Use of fungal technology in soil remediation: A Case Study

Two white rot fungi Irpex lacteus and Pleurotus ostreatus and a PAH-degrading bacterial strain of Pseudomonas putida were used as inoculum for bioremediation of petroleum hydrocarbon-contaminated soil from a manufactured-gas-plant-area. Also two cocultures comprising a fungus with Pseudomonas putida were applied. After 10-week treatment out of 12 different PAHs, concentration of phenanthrene, anthracene, fluoranthene and pyrene decreased up to 66%. The ecotoxicity of the soil after bioremediation did not reveal any effect on the survival of Daphnia magna, a crustacian. However, the toxic effect on seed germination of plant Brassica alba and oxidoreductase activity of bacterium Bacillus cereus decreased after 5 and 10 weeks of treatment.     

Changes in Extractability of Cr and Pb in a Polycontaminated Soil After Bioaugmentation With Microbial Producers of Biosurfactants, Organic Acids and Siderophores

Partly because of the low bioavailability of metals, the soil cleaning-up using phytoremediation is usually time-consuming. In order to enhance the amount of metals at the plant's disposal, the soil bioaugmentation coupled together with phytoextraction is an emerging technology. In this preliminary work, two agricultural soils which mainly differed in their Cr, Hg and Pb contents (LC, low-contaminated soil; HC, high-contaminated soil) were bioaugmented in laboratory conditions by either bacterial (Bacillus subtilis, Pseudomonas aeruginosa, Pseudomonas fluorescens or Ralstonia metallidurans) or fungal inocula (Aspergillus niger or Penicillium simplicissimum) and incubated during three weeks. The LC soil pots bioaugmented with A. niger and P. aeruginosa contained higher concentrations of Cr (0.08 and 0.25 mg.kg⁻¹ dw soil) and Pb (0.25 and 0.3 mg.kg⁻¹ dw soil) in the exchangeable fraction F1 (extraction with MgCl₂) by comparison with the non-bioaugmented soil where neither Cr nor Pb was detected. Conversely, immobilization of Cr and Pb in the soil were observed with the other microorganisms. The soil bioaugmentation not only modified the metal speciation for the most easily extractable fractions but also modified the distribution of metals in the other fractions, to a lesser extent nevertheless. The difference in microbial concentrations between the bioaugmented or not HC soils reached up to 1.8 log units. Thus the microorganisms that we chose for the soil bioaugmentation were competitive towards the indigenous microflora. The PCA analysis showed close positive relationships between the microorganisms which potentially produced siderophores in the soil and the amount of Cr and Pb in the fraction F1.     

Ethanol washing of PAH-contaminated soil and Fenton oxidation of washing solution

As a means to remediate soil contaminated by polycyclic aromatic hydrocarbons, we investigated a combined process involving ethanol washing followed by a Fenton oxidation reaction. Artificial loamy soil was contaminated with various representative polycyclic aromatic hydrocarbons (i.e., fluorene, anthracene, pyrene, benzo(b)fluoranthene, or benzo(a)pyrene) at concentrations ten times higher than regulatory soil standards of The Netherlands or Canada, and then washed four times in ethanol, which reduced the concentration of polycyclic aromatic hydrocarbon contamination to below the regulatory standard. Fenton oxidation of ethanol solutions containing anthracene, benzo(a)pyrene, pyrene, acenaphthylene, acenaphthene, benz(a)anthracene, benzo(j)fluoranthene, or indeno(1,2,3-cd)pyrene showed a removal efficiency of 73.3%–99.0%; by contrast, solutions containing naphthalene, fluorene, fluoranthene, phenanthrene, or benzo(b)fluoranthene showed a removal efficiency of 9.6%–27.6%. Since each of the nonremediated polycyclic aromatic hydrocarbons, excluding benzo(b)fluoranthene, are easily biodegradable, these results indicate that the proposed treatment can be successfully applied to polycyclic aromatic hydrocarbon-contaminated soil that does not contain high concentrations of benzo(b)fluoranthene. The main reaction products resulting from Fenton oxidation of ethanol solutions containing anthracene or benz(a)anthracene were anthraquinon or benz(a)anthracene-7,12-dione, respectively; while 1,8-naphthalic anhydride was produced by solutions of acenaphthylene and acenaphthene, and 9-fluorenone by a fluorene solution. Received: June 9, 1998 / Accepted: March 24, 1999     

