Caerulescens Plants Research Articles

Cadmium and Zn hyperaccumulation provide efficient constitutive defense against Turnip yellow mosaic virus infection in Noccaea caerulescens

To understand the role of Zn and Cd in anti-viral defence, Zn/Cd hyperaccumulator Noccaea caerulescens plants grown with deficient (0.3 µM), replete (10 µM) and excess (100 µM) Zn2+ and Cd (10 µM Zn2+ + 1 µM Cd2+) were infected with Turnip yellow mosaic virus (TYMV). Gas exchange and chlorophyll fluorescence kinetics analyses demonstrated direct TYMV effects on photosynthetic light reactions but N. caerulescens was more resistant against TYMV than the previously studied non-hyperaccumulator N. ochroleucum. Virus abundance and photosynthesis inhibition were the lowest in the high Zn and Cd treatments. RNAseq analysis of 10 µM Zn2+ plants revealed TYMV-induced upregulation of Ca transporters, chloroplastic ZTP29 and defence genes, but none of those that are known to be strongly involved in hyperaccumulation. Synchrotron µ-XRF tomography, however, showed that Zn hyperaccumulation remained strongest in vacuoles of epidermal storage cells regardless of infection. This was in contrast to N. ochroleucum, where apoplastic Zn drastically increased in response to TYMV. These results suggest that the antiviral response of N. caerulescens is less induced by the onset of this biotic stress, but it is rather a permanent resistant state of the plant. Real-time qPCR revealed upregulation of ferritin in Zn10 infected plants, suggesting Fe deprivation as a virus defence strategy under suboptimal Zn supply.

Interactions between the Hyperaccumulator Noccaea caerulescens and Brassica juncea or Lupinus albus for Phytoextraction

Trace-element-contaminated soils cause environmental concern and represent a source of contamination for surrounding areas. Phytoremediation uses plants to diminish the environmental risks associated with this contamination. When the final aim is the extraction of the pollutants, this technique requires the use of plants that are able to accumulate high concentrations of the target elements in their aerial part, while producing high plant biomass. Here, pot experiments were carried out in order to determine the interaction between a hyperaccumulator (Nocaea caerulescens) and a metal excluder (Lupinus albus) or an accumulator (Brassica juncea) species regarding their trace element accumulation/exclusion capacity when sharing the rhizosphere. The plants were grown alone or were cocultivated in soils with different levels of trace element contamination. The Zn concentration in N. caerulescens plants was lower in cocultivation with B. juncea than when they were grown alone, indicating competition between the two species for Zn uptake. Contrastingly, when grown with L. albus, the Zn concentrations in N. caerulescens plants were higher than when grown alone. Therefore, under climatic conditions adequate for N. caerulescens growth, cocultivation with L. albus could favor Zn phytoextraction, while in the case of B. juncea, crop rotation rather than cocultivation is recommended for efficient phytoextraction.

Chlorophyll Fluorescence Imaging Analysis for Elucidating the Mechanism of Photosystem II Acclimation to Cadmium Exposure in the Hyperaccumulating Plant Noccaea caerulescens.

We provide new data on the mechanism of Noccaea caerulescens acclimation to Cd exposure by elucidating the process of photosystem II (PSII) acclimation by chlorophyll fluorescence imaging analysis. Seeds from the metallophyte N. caerulescens were grown in hydroponic culture for 12 weeks before exposure to 40 and 120 μM Cd for 3 and 4 days. At the beginning of exposure to 40 μM Cd, we observed a spatial leaf heterogeneity of decreased PSII photochemistry, that later recovered completely. This acclimation was achieved possibly through the reduced plastoquinone (PQ) pool signaling. Exposure to 120 μM Cd under the growth light did not affect PSII photochemistry, while under high light due to a photoprotective mechanism (regulated heat dissipation for protection) that down-regulated PSII quantum yield, the quantum yield of non-regulated energy loss in PSII (ΦNO) decreased even more than control values. Thus, N. caerulescens plants exposed to 120 μM Cd for 4 days exhibited lower reactive oxygen species (ROS) production as singlet oxygen (1O2). The response of N. caerulescens to Cd exposure fits the ‘Threshold for Tolerance Model’, with a lag time of 4 d and a threshold concentration of 40 μM Cd required for the induction of the acclimation mechanism.

