Enhanced dissipation of phenanthrene in spiked soil by arbuscular mycorrhizal alfalfa combined with a non-ionic surfactant amendment

Experiments were conducted to assess the role of colonization of alfalfa roots by an arbuscular mycorrhizal (AM) fungus (Glomus etunicatum) in conjunction with a non-ionic surfactant (Triton X-100) in dissipation of phenanthrene in a soil spiked with phenanthrene at 0, 2.5, 5.0 and 10.0 mg kg(-1). After plant harvest the residual phenanthrene concentration in the soil decreased markedly. Mycorrhizal treatment enhanced phenanthrene dissipation in the rhizosphere and bulk soils irrespective of phenanthrene application rate. Addition of Triton X-100 resulted in the highest phenanthrene concentration in the rhizosphere soil among the treatments, while the lowest phenanthrene concentration in the bulk soil was obtained by AM inoculation and amendment with Triton X-100. AM inoculation and addition of the surfactant consistently promoted phenanthrene dissipation in the soil and decreased the microbial biomass based on phospholipid fatty acid (PLFA) analysis. PLFA profiles demonstrated that AM inoculation together with addition of Triton X-100 altered the microbial community structure in the rhizosphere soil. The results of this study provide a reference value for phytoremediation of soil contaminated by organic pollutants.     

Comparative effects of native filamentous and arbuscular mycorrhizal fungi in the establishment of an autochthonous, leguminous shrub growing in a metal-contaminated soil

The aim of this study was to assess the effectiveness of inoculation with a native arbuscular mycorrhizal (AM) fungus, Glomus mosseae (Nicol. and Gerd.) Gerd. and Trappe, or a filamentous fungus, Penicillium aurantiogriseum Dierckx 1901, on the establishment of Coronilla juncea L. seedlings grown in a polluted, semiarid soil. For that, root and shoot biomass, nutrient uptake, mycorrhizal colonisation and nitrate reductase (NR) and phosphatase activities were analysed. Six months after planting, the shoot biomass of C. juncea was increased only by the inoculation with G. mosseae (by about 62% compared with non-mycorrhizal plants). The shoot NR and root acid phosphatase activities were increased more by inoculation with G. mosseae than with P. aurantiogriseum inoculation. The root NR activity and foliar nutrient contents were increased only by the inoculation with the AM fungus. The root Zn and Cu decreased with the AM fungus. In conclusion, the autochthonous AM fungus was an effective inoculant with regard to stimulating growth and alleviating heavy metal toxicity for plants growing on a soil contaminated by multiple heavy metals. Inoculation with an autochthonous, filamentous fungus does not seem to be a good strategy for phytoremediation of such problematic sites.     

Influence of organic matter on the uptake of cadmium, zinc, copper and iron by sorghum plants

This article describes an experiment, carried out under controlled environment conditions, to investigate the effects of a fulvic acid fraction of soil organic matter on growth, cadmium (Cd) uptake and redistribution by sorghum. In addition the uptake of copper (Cu), zinc (Zn) and iron (Fe) was also determined. Sorghum was grown in nutrient solutions with 0, 0.1, 1 and 10 mg Cd dm(-3), in the absence and presence of organic matter (32 mg C dm(-3)), for various periods up to 20 days. A decrease in sorghum biomass due to Cd toxicity was observed at 10 mg Cd dm(-3), but for concentrations of 0.1 and 1 mg Cd dm(-3) the biomass was increased compared with control, without visual toxicity symptoms. The presence of organic matter (OM) further increased biomass production. Cadmium was mainly retained in sorghum roots, as usually found in tolerant plants, but Cd accumulation in sorghum was greater than in other Gramineae, or even more tolerant plants such as lettuce. The presence of OM decreased the bioavailability of Cd that was partially retained in solution by the OM ligands. However, OM promoted the translocation of Cd to shoots, an effect that may pose a risk to public health because plant-animal transfer of Cd could be enhanced. The presence of OM decreased the uptake of Cu, Zn and Fe. The presence (vs. absence) of 0.1 mg Cd dm(-3) enhanced the uptake of Fe, both in the absence and presence of OM.     