有机溶剂浸取石油污染土壤中的多环芳烃

采用索氏提取法和气相色谱分析了某炼油厂周边土壤中的多环芳烃的种类及含量,考察了6种有机溶剂对土壤中多环芳烃的浸取效果,探讨了溶剂与溶质溶解度参数差异对浸取效果的影响。结果表明：土壤试样中含有蒽、荧蒽、芘、 、苯并[a]蒽、苯并[b]荧蒽、苯并[k]荧蒽和苯并[a]芘8种多环芳烃,其含量分别为4.20,22.05,10.62,4.26,5.54,0.80,0.94,4.18 mg/kg;筛选出二氯甲烷作为土壤中多环芳烃的浸取溶剂;最佳浸取条件为浸取时间10 min、浸取温度30 ℃、溶剂与土壤的液固比5:1（mL/g）、土壤含水量8%,在此条件下,总多环芳烃浸出率为83.0%,各种多环芳烃的浸出率分别为蒽97.8%、荧蒽78.2%、芘99.9%、 98.5%、苯并[a]蒽81.1%、苯并[b]荧蒽47.6%、苯并[k]荧蒽14.8%、苯并[a]芘58.7%。     

Permanganate oxidation of sorbed polycyclic aromatic hydrocarbons

The polycyclic aromatic hydrocarbons (PAH) that contaminate soils at many industrial and government sites are resistant to natural biotic and abiotic degradation processes. The recalcitrant nature of these compounds may require aggressive chemical treatment to effectively remediate these sites. This study was conducted to assess the viability of permanganate oxidative treatment as a method to reduce PAH concentration in contaminated soils. Study results demonstrated a reduction in soil sorbed concentration for a mixture of six PAHs that included anthracene, benzo(a)pyrene, chrysene, fluoranthene, phenanthrene, and pyrene by potassium permanganate (KMnO4) oxidative treatment. The greatest reduction in soil concentration was observed for benzo(a)pyrene, pyrene, phenanthrene, and anthracene at 72.1, 64.2, 56.2, and 53.8%, respectively, in 30 min at a KMnO4 concentration of 160 mM. Minimal reductions in fluoranthene and chrysene concentration were observed at 13.4 and 7.8%, respectively, under the same conditions. A relative chemical reactivity order of benzo(a)pyrene>pyrene>phenanthrene>anthracene>fluoranthene>chrysene towards permanganate ion was observed. Aromatic sextet theory was applied to the degradation results to explain the highly variable and compound-specific chemical reactivity order.     

Factors affecting polycyclic aromatic hydrocarbon biodegradation by Aspergillus flavus

This study investigated the ability of fungi isolated from highly contaminated soil to biodegrade polycyclic aromatic hydrocarbon (PAH) compounds, as well as the effect of several parameters on the biodegradation ability of these fungi. The isolated fungi were identified using ITS rDNA sequencing and tested using 2,6‐dichlorophinolendophenol to determine their preliminary ability to degrade crude oil. The top‐performing fungi, Aspergillus flavus and Aspergillus fumigatus, were selected to test their ability to biodegrade PAH compounds as single isolates. After 15 days of incubation, A. flavus degraded 82.7% of the total PAH compounds, with the complete degradation of six compounds, whereas A. fumigatus degraded 68.9% of the total PAHs, with four aromatic compounds completely degraded. We also tested whether different temperatures, pH, and nitrogen sources influenced the growth of A. flavus and the degradation rate. The degradation process was optimal at a temperature of 30°C, pH of 5.5, and with nitrogen in the form of yeast extract. Finally, the ability of the fungal candidate, A. flavus, to degrade PAH compounds under these optimum conditions was studied. The results showed that 95.87% of the total PAHs, including 11 aromatic compounds, were completely degraded after 15 days of incubation. This suggests that A. flavus is a potential microorganism for the degradation of PAH compounds in aqueous cultures.     

Literature overview: Microbial metabolism of high molecular weight polycyclic aromatic hydrocarbons

High molecular weight polycyclic aromatic hydrocarbons (HMW PAHs) increase in hydrophobicity with increases in their molecular weight and ring angularity. Microbial strategies to deal with PAH hydrophobicity include biofilm formation, enzyme induction, and biosurfactants, the effect of which is variable on PAH metabolism depending on the surfactant type and concentration, substrate, and microbial strain(s). Aerobic HMW PAH metabolism proceeds via mineralization, partial degradation, and cometabolic transformations. Generally, bacteria and nonlignolytic fungi metabolize PAHs via initial PAH ring oxidation by dioxygenases to form cis‐dihydrodiols, which are transformed to catechol compounds by dehydrogenases and other mono‐ and dioxygenases to substituted catechol and noncatechol compounds, all ortho‐ or metacleaved and further oxidized to simpler compounds. However, lignolytic fungi form quinones and acids to CO₂. This review discusses the pathways for HMW PAH microbial metabolism. © 2008 Wiley Periodicals, Inc.     

Degradation of L- and DL-lactic acid oligomers in the presence ofFusarium moniliforme andPseudomonas putida

Poly(-alkanoates) derived from lactic acid enantiomers are known to degrade easily hydrolytically in aqueous media. The ability of two microorganisms, a filamentous fungus,Fusarium moniliforme, and a bacterium,Pseudomonas putida, to assimilate the degradation by-products of poly(lactic acid) (PLA), namely, lactic acid, lactyllactic acid dimers, and higher oligomers, was investigated in liquid culture. To distinguish the influence of chirality on bioassimilation, two series of substrates were considered which derived from the racemic and the L-form of lactic acid, respectively. The fate of these compounds was monitored by HPLC. Under the selected conditions,DL- andL-lactic acids were totally used by the two microorganisms regardless of the enantiomeric composition. Both microorganisms degraded the LL-dimer rather rapidly. However,F. moniliforme acted more rapidly thanP. putida. It is likely that the DD-dimer also biodegraded but at a slower rate, especially in the case of the fungi. Higher racemic oligomers were slowly assimilated by the two microorganisms, whereas higher L-oligomers appeared biostable probably because of their crystallinity. A synergistic effect was observed when both microorganisms were present in the same culture medium containing racemic oligomers.Presented at the 4th International Workshop on Biodegradable Plastics and Polymers, October 11–14, 1995. Durham, New Hampshire.     

Enhanced biodegradation of creosote-contaminated soil

Bioremediation, a viable option for treatment of cresote-contaminated soil, can be enhanced by the use of surfactant. A study was conducted to investigate the effect of a non-ionic surfactant, Triton X-100, on biodegradation of creosote-contaminated soil. Abiotic soil desorption experiments were performed to determine the kinetics of release of selected polynuclear aromatic hydrocarbon (PAH) compounds. Respirometric experiments were also conducted to evaluate the effect of nonionic surfactant on biodegradation. The N-Con system respirometer was used to monitor the oxygen uptake by the microorganisms. The abiotic experiments results indicated that the addition of surfactant to soil/water systems increased the desorption of PAH compounds. It was also observed that the desorption rate of PAH compounds depended on their molecular weight. The 3- and 4-ring PAH compounds showed higher and faster desorption rates than the 5- and 6-ring PAHs. The respirometric experiments indicated that an increase in soil contamination level from 112.5 to 771.8 mg/kg showed an increase in oxygen uptake. But for a soil contamination level of 1102.5 mg/kg, the oxygen uptake was similar to the contamination level of 771.8 mg/kg. This might be due to toxicity by the surfactant or the solubilized PAHs at high concentration or interference with contaminant transport into the cell or to reversible physical-chemical interferences with the activity of enzymes involved in the PAH degradation. The increase in PAH availability to the microorganisms in the aqueous phase produced an increase in oxygen consumption that is proportional to the biodegradation of organic compounds.     

Evaluation of white‐rot fungi for the remediation of creosote‐contaminated soil

Application of fungal‐based bioaugmentation was evaluated for the remediation of creosote‐contaminated soil at a wood‐preserving site in West Virginia. Soil at the site contained creosote‐range polycyclic aromatic hydrocarbons (PAHs) at concentrations in some areas that exceed industrial risk‐based levels. Two white‐rot fungi (Pleurotus ostreatus and Irpex lacteus) were evaluated for remediation effectiveness in a two‐month bench‐scale treatability test. Both fungi produced similar results, with up to 67.3 percent degradation of total PAHs in 56 days. Pilot‐scale testing was performed at the site using Pleurotus ostreatus grown on two local substrate mixtures. During the 276‐day field trial, total PAHs were degraded by up to 93.2 percent, with all individual PAHs except one achieving industrial risk‐based concentrations. It was recommended that fungal‐based remediation be applied to all contaminated soil at the site. © 2002 Wiley Periodicals, Inc.     

High Immobilization of NH4+ in Danish Heath Soil Related to Succession,Soil and Nutrients: Implications for Critical Loads of N

During recent decades heathlands havechanged into grasslands in regions with high atmosphericnitrogen deposition. In regions with intermediatedeposition level (e.g., Denmark) changes have been lesspronounced which may be due to delay or decrease inresponse of the ecosystem. The mor layer (O horizon) mayplay an important role for this delay due to high sinkstrength for N. In this study, the capacity for netNH₄ ⁺ immobilization and mineralization wasstudied during short- and long-term incubations (2–36 days)of mor samples from Danish dry inland heaths. High short-term capacity for net NH₄ ⁺ immobilization wasfound to be a general characteristic of Danish heath morlayers both under heather (Calluna vulgaris) andcrowberry (Empetrum nigrum ssp nigrum), the latterdominating late stages in heathland succession. The netNH₄ ⁺ immobilization was higher under youngcompared to old or dead vegetation, and higher on lessnutrient poor soils than on extremely nutrient poor soils.The addition of N, P and C stimulated CO₂ productionand net NH₄ ⁺ immobilization, but not net Nmineralization. The immobilization of ¹⁵NH₄ ⁺caused release of dissolved organic N, increased N anddecreased C/N ratio in the microbial biomass, and indicatedgrowth of microorganisms with other metabolic abilitiesthan the indigenous population. No evidence was obtained ofstabilization of immobilized ¹⁵NH₄ ⁺ intosoil organic matter during the experiment. On background ofthe results and current knowledge it was concluded that therecognition of the high capacity for net NH₄ ⁺immobilization in mor layers does not allow for a raiseof critical loads for N for northern dry inland heaths.     

Distribution of Cd,Ck, Pb and Zn in Soil and Vegetation Compartments in Stands of Five Boreal Tree Species in N.E. Sweden

Concentrations and total quantity of cadmium (Cd), cupper (Cu),lead (Pb) and zink (Zn) were determined in biomass and soil compartments in a replicated tree species experiment with 27-yr-old stands growing on former farmland in N.E. Sweden. Sequentialextractions of soil samples were performed in order to estimate the exchangeable and an organically bound fraction of each element. The tree species included were Picea abies (L.)H. Karst., Pinus sylvestris L., Pinus contorta Dougl., Larix sibirica Ledeb., and Betula pendula Roth.Tree species influenced the rate of removal of Cu, Pb and Zn incase of stemwood harvesting, and of Cd, Cu and Zn in the case ofwhole-tree harvesting. B. pendula and P. abies had higher quantities and average concentrations of Zn in the biomass. For all species, >50% of the Zn in the stems was found in the bark. P. abies and L. sibirica had higher quantities of Cu in the biomass than the other species.P. abies and P. contorta had high quantities of Cd inthe biomass in relation to the other species. Branches and stembark contained high concentrations of Cd and Pb in relation to foliage and stemwood. Dead branches had especially high concentrations of Pb. The high accumulation rate of Zn in thebiomass of B. pendula was related to a low exchangeable amount of Zn in the A horizon. In the superficial centimeters ofthe A horizon, a depletion similar to that found for Zn was detected for Cu, whereas for Cd and Pb, no correlations were found between quantities of elements in the trees and element pools in the soil.     

Degradation of Poly(3-hydroxybutyrate) and Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) by Some Soil Aspergillus spp.

Systematic screening of 45 soil fungi for degradation polyhydroxyalkanoic acids (PHAs) has led to the selection of 6 potent Aspergillus isolates belonging to A. flavus, A. oryzae, A. parasiticus, and A. racemosus. Degradation of PHAs as determined by tube assay method revealed that these Aspergillus spp. were more efficient in degrading poly(3-hydroxybutyrate) [P(3HB)] compared to copolymer of 3-hydroxybutyric acid and 3-hydroxyvaleric acid (P3HB-co-16% 3HV). Moreover, the extent of degradation in mineral base medium was much better than those in complex organic medium. For all the Aspergillus spp. tested, maximum degradation was recorded at a temperature of 37°C with significant inhibition of growth. The optimum pH range for degradation was 6.5–7.0 with degradation being maximum at pH 6.8. The extent of polymer degradation increased with increase in substrate concentration, the optimum concentration for most of the cultures being 0.4% and 0.2% (w/v) for P(3HB) and P(3HB-co-16%3HV) respectively. Supplementation of the degradation medium with additional carbon sources exerted significant inhibitory effect on both P(3HB) and P(3HB-co-16%3HV) degradation.     

A Study of the Use of Alfalfa (Medicago sativa L.) for the Phytoremediation of Organic Contaminants in Soil

This article presents the results of a study that was conducted to determine the effectiveness of using alfalfa (Medicago sativa L.) to enhance the phytoremediation of three different types of chemical contaminants. The chemicals studied were trinitrotoluene (TNT), the polycyclic aromatic hydrocarbon (PAH) pyrene, and the polychlorinated biphenyl (PCB) Aroclor 1248. Experiments were conducted using soils that contained high and low organic matter content. The results indicated that recoveries of pyrene and TNT from soil were highly dependent on the soil organic matter content, while the recovery of PCB was not. Significantly low levels of pyrene and TNT were recovered from all treatments in the soil with 6.3 percent organic matter content compared to recovery levels found in soil with 2.6 percent organic matter. The presence of alfalfa plants had a significant effect on the transformation of TNT and PCB in the low organic matter content soil only and had no effect on the fate of pyrene. In the low organic matter soil, only 15 percent and 17 percent of the initial TNT and PCB levels, respectively, were transformed in the unplanted control soils compared to 66 percent and 77 percent in the alfalfa planted pots. In both soil types, pyrene dissipation could not be attributed to the presence of alfalfa plants. Overall, it was concluded that under high soil organic matter conditions, adsorption and covalent binding to the soil organic matter appeared to be the dominant force of pyrene and TNT removal. The effectiveness of using alfalfa to enhance PCB and TNT transformations was more significant in the lower organic matter soil; thus phytoremediation had a greater effect in soils with lower organic matter content. © 2001 John Wiley & Sons, Inc.     

Biodegradation of Aliphatic and Aromatic Hydrocarbons: Specificity among Bacteria Isolated from Refinery Waste Sludge

Indigenous microorganisms, enriched and isolated from refinery waste sludge, were observed to possess a broad range of metabolic activities for mixtures of several classes of substrates of petroleum hydrocarbons, such as monoaromatic and polycyclic aromatic hydrocarbons (PAHs) and n- and branched alkanes. Three of the best-growing bacterial isolates selectively enriched with these compounds were identified by 16S rDNA sequencing as belonging to the genera Enterobacter and Ochrobactrum. Two of them, Enterobacter sp. strain EK3.1 and Ochrobactrum sp. strain EK6 utilise a hydrocarbon mixture of the branched alkane 2,6,10,14-tetramethylpentadecane and the PAHs acenaphthylene and acenaphthene. Enterobacter sp. strain EK4 can grow with a mixture of 2,6,10,14-tetramethylpentadecane, toluene, acenaphthylene and acenaphthene as carbon sources. Nucleic acid fingerprint analysis, by terminal restriction fragment length polymorphism (T-RFLP) of the PCR-amplified 16S rRNA genes, of the autochthonous bacterial community in contaminated soil samples showed complex and different community structures under different treatments of refinery waste sludge in landfarm areas. The characteristic peaks of the T-RFLP profiles of the individual, isolated degrading bacteria Enterobacter spp. and Ochrobactrum sp. were detected in the T-RFLP fingerprint of the bacterial community of the four months old treated landfarm soil, suggesting the enrichment of bacteria belonging to the same operational taxonomic units, as well as their importance in degrading activity.     

Long-Term Maintenance of Rapid Atrazine Degradation in Soils Inoculated with Atrazine Degraders

Two different microbial communities able to degrade atrazine (atz) were inoculated in four different soils. The most critical factor affecting the success of inoculation was the soil pH and its organic matter (OM) content. In two alkaline soils (pH > 7), some inoculations led immediately to a strong increase of the biodegradation rate. In a third slightly acidic soil (pH = 6.1), only one inoculum could enhance atz degradation. In a soil amended with organic matter and straw (pH = 5.7, OM = 16.5%), inoculation had only little effect on atz dissipation on the short as well as on the long-term. Nine months after the microflora inoculations, atz was added again and rapid biodegradation in all alkaline inoculated soils was recorded, indicating the long-term efficiency of inoculation. In these soils, the number of atz degraders was estimated at between 6.5 × 10³ and 1.5 × 10⁶ (g of soil)^-1, using the most probable number (MPN) method. Furthermore, the presence of the atz degraders was confirmed by the detection of the gene atzA in these soils. Denaturing gradient gel electrophoresis (DGGE) analysis of the 16S rDNA genes indicated that the inoculated bacterial communities had little effect on the patterns of the indigenous soil microflora.     

