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Decomposer animals and bioremediation of soils.

Although microorganisms are degrading the contaminants in bioremediation processes, soil animals can also have important--while usually an indirect--role in these processes. Soil animals are useful indicators of soil contamination, both before and after the bioremediation. Many toxicity and bioavailability assessment methods utilizing soil animals have been developed for hazard and risk-assessment procedures. Not only the survival of the animals, but also more sensitive parameters like growth, reproduction and community structure have often been taken into account in the assessment. The use of bioassays together with chemical analyses gives the most reliable results for risk analyses. This is because physical, chemical and biological properties of the remediated soil may be changed during the process, and it is possible that transformation rather than mineralization of the contaminants has taken place. In addition, the soil may contain other harmful substances than those searched in chemical analyses. Finally, because the ultimate goal of the bioremediation should be--together with mineralization of the harmful substances--the ecological recovery of the soil, development of diverse decomposer community as a basis of the functioning ecosystem should be ensured. Soil animals, especially the large ones, can also actively take part in the ecological recovery processes through their own activity. The potential risk of transfer of contaminants accumulated in soil animals to the above-ground food webs should be borne in mind.

Journal Article↗

Field applications of genetically engineered microorganisms for bioremediation processes.

Genetically engineered microorganisms (GEMs) have shown potential for bioremediation applications in soil, groundwater, and activated sludge environments, exhibiting enhanced degradative capabilities encompassing a wide range of chemical contaminants. However, the vast majority of studies pertaining to genetically engineered microbial bioremediation are supported by laboratory-based experimental data. In general, relatively few examples of GEM applications in environmental ecosystems exist. Unfortunately, the only manner in which to fully address the competence of GEMs in bioremediation efforts is through long-term field release studies. It is therefore essential that field studies be performed to acquire the requisite information for determining the overall effectiveness and risks associated with GEM introduction into natural ecosystems.

Biodegradation, Environmental↗

Intrinsic bioremediation: an environmental restoration technology.

In the past year, there has been a dramatic maturation of intrinsic bioremediation technologies. As methods pertinent to the intrinsic bioremediation of fuel hydrocarbons have become better established, a comprehensive protocol has been developed for the validation and prediction of these processes. Study of intrinsic bioremediation of other prevalent contaminants, such as chlorinated aliphatics, polychlorinated biphenyls, and high explosives, has also been undertaken.

Biodegradation, Environmental↗

A bioengineering system for in situ bioremediation of contaminated groundwater.

Much of the past and current focus of bioremediation has been on laboratory studies of microbial processes. By necessity, early studies have ignored important field properties, parameters, and processes that control the ultimate success of in situ bioremediation of contaminated groundwater. This paper presents a bioengineering systems approach that examines the impact of some of these field variables on common bioremediation practices. Using simple systems, the niche of biostimulation is shown to be aquifers with high contaminant sorption. A novel gas-phase biostimulation filter and a novel resting-state bioaugmentation/biofilter approach which show promise for effective field implementation are discussed.

Bacteria↗

Response of bacterial community during bioremediation of an oil-polluted soil.

AIM: To study the response of the bacterial community to bioremediation of a soil with an aged contamination of crude oil. METHODS AND RESULTS: The bacterial community in laboratory soil columns during a 72-day biostimulation treatment was followed by analysing the number of total cultivable hydrocarbon-degrading bacteria, soil respiratory activity and the 16S-23S rDNA internal transcribed spacer homoduplex heteroduplex polymorphisms (ITS-HHP) of total soil bacterial DNA. ITS-HHP permits an estimate of both length and sequence polymorphism in a 16S-23S rDNA spacer population, using to advantage the homoduplex and heteroduplex fragments that are generated during PCR. The treatment, made by air sparging and biostimulation with a mineral nutrient and surfactant solution, resulted in a 39.5% decrease of the total hydrocarbon content. Within 4 days of treatment onset the bacterial community underwent a first phase of activation that led to a substantial increase in the observable diversity. Subsequently, after a 12-day period of stability, another activation phase was observed with further shifts of the community structure and an increase in the abundance and diversity of catechol-2,3-dioxygenase (C23O) genes. CONCLUSIONS: The overall data suggest an important contribution of uncultivable bacteria to the soil bioremediation, since, during the second activation phase, the increases of the respiratory activity, bacterial diversity and C23O gene abundance and diversity were not accompanied by a corresponding increase of the cultivable bacteria number. SIGNIFICANCE AND IMPACT OF THE STUDY: This study shows that successive phases of activation of bacterial populations occur during a bioremediation treatment of oil-polluted soil.

