Plant-Soil-Contaminant Specificity Affects Phytoremediation of Organic Contaminants

The objective of this study was the evaluation of seven forage and conservation crop species for phytoremediation of trinitrotoluene (TNT) and pyrene-contaminated soils. TNT and pyrene were added to soil at 100 mg kg^-1. Crop species screening studies were conducted in a greenhouse and growth chambers on two soil types with different organic matter contents. Under high soil organic matter conditions, adsorption or covalent binding to the soil organic matter appeared to be a dominant force of removal limiting TNT and pyrene availability. In both soil types, pyrene dissipation could not be attributed to the presence of plants. However, in soils with lower organic matter content, all of the plant species treatments showed a significantly higher degree of TNT transformation compared with the unplanted control. Statistically significant differences in TNT transformation were observed among crop species grown in the low OM soil. Reed canary grass (Phalaris arundinacea L.) and switchgrass (Panicum virgatum L.) were the most effective species in enhancing TNT transformation. Our data indicated that use of plants was effective for phytoremediation of TNT-contaminated low OM content soils, but did not have any significant effect on pyrene dissipation. Based on these observations, it appears that plant-soil-contaminant interactions are very specific, and this specificity determines the effectiveness of phytoremediation schemes.     

Long-Term Explosive Contamination in Soil: Effects on Soil Microbial Community and Bioremediation

Explosive contamination in soil is a great concern for environmental health. Following 50 years of munitions manufacturing and loading, soils from two different sites contained ≥ 6,435 mg 2,4,6-trinitrotoluene (TNT), 2,933 mg hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) and 2,135 mg octahydrol-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX) kg^{? 1} soil. Extractable nitrate-N was as high as 315 and ammonium-N reached 150 mg N kg^{? 1} soil. Water leachates in the highly contaminated soils showed near saturation levels of TNT and RDX, suggesting great risk to water quality. The long-term contamination resulted in undetectable fungal populations and as low as 180 bacterial colony forming units (CFU) g^–1 soil. In the most severely contaminated soil, dehydrogenase activity was undetectable and microbial biomass carbon was very low (< 3.4 mg C _mic kg^–1 soil). The diminished biological activity was a consequence of long-term contamination because short-term (14 d) contamination of TNT at up to 5000 mg TNT kg^–1 soil did not cause a decline in the culturable bacterial population. Natural attenuation may not be a feasible remediation strategy in soils with long-term contamination by high concentrations of explosives.     

Effect of 2,4,6-trinitrotoluene on soil bacterial communities

To gain insight into the impact of 2,4,6-trinitrotoluene (TNT) on soil microbial communities, we characterized the bacterial community of several TNT-contaminated soils from two sites with different histories of contamination and concentrations of TNT. The amount of extracted DNA, the total cell counts and the number of CFU were lower in the TNT-contaminated soils. Analysis of soil bacterial diversity by DGGE showed a predominance of Pseudomonadaceae and Xanthomonadaceae in the TNT-contaminated soils, as well as the presence of Caulobacteraceae. CFU from TNT-contaminated soils were identified as Pseudomonadaceae, and, to a lesser extent, Caulobacteraceae. Finally, a pristine soil was spiked with different concentrations of TNT and the soil microcosms were incubated for 4 months. The amount of extracted DNA decreased in the microcosms with a high TNT concentration [1.4 and 28.5 g TNT/kg (dry wt) of soil] over the incubation period. After 7 days of incubation of these soil microcosms, there was already a clear shift of their original flora towards a community dominated by Pseudomonadaceae, Xanthomonadaceae, Comamonadaceae and Caulobacteraceae. These results indicate that TNT affects soil bacterial diversity by selecting a narrow range of bacterial species that belong mostly to Pseudomonadaceae and Xanthomonadaceae.     

