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The role of symbiotic rhizobia in the alleviation of polychlorinated biphenyl (PCB)-induced phytotoxicity in alfalfa and the brassinosteroid (BR) hormone signaling involved were investigated during phytoremediation. The association between alfalfa and Sinorhizobium meliloti was adopted as a remediation model. Phytotoxicity due to PCB 77 (3,3',4,4'-tetrachlorobiphenyl) exerted adverse impacts on plant performance (biomass accumulation and photosynthesis) and elicited cellular oxidative stress (overproduction of reactive oxygen species, lipid peroxidation, and cell necrosis) which was largely attenuated by pre-inoculation with S. meliloti strain NM. The protective role may have been achieved as a result of strengthening of basic antioxidant defense before stress as evidenced by the augmented activity and gene expression of antioxidative enzymes (peroxidase, glutathione reductase, superoxide dismutase, catalase, and ascorbate peroxidase) of both leaves and roots. In nodulated seedlings peroxidase showed additive increased activity following PCB exposure but the activities of the other four enzymes tended to remain stable after stress. Furthermore, application of strain NM and brassinolide both triggered the accumulation of endogenous BRs and the antioxidant network, while pre-treatment of seedlings with a biosynthetic inhibitor of BRs, brassinazole, abolished the rhizobia-induced activation of detoxification responses towards PCB. These observations indicate that association with S. meliloti NM enhanced the systemic antioxidant defenses of alfalfa to detoxify PCB, at least in part, via BR-dependent signaling pathways. These results contribute to our knowledge of the 'logistic role' played by rhizobia in assisting the phytoremediation of PCB-contaminated soils and suggest an optimum manipulation strategy for bioremediation.

Pentachloronitrobenzene (PCNB) is a toxic chlorinated nitroaromatic compound. However, only a few bacteria have been reported to be able to utilize PCNB. In the present study, one pentachloronitrobenzene (PCNB)-degrading bacterium, Cupriavidus sp. YNS-85, was isolated from a contaminated Panax notoginseng plantation. The strain co-metabolized 200 mg L−1 PCNB in aqueous solution with a removal rate of 73.8% after 5 days. The bacterium also degraded PCNB effectively under acid conditions (pH 4–6) and showed resistance to toxic trace elements (arsenic, copper, and cadmium). Its ability to utilize proposed PCNB intermediates as sole carbon sources was also confirmed. The soil microcosm experiment further demonstrated that bacterial bioaugmentation enhanced the removal of PCNB (37.8%) from soil and the accumulation of pentachloroaniline (89.3%) after 30 days. Soil enzyme activity and microbial community functional diversity were positively influenced after bioremediation. These findings indicate that Cupriavidus sp. YNS-85 may be a suitable inoculant for in situ bioremediation of PCNB-polluted sites, especially those with acid soils co-contaminated with heavy metal(loid)s.

A bacterial strain ZY1 capable of utilizing PCBs as its carbon source was isolated from the root nodules of Chinese milk vetch (Astragalus sinicus L.). The strain was identified as Mesorhizobium sp. according to its physiological-biochemical properties and the analysis of its 16S rRNA gene sequence. When the initial OD600 was 0.15, 62.7% of 15 mg L(-1) 3,3',4,4'-TCB in a liquid culture was degraded by Mesorhizobium sp. ZY1 within 10 days. Mesorhizobium sp. ZY1 also greatly increased the biotransformation of soil PCBs. Pot experiments indicated that the soil PCB concentrations of a single incubation of strain ZY1 (R) and a single planting of A. sinicus (P) decreased by 20.5% and 23.0%, respectively, and the concentration of PCBs in soil treated with A. sinicus and strain ZY1 decreased by 53.1%. We also observed that A. sinicus-Mesorhizobium sp. ZY1 treatment (PR) improved plant biomass and the concentration of PCBs in plants compared with a single A. sinicus planting treatment (P). The results suggest that the synergistic association between A. sinicus and PCBs-degrading Mesorhizobium sp. ZY1 can stimulate the phytoextraction of PCBs and the rhizosphere microflora to degrade PCBs, and might be a promising bioremediation strategy for PCB-contaminated soil.

