Correction to: Recalibration and validation of the Swiss lichen bioindication methods for air quality assessment.

The correct fig. 2 is presented in this paper.

Recalibration and validation of the Swiss lichen bioindication methods for air quality assessment.

The aim of this study was to recalibrate the Swiss lichen bioindication methods, developed and calibrated with air pollution data 30 years ago. Since then, levels of air pollution have considerably decreased, and the mix of pollutants has changed due to successful emission control in Switzerland and neighboring countries. In particular, there has been a change from a sulfur- and acid-dominated to a more nitrogen-dominated pollution mix of NH3/NOx and ozone, resulting in increased pH levels. This allowed a recolonization and change in abundance and composition of the epiphytic lichen vegetation, indicating an improved air quality in Switzerland. The existing indices of atmospheric pollution or purity IAP18 and IAPBR developed 30 years ago showed good longitudinal correlations with air pollutant levels until the end of the last century, but a growing drift was observed in some regions over the last 15 years. This called for a method recalibration with more recent air pollution data. Data from a total of 7178 trees from 22 Swiss regions grouped into 1331 homogenous plots and covering the period 1994 to 2017 were averaged by year within plots. Three pollutant-specific lichen indices were newly established, one for primary pollutants (NO2, PM10, SO2), one for ozone (AOT40f), and one for ammonia (NH3). These pollutant-specific lichen indices were derived from linear regression models with lichen variables and a linear time trend variable as predictors, using time-dependent coefficients. Parameters were selected using the Lasso method. The primary pollutant lichen index showed a coefficient of determination R2 of 0.86 in the model with NO2, PM10, and SO2 as predictor variables, whereas corresponding models with other predictor variables (i.e., NH3, AOT40f, and meteorological variables) were of considerably lower fit. Regionalized lichen models for three larger Swiss regions revealed even better results, compared with the unified Swiss models. The best regionalized ozone and ammonia lichen indices reached an R2 of 0.88 and 0.71, respectively.

Field evaluation of one Cu-resistant somaclonal variant and two clones of tobacco for copper phytoextraction at a wood preservation site.

A Cu-resistant somaclonal tobacco variant (NBCu 10-8-F1, C1), its BaG mother clone (C3), and the FoP tobacco clone (C2) were cultivated at a wood preservation site on Cu-contaminated soils (239-1290 mg Cu kg-1 soil range) and an uncontaminated control site (CTRL, 21 mg Cu kg-1) to assess their shoot DW yields and potential use for bioavailable Cu stripping. The Cu concentration in the soil pore water varied between 0.15 and 0.84 mg L-1. Influences of Cu exposure and soil treatments, i.e., untreated soil (Unt), soils amended with compost and either dolomitic limestone (OMDL) or zerovalent iron grit (OMZ), on plant growth and shoot ionome were determined. All transplants survived and grew even at high total soil Cu. Shoots were harvested after 3 months (cut 1). Subsequently, bottom suckers developed and were harvested after 2 months (cut 2). Total shoot DW yield (cuts 1 + 2) varied between 0.8 and 9.9 t DW ha-1 year-1 depending on tobacco cultivars, soil treatments, and soil Cu exposure. It peaked for all cultivars in the OMDL plots at moderate Cu exposure (239-518 mg kg-1 soil), notably for the C2 plants. Cut 2 contributed for 11-43% to total shoot DW yield. Increase in shoot DW yield diluted shoot Cu concentration. At low Cu exposure, total shoot Cu removal peaked for the variant. At moderate Cu exposure, shoot Cu concentrations were similar in all cultivars, but total shoot Cu removal was highest for the C2 plants. At high Cu exposure (753-1140 mg kg-1), shoot Cu concentrations peaked for the C2 plants in the Unt plots, the C1 and C2 plants in the OMZ plot, and the C3 ones in the OMDL plots. Shoot Cu removal (in g Cu ha-1 year-1) ranged from 15.4 (C2 on the CTRL soil) to 261.3 (C2 on moderately contaminated OMDL soils). The C2 plants phytoextracted more Cu than the C1 and C3 ones in the Unt plots and in the OMDL plots at moderate Cu exposure. In the OMDL plots with high Cu exposure, shoot Cu removal was highest for the C1 plants. Soil amendments improved shoot Cu removal through increase in either shoot DW yield (OMDL-3-fold) or shoot Cu concentration (OMZ-1.3-fold). Increased shoot Cu concentration induced an ionome imbalance with increased shoot Al, Fe, B, and Mg concentrations and decreased P and K ones. Copper concentrations in plant parts varied in decreasing order: roots > leaves > inflorescence (cymes including seeds) > stem, whereas Cu removal ranked as roots > stem = leaves > inflorescence.

