Aged PAH-contaminated Soils Research Articles

Effects of different organic substrate compositions on the decontamination of aged PAH-polluted soils through outdoor co-composting

The effects of different organic substrate compositions on the efficiency of outdoor co-composting as a bioremediation technology for decontaminating soil polluted by polycyclic aromatic hydrocarbons (PAHs) were investigated. Four different substrate mixtures and two different aged PAH-contaminated soils were used in a semi-pilot-scale experiment that lasted nearly 700 days. The two soils (A and B) differed concerning both the initial concentrations of the Ʃ16 US EPA PAHs (5926 vs. 369 mg kg−1, respectively) and the type of predominant PAH group by molecular weight.The experiments revealed that while the composition of the organic substrate had an impact on the rate of PAH degradation, it did not significantly influence the final extent of PAH degradation. Notably, the organic substrate consisting of green waste and wood chips (GW) was found to facilitate the most rapid rate of PAH degradation (first-order rate constant k = 0.033 ± 0.000 d−1 with soil A over the initial 42 days of the experiment and k = 0.036 ± 0.000 d−1 with soil B over the initial 56 days). Despite the differences in organic substrate compositions and types of soil being treated, PAH degradation levels exceeded at least 95% in all the treatments after more than 680 days of co-composting.Regardless of the composition, the removal of low- and medium- molecular-weight (2-4 rings) PAHs was nearly complete by the end of the experiment. Furthermore, high-molecular-weight PAHs (5 rings and more) were significantly degraded during co-composting, with reductions ranging from 54% to 79% in soil A and from 59% to 68% in soil B. All composts were dominated by Proteobacteria, Firmicutes, and Actinobacteria, with significant differences in abundance between soils. Genera with PAH degradation potentials were detected in all samples. The results of a battery of toxicity tests showed that there was almost no toxicity associated with the final composts.

Changes of microbial community and activity under different electric fields during electro-bioremediation of PAH-contaminated soil

Electro-bioremediation is a promising technology for remediation of soil contaminated with persistent organic compounds such as polycyclic aromatic hydrocarbons (PAHs). During electro-bioremediation, electrical fields have been shown to increase pollutant degradation. However, it remains unclear whether there is an optimal strength for the electrical field applied that is conductive to the maximum role played by microbes. This study aimed to determine the optimal strength of electric field through the analysis of the effects of different voltages on the microbial community and activity. Four bench-scale experiments with voltages of 0, 1, 2 and 3 V cm−1 were conducted for 90 days in an aged PAH-contaminated soil. The spatiotemporal changes of the soil pH, moisture content and temperature, microbial biomass and community structure, and the degradation extent of PAHs were researched over 90 days. The results indicated that the total microbial biomass and degradation activity were highest at voltages of 2 V cm−1. The concentration of total phospholipid fatty acids, used to quantify soil microbial biomass, reached 65.7 nmol g−1 soil, and the mean degradation extent of PAHs was 44.0%. Similarly, the maximum biomass of actinomycetes, bacteria and fungus also occurred at the voltage of 2 V cm−1. The Gram-positive/Gram-negative and (cy17:0+cy 19:0)/(16:1ω7+18:1ω7) ratios also showed that the intensity of electric field and electrode reactions strongly influenced the microbial community structure. Therefore, to optimize the electro-bioremediation of PAH-contaminated soil, the strength of electric field needs to be selected carefully. This work provides reference for the development of novel electrokinetically enhanced bioremediation processes.