Epigenetic modifications preserve the hyperaccumulator Noccaea caerulescens from Ni geno-toxicity.

The Ni hyperaccumulator Noccaea caerulescens has adapted to live in a naturally stressed environment, evolving a complex pattern of traits to cope with adverse conditions. Evidence is accumulating regarding the important role of epigenetic modifications in regulating plant responses to stress. In this study, we present data from the natural "open-field" adaptation of the Ni hyperaccumulator N. caerulescens to serpentine soil and provide the first evidence of the involvement of epigenetic changes in response to the high Ni content present in plant leaves. The alkaline comet assay revealed the integrity of the nuclei of leaf cells of N. caerulescens grown in a Ni-rich environment, while in the non-tolerant Arabidopsis thaliana exposed to Ni, the nuclei were severely damaged. DNA of N. caerulescens plants grown in situ were considerably hyper-methylated compared to A. thaliana plants exposed to Ni. In addition, qRT-PCR revealed that N. caerulescens MET1, DRM2, and HDA8 genes involved in epigenetic DNA and histone modification were up-regulated in the presence of high Ni content in leaves. Such epigenetic modifications may constitute a defense strategy that prevents genome instability and direct damage to the DNA structure by Ni ion, enabling plants to survive in an extreme environment. Further studies will be necessary to analyze in detail the involvement of DNA methylation and other epigenetic mechanisms in the complex process of metal hyperaccumulation and plants' adaptive response. Environ. Mol. Mutagen. 59:464-475, 2018. © 2018 Wiley Periodicals, Inc.

Zinc Isotope Fractionation in the Hyperaccumulator Noccaea caerulescens and the Nonaccumulating Plant Thlaspi arvense at Low and High Zn Supply.

On the basis of our previous field survey, we postulate that the pattern and degree of zinc (Zn) isotope fractionation in the Zn hyperaccumulator Noccaea caerulescens (J. & C. Presl) F. K. Mey may reflect a relationship between Zn bioavailability and plant uptake strategies. Here, we investigated Zn isotope discrimination during Zn uptake and translocation in N. caerulescens and in a nonaccumulator Thlaspi arvense L. with a contrasting Zn accumulation ability in response to low (Zn-L) and high (Zn-H) Zn supplies. The average isotope fractionations of the N. caerulescens plant as a whole, relative to solution (Δ(66)Znplant-solution), were -0.06 and -0.12‰ at Zn-L-C and Zn-H-C, respectively, indicative of the predominance of a high-affinity (e.g., ZIP transporter proteins) transport across the root cell membrane. For T. arvense, plants were more enriched in light isotopes under Zn-H-A (Δ(66)Znplant-solution = -0.26‰) than under Zn-L-A and N. caerulescens plants, implying that a low-affinity (e.g., ion channel) transport might begin to function in the nonaccumulating plants when external Zn supply increases. Within the root tissues of both species, the apoplast fractions retained up to 30% of Zn mass under Zn-H. Moreover, the highest δ(66)Zn (0.75‰-0.86‰) was found in tightly bound apoplastic Zn, pointing to the strong sequestration in roots (e.g., binding to high-affinity ligands/precipitation with phosphate) when plants suffer from high Zn stress. During translocation, the magnitude of isotope fractionation was significantly greater at Zn-H (Δ(66)Znroot-shoot = 0.79‰) than at Zn-L, indicating that fractionation mechanisms associated with root-shoot translocation might be identical to the two plant species. Hence, we clearly demonstrated that Zn isotope fractionation could provide insight into the internal sequestration mechanisms of roots when plants respond to low and high Zn supplies.

Local adaptation is associated with zinc tolerance in Pseudomonas endophytes of the metal-hyperaccumulator plant Noccaea caerulescens.

Metal-hyperaccumulating plants, which are hypothesized to use metals for defence against pests and pathogens, provide a unique context in which to study plant-pathogen coevolution. Previously, we demonstrated that the high concentrations of zinc found in leaves of the hyperaccumulator Noccaea caerulescens provide protection against bacterial pathogens, with a potential trade-off between metal-based and pathogen-induced defences. We speculated that an evolutionary arms race between zinc-based defences in N. caerulescens and zinc tolerance in pathogens might have driven the development of the hyperaccumulation phenotype. Here, we investigate the possibility of local adaptation by bacteria to the zinc-rich environment of N. caerulescens leaves and show that leaves sampled from the contaminated surroundings of a former mine site harboured endophytes with greater zinc tolerance than those within plants of an artificially created hyperaccumulating population. Experimental manipulation of zinc concentrations in plants of this artificial population influenced the zinc tolerance of recovered endophytes. In laboratory experiments, only endophytic bacteria isolated from plants of the natural population were able to grow to high population densities in any N. caerulescens plants. These findings suggest that long-term coexistence with zinc-hyperaccumulating plants leads to local adaptation by endophytic bacteria to the environment within their leaves.

Noccaea caerulescens populations adapted to grow in metalliferous and non-metalliferous soils: Ni tolerance, accumulation and expression analysis of genes involved in metal homeostasis

Three populations of the Brassicaceae Noccaea caerulescens: (i) the metallicolous Mt. Prinzera population (Italy), which have adapted to grow on serpentinite, a soil naturally rich in Ni, Co, Cr; (ii) the metallicolous La Calamine population (Belgium), which have adapted to growth on soil highly contaminated by Cd, Zn and Pb, and (iii) the non-metallicolous population growing near the town of Rožnov pod Radhoštěm (Czech Republic), were grown in hydroponics and treated with different Ni concentrations (0, 10 and 100μM NiSO4). Ni tolerance and accumulation were analysed along with the expression of genes belonging to different families involved in plant metal homeostasis: ZNT1, ZNT2, NRAMP3, NRAMP4 coding for non-ATP-hydrolysing plasma membrane and vacuolar metal transporters, HMA3, HMA4 coding for ATP-hydrolising metal transporters, NAS1, NAS3, NAS4 and MT1B involved in metal chelation. The three populations showed different levels of expression of some of the tested genes in condition of 0μM Ni. In addition, the Ni hyperaccumulator Mt. Prinzera showed the highest Ni translocation capacity at 10μM Ni and a specific up-regulation of ZNT1, ZNT2, NAS3, NRAMP3 and NRAMP4 genes. With the same Ni treatment, the La Calamine population induced HMA4 and MT1B genes, while the Rožnov pod Radhoštěm population only displayed an over-expression of all the genes at 100μM Ni, the condition in which this population suffers heavily from Ni stress. The expression of ZNT1, ZNT2, NRAMP3, NRAMP4 was also tested on N. caerulescens plants growing in the natural environment on Mt. Prinzera. Higher transcript levels of ZNT1 and ZNT2 were associated with higher levels of plant's total Ni content. Overall these results suggested that some of the genes considered can have a role in determining metal tolerance and accumulation in metallicolous N. caerulescens populations, while they were not involved in metal homeostasis in the non-metallicolous population.

