The effect of sulphur supplementation on cadmium phytotoxicity in wheat and lettuce: changes in physiochemical properties of roots and accumulation of phytochelatins.

Intensive sulphur fertilisation has been reported to improve the nutrient balance and growth of Cd-exposed plants, but the reasons of this phenomenon and the role of sulphur compounds in the resistance to cadmium are unclear. We investigated sulphur supplementation-induced changes in the surface properties of roots and the level of thiol peptides (PCs) in Cd-stressed Triticum aestivum L. (monocots clade) and Lactuca sativa L. (dicots clade) grown in nutrient solution. The combination of three sulphur (2 mM S-basic level, 6 or 9 mM S-elevated levels) and four cadmium (0, 0.0002, 0.02 or 0.04 mM Cd) concentrations was used. The physicochemical parameters of the roots were determined based on the apparent surface area (Sr), total variable surface charge (Q), cation exchange capacity (CEC) and surface charge density (SCD). In Cd-exposed plants supplied with sulphur, a different character and trend in the physicochemical changes (adsorption and ion exchange) of roots were noted. At the increased sulphur levels, as a rule, the Sr, CEC, Q and SCD values clearly increased in the lettuce but decreased in the wheat in the entire range of the Cd concentrations, except the enhanced Sr of wheat supplied with 6 mM S together with elevated (0.0002 mM) and unchanged (0.02, 0.04 mM Cd) value of this parameter at 9 mM S. This indicates a clade-specific and/or species-specific plant reaction. The 6 mM S appears to be more effective than 9 mM S in alleviation of the cadmium's toxic effects on roots. It was found that at 0.02 and 0.04 mM Cd, the use of 6 mM S limits the Cd accumulation in the roots of both species in comparison with the basic S fertilisation. Moreover, PC accumulation was much more efficient in wheat than in lettuce, and intensive sulphur nutrition generally induced biosynthesis of these chelating compounds. Physicochemical parameters together with quantitative and qualitative assessment of thiol peptides can be important indicators of the efficiency of root system functioning under cadmium stress. The differences between the species and the multidirectional character of the changes are a result of the involvement of a number of multi-level mechanisms engaged in the defence against metal toxicity.

Zinc- and cadmium-tolerant endophytic bacteria from Murdannia spectabilis (Kurz) Faden. studied for plant growth-promoting properties, in vitro inoculation, and antagonism.

This research aims to isolate and identify Zn- and Cd-tolerant endophytic bacteria from Murdannia spectabilis, identify their properties with and without Zn and Cd stress, and to investigate the effect of bacterial inoculation in an in vitro system. Twenty-four isolates could survive on trypticase soya agar (TSA) supplemented with Zn (250-500 mg L-1) and/or Cd (20-50 mg L-1) that belonged to the genera Bacillus, Pantoea, Microbacterium, Curtobacterium, Chryseobacterium, Cupriavidus, Siphonobacter, and Pseudomonas. Each strain had different indole-3-acetic acid (IAA), 1-aminocyclopropane-1-carboxylate (ACC) deaminase and siderophore production, nitrogen fixation, phosphate solubilization, and lignocellulosic enzyme characteristics. Cupriavidus plantarum MDR5 and Chryseobacterium sp. MDR7 were selected for inoculation into plantlets that were already occupied by Curtobacterium sp. TMIL due to them have a high tolerance for Zn and Cd while showing no pathogenicity. As determined via an in vitro system, Cupriavidus plantarum MDR5 remained in the plants to a greater extent than Chryseobacterium sp. MDR7, while Curtobacterium sp. TMIL was the dominant species. The Zn plus Cd treatment supported the persistence of Cupriavidus plantarum MDR5. Dual and mixed cultivation showed no antagonistic effects between the endophytes. Although the plant growth and Zn/Cd accumulation were not significantly affected by the Zn-/Cd-tolerant endophytes, the inoculation did not weaken the plants. Therefore, Cupriavidus plantarum MDR5 could be applied in a bioaugmentation process.

Effect of drill cuttings addition on physicochemical and chemical properties of soil and red clover (Trifolium pretense L.) growth.

The most economical method of drill cuttings disposal may be their application in land reclamation which allows for the wastes recovery. However, the wastes application into the soil should ensure that the quality of the environment would not be deteriorated. These investigations were aimed at identifying the effect of drill cuttings, which were the mixture of different types of drilling wastes, on the physicochemical properties of acidic soil and growth of red clover (Trifolium pratense L.). The experimental design comprised 5 treatments, which differed in a dose of the drill cuttings: 0% (control), 2.5%, 5%, 10% and 15% of dry weight. A six-week pot experiment was conducted to determine the influence of the wastes on the plant growth. The results showed that the drill cuttings addition significantly changed the chemical and physicochemical properties of the soil, such as: electrical conductivity (EC), pH, base saturation, content of carbonate, alkaline cations (Ca2+, Na+, K+, Mg2+), organic matter, total organic carbon (TOC), and available phosphorus form. However, the most important factors that influenced the growth of red clover were pH, base saturation, content of Mg2+ and plant available phosphorus. The red clover biomass was increased from 1.5 to 2.5 times depending on the dose of wastes. We concluded that the examined wastes can be used for reclamation of the acid and unfertile degraded soils, but the amount of wastes should not exceed 5% of the soil, because the highest total clover biomass was observed just at this dose.

Rice straw biochar and phosphorus inputs have more positive effects on the yield and nutrient uptake of Lolium multiflorum than arbuscular mycorrhizal fungi in acidic Cd-contaminated soils.

The purpose of the study was to examine biochar amendment, phosphorus (P) fertilizer and arbuscular mycorrhizal fungi (AMF) on the yield, nutrient and cadmium (Cd) absorption of Lolium multiflorum in acidic soil. It was shown that mycorrhizal inoculation had no positive influence on the plant shoot biomass and the contents of nitrogen (N), P, potassium (K), calcium (Ca) and magnesium (Mg) in plants at all biochar and P level treatments. Irrespective of mycorrhizal inoculation and P level, biochar amendments markedly elevated the soil available P and K uptake in plant tissues. In contrast, biochar significantly decreased the translocation factor of plants, soil exchangeable Cd, and acid and neutral phosphatase activities, regardless of the mycorrhizal inoculation and P fertilizer. Without P fertilization, biochar amendments significantly promoted shoot P content, while biochar amendments significantly reduced shoot P content when P fertilizer was applied. Without biochar application, P fertilizer application significantly promoted the biomass and N uptake of shoots in both AMF inoculation treatments, while P fertilizer increased these only in the presence of biochar and mycorrhizal inoculation. The increased N content induced by the biochar amendment elevated the shoot N:P ratio and alleviated the N deficiency with P fertilizer input. Thus, we concluded that the addition of biochar and P fertilizer showed more positive effects on the promotion of growth and nutrient uptake of L. multiflorum than AMF grown in acidic Cd-contaminated soils.

Pectin/Chitosan/Tripolyphosphate Nanoparticles: Efficient Carriers for Reducing Soil Sorption, Cytotoxicity, and Mutagenicity of Paraquat and Enhancing Its Herbicide Activity.

As a potent herbicide capable of contaminating water and soil environments, paraquat, which is still widely used worldwide, is toxic to mammals, algae, aquatic animals, etc. Paraquat was loaded on novel nanoparticles composed of pectin, chitosan, and sodium tripolyphosphate (PEC/CS/TPP). The size, polydispersity index, and ζ potential of nanoparticles were characterized. Further assessments were carried out by SEM, AFM, FT-IR, and DSC. The encapsulation was highly efficient, and there was a delayed release pattern of paraquat. The encapsulated herbicide was less toxic to alveolar and mouth cell lines. Moreover, the mutagenicity of the formulation was significantly lower than those of pure or commercial forms of paraquat in a Salmonella typhimurium strain model. The soil sorption of paraquat and the deep soil penetration of the nanoparticle-associated herbicide were also decreased. The herbicidal activity of paraquat for maize or mustard was not only preserved but also enhanced after encapsulation. It was concluded that paraquat encapsulation with PEC/CS/TPP nanoparticles is highly efficient and the formulation has significant herbicide activity. It is less toxic to human environment and cells, as was evidenced by less soil sorption, cytotoxicity, and mutagenicity. Hence, paraquat-loaded PEC/CS/TPP nanoparticles have potential advantages for future use in agriculture.

Effects of boron, silicon and their interactions on cadmium accumulation and toxicity in rice plants.

