Can lettuce plants grow in saline soils supplemented with biochar?

Salt stress is presently a major environmental concern, given the huge number of soils affected by the presence of dissolved salts. Therefore, it is necessary to find solutions, preferably nature-based ones, to deal with this problem. In this study, biochar, a product made from plant biomass residues through the process of pyrolysis, was tested to alleviate salt stress on lettuce (Lactuca sativa L.) plants. Six different concentrations of NaCl were tested: 0, 50, 100, 200, 300 and 400 mM with and without the addition of 5% (w/w) biochar. Biochar ability to mitigate salinity damage was assessed by means of both biometric (fresh weight), physiological (chlorophyll content), and biochemical (i.e., electrolyte leakage, total antioxidant power, total soluble proteins, free amino acids, and mineral content) parameters. The experiment lasted four weeks. The results showed that NaCl has a negative effect from the concentration of 100-200 mM and that biochar was to some extent effective in mitigating the negative effects of salt on plant physiology; nevertheless, biochar failed to counteract Na accumulation. Similarly, biochar did not influence the content of free amino acids in lettuce leaves, but enhanced the expression of several parameters, such as total antioxidant power, fresh weight, chlorophyll content, total soluble protein, K content, although only clearly evident in some cases. Overall, the present study showed that biochar is a viable solution to counteract the damage caused by high salt concentrations on plant growth.

Dose-Dependent Effects of a Corn Starch-Based Bioplastic on Basil (Ocimum basilicum L.): Implications for Growth, Biochemical Parameters, and Nutrient Content.

Plastic pollution is a pressing global issue, prompting the exploration of sustainable alternatives such as bioplastics (BPs). In agriculture, BPs have gained relevance as mulching films. This study investigated the effect of the presence in the soil of different concentrations (0-3%, w/w) of a corn starch-based bioplastic on basil (Ocimum basilicum L.). The results showed that increasing bioplastic concentration reduced shoot fresh biomass production. Biochemical analyses revealed changes in the shoot in soluble protein content, biomarkers of oxidative and osmotic stress (malondialdehyde and proline, respectively), anti-radical activity, and antioxidant compounds (phenols, flavonoids, and ascorbic acid), which are indicative of plant adaptive mechanisms in response to stress caused by the presence of the different concentrations of bioplastic in the soil. Macro- and micronutrient analysis showed imbalances in nutrient uptake, with a decreased content of potassium, phosphorus, and manganese, and an increased content of magnesium, iron, and copper in the shoot at high BP concentrations.

Transcriptomic and physiological approaches to decipher cold stress mitigation exerted by brown-seaweed extract application in tomato.

Chilling temperatures represent a challenge for crop species originating from warm geographical areas. In this situation, biostimulants serve as an eco-friendly resource to mitigate cold stress in crops. Tomato (Solanum lycopersicum L.) is an economically important vegetable crop, but quite sensitive to cold stress, which it encounters in both open field and greenhouse settings. In this study, the biostimulant effect of a brown-seaweed extract (BSE) has been evaluated in tomato exposed to low temperature. To assess the product effects, physiological and molecular characterizations were conducted. Under cold stress conditions, stomatal conductance, net photosynthesis, and yield were significantly (p ≤ 0.05) higher in BSE-treated plants compared to the untreated ones. A global transcriptomic survey after BSE application revealed the impact of the BSE treatment on genes leading to key responses to cold stress. This was highlighted by the significantly enriched GO categories relative to proline (GO:0006560), flavonoids (GO:0009812, GO:0009813), and chlorophyll (GO:0015994). Molecular data were integrated by biochemical analysis showing that the BSE treatment causes greater proline, polyphenols, flavonoids, tannins, and carotenoids contents.The study highlighted the role of antioxidant molecules to enhance tomato tolerance to low temperature mediated by BSE-based biostimulant.

Exploring sustainable alternatives: Wood distillate alleviates the impact of bioplastic in basil plants.