Bacteria↗

Bioremediation of crude oil polluted soil by the white rot fungus, Pleurotus tuberregium (Fr.) Sing.

Bioremediation has become an attractive alternative to physicochemical methods of remediation of polluted sites. White rot fungi (WRF) are increasingly being investigated and used in bioremediation, because of their ability to degrade an extremely diverse range of very persistent or toxic environmental pollutants. The white rot fungus, Pleurotus tuberregium, was examined for its ability to ameliorate crude oil polluted soil. This was inferred from the ability of the polluted soil to support seed germination and seedling growth in Vigna unguiculata, at 0, 7 and 14 days post treatment. Results obtained from the present study showed that bioremediation of soil contaminated with crude oil was possible, especially when the fungus had been allowed to establish and fully colonize the substrate mixed with the soil. There were significant improvements in % germination, plant height and root elongation values of test plants, when seeds were planted 14 days post soil treatment. At 1 to 5% crude oil pollution, % germination values were comparable with the values in control plants in the 14 days treatment, and significantly higher than values obtained in the day 0 treatment. Also, at the highest level of crude oil pollution (15%), there was about 25% improvement in % germination value over the 0 day treatment. This trend of improvement in values was also observed for plant height, root elongation and biomass accumulation as well as decreased total hydrocarbon content.

Biodegradation, Environmental↗

Mechanisms prevalent during bioremediation of wastewaters from the pulp and paper industry.

Bioremediation of wastewaters represents an important treatment methodology, especially when examined against the backdrop of ever-stricter legislation that is evolving in order to regulate effluent release into the environment. It has been reported that bioremediation specifically holds promise in solving environmental problems. Crucial questions surrounding the treatment of effluents include: efficiency of the process, economic feasibility, legal requirements, and the mechanisms involved in the remediation process. Of all these issues mentioned, the last requires special attention. This paper investigates these matters and focuses on techniques that are currently employed to determine the efficiency of bioremediation and mechanisms involved therein. The physiological significance of biosorption is also examined, as this subject has not been fully addressed in previous publications.

Basidiomycota↗

Comparison of ex situ and in situ bioremediation of unsaturated soils contaminated by petroleum.

The most feasible bioremediation technologies of unsaturated soils contaminated by Petroleum are (1) the on-site controlled remediation of the excavated contaminated plume in the contained system; and (2) the in situ sub-surface, highly controlled process in which nutrients and microbes or air are injected into the plume volume via a grid of wells and reversing the direction of the pumping. The detailed results of ex situ (controlled aerobic bioremediation) of ten sites with differing petroleum contamination and four in situ cases are reported and compared. It is concluded, that bioremediation of petroleum contaminated soil is an effective, safe and economically feasible remediation technique for the remediation of such contaminated sites, contingent on the providing of the site-specific appropriate conditions. However, the biodegradation rate of the NAPL is more variable in the in situ than in the ex situ aerobic process, correlating best in the latter with the molecular weight of the petroleum contaminants.

Bacteria, Aerobic↗

Liposomes enhance bioremediation of oil-contaminated soil.

Liposomes (composed of soy phosphatides) in the form of small unilamellar vesicles (SUV), when added to soil contaminated by crude oil, accelerate bioremediation. After three weeks incubation at 30 degrees C, using soil experimentally contaminated (with 10,000 ppm crude oil), level of bioremediation increased from 40% without SUV to 75% with SUV (0.1 wt% phospholipids per dry weight soil). Similarly, for accidentally contaminated soil (with approximately 17,000 ppm crude oil), addition of 0.1 wt% SUV to the soil increased the bioremediation level from 55 to 80%. The enhancing effect of liposomes is explained by two interrelated phenomena: a large increase both in total bacteria number and in diversity of bacterial species in the soil. Comparison after four weeks revealed 21 bacterial species in the presence of liposomes (many being oil-degrading bacterial species) and only nine species in the absence of liposomes. Both effects may be related to the physical effects of liposome phospholipids, which modify the crude oil by wetting it, thereby making it more accessible to the microorganisms. In addition, liposome phospholipids serve as phosphate and nitrogen sources for the bacteria.

Bacteria↗

Analysis of the genetic potential and gene expression of microbial communities involved in the in situ bioremediation of uranium and harvesting electrical energy from organic matter.

The proposed research will investigate two microbial communities that are of direct relevance to Department of Energy interests. One is the microbial community associated with the in situ bioremediation of uranium-contaminated groundwater. The second is a microbial community that harvests energy from waste organic matter in the form of electricity. These studies will address DOE needs for (1) remediation of metals and radionuclides at DOE sites and (2) the development of cleaner forms of energy and biomass conversion to energy. Our previous studies have demonstrated that the microbial communities involved in uranium bioremediation and energy harvesting are both dominated by microorganisms in the family Geobacteraceae and that the organisms in this family are responsible for uranium bioremediation and electron transfer to electrodes. The initial objectives of this study are to (1) describe the genetic potential of the Geobacteraceae that predominate in the environments of interest; (2) identify conserved patterns of gene expression within the Geobacteraceae family in response to a range of environmental conditions; (3) begin to identify mechanisms controlling the expression of key genes related to survival, growth, and activity in subsurface environments and on electrodes; and (4) use the results from subobjectives 1-3 to develop a conceptual model for predicting gene expression of Geobacteraceae in the environments of interest. This will serve as the basis for a subsequent simulation model of the growth and activity of Geobacteraceae in the subsurface and on electrodes.

Biodegradation, Environmental↗

Bioremediation of creosote-contaminated soil in South Africa by landfarming.

AIMS: To determine the combined effects of biostimulation and bioaugmentation in the landfarming of a mispah form (lithosol; food and Agriculture Organisation (FAO)) soil contaminated with >310000 mg kg-1 creosote with a view to developing a bioremediation technology for soils heavily contaminated with creosote. METHODS AND RESULTS: The excavated soil was mixed with 2500 kg ha-1 dolomitic lime and 2000 kg ha-1 mono-ammonium phosphate (MAP) before spreading over a treatment bed of shale reinforced with clay. Sewage sludge (500 kg) was ploughed into 450 m3 of contaminated soil in the second and sixth months of treatment. A further 1000 kg ha-1 MAP was added to the soil at the end of the fifth month. Moisture was maintained at 70% field capacity. Total creosote was determined by the US Environmental Protection Agency (EPA) method 418.1 and concentrations of selected creosote components were determined by gas chromatography/flame ionisation detection (GC/FID). Total creosote was reduced by more than 90% by the 10th month of landfarming. The rate of reduction in creosote concentration was highest after the addition of sewage sludge. The three-ring PAHs were more slowly removed than naphthalene and the phenolic compounds. The four- and five-ring PAHs, although persist until the end of treatment, were reduced by 76-87% at the end of the experiment. CONCLUSIONS: A combination of biostimulation and bioaugmentation during landfarming could enhance the bioremediation of soils heavily contaminated with creosote. SIGNIFICANCE AND IMPACT OF THE STUDY: The study provides information on the management of a combination of biostimulation and bioaugmentation during landfarming, and contributes to the knowledge and database necessary for the development of a technology for bioremediating creosote-contaminated land.

Agriculture↗

Low temperature bioremediation of oil-contaminated soil using biostimulation and bioaugmentation with a Pseudomonas sp. from maritime Antarctica.