Cadmium (Cd) is a highly toxic heavy metal for both animals and plants. Rice consumption is a major source of Cd intake for human. Minimization of Cd accumulation in rice is key to reduce Cd hazard to human. Here we showed alleviating effects of boron (B), silicon (Si) and their mixture on Cd accumulation and toxicity in hydroponically-cultured rice plants. Cd treatment (100 µM) led to Cd accumulation in roots and shoots, as well as significant reduction in plant growth. However, amendment of either B or Si significantly alleviated Cd accumulation and toxicity. Moreover, simultaneous supply of B and Si showed better alleviating effect. However, addition of B and Si alleviated Cd-induced oxidative stress in Cd-treated plants as reflected by reduced MDA, H2O2 and O2-, as well as increased activities of major antioxidant enzymes. Cd exposure induced the expression of Cd transporter genes of OsHMA2, OsHMA3, OsNramp1 and OsNramp5. In contrast, simultaneous supplement of B and Si in Cd-treated plants compromised the gene expression. Our results show that both B and Si alleviate Cd accumulation and toxicity by improving oxidative stress and suppressing Cd uptake and transport, and the two elements display joint effect.

Glyphosate application increased catabolic activity of gram-negative bacteria but impaired soil fungal community.

Glyphosate is a non-selective organophosphate herbicide that is widely used in agriculture, but its effects on soil microbial communities are highly variable and often contradictory, especially for high dose applications. We applied glyphosate at two rates: the recommended rate of 50 mg active ingredient kg-1 soil and 10-fold this rate to simulate multiple glyphosate applications during a growing season. After 6 months, we investigated the effects on the composition of soil microbial community, the catabolic activity and the genetic diversity of the bacterial community using phospholipid fatty acids (PLFAs), community level catabolic profiles (CLCPs), and 16S rRNA denaturing gradient gel electrophoresis (DGGE). Microbial biomass carbon (Cmic) was reduced by 45%, and the numbers of the cultivable bacteria and fungi were decreased by 84 and 63%, respectively, under the higher glyphosate application rate. According to the PLFA analysis, the fungal biomass was reduced by 29% under both application rates. However, the CLCPs showed that the catabolic activity of the gram-negative (G-) bacterial community was significantly increased under the high glyphosate application rate. Furthermore, the DGGE analysis indicated that the bacterial community in the soil that had received the high glyphosate application rate was dominated by G- bacteria. Real-time PCR results suggested that copies of the glyphosate tolerance gene (EPSPS) increased significantly in the treatment with the high glyphosate application rate. Our results indicated that fungi were impaired through glyphosate while G- bacteria played an important role in the tolerance of microbiota to glyphosate applications.

Effects of Rhizophagus clarus and P availability in the tolerance and physiological response of Mucuna cinereum to copper.

Arbuscular mycorrhizal fungi (AMF) improve plant ability to uptake P and tolerate heavy metals. This study aimed to evaluate the effect of available P and the inoculation of Rhizophagus clarus in a Cu-contaminated soil (i) on the activity of acid phosphatases (soil and plant), the presence of glomalin, and (ii) in the biochemical and physiological status of Mucuna cinereum. A Typic Hapludalf soil artificially contaminated by adding 60 mg kg-1 Cu was used in a 3 × 2 factorial design with three replicates. Treatments consisted of three P levels: 0, 40, and 100 mg kg-1 P. Each P treatment level was inoculated (+AMF)/non-inoculated (-AMF) with 200 spores of R. clarus per pot, and plants grown for 45 days. The addition of at least 40 mg kg-1 P and the inoculation of plants with R. clarus proved to be efficient to reduce Cu phytotoxicity and increase dry matter yield. Mycorrhization and phosphate fertilization reduced the activity of enzymes regulating oxidative stress (SOD and POD), and altered the chlorophyll a fluorescence parameters, due to the lower stress caused by available Cu. These results suggest a synergism between the application of P and the inoculation with R. clarus, favoring the growth of M. cinereum in a Cu-contaminated soil. This study shows that AMF inoculation represents an interesting alternative to P fertilization to improve plant development when exposed to excess Cu.

Effects and mechanisms of meta-sodium silicate amendments on lead uptake and accumulation by rice.