Promising Strategies for the Mineralisation of 2,4,6-trinitrotoluene

2,4,6-trinitrotoluene (TNT) is known to be one of the most common military explosives. In spite of its established toxicity and mutagenicity for many organisms, soils and groundwater are still being frequently contaminated at manufacturing, disposal and TNT destruction sites. The inability of natural aquatic and soil biota to use TNT as growth substrate has been recognized as the primary limitation in the application of bioremediation processes to contaminated environments. However, promising degradation pathways have been recently discovered which may lead to the mineralisation of TNT. Significant advances have been made in studying the mechanism of TNT denitration, which can be considered as the major reaction and the driving force towards beneficial biodegradation. The possibilities to favour TNT denitration are discussed based on current knowledge of the enzymology and genetics of denitration in nitroaromatic degrading organisms. The literature survey demonstrates that the only enzymes characterized so far for their denitrase activity towards TNT belong to the class I flavin-dependent β/α barrel oxidoreductases, also known as the “Old Yellow Enzyme” family. In addition, this review provides an overview of strategies and future directions towards a rational search for new catabolic activities, including metagenomic library screening, plus new possibilities to improve the activity of known catabolic enzymes acting on TNT, such as DNA shuffling.     

Fate and Transport of 2,4,6-Trinitrotoluene in Loams at a Former Explosives Factory

Natural attenuation processes affecting 2,4,6-trinitrotoluene (TNT) were determined within loams for two study areas at the former Explosives Factory Maribyrnong, Australia. TNT fate and transport was investigated through spectrophotometric/High Performance Liquid Chromatography (HPLC) analyses of soil and groundwater, adsorption and microcosm testwork. A five tonne crystalline TNT source zone delineated within near surface soils at the base of a TNT process waste lagoon was found to be supplying aqueous TNT loading (7 ppm) to subsurface soils and groundwater. The resultant plume was localized within the loam aquitard due to a combination of natural attenuation processes and hydrogeological constraints, including low hydraulic conductivity and upward hydraulic gradients. Freundlich described sorptive partitioning was the main TNT sink (K_F = 29 mL/g), while transformation rates were moderate (1.01 × 10^-4 h^-1) under the aerobic conditions. Increasing 2-amino-4,6-dinitrotoluene predominance over 4-amino-2,6-dinitrotoluene was discovered with depth (in situ) and time (microcosms). Simplified dissolution rate calculations indicate that without mitigation of the TNT source, contaminant persistence within the vadose zone may approach 2000 years, while ATRANS20 simulations demonstrate that the TNT plume propagates very slowly along the flow path within the aquitard.     

Transformation of 2,4,6-trinitrotoluene (TNT) by <Emphasis Type="Italic">Raoultella</Emphasis> <Emphasis Type="Italic">terrigena</Emphasis>

Manufacture of nitroorganic explosives generates toxic wastes leading to contamination of soils and waters, especially groundwater. For that reason bacteria living in environments highly contaminated with 2,4,6-trinitrotoluene (TNT) and other nitroorganic compounds were investigated for their capacity for TNT degradation. One isolate, Raoultella terrigena strain HB, removed TNT at concentrations between 10 and 100 mg l⁻¹ completely from culture supernatants under optimum aerobic conditions within several hours. Only low concentrations of nutrient supplements were needed for the cometabolic transformation process. Radioactivity measurements with ring-labelled ¹⁴C–TNT detected about 10–20% of the initial radioactivity in the culture supernatant and the residual 80–90% as water-insoluble organic compounds in the cellular pellet. HPLC analysis identified aminodinitrotoluenes (2-ADNT, 4-ADNT) and diaminonitrotoluenes (2,4-DANT) as the metabolites which remained soluble in the culture medium and azoxy-dimers as the main products in the cell extracts. Hence, the new isolate could be useful for the removal of TNT from contaminated waters.     