Temporal change of the accumulation of persistent organic pollutants (POPs) and polycyclic aromatic hydrocarbons (PAHs) in lichens in Switzerland between 1995 and 2014.

The aim of this study was to assess the temporal change of atmospheric deposition patterns of persistent organic pollutants (POPs) and polycyclic aromatic hydrocarbons (PAHs) in Switzerland between 1995 and 2014 by a passive biomonitoring with lichens. Lichen tissues sampled at 16 representative sites in the same season of 1995 and 2014 were analyzed for a total of 94 individual and 27 sum parameters of POPs and PAHs by means of gas chromatography-mass spectrometry (GC/MS). The comparative analyses showed a decrease of 40 to 80% (medians) for most of the POPs and PAHs concentration in lichens at all site categories. Reduction in tissue concentration of the polychlorinated dibenzo-p-dioxins/furans (PCDD/PCDFs), such as the highly toxic 2,3,7,8-TetraCDD and the TEQ according to WHO (2005) were 66% and 73%, respectively. For the dioxin- and non-dioxin-like PCBs, a decrease of 67% and 58% was found. The average decrease of 30 organochlorine pesticides and insecticides (OCPs) was 65%, with a 94% decrease for lindane. For the 27 PAHs and for benzo(a)pyrene, an average decrease of 58% and 59% was found. Polybrominated diphenyl ethers (PBDE) showed reduced concentrations in lichens at rural and agglomeration sites, but an increase of contamination was observed at industrial and road traffic sites. The direct comparison of changes of POPs and PAHs concentrations in lichens and of emissions between 1995 and 2014 revealed consistent results. The results of this study highlight for the first time in biota the positive effect of emission regulation of POPs in Switzerland.

Trace elements bioavailability to winter wheat (Triticum aestivum L.) grown subsequent to high biomass plants in a greenhouse study.

Multielement-contaminated agricultural land requires the adaptation of agronomic practices to meet legal requirements for safe biomass production. The incorporation of bioenergy plants with, at least, moderate phytoextraction capacity into crop rotations with cereals can affect trace elements (TE) phytoavailability and, simultaneously, constitute economic revenues for farmers outside the food or forage sector. Hence, in a crop rotation pot study sunflower (Helianthus annuus L.), modified for high biomass and TE accumulation by chemical mutagenesis, was compared to winter oilseed rape (Brassica napus L.) as pre-crop. On two agricultural soils with different TE loads, the crops´ potential for phytoextraction and for impacts on TE uptake by subsequent winter wheat (Triticum aestivum L.) was studied. The results showed that rape tolerated high-level mixed contamination with metals (Cd, Pb and Zn) and As more than sunflower. In both soils, labile metals concentration increased and soil acidity remained high following sunflower. Furthermore, enhanced grain As accumulation in subsequent wheat was observed. By contrast, soil acidity and Cd or Zn accumulation of subsequent wheat decreased following rape. In the short term, moderate phytoextraction was superimposed by nutrient use or rhizosphere effects of pre-crops, which should be carefully monitored when designing crop rotations for contaminated land.

Morphological and functional responses of a metal-tolerant sunflower mutant line to a copper-contaminated soil series.

