A decision framework for selecting remediation technologies at hydrocarbon‐contaminated sites

A variety of remediation technologies are available to address hydrocarbon contamination, including free product recovery, soil venting, air sparging, groundwater recovery and treatment, and in situ bioremediation. These technologies address hydrocarbon contamination distributed between free, adsorbed, and dissolved phases in both the vadose and saturated zones. Selection of appropriate technologies is dependent on a number of factors, including contaminants, site‐specific characteristics, clean‐up goals, technology feasibility, cost, and regulatory and time requirements. This article describes a decision framework for selecting appropriate remediation technologies at hydrocarbon‐contaminated sites in a structured and tiered manner. Decision modules include (1) site characterization and product recovery; (2) vadosezone treatment: soil venting, bioremediation, and excavation; (3) saturated zone treatment: sparging, bioremediation, groundwater recovery, and excavation; and (4) groundwater treatment: carbon, air stripping, advanced oxidation, and bioreactors. Selection criteria for treatment technologies that address vadose‐ and saturated‐zone soils, as well as recovered groundwater, are described. The decision framework provides a systematic process to formulate solutions to complex problems and documents the rationale for selecting remediation systems designed to achieve closure at hydrocarbon‐contaminated sites.     

Effects of electron donor and acceptor conditions on reductive dehalogenation of tetrachloromethane by Shewanella putrefaciens 200.

Shewanella putrefaciens 200 is a nonfermentative bacterium that is capable of dehalogenating tetrachloromethane to chloroform and other, unidentified products under anaerobic conditions. Since S. putrefaciens 200 can respire anaerobically by using a variety of terminal electron acceptors, