Epigenetic changes induced by chronic and acute chromium stress treatments in Arabidopsis thaliana identified by the MSAP-Seq.

Chromium (Cr) is an highly toxic metal to plants and causes severe damage to their growth, development, and reproduction. Plant exposure to chronic and acute Cr stress treatments results in significant changes at short time in the gene expression profile and at long time in the genomic DNA methylation profile at a transgenerational level and, consequently, in gene expression. These epigenetic modifications and their implications imposed by the Cr stress are not yet completely known in plants. Herein, were identified the epigenetic changes induced by chronic and acute Cr stress treatments in Arabidopsis thaliana plants using Methylation Sensitive Amplification Polymorphism coupled with next-generation sequencing (MSAP-Seq). First-generation Arabidopsis plants (termed F0 plants) kept under hoagland solution were subjected to Cr stress treatments. For chronic Cr stress, plants were treated through hoagland solution with 2.5 µM Cr during the entire cultivation period until seed harvest. Meanwhile, for acute Cr stress, plants were treated with 5 µM Cr during the first three weeks and returned to unstressful control condition until seed harvest. Seeds from F0 plants were sown and F1 plants were re-submitted to the same Cr stress treatments. The seed germination rate was evaluated from F-2 seeds harvested of F1 plants kept under different Cr stress treatments (0, 10, 20, and 40 µM) compared to the unstressful control condition. These data showed significant changes in the germination rate of F-2 seeds originating from stressed F1 plants compared to F-2 seeds harvested from unstressful control plants. Given this data, F1 plants kept under these chronic and acute Cr stress treatments and unstressful control condition were evaluated for the transgenerational epigenetic modifications using MSAP-Seq. The MSAP-Seq data showed that several genes were modified in their methylation status as a consequence of chronic and acute Cr stress treatment to maintain plant defenses activated. In particular, RNA processing, protein translation, photorespiration, energy production, transmembrane transport, DNA transcription, plant development, and plant resilience were the major biological processes modulated by epigenetic mechanisms identified in F1 plants kept under chronic and acute Cr stress. Therefore, collective data suggested that Arabidopsis plants kept under Cr stress regulate their epigenetic status over generations based on DNA methylation to modulate defense and resilience mechanisms.

Impact of airborne iron oxide nanoparticles on Tillandsia usneoides as a model plant to assess pollution in heavy traffic areas.

Due to the increasing evidence of widespread sub-micron pollutants in the atmosphere, the impact of airborne nanoparticles is a subject of great relevance. In particular, the smallest particles are considered the most active and dangerous, having a higher surface/volume ratio. Here we tested the effect of iron oxide (Fe3O4) nanoparticles (IONPs) with different mean diameter and size distribution on the model plant Tillandsia usneoides. Strands were placed in home-built closed boxes and exposed to levels of airborne IONPs reported for the roadside air, i.e. in the order of 107 - 108 items m-2. Plant growth and other morpho-physiological parameters were monitored for two weeks, showing that exposure to IONPs significantly reduced the length increment of the treated strands with respect to controls. A dose-dependence of this impairing effect was found only for particles with mean size of a few tens of nanometers. These were also proved to be the most toxic at the highest concentration tested. The IONP-induced hamper in growth was correlated with altered concentration of macro- and micronutrients in the plant, while no significant variation in photosynthetic activity was detected in treated samples. Microscopy investigation showed that IONPs could adhere to the plant surface and were preferentially located on the trichome wings. Our results report, for the first time, evidence of the negative effects of airborne IONP pollution on plant health, thus raising concerns about related environmental risks. Future research should be devoted to other plant species and pollutants to assess the impact of airborne pollution on plants and devise suitable attenuation practices.

Biochar from co-pyrolysis of biological sludge and sawdust in comparison with the conventional filling media of vertical-flow constructed wetlands for the treatment of domestic-textile wastewater.