The objectives of this research were to study the effects of Na2SiO3 application on the uptake, translocation, and accumulation of Pb in rice and to investigate the mechanisms of Pb immobilization by Na2SiO3 in paddy rice soils and rice plants. Pot experiments were conducted using a Cd-Pb-Zn-polluted soil and Oryza sativa L. ssp. indica cv. Donglian 5. L3-edge X-ray absorption spectroscopy was used to identify Pb species in soils and roots. The results showed that the application of Na2SiO3 increased soil pH and available soil Si but decreased DTPA-extractable Pb in the soil. High dose of Na2SiO3 (12.5 g/kg) reduced the Pb level in brown rice as it inhibited Pb transfer from soil to rice grains, especially Pb transfer from the root to the stem. The Pb X-ray absorption near-edge spectroscopic analysis revealed that application of high dose of Na2SiO3 increased Pb-ferrihydrite and PbSiO3 precipitates in the soil and in the root while it reduced Pb-humic acids (Pb-HAs) in the soil and Pb-pectin in the root. The decrease in Pb availability in the soil can be partly attributed to increase the precipitation of PbSiO3 and the association of Pb2+ with Fe oxides in the soil. The inhibition of the root-to-stem translocation of Pb was partially due to the precipitation of PbSiO3 on the root surfaces or inside the roots.

Leaf defense system of Robinia pseudoacacia L. seedlings exposed to 3years of elevated atmospheric CO2 and Cd-contaminated soils.

Short-term exposure to elevated CO2 increases cadmium (Cd) uptake in some plant species (wheat, poplars, and willows), which triggers an increase in antioxidative system activity to deal with additional reactive oxygen species that are generated. Here, we examined leaf defenses in Robinia pseudoacacia L. seedlings exposed to elevated CO2+Cd for 3years. Three years of elevated CO2 decreased Cd uptake into leaves and the Cd content in soils and increased the pH of rhizosphere soil relative to ambient CO2. In plants exposed to Cd stress, leaf chlorophyll content was greater under elevated CO2 than under ambient CO2. Superoxide dismutase, peroxidase, and catalase activity increased, glutathione content increased, and malondialdehyde and phytochelatins contents decreased under elevated CO2+Cd relative to Cd alone. Proline, soluble sugars, flavonoids, saponins, and phenolic acids contents were greater under elevated CO2+Cd than under Cd alone, and condensed tannin content was lower. Overall, long-term elevation of CO2 enhanced the leaf defense system of R. pseudoacacia exposed to Cd by stimulating antioxidant enzyme activity, osmotic adjustment, and the production of glutathione, flavonoids and phenolic acids. Future research should focus on understanding the mechanisms involved in the decrease in Cd uptake into leaves and Cd content in soils and the increase in rhizosphere soil pH under long-term exposure to elevated CO2.

A pathway of bisphenol A affecting mineral element contents in plant roots at different growth stages.

Bisphenol A (BPA), an environmental endocrine disruptor, is an important industrial raw material. The wide use of BPA has increased the risk of BPA release into the environment, and it has become a new environmental pollutant. In this work, the ecological deleterious effects of this new pollutant on soybean roots at different growth stages were investigated by determining the contents of mineral elements (P, K, Ca, and Mg) and analyzing root activity and the activities of critical respiratory enzymes (hexokinase, phosphofructokinase, pyruvate kinase, and isocitrate dehydrogenase). Our results revealed that low dose (1.5mg/L) of BPA increased the levels of P, K, Mg, and Ca in soybean roots at different growth stages. Whereas, high doses (6.0 and 12.0mg/L) of BPA decreased the levels of P, K, and Mg contents in a dose-dependent manner. BPA had a promotive effect on the content of Ca in soybean roots. Synchronous observation showed that the aforementioned dual response to BPA were also observed in the root activity and respiratory enzyme activities. The effects of BPA on the mineral element contents, root activity and respiratory enzyme activities in soybean roots at different growth stages followed the order: flowering and podding stage>seed-filling stage>seedling stage (mineral element contents); seedling stage>flowering and podding stage>seed-filling stage (root activity and respiratory enzyme activities). In a word, the response of plant root activity and respiratory enzyme activities to BPA pollution is a pathway of BPA affecting mineral element contents in plant roots.

Selenium exposure results in reduced reproduction in an invasive ant species and altered competitive behavior for a native ant species.