Improvement of growth of Eucalyptus globulus and soil biological parameters by amendment with sewage sludge and inoculation with arbuscular mycorrhizal and saprobe fungi

Sewage sludge is widely used as an organic soil amendment to improve soil fertility. We investigated the effects of sewage sludge (SS) application on certain biological parameters of Eucalyptus globulus Labill. The plant was either uninoculated or inoculated with saprobe fungi (Coriolopsis rigida and Trichoderma harzianum) or arbuscular mycorrhizal (AM) fungi (Glomus deserticola and Gigaspora rosea). Sewage sludge was applied to the surface of experimental plots at rates of 0, 2, 4, 6 and 8 g 100 g^− 1 of soil. Inoculation with both AM and saprobe fungi in the presence of SS was essential for the promotion of plant growth. The AM, saprobe fungi and SS significantly increased dry shoot weight. The AM fungi induced a significant increase in Fluorescein diacetate (FDA) activity but did not increase β-glucosidase activity. Addition of SS to AM-inoculated soil did not affect either FDA or α-glucosidase activities in plants from soil that was either uninoculated or inoculated with the saprobe fungi. SS increased β-glucosidase activity when it was applied at 4 g 100 g^− 1. SS negatively affected AM colonization as well as the mycelium SDH activity for both mycorrhizal fungi. SS increased Eucalyptus shoot biomass and enhanced its nutrient status. Inoculation of the soil with G. deserticola stimulated significant E. globulus growth and increases in shoot tissue content of N, P, K, Ca, Mg and Fe. Dual inoculation with G. deserticola and either of the saprobe fungi had positive effects on K, Ca, Mg and Fe contents. The application of 8 g 100 g^− 1 of SS had no positive effects on plant nutrition. The experimental setup provided a suitable tool for evaluating SS in combination with saprobe and AM fungi as a biological fertiliser for its beneficial effects on E. globulus plant growth.     

Impacts of chelate-assisted phytoremediation on microbial community composition in the rhizosphere of a copper accumulator and non-accumulator

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.     

Accumulation of heavy metals in sunflower and sorghum plants affected by the Guadiamar spill

The collapse of a pyrite-mining, tailing dam on 25 April 1998 contaminated approximately 2000 ha of croplands along the Agrio and Guadiamar river valleys in southern Spain. This paper reports the accumulation of chemical elements in soil and in two crops--sunflower and sorghum--affected by the spill. Total concentrations of As, Bi, Cd, Cu, Mn, Pb, Sb, Tl and Zn in spill-affected soils were greater than in adjacent, unaffected soils. Leaves of spill-affected crop plants had higher nutrient (K, Ca and Mg for sunflower and N and K for sorghum) concentrations than controls, indicating a 'fertilising' effect caused by the sludge. Seeds of spill-affected sunflower plants did accumulate more As, Cd, Cu and Zn than controls, but values were below toxic levels. Leaves of sorghum plants accumulated more As, Bi, Cd, Mn, Pb, Tl and Zn than controls, but these values were also below toxic levels for livestock consumption. In general, none of the heavy metals studied in both crops reached either phytotoxic or toxic levels for humans or livestock. Nevertheless, a continuous monitoring of heavy metal accumulation in soil and plants must be established in the spill-affected area.     

Alkaline biosolids and EDTA for phytoremediation of an acidic loamy soil spiked with cadmium