The growing interest in bioplastics and bio-based crop management products in agriculture is driven by the Sustainable Development Goals of the 2030 Agenda. However, recent research has raised concerns about the sustainability of bioplastics due to their potential negative impact on crop growth and yield, with implications for the environment and human health. In this study, wood distillate (WD) was evaluated as a natural enhancer of plant growth and defence system to mitigate the negative impact of a starch-based bioplastic on basil (Ocimum basilicum L.) plants. The study analyzed physiological and biochemical changes in basil plants subjected for 35 days to single or combined treatments of WD and bioplastic by measuring biomarkers of healthy growth, such as soluble proteins, sugars, vitamin C, and malondialdehyde (MDA). The results showed that WD promoted basil development, whereas the presence of bioplastic hindered it. Interestingly, WD did not affect sugars but increased vitamin C by 12 %, which is considered a positive effect as changes in sugar levels could indicate plant stress. In contrast, bioplastic resulted in reduced sugars (-41 %) and increased (+17 %) MDA level, while vitamin C content remained unchanged. However, when WD was added to plants grown with bioplastic, it elevated the levels of all examined parameters, except for sugars and vitamin C, which experienced reductions (-66 % and 33 %, respectively). Intriguingly, despite this reduction, the observed direct correlation between sugar and vitamin C contents was maintained, indicating that the decrease in sugar content may have reached a critical threshold. This study suggests that the use of WD has the potential to alleviate the adverse effects of bioplastic on basil growth and development and highlights the importance of adopting sustainable practices in agriculture, as well as the need for a critical assessment of the environmental impact of new technologies and products.

Effect of Biochar Type, Concentration and Washing Conditions on the Germination Parameters of Three Model Crops.

Biochar has been recognized as a promising and efficient material for soil amendment. However, its effects on seed germination are variable due to its alkaline pH and/or the presence of phytotoxic substances. In this study, two types of biochar (B1 and B2) were mixed with soil at different concentrations (0%, 5%, 10%, 25%, 50% and 100%, w:w), and both the solid and liquid fractions of these mixtures were tested on the germination of basil, lettuce and tomato seeds. Furthermore, solid fractions subjected to a pre-washing treatment (B1W and B2W) were also investigated for their effects on seed germination. Three germination parameters were then measured: seed germination number (GN), radicle length (RL) and germination index (GI). Biochar B2W at 10% increased both RL and GI in basil by 50% and 70%, respectively, while B1 at 25% increased these parameters in tomato by 25%. No effects or negative effects were recorded for lettuce. Liquid fractions (L1 and L2) generally hampered seed germination, suggesting the presence of potentially water-soluble phytotoxic compounds in biochar. These results point to biochar as a suitable component for germination substrates and highlight that germination tests are critical to select the best performing biochar according to the target crop.

Interaction between Sulfate and Selenate in Tetraploid Wheat (Triticum turgidum L.) Genotypes.

Selenium (Se) is an essential micronutrient of fundamental importance to human health and the main Se source is from plant-derived foods. Plants mainly take up Se as selenate (SeO42-), through the root sulfate transport system, because of their chemical similarity. The aims of this study were (1) to characterize the interaction between Se and S during the root uptake process, by measuring the expression of genes coding for high-affinity sulfate transporters and (2) to explore the possibility of increasing plant capability to take up Se by modulating S availability in the growth medium. We selected different tetraploid wheat genotypes as model plants, including a modern genotype, Svevo (Triticum turgidum ssp. durum), and three ancient Khorasan wheats, Kamut, Turanicum 21, and Etrusco (Triticum turgidum ssp. turanicum). The plants were cultivated hydroponically for 20 days in the presence of two sulfate levels, adequate (S = 1.2 mM) and limiting (L = 0.06 mM), and three selenate levels (0, 10, 50 µM). Our findings clearly showed the differential expression of genes encoding the two high-affinity transporters (TdSultr1.1 and TdSultr1.3), which are involved in the primary uptake of sulfate from the rhizosphere. Interestingly, Se accumulation in shoots was higher when S was limited in the nutrient solution.