Evaluation of fatty acid derivatives in the remediation of aged PAH-contaminated soil and microbial community and degradation gene response

In this study, derivatives of two common fatty acids in plant root exudates, sodium palmitate and sodium linoleate (sodium aliphatates), were added to an aged Polycyclic aromatic hydrocarbons (PAHs) contaminated soil to estimate their effectiveness in the removal of PAHs. Sodium linoleate was more effective in lowering PAHs and especially high-molecular-weight (4–6 ring) PAHs (HMW-PAHs). Principal coordinates analysis (PCoA) indicates that both amendments led to a shift in the soil bacterial community. Moreover, linear discriminant effect size (LEfSe) analysis demonstrates that the specific PAHs degraders Pseudomonas, Arenimonas, Pseudoxanthomonas and Lysobacter belonging to the γ-proteobacteria and Nocardia and Rhodococcus belonging to the Actinobacteria were the biomarkers of, respectively, sodium linoleate and sodium palmitate amendments. Correlation analysis suggests that four biomarkers in the sodium linoleate amendment treatment from γ-proteobacteria were all highly linearly negatively related to HMW-PAHs residues (p < 0.01) while two biomarkers in the sodium palmitate amendment treatment from Actinobacteria were highly linearly negatively related to LMW-PAHs residues (p < 0.01). Higher removal efficiency of PAHs (especially HMW-PAHs) in the sodium linoleate amendment treatment than in the sodium palmitate amendment treatment might be ascribed to the specific enrichment of microbes from the γ-proteobacteria. The bacterial functional KEGG orthologs (KOs) assigned to PAHs metabolism and functional C23O and C12O genes related to cleavage of the benzene ring were both up-regulated. These results provide new insight into the mechanisms of the two sodium aliphatate amendments in accelerating PAHs biodegradation and have implications for practical application in the remediation of PAHs-contaminated soils.

PAH biodegradation by telluric saprotrophic fungi isolated from aged PAH-contaminated soils in mineral medium and historically contaminated soil microcosms

Remediation of contaminated soils is of high relevance considering losses of this limited resource in most countries through erosion or through destruction for societal purposes. Most physicochemical remediation techniques lead to soil destruction avoiding reuse of the sites and loss of soil functions including ecological services like water retention/filtration, plant needs, carbon sequestration, or atmosphere restoration. This study aims to find out efficient telluric PAH-degrading fungi to both remediate soils and preserve their functioning. Fifty telluric saprotrophic fungi were thus isolated from different aged PAH-contaminated soils sampled from four brownfields in the North of France. Thirty of these isolates were screened in a mineral medium for their ability to degrade benzo[a]pyrene (BaP) used as a model of HMW PAH. After 9 days of incubation, the remaining BaP was quantified through HPLC analysis. Then, a set of microcosms was performed with an aged PAH-contaminated non-sterile soil encompassing different approaches with bioaugmentation using mycelia of three strains pre-established on expanded clay particles and, for one of these strains, biostimulation strategies including nitrogen or nitrogen/phosphorous supplementations or aeration by stirring. After an incubation time of 30 days, remaining PAH were quantified through GC-MS and evolution of fungal populations evaluated through qPCR. Isolated Penicillium canescens, Cladosporium cladosporioides, Fusarium solani, and Talaromyces helicus degraded more than 30% of the initial 500 μg of BaP after 9 days of incubation. The three latest strains were thus inoculated in aged PAH-contaminated soil microcosms. After 30 days, PAH quantitative analyses showed that the highest degradation was obtained by bioaugmentation with T. helicus (26% of the initial total PAH content: 321.7 mg kg−1), which seems high considering an aged industrial contamination. Biostimulation approaches coupled to the inoculation of this strain did not improve the degradation. DNA quantification of the inoculated species confirmed their enrichment in the soil revealing the interest of the used inoculation strategy. We discuss bioaugmentation and biostimulation approaches in the case of the considered pollutants and soil. A strain identified as T. helicus appears interesting for its HMW PAH degradation capacities both in mineral medium with pure BaP and in industrial non-sterile soil microcosms contaminated with a hydrocarbon complex mixture. This study also confirms the efficiency of a well-studied BaP-degrading strain of F. solani. These results indicate the potentialities of the used bioaugmentation approach and underline the necessity of scale-up strategies to apply this kind of technique on site.

