Bioremediation of petroleum hydrocarbons polluted soil by spent mushroom substrates: Microbiological structure and functionality.

Spent mushroom substrate (SMS) holds valuable microbiota that can be useful in remediating polluted soils with hydrocarbons. However, the microorganisms behind the bioremediation process remain uncertain. In this work, a bioremediation assay of total petroleum hydrocarbons (TPHs) polluted soil by SMS application was performed to elucidate the microorganisms and consortia involved in biodegradation by a metabarcoding analysis. Untreated polluted soil was compared to seven bioremediation treatments by adding SMS of Agaricus bisporus, Pleurotus eryngii, Pleurotus ostreatus, and combinations. Soil microbial activity, TPH biodegradation, taxonomic classification, and predictive functional analysis were evaluated in the microbiopiles at 60 days. Different metagenomics approaches were performed to understand the impact of each SMS on native soil microbiota and TPHs biodegradation. All SMSs enhanced the degradation of aliphatic and aromatic hydrocarbons, being A. bisporus the most effective, promoting an efficient consortium constituted by the bacterial families Alcanivoraceae, Alcaligenaceae, and Dietziaceae along with the fungal genera Scedosporium and Aspergillus. The predictive 16 S rRNA gene study partially explained the decontamination efficacy by observing changes in the taxonomic structure of bacteria and fungi, and changes in the potential profiles of estimated degradative genes across the different treatments. This work provides new insights into TPHs bioremediation.

Main Factors Determining the Scale-Up Effectiveness of Mycoremediation for the Decontamination of Aliphatic Hydrocarbons in Soil.

Soil contamination constitutes a significant threat to the health of soil ecosystems in terms of complexity, toxicity, and recalcitrance. Among all contaminants, aliphatic petroleum hydrocarbons (APH) are of particular concern due to their abundance and persistence in the environment and the need of remediation technologies to ensure their removal in an environmentally, socially, and economically sustainable way. Soil remediation technologies presently available on the market to tackle soil contamination by petroleum hydrocarbons (PH) include landfilling, physical treatments (e.g., thermal desorption), chemical treatments (e.g., oxidation), and conventional bioremediation. The first two solutions are costly and energy-intensive approaches. Conversely, bioremediation of on-site excavated soil arranged in biopiles is a more sustainable procedure. Biopiles are engineered heaps able to stimulate microbial activity and enhance biodegradation, thus ensuring the removal of organic pollutants. This soil remediation technology is currently the most environmentally friendly solution available on the market, as it is less energy-intensive and has no detrimental impact on biological soil functions. However, its major limitation is its low removal efficiency, especially for long-chain hydrocarbons (LCH), compared to thermal desorption. Nevertheless, the use of fungi for remediation of environmental contaminants retains the benefits of bioremediation treatments, including low economic, social, and environmental costs, while attaining removal efficiencies similar to thermal desorption. Mycoremediation is a widely studied technology at lab scale, but there are few experiences at pilot scale. Several factors may reduce the overall efficiency of on-site mycoremediation biopiles (mycopiles), and the efficiency detected in the bench scale. These factors include the bioavailability of hydrocarbons, the selection of fungal species and bulking agents and their application rate, the interaction between the inoculated fungi and the indigenous microbiota, soil properties and nutrients, and other environmental factors (e.g., humidity, oxygen, and temperature). The identification of these factors at an early stage of biotreatability experiments would allow the application of this on-site technology to be refined and fine-tuned. This review brings together all mycoremediation work applied to aliphatic petroleum hydrocarbons (APH) and identifies the key factors in making mycoremediation effective. It also includes technological advances that reduce the effect of these factors, such as the structure of mycopiles, the application of surfactants, and the control of environmental factors.

The role of biochar and green compost amendments in the adsorption, leaching, and degradation of sulfamethoxazole in basic soil.