The suppression of TdMRP3 genes reduces the phytic acid and increases the nutrient accumulation in durum wheat grain.

Micronutrient malnutrition affects more than half of the world population. Reduced bioavailability of microelements in the raw materials is considered one of the main causes of mineral deficiency in populations whose diet is largely based on the consumption of staple crops. In this context, the production of low phytic acid (lpa) cereals is a main goal of the breeding programs, as phytic acid (PA) binds essential mineral cations such as iron (Fe), zinc (Zn), manganese (Mn), potassium (K), calcium (Ca) and magnesium (Mg) precipitating in the form of phytate salts poorly digested by monogastric animals, including humans, due to the lack of phytases in the digestive tract. Since PA limits the bioavailability of microelements, it is widely recognized as an anti-nutritional compound. A Targeting Induced Local Lesions IN Genomes (TILLING) approach has been undertaken to silence the genes encoding the TdABCC13 proteins, known as Multidrug-Resistance associated Proteins 3 (TdMRP3), transporters involved in the accumulation of PA inside the vacuole in durum wheat. The TdMRP3 complete null genotypes showed a significant reduction in the content of PA and were able to accumulate a higher amount of essential micronutrients (Fe, Zn, Mn) compared to the control. The number of spikelets and seeds per spike, traits associated with the agronomic performances, were reduced compared to the control, but the negative effect was in part balanced by the increased grain weight. The TdMRP3 mutant lines showed morphological differences in the root apparatus such as a significant decrease in the number of root tips, root length, volume and surface area and an increase in root average diameter compared to the control plants. These materials represent a promising basis for obtaining new commercial durum wheats with higher nutritional value.

Biochar improves the performance of Avena sativa L. grown in gasoline-polluted soils.

This study investigated the effect of soil contamination by different concentrations of gasoline on oat (Avena sativa L.) and tested the effect of biochar supply to the polluted soils on the performance of oat plants. Oat seeds were sowed in contaminated soils with different concentrations of gasoline: 0% (control), 1%, 2%, 6%, and 10% (v/w), and grown for 2 weeks. Germination, fresh weight, root and stem length, photosynthetic parameters (i.e., chlorophyll content, PIABS, FV/FM, and NDVI), and total antioxidant power were analyzed. The results showed a remarkable negative effect on almost all the investigated parameters starting from the gasoline concentration of 6%. Based on these results, a new experiment was run by adding 5% (w/w) biochar (a carbon-rich byproduct of wood biomass pyrolysis) to the 6% and 10% polluted soils to test whether adding biochar had a beneficial effect on oat performance. The results showed that biochar supply greatly reduced the negative effects caused by gasoline on all the investigated parameters.

Fire-induced effects on the bioavailability of potentially toxic elements in a polluted agricultural soil: implications for Cr uptake by durum wheat plants.

Fire events can modify the distribution and speciation of potentially toxic elements (PTEs) in soil, especially if they are associated to organic matter (OM). In fact, OM can undergo substantial structural modifications at high temperatures, up to the complete mineralization. The present study aims to investigate the changes of PTEs' bioavailability to durum wheat (Triticum durum Desf.) plants after simulating fire events (up to 300 °C and 500 °C) in an agricultural soil polluted by Cr, Zn, Cu, and Pb. The PTEs' uptake and allocation in plant tissues were assessed using the RHIZOtest system. After the fire simulations, no evident risk of accumulation and translocation in plants was observed for Zn, Pb, and Cu. Conversely, a high accumulation in roots and a significant translocation to shoots were observed for Cr, which reached concentrations of 829 mg kg-1 in roots and 52 mg kg-1 in shoots at 500 °C. Additional experimental evidence suggested that Cr was taken up by plants grown on heated soils as Cr(VI). Once acquired by roots, only a small part of Cr (up to 6%) was translocated to shoots where it was likely present as mobile forms, as evidenced by micro X-ray fluorescence (µ-XRF) analyses. Overall, the results obtained provide evidence that the high temperatures occurring during fire events can increase the mobility and bioavailability of certain PTEs transforming apparently safe environments into potentially dangerous sources of pollution. These processes can ultimately affect the human health through the food chain transfer of PTEs or their migration into surface water and groundwater.