Effective remediation of aged HMW-PAHs polluted agricultural soil by the combination of Fusarium sp. and smooth bromegrass (Bromus inermis Leyss.)

Fusarium sp. strain ZH-H2 is capable to degrade high molecular weight polycyclic aromatic hydrocarbons (HMW-PAHs), smooth bromegrass (Bromus inermis Leyss.) can also degrade 4- to 6-ring PAHs. Pot experiments were conducted to investigate how bromegrass and different inoculum sizes of ZH-H2 clean up HMW-PAHs in agricultural soil derived from a coal mine area. The results showed that, compared with control, different sizes of inocula of ZH-H2 effectively degraded HMW-PAHs, with removal rates of 19.01, 34.25 and 29.26% for 4-, 5- and 6-ring PAHs in the treatment with 1.0 g kg−1 ZH-H2 incubation after 90 d. After 5 mon of cultivation, bromegrass reached degradation rate of these compounds by 12.66, 36.26 and 36.24%, respectively. By adding strain ZH-H2 to bromegrass, HMW-PAHs degradation was further improved up to 4.24 times greater than bromegrass (W), in addition to the degradation rate of Bbf decrease. For removal rates of both 5- and 6-ring PAHs, addition of 0.5 g kg−1Fusarium ZH-H2 to pots with bromegrass performed better than addition of 0.1 g kg−1, while the highest concentration of 1.0 g kg−1Fusarium ZH-H2 did not further improve degradation. Degradation of 4-ring PAHs showed no significant difference among different ZH-H2 incubations with bromegrass treatments. We found that the degradation rates of 4-, 5- and 6-ring PAHs in all treatments are significantly correlated in a positive, linear manner with activity of lignin peroxidase (LiP) (r=0.8065, 0.9350 and 0.9165, respectively), while degradation of 5- and 6-ring PAHs is correlated to polyphenoloxidase (PPO) activity (r=0.7577 and 07806). Our findings suggest that the combination of Fusarium sp. ZH-H2 and bromegrass offers a suitable alternative for phytoremediation of aged PAH-contaminated soil in coal mining areas, with a recommended inoculation size of 0.5 g Fusarium sp. ZH-H2 per kg soil.

Starch-enhanced degradation of HMW PAHs by Fusarium sp. in an aged polluted soil from a coal mining area

The present study used strain ZH-H2 (Fusarium sp.) isolated by our group as the PAH-degrading strain and 5-6-rings PAHs as degradation objects. The soil incubation experiment was carried out to investigate the starch-enhanced degradation effects of HMW PAHs by Fusarium sp. in an Aged Polluted Soil from a Coal Mining Area. The results showed that the removal rates of BaP, InP and BghiP increased with increasing inoculation rate of ZH-H2 in the unsterile aged polluted soil of coal mining area, with the exception of BbF degradation which increased in the H2 treatment and then decreased. Different addition dosage of starch apparently resulted in degradation of 4 PAHs in soil, with removal rates of 14.47% for BaP, 23.83% for DbA, 30.77% for BghiP and 31.00% for InP obtained with treatment D2, respectively higher than in treatment D1. So starch addition apparently enhanced the degradation of the 4 PAHs, especially InP and BghiP, by native microbes in the aged HMW PAH-polluted soil. By adding starch to these aged polluted soils with inoculated strain ZH-H2, HMW-PAHs degradation was further improved and addition of 0.5 g kg−1 starch to soils with 1.0 g kg−1 Fusarium ZH-H2 (D2 + H2) performed best to the 4 HMW-PAHs in all of these combination treatments by a factor of up to 3.09, depending on the PAH. We found that the highest polyphenol oxidase activities under D2 + H2 treatments are consistent with the results of removal rates of 4 PAHs. Our findings suggest that the combination of Fusarium sp. ZH-H2 and starch offers a suitable alternative for bioremediation of aged PAH-contaminated soil in coal mining areas, with a recommended inoculation size of 0.5 g Fusarium sp. ZH-H2 and addition of 0.5 g kg−1 starch per kg soil.

The role of lipopeptide biosurfactant on microbial remediation of aged polycyclic aromatic hydrocarbons (PAHs)-contaminated soil

Polycyclic aromatic hydrocarbons (PAHs) have been recognised as priority pollutants due to their carcinogenic, mutagenic and teratogenic properties. Bioremediation of PAHs is limited by low bioavailability of soil-bound PAHs due to their low aqueous solubility, high hydrophobicity and strong sorption to soil. The purpose of this study was to investigate the biodegradation enhancement potential of a heavy molecular mass lipopeptidal biosurfactant produced by Bacillus cereus SPL-4 on an aged wood treatment plant soil heavily contaminated with PAHs. In the laboratory scale batch treatments conducted (concentration of 6745.5mgkg−1, sum of 13PAHs) the high molecular weight PAHs removal rate was significantly enhanced in the biosurfactant supplemented microcosms than in the surfactant free controls. In the microcosms supplemented with 0.2 and 0.6% (w/w) lipopeptide, 51.2% of 4-ring and 55% of 5- and 6-ring PAHs; 64.1% of 4-ring and 79% of 5- and 6-ring PAHs were removed respectively, compared to 29% of 4-ring and 25.5% of 5- and 6-ring PAHs removal in the surfactant free control after 64daysof incubation. However, there was no statistically significant change in the degradation rates of low molecular weight PAHs in surfactant amended and surfactant free controls. Fitting the experimental data to two compartment model showed that the biosurfactant addition at 0.2 and 0.6% (w/w) increased the fast degradation fraction to 34.2 and 63% respectively, compared to 6% at the surfactant free controls. The degradation of 5 and 6 ring PAHs was significantly enhanced (p<0.05) in the higher surfactant dosage compared to the lower dosage. The results suggest that biosurfactant assisted degradation by microbial consortium may be a promising practical bioremediation strategy for aged PAH-contaminated soils.

Comparison of rapid solvent extraction systems for the GC–MS/MS characterization of polycyclic aromatic hydrocarbons in aged, contaminated soil

Polycyclic aromatic hydrocarbons (PAHs) are a major class of organic hydrocarbons with high molecular weight that originate from both natural and anthropogenic sources. Sixteen PAHs are included in the U.S Environmental Protection agency list of priority pollutants due to their mutagenic, carcinogenic, toxic and teratogenic properties. In this study, the development and optimization of a simplified and rapid solvent extraction for the characterisation of 16 USEPA priority poly aromatic hydrocarbons (PAHs) in aged contaminated soils was established with subsequent analysis by GC–MS/MS.•Five different extraction solvent systems: dichloromethane: acetone, chloroform: methanol, dichloromethane, acetone: hexane and hexane were assessed in terms of their ability to extract PAHs from aged PAH-contaminated soils.•Highest PAH concentrations were extracted using acetone: hexane and chloroform: methanol. Given the greater toxicity associated with chloroform: methanol, acetone: hexane appears the best choice of solvent extraction system.•This protocol enables efficient extraction of PAHs from aged weathered soils.

Impact of clay mineral, wood sawdust or root organic matter on the bacterial and fungal community structures in two aged PAH-contaminated soils.