Bioavailability of 2, 4, 6-Trinitrotoluene (TNT) to Earthworms in Three Different Types of Soils in China

To study the effects of aging time (the length of time when contaminants are sequestered in soil) and soil properties on TNT bioavailability in soil, earthworms (Eisenia fetida) were exposed to three types of soils (fluvo-aquic soil, loessal soil, and black soil) contaminated by TNT for 7, 14, 21, 28, 35, and 42 days. The Earthworm-Soil Accumulation Factor (ESAF) of TNT and soil properties were analyzed. The ESAFs in black soil were significantly lower than those in fluvo-aquic soil and loessal soil (P < 0.05). In loessal soils, the ESAF increased with aging time, while that in black soils decreased. The ESAF of TNT had a significantly negative correlation with soil organic matter content, clay contents, and cation exchange capacity, which were the main factors affecting the TNT bioavailability in soils (P < 0.01). There was more quartz and feldspar in black soil, as well as more particles and micropores on the surface, which resulted in the easy adsorption and lower bioavailability of TNT. In conclusion, TNT bioavailability in soils is affected by aging time, soil physical and chemical properties, and mineral and surface properties, which must be considered when biotreatment for TNT in soils is applied.     

Microbial Transformation of Nitroaromatics in Surface Soils and Aquifer Materials

Microorganisms indigenous to surface soils and aquifer materials collected at a munitions-contaminated site transformed 2,4,6-trinitrotoluene (TNT), 2,4-dinitrotoluene (2,4-DNT), and 2,6-dinitrotoluene (2,6-DNT) to amino-nitro intermediates within 20 to 70 days. Carbon mineralization studies with both unlabeled (TNT, 2,4-DNT, and 2,6-DNT) and radiolabeled ([¹⁴C]TNT) substrates indicated that a significant fraction of these source compounds was degraded to CO₂.     

Detection of 2,4,6-Trinitrotoluene-Utilizing Anaerobic Bacteria by 15N and 13C Incorporation

2,4,6-Trinitrotoluene (¹⁵N or ¹³C labeled) was added to Norfolk Harbor sediments to test whether anaerobic bacteria use TNT for growth. Stable-isotope probing (SIP)-terminal restriction fragment length polymorphism (TRFLP) detected peaks in the [¹⁵N]TNT cultures (60, 163, and 168 bp). The 60-bp peak was also present in the [¹³C]TNT cultures and was related to Lysobacter taiwanensis.It has been estimated that there are over 1 million cubic yards of material contaminated with 2,4,6-trinitrotoluene (TNT) in the United States at concentrations as high as 600,000 to 700,00 mg/kg of material (9). Marine and estuarine sediments have also been impacted through the manufacturing, use, and/or disposal of TNT. Microbial biodegradation of these pollutants in situ is preferable due to the large volume of contaminated soils/sediments. However, it is unclear whether in situ bacteria can utilize TNT as a nitrogen or carbon source. Under aerobic conditions, TNT appears to be largely unavailable to bacteria but can be used by a variety of fungi as a carbon and nitrogen source (7). Under anaerobic conditions, only a few bacterial strains (Clostridium and Desulfovibrio strains and Pseudomonas sp. strain JLR11) have been reported to utilize TNT as a sole nitrogen source (6, 7). It is widely believed that nitroaromatic compounds cannot serve as growth substrates under anaerobic conditions in situ (11), and coamendment strategies are suggested for stimulating TNT transformation to 2,4,6-triaminotoluene (TAT) (1, 7, 18). Given these difficulties, there is no direct evidence that TNT can be biodegraded in situ and there is little proof that anaerobic bacteria can utilize TNT as a sole carbon or nitrogen source in organic-rich sediments. This study tested whether bacteria in Norfolk Harbor sediment are able to incorporate nitrogen (N) or carbon (C) from TNT into biomass under sulfidogenic conditions using stable-isotope probing (SIP). The findings indicate that bacteria assimilate ¹⁵N and ¹³C from TNT into their genomes during anaerobic incubations (2 to 35 days). Interestingly, one small-subunit (SSU) gene, related to Lysobacter taiwanensis, was observed in both the ¹⁵N and the ¹³C incubations.     