The fate of the antibiotic sulfamethoxazole in amended soils remains unclear, moreover in basic soils. This work aimed to assess the adsorption, leaching, and biodegradation of sulfamethoxazole in unamended and biochar from holm oak pruning (BC)- and green compost from urban pruning (CG)-amended basic soil. Adsorption properties of the organic amendments and soil were determined by adsorption isotherms of sulfamethoxazole. The leachability of this antibiotic from unamended (Soil) and BC- (Soil + BC) and GC- (Soil + GC) amended soil was determined by leaching columns using water as solvent up to 250 mL. Finally, Soil, Soil + BC, and Soil + GC were spiked with sulfamethoxazole and incubated for 42 days. The degradation rate and microbial activity were periodically monitored. Adsorption isotherms showed poor adsorption of sulfamethoxazole in unamended basic soil. BC and CG showed good adsorption capacity. Soil + BC and Soil + GC increased the sulfamethoxazole adsorption capacity of the soil. The low sulfamethoxazole adsorption of Soil produced quick and intense sulfamethoxazole leaching. Soil + BC reduced the sulfamethoxazole leaching, unlike to Soil + GC which enhanced it concerning Soil. The pH of adsorption isotherms and leachates indicate that the anion of sulfamethoxazole was the major specie in unamended and amended soil. CG enhanced the microbial activity of the soil and promoted the degradability of sulfamethoxazole. In contrast, the high adsorption and low biostimulation effect of BC in soil reduced the degradation of sulfamethoxazole. The half-life of sulfamethoxazole was 2.6, 6.9, and 11.9 days for Soil + GC, Soil, and Soil + BC, respectively. This work shows the benefits and risks of two organic amendments, BC and GC, for the environmental fate of sulfamethoxazole. The different nature of the organic carbon of the amendments was responsible for the different effects on the soil.

Synergistic effects of biochar and biostimulants on nutrient and toxic element uptake by pepper in contaminated soils.

BACKGROUND: Nowadays a significant amount of land contaminated with toxic elements is being used for agriculture, posing a serious risk of crop contamination and toxicity. Several methodologies are being used to remediate soil contamination, including the use of amendments such as biochar. This work evaluated the effects of biochar combined with different fertirrigations (water, a conventional fertilizer solution, or a fertilizer solution with a commercial biostimulant derived from leonardite) on the availability of toxic elements and nutrients for pepper cultivated in a soil contaminated with As, Cd, Pb, and Zn. RESULTS: Irrigation with fertilizer solutions improved plant growth regardless of the biochar amendment. Biochar decreased the bioavailability of Cu and Pb in soil and the Cu content in pepper leaves. Combined with fertilization, biochar also decreased plant As and Pb content. Biochar combined with biostimulant decreased the bioavailable content of Cd in soil and its uptake by pepper plants. CONCLUSION: The use of biochar and biostimulant presented advantages for plant production in a non-suitable scenario of nutrient scarcity and contamination. © 2021 The Authors. Journal of The Science of Food and Agriculture published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

An assessment of Pleurotus ostreatus to remove sulfonamides, and its role as a biofilter based on its own spent mushroom substrate.

A double strategy based on the removal of sulfonamide antibiotics by Pleurotus ostreatus and adsorption on spent mushroom substrate was assessed to reclaim contaminated wastewater. P. ostreatus was firstly tested in a liquid medium fortified with five sulfonamides: sulfamethoxazole, sulfadiazine, sulfathiazole, sulfapyridine and sulfamethazine, to evaluate its capacity to remove them and to test for any adverse effects on fungal growth and for any reduction in residual antibiotic activity. P. ostreatus was effective in removing sulfonamides up to 83 to 91% of the applied doses over 14 days. The antibiotic activity of the sulfonamide residues was reduced by 50%. Sulfamethoxazole transformation products by laccase were identified, and the degradation pathway was proposed. In addition, P. ostreatus growth on a semi-solid medium of spent mushroom substrate and malt extract agar was used to develop a biofilter for the removal of sulfonamides from real wastewater. The biofilter was able to remove more than 90% of the sulfonamide concentrations over 24 h by combining adsorption and biodegradation mechanisms.

Holistic assessment of the microbiome dynamics in the substrates used for commercial champignon (Agaricus bisporus) cultivation.

Microorganisms strongly influence and are required to generate the selective substrate that provides nutrients and support for fungal growth, and ultimately to induce mushroom fructification under controlled environmental conditions. In this work, the fungal and bacterial microbiota living in the different substrates employed in a commercial crop (compost phase I, II and III, flush 1 and 2, and casing material on day 1, 6 and 8 after compost casing and during flush 1 and 2) have been characterized along the different stages of cultivation by metataxonomic analysis (16S rRNA and ITS2), analysis of phospholipid fatty acid content (PLFAs) and RT-qPCR. Additionally, laccase activity and the content of lignin and complex carbohydrates in compost and casing have been quantified. The bacterial diversity in compost and casing increased throughout the crop cycle boosted by the connection of both substrates. As reflected by the PLFAs, the total living bacterial biomass appears to be negatively correlated with the mycelium of the crop. Agaricus bisporus was the dominant fungal species in colonized substrates, displacing the pre-eminent Ascomycota, accompanied by a sustained increase in laccase activity, which is considered to be a major product of protein synthesis during the mycelial growth of champignon. From phase II onwards, the metabolic machinery of the fungal crop degrades lignin and carbohydrates in compost, while these components are hardly degraded in casing, which reflects the minor role of the casing for nourishing the crop. The techniques employed in this study provide a holistic and detailed characterization of the changing microbial composition in commercial champignon substrates. The knowledge generated will contribute to improve compost formulations (selection of base materials) and accelerate compost production, for instance, through biotechnological interventions in the form of tailored biostimulants and to design environmentally sustainable bio-based casing materials.

Mobility monitoring of two herbicides in amended soils: A field study for modeling applications.

This paper reports the mobility and total balance of chlorotoluron (CTL), flufenacet (FNC) and bromide ion (Br-) throughout a sandy soil profile after the application of spent mushroom substrate (SMS) and green compost (GC). Obtaining mobility dataset is crucial to simulate the herbicides' fate under amended soil scenarios by application pesticide leaching models with regulatory application (FOCUS models). The application of organic residues is nowadays increased to improve the crop yields and there is a gap in the simulations of this kind of amended scenarios. A two-year field experiment involving unamended soil (S) and SMS- or GC-amended soil plots was conducted. CTL, FNC, and Br- were annually applied and their residual concentrations were determined in soil profiles (0-100 cm) regularly sampled. In all the treatments the order of mobility is followed as FNC < CTL < Br-. SMS and GC increased herbicide retention in the top 10 cm by the higher organic carbon (OC) content than the unamended soil, and their ability to increase the soil's water-holding capacity and to decrease water percolation. Simultaneously dissolved organic carbon (DOC) content facilitated herbicide transport being it favoured by the initial soil moisture content and the rainfall shortly after the chemicals' initial application. Over the first year, residual amounts (<2.6%) of Br-, CTL and FNC were leached down to 90-100 cm depth in the three treatments. However, over the second year low CTL and FNC amounts (<1.0%) reached the bottom layer only in S + SMS although high Br- concentrations did so in the three treatments (<20%). According to the total balance of Br-, CTL, and FNC in the soil profiles other processes (degradation, mineralisation, bound residues formation, and/or crop uptake) different from leaching below 1 m depth might play a key role in their dissipation especially in the amended soil profiles. SMS and GC are likely to be used as organic amendments to preserve the soil and water quality but in the case of SMS, its higher DOC content could imply a higher potential risk for groundwater contamination than GC.

Organic carbon nature determines the capacity of organic amendments to adsorb pesticides in soil.

The spread of organic pollutants from soil to other environments is one important source of environmental pollution. The addition of organic amendments to soil is an interesting strategy to control pollutants leaching. However, the contribution of different carbon types of organic amendments to organic pollutants adsorption is not clear. Hence, the objective of this work was to determine the role of carbon types of organic amendments into the adsorption of four herbicides. To this extent, organic amendments were characterized by elemental analysis and 13C-NMR and adsorption-desorption isotherms of herbicides by the organic amendments and two soils amended with them were obtained. Adsorption coefficients were correlated with the organic carbon content of the organic amendments and the adsorption process was enhanced by the hydrophobicity of herbicides and the aliphatic and aromatic carbon of amendments. Organic amendments increased the adsorption of herbicides by soils but it is not possible to extrapolate results from one soil to another because organo-mineral interactions between soils and organic amendments can modify this process. Desorption isotherms of herbicides from organic amendments and/or amended soils presented hysteresis indicating the irreversible adsorption of herbicides. Desorption results indicated, the abundance of O-alkyl and N-alkyl groups in organic amendments enhanced the hysteresis in amended soils.

Impact of a tire fire accident on soil pollution and the use of clay minerals as natural geo-indicators.

Following the occurrence of a fire at a tire landfill in the surrounding area of Madrid City (Spain), polycyclic aromatic hydrocarbons (PAHs) and trace elements present in soils were analyzed to assess the impact of the fire. The capacity of the soils' clay mineral fraction to reflect this air pollution incident was studied. Fourteen soil samples were collected at different distances under the smoke plume, and they were subjected to high-performance liquid chromatography-photodiode array detection, inductively coupled plasma mass spectrometry and X-ray diffraction analyses. Clay minerals content showed a strong correlation with the pollutants potentially released in the tire fire, acenaphthene, pyrene, benzo(a)pyrene and benzo(a)fluoranthene. Trace metals Zn and Se were related to the proximity of the tire fire without any relationship with clay minerals content. This work suggests the use of natural clay minerals as potential PAHs geo-indicators in response to air pollution, complementary to current air and biological analyses.

