by reducing the binding of 14-3-3 proteins. This effect was associated with both a reduction in H2O2 and downregulation of genes coding for enzymes involved in ROS production and scavenging, as well as calcium homeostasis. These early events were followed by the downregulation of IAA synthesis and gene expression and the increase in both cytokinin and ABA, which were associated with a reduction in root growth. The expression of the homolog of the MagR gene, ZmISCA2, paralleled that of CRY1, suggesting a possible role of ISCA in maize magnetic induction. Interestingly, melatonin, a widespread molecule present in many kingdoms, was increased by the GMF reduction, suggesting a still unknown role of this molecule in magnetoreception.
The Salt Tolerance–Related Protein (STRP) Is a Positive Regulator of the Response to Salt Stress in Arabidopsis thaliana Anna Fiorillo, Michela Manai, Sabina Visconti, Lorenzo Camoni Plants, 2023 Salt stress is a major abiotic stress limiting plant survival and crop productivity. Plant adaptation to salt stress involves complex responses, including changes in gene expression, regulation of hormone signaling, and production of stress-responsive proteins. The Salt Tolerance–Related Protein (STRP) has been recently characterized as a Late Embryogenesis Abundant (LEA)–like, intrinsically disordered protein involved in plant responses to cold stress. In addition, STRP has been proposed as a mediator of salt stress response in Arabidopsis thaliana, but its role has still to be fully clarified. Here, we investigated the role of STRP in salt stress responses in A. thaliana. The protein rapidly accumulates under salt stress due to a reduction of proteasome–mediated degradation. Physiological and biochemical responses of the strp mutant and STRP–overexpressing (STRP OE) plants demonstrate that salt stress impairs seed germination and seedling development more markedly in the strp mutant than in A. thaliana wild type (wt). At the same time, the inhibitory effect is significantly reduced in STRP OE plants. Moreover, the strp mutant has a lower ability to counteract oxidative stress, cannot accumulate the osmocompatible solute proline, and does not increase abscisic acid (ABA) levels in response to salinity stress. Accordingly, the opposite effect was observed in STRP OE plants. Overall, obtained results suggest that STRP performs its protective functions by reducing the oxidative burst induced by salt stress, and plays a role in the osmotic adjustment mechanisms required to preserve cellular homeostasis. These findings propose STRP as a critical component of the response mechanisms to saline stress in A. thaliana.
A Phage Therapy Model for the Prevention of Pseudomonas syringae pv. actinidiae Infection of Kiwifruit Plants Anna Fiorillo, Domenico Frezza, Gustavo Di Lallo, Sabina Visconti Plant Disease, 2023 Great efforts have been made with chemicals and pesticides to contain the spread of Pseudomonas syringae pv. actinidiae (Psa) responsible for kiwifruit canker. Unfortunately, only partial results were obtained for this bacterial pandemic, and alternative remedies were proposed to avoid soil pollution and the onset of antibiotic resistance. Among these, phage therapy represents a possible tool with low environmental impact and high specificity. Several phages have been isolated and tested for the capacity to kill Psa in vitro, but experiments to verify their efficacy in vivo are still lacking. In the present study, we demonstrated that the phage φPSA2 (previously characterized) contains the spread of Psa inside plant tissue and reduces the symptoms of the disease. Our data are a strong indication for the efficiency of this phage and open the possibility of developing a phage therapy based on φPSA2 to counteract the bacterial canker of kiwifruit.
Comparative Analysis of the Response to Polyethylene Glycol-Simulated Drought Stress in Roots from Seedlings of “Modern” and “Ancient” Wheat Varieties Ilva Licaj, Maria Chiara Di Meo, Anna Fiorillo, Simone Samperna, Mauro Marra, Mariapina Rocco Plants, 2023 Durum wheat is widely cultivated in the Mediterranean, where it is the basis for the production of high added-value food derivatives such as pasta. In the next few years, the detrimental effects of global climate change will represent a serious challenge to crop yields. For durum wheat, the threat of climate change is worsened by the fact that cultivation relies on a few genetically uniform, elite varieties, better suited to intensive cultivation than “traditional” ones but less resistant to environmental stress. Hence, the renewed interest in “ancient” traditional varieties are expected to be more tolerant to environmental stress as a source of genetic resources to be exploited for the selection of useful agronomic traits such as drought tolerance. The aim of this study was to perform a comparative analysis of the effect and response of roots from the seedlings of two durum wheat cultivars: Svevo, a widely cultivated elite variety, and Saragolla, a traditional variety appreciated for its organoleptic characteristics, to Polyethylene glycol-simulated drought stress. The effect of water stress on root growth was analyzed and related to biochemical data such as hydrogen peroxide production, electrolyte leakage, membrane lipid peroxidation, proline synthesis, as well as to molecular data such as qRT-PCR analysis of drought responsive genes and proteomic analysis of changes in the protein repertoire of roots from the two cultivars.