Polycyclic aromatic hydrocarbon (PAH) and metal-polluted sites caused by abandoned coking plants are receiving wide attention. To address the associated environmental concerns, innovative remediation technologies are urgently needed. This study was initiated to investigate the feasibility of a cleanup strategy that employed an initial phase, using methyl-β-cyclodextrin (MCD) solution to enhance ex situ soil washing for extracting PAHs and metals simultaneously, followed by the addition of PAH-degrading bacteria (Paracoccus sp. strain HPD-2) and supplemental nutrients to treat the residual soil-bound PAHs. Elevated temperature (50 °C) in combination with ultrasonication (35 kHz, 30 min) at 100 g MCD L(-1) was effective in extracting PAHs and metals to assist soil washing; 93 % of total PAHs, 72 % of Cd, 78 % of Ni, 93 % of Zn, 84 % of Cr, and 68 % of Pb were removed from soil after three successive washing cycles. Treating the residual soil-bound PAHs for 20 weeks led to maximum biodegradation rates of 34, 45, 36, and 32 % of the remaining total PAHs, 3-ring PAHs, 4-ring PAHs, and 5(+6)-ring PAHs after washing procedure, respectively. Based on BIOLOG Ecoplate assay, the combined treatment at least partially restored microbiological functions in the contaminated soil. The ex situ cleanup strategy through MCD-enhanced soil washing followed by microbial augmentation can be effective in remediating PAH and metal-contaminated soil.

The contamination of soils by polycyclic aromatic hydrocarbons (PAHs) is a widespread environmental problem and the remediation of PAHs from these areas has been a major concern. The effectiveness of many in situ bioremediation systems may be constrained by low contaminant bioavailability due to limited aqueous solubility or a large magnitude of sorption. The objective of this research was to evaluate the effect of methyl-beta-cyclodextrin (MCD) on bioaugmentation by Paracoccus sp. strain HPD-2 of an aged PAH-contaminated soil. When 10% (W/W) MCD amendment was combined with bioaugmentation by the PAH-degrading bacterium Paracoccus sp. strain HPD-2, the percentage degradation of total PAHs was significantly enhanced up to 34.8%. Higher counts of culturable PAH-degrading bacteria and higher soil dehydrogenase and soil polyphenol oxidase activities were observed in 10% (W/W) MCD-assisted bioaugmentation soil. This MCD-assisted bioaugmentation strategy showed significant increases (p0.05) in the average well color development (AWCD) obtained by the BIOLOG Eco plate assay, Shannon-Weaver index (H) and Simpson index (lambda) compared with the controls, implying that this strategy at least partially restored the microbiological functioning of the PAH-contaminated soil. The results suggest that MCD-aided bioaugmentation by Paracoccus sp. strain HPD-2 may be a promising practical bioremediation strategy for aged PAH-contaminated soils.

A field experiment was conducted to study the effects of inoculation with the arbuscular mycorrhizal fungus Glomus caledonium and/or Rhizobium meliloti on phytoremediation of an agricultural soil contaminated with weathered PCBs by alfalfa grown for 180 days. Planting alfalfa (P), alfalfa inoculated with G. caledonium (P + AM), alfalfa inoculated with R. meliloti (P + R), and alfalfa co-inoculated with R. meliloti and G. caledonium (P+AM+R) decreased significantly initial soil PCB concentrations by 8.1, 12.0, 33.8, and 43.5%, respectively. Inoculation with R. meliloti and/or G. caledonium (P+AM+R) increased the yield of alfalfa, and the accumulation of PCBs in the shoots. Soil microbial counts and the carbon utilization ability of the soil microbial community increased when alfalfa was inoculated with R. meliloti and/or G. caledonium. Results of this field study suggest that synergistic interactions between AMF and Rhizobium may have great potential to enhance phytoremediation by alfalfa of an agricultural soil contaminated with weathered PCBs.