Assessment of 14C-prosulfocarb dissipation mechanism in soil after amendment and its impact on the microbial community.

Adding organic amendments to soil could modify the bioavailability of herbicides and lead to changes in the microbial community's activity and structure. The objective here was to study the dissipation and total mass balance of 14C-labeled prosulfocarb applied at two rates (4 and 10 mg kg-1) in unamended and green compost (GC)-amended soil. Soil dehydrogenase activity (DHA) and phospholipid fatty acid (PLFA) profile analysis were determined to evaluate the effect of herbicide residues on microbial community's activity and structure over the dissipation period. The dissipation rate of prosulfocarb decreased after soil amendment due to higher herbicide adsorption by the amended soil. The 50% dissipation time (DT50) increased 1.7 times in the unamended soil when the concentration of prosulfocarb increased 2.5 times. The mass balance results indicate that the sum of water and organic extractable fractions represented the highest amounts up to the dissipation of 50% 14C-prosulfocarb. The 14C-herbicide was then mainly mineralized (up to 11%-31%) or formed non-extractable residues (up to 35%-44%). The amount of 14C-prosulfocarb residues extracted with methanol was slightly higher in amended soils than in unamended ones. 14C-prosulfocarb mineralization was higher in unamended soils than in amended ones. The formation of non-extractable residues was continuous, and increased over time. Soil DHA decreased in the unamended soil and was maintained in the GC-amended soil at the end of the assay. The microbial structure was barely disturbed over the prosulfocarb degradation process, although it was clearly influenced by the application of GC. The results obtained reveal the influence organic amendment has on herbicide bioavailability to decrease its biodegradation and buffer its impact on the soil microbial structure.

Influence of different agricultural management practices on soil microbial community over dissipation time of two herbicides.

Soil microbiology could be affected by the presence of pesticide residues during intensive farming, potentially threatening the soil environment. The aim here was to assess the dissipation of the herbicides triasulfuron and prosulfocarb, applied as a combined commercial formulation, and the changes in soil microbial communities (through the profile of phospholipid fatty acids (PLFAs) extracted from the soil) during the dissipation time of the herbicides under field conditions. The dissipation of herbicides and the soil microbial structure were assessed under different agricultural practices, such as the repeated application of herbicides (twice), in unamended and amended soils with two organic amendments derived from green compost (GC1 and GC2) and with non-irrigation and irrigation regimes. The results obtained indicate slower dissipation for triasulfuron than for prosulfocarb. The 50% dissipation time (DT50) decreased under all conditions for the second application of triasulfuron, although not for prosulfocarb. The DT50 values for both herbicides increased in the GC2 amended soil with the highest organic carbon (OC) content. The DT50 values decreased for prosulfocarb with irrigation, but not for triasulfuron, despite its higher water solubility. The herbicides did not have any significant effects on the relative population of Gram-negative and Gram-positive bacteria during the assay, but the relative abundance of Actinobacteria increased in all the soils with herbicides. At the end of the assay (215 days), the negative effects of herbicides on fungi abundance were significant (p < 0.05) for all the treatments. These microbiological changes were detected in non-irrigated and irrigated soils, and were more noticeable after the second application of herbicides. Actinobacteria could be responsible for the modification of herbicide degradation rates, which tend to be faster after the second application. This study makes a useful contribution to the evaluation of the soil environment and microbiological risks due to the long-term repeated application of herbicides under different agricultural management practices.

Stevensite-based geofilter for the retention of tetracycline from water.

The antibiotic tetracycline, is considered a contaminant of emerging concern due to its presence in wastewater effluents, surface waters and groundwaters. Adsorption of tetracycline on soils and clays has been extensively studied to remove the contaminant from the water. A decreasing adsorption as the pH increases is normally reported in the pH range 3-9. However, adsorption isotherms performed on a commercial stevensite presented increasing adsorption with the increasing pH, in the pH range 2-8. This is very interesting since the pH in natural and wasterwaters are normally in the range 6-8. A laboratory design of a geofilter using a mixture of sand and stevensite was tested against an inflow solution of tetracycline 1 g/L, NaNO3 0.1 M and pH = 7 in an advective transport cell experiment. The number of tetracycline molecules exceed by >3 times the number exchangeable positions in the stevensite geofilter. Under these conditions, the TC adsorption on the geofilter reaches 590 mg/g, surpassing the retention capacity of most adsorbents found in literature. Besides, the tetracycline is completely desorbed by the inflow of a saline solution (Mg(NO3)2 0.5 M, at pH = 2) with capacity to replace the exchangeable positions, thus, recovering the geofilter and the tetracycline.