The high organic pollutant concentration of aged polycyclic aromatic hydrocarbon (PAH)-contaminated wasteland soils is highly recalcitrant to biodegradation due to its very low bioavailability. In such soils, the microbial community is well adapted to the pollution, but the microbial activity is limited by nutrient availability. Management strategies could be applied to modify the soil microbial functioning as well as the PAH contamination through various amendment types. The impact of amendment with clay minerals (montmorillonite), wood sawdust and organic matter plant roots on microbial community structure was investigated on two aged PAH-contaminated soils both in laboratory and 1-year on-site pot experiments. Total PAH content (sum of 16 PAHs of the US-EPA list) and polar polycyclic aromatic compounds (pPAC) were monitored as well as the available PAH fraction using the Tenax method. The bacterial and fungal community structures were monitored using fingerprinting thermal gradient gel electrophoresis (TTGE) method. The abundance of bacteria (16S rRNA genes), fungi (18S rRNA genes) and PAH degraders (PAH-ring hydroxylating dioxygenase and catechol dioxygenase genes) was followed through qPCR assays. Although the treatments did not modify the total and available PAH content, the microbial community density, structure and the PAH degradation potential changed when fresh organic matter was provided as sawdust and under rhizosphere influence, while the clay mineral only increased the percentage of catechol-1,2-dioxygenase genes. The abundance of bacteria and fungi and the percentage of fungi relative to bacteria were enhanced in soil samples supplemented with wood sawdust and in the plant rhizospheric soils. Two distinct fungal populations developed in the two soils supplemented with sawdust, i.e. fungi related to Chaetomium and Neurospora genera and Brachyconidiellopsis and Pseudallescheria genera, in H and NM soils respectively. Wood sawdust amendment favoured the development of PAH-degrading bacteria holding Gram-negative PAH-ring hydroxylating dioxygenase, catechol-1,2-dioxygenase and catechol-2,3-dioxygenase genes. Regarding the total community structure, bacteria closely related to Thiobacillus (β-Proteobacteria) and Steroidobacter (γ-Proteobacteria) genera were favoured by wood sawdust amendment. In both soils, plant rhizospheres induced the development of fungi belonging to Ascomycota and related to Alternaria and Fusarium genera. Bacteria closely related to Luteolibacter (Verrucomicrobia) and Microbacterium (Actinobacteria) were favoured in alfalfa and ryegrass rhizosphere.

Biodegradation of Polycyclic Aromatic Hydrocarbons (PAHs) by Trichoderma reesei FS10-C and Effect of Bioaugmentation on an Aged PAH-Contaminated Soil

ABSTRACT The co-metabolism of benzo[a]pyrene (B[a]P) and the capacity of the fungus Trichoderma reesei FS10-C to bioremediate an aged polycyclic aromatic hydrocarbon (PAH)-contaminated soil were investigated. The fungal isolate removed about 54% of B[a]P (20 mg L−1) after 12 days of incubation with glucose (10 g L−1) supplementation as a co-metabolic substrate. Bioaugmented microcosms showed a 25% decrease in total PAH concentrations in soil after 28 days, and the degradation percentages of 3-, 4-, and 5(+6)-ring PAHs were 36%, 35%, and 25%, respectively. In addition, bioaugmented microcosms exhibited higher dehydrogenase (DHA) and fluorescein diacetate hydrolysis (FDAH) activities and increased average well-color development (AWCD), Shannon-Weaver index (H), and Simpson index (D) significantly. Principal component analysis (PCA) also distinguished clear differentiation between treatments, indicating that bioaugmentation restored the microbiological function of the PAH-contaminated soil. The results suggest that bioaugmentation by T. reesei FS10-C might be a promising bioremediation strategy for aged PAH-contaminated soils.

Enzyme Responses to Phytoremediation of PAH-Contaminated Soil Using Echinacea purpurea (L.)

Pot-culture experiments were performed to study the relationship between enzyme activity and polycyclic aromatic hydrocarbon (PAH) degradation by sowing Echinacea purpurea (L.) in aged PAH-contaminated soils. E. purpurea (L.) was cultivated in PAH-contaminated soils for 60, 120, and 150 days. The data showed that the rate of removal of fluoranthene, pyrene, benzo(a)anthracene, chrysene, benzo(b)fluoranthene, benzo(k)fluoranthene, benz(a)pyrene, and dibenzo(a,h)anthracene was 92.92 % at 150 days. Furthermore, there was a better recovery of these eight PAHs from the PAH-contaminated soils in comparison with the unplanted soils at 120 days, and the recovery from the PAH-contaminated soils was higher than that from the control soils. Significant reductions were detected in the levels of these eight PAHs, corresponding to 96.21, 94.53, 97.35, 90.70, 99.64, 97.34, 86.67, and 89.90 %, respectively. The activities of dehydrogenase, polyphenol oxidase, and urease increased as a result of PAH treatment, and these effects were suppressed over time. The activities of peroxidase and alkaline phosphatase were reduced, which suggests that these enzymes are sensitive to PAHs. The data in this study revealed that enzyme activities were strongly correlated to the higher degradation performance of the E. purpurea (L.) plant on PAH-contaminated soil. More researches are required to elucidate the mechanisms underlying PAH detoxification and degradation in contaminated soil.

Methyl-β-cyclodextrin enhanced biodegradation of polycyclic aromatic hydrocarbons and associated microbial activity in 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-β-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 (p < 0.05) in the average well color development (AWCD) obtained by the BIOLOG Eco plate assay, Shannon-Weaver index (H) and Simpson index (γ) 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.

Influence of Rhizobium meliloti on phytoremediation of polycyclic aromatic hydrocarbons by alfalfa in an aged contaminated soil

Microbe-assisted phytoremediation is emerging as one of the most effective means by which plants and their associated rhizosphere microbes degrade organic contaminants in soils. A pot study was conducted to examine the effects of inoculation with Rhizobium meliloti on phytoremediation by alfalfa grown for 90 days in an agricultural soil contaminated with weathered polycyclic aromatic hydrocarbons (PAHs). Planting with uninoculated alfalfa (P) and alfalfa inoculated with R. meliloti (PR) significantly lowered the initial soil PAH concentrations by 37.2 and 51.4% respectively compared with unplanted control soil. Inoculation with R. meliloti significantly increased the counts of culturable PAH-degrading bacteria, soil microbial activity and the carbon utilization ability of the soil microbial community. The results suggest that the symbiotic association between alfalfa and Rhizobium can stimulate the rhizosphere microflora to degrade PAHs and its application may be a promising bioremediation strategy for aged PAH-contaminated soils.

Effect of bioaugmentation by Paracoccus sp. strain HPD-2 on the soil microbial community and removal of polycyclic aromatic hydrocarbons from an aged contaminated soil

A microcosm study was conducted to test the bioremediation potential of Paracoccus sp. strain HPD-2 on an aged PAH-contaminated soil. Bioaugmented microcosms showed a 23.2% decrease in soil total PAH concentrations after 28 days, with a decline in average concentration from 9942 to 7638 μg kg −1 dry soil. The percentage degradation of 3-, 4- and 5(+6)-ring PAHs was 35.1%, 20.7% and 24.3%, respectively. Higher counts of culturable PAH-degrading bacteria, microbial biomass and enzyme activities were observed in bioaugmented soil. The bioaugmented microcosms showed significant increases ( p < 0.05) in the average well-color development (AWCD) obtained by the BIOLOG ecoplate assay and Shannon–Weaver index ( H) compared to the controls. Principal component analysis of BIOLOG data clearly differentiated between the bioaugmented and control microcosms, implying that bioaugmentation restored the microbiological functioning of the PAH-contaminated soil. The results suggest that bioaugmentation by Paracoccus sp. strain HPD-2 may be a promising bioremediation strategy for aged PAH-contaminated soils.

Effects of soil amendment with different carbon sources and other factors on the bioremediation of an aged PAH-contaminated soil

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.