Effective biodegradation of 2,4,6-trinitrotoluene using a novel bacterial strain isolated from TNT-contaminated soil

In this environmental-sample based study, rapid microbial-mediated degradation of 2,4,6-trinitrotoluene (TNT) contaminated soils is demonstrated by a novel strain, Achromobacter spanius STE 11. Complete removal of 100 mg L⁻¹ TNT is achieved within only 20 h under aerobic conditions by the isolate. In this bio-conversion process, TNT is transformed to 2,4-dinitrotoluene (7 mg L⁻¹), 2,6-dinitrotoluene (3 mg L⁻¹), 4-aminodinitrotoluene (49 mg L⁻¹) and 2-aminodinitrotoluene (16 mg L⁻¹) as the key metabolites. A. spanius STE 11 has the ability to denitrate TNT in aerobic conditions as suggested by the dinitrotoluene and NO₃ productions during the growth period. Elemental analysis results indicate that 24.77 mg L⁻¹ nitrogen from TNT was accumulated in the cell biomass, showing that STE 11 can use TNT as its sole nitrogen source. TNT degradation was observed between pH 4.0–8.0 and 4–43 °C; however, the most efficient degradation was at pH 6.0–7.0 and 30 °C.     

Sensitive biomarker of polycyclic aromatic hydrocarbons (PAHs): urinary 1-hydroxyprene glucuronide in relation to smoking and low ambient levels of exposure

The lysosomal neutral red retention time (NRRT) assay, a biomarker for lysosomal membrane stability, and the total immune activity (TIA) assay, a measure of non-specific immune system activity, were used in laboratory studies to assess the toxic effects of 2,4,6-trinitrotoluene (TNT) on earthworms (Eisenia andrei) in vivo. The results were compared with the concentration of TNT and its metabolites in earthworm tissue, as well as standard sublethal toxicity endpoints including growth (i.e. weight change) and reproduction effects from previously published studies. Filter paper experiments indicated a significant decrease in NRRT at ≥1.8 μg TNT cm^-2, whereas sublethal (weight loss) and lethal effects to earthworms were detected at ≥3.5 and 7.1 μg TNT cm^-2, respectively. Experiments in artificial soil showed that NRRT effects could be detected at lower TNT concentrations (≥55 mg TNT kg^-1 soil dry weight) compared with other sublethal endpoints (effects on growth and reproduction). The TIA biomarker did not significantly respond to TNT. Copper (as CuSO₄, filter paper contact tests) and 2-chloroacetamide (soil tests), which were used as reference toxicants, also decreased the NRRT. The use of the NRRT assay linked with tissue concentrations of TNT metabolites in earthworms was identified as a potentially appropriate biomarker approach for TNT exposure assessment under laboratory conditions and a novel tool for effects-based risk assessment.     

Mass Balance Studies with 14C-Labeled 2,4,6-Trinitrotoluene (TNT) Mediated by an Anaerobic Desulfovibrio Species and an Aerobic Serratia Species

Investigations were carried out to evaluate the level of incorporation of radiolabeled 2,4,6-trinitrotoluene (TNT) and metabolites into the bacterial biomass of two different bacterial species after cometabolically mediated TNT transformation. Biotransformation experiments with ¹⁴C-TNT indicated that TNT was not mineralized; however, carbon derived from TNT became associated with the cells. It was found that more than 42% of the initially applied radiolabel was associated with the cell biomass after cometabolic ¹⁴C-TNT transformation with the strictly anerobic Desulfovibrio species strain SHV, whereas with the strictly aerobic Serratia plymuthica species strain B7, 32% of cell-associated ¹⁴C activity was measured. The remainder of the radiolabel was present in the supernatants of the liquid cultures in the form of different TNT metabolites. Under anoxic conditions with the Desulfovibrio species, TNT was ultimately transformed to 2,4,6-triaminotoluene (TAT) and both diaminonitrotoluene isomers, whereas under oxic conditions with the Serratia species, TNT was converted to hydroxylaminodinitrotoluenes and aminodinitrotoluenes, with 4-amino-2,6-dinitrotoluene (4ADNT) being the major end product. In both culture supernatants, small amounts of very polar, radiolabeled, but unidentified metabolites were detected. At the end of the experiments approximately 92% and 96% of the originally applied radioactivity was recovered in the studies with the Serratia and Desulfovibrio species, respectively. Received: 21 May 1998 / Accepted: 6 July 1998     