A biochar from co-pyrolysis of a mixture of sawdust and biological sludge (70/30, w/w), providing a high environmental compatibility in terms of water leachable polycyclic aromatic hydrocarbons and inorganic elements, together with a remarkable surface area (389 m2/g), was integrated into laboratory-scale vertical-flow constructed wetlands (VF-CWs), planted with Phragmites australis and unplanted. Biochar-filled VF-CWs have been tested for 8 months for the refining of effluents from the tertiary clariflocculation stage of a wastewater treatment plant operating in a mixed domestic-industrial textile context, in comparison with systems filled with gravel. VF-CW influents and effluents were monitored for chemical oxygen demand (COD), nitrogen and phosphorus cycles, and absorbance values at 254 and 420 nm, the latter as rapid and reliable screening parameters of the removal of organic micropollutants containing aromatic moieties and/or chromophores. Biochar-based systems provided a statistically significant improvement in COD (Δ = 22%) and ammonia (Δ = 35%) removal, as well as in the reduction of UV-Vis absorbance values (Δ = 32-34% and Δ = 28% for 254 and 420 nm, respectively), compared to gravel-filled microcosms. The higher removal of organic was mainly attributed to the well-known adsorption properties of biochars, while for nitrogen the biological mechanisms seem to play a predominant role.

Growth, physiological parameters and DNA methylation in Spirodela polyrhiza (L.) Schleid exposed to PET micro-nanoplastic contaminated waters.

The effects of polyethylene terephthalate micro-nanoplastics (PET-MNPs) were tested on the model freshwater species Spirodela polyrhiza (L.) Schleid., with focus on possible particle-induced epigenetic effects (i.e. alteration of DNA methylation status). MNPs (size âˆ¼ 200-300 nm) were produced as water dispersions from PET bottles through repeated cycles of homogenization and used to prepare N-medium at two environmentally relevant concentrations (∼0.05 g L-1 and ∼0.1 g L-1 of MNPs). After 10 days of exposure, a reduction in fresh and dry weight was observed in treated plants, even if the average specific growth rate for both frond number and area was not altered. Impaired growth was coupled with a MNP-induced decrease of chlorophyll fluorescence parameters (i.e. ΨETo and Piabs, indicators of photochemical efficiency) and starch concentration, as well as with alterations in plant ionomic profile and oxidative status. The methylation-sensitive amplification polymorphism (MSAP) technique was used to assess possible changes in DNA methylation levels induced by plastic particles. The analysis showed unusual hypermethylation in 5'-CCGG sites that could be implicated in DNA protection from dangerous agents (i.e. reactive oxygen species) or in the formation of new epialleles. This work represents the first evidence of MNP-induced epigenetic modifications in the plant world.

Adverse Effects of Micro- and Nanoplastics on Humans and the Environment.

The pervasive pollution caused by nano- and microplastics (N/MPLs) is a pressing concern, and was exacerbated during the COVID-19 pandemic due to the substantial release of disposable Personal Protective Equipment (PPE) into the environment [...].

Metabolic Responses in Leaves of 15 Italian Olive Cultivars in Correspondence to Variable Climatic Elements.

This study aims to evaluate the metabolic changes that occurred in olive leaves as responses over time to variations in climatic elements. Rainfall, temperature, and solar radiation data were collected over 4 months (August-November) to assess the impact of different climatic trends on the metabolism of the leaves of 15 Italian olive cultivars, cultivated at the experimental farm of the University of Florence. The net photosynthetic rate (AN) and stomatal conductance (gs), measured as main indicators of primary metabolism, were mainly influenced by the "cultivar" effect compared to the "climate" effect. The lowest AN value was showed by "Bianchera", while "Ascolana" recorded the highest (8.6 and 13.6 µmol CO2 m-2s-1, respectively). On the other hand, the secondary metabolism indicators, volatile organic compound (VOC) and oleuropein (OL) content, were much more influenced by climate trends, especially rainfall. A phase of high rainfall caused a significant increase in the VOCs emission from leaves, even with different behaviors among the genotypes. The highest differences were observed between "Maiatica di Ferrandina", with the highest average values (~85,000 npcs), and "Frantoio", which showed the lowest (~22,700 npcs). The OL content underwent considerable fluctuations in relation to the rainfall but also appeared to be controlled by the genotype. "Coratina" always showed the highest OL concentration (reaching the maximum ~98 mg g-1), indicating the great potential of this cultivar for the industrial recovery of OL.