Influence of Vegetation on the In Situ Bacterial Community and Polycyclic Aromatic Hydrocarbon (PAH) Degraders in Aged PAH-Contaminated or Thermal-Desorption-Treated Soil

The polycyclic aromatic hydrocarbon (PAH) contamination, bacterial community, and PAH-degrading bacteria were monitored in aged PAH-contaminated soil (Neuves-Maisons [NM] soil; with a mean of 1,915 mg of 16 PAHs.kg(-1) of soil dry weight) and in the same soil previously treated by thermal desorption (TD soil; with a mean of 106 mg of 16 PAHs.kg(-1) of soil dry weight). This study was conducted in situ for 2 years using experimental plots of the two soils. NM soil was colonized by spontaneous vegetation (NM-SV), planted with Medicago sativa (NM-Ms), or left as bare soil (NM-BS), and the TD soil was planted with Medicago sativa (TD-Ms). The bacterial community density, structure, and diversity were estimated by real-time PCR quantification of the 16S rRNA gene copy number, temporal thermal gradient gel electrophoresis fingerprinting, and band sequencing, respectively. The density of the bacterial community increased the first year during stabilization of the system and stayed constant in the NM soil, while it continued to increase in the TD soil during the second year. The bacterial community structure diverged among all the plot types after 2 years on site. In the NM-BS plots, the bacterial community was represented mainly by Betaproteobacteria and Gammaproteobacteria. The presence of vegetation (NM-SV and NM-Ms) in the NM soil favored the development of a wider range of bacterial phyla (Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, Verrucomicrobia, Actinobacteria, Firmicutes, and Chloroflexi) that, for the most part, were not closely related to known bacterial representatives. Moreover, under the influence of the same plant, the bacterial community that developed in the TD-Ms was represented by different bacterial species (Alphaproteobacteria, Betaproteobacteria, Gammaproteobacteria, and Actinobacteria) than that in the NM-Ms. During the 2 years of monitoring, the PAH concentration did not evolve significantly. The abundance of gram-negative (GN) and gram-positive (GP) PAH-degrading bacteria was estimated by real-time PCR quantification of specific functional genes encoding the alpha subunit of PAH-ring hydroxylating dioxygenase (PAH-RHD(alpha)). The percentage of the PAH-RHD(alpha) GN bacterial genes relative to 16S rRNA gene density decreased with time in all the plots. The GP PAH-RHD(alpha) bacterial gene proportion decreased in the NM-BS plots but stayed constant or increased under vegetation influence (NM-SV, NM-Ms, and TD-Ms).

Effects of Agronomic Practices on Phytoremediation of an Aged PAH‐Contaminated Soil

Phytoremediation offers an ecologically and economically attractive remediation technique for soils contaminated with polycyclic aromatic hydrocarbons (PAHs). In addition to the choice of plant species, agronomic practices may affect the efficiency of PAH phytoremediation. Inorganic nutrient amendments may stimulate plant and microbial growth, and clipping aboveground biomass might stimulate root turnover, which has been associated with increases in soil microbial populations. To assess the influence of fertilization and clipping on PAH dissipation in a nutrient-poor, aged PAH-contaminated soil, a 14-mo phytoremediation study was conducted using perennial ryegrass (Lolium perenne) as a model species. Six soil treatments were performed in replicate: unplanted; unplanted and fertilized; planted; planted and fertilized; planted and clipped; and planted, clipped, and fertilized. Plant growth, soil PAH concentrations, and the concentrations of total and PAH-degrading microorganisms were measured after 7 and 14 mo. Overall, planting (with nearly 80% reduction in total PAHs) and planting + clipping (76% reduction in total PAHs) were the most effective treatments for increased PAH dissipation after 14 mo. Fertilization greatly stimulated plant and total microbial growth, but negatively affected PAH dissipation (29% reduction in total PAHs). Furthermore, unplanted and fertilized soils revealed a similar negative impact (25% reduction) on PAH dissipation after 14 mo. Clipping did not directly affect PAH dissipation, but when combined with fertilization (61% reduction in total PAHs), appeared to mitigate the negative impact of fertilization on PAH dissipation. Therefore, fertilization and clipping may be included in phytoremediation design strategies, as their combined effect stimulates plant growth while not affecting PAH dissipation.