Competitive ability and numerical dominance are important factors contributing to the ability of invasive ant species to establish and expand their ranges in new habitats. However, few studies have investigated the impact of environmental contamination on competitive behavior in ants as a potential factor influencing dynamics between invasive and native ant species. Here we investigated the widespread contaminant selenium to investigate its potential influence on invasion by the exotic Argentine ant, Linepithema humile, through effects on reproduction and competitive behavior. For the fecundity experiment, treatments were provided to Argentine ant colonies via to sugar water solutions containing one of three concentrations of selenium (0, 5 and 10 µg Se mL(-1)) that fall within the range found in soil and plants growing in contaminated areas. Competition experiments included both the Argentine ant and the native Dorymyrmex bicolor to determine the impact of selenium exposure (0 or 15 µg Se mL(-1)) on exploitation- and interference-competition between ant species. The results of the fecundity experiment revealed that selenium negatively impacted queen survival and brood production of Argentine ants. Viability of the developing brood was also affected in that offspring reached adulthood only in colonies that were not given selenium, whereas those in treated colonies died in their larval stages. Selenium exposure did not alter direct competitive behaviors for either species, but selenium exposure contributed to an increased bait discovery time for D. bicolor. Our results suggest that environmental toxins may not only pose problems for native ant species, but may also serve as a potential obstacle for establishment among exotic species.

Response and resilience of soil microbial communities inhabiting in edible oil stress/contamination from industrial estates.

BACKGROUND: Gauging the microbial community structures and functions become imperative to understand the ecological processes. To understand the impact of long-term oil contamination on microbial community structure soil samples were taken from oil fields located in different industrial regions across Kadi, near Ahmedabad, India. Soil collected was hence used for metagenomic DNA extraction to study the capabilities of intrinsic microbial community in tolerating the oil perturbation. RESULTS: Taxonomic profiling was carried out by two different complementary approaches i.e. 16S rDNA and lowest common ancestor. The community profiling revealed the enrichment of phylum "Proteobacteria" and genus "Chromobacterium," respectively for polluted soil sample. Our results indicated that soil microbial diversity (Shannon diversity index) decreased significantly with contamination. Further, assignment of obtained metagenome reads to Clusters of Orthologous Groups (COG) of protein and Kyoto Encyclopedia of Genes and Genomes (KEGG) hits revealed metabolic potential of indigenous microbial community. Enzymes were mapped on fatty acid biosynthesis pathway to elucidate their roles in possible catalytic reactions. CONCLUSION: To the best of our knowledge this is first study for influence of edible oil on soil microbial communities via shotgun sequencing. The results indicated that long-term oil contamination significantly affects soil microbial community structure by acting as an environmental filter to decrease the regional differences distinguishing soil microbial communities.

Performance of rose scented geranium (Pelargonium graveolens) in heavy metal polluted soil vis-à-vis phytoaccumulation of metals.

An investigation was carried out to evaluate the effect of heavy metal toxicity on growth, herb, oil yield and quality and metal accumulation in rose scented geranium (Pelargonium graveolens) grown in heavy metal enriched soils. Four heavy metals (Cd, Ni, Cr, and Pb) each at two levels (10 and 20 mg kg-1 soil) were tested on geranium. Results indicated that Cr concentration in soil at 20 mg kg-1 reduced leaves, stem and root yield by 70, 83, and 45%, respectively, over control. Root growth was significantly affected in Cr stressed soil. Nickel, Cr, and Cd concentration and accumulation in plant increased with higher application of these metals. Chromium, nickel and cadmium uptake was observed to be higher in leaves than in stem and roots. Essential oil constituents were generally not significantly affected by heavy metals except Pb at 10 and 20 ppm, which significantly increased the content of citronellol and Ni at 20 ppm increased the content of geraniol. Looking in to the higher accumulation of toxic metals by geranium and the minimal impact of heavy metals on quality of essential oil, geranium can be commercially cultivated in heavy metal polluted soil for production of high value essential oil.

Effect of Arbuscular Mycorrhizal Fungi On Yield and Phytoremediation Performance of Pot Marigold (Calendula officinalis L.) Under Heavy Metals Stress.