Impact of starch-based bioplastic on growth and biochemical parameters of basil plants.

The recent use of bioplastics in agriculture is considered an ecological choice, aimed at limiting the environmental impact of plastics, in line with the Sustainable Development Goals of the United Nations. However, the impact of bioplastic residues on the environment is unclear as knowledge is lacking. This is the first study investigating the effect of a starch-based bioplastic on the growth and biochemical parameters of basil. Bioplastic was experimentally prepared and added to the soil at 2.5 % (w/w), corresponding to twice the concentration of plastic mulch film residues currently found in cultivated soils, in view of the increasing agricultural use of bioplastics. Basil plants were grown without (controls) and with bioplastic addition for 35 days, under controlled experimental conditions. Compared to the control, plants exposed to bioplastic showed stunted growth (in terms of shoot fresh weight, height, and number of leaves). Significant reductions in the content of chlorophyll, protein, ascorbic acid, and glucose were also observed. Finally, the treatment caused oxidative stress, as evidenced by the increased content of malondialdehyde in the shoots. The addition of bioplastic increased the electrical conductivity and reduced the cation exchange capacity of the cultivation soil. These results suggest that bioplastic in soil may promote the onset of stressful conditions for plant growth in a similar manner to plastic. They will be complemented by further investigations to unravel the mechanisms underlying these responses, involving different doses and types of bioplastics and other crop species.

Interaction Between Sulfur and Iron in Plants.

It is well known that S interacts with some macronutrients, such as N, P, and K, as well as with some micronutrients, such as Fe, Mo, Cu, Zn, and B. From our current understanding, such interactions could be related to the fact that: (i) S shares similar chemical properties with other elements (e.g., Mo and Se) determining competition for the acquisition/transport process (SULTR transporter family proteins); (ii) S-requiring metabolic processes need the presence of other nutrients or regulate plant responses to other nutritional deficiencies (S-containing metabolites are the precursor for the synthesis of ethylene and phytosiderophores); (iii) S directly interacts with other elements (e.g., Fe) by forming complexes and chemical bonds, such as Fe-S clusters; and (iv) S is a constituent of organic molecules, which play crucial roles in plants (glutathione, transporters, etc.). This review summarizes the current state of knowledge of the interplay between Fe and S in plants. It has been demonstrated that plant capability to take up and accumulate Fe strongly depends on S availability in the growth medium in both monocots and dicot plants. Moreover, providing S above the average nutritional need enhances the Fe content in wheat grains, this beneficial effect being particularly pronounced under severe Fe limitation. On the other hand, Fe shortage induces a significant increase in the demand for S, resulting in enhanced S uptake and assimilation rate, similar to what happens under S deficiency. The critical evaluation of the recent studies on the modulation of Fe/S interaction by integrating old and new insights gained on this topic will help to identify the main knowledge gaps. Indeed, it remains a challenge to determine how the interplay between S and Fe is regulated and how plants are able to sense environmental nutrient fluctuations and then to adapt their uptake, translocation, assimilation, and signaling. A better knowledge of the mechanisms of Fe/S interaction might considerably help in improving crop performance within a context of limited nutrient resources and a more sustainable agriculture.

Evaluating the Aqueous Phase From Hydrothermal Carbonization of Cow Manure Digestate as Possible Fertilizer Solution for Plant Growth.