Functional diversity as indicator of the recovery of soil health derived from Thlaspi caerulescens growth and metal phytoextraction

Continuous phytoextraction has lately drawn a lot of attention due to its potential for the remediation of metal polluted soils. Although when assessing the success of a phytoextraction process, up till now, emphasis has mostly been placed on metal removal, it is important to highlight that the ultimate objective of a phytoextraction process must be to restore soil health. Consequently, a short-term microcosm study was carried out to evaluate the capacity of an actively growing ecotype of the Zn and Cd hyperaccumulator Thlaspi caerulescens (Lanestosa ecotype) to phytoextract metals from soil and, above all, to assess the potential of soil functional diversity (through the determination of soil enzyme activities and community level physiological profiles) to both determine the toxic effect of metals on soil condition and to monitor the efficiency of a metal phytoextraction process. T. caerulescens plants grown in metal polluted soils showed a shoot metal concentration of 337 mg of Cd, 5670 mg of Zn and 76.6 mg of Pb per kg of dry weight tissue. Apart from confirming its great potential for Zn and Cd phytoextraction, the presence of T. caerulescens, as compared to the metal phytoextraction itself, had the major effect on soil biological parameters. Actually, in metal polluted soils, the presence of T. caerulescens led to a 154, 115, 140, 37 and 164% increase in the activity of β-glucosidase, arylsulphatase, acid phosphatase, alkaline phosphatase and urease, respectively. Metal pollution did not cause a clear inhibition of soil enzyme activities. Contrasting results were obtained with EcoPlates™ versus soil enzyme activities. Actually, the presence of metals led to significantly lower values of Shannon's index calculated from enzyme activities and non-significant higher values of this same index when calculated from EcoPlates™ data. It was concluded that biological indicators of soil health are valid tools to evaluate the success of a metal phytoextraction process.

Arabidopsis thaliana and Thlaspi caerulescens respond comparably to low zinc supply

The main objective of this research was to study the response of Arabidopsis thaliana L. and Thlaspi caerulescens J. & C. Presl to different Zn supplies. The A. thaliana plants were exposed to Zn-deficiency (0 and 0.05 μM Zn) and compared to the plants grown on media containing standard Zn (2 μM). T. caerulescens plants were also exposed to Zn-deficiency (0.05 μM Zn), but as this is a Zn hyperaccumulator species, also to high Zn (1,000 μM Zn). Plants were compared to plants grown on standard Zn media (100 μM Zn). Both A. thaliana and T. caerulescens were found to be heavily affected by Zn deficiency, showing similar retarded growth and reduced reproduction phenotypes, and even less reduction in biomass production in T. caerulescens than in A. thaliana. T. caerulescens plants were similarly affected when grown on high Zn concentrations, with comparable effects on reproductive tissues as seen on low Zn supply.

Testing of Outstanding Individuals of Thlaspi Caerulescens for Cadmium Phytoextraction

Thlaspi caerulescens is known to hyperaccumulate high quantities of Cd with Cd concentrations up to 3000 mg kg−1 in some populations from south of France. However, within these populations, the Cd concentrations can vary widely from plant to plant in a way that appears to be not entirely due to variations in soil Cd. The aim of this work was to investigate the variability in the Cd uptake ability of individual plants within a population and among seedlings grown from seeds from a single plant. Ten populations of T. caerulescens plants were selected from four locations (V: Viviez; SF: Saint Félix-de-Pallières; LB: Le Bleymard; CMA: Col du Mas de l’Air) depending of the extent and soil homogeneity of the site. One population from CMA consisted of the progeny of a single maternal plant. Hundred plants of each population were grown for three months in the same homogeneous and lightly Cd-polluted soil (about 20 mg total Cd kg−1 dry soil). Cadmium uptake behavior of the plants was monitored by labeling the soil with 109Cd. To allow partial plant destruction, radioanalysis was performed on the largest leaf of each plant as an indicator of the total Cd concentration in plant shoots. Results showed significant differences in biomass production and Cd uptake by T. caerulescens between sites and between populations within sites. We observed a wide intra-population variation in biomass, Cd concentration and total Cd uptake. For these properties, 1 to 5 percents of the plants in each population varied by more than a factor of two from the mean values. The mean Cd uptake by the single-plant population from CMA was more than 40% higher than for the population at large. T. caerulescens would respond to traditional selection methods, which would significantly improve the phytoextraction of Cd.