In order to study the effect of mycorrhizal fungi (inoculated and non-inoculated) and heavy metals stress [0, Pb (150 and 300 mg/kg) and Cd (40 and 80 mg/kg)] on pot marigold (Calendula officinalis L.), a factorial experiment was conducted based on a randomized complete block design with 4 replications in Research Greenhouse of Department of Horticultural Sciences, University of Tehran, Iran, during 2012-2013. Plant height, herbal and flower fresh and dry weight, root fresh and dry weight and root volume, colonization percentage, total petal extract, total petal flavonoids, root and shoot P and K uptakes, and Pb and Cd accumulations in root and shoot were measured. Results indicated that with increasing soil Pb and Cd concentration, growth and yield of pot marigold was reduced significantly; Cd had greater negative impacts than Pb. However, mycorrhizal fungi alleviated these impacts by improving plant growth and yield. Pot marigold concentrated high amounts of Pb and especially Cd in its roots and shoots; mycorrhizal plants had a greater accumulation of these metals, so that those under 80 mg/kg Cd soil(-1) accumulated 833.3 and 1585.8 mg Cd in their shoots and roots, respectively. In conclusion, mycorrhizal fungi can improve not only growth and yield of pot marigold in heavy metal stressed condition, but also phytoremediation performance by increasing heavy metals accumulation in the plant organs.

Metals other than uranium affected microbial community composition in a historical uranium-mining site.

To understand the links between the long-term impact of uranium and other metals on microbial community composition, ground- and surface water-influenced soils varying greatly in uranium and metal concentrations were investigated at the former uranium-mining district in Ronneburg, Germany. A soil-based 16S PhyloChip approach revealed 2358 bacterial and 35 archaeal operational taxonomic units (OTU) within diverse phylogenetic groups with higher OTU numbers than at other uranium-contaminated sites, e.g., at Oak Ridge. Iron- and sulfate-reducing bacteria (FeRB and SRB), which have the potential to attenuate uranium and other metals by the enzymatic and/or abiotic reduction of metal ions, were found at all sites. Although soil concentrations of solid-phase uranium were high, ranging from 5 to 1569 µg·g (dry weight) soil(-1), redundancy analysis (RDA) and forward selection indicated that neither total nor bio-available uranium concentrations contributed significantly to the observed OTU distribution. Instead, microbial community composition appeared to be influenced more by redox potential. Bacterial communities were also influenced by bio-available manganese and total cobalt and cadmium concentrations. Bio-available cadmium impacted FeRB distribution while bio-available manganese and copper as well as solid-phase zinc concentrations in the soil affected SRB composition. Archaeal communities were influenced by the bio-available lead as well as total zinc and cobalt concentrations. These results suggest that (i) microbial richness was not impacted by heavy metals and radionuclides and that (ii) redox potential and secondary metal contaminants had the strongest effect on microbial community composition, as opposed to uranium, the primary source of contamination.

Effect of phosphogypsum on growth, physiology, and the antioxidative defense system in sunflower seedlings.

Phosphogypsum (PG) is the solid waste product of phosphate fertilizer production and is characterized by high concentrations of salts, heavy metals, and certain natural radionuclides. The work reported in this paper examined the influence of PG amendment on soil physicochemical proprieties, along with its potential impact on several physiological traits of sunflower seedlings grown under controlled conditions. Sunflower seedlings were grown on agricultural soil substrates amended with PG at rates of 0, 2.5, and 5 %. The pH of the soil decreased but electrical conductivity and organic matter, calcium, phosphorus, sodium, and heavy metal contents increased in proportion to PG concentration. In contrast, no variations were observed in magnesium content and small increases were recorded in potassium content. The effects of PG on sunflower growth, leaf chlorophyll content, nutritional status, osmotic regulator content, heavy metal accumulation, and antioxidative enzymes were investigated. Concentrations of trace elements in sunflower seedlings grown in PG-amended soil were considerably lower than ranges considered phytotoxic for vascular plants. The 5 % PG dose inhibited shoot extension and accumulation of biomass and caused a decline in total protein content. However, chlorophyll, lipid peroxidation, proline and sugar contents, and activities of antioxidant enzymes such as superoxide dismutase and catalase increased. Collectively, these results strongly support the hypothesis that enzymatic antioxidation capacity is an important mechanism in tolerance of PG salinity in sunflower seedlings.

Copper tolerance mechanisms of Mesorhizobium amorphae and its role in aiding phytostabilization by Robinia pseudoacacia in copper contaminated soil.