Improving the agronomic use of recycled nutrients derived from organic waste is one of the priorities within the measures adopted by the European community to reduce environmental issues but remains an unexplored area of research. This study focused on investigating the possibility of using innovative fertilizer solutions in hydroponic systems for the growth of agricultural plants. To this purpose, a liquid fraction [aqueous hydrothermal carbonization (HTC) liquid (AHL)] derived from HTC of cow manure digestate was chemically characterized (pH, electrical conductivity, mineral elements, and organic compounds such as phytotoxins), diluted with distilled water (1:30, 1:60, and 1:90, v/v) to reduce its potential phytotoxicity, and used to grow hydroponic maize (Zea mays L.) plants instead of the classical full-strength nutrient solution. The results indicated that the dilution ratio 1:30 of the AHL solution maintained a high level of toxicity for the plants (phytotoxic substances, especially Na and alkalinity), inducing the arrest of their growth. Differently, the two other dilution ratios (i.e., 1:60 and 1:90) seemed to considerably limit the levels of toxicity, since they allowed the plants to develop. However, these dilution ratios were poor in nutrient elements, inducing alteration in photosynthesis and an onset of deficiency symptoms such as pronounced leaf chlorosis. In view of an eco-friendly approach, future studies are, therefore, needed to identify the correct species-specific dilution ratio to supply both low levels of phytotoxins and adequate content of essential nutrients for appropriate plant growth and development. Furthermore, in order to lower specific Na phytotoxicity, treatments are of utmost importance before using AHL as a fertilizer solution.

Corrigendum: Root Handling Affects Carboxylates Exudation and Phosphate Uptake of White Lupin Roots.

[This corrects the article DOI: 10.3389/fpls.2020.584568.].

Phytotoxicity of hydrochars obtained by hydrothermal carbonization of manure-based digestate.

The management of digestate, the main by-product of the anaerobic digestion (AD) process, is one of the most serious environmental issues. Although digestate is used on arable land as a fertilizer, it can have a negative impact on the environment due to nitrate leaching into the groundwater and ammonia volatilization into the atmosphere, with high economic and environmental disposal costs. Therefore, hydrothermal carbonization (HTC), a thermochemical biomass conversion process, could represent a sustainable and efficient alternative for digestate management. Hydrochar, the solid product of the HTC process, has been recently proposed as a plant growing medium in soilless culture systems (SCS). Here, using cow manure digestate as feedstock, we investigated the influence of the HTC process reaction temperature (180, 220 and 250 °C) and residence time (1 and 3 h) on the physical-chemical properties (pH, electrical conductivity, and mineral element concentrations) of the resulting hydrochars. Furthermore, in order to fully valorize hydrochar as a growing medium, their possible phytotoxic effects and those of their water extracts (prepared at two different concentrations and at different pHs) were tested in germination tests with cress seeds (Lepidium sativum L.). Concentrations of nutrients, heavy metals, organic acids, sugars and furan compounds were determined in the water extracts. Characterization analysis of these hydrochars revealed that they can be distinguished from each other by their physical-chemical properties, which were significantly affected by the two process parameters. Specifically, the HTC temperature had a greater effect on the composition of hydrochars than the residence time. Germination tests found hydrochar water extracts to show significantly lower phytotoxicity than the hydrochars themselves. Notably, the phytotoxic effect of the extracts decreased with increasing extraction ratio and decreasing pH. The chromatographic characterization of extracts identified the presence of potential phytotoxins, such as furan compounds (i.e., hydroxymethylfurfural and furfural). However, before using hydrochars as potential and innovative growing media for plants, their phytotoxicity should be limited, for example through their dilution with other substrates. Overall, AD-HTC coupling could represent a valuable eco-sustainable expedient in the field of biomasses, green economy and waste conversion and, therefore, further investigations in this direction are necessary.

Root Handling Affects Carboxylates Exudation and Phosphate Uptake of White Lupin Roots.