The potential use of a metal-tolerant sunflower mutant line for biomonitoring Cu phytoavailability, Cu-induced soil phytotoxicity, and Cu phytoextraction was assessed on a Cu-contaminated soil series (13-1020 mg Cu kg-1) obtained by fading a sandy topsoil from a wood preservation site with a similar uncontaminated soil. Morphological and functional plant responses as well as shoot, leaf, and root ionomes were measured after a 1-month pot experiment. Hypocotyl length, shoot and root dry weight (DW) yields, and leaf area gradually decreased as soil Cu exposure rose. Their dose-response curves (DRC) plotted against indicators of Cu exposure were generally well fitted by sigmoidal curves. The half-maximal effective concentration (EC50) of morphological parameters ranged between 203 and 333 mg Cu kg-1 soil, corresponding to 290-430 µg Cu L-1 in the soil pore water, and 20 ± 5 mg Cu kg-1 DW in the shoots. The EC10 for shoot Cu concentration (13-15 mg Cu kg-1 DW) coincided to 166 mg Cu kg-1 soil. Total chlorophyll content and total antioxidant capacity (TAC) were early biomarkers (EC10: 23 and 51 mg Cu kg-1 soil). Their DRC displayed a biphasic response. Photosynthetic pigment contents, e.g., carotenoids, correlated with TAC. Ionome was changed in Cu-stressed roots, shoots, and leaves. Shoot Cu removal peaked roughly at 280 µg Cu L-1 in the soil pore water.

Tobacco, Sunflower and High Biomass SRC Clones Show Potential for Trace Metal Phytoextraction on a Moderately Contaminated Field Site in Belgium.

Phytoextraction could be a potential management option for diffusely Cd-Zn-Pb-polluted agricultural land in Northeast Belgium. The use of high yielding crops with a sufficiently high metal accumulation is preferred as these are expected to both gradually decontaminate the soil while generating an income through biomass valorization. To find out which high biomass crop possessed the highest and most constant (in time) phytoextraction potential on these soils, different plant species and different mutants or clones of each species, were evaluated during consecutive years. Biomass production and metal accumulation of pre-selected tobacco somaclonal variants (Nicotiana tabacum L.) and pre-selected sunflower mutants (Helianthus annuus L.) were investigated for two productivity years, while the phytoextraction potential of experimental poplar (Populus) and willow (Salix) in short rotation coppice (SRC) was assessed at the end of the second cutting cycle (after two times four growing seasons). The tobacco clones and the sunflower mutants showed efficient extraction of, respectively, Cd and Zn, while the highest simultaneous extractions of Cd and Zn were gained with some SRC clones. Variation in biomass production and metal accumulation were high for all crops over the years. The highest biomass production was observed for the experimental poplar clone of the crossing type Populus deltoides (P. maximowiczii x P. trichocarpa) with 9.9 ton DW per ha per year. The remediation period to reach legal threshold values for the pseudo-total content of Cd in this specific soil was estimated to be at least 60 years. Combining estimated phytoextraction potential and economic and environmental aspects, the SRC option is proposed as the most suitable crop for implementing metal phytoextraction in the investigated area.

Assessing phytotoxicity of trace element-contaminated soils phytomanaged with gentle remediation options at ten European field trials.

Gentle remediation options (GRO), i.e. in situ stabilisation, (aided) phytoextraction and (aided) phytostabilisation, were implemented at ten European sites contaminated with trace elements (TE) from various anthropogenic sources: mining, atmospheric fallout, landfill leachates, wood preservatives, dredged-sediments, and dumped wastes. To assess the performance of the GRO options, topsoil was collected from each field trial, potted, and cultivated with lettuce (Lactuca sativa L.) for 48days. Shoot dry weight (DW) yield, photosynthesis efficiency and major element and TE concentrations in the soil pore water and lettuce shoots were measured. GRO implementation had a limited effect on TE concentrations in the soil pore water, although use of multivariate Co-inertia Analysis revealed a clear amelioration effect in phytomanaged soils. Phytomanagement increased shoot DW yield at all industrial and mine sites, whereas in agricultural soils improvements were produced in one out of five sites. Photosynthesis efficiency was less sensitive than changes in shoot biomass and did not discriminate changes in soil conditions. Based on lettuce shoot DW yield, compost amendment followed by phytoextraction yielded better results than phytostabilisation; moreover shoot ionome data proved that, depending on initial soil conditions, recurrent compost application may be required to maintain crop production with common shoot nutrient concentrations.

Biochar and compost amendments enhance copper immobilisation and support plant growth in contaminated soils.

Contamination of soil with trace elements, such as Cu, is an important risk management issue. A pot experiment was conducted to determine the effects of three biochars and compost on plant growth and the immobilisation of Cu in a contaminated soil from a site formerly used for wood preservation. To assess Cu mobility, amended soils were analysed using leaching tests pre- and post-incubation, and post-growth. Amended and unamended soils were planted with sunflower, and the resulting plant material was assessed for yield and Cu concentration. All amendments significantly reduced leachable Cu compared to the unamended soil, however, the greatest reductions in leachable Cu were associated with the higher biochar application rate. The greatest improvements in plant yields were obtained with the higher application rate of biochar in combination with compost. The results suggest joint biochar and compost amendment reduces Cu mobility and can support biomass production on Cu-contaminated soils.

Agronomic Practices for Improving Gentle Remediation of Trace Element-Contaminated Soils.

The last few decades have seen the rise of Gentle soil Remediation Options (GRO), which notably include in situ contaminant stabilization ("inactivation") and plant-based (generally termed "phytoremediation") options. For trace element (TE)-contaminated sites, GRO aim to either decrease their labile pool and/or total content in the soil, thereby reducing related pollutant linkages. Much research has been dedicated to the screening and selection of TE-tolerant plant species and genotypes for application in GRO. However, the number of field trials demonstrating successful GRO remains well below the number of studies carried out at a greenhouse level. The move from greenhouse to field conditions requires incorporating agronomical knowledge into the remediation process and the ecological restoration of ecosystem services. This review summarizes agronomic practices against their demonstrated or potential positive effect on GRO performance, including plant selection, soil management practices, crop rotation, short rotation coppice, intercropping/row cropping, planting methods and plant densities, harvest and fertilization management, pest and weed control and irrigation management. Potentially negative effects of GRO, e.g., the introduction of potentially invasive species, are also discussed. Lessons learnt from long-term European field case sites are given for aiding the choice of appropriate management practices and plant species.

Endophytic bacteria take the challenge to improve Cu phytoextraction by sunflower.

Endophytic bacteria from roots and crude seed extracts of a Cu-tolerant population of Agrostis capillaris were inoculated to a sunflower metal-tolerant mutant line, and their influence on Cu tolerance and phytoextraction was assessed using a Cu-contaminated soil series. Ten endophytic bacterial strains isolated from surface-sterilized A. capillaris roots were mixed to prepare the root endophyte inoculant (RE). In parallel, surface-sterilized seeds of A. capillaris were crushed in MgSO4 to prepare a crude seed extract containing seed endophytes (SE). An aliquot of this seed extract was filtered at 0.2 µm to obtain a bacterial cell-free seed extract (SEF). After surface sterilization, germinated sunflower seeds were separately treated with one of five modalities: no treatment (C), immersion in MgSO4 (CMg) or SEF solutions and inoculation with RE or SE. All plants were cultivated on a Cu-contaminated soil series (13-1020 mg Cu kg(-1)). Cultivable RE strains were mostly members of the Pseudomonas genera, and one strain was closely related to Labrys sp. The cultivable SE strains belonged mainly to the Bacillus genera and some members of the Rhodococcus genera. The treatment effects depended on the soil Cu concentration. Both SE and SEF plants had a higher Cu tolerance in the 13-517 mg Cu kg(-1) soil range as reflected by increased shoot and root DW yields compared to control plants. This was accompanied by a slight decrease in shoot Cu concentration and increase in root Cu concentration. Shoot and root DW yields were more promoted by SE than SEF in the 13-114 mg Cu kg(-1) soil range, which could reflect the influence of seed-located bacterial endophytes. At intermediate soil Cu (416-818 mg Cu kg(-1) soil), the RE and CMg plants had lower shoot Cu concentrations than the control, SE and SEF plants. At high total soil Cu (617-1020 mg Cu kg(-1)), root DW yield of RE plants slightly increased and their root Cu concentration rose by up to 1.9-fold. In terms of phytoextraction efficiency, shoot Cu removal was increased for sunflower plants inoculated with crude and bacterial cell-free seed extracts by 1.3- to 2.2-fold in the 13-416 mg Cu kg(-1) soil range. Such increase was mainly driven by an enhanced shoot DW yield. The number and distribution of endophytic bacteria in the harvested sunflower tissues must be further examined.

Selecting chemical and ecotoxicological test batteries for risk assessment of trace element-contaminated soils (phyto)managed by gentle remediation options (GRO).