Large Expression Differences in Genes for Iron and Zinc Homeostasis, Stress Response, and Lignin Biosynthesis Distinguish Roots of Arabidopsis thaliana and the Related Metal Hyperaccumulator Thlaspi caerulescens

The micronutrient zinc has an essential role in physiological and metabolic processes in plants as a cofactor or structural element in 300 catalytic and noncatalytic proteins, but it is very toxic when available in elevated amounts. Plants tightly regulate their internal zinc concentrations in a process called zinc homeostasis. The exceptional zinc hyperaccumulator species Thlaspi caerulescens can accumulate up to 3% of zinc, but also high amounts of nickel and cadmium, without any sign of toxicity. This should have drastic effects on the zinc homeostasis mechanism. We examined in detail the transcription profiles of roots of Arabidopsis thaliana and T. caerulescens plants grown under deficient, sufficient, and excess supply of zinc. A total of 608 zinc-responsive genes with at least a 3-fold difference in expression level were detected in A. thaliana and 352 in T. caerulescens in response to changes in zinc supply. Only 14% of these genes were also zinc responsive in A. thaliana. When comparing A. thaliana with T. caerulescens at each zinc exposure, more than 2,200 genes were significantly differentially expressed (>or=5-fold and false discovery rate < 0.05). While a large fraction of these genes are of yet unknown function, many genes with a different expression between A. thaliana and T. caerulescens appear to function in metal homeostasis, in abiotic stress response, and in lignin biosynthesis. The high expression of lignin biosynthesis genes corresponds to the deposition of lignin in the endodermis, of which there are two layers in T. caerulescens roots and only one in A. thaliana.

Potential of Thlaspi caerulescens for Cadmium Phytoremediation: Comparison of Two Representative Soil Types in Japan under Different Planting Frequencies

To assess the potential of the Cd hyperaccumulator Thlaspi caerulescens (Ganges ecotype) for Cd phytoremediation in Japan, we compared the changes in the soil Cd concentration between a Fluvisol and an Andosol and the efficiency of Cd removal under different planting frequencies in a pot experiment. The soils were artificially contaminated with Cd(NO3)2 to the level of about 5 mg Cd kg−1. The cultivation system of Thlaspi caerulescens applied to both soil types continued for 4 months (4 × 1 treatment) and the other system only applied to the Fluvisol consisted of two plantings, at the beginning and after 2 months (2 × 2 treatment). Control without plants was also prepared. The cumulative amount of Cd absorbed by shoots for 4 months was 385 μg pot−1 for the 4 × 1 treatment of the Fluvisol, 582 μg pot−1 for the 2 × 2 treatment of the Fluvisol, and 112 μg pot−1 for the Andosol. In the Fluvisol, Thlaspi caerulescens plants in the 2 × 2 treatment could remove 1.5 times more Cd than in the 4 × 1 treatment. A much higher Cd concentration in the shoots of the second cropping in the 2 × 2 treatment mainly led to this difference in efficiency. The total Cd concentration of the unplanted Fluvisol and Andosol was 5.0 and 6.1 mg kg−1, and after 4 months, that of the Fluvisol in the 4 × 1 and 2 × 2 treatments and the Andosol reached 4.0, 2.9 and 5.1 mg kg−1, respectively. The difference between the rhizosphere and the non-rhizosphere soils was not evident in the Fluvisol, while the concentration in the rhizosphere was lower than that in the non-rhizosphere soil in the Andosol. In the Fluvisol, the amount of available (1 M NH4NO3-extractable) Cd decreased only in the rhizosphere, while in the Andosol, it was halved in both rhizosphere and non-rhizosphere soils. Furthermore, the ratio of the amount of available Cd to total Cd did not decrease in the Fluvisol, whereas it decreased in the Andosol, which indicated the presence of a smaller pool of potentially available Cd in the Andosol. Based on the shoot biomass and bioaccumulation factor obtained in the 4 × 1 treatment, it was estimated that only about 2 crops for the Fluvisol and 6 crops for the Andosol could decrease the Cd concentration to 50%. In conclusion, 1) short-rotation is recommended because of the high Cd concentration in the shoots of the second cropping and the increase in the volume of the rhizosphere soil, 2) the amount of available Cd in soil should be monitored in addition to the concentration of total Cd to evaluate the effectiveness of phytoremediation, and 3) the potential of Thlaspi caerulescens was clearly demonstrated and the use of this hyperaccumulator could become a promising alternative for the remediation of Cd-contaminated soils in Japan.