Biodegradation of 2,4,6-trinitrotoluene byPhanerochaete chrysosporium: Identification of initial degradation products and the discovery of a TNT metabolite that inhibits lignin peroxidases

Biodegradation of 2,4,6-trinitrotoluene (TNT) by the wood-rotting BasidiomycetePhanerochaete chrysosporium was studied in a fixed-film silicone membrane bioreactor and in agitated pellected cultures. The initial intermediate products of TNT biodegradation were shown to be 2-amino-4,6-dinitrotoluene (2amDNT) and 4-amino-2,6-dinitrotoluene (4amDNT). These intermediates were also degraded byP. chrysosporium. However, their rates of degradation were slow and appeared to represent rate-limiting steps in TNT degradation. The fact that 2amDNT and 4amDNT were further degraded is of importance. In most other microbial systems these compounds are typically not further degraded or are dimerized to even more persistent azo and azoxydimers. Similar to previous studies performed in stationary cultures, it was shown that substantial amounts of [¹⁴C]-TNT were degrade to [¹⁴C]-carbon dioxide in agitated pelleted cultures. Lignin peroxidase activity (assayed by veratryl alcohol oxidation) virtually disappeared upon addition of TNT to ligninolytic cultures ofP. chrysosporium. However, TNT, 2amDNT, and 4amDNT did not inhibit lignin peroxidase activity, nor were they substrates for this enzyme. Subsequent studies revealed that 4-hydroxylamino-2,6-dinitrotoluene, an intermediate in TNT reduction, was a potent lignin peroxidase inhibitor. Further studies revealed that this compound was also a substrate for lignin peroxidase H8.     

Remediation of Explosive-Contaminated Soils: Alkaline Hydrolysis and Subcritical Water Degradation

Alkaline hydrolysis and subcritical water degradation were investigated as ex-situ remediation processes to treat explosive-contaminated soils from military training sites in South Korea. The addition of NaOH solution to the contaminated soils resulted in rapid degradation of the explosives. The degradation of explosives via alkaline hydrolysis was greatly enhanced at pH ≥12. Estimated pseudo-first-order rate constants for the alkaline hydrolysis of 2,4-dinitrotoluene (DNT), 2,4,6-trinitrotoluene (TNT) and hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) in contaminated soil at pH 13 were (9.6?±?0.1)×10^?2, (2.2?±?0.1)×10^?1, and (1.7?±?0.2)×10^?2 min^?1, respectively. In the case of subcritical water degradation, the three explosives were completely removed at 200–300°C due to oxidation at high temperatures and pressures. The degradation rate increased as temperature increased. The pseudo-first-order rate constants for DNT, TNT, and RDX at 300°C were (9.4?±?0.8)×10^?2, (22.8?±?0.3)×10^?2, and (16.4?±?1.0)×10^?2, respectively. When the soil-to-water ratio was more than 1:5, the extent of alkaline hydrolysis and subcritical water degradation was significantly inhibited.     

Immunogenicity of Iodine 131 chimeric tumor necrosis therapy monoclonal antibody in advanced lung cancer patients

Purpose Iodine-131 radiolabeled chimeric tumor necrosis therapy monoclonal antibody (¹³¹I-TNT) has been approved for the treatment of advanced lung cancer in China. In the present study, the immunogenicity of TNT was studied in advanced lung cancer patients using BIACORE and enzyme linked immunosorbent assay (ELISA) methods. Experimental design Serum samples from 78 advanced lung cancer patients were analyzed for antibody development to TNT after systemic or intratumoral administration of two doses of ¹³¹I-TNT. Patients’ sera were obtained before, and 2 weeks and 2 months after ¹³¹I-TNT radioimmunotherapy. Results Four of 78 lung cancer patients (4/78 or 5.13%) developed antibodies to TNT as measured by ELISA method, and 7 of 78 patients (8.97%) development anti-TNT antibody as measured by BIACORE biosensor after 2 doses of ¹³¹I-TNT administration (P > 0.05). All the 4 ELISA-positive patients were also BIACORE-positive. Among the 7 BIACORE-positive patients, 5 (of 42, 11.9%) patients receiving intravenous TNT injection developed antibodies to TNT, and 2 (of 36, 5.56%) patients, receiving intratumoral therapy developed antibodies to TNT. The route of administration of the radiolabeled TNT antibody was not a statistically significant factor in the incidence of anti-TNT antibody. Detailed BIACORE serological analysis showed that the induced antibodies were mostly of the IgG1 subclass. Conclusions ¹³¹I-TNT was immunogenic in only a small minority of advanced lung cancer patients (8.97%). The route of administration did not statistically influence the incidence of anti-TNT antibody after TNT radioimmunotherapy in lung cancer patients.     

Transformation of 2,4,6-trinitrotoluene by white rot fungus Irpex lacteus

Up to 200 mg 2,4,6-trinitrotoluene (TNT) l^–1 was removed within 12 h after adding it to a 5-day old culture of Irpex lacteus. The initial formation of hydroxylamino-dinitrotoluenes (2- and 4-OHAmDNT) from TNT was detected, followed by their successive transformation to aminodinitrotoluenes (2- and 4-AmDNT). Transformation of TNT to AmDNT via OHAmDNT was fast, but the next step was slow and seemed to be a rate-limiting step in TNT degradation. OHAmDNT isomers were also rapidly transformed by an in vitro enzymatic system. Both the mycelium and extracellular enzymes of I. lacteus were required for the TNT degradation.     

Influence of ionic strength on sodium-calcium exchange of two temperate climate soils

Salt-affected soils have been studied extensively with respect to their Na–Ca exchange properties. These studies have focused on soil environments of the arid West. However, because of irrigation and oil well brine discharges in the temperate region of the U.S. there is need to understand sodicity behavior of such soils. In this study, two Kentucky soils (Pembroke and Uniontown) at the 0–10 cm depth were studied to evaluate the influence of ionic strength (I) and sodium adsorption ratio (SAR) on cation selectivity coefficients. The data showed that both soils exhibit at least two classes of exchange sites and in general the apparent affinity for Na⁺ increased when solution ionic strength increased. Furthermore, both soils under all three ionic strengths tested showed greater affinity for Na⁺ than the average agricultural saline soil of the arid West. The data suggested the need for establishing critical salt dispersion thresholds for temperate climate soils and developing effective brine management approaches.     

Chaotropic effects on 2,4,6-trinitrotoluene uptake by wheat (Triticum aestivum)

Previous research in our laboratory investigated the effectiveness of a common agrochemical, urea used as a chaotropic agent to facilitate 2,4,6-trinitrotoluene (TNT) removal by vetiver grass (Vetiveria zizanioides L.). Chaotropic agents disrupt water structure, increasing solubilization of hydrophobic compounds (TNT), and enhancing plant TNT uptake. Our findings showed that urea significantly enhanced TNT uptake kinetics by vetiver. We hypothesized that the beneficial effect of urea on the overall TNT uptake by vetiver grass was not plant-specific. We explored this hypothesis by testing the ability of wheat (Triticum aestivum L.) in removing TNT from aqueous media in the presence of urea. Results showed that untreated (no urea) wheat exhibited a slow, kinetically limited TNT uptake that was nearly half of the urea-treated wheat TNT capacity (250 mg kg⁻¹). Chaotropic effects of urea were illustrated by the significant (P < 0.001) increase in the TNT second-order reaction rate constants over those of the untreated (no urea) controls. Plant TNT speciation showed that TNT and several of its metabolites were detected in both root and shoot compartments of the plant, allowing for 110 and 36% recovery for the untreated and 0.1% urea treated plants. The lower % recovery of the urea-treated plants was attributed to a number of unknown polar TNT metabolites. Responsible Editor: Hans Lambers.     

