| The environmental risk of heavy metal pollution is pronounced in soils adjacent to largeindustrial complexes. It is important that the functioning of soil microorganisms inecosystems exposed to long-term contamination by heavy metals. We studied the potentialeffects of heavy metals on microbial biomass, activity and community composition in soilnear a Copper Smelter in China. The results showed microbial biomass C was negativelyaffected by the elevated metal levels and was closely correlated to heavy metal stress. Enzymeactivity was greatly depressed by conditions in the heavy-metal-contaminated sites. Goodcorrelation was observed between enzyme activity and the distance from the smelter. Elevatedmetal loadings resulted in changes in the activity of the soil microbe, as indicated by changesin their metabolic profiles from correlation analysis. Significant decrease of soil phosphataseactivities was found in the soils 200 m away from the smelter. Polymerase chainreaction-denaturing gradient gel electrophoresis (PCR-DGGE) analysis demonstrated thatheavy metals pollution had a significant impact on bacterial and actinomycetic communitystructure. There were negative correlations between soil microbial biomass, phosphataseactivity and NH4NO3 extractable heavy metals. Several species (Vetiveria, rape, mustard)were selected in the field study. Results showed that the availability of heavy metals inrhizosphere soils was decreased compared to the control (no plant). Among these plants,Vetiveria had the best effect on decreasing the availability of heavy metals in rhizosphere. Itwas found that the change of chemical forms heavy metals would result in the diversity ofmicroorganism in soil. The soil microorganism activity and community composition could bepredicted significantly using the availability of Cu and Zn. By combining different monitoringapproaches from different viewpoints, the set of methods applied in this study were sensitiveto site differences and contributed to a better understanding of heavy metals effects on thestructure, size and activity of microbial communities in soils. The data presented demonstratethe role of heavy metals pollution in understanding the heavy metal toxicity to soilmicroorganism near a copper smelter in China.The biomass of plants in the experiment was reduced significantly after copper addition. E.splendens showed a Cu-tolerant plant. The biomass of E.splendens was much higher thanthat of T.repens or C.communius when Cu treatment concentration was 500 mg/kg. Rhizozphere pH was decreased after Cu addition. NHUNOs-extracted Cu in the rhizosphere of E.splendens was lower than that of unplanted, T.repens or C.communius. In addition, rhizophere soil bacteria biomass, enzyme and community composition induced by Cu treatments. It was indicated that soil phosphatase activity was inhibited by Cu, while E.splendens showed a character of Cu-tolerant plant Soil phosphatase activity in the rhizosphere of E.splendens was higher than that of unplanted or other plants. It was concluded from the results of PCR-DGGE that rhzisophere soil of T.repens or C.communius bacteria community composition was reduced after Cu addition. However, the rhizosphere soil of E.splendens bacteria community composition was highest when the Cu treatment concentration was 200 mg/kg. Cloning and DNA sequencing results showed soil bacteria contained a>p, y-Proteobacteria^ actinomycete and so on in the rhziosphere ofE.splendens. Further experiment was done to determine the quantity of specific soil bacteria by Real-time PCR. It was showed that the copy number of $-Proteobacteria in the rhizosphere of E.splendens has an inflection. The copy number was highest in rhizosphere of E.splendens when 200mg/kg Cu addition. In this paper, we present data indicating that rhizosphere microbial activity and community may play an important role in increasing the copper tolerance of plant.The role of rhizosphere bacteria in facilitating the solubility of copper (Cu) in contaminated soil and Cu accumulation in plant were studied. The bacteria strains were isolated from the rhizosphere of Elsholtzia splendens, a Cu accumulator growing on Tonglu Mountain copper mines. After the sandy soils containing 237 mg Kg-1 were incubated with the bacteria strains, it was indicated that rhizosphere microbes played an important role in influencing the availability of water-soluble Cu in soils. Soils had greater concentrations of water-extractable Cu compared with axenic soils inoculated different bacterial strains. Further evidence for bacterial facilitation of increased solubility of Cu in the soil was obtained using the antibiotic ampicillin (O.lmg g'1). There were 36% decreases in Cu concentration in the presence of bacterial strain MS 12 and ampicillin together compared with bacterial inoculation alone. Different bacterial strains had different abilities on soil water-soluble Cu. To achieve the highest rates of plant Cu accumulation, it was necessary for bacteria to be present in therhizosphere of E. splendens. Inoculated plants supplied with 20 umol L'1 CuSO4 had significantly greater concentrations of Cu in shoots and roots than uninoculated plants and bacterial strain MS2 was the most effective strain in promoting plant Cu uptake. There were 2.2-fold and 2.5-fold increases in Cu accumulation in the shoots and roots of plants inoculated with strain MS2 compared to axenic controls. Furthermore, when ampicillin and the bacterial strains were added together to the nutrient solution, the Cu concentrations in roots and shoots of ampicillin-treated plants were lower than those in inoculated plants. When ampicillin was added to the nutrient solution, Cu accumulation was inhibited by about 24-44% in shoots and 20-44% in roots. The above results provided a new insight into the phytoremediation of Cu-contaminated soil.Chelate-assisted phytoremediation has been proposed as an effective tool for the extraction of heavy metals from soil by plants. However, side-effects of the addition of chelate to soil microbial community are usually neglected. We studied the potential effects of chelate (glucose and citric acid) amendment on phytoextraction of copper and microbial community composition in soil under laboratory conditions. A copper (Cu) accumulator, Elsholtzia splendens, and a non-accumulator, Trifolium repens, were grown on a sandy loam soil containing 317 mg kg-1 Cu. Microbial community compositions were analyzed by using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE). The results showed that the biomass of E. splendens grown with the chelate did not differ from that of the control. Addition of citric acid decreased the biomass of T. repens in comparison to that of glucose treatment. Application of glucose or citric acid significantly increased the extractable Cu concentration in planted and unplanted soils. Concentrations of Cu in the shoots of E. splendens were 2.6, 1.9 and 2.9 times of those of T. repens under no chelate, citric acid and glucose treatments respectively. PCR-DGGE fingerprint analysis revealed that there were negative correlations between bacteria diversity and NH4NO3 extractable-Cu under glucose or citric acid treatment. It was indicated the amendment of glucose to the plant T. repens increased the bacteria diversity in soil as compared to that in soils of non-chelate treatment. The above results indicated chelate addition facilitated phytoremediation of soil Cu and did not have a negative effect on microbial community.Laboratory incubation and a pot experiment in greenhouse were conducted undercontrolled environment conditions to study effects of different sulfur compounds on soil solution pH and solubilization of heavy metals in heavy metal-contaminated soils (Cu and Zn) . The results showed that elemental sulfur has potential to solubilize soil metals and thus enhance phytoextraction of these metals from soils. The change of microbial community structures in soils is analyzed by using by using PCR-DGGE. Resuts showed that soil microbial community composition was changed after S addition. The bands were reduced for elevated heavy metals and low pH. The following cloning and sequence results showed the band 1 was correlation with the S oxidation. The band 4 standed for microorganism isolated from metal-rich and acid soil. The aboved results indicated that S was oxidizing by soil microorganism, then reduced the soil pH and facilited heavy metals accumulation in Elsholtzia splendens.
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