Microbial Inoculants on Woody Legumes to Recover a Municipal Landfill Site

Tree and shrubby legumes have great potential in degraded land rehabilitation because of their ability to form symbiotic associations with nitrogen fixing rhizobia and mycorrhizal fungi. Extensive soil disturbance reduces natural microbial propagules thus preventing the formation of beneficial plant-microbes symbiosis. Reintroduction of selected microbial symbionts may improve the recovery rate of disturbed ecosystems. We inoculated selected rhizobia and arbuscular mycorrhizal fungi on two woody legume species, the mediterranean shrub Spartium junceumL. and the exotic tree Acacia cyanophylla Lindl. in order to recover a sealed municipal landfill (Palermo, Sicily, Italy). Inoculated plants showed shoot growth parameters 2 to 12-fold higher than uninoculated plants. After transplanting on the municipal landfill site, inoculated plants showed no transplant shock and low mortality (6–15%). The chemical analysis of P and N plant content showed no differences between inoculated and uninoculated plants suggesting that a dilution effect occurred due to higher biomass production of the inoculated plants. The beneficial effects of mycorrhization and rhizobium inoculum on growth parameters were still detectable one year after transplanting in S. junceum.     

Effects of physicochemical factors on PAH degradation by Planomicrobium alkanoclasticum

Polycyclic aromatic hydrocarbons (PAHs) are environmental pollutants that are mutagenic, carcinogenic, and toxic to living organisms. Here, the ability and effectiveness of selected bacteria isolated from an oil‐contaminated area in biodegrading PAHs were evaluated, and the optimal conditions conducive to bacterial PAH biodegradation were determined. Of six bacterial isolates identified based on their 16S rRNA sequences, Planomicrobium alkanoclasticum could subsist on and consume nearly all hydrocarbons according to the 2,6‐dichlorophenolindophenol assay. The efficacy of this isolate at PAH biodegradation was then empirically confirmed. After 30 days of incubation, P. alkanoclasticum degraded 90.8% of the 16 PAH compounds analyzed and fully degraded eight of them. The optimum P. alkanoclasticum growth conditions were 35°C, pH 7.5, and NaNO₃ as the nitrogen source. Under these biostimulant conditions, P. alkanoclasticum degraded 91.4% of the total PAH concentration and completely decomposed seven PAHs after 15 days incubation. Hence, P. alkanoclasticum is an apt candidate for the biodegradation of PAHs and the bioremediation of sites contaminated by them.     

Composition of the Saprotrophic Fungi in Calluna Heathland Soil and the Influence of Ammonium Nitrate Application

The composition of saprotrophic soil fungi in the mor layerof a Calluna-dominated, Danish heathland wasinvestigated after two years of fertilization with ammoniumnitrate (0, 35, 50 and 70 kg N ha^1- yr^-1) using asoil washing technique. The most frequently isolated generafrom the soil particles plated were Penicillium, Trichoderma, Mortierella and Mucor. Eightspecies of Penicillium were identified and Penicillium spinulosum was the most frequently isolated.The occurrence of dark, sterile fungi on the soil particleswas low. There were no measurable changes in the specificcomposition of the saprotrophic soil fungal groups due tothe nitrogen treatments, beside from an increasedoccurrence of Absidia californica. I conclude that adirect impact on the composition of the saprotrophic fungiin heathland soil is unlikely under enhanced nitrogen input.     

Managing bioremediation of a creosote‐contaminated superfund site by optimizing moisture and temperature in a biopile

Soil moisture content and temperature in a contaminated soil biopile equipped with immobilized microbe bioreactors (IMBRs) were optimized during ex situ bioremediation at a creosote‐contaminated Superfund site. Efficiency of remediation during warm summer months without soil‐temperature and moisture optimization was compared with that of cold winter months when corrective measures were applied. Significant reduction (35 percent) in total polycyclic aromatic hydrocarbons (PAHs) was observed, compared to 3.97 percent without corrective measures (p < 0.05). Kinetic rates (KRs) for total PAH removal were significantly enhanced from 3.93 to 50.95 mg/kg/day. KRs for removal of high molecular mass four‐to‐six‐ring PAHs were also significantly enhanced from 70.29 mg/kg/day to 97.45 mg/kg/day ( p < 0.05). Bioremediation of two‐ and three‐ring PAHs increased significantly from 15 percent to 40 percent. Benzo[a]pyrene toxicity equivalent mass (BaP_equiv) was significantly reduced by 48 percent with KR of 0.47 mg/kg/day as compared to 22 percent with KR of 0.14 mg/kg/day (p < 0.05). Soil moisture content was enhanced from 15.7 percent to 41.4 percent. © 2007 Wiley Periodicals, Inc.