Metabolism of 2,4,6-trinitrotoluene by the white-rot fungus Bjerkandera adusta DSM 3375 depends on cytochrome P-450

Degradation of 2,4,6-trinitrotoluene (TNT) by the white-rot fungus Bjerkandera adusta DSM 3375 was studied in relation to extracellular ligninolytic activities. The Mn(II)-dependent peroxidase, the only ligninolytic enzyme detectable, reached a maximum activity of 600 ± 159 U/l after incubation in mineral medium with a sufficient nitrogen source. In contrast, the highest extent of [¹⁴C]TNT mineralization was detected in malt extract broth, so that the ability of B. adusta to mineralize TNT did not parallel ligninolytic activity. The microsomal fraction of cells grown in the presence of TNT was found to contain 11 pmol cytochrome P-450/mg protein. In cells grown without TNT, no microsomal cytochrome P-450 could be found. Instead, 14 pmol P-450/mg protein was present in the cytosolic fraction of these cells. Cytochrome P-450 apparently affected the TNT metabolism, as shown by inhibitory studies. Addition of the cytochrome P-450 inhibitor piperonyl butoxide diminished the ¹⁴CO₂ release from 21% to 0.9%, as determined after 23 days of incubation, while 1-aminobenzotriazole and metyrapone decreased the mineralization to 8.6% and 6.3% respectively. Mass-balance analysis of TNT degradation in liquid cultures revealed that, by inhibition of cytochrome P-450, the TNT-derived radioactivity associated with biomass and with polar, water-soluble metabolites decreased from 93.9% to 15.0% and the fraction of radiolabelled metabolites extractable with organic solvents fell to 92.6%. The TNT metabolites of this fraction were identified as aminodinitrotoluenes, indicating that this initial transformation product of TNT may function as a substrate for cytochrome-P-450-dependent reactions in B. adusta. Received: 27 May 1999 / Received revision: 19 August 1999 / Accepted: 19 August 1999     

Characterization of Bacteria in the Rhizosphere Soils of Polygonum Pubescens and Their Potential in Promoting Growth and Cd,Pb, Zn Uptake by Brassica napus

Microbe-enhanced phytoremediation has been considered as a promising measure for the remediation of metal-contaminated soils. In this study, two bacterial strains JYX7 and JYX10 were isolated from rhizosphere soils of Polygonum pubescens grown in metal-polluted soil and identified as of Enterobacter sp. and Klebsiella sp. based on 16S rDNA sequences, respectively. JYX7 and JYX10 showed high Cd, Pb and Zn tolerance and increased water-soluble Cd, Pb and Zn concentrations in culture solution and metal-added soils. Two isolates produced plant growth-promoting substances such as indole acetic acid, siderophore, 1-aminocyclopropane-1-carboxylic deaminase, and solubilized inorganic phosphate. Based upon their ability in metal tolerance and solubilization, two isolates were further studied for their effects on growth and accumulation of Cd, Pb, and Zn in Brassica napus (rape) by pot experiments. Rapes inoculated with JYX7 and JYX10 had significantly higher dry weights, concentrations and uptakes of Cd, Pb, Zn in both above-ground and root tissues than those without inoculation grown in soils amended with Cd (25 mg kg^?1), Pb (200 mg kg^?1) or Zn (200 mg kg^?1). The present results demonstrated that JYX7 and JYX10 are valuable microorganism, which can improve the efficiency of phytoremediation in soils polluted by Cd, Pb, and Zn.