A greenhouse experiment was conducted to investigate the growth of Brassica juncea and Cd phytoextraction in a mimicked Cd contaminated acidic loamy soil amended with alkaline biosolids, prepared from sewage sludge and coal fly ash, in the presence and absence of EDTA at 2 mmol kg(-1). The acidic loamy soil was spiked with 0, 5, 20, 50 and 100 mg Cd kg(-1) in the form of CdCO(3) and then amended with 4% alkaline biosolids (w/w). Alkaline biosolids and 0.12% CaCO(3) amendments resulted in a higher biomass than unamended soil spiked with 20 mg kg(-1) Cd where plants did not survive and of the two amendments, alkaline biosolids amendment had higher plant dry weight yield and phytoextraction of Cd. Adding 2 mmol kg(-1) EDTA to alkaline biosolids amended soil significantly increased the solubility of Cd ions by 9- to 29-fold, but plant Cd accumulation decreased by a factor of 24-48%. The results indicate that alkaline biosolids amendment is an effective approach for assisting growth of B. juncea and phytoextraction of Cd from the contaminated acidic loamy soil, but further application of chelating agents did not enhance the phytoextraction efficiency of Cd.     

Isolation, identification and utilization of thermophilic strains in aerobic digestion of sewage sludge

Two representative thermophilic bacterial strains (T1 and T2) were isolated from a one-stage autothermal thermophilic aerobic digestion pilot-scale reactor. 16S rRNA gene analysis indicated that they were Hydrogenophilaceae and Xanthomonodaceae. These isolated strains were inoculated separately and/or jointly in sewage sludge, to investigate their effects on sludge stabilization under thermophilic aerobic digestion condition. Four digestion conditions were tested for 480 h. Digestion without inoculation and inoculation with strain T2, as well as joint- inoculation with strains T1 and T2, achieved 32.6%, 43.0%, and 38.2% volatile solids (VS) removal, respectively. Removal in a digester inoculated with stain T1 only reached 27.2%. For the first 144 h, the three inoculated digesters all experienced higher VS removal than the digester without inoculations. Both specific thermophilic strains and micro-environment significantly affected the VS removal. DGGE profiles revealed that the isolated strains T1 and T2 can successfully establish in the thermophilic digesters. Other viable bacteria (including anaerobic or facultative microbes) also appeared in the digestion system, enhancing the microbial activity.     

Enhanced removal of polychlorinated biphenyls from alfalfa rhizosphere soil in a field study: The impact of a rhizobial inoculum

Polychlorinated biphenyls (PCB) are persistent pollutants in soil environments where they continue to present considerable human health risks. Successful strategies to remediate contaminated soils are needed that are effective and of low cost. Bioremediation approaches that include the use of plants and microbial communities to promote degradation of PCB have significant potential but need further assessment under field conditions. The effects of growth of alfalfa (Medicago sativa L.) and inoculation with a symbiotic nitrogen fixing bacterium (Rhizobium meliloti) on the removal of polychlorinated biphenyls (PCB) from rhizosphere soil were evaluated in a field experiment. The initial PCB content of the soil ranged from 414 to 498 µg kg⁻¹. PCB removal for the rhizosphere soil was enhanced in the planted treatments, an average of 36% decrease in PCB levels compared to a 5.4% decrease in the unplanted soil, and further enhanced when plants were inoculated with the symbiotic Rhizobium (an average of 43% decrease) when evaluated at 90 days after planting. Plant biomass production was higher in the inoculated treatment. The total PCB content was increased from 3.30 µg kg⁻¹ to 26.72 µg kg⁻¹ in plant shoots, and from 115.07 µg kg⁻¹ to 142.23 µg kg⁻¹ in roots in the inoculated treatment compared to the planted treatment. Increased colony forming units (cfu) of total heterotrophic bacteria, biphenyl-degrading bacteria and fungi were observed in the rhizosphere of inoculated plants. PCB removal from the rhizosphere soil was not significantly correlated with the direct PCB uptake by the plants in any of the treatments but was significantly correlated with the stimulation of rhizosphere microflora. Changes in the soil microbial community structure in the planted and inoculated treatment were observed by profiling of bacterial ribosomal sequences. Some bacteria, such as Flavobacterium sp., may have contributed to the effective degradation of PCB and deserve further investigation.     