Soil element fractions affect phytotoxicity, microbial biomass and activity in volcanic areas.

Soil quality is strongly affected by microbial biomass that is involved in organic matter mineralization and the supply of nutrients to plants. The effects of trace elements on soil microbial biomass and activity are still controversial, and the contents of the elements in different forms, more than the total amounts, may affect soil microbial community. Volcanic soils are peculiar environments because of their chemical characteristics. Therefore, the aims of this research were to evaluate in volcanic soils: i) the elemental composition; ii) the elemental availability; and iii) the effects of elemental fractions on soil microbial biomass and activity. In order to reach the aims, the BCR sequential extraction method was applied in order to separate 22 elements in different soil fractions: acid soluble, reducible, oxidisable and residual. The studied biological parameters were: microbial and fungal biomasses, soil respiration, metabolic quotient, coefficient of endogenous mineralization, dehydrogenase and hydrolase activities, and phytotoxicity. Among the investigated elements, Al, B, Ba, Ca, Cr, Fe, K, Mg, Mn, Na, Ni, Ti, V and Zn were scarcely available; Cd appeared to be the most ready available element; Zn was mainly present in the acid soluble and in the residual fractions. Microbial biomass and activity appeared to be mainly affected by the reducible and oxidasable fractions of the investigated elements more than the acid soluble or residual ones. With the exception of La and V, the elemental content in the various fractions would seem to stimulate the microbial biomass and activity. Finally, the investigated volcanic soils showed phytotoxic properties.

Total and fraction content of elements in volcanic soil: Natural or anthropogenic derivation.

Soil element composition derives from parent material disaggregation during pedogenesis and weathering processes but also by anthropogenic inputs. Elements are present in soils in different chemical forms that affect their availability and mobility. The aim of the study was to evaluate the main derivation, natural or anthropogenic, of elements in the soils of the Vesuvius National Park (a natural environment strongly affected by human impacts). Besides, the effects of age of the lava from which soils derive, different vegetation covers, traffic fluxes along the two roads connecting the Vesuvius crater and altitudes of the sites on the pseudo-total element concentrations and on their contents in different fraction of soil were investigated. To reach the aims, BCR (Bureau Commun de Référence) sequential extraction was performed in order to determine the distribution of elements into: acid-soluble, reducible, oxidizable and residual fractions. The relationship between the main environmental media and distribution of elements was discussed using non-metric multidimensional scaling (NMDS). The findings showed that, with the exception of Cd, Cu, Pb and Zn that would seem to derive also from human activities, the other investigated elements (Al, As, B, Ba, Ca, Cd, Cr, Cu, Fe, K, La, Mg, Mn, Na, Ni, P, Pb, Si, Ti, V, W and Zn) mainly had a natural derivation. Among the investigated elements, only Cd could represent a potential high risk for the studied andosols. The highest element accumulations in the soils at low altitude could be attributable to an integrated effect of plant cover, vicinity of downtowns and traffic flux. The acid-soluble fraction of elements appeared more linked to lava age; the reducible and oxidizable ones to plant cover; the residual one to the chemical composition of the parent material that gave origin to the soils.

The use of spent mushroom compost to enhance the ability of Atriplex halimus to phytoremediate contaminated mine soils.

The mushroom cultivation industry produces a huge amount of spent mushroom compost (SMC), a wide world agricultural organic waste which causes serious environmental problems. However, this cheap organic waste could be useful in the remediation of contaminated soils. The aim of this work was to assess the potential of SMC in combination with the native shrub Atriplex halimus, to phytoremediate two mine soils contaminated with Cd, Pb and Cu. Firstly, to minimize metal availability in the soil, the optimal doses of SMC were determined. Secondly, a phytoremediation assay in greenhouse conditions was carried out to test the effects of A. halimus in combination with SMC at different doses. The results showed the ability of SMC to reduce soil acidity, the mobility of the metals and the enhancement of A. halimus growth. SMC promoted metal immobilization in the root of A. halimus and decreased the translocation from the roots to the shoots. The combination of SMC amendment and A. halimus produced phytostabilization of the metals in the mine soils assayed. In conclusion, SMC represents an adequate organic solid waste which in combination with A. halimus can reduce the adverse impact caused by the high mobility of metals in acid mine soils.

Bioremediation of multi-polluted soil by spent mushroom (Agaricus bisporus) substrate: Polycyclic aromatic hydrocarbons degradation and Pb availability.

This study investigates the effect of three spent Agaricus bisporus substrate (SAS) application methods on bioremediation of soil multi-polluted with Pb and PAH from close to a shooting range with respect natural attenuation (SM). The remediation treatments involve (i) use of sterilized SAS to biostimulate the inherent soil microbiota (SSAS) and two bioaugmentation possibilities (ii) its use without previous treatment to inoculate A. bisporus and inherent microbiota (SAS) or (iii) SAS sterilization and further A. bisporus re-inoculation (Abisp). The efficiency of each bioremediation microcosm was evaluated by: fungal activity, heterotrophic and PAH-degrading bacterial population, PAH removal, Pb mobility and soil eco-toxicity. Biostimulation of the native soil microbiology (SSAS) achieved similar levels of PAH biodegradation as SM and poor soil detoxification. Bioaugmented microcosms produced higher PAH removal and eco-toxicity reduction via different routes. SAS increased the PAH-degrading bacterial population, but lowered fungal activity. Abisp was a good inoculum carrier for A. bisporus exhibiting high levels of ligninolytic activity, the total and PAH-degrading bacteria population increased with incubation time. The three SAS applications produced slight Pb mobilization (<0.3%). SAS sterilization and further A. bisporus re-inoculation (Abisp) proved the best application method to remove PAH, mainly BaP, and detoxify the multi-polluted soil.

Implications of polluted soil biostimulation and bioaugmentation with spent mushroom substrate (Agaricus bisporus) on the microbial community and polycyclic aromatic hydrocarbons biodegradation.

Different applications of spent Agaricus bisporus substrate (SAS), a widespread agro-industrial waste, were investigated with respect to the remediation of a historically polluted soil with Polycyclic Aromatic Hydrocarbons (PAH). In one treatment, the waste was sterilized (SSAS) prior to its application in order to assess its ability to biostimulate, as an organic amendment, the resident soil microbiota and ensuing contaminant degradation. For the other treatments, two bioaugmentation approaches were investigated; the first involved the use of the waste itself and thus implied the application of A. bisporus and the inherent microbiota of the waste. In the second treatment, SAS was sterilized and inoculated again with the fungus to assess its ability to act as a fungal carrier. All these treatments were compared with natural attenuation in terms of their impact on soil heterotrophic and PAH-degrading bacteria, fungal growth, biodiversity of soil microbiota and ability to affect PAH bioavailability and ensuing degradation and detoxification. Results clearly showed that historically PAH contaminated soil was not amenable to natural attenuation. Conversely, the addition of sterilized spent A. bisporus substrate to the soil stimulated resident soil bacteria with ensuing high removals of 3-ring PAH. Both augmentation treatments were more effective in removing highly condensed PAH, some of which known to possess a significant carcinogenic activity. Regardless of the mode of application, the present results strongly support the adequacy of SAS for environmental remediation purposes and open the way to an attractive recycling option of this waste.

Combination of biochar amendment and mycoremediation for polycyclic aromatic hydrocarbons immobilization and biodegradation in creosote-contaminated soil.

Soils impregnated with creosote contain high concentrations of polycyclic aromatic hydrocarbons (PAH). To bioremediate these soils and avoid PAH spread, different bioremediation strategies were tested, based on natural attenuation, biochar application, wheat straw biostimulation, Pleurotus ostreatus mycoremediation, and the novel sequential application of biochar for 21 days and P. ostreatus 21 days more. Soil was sampled after 21 and 42 days after the remediation application. The efficiency and effectiveness of each remediation treatment were assessed according to PAH degradation and immobilization, fungal and bacterial development, soil eco-toxicity and legal considerations. Natural attenuation and biochar treatments did not achieve adequate PAH removal and soil eco-toxicity reduction. Biostimulation showed the highest bacterial development but low PAH degradation rate. Mycoremediation achieved the best PAH degradation rate and the lowest bioavailable fraction and soil eco-toxicity. This bioremediation strategy achieved PAH concentrations below Spanish legislation for contaminated soils (RD 9/2005). Sequential application of biochar and P. ostreatus was the second treatment most effective for PAH biodegradation and immobilization. However, the activity of P. ostreatus was increased by previous biochar application and PAH degradation efficiency was increased. Therefore, the combined strategy for PAH degradation have high potential to increase remediation efficiency.