During the past decades a number of field trials with gentle remediation options (GRO) have been established on trace element (TE) contaminated sites throughout Europe. Each research group selects different methods to assess the remediation success making it difficult to compare efficacy between various sites and treatments. This study aimed at selecting a minimum risk assessment battery combining chemical and ecotoxicological assays for assessing and comparing the effectiveness of GRO implemented in seven European case studies. Two test batteries were pre-selected; a chemical one for quantifying TE exposure in untreated soils and GRO-managed soils and a biological one for characterizing soil functionality and ecotoxicity. Soil samples from field studies representing one of the main GROs (phytoextraction in Belgium, Sweden, Germany and Switzerland, aided phytoextraction in France, and aided phytostabilization or in situ stabilization/phytoexclusion in Poland, France and Austria) were collected and assessed using the selected test batteries. The best correlations were obtained between NH4NO3-extractable, followed by NaNO3-extractable TE and the ecotoxicological responses. Biometrical parameters and biomarkers of dwarf beans were the most responsive indicators for the soil treatments and changes in soil TE exposures. Plant growth was inhibited at the higher extractable TE concentrations, while plant stress enzyme activities increased with the higher TE extractability. Based on these results, a minimum risk assessment battery to compare/biomonitor the sites phytomanaged by GROs might consist of the NH4NO3 extraction and the bean Plantox test including the stress enzyme activities.

Feasibility of labile Zn phytoextraction using enhanced tobacco and sunflower: results of five- and one-year field-scale experiments in Switzerland.

Phytoextraction with somaclonal variants of tobacco and sunflower mutant lines (non-GMs) with enhanced metal uptake and tolerance can be a sustainable alternative to conventional destructive decontamination methods, especially for stripping bioavailable zinc excess in topsoil. The overall results of a 5-year time series experiment at field scale in north-eastern Switzerland confirm that the labile Zn pool in soil can be lowered by 45-70%, whereas subplots without phytoextraction treatment maintained labile Zn concentrations. In 2011, the phytoextraction experiment site was enlarged by a factor of 3, and the labile 0.1 M NaNO3 extractable Zn concentration in the soil was reduced up to 58% one period after harvest. A Mass Balance Analysis confirmed soil Zn decontamination in line with plant Zn uptake. The plants partially take Zn from the non-labile pool of the totaL The sustainability of Zn phytoextraction in subplots that no longer exceed the Swiss trigger value is now assessed over time. In contrary to the phytoextraction of total soil Zn which needs a long cleaning up time, the bioavailable Zn stripping is feasible within a few years period.

Phytoremediation of contaminated soils and groundwater: lessons from the field.

BACKGROUND, AIM, AND SCOPE: The use of plants and associated microorganisms to remove, contain, inactivate, or degrade harmful environmental contaminants (generally termed phytoremediation) and to revitalize contaminated sites is gaining more and more attention. In this review, prerequisites for a successful remediation will be discussed. The performance of phytoremediation as an environmental remediation technology indeed depends on several factors including the extent of soil contamination, the availability and accessibility of contaminants for rhizosphere microorganisms and uptake into roots (bioavailability), and the ability of the plant and its associated microorganisms to intercept, absorb, accumulate, and/or degrade the contaminants. The main aim is to provide an overview of existing field experience in Europe concerning the use of plants and their associated microorganisms whether or not combined with amendments for the revitalization or remediation of contaminated soils and undeep groundwater. Contaminations with trace elements (except radionuclides) and organics will be considered. Because remediation with transgenic organisms is largely untested in the field, this topic is not covered in this review. Brief attention will be paid to the economical aspects, use, and processing of the biomass. CONCLUSIONS AND PERSPECTIVES: It is clear that in spite of a growing public and commercial interest and the success of several pilot studies and field scale applications more fundamental research still is needed to better exploit the metabolic diversity of the plants themselves, but also to better understand the complex interactions between contaminants, soil, plant roots, and microorganisms (bacteria and mycorrhiza) in the rhizosphere. Further, more data are still needed to quantify the underlying economics, as a support for public acceptance and last but not least to convince policy makers and stakeholders (who are not very familiar with such techniques).

Response of antioxidant enzymes in Nicotiana tabacum clones during phytoextraction of heavy metals.

BACKGROUND, AIM, AND SCOPE: Tobacco, Nicotiana tabacum, is a widely used model plant for growth on heavy-metal-contaminated sites. Its high biomass and deep rooting system make it interesting for phytoextraction. In the present study, we investigated the antioxidative activities and glutathione-dependent enzymes of different tobacco clones optimized for better Cd and Zn accumulation in order to characterize their performance in the field. MAIN FEATURES: The improved heavy metal resistance also makes the investigated tobacco clones interesting for understanding the plant defense enzyme system in general. Freshly harvested plant material (N. tabacum leaves) was used to investigate the antioxidative cascade in plants grown on heavy metal contaminated sites with and without amendments of different ammonium nitrate and ammonium sulfate fertilizers. MATERIALS AND METHODS: Plants were grown on heavily polluted soils in north-east Switzerland. Leaves were harvested at the field site and directly deep frozen in liquid N(2). Studies were concentrated on the antioxidative enzymes of the Halliwell-Asada cycle, and spectrophotometric measurements of catalase (CAT, EC 1.11.1.6), ascorbate peroxidase (APX, EC 1.11.1.11), superoxide dismutase (SOD, EC 1.15.1.1), glutathione peroxidase (GPX, EC 1.11.1.9), glutathione reductase (GR, EC 1.6.4.2), glutathione S-transferase (GST, EC 2.5.1.18) were performed. RESULTS AND DISCUSSION: We tried to explain the relationship between fertilizer amendments and the activity of the enzymatic defense systems. When tobacco (N. tabacum) plants originating from different mutants were grown under field conditions with varying fertilizer application, the uptake of cadmium and zinc from soil increased with increasing biomass. Depending on Cd and Zn uptake, several antioxidant enzymes showed significantly different activities. Whereas SOD and CAT were usually elevated, several other enzymes, and isoforms of GST were strongly inhibited. CONCLUSIONS: Heavy metal uptake represents severe stress to plants, and specific antioxidative enzymes are induced at the cost of more general reactions of the Halliwell-Asada cycle. In well-supplied plants, the glutathione level remains more or less unchanged. The lack of certain glutathione S-transferases upon exposure to heavy metals might be problematic in cases when organic pollutants coincide with heavy metal pollution. When planning phytoremediation of sites, mixed pollution scenarios have to be foreseen and plants should be selected according to both, their stress resistance and hyperaccumulative capacity.

Bioenergy to save the world. Producing novel energy plants for growth on abandoned land.

BACKGROUND AND AIM: Following to the 2006 climate summit, the European Union formally set the goal of limiting global warming to 2 degrees Celsius. But even today, climate change is already affecting people and ecosystems. Examples are melting glaciers and polar ice, reports about thawing permafrost areas, dying coral reefs, rising sea levels, changing ecosystems and fatal heat periods. Within the last 150 years, CO2 levels rose from 280 ppm to currently over 400 ppm. If we continue on our present course, CO2 equivalent levels could approach 600 ppm by 2035. However, if CO2 levels are not stabilized at the 450-550 ppm level, the consequences could be quite severe. Hence, if we do not act now, the opportunity to stabilise at even 550 ppm is likely to slip away. Long-term stabilisation will require that CO2 emissions ultimately be reduced to more than 80% below current levels. This will require major changes in how we operate. RESULTS: Reducing greenhouse gases from burning fossil fuels seems to be the most promising approach to counterbalance the dramatic climate changes we would face in the near future. It is clear since the Kyoto protocol that the availability of fossil carbon resources will not match our future requirements. Furthermore, the distribution of fossil carbon sources around the globe makes them an even less reliable source in the future. We propose to screen crop and non-crop species for high biomass production and good survival on marginal soils as well as to produce mutants from the same species by chemical mutagenesis or related methods. These plants, when grown in adequate crop rotation, will provide local farming communities with biomass for the fermentation in decentralized biogas reactors, and the resulting nitrogen rich manure can be distributed on the fields to improve the soil. DISCUSSION: Such an approach will open new economic perspectives to small farmers, and provide a clever way to self sufficient and sustainable rural development. Together with the present economic reality, where energy and raw material prices have drastically increased over the last decade, they necessitate the development and the establishment of alternative concepts. CONCLUSIONS: Biotechnology is available to apply fast breeding to promising energy plant species. It is important that our valuable arable land is preserved for agriculture. The opportunity to switch from low-income agriculture to biogas production may convince small farmers to adhere to their business and by that preserve the identity of rural communities. PERSPECTIVES: Overall, biogas is a promising alternative for the future, because its resource base is widely available, and single farms or small local cooperatives might start biogas plant operation.