Can microplastics threaten plant productivity and fruit quality? Insights from Micro-Tom and Micro-PET/PVC.

Solanum lycopersicum L., a crop grown worldwide with a high nutritional value for the human diet, was used to test the impact of microplastics on plant growth, productivity, and fruit quality. Two of the most represented microplastics in soils, polyethylene terephthalate (PET) and polyvinyl chloride (PVC), were tested. Plants were grown in pots with an environmentally realistic concentration of microplastics and, during the whole crop life cycle, photosynthetic parameters, number of flowers and fruits were monitored. At the end of the cultivation, plant biometry and ionome were evaluated, along with fruit production and quality. Both pollutants had negligible effects on shoot traits, with only PVC causing a significant reduction in shoot fresh weight. Despite an apparent low or no toxicity during the vegetative stage, both microplastics decreased the number of fruits and, in the case of PVC, also their fresh weights. The plastic polymer-induced decline in fruit production was coupled with wide variations in fruit ionome, with marked increases in Ni and Cd. By contrast there was a decline in the nutritionally valuable lycopene, total soluble solids, and total phenols. Altogether, our results reveal that microplastics can not only limit crop productivity but also negatively impact fruit quality and enhance the concentration of food safety hazards, thus raising concerns for their potential health risks for humans.

In vitro and in vivo copper-treated Myrtus communis L.: terpene profiles and evidence for potential cultivation on metal-contaminated soils.

The intensive application of agro-chemicals, and in particular of copper (Cu)-based compounds, causes increasing pollution of agricultural land, with serious risks for human health. Aromatic plants used for purposes other than food, can be considered for the exploitation and/or remediation of metal-polluted substrates, since contamination of the essential oils by these metals was not significant. Myrtle (Myrtus communis L.) is a Mediterranean evergreen shrub whose essential oil has many commercial applications. In this work, the effect of an excess of Cu in respect to control conditions was assessed on M. communis growth and foliar terpene composition. Metal accumulation in roots and shoots was also evaluated for the possible use of this species in phytoremediation. The amount of Cu applied in our experiments minimally affected the terpene profiles of in vitro grown plants, whereas no variations were detected in in vivo plants. The presence of the metal in the soil did not significantly impair plant growth, thus allowing its cultivation on polluted substrates. On the other hand, the amount of Cu in the plant was not enough to result in a significant reduction of Cu levels in the soil. Therefore, myrtle plants proved to be good candidates for the re-vegetation of Cu-contaminated lands.

Transgenerational effects of chromium stress at the phenotypic and molecular level in Arabidopsis thaliana.

In this study, we describe the results obtained in a study of the transgenerational phenotypic effects of chromium (Cr) stress on the model plant species Arabidopsis thaliana. The F1 generation derived from parents grown under chronic and medium chronic stress showed significantly higher levels of the maximal effective concentration (EC50) compared with F1 plants generated from unstressed parents. Moreover, F1 plants from Cr-stressed parents showed a higher germination rate when grown in the presence of Cr. F1 plants derived from parents cultivated under chronic Cr stress displayed reduced hydrogen peroxide levels under Cr stress compared to controls. At lower Cr stress levels, F1 plants were observed to activate promptly more genes involved in Cr stress responses than F0 plants, implying a memory effect linked to transgenerational priming. At higher Cr levels, and at later stages, F1 plants modulated significantly fewer genes than F0 plants, implying a memory effect leading to Cr stress adaptation. Several bHLH transcription factors were induced by Cr stress in F1 but not in F0 plants, including bHLH100, ORG2 and ORG3. F1 plants optimized gene expression towards pathways linked to iron starvation response. A model of the transcriptional regulation of transgenerational memory to Cr stress is presented here, and could be applied for other heavy metal stresses.