A field-scale experiment was carried out to compare the effectiveness of five plant species in the remediation of oily sludge. Alfalfa, tall fescue, and soybean substantially increased the removal rate of oil and grease (O&G) after 120 days of remediation. Of these, soybean treatment showed the highest removal rate of 34.2% compared with only 13.7% in the unplanted control plot. In addition to assisting with phytoremediation, soybean can also be used for energy production, for example in the production of biodiesel. Furthermore, microbial counts and community level physiological profiling using Biolog sole carbon source utilization tests were used to investigate the effect of plants on the microbial community of oily sludge. Plants selectively increased microbial counts in the rhizosphere and O&G concentrations of the sludge were negatively correlated with counts of TPH (total petroleum hydrocarbon) degraders. The acute biotoxicity of the sludge was also tested by the bacterial luminescence method and alfalfa treatments decreased the biological toxicity of the sludge compared with the unplanted control.

In situ bioremediation of oily sludge-contaminated soil by biostimulation of indigenous microbes through adding manure was conducted at the Shengli oilfield in northern China. After bioremediation for 360 days, total petroleum hydrocarbon (TPH) content was reduced by 58.2% in the treated plots compared with only 15.6% in the control plot. Moreover, bioremediation significantly improved the physicochemical properties of the soil in the treated plot. Soil microbial counts and community-level physiological profiling were also examined. Manure addition increased TPH degraders and polycyclic aromatic hydrocarbon (PAH) degraders in the contaminated soil by one to two orders of magnitude. The activity and biodiversity of soil microbial communities also increased markedly in the treated plot compared with that of the control. Finally, biotoxicity was used to evaluate the soils and a sharp increase in the EC50 of the soil after bioremediation was observed, indicating that bioremediation had reduced the toxicity of the soil.

Microcosms were set up with a PAHs-contaminated soil using biostimulation (addition of ground corn cob) and bioaugmentation (inoculated with Monilinia sp. W5-2). Degradation of polycyclic aromatic hydrocarbons and microbial community were examined at the end of incubation period. After 30 days, bioaugmented microcosms showed a 35+/-0% decrease in total PAHs, while biostimulated and control microcosms showed 16+/-9% and 3+/-0% decrease in total PAHs, respectively. Bioaugmented microcosms also revealed 70+/-8% and 72+/-2% decreases in benzo[a]pyrene and anthracene, respectively, while the values for biostimulated and control microcosms were much lower. Detoxification of soils in bioaugmented microcosms was confirmed by genetic toxicity assay, suggesting important role of fungal remediation. Molecular fingerprint profiles and selective enumeration showed biostimulation with ground corn cob increased both number and abundance of indigenous aromatic hydrocarbons degraders and changed microbial community composition in soil, which is beneficial to natural attenuation of PAHs. At the same time, bioaugmentation with Monilinia strain W5-2 imposed negligible effect on indigenous microbial community. This study suggests that fungal remediation is promising in eliminating PAHs, especially the part of recalcitrant and highly toxic benzo[a]pyrene, in contaminated soil. It is also the first description of soil bioremediation with Monilinia sp.

The effectiveness of many bioremediation systems for PAH-contaminated soil may be constrained by low contaminant bioaccessibility due to limited aqueous solubility or large sorption capacity. Information on the extent to which PAHs can be readily biodegraded is of vital importance in the decision whether or not to remediate a contaminated soil. In the present study the rate-limiting factors in methyl-β-cyclodextrin (MCD)-enhanced bioremediation of PAH-contaminated soil were evaluated. MCD amendment at 10 % (w/w) combined with inoculation with the PAH-degrading bacterium Paracoccus sp. strain HPD-2 produced maximum removal of total PAHs of up to 35 %. The desorption of PAHs from contaminated soil was determined before and after 32 weeks of bioremediation. 10 % (w/w) MCD amendment (M2) increased the Tenax extraction of total PAHs from 12 to 30 % and promoted degradation by up to 26 % compared to 6 % in the control. However, the percentage of Tenax extraction for total PAHs was much larger than that of degradation. Thus, in the control and M2 treatment it is likely that during the initial phase the bioaccessibility of PAHs is high and biodegradation rates may be limited by microbial processes. On the other hand, when the soil was inoculated with the PAH-degrading bacterium (CKB and MB2), the slowly and very slowly desorbing fractions (F sl and F vl ) became larger and the rate constants of slow and very slow desorption (k sl and k vl ) became extremely small after bioremediation, suggesting that desorption is likely rate limiting during the second, slow phase of biotransformation. These results have practical implications for site risk assessment and cleanup strategies.