Addition of microbially-treated sugar beet residue and a native bacterium increases structural stability in heavy metal-contaminated Mediterranean soils

A mesocosm experiment was conducted to investigate the effect of the addition of Aspergillus niger-treated sugar beet waste, in the presence of rock phosphate, and inoculation with a native, metal-tolerant bacterium, Bacillus thuringiensis, on the stabilisation of soil aggregates of two mine tailings, with differing pH values, from a semiarid Mediterranean area and on the stimulation of growth of Piptatherum miliaceum. Bacterium combined with organic amendment enhanced structural stability (38% in acidic soil and 106% in neutral soil compared with their corresponding controls). Only the organic amendment increased pH, electrical conductivity, water-soluble C, water-soluble carbohydrates and plant growth, in both soils. While in neutral soil both organic amendment and bacterium increased dehydrogenase activity, only organic amendment had a significant effect in acidic soil. This study demonstrates that the use of P. miliaceum in combination with organic amendment and bacterium is a suitable tool for the stabilisation of the soil structure of degraded mine tailings, although its effectiveness is dependent on soil pH.     

Microbial population in a hydrogen-dependent denitrification reactor

The bacterial population in an H2-dependent denitrification system was studied. The laboratory set-up was designed for the treatment of potable water and consisted of an electrochemical cell, where the water to be treated was enriched with H2 prior to entering a bioreactor. Bioreactors (columns packed with granulated active carbon) were inoculated with denitrifying bacterial strains isolated from a previous reactor, then sampled immediately after inoculation, or after 1 or 3 months of continuous operation. Total number of the bacteria and numbers of each different strain were determined at various levels of the bioreactor. The strains present in the inoculum were identified as Ochrobactrum anthropi, Pseudomonas stutzeri, Paracoccus panthotrophus and Paracoccus denitrificans. Numbers of the latter declined markedly with time with the other three strains being responsible for nitrate removal. A correlation was found between the relative abundance of each strain and its specific denitrification activity.     

Genetic Engineering for Wastewater Treatment

This paper discusses the application of genetic engineering to improved wastewater treatment, with particular emphasis on the enhanced breakdown of recalcitrant pollutants. The genetic construction of bacterial strains capable of degrading problem compounds, especially chlorinated aromatics, is described. In addition, the problems affecting the functioning of these 'genetically engineered'laboratory strains in situ is addressed. The results of recent studies into the survival and function of both 'natural'and 'genetically engineered'bacteria inoculated into laboratory-scale activated-sludge units are presented. These results indicate that several factors may affect the expression of catabolic activities in situ , and that those bacteria which are well adapted to growth and survival in target ecosystems are more likely to be successful inoculants than are laboratory strains.     

Phytoprotective influence of bacteria on growth and cadmium accumulation in the aquatic plant Lemna minor

Certain plants are known to accumulate heavy metals, and can be used in remediation of polluted soil or water. Plant-associated bacteria, especially those that are metal tolerant, may enhance the total amount of metal accumulated by the plant, but this process is still unclear. In this study, we investigated metal enhancement vs. exclusion by plants, and the phytoprotective role plant-associated bacteria might provide to plants exposed to heavy metal. We isolated cadmium-tolerant bacteria from the roots of the aquatic plant Lemna minor grown in heavy metal-polluted waters, and tested these isolates for tolerance to cadmium. The efficiency of plants to accumulate heavy metal from their surrounding environment was then tested by comparing L. minor plants grown with added metal tolerant bacteria to plants grown axenically to determine, whether bacteria associated with these plants increase metal accumulation in the plant.Unexpectedly, cadmium tolerance was not seen in all bacterial isolates that had been exposed to cadmium. Axenic plants accumulated slightly more cadmium than plants inoculated with bacterial isolates. Certain isolates promoted root growth, but overall, addition of bacterial strains did not enhance plant cadmium uptake, and in some cases, inhibited cadmium accumulation by plants. This suggests that bacteria serve a phytoprotective role in their relationship with Lemna minor, preventing toxic cadmium from entering plants.     