Bioaugmentation and biostimulation effects on PAH dissipation and soil ecotoxicity under controlled conditions

The degradation of spiked anthracene (ANT), pyrene (PYR) and benzo[ a]pyrene (B[ a]P) in soil (3000 mg ∑ 3 PAHs kg −1 dry soil) was studied in aerobically incubated microcosms for 120 d. The applied treatments aimed at enhancing PAH removal from the heavily contaminated soils are: (i) bioaugmentation by adding aged PAH-contaminated soil (ACS) containing activated indigenous degraders; and (ii) combined bioaugmentation/biostimulation by incorporating sewage sludge compost (SSC) and decaying rice straw (DRS). The adopted treatments produced higher PAH dissipation rates than those observed in unamended PAH-spiked soils, especially for ANT and PYR in the presence of DRS or ACS (>96%). However, B[ a]P was the most recalcitrant hydrocarbon to biodegradation. Extracellular enzyme investigation revealed the existence of ligninolytic activities in all soil treatments, including control but no relationship could be found with PAH dissipation. The ecotoxicological assessment indicated that regardless of applied treatment, PAH-spiked soils were chronically lethal to ostracod Heterocypris incongruens and confirmed the sensitivity of the microcrustacean to the concomitant presence of these three hydrocarbons. Lettuce root elongation inhibition was correlated with PAH level but the presence of SSC conferred a strong phytotoxic capacity to PAH-spiked soils. DRS amendment may constitute a cost-effective alternative for hydrocarbon bioremediation as it has impacted positively on soil microbial activity and enhanced PAH removal with no apparent changes in soil physico-chemical properties.

Effect of inoculum pretreatment on survival, activity and catabolic gene expression of Sphingobium yanoikuyae B1 in an aged polycyclic aromatic hydrocarbon-contaminated soil

The survival and effectiveness of a bioaugmentation strain in its target environment depend not only on physicochemical parameters in the soil but also on the physiological state of the inoculated organism. This study examined the effect of variations in inoculum pretreatment on the survival, metabolic activity (measured as rRNA content) and polycyclic aromatic hydrocarbon (PAH)-catabolic gene expression of Sphingobium yanoikuyae B1 in an aged PAH-contaminated soil. RNA denaturing gradient gel electrophoresis analysis showed stable colonization of PAH-contaminated soil by S. yanoikuyae B1 after four pretreatments (growth in complex or minimal medium, starvation, or acclimation to phenanthrene). By contrast, extractable CFUs decreased with time for all four treatments, and significantly faster for Luria Bertani-grown inocula, suggesting that these cells adhered strongly to soil particles while remaining metabolically active. Pretreatment of the inoculum had a dramatic effect on the expression of genes specific to the PAH-degradation pathway. The highest levels of bphC and xylE expression were seen for inocula that had been precultivated on complex medium, and degradation of PAHs was significantly enhanced in soils treated with these inocula. The results suggest that using complex media instead of minimal media for cultivating bioaugmentation inocula may improve the subsequent efficiency of contaminant biodegradation in the soil.

Effect of Sphingobium yanoikuyae B1 inoculation on bacterial community dynamics and polycyclic aromatic hydrocarbon degradation in aged and freshly PAH-contaminated soils

Sphingobium yanoikuyae B1 is able to degrade a range of polycyclic aromatic hydrocarbons (PAHs) and as a sphingomonad belongs to one of the dominant genera found in PAH-contaminated soils. We examined the ecological effect that soil inoculation with S. yanoikuyae B1 has on the native bacterial community in three different soils: aged PAH-contaminated soil from an industrial site, compost freshly contaminated with PAHs and un-contaminated compost. Survival of S. yanoikuyae B1 was dependent on the presence of PAHs, and the strain was unable to colonize un-contaminated compost. Inoculation with S. yanoikuyae B1 did not cause extensive changes in the native bacterial community of either soil, as assessed by denaturing gel electrophoresis, but its presence led to an increase in the population level of two other species in the aged contaminated soil community and appeared to have an antagonistic affect on several members of the contaminated compost community, indicating niche competition.