Carbon supplementation, soil moisture and soil aeration are believed to enhance in situ bioremediation of PAH-contaminated soils by stimulating the growth of indigenous microorganisms. However, the effects of added carbon and nitrogen together with soil moisture and soil aeration on the dissipation of PAHs and on associated microbial counts have yet to be fully assessed. In this study the effects on bioremediation of carbon source, carbon-to-nitrogen ratio, soil moisture and aeration on an aged PAH-contaminated agricultural soil were studied in microcosms over a 90-day period. Additions of starch, glucose and sodium succinate increased soil bacterial and fungal counts and accelerated the dissipation of phenanthrene and benzo(a)pyrene in soil. Decreases in phenanthrene and benzo(a)pyrene concentrations were effective in soil supplemented with glucose and sodium succinate (both 0.2 g C kg(-1) dry soil) and starch (1.0 g C kg(-1) dry soil). The bioremediation effect at a C/N ratio of 10:1 was significantly higher (P < 0.05) than at a C/N of either 25:1 or 40:1. Soil microbial counts and PAH dissipation were lower in the submerged soil but soil aeration increased bacterial and fungal counts, enhanced indigenous microbial metabolic activities, and accelerated the natural degradation of phenanthrene and benzo(a)pyrene. The results suggest that optimizing carbon source, C/N ratio, soil moisture and aeration conditions may be a feasible remediation strategy in certain PAH contaminated soils with large active microbial populations.

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 microg 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 microg kg(-)(1) to 26.72 microg kg(-)(1) in plant shoots, and from 115.07 microg kg(-)(1) to 142.23 microg 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.

Field-scale bioremediation of oily sludge in prepared beds was studied at Shengli oilfield in northern China. The influence of manure, coarse sand, sawdust, a specialized microbial preparation and greenhouse conditions on the efficiency of removal of oil and grease was evaluated. After bioremediation for 230d, oil and grease content fell by 32-42gkg(-1)dry sludge in treated plots, indicating removal of 27-46% compared with only 15% in the control plot. Addition of manure, coarse sand, sawdust and greenhouse conditions significantly (p0.05) increased the amount removed. Moreover, the physico-chemical properties of the sludge in all treated plots improved significantly after bioremediation. Microbial biomass in sludge and community-level physiological profiling examined using BIOLOG microplates was also studied. Total petroleum hydrocarbon degraders and polycyclic aromatic hydrocarbon degraders increased in all treated oily sludge. The activity of sludge microbial communities increased markedly in the treated plots compared with the control. Canonical correspondence analysis showed that differences in substrate utilization patterns were highly correlated (p0.05) with sludge hydrolyzable N and oil and grease content. The biological toxicity of the oily sludge was lower following bioremediation in most of the treated plots as evaluated using Photobacterium phosphoreum T3.

Background A climate-controlled pot experiment was conducted to investigate the effects of planting alfalfa and applying organic fertilizer on the dissipation of benzo[a]pyrene from an aged contaminated agricultural soil. Results Short-term planting of alfalfa inhibited the dissipation of benzo[a]pyrene from the soil by 8.9%, and organic fertilizer enhanced benzo[a]pyrene removal from the soil by 11.6% compared with the unplanted and unfertilized treatments, respectively. No significant interaction was observed between alfalfa and organic fertilizer on benzo[a]pyrene dissipation. Sterilization completely inhibited the removal of benzo[a]pyrene from the soil indicating that its degradation by indigenous microorganisms may have been the main mechanism of dissipation. Furthermore, significant positive relationships were observed between benzo[a]pyrene removal and the contents of soil ammonium nitrogen, nitrate nitrogen, and total mineral nitrogen at the end of the experiment, suggesting that competition between plants and microorganisms for nitrogen may have inhibited benzo[a]pyrene dissipation in the rhizosphere of alfalfa and the addition of organic fertilizer may facilitate microbial degradation of benzo[a]pyrene in the soil.