The reliable quantification of root exudation and nutrient uptake is a very challenging task, especially when considering single root segments. Most methods used necessitate root handling e.g. root dissecting/cutting. However, there is a knowledge gap on how much these techniques affect root physiology. Thus, this study aimed at assessing the effect of different root handling techniques on the phosphate (Pi) uptake and carboxylate exudation of white lupin roots. White lupin plants were grown hydroponically in a full and Pi-deficient nutrient solution for 60 days. Phosphate uptake and carboxylate exudation of cluster and non-cluster roots were measured using custom made cells 1, 4, and 8 h after the onset of light. Three different experimental set-ups were used: i) without cutting the root apparatus from the shoots, nor dissecting the root into smaller root sections - named intact plant (IP); ii) separating the roots from the shoots, without dissecting the root into smaller sections - named intact root (IR); iii) separating the roots form the shoots and dissecting the roots in different sections-named dissected roots (DR). The sampling at 8 h led to the most significant alterations of the root Pi uptake induced by the sampling method. Generally, roots were mainly affected by the DR sampling method, indicating that results of studies in which roots are cut/dissected should be interpreted carefully. Additionally, the study revealed that the root tip showed a very high Pi uptake rate, suggesting that the tip could act as a Pi sensor. Citrate, malate and lactate could be detected in juvenile, mature and senescent cluster root exudation. We observed a significant effect of the handling method on carboxylate exudation only at sampling hours 1 and 8, although no clear and distinctive trend could be observed. Results here presented reveal that the root handling as well as the sampling time point can greatly influence root physiology and therefore should not be neglected when interpreting rhizosphere dynamics.

Phosphorus deficiency changes carbon isotope fractionation and triggers exudate reacquisition in tomato plants.

Plant roots are able to exude vast amounts of metabolites into the rhizosphere in response to phosphorus (P) deficiency. Causing noteworthy costs in terms of energy and carbon (C) for the plants. Therefore, it is suggested that exudates reacquisition by roots could represent an energy saving strategy of plants. This study aimed at investigating the effect of P deficiency on the ability of hydroponically grown tomato plants to re-acquire specific compounds generally present in root exudates by using 13C-labelled molecules. Results showed that P deficient tomato plants were able to take up citrate (+ 37%) and malate (+ 37%), particularly when compared to controls. While glycine (+ 42%) and fructose (+ 49%) uptake was enhanced in P shortage, glucose acquisition was not affected by the nutritional status. Unexpectedly, results also showed that P deficiency leads to a 13C enrichment in both tomato roots and shoots over time (shoots-+ 2.66‰, roots-+ 2.64‰, compared to control plants), probably due to stomata closure triggered by P deficiency. These findings highlight that tomato plants are able to take up a wide range of metabolites belonging to root exudates, thus maximizing C trade off. This trait is particularly evident when plants grew in P deficiency.

Selected Plant-Related Papers from the First Joint Meeting on Soil and Plant System Sciences (SPSS 2019)-"Natural and Human-Induced Impacts on the Critical Zone and Food Production".

The First Joint Meeting on Soil and Plant System Sciences (SPSS 2019), titled "Natural and Human-Induced Impacts on the Critical Zone and Food Production", aimed at integrating different scientific backgrounds and topics flowing into the Critical Zone, where chemical, biological, physical, and geological processes work together to support life on the Earth's surface. The SPSS 2019 meeting gathered the thoughts and findings of scientists, professionals and individuals from different countries working in different research fields. This Special Issue comprises a selection of original works on the plant-related topics presented during this international meeting.

Single and Combined Fe and S Deficiency Differentially Modulate Root Exudate Composition in Tomato: A Double Strategy for Fe Acquisition?