Interplay of plastic pollution with algae and plants: hidden danger or a blessing?

In the era of plastic pollution, plants have been discarded as a system that is not affected by micro and nanoplastics, but contrary to beliefs that plants cannot absorb plastic particles, recent research proved otherwise. The presented review gives insight into known aspects of plants' interplay with plastics and how plants' ability to absorb plastic particles can be utilized to remove plastics from water and soil systems. Microplastics usually cannot be absorbed by plant root systems due to their size, but some reports indicate they might enter plant tissues through stomata. On the other hand, nanoparticles can enter plant root systems, and reports of their transport via xylem to upper plant parts have been recorded. Bioaccumulation of nanoplastics in upper plant parts is still not confirmed. The prospects of using biosystems for the remediation of soils contaminated with plastics are still unknown. However, algae could be used to degrade plastic particles in water systems through enzyme facilitated degradation processes. Considering the amount of plastic pollution, especially in the oceans, further research is necessary on the utilization of algae in plastic degradation. Special attention should be given to the research concerning utilization of algae with restricted algal growth, ensuring that a different problem is not induced, "sea blooming", during the degradation of plastics.

Plastic is in the air: Impact of micro-nanoplastics from airborne pollution on Tillandsia usneoides (L.) L. (Bromeliaceae) as a possible green sensor.

Due to the increasing evidence of widespread plastic pollution in the air, the impact on plants of airborne particles of polycarbonate (PC), polyethyleneterephthalate (PET), polyethylene (PE), and polyvinylchloride (PVC) was tested by administering pristine and aged airborne micro-nanoplastics (MNPs) to Tillandsia usneoides for two weeks. Here we showed that exposure to pristine MNPs, significantly reduced plant growth with respect to controls. Particularly, PVC almost halved plant development at the end of the treatment, while the other plastics exerted negative effects on growth only at the beginning of the exposure, with final stages comparable to those of controls. Plants exposed to aged MNPs showed significantly decreased growth at early stages with PC, later in the growth with PE, and even later with PET. Aged PVC did not exert a toxic effect on plants. When present, the plastic-mediated reduction in plant growth was coupled with a decrease in photosynthetic activity and alterations in the plant concentration of macro- and micronutrients. The plastic particles were showed to adhere to the plant surface and, preferentially, on the trichome wings. Our results reported, for the first time, evidence of negative effects of airborne plastic pollution on plant health, thus raising concerns for related environmental risks.

Improving plant-based genotoxicity bioassay through AFLP technique for trace metal-contaminated water: insights from Myriophyllum aquaticum (Vell.) Verdc. and Cd.

In this work, we evaluated whether the species Myriophyllum aquaticum (Vell.) Verdc. can be a promising material for devising reliable eco-toxicological tests for Cd-contaminated waters. Plants of M. aquaticum were exposed to Cd, using different concentrations (1 mg L-1, 2.5 mg L-1, 5 mg L-1, and 10 mg L-1; experiment 1) and exposure times (2.5 mg L-1 for 3 days, 7 days, 14 days, and 21 days; experiment 2). Plant growth and Cd accumulation were monitored during the treatment period, and Cd genotoxicity was assessed by analyzing Cd-induced changes in the AFLP fingerprinting profiles using famEcoRI(TAC)/MseI(ATG) and hexEcoRI(ACG)/MseI(ATG) pairs of primers. Root and shoot growth was reduced already at the lowest Cd concentration used (about 20% reduction for roots and 60% for shoots at 1 mg L-1; experiment 1) and after 7 days (about 50% reduction for roots and 70% for shoots; experiment 2). The primer combinations produced 154 and 191 polymorphic loci for experiments 1 and 2, respectively. Mean genetic diversity (He) reduction among the treatment groups was observed starting from 2.5 mg L-1 (He 0.211 treated vs 0.236 control; experiment 1) and after 3 days (He 0.169 treated vs 0.261 control; experiment 2), indicating that results obtained from AFLP profiles did not match with plant growth measurements. Therefore, our results showed that M. aquaticum proved to be a suitable model system for the investigation of Cd genotoxicity through AFLP fingerprinting profile, whereas the more classic eco-toxicological tests based only on biometric parameters could not correctly estimate the risk associated with undetected Cd genotoxicity.

Photosynthesizing while hyperaccumulating nickel: Insights from the genus Odontarrhena (Brassicaceae).

Nickel-induced changes in photosynthetic activity were investigated in three Ni-hyperaccumulating Odontarrhena species with increasing Ni tolerance and accumulation capacity, O. muralis, O. moravensis, and O. chalcidica. Plantlets were grown in hydroponics at increasing NiSO4 concentrations (0, 0.25, and 1 mM) for one week, and the effects of Ni on growth, metal accumulation, photosynthesis, and nitrogen (N) allocation to components of the photosynthetic apparatus were analysed. Nickel treatments in O. chalcidica, and O. moravensis to a lesser extent, increased not only the photochemical efficiency of photosystem II (PSII) and the CO2 assimilation rate, but also CO2 diffusion from the atmosphere to the carboxylation sites. These two species displayed a specific increase and/or rearrangement of the photosynthetic pigments and a higher leaf N allocation to the photosynthetic components in the presence of the metal. Odontarrhena muralis displayed a decrease in photosynthetic performance at the lowest Ni concentration due to a combination of both stomatal and non-stomatal limitations. Our data represent the first complete investigation of the effects of Ni on the photosynthetic machinery in Ni hyperaccumulating plants. Our findings clearly indicate a stimulatory, hormetic-like, effect of the metal on both biophysics and biochemistry of photosynthesis in the species with the highest hyperaccumulation capacity.

Impact of microplastics on growth, photosynthesis and essential elements in Cucurbita pepo L.

In this study, Cucurbita pepo L., one of the most cultivated, consumed and economically important crop worldwide, was used as model plant to test the toxic effects of the four most abundant microplastics identified in contaminated soils, i.e. polypropylene (PP), polyethylene (PE), polyvinylchloride (PVC), and polyethyleneterephthalate (PET). Cucurbita plants were grown in pots with increasing concentrations of the microplastics, then plant biometry, photosynthetic parameters and ionome of treated vs. untreated samples were compared to evaluate the toxicity of each plastic. All the pollutants impaired root and, especially, shoot growth. Specific and concentration-dependant effects of the different microplastics were found, including reduction in leaf size, chlorophyll content and photosynthetic efficiency, as well as changes in the micro- and macro-elemental profile. Among all the microplastics, PVC was identified as the most toxic and PE as the less toxic material. PVC decreased the dimensions of the leaf lamina, the values of the photosynthetic performance index and the plant iron concentration to a higher extent in respect to the other treatments. Microplastic toxicity exerted on the growth of C. pepo raises concerns about possible yield and economic loss, as well as for risks of a possible transfer into the food chain.

Agronomic performance and food safety of strawberry cultivated on a remediated sediment.

A marine sediment phytoremediated and homogenized by landfarming was tested for its potential recycle as growing media in horticulture. Two strawberry cultivars, Camarosa and Monterey, were grown on remediated sediment alone (TS100), commercial peat/pumice based growing medium (TS0) and a mixture 1:1 in volume of sediment and peat (TS50). Chemical fertility and strawberry production and safety of produced food were monitored for three consecutive productive seasons on the same growing media. During the first year of cultivation, plants grown on sediment-based media showed a significantly lower biomass production and fruit yield compared with peat, mainly due to the sediment low fertility. In the subsequent two years, the plant re-cultivation improved the sediment structure and N mineralization, and on the third cultivation year both strawberry cultivars showed higher fruit productivity and no accumulation of potentially toxic trace metals. The produced fruits did non accumulate high concentrations of trace metals, and risk assessment showed no risks for human health related to the consumption of strawberry produced on sediment-based growing media. We concluded that a phytoremediated sediment could be recycled as an ingredient of soilless growing media for reducing the environmental impact of plant nursery production and posing no risks for human health. These results show that reclaimed sediments could be reconsidered as a component material category in the new EU regulation on fertilizers.

Green and cost-effective synthesis of copper nanoparticles by extracts of non-edible and waste plant materials from Vaccinium species: Characterization and antimicrobial activity.

The aim of this work was the green synthesis of copper nanoparticles (Cu-NPs) using aqueous extracts of (i) bilberry (Vaccinium myrtillus L.) waste residues from the production of fruit juices and (ii) non-edible "false bilberry" fruits (Vaccinium uliginosum L. subsp. gaultherioides). Different cupric salts (CuCl2, Cu(CH3COO)2 and Cu(NO3)2) were used for the synthesis. The formation of stable nanoparticles (CuNPs) was assessed by transmission electron microscopy and the oxidation state of copper in these aggregates was followed by X-ray photoelectron spectroscopy. The polyphenol composition of the extracts was characterized, before and after the synthesis, using spectrophotometric methods (i.e. total soluble polyphenols and total monomeric anthocyanins) and high-performance liquid chromatography coupled with tandem mass spectrometry (i.e. individual anthocyanins). Polyphenol concentration in the extracts was found to decrease after the synthesis, indicating their active participation to the processes, which led to the formation of Cu-NPs. The antimicrobial activity of Cu-NPs, berry extracts, and cupric ion solutions were analysed by broth microdilution and time-kill assays, on prokaryotic and eukaryotic models. The antimicrobial activity of Cu-NPs, especially those derived from bilberry waste residues, appeared to be higher for both Gram-negative and Gram-positive bacteria, and for fungi, compared to the ones of its single components (cupric salts and berry extracts). Therefore, Cu-NPs from the green synthesis here proposed can be considered as a cost-effective sanitization tool with a wide spectrum of action.

Closing the loop in a constructed wetland for the improvement of metal removal: the use of Phragmites australis biomass harvested from the system as biosorbent.

Among the numerous clean-up techniques for water treatment, sorption methods are widely used for the removal of trace metals. Phragmites australis is a macrophyte commonly used in constructed wetlands for water purification, and in the last decades, its use as biosorbent has attracted increasing attention. In view of a circularly economy approach, this study investigated improvement of trace metal removal by recycling the biomass of P. australis colonizing a constructed wetland, which operates as post-treatment of effluent wastewater from an activated sludge plant serving the textile industrial district of Prato (Italy). After the annual mowing of the reed plants, the biomass was dried and blended to derive a sustainable and eco-friendly biosorbent and its sorption capacity for Fe, Cu, and Zn was investigated comparing the batch system with the easier-to-handle column technique. The possibility of regeneration and reuse of the biosorbent was also evaluated. The biomaterial showed an interesting sorption capacity for Cu, Fe, and Zn, both in batch and in column experiments, especially for Fe ions. The immobilization of the biosorbent in column filters induced some improvement in the removal efficiency, and, in addition, this operation mode has the advantage of being much more suitable for practical applications than the batch process.

Extracting cadmium in the presence of salt: a study on three poplar clones under controlled conditions.