Cadmium tolerance in Thlaspi caerulescens: I. Growth parameters, metal accumulation and phytochelatin synthesis in response to cadmium

A population of the metallophyte, Thlaspi caerulescens, originating from a Cd–Pb–Zn old mining and smelter site at Plombières (Belgium) was studied. T. caerulescens was cultivated hydroponically to investigate Cd uptake and tolerance. Cd was added to Hoagland’s medium at concentration range from 5 to 500 μM. The plants could tolerate 500 μM Cd in the solution showing only minor visible symptoms of toxicity but with a 32% decrease in fresh weight. After 14 days at 500 μM, Cd content in roots and shoots was 707 and 602 mg kg −1 of dry weight (d.w.), respectively. Application of Cd to hydroponically cultivated T. caerulescens induced the accumulation of PCs in plant roots and shoots. Buthionine sulfoximine (BSO) application almost completely reduced (by 98–100%) the accumulation of PCs without simultaneous increase in plants sensitivity to Cd. These results suggest a minor if any role of PCs in tolerance to Cd of the studied population of T. caerulescens in hydroponics. On the other hand, no PC accumulation was detected either in T. caerulescens plants growing in their natural environment at Plombierès or in plants growing in their native soil in a greenhouse. These results suggest that naturally selected tolerance in T. caerulescens population from Plombières is not associated with enhanced PCs synthesis.

Cadmium tolerance in Thlaspi caerulescensI. Growth parameters, metal accumulation and phytochelatin synthesis in response to cadmium

A population of the metallophyte, Thlaspi caerulescens , originating from a Cd–Pb–Zn old mining and smelter site at Plombières (Belgium) was studied. T. caerulescens was cultivated hydroponically to investigate Cd uptake and tolerance. Cd was added to Hoagland’s medium at concentration range from 5 to 500 μM. The plants could tolerate 500 μM Cd in the solution showing only minor visible symptoms of toxicity but with a 32% decrease in fresh weight. After 14 days at 500 μM, Cd content in roots and shoots was 707 and 602 mg kg −1 of dry weight (d.w.), respectively. Application of Cd to hydroponically cultivated T. caerulescens induced the accumulation of PCs in plant roots and shoots. Buthionine sulfoximine (BSO) application almost completely reduced (by 98–100%) the accumulation of PCs without simultaneous increase in plants sensitivity to Cd. These results suggest a minor if any role of PCs in tolerance to Cd of the studied population of T. caerulescens in hydroponics. On the other hand, no PC accumulation was detected either in T. caerulescens plants growing in their natural environment at Plombierès or in plants growing in their native soil in a greenhouse. These results suggest that naturally selected tolerance in T. caerulescens population from Plombières is not associated with enhanced PCs synthesis.

Determination of glucosinolates in rapeseed and Thlaspi caerulescens plants by liquid chromatography–atmospheric pressure chemical ionization mass spectrometry

Liquid chromatography–atmospheric pressure chemical ionization mass spectrometry was used to identify glucosinolates in plant extracts. Optimization of the analytical conditions and the determination of the method detection limit was performed using commercial 2-propenylglucosinolate (sinigrin). Optimal values for the following parameters were determined: nebulization pressure, gas temperature, flux of drying gas, capillar voltage, corona current and fragmentor conditions. The method detection limit for sinigrin was 2.85 ng. For validation of the method the glucosinolates in reference material (rapeseed) from the Community Bureau of Reference Materials (BCR) were analyzed. The method was applied for the determination of glucosinolates in Thlaspi caerulescens plants.