The legume-rhizobium symbiosis has been proposed as an important system for phytoremediation of heavy metal contaminated soils due to its beneficial activity of symbiotic nitrogen fixation. However, little is known about metal resistant mechanism of rhizobia and the role of metal resistance determinants in phytoremediation. In this study, copper resistance mechanisms were investigated for a multiple metal resistant plant growth promoting rhizobium, Mesorhizobium amorphae 186. Three categories of determinants involved in copper resistance were identified through transposon mutagenesis, including genes encoding a P-type ATPase (CopA), hypothetical proteins, and other proteins (a GTP-binding protein and a ribosomal protein). Among these determinants, copA played the dominant role in copper homeostasis of M. amorphae 186. Mutagenesis of a hypothetical gene lipA in mutant MlipA exhibited pleiotropic phenotypes including sensitivity to copper, blocked symbiotic capacity and inhibited growth. In addition, the expression of cusB encoding part of an RND-type efflux system was induced by copper. To explore the possible role of copper resistance mechanism in phytoremediation of copper contaminated soil, the symbiotic nodulation and nitrogen fixation abilities were compared using a wild-type strain, a copA-defective mutant, and a lipA-defective mutant. Results showed that a copA deletion did not affect the symbiotic capacity of rhizobia under uncontaminated condition, but the protective role of copA in symbiotic processes at high copper concentration is likely concentration-dependent. In contrast, inoculation of a lipA-defective strain led to significant decreases in the functional nodule numbers, total N content, plant biomass and leghemoglobin expression level of Robinia pseudoacacia even under conditions of uncontaminated soil. Moreover, plants inoculated with lipA-defective strain accumulated much less copper than both the wild-type strain and the copA-defective strain, suggesting an important role of a healthy symbiotic relationship between legume and rhizobia in phytostabilization.

Arbuscular mycorrhizal symbiosis influences arsenic accumulation and speciation in Medicago truncatula L. in arsenic-contaminated soil.

In two pot experiments, wild type and a non-mycorrhizal mutant (TR25:3-1) of Medicago truncatula were grown in arsenic (As)-contaminated soil to investigate the influences of arbuscular mycorrhizal fungi (AMF) on As accumulation and speciation in host plants. The results indicated that the plant biomass of M. truncatula was dramatically increased by AM symbiosis. Mycorrhizal colonization significantly increased phosphorus concentrations and decreased As concentrations in plants. Moreover, mycorrhizal colonization generally increased the percentage of arsenite in total As both in shoots and roots, while dimethylarsenic acid (DMA) was only detected in shoots of mycorrhizal plants. The results suggested that AMF are most likely to get involved in the methylating of inorganic As into less toxic organic DMA and also in the reduction of arsenate to arsenite. The study allowed a deeper insight into the As detoxification mechanisms in AM associations. By using the mutant M. truncatula, we demonstrated the importance of AMF in plant As tolerance under natural conditions.

Transformation of heavy metal fractions on soil urease and nitrate reductase activities in copper and selenium co-contaminated soil.

This study aims to explore the effects of the distribution, transformation and bioavailability of different fractions of copper (Cu) and selenium (Se) in co-contaminated soils on soil enzymes, providing references for the phytoremediation of contaminated areas and agriculture environmental protection. Pot experiments and laboratory analysis were used to investigate the transformation and bioavailability of additional Cu and Se for pakchoi (Brassica chinensis) in co-contaminated soil. In the uncontaminated soil, Cu mainly existed in residual form, whereas Se was present in residual form and in elemental and organic-sulfide matter-bound form. In the contaminated soil, Cu mainly bound to Fe-Mn oxidates, whereas Se was in exchangeable and carbonates forms. After a month of pakchoi growth, Cu tended to transfer into organic matter-bound fractions, whereas Se tended to bound to Fe-Mn oxidates. The IR (reduced partition index) value of Cu decreased as the concentrations of Cu and Se gradually increased, whereas the IR value of Se decreased as the concentration of Se increased. The IR value before pakchoi planting and after it was harvested was not affected by the concentration of exogenous Cu. Soil urease and nitrate reductase activities were inhibited by Cu and Se pollution either individually or combined in different degrees, following the order nitrate reductase>urease. The significant correlation between the IR value and soil enzyme activities suggests that this value could be used to evaluate the bioavailability of heavy metals in soil. Path analysis showed that the variations in exchangeable Cu and organic-sulfide matter-bound and elemental Se had direct effects on the activities of the two enzymes, suggesting their high bioavailability. Therefore, the IR value and the transformation of metals in soil could be used as indicators in evaluating the bioavailability of heavy metals.