Effect of alkaline pH and associated Zn on the concentration and total uptake of Cd by lettuce: comparison with predictions from the CLEA model

An eight-fold underestimate of the potential Cd exposure to humans via ingestion of lettuce grown in moderately alkaline soil has been measured experimentally. Current models of Cd uptake by leafy vegetables, which are used in risk assessment (e.g. CLEA in UK) predict higher concentration factors in acid than in alkaline soils. Experimental evidence shows that Cd uptake, although it decreases with increasing pH from acid to neutral soils, increases again in alkaline soils, confirming recent finding from other workers. The concentration of Zn in the soil also significantly affects the uptake of Cd, although this is not included in the current prediction models either. The effect of Zn on the uptake of Cd by plants is greater in slightly alkaline soils (pH 7.7) than in slightly acidic or neutral soils. High concentrations of Zn in soil (1000 mg/kg), which are often associated with elevated Cd levels, further increase the Cd concentration factor to values 12 times higher than that predicted by the CLEA model. This is due in part to the effect of the high soil Zn on reducing the above-ground biomass of the plants.     

Heavy metal and micronutrient uptake in soybeans as influenced by sewage sludge amendment

Soybean (Glycine max (L.) Merr.) uptake of the elements, Cd, Ni, Pb, Cu, Zn and Mn, from a sewage sludge-amended Mecklenburg soil was conducted in the greenhouse. “Bragg” soybeans were grown in pots for five weeks at which time the tops and roots were sampled separately for elemental analysis. Soil samples from each pot were extracted with DTPA (diethylenetriaminepentaacetic acid) and the concentration of extractable elements correlated with the elemental content in the soybean plant. There was a significant increase in dry matter production with sludge treatment. Concentrations of Cd, Ni and Pb in the soybean shoots and roots increased from sludge-amended soil as compared to the control. The metal concentration in the soybean tissue increased with increasing levels of sludge amendment. Uptake of the heavy metals was greater by the roots than by the shoots indicating some barrier to movement of the metals from roots to shoots. The DTPA extractable Cd in sludge-amended soil increased significantly, and showed correlation to the soybean tissue metal concentrations. As for the micronutrients, Cu increased in the soybean shoot as the extractable Cu increased. There was no significant relationship between soybean tissue Zn and Mn and extractable Zn and Mn.     

Using phosphate rock to immobilize metals in soil and increase arsenic uptake by hyperaccumulator Pteris vittata

This greenhouse experiment evaluated the effects of phosphate rock (PR) on arsenic and metal uptake by the arsenic hyperaccumulator Pteris vittata in a soil spiked with arsenic and heavy metals Cd, Pb and Zn. Five soil treatments were used, 1) control with no arsenic, 2) spiked with 50 mg kg(-1) As (As) as Na2H AsO4, 3) spiked with 50 mg kg(-1) As and P as PR (AsP), 4) spiked with 50 mg kg(-1) As, Pb, Cd, and Zn (AsM), and 5) spiked with 50 mg kg(-1) As, Pb, Cd, Zn and P (AsMP). The plants were harvested after growing in the soil for five weeks. Compared to the As treatment, the presence of heavy metals (AsM) reduced arsenic concentrations in the fronds from 1631 to 608 mg kg(-1). However, this effect was mitigated by PR (AsMP), with arsenic concentrations in the fronds increased from 608 to 1046 mg kg(-1). Phosphate rock also significantly reduced Pb (13.5 to 4.10 mg kg(-1)) and Cd (13.0 to 3.45 mg kg(-1)) concentrations in the fronds. Most of the arsenic in P. vittata was accumulated in the fronds (89-93%). Compared to the control, P was more concentrated in the roots along with less P being translocated to the fronds in the treatments with arsenic. While in those same treatments higher Ca concentrations in both the fronds and roots were observed. This research shows that PR was effective in increasing arsenic uptake and decreasing metal uptake by P. vittata and thus can be used as a cost-effective amendment for phytoremediation of arsenic and metal polluted soils.     