Fe chlorosis is considered as one of the major constraints on crop growth and yield worldwide, being particularly worse when associated with S shortage, due to the tight link between Fe and S. Plant adaptation to inadequate nutrient availabilities often relies on the release of root exudates that enhance nutrients, mobilization from soil colloids and favour their uptake by roots. This work aims at characterizing the exudomic profile of hydroponically grown tomato plants subjected to either single or combined Fe and S deficiency, as well as at shedding light on the regulation mechanisms underlying Fe and S acquisition processes by plants. Root exudates have been analysed by untargeted metabolomics, through liquid chromatography-mass spectrometry as well as gas chromatography-mass spectrometry following derivatization. More than 200 metabolites could be putatively annotated. Venn diagrams show that 23%, 10% and 21% of differential metabolites are distinctively modulated by single Fe deficiency, single S deficiency or combined Fe-S deficiency, respectively. Interestingly, for the first time, a mugineic acid derivative is detected in dicot plants root exudates. The results seem to support the hypothesis of the co-existence of the two Fe acquisition strategies in tomato plants.

Revisiting Fe/S interplay in tomato: A split-root approach to study the systemic and local responses.

Based on our previous studies demonstrating an intriguing interplay between sulfur (S) and iron (Fe), a split-root experiment was performed to determine whether plant S status and/or S external concentration could modify plant capability to take up and accumulate Fe. This split-root system allowed the roots of each tomato plant to grow in two different compartments, both Fe-deficient, but one S-sufficient, and the other one S-free. Although S was freely available to half root system and thus plant S status was preserved, S-deficient part of root apparatus exhibited a decrease of total S, thiols and protein content, an enhanced activity of both ATPsulfurylase and O-acetylserine(thiol)lyase, and a higher expression of SlST1.1, as occurring under S deficiency. The side of the root apparatus exposed to combined S and Fe deficiency, showed an over induction of the FeIII-reducing capacity (+40%) and of the expression levels of the gene codifying for this protein (SlFRO1), with respect to the Fe-deficient part of the root system. Interestingly, the regulation pattern of the bHLH transcription factor SlFER, controlling the expression of both SlFRO1 and SlIRT1 genes, was very close to that of SlFRO1. SlIRT1 expression levels appeared unaffected by S supply, suggesting distinct regulatory processes targeting SlFRO1 and SlIRT1.

Mitochondria dysfunctions under Fe and S deficiency: is citric acid involved in the regulation of adaptive responses?

Within the last years, extensive information has been accumulated on the reciprocal influence between S and Fe nutrition at both physiological and molecular level in several plant species, but the mechanisms regulating S and Fe sensing and signaling are not fully understood. Fe and S interact for the building of Fe-S clusters, and mitochondria is one of the cellular compartments where Fe-S cluster assembly takes place. Therefore, it would be expected that mitochondria might play a central role in the regulation of Fe and S interaction. The Fe deficiency-induced alteration in the synthesis of mitochondria-derived carboxylic acids, such as citric acid, and the evidence that such molecules have already been identified as important players of metabolite signaling in several organisms, further support this hypothesis. Tomato plants were grown under single or combined Fe and S deficiency with the aim of verifying whether mitochondria activities played a role in Fe/S interaction. Both Fe and S deficiencies determined similar alteration of respiratory chain activity: a general decrease of Fe-S containing complexes as well as an increase of alternative NAD(P)H activities was observed in both Fe and S deficient-plants. However, the content of Krebs cycle-related organic acids in roots was substantially different in response to treatments, being the accumulation of citric acid always increased, while the others (i.e. succinic, malic, fumaric acids) always decreased. Interestingly, citric acid levels significantly correlated with the expression of some Fe and S deficiency induced genes. Our results contribute to existing knowledge on the complexity of the S/Fe interaction, suggesting a model in which endogenous alteration of citric acid content in plant tissues might act as signal molecule for the regulation of some nuclear-encoded and nutrient-responsive genes and also provide a basis for further study of the mechanism underlying S and Fe sensing and signalling.