In vitro breeding of Brassica juncea L. to enhance metal accumulation and extraction properties.

In vitro breeding and somaclonal variation were used as tools to improve the potential of Indian mustard (Brassica juncea L.) to extract and accumulate toxic metals. Calli from B. juncea were cultivated on a modified MS medium supplemented with 10-200 microM Cd or Pb. Afterwards, new B. juncea somaclones were regenerated from metal-tolerant callus cells. Three different phenotypes with improved tolerance of Cd, Zn and Pb were observed under hydroponic conditions: enhanced metal accumulation in both shoots and roots; limited metal translocation from roots to shoots; reduced accumulation in shoots and roots. Seven out of thirty individual variants showed a significantly higher metal extraction than the control plants. The improvement of metal shoot accumulation of the best regenerant (3 x Cd, 1.6 x Zn, 1.8 x Pb) and metal extraction (6.2 x Cd, 3.2 x Zn, 3.8 x Pb) indicated a successful breeding and selection of B. juncea, which could be used for phytoremediation purpose.

Chemical mutagenesis--a promising technique to increase metal concentration and extraction in sunflowers.

Since most of the metal-hyperaccumulating wild plants only produce very low biomass and many high-yielding crops accumulate only moderate amounts of metals, the current research is mainly focused on overcoming these limitations and the optimization of metal phytoextraction. The main goal of the present study was the improvement of metal concentration and extraction properties of Helianthus annuus L by chemical mutagenesis (the non-GMO approach). Sunflowers--hybrid cultivar Salut and inbred lines-were treated with the chemical mutagen ethyl methanesulfonate (EMS). The effect of chemical mutagenesis on metal concentration in and extraction by new sunflower M1 and M2 mutants was directly assessed on a metal-contaminated field in Raft, Switzerland. Mutants of the M2 generation showed a 2-3 times higher metal shoot concentration than the control plants. The best M2 sunflower "giant mutant" 14/185/04 showed a significantly enhanced metal extraction ability: 7.5 times for Cd, 9.2 times for Zn, and 8.2 times for Pb in aboveground parts, as compared to the control plants. Theoretical calculations for the phytoextraction potential of new sunflower variants note that the best sunflower mutant can produce up to 26 t dry matter per hectare and remove 13.3 kg Zn per hectare and year at the sewage sludge contaminated site of Raft; that is a gain factor of 9 compared to Zn extraction by sunflower controls. Furthermore, the use of sunflower oil and biomass for technical purposes (lubricants, biodiesel, biogas) should produce an additional value and improve the economical balance of phytoextraction.

Screening of sunflower cultivars for metal phytoextraction in a contaminated field prior to mutagenesis.

Sunflower can be used for the remediation of metal-contaminated soils. Its high biomass production makes this plant species interestingfor phytoextraction and using sunflower oil for a technical purpose may improve the economic balance of phytoremediation. The aim of the present field study was to screen 15 commercial cultivars of Helianthus annuus L. grown on metal-contaminated soil, to find out the variety with the highest metal extraction, which can be further improved by mutation or in vitro breeding procedures. Two different fertilizers (ammonium sulphate and ammonium nitrate) were also used to enhance the bioavailability of metals in soil Highly significant differences were observed within tested varieties for metal accumulation and extraction efficiency. Furthermore, ammonium nitrate increased cadmium extraction, whereas ammonium sulphate enhanced zinc and lead uptake in most tested cultivars. In this field-based sunflower screening, we found enhanced cumulative Cd, Zn, and Pb extraction efficiency by a factor 4.4 for Salut cultivar. We therefore emphasize that prior to any classical breeding or genetic engineering enhancing metal uptake potential, a careful screening of various genotypes should be done to select the cultivar with the naturally highest metal uptake and to start the genetic improvement with the best available plant material.