This study aimed at determining the cadmium phytoextraction potential of three Populus alba L. clones cultivated in the presence of increasing sodium chloride concentrations. Plantlets of a commercial and two autochthonous poplar clones were grown in perlite with nutrient solution enriched in CdSO4 (50 and 100 µM) and NaCl (25 and 50 mM), administered either alone or in combination. The three clones showed significant variation not only in cadmium and salt tolerance, accumulation and content, but also in the effect of the interaction between the two elements on these parameters. The toxic effect of Cd and salt excess on plants was mutually exacerbated by the presence of both. Even though the outcome of the joint treatment was always a decrease in shoot Cd or Na accumulation, the three clones showed variation in the extent of such reduction. Evaluating the total element content per plant shoot, the fast-growing commercial clone displayed the highest phytoextraction potential for Cd and Na, either alone or in mixture. Our results demonstrated for the first time that the Cd response in presence of salt can vary in the different clones.

Population Genetics of Odontarrhena (Brassicaceae) from Albania: The Effects of Anthropic Habitat Disturbance, Soil, and Altitude on a Ni-Hyperaccumulator Plant Group from a Major Serpentine Hotspot.

Albanian taxa and populations of the genus Odontarrhena are most promising candidates for research on metal tolerance and Ni-agromining, but their genetic structure remains unknown. We investigated phylogenetic relationships and genetic differentiation in relation to distribution and ploidy of the taxa, anthropic site disturbance, elevation, soil type, and trace metals at each population site. After performing DNA sequencing of selected accessions, we applied DNA-fingerprinting to analyze the genetic structure of 32 populations from ultramafic and non-ultramafic outcrops across Albania. Low sequence divergence resulted in poorly resolved phylograms, but supported affinity between the two diploid serpentine endemics O. moravensis and O. rigida. Analysis of molecular variance (AMOVA) revealed significant population differentiation, but no isolation by distance. Among-population variation was higher in polyploids than in diploids, in which genetic distances were lower. Genetic admixing at population and individual level occurred especially in the polyploids O. chalcidica, O. decipiens, and O. smolikana. Admixing increased with site disturbance. Outlier loci were higher in serpentine populations but decreased along altitude with lower drought and heat stress. Genetic variability gained by gene flow and hybridization at contact zones with "resident" species of primary ultramafic habitats promoted expansion of the tetraploid O. chalcidica across anthropogenic sites.

Inability to accumulate Ni in a genus of hyperaccumulators: the paradox of Odontarrhena sibirica (Brassicaceae).

MAIN CONCLUSION: Odontarrhena is a highly diverse genus of Ni-hyperaccumulators. Here, we demonstrate substantial inability to accumulate Ni in the facultative serpentinophyte O. sibirica, which seems a unique case among the numerous species of the genus that grow on ultramafic soils. Odontarrhena is the most diverse genus of Ni-accumulating plants in W Eurasia, with most taxa growing obligatorily or facultatively on ultramafic soils. A notable exception may be O. sibirica, a facultative serpentinophyte from the E Mediterranean and W Asia in which accumulation ability is still enigmatic. We addressed this issue using observational and experimental methods. Atomic Absorption Analysis of 33 herbarium specimens and plant and soil samples from seven ultramafic and non-ultramafic sites in Greece revealed shoot Ni values always much lower than 1000 µg g-1, non-significant differences between plants from the two soil types and no relationship with soil pH. Only two Turkish specimens from waste mines had shoot Ni concentration > 1000 µg g-1. The reasons for this deviating result remain obscure, but may be associated with inherent peculiarities of the local populations. When cultivated together with congeneric Ni-accumulating species on the same natural ultramafic soil, only O. sibirica was unable to accumulate the metal. Although plant growth was stimulated in hydroponics at relatively low NiSO4 levels (50-150 µM), as typical for hyperaccumulators, Ni-accumulation occurred only at higher concentrations which had a toxic effect. This peculiar combination of Ni-response traits could be the result of a partial evolutionary loss of ability with respect to all other Ni-accumulating congeneric species. For this, O. sibirica could represent a unique model system for further studies on the evolutionary dynamics, physiological mechanisms and genetic control of metal accumulation and homeostasis.