AIMS: To identify native Antarctic bacteria capable of oil degradation at low temperatures. METHODS AND RESULTS: Oil contaminated and pristine soils from Signy Island (South Orkney Islands, Antarctica) were examined for bacteria capable of oil degradation at low temperatures. Of the 300 isolates cultured, Pseudomonas strain ST41 grew on the widest range of hydrocarbons at 4 degrees C. ST41 was used in microcosm studies of low temperature bioremediation of oil-contaminated soils. Microcosm experiments showed that at 4 degrees C the levels of oil degradation increased, relative to the controls, with (i) the addition of ST41 to the existing soil microbial population (bioaugmentation), (ii) the addition of nutrients (biostimulation) and to the greatest extent with (iii) a combination of both treatments (bioaugmentation and biostimulation). Addition of water to oil contaminated soil (hydration) also enhanced oil degradation, although less than the other treatments. Analysis of the dominant species in the microcosms after 12 weeks, using temporal temperature gradient gel electrophoresis, showed Pseudomonas species to be the dominant soil bacteria in both bioaugmented and biostimulated microcosms. CONCLUSIONS: Addition of water and nutrients may enhance oil degradation through the biostimulation of indigenous oil-degrading microbial populations within the soil. However, bioaugmentation with Antarctic bacteria capable of efficient low temperature hydrocarbon degradation may enhance the rate of bioremediation if applied soon after the spill. SIGNIFICANCE AND IMPACT OF THE STUDY: In the future, native soil bacteria could be of use in bioremediation technologies in Antarctica.

Alkanes↗

Actinide and metal toxicity to prospective bioremediation bacteria.

Bacteria may be beneficial for alleviating actinide contaminant migration through processes such as bioaccumulation or metal reduction. However, sites with radioactive contamination often contain multiple additional contaminants, including metals and organic chelators. Bacteria-based bioremediation requires that the microorganism functions in the presence of the target contaminant, as well as other contaminants. Here, we evaluate the toxicity of actinides, metals and chelators to two different bacteria proposed for use in radionuclide bioremediation, Deinococcus radiodurans and Pseudomonas putida, and the toxicity of Pu(VI) to Shewanella putrefaciens. Growth of D. radiodurans was inhibited at metal concentrations ranging from 1.8 microM Cd(II) to 32 mM Fe(III). Growth of P. putida was inhibited at metal concentrations ranging from 50 microM Ni(II) to 240 mM Fe(III). Actinides inhibited growth at mM concentrations: chelated Pu(IV), U(VI) and Np(V) inhibit D. radiodurans growth at 5.2, 2.5 and 2.1 mM respectively. Chelated U(VI) inhibits P. putida growth at 1.7 mM, while 3.6 mM chelated Pu(IV) inhibits growth only slightly. Pu(VI) inhibits S. putrefaciens growth at 6 mM. These results indicate that actinide toxicity is primarily chemical (not radiological), and that radiation resistance does not ensure radionuclide tolerance. This study also shows that Pu is less toxic than U and that actinides are less toxic than other types of metals, which suggests that actinide toxicity will not impede bioremediation using naturally occurring bacteria.

Actinoid Series Elements↗

First production-level bioremediation of explosives-contaminated soil in the United States.

Umatilla Army Depot Activity (UMDA) near Hermiston, Oregon was the location of the first production-level bioremediation of explosives-contaminated soil in the U.S. Soil from munitions washout lagoons contained high concentrations of TNT (2,4,6-Trinitrotoluene) and RDX (Hexahydro-1,3,5-trinitro-1,3,5- triazine) as well as HMX (Octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine). In addition to these primary contaminants, laboratory tests were performed for Tetryl (Methyl-2,4,6-trinitrophenylnitramine), 4-Am-DNT (4-Amino-2,6-dinitrotoluene), 2-Am-DNT (2-Amino-4,6-dinitrotoluene), 2,4 DNT (2,4-Dinitrotoluene), 2,6 DNT (2,6-Dinitrotoluene), 1,3,5-TNB (1,3,5-Trinitrobenzene), 1,3,-DNB (1,3-Dinitrobenzene) and NB (Nitrobenzene) during the pilot-scale treatability tests. The clean-up goal established by the Record of Decision (ROD) was 30 mg/kg each for TNT and RDX. Degradation progress was monitored using immunoassay field screening Methods SW 846,4050 and 4051. Confirmational analysis consisted of EPA Method 8330. Treatment time on a 2,700 cubic yard batch (810 cubic yards of soil) was 10-12 days. A composting technique developed by the Army Environmental Center and implemented by Bioremediation Service, Inc., Portland, Oregon was used at the site. Agricultural waste products (or amendments including cow manure, chicken manure, potato waste, sawdust and alfalfa) were blended with the contaminated soil during treatment. Specialized soil turning equipment mixed the compost for optimum biological action and homogeneity. Homogeneity of the compost mix ensured rapid degradation of all contaminants. Physical and chemical properties were closely monitored to ensure that thermophilic bacteria played a dominant role in the degradation process. Nearly 5,000 cubic yards of soil have been successfully treated, and more than 70% of all analyses indicate non-detectable levels of both TNT and RDX. The U.S. Army Corps of Engineers estimates that over $2.6 million is being saved using bioremediation at Umatilla.

Azocines↗

Evaluation of inoculum addition to stimulate in situ bioremediation of oily-sludge-contaminated soil.

A full-scale study evaluating an inoculum addition to stimulate in situ bioremediation of oily-sludge-contaminated soil was conducted at an oil refinery where the indigenous population of hydrocarbon-degrading bacteria in the soil was very low (10(3) to 10(4) CFU/g of soil). A feasibility study was conducted prior to the full-scale bioremediation study. In this feasibility study, out of six treatments, the application of a bacterial consortium and nutrients resulted in maximum biodegradation of total petroleum hydrocarbon (TPH) in 120 days. Therefore, this treatment was selected for the full-scale study. In the full-scale study, plots A and B were treated with a bacterial consortium and nutrients, which resulted in 92.0 and 89.7% removal of TPH, respectively, in 1 year, compared to 14.0% removal of TPH in the control plot C. In plot A, the alkane fraction of TPH was reduced by 94.2%, the aromatic fraction of TPH was reduced by 91.9%, and NSO (nitrogen-, sulfur-, and oxygen-containing compound) and asphaltene fractions of TPH were reduced by 85.2% in 1 year. Similarly, in plot B the degradation of alkane, aromatic, and NSO plus asphaltene fractions of TPH was 95.1, 94.8, and 63.5%, respectively, in 345 days. However, in plot C, removal of alkane (17.3%), aromatic (12.9%), and NSO plus asphaltene (5.8%) fractions was much less. The population of introduced Acinetobacter baumannii strains in plots A and B was stable even after 1 year. Physical and chemical properties of the soil at the bioremediation site improved significantly in 1 year.

Acinetobacter↗

Robust hydrocarbon degradation and dynamics of bacterial communities during nutrient-enhanced oil spill bioremediation.

Degradation of oil on beaches is, in general, limited by the supply of inorganic nutrients. In order to obtain a more systematic understanding of the effects of nutrient addition on oil spill bioremediation, beach sediment microcosms contaminated with oil were treated with different levels of inorganic nutrients. Oil biodegradation was assessed respirometrically and on the basis of changes in oil composition. Bacterial communities were compared by numerical analysis of denaturing gradient gel electrophoresis (DGGE) profiles of PCR-amplified 16S rRNA genes and cloning and sequencing of PCR-amplified 16S rRNA genes. Nutrient amendment over a wide range of concentrations significantly improved oil degradation, confirming that N and P limited degradation over the concentration range tested. However, the extent and rate of oil degradation were similar for all microcosms, indicating that, in this experiment, it was the addition of inorganic nutrients rather than the precise amount that was most important operationally. Very different microbial communities were selected in all of the microcosms. Similarities between DGGE profiles of replicate samples from a single microcosm were high (95% +/- 5%), but similarities between DGGE profiles from replicate microcosms receiving the same level of inorganic nutrients (68% +/- 5%) were not significantly higher than those between microcosms subjected to different nutrient amendments (63% +/- 7%). Therefore, it is apparent that the different communities selected cannot be attributed to the level of inorganic nutrients present in different microcosms. Bioremediation treatments dramatically reduced the diversity of the bacterial community. The decrease in diversity could be accounted for by a strong selection for bacteria belonging to the alkane-degrading Alcanivorax/Fundibacter group. On the basis of Shannon-Weaver indices, rapid recovery of the bacterial community diversity to preoiling levels of diversity occurred. However, although the overall diversity was similar, there were considerable qualitative differences in the community structure before and after the bioremediation treatments.

Bacteria↗

Intensification of the aerobic bioremediation of an actual site soil historically contaminated by polychlorinated biphenyls (PCBs) through bioaugmentation with a non acclimated, complex source of microorganisms.

BACKGROUND: The biotreatability of actual-site polychlorinated biphenyl (PCB)-contaminated soils is often limited by their poor content of autochthonous pollutant-degrading microorganisms. In such cases, inoculation might be the solution for a successful bioremediation. Some pure and mixed cultures of characterized PCB degrading bacteria have been tested to this purpose. However, several failures have been recorded mostly due to the inability of inoculated microbes to compete with autochthonous microflora and to face the toxicity and the scarcity of nutrients occurring in the contaminated biotope. Complex microbial systems, such as compost or sludge, normally consisting of a large variety of robust microorganisms and essential nutrients, would have better chances to succeed in colonizing degraded contaminated soils. However, such sources of microorganisms have been poorly applied in soil bioremediation and in particular in the biotreatment of soil with PCBs. Thus, in this study the effects of Enzyveba, i.e. a consortium of non-adapted microorganisms developed from composted material, on the slurry- and solid-phase aerobic bioremediation of an actual-site, aged PCB-contaminated soil were studied. RESULTS: A slow and only partial biodegradation of low-chlorinated biphenyls, along with a moderate depletion of initial soil ecotoxicity, were observed in the not-inoculated reactors. Enzyveba significantly increased the availability and the persistence of aerobic PCB- and chlorobenzoic acid-degrading cultivable bacteria in the bioreactors, in particular during the earlier phase of treatment. It also markedly enhanced PCB-biodegradation rate and extent (from 50 to 100%) as well as the final soil detoxification, in particular under slurry-phase conditions. Taken together, data obtained suggest that Enzyveba enhanced the biotreatability of the selected soil by providing exogenous bacteria and fungi able to remove inhibitory or toxic intermediates of PCB biodegradation and/or exogenous nutrients able to sustain microorganisms in charge for PCB mineralization. CONCLUSION: Enzyveba appears a promising agent for bioaugmenting actual-site PCB-polluted soils with a native low content of indigenous specialized microflora. This not only for its positive effects on the soil biotreatability but also for its availability on the market at a relatively low cost.

Journal Article↗

Genotoxicity of bioremediated soils from the Reilly Tar site, St. Louis Park, Minnesota.

An in vitro approach was used to measure the genotoxicity of creosote-contaminated soil before and after four bioremediation processes. The soil was taken from the Reilly Tar site, a closed Superfund site in Saint Louis Park, Minnesota. The creosote soil was bioremediated in bioslurry, biopile, compost, and land treatment, which were optimized for effective treatment. Mutagenicity profiles of dichloromethane extracts of the five soils were determined in the Spiral technique of the Salmonella assay with seven tester strains. Quantitative mutagenic responses in the plate incorporation technique were then determined in the most sensitive strains, YG1041 and YG1042. Mutagenic potency (revertants per microgram extract) in YG1041 suggested that compost, land treatment, and untreated creosote soil extracts were moderately mutagenic with Arochlor-induced rat liver (S9) but were nonmutagenic without S9. However, the bioslurry extract was strongly mutagenic and the biopile extract was moderately mutagenic either with or without S9. A similar trend was obtained in strain YG1042. The strong mutagenic activity in the bioslurry extract was reduced by 50% in TA98NR, which suggested the presence of mutagenic nitrohydrocarbons. Variation in reproducibility was 15% or less for the bioassay and extraction procedures. Bioavailability of mutagens in the biopile soil was determined with six solvents; water-soluble mutagens accounted for 40% of the total mutagenic activity and they were stable at room temperature. The mutagenic activity in the bioslurry and biopsile samples was due to either the processes themselves or to the added sludge/manure amendments. The in vitro approach was effective in monitoring bioremediated soils for genotoxicity and will be useful in future laboratory and in situ studies.

Animals↗