Pine forest and grassland differently influence the response of soil microbial communities to metal contamination

Metal pollution can affect soil microbial communities, and vegetation potentially influences this relationship. It can, for example, modify the toxicity of metal to soil microbes by controlling its input to the ground or by altering soil physicochemical properties. This study examined metal effects on soil respiration, potentially active microbial biomass (SIR) and catabolic abilities of culturable heterotrophic bacterial communities (Biolog GN) in pine forest and grassland ecosystems developed on soils contaminated with Zn, Pb and Cd. In samples from non-forested areas we found that metal pollution reduced the microbial biomass and functional diversity of bacteria, while increasing the metabolic quotient. In samples from pine forests we found no relationship between metal pollution and microbial parameters. Metals induced changes in soil respiration neither in forest nor in grassland sites. Generally, microbial performance was determined predominantly by soil physicochemical properties (nutrient content, acidity, contamination level). Vegetation type seemed a minor but important factor influencing microbial communities. More work is needed to determine why even relatively high metal concentrations do not significantly affect microbial communities in forest soils.     

Trace element availability and plant growth in a mine-spill contaminated soil under assisted natural remediation I. Soils

We evaluated the effects of different amendments and/or a plant cover on reclamation of a trace element contaminated soil. Seven treatments were established: four organic (leonardite (LEO), litter (LIT), municipal waste compost (MWC), biosolid compost (BC)), one inorganic (sugar beet lime (SL)) and two controls (control without amendment but with Agrostis (CTRP) and control without amendment and without Agrostis (CTR)). Results showed that total organic C was significantly higher in organic treatments in all samplings. Water-soluble C was lower in CTR compared to other treatments, but no significant differences were observed between organic treatments and SL and CTR. SL, BC and MWC treatments increased soil pH and reduced 0.01 M CaCl2-extractable Cd, Cu and Zn concentrations more efficiently, especially in the first 2 years. At the end of the experiment 0.01 M CaCl2-extractable trace element concentrations were similar in all treatments. 0.01 M CaCl2-extractable As and Pb were below the detection limit. Addition of amendments showed no clear reduction in 0.05 M EDTA-extractable trace element concentrations and some amendments even increased 0.05 M EDTA-extractable As and Cu with time. Pseudo-total trace element concentrations were higher for As in controls. On the other hand, mean values of Cu and Zn were higher in MWC treatment. BC and SL treatments also showed higher Zn mean concentration than controls. No amendment effect was observed for Cd and Pb.     

The assessment of air and soil as contributors of some trace metals to vegetable plants. II. Translocation of atmospheric and laboratory-generated cadmium aerosols to and within vegetable plants

Radish, spinach and lettuce were grown in the field on 109Cd-labelled soil. Efficient transfer to the plant was observed with concentration factors [Cd concn. in plant (dry wt) divided by Cd concn. in soil (dry wt)] of between 0.30 and 2.00. Soil uptake of Cd accounted for only 69-64% of Cd in the plants, the remainder arising from atmospheric deposition. By fumigating radish, lettuce and carrot with Cd aerosol in a wind tunnel and harvesting and analysis subsequent to further growth, translocation of Cd to non-exposed parts of plants was demonstrated. The percentage of deposited Cd which is translocated in the wind-tunnel-fumigated plants is small (0.6-4%), but is larger in plants grown in the field (28-36%) or in a laboratory growth cabinet (12-35%).     

Heterogeneity of cadmium concentration in soil as a source of uncertainty in plant uptake and its implications for human health risk assessment

The major route of exposure of humans to the toxic element cadmium (Cd) is via the consumption of vegetables homegrown on Cd contaminated soil. It is well known that soil pH is one of the main soil properties controlling bioavailability of Cd in plants. This is acknowledged in human health risk assessment models that incorporate pH dependant concentration factors (CF=plant Cd/soil Cd). However, variation in spatial heterogeneity of nutrients and heavy metals in soil can have a profound effect on plant performance and uptake of nutrients and heavy metals. Here we show for lettuce (variety Crispino) that variation in plant-scale heterogeneity of Cd in soil affects bioavailability and hence CF by a factor of 2. Plant yield is also significantly affected. This has important implications for both human health risk assessment, as variation in CF affects predicted exposure, and for phytoremediation where an optimal combination of plant yield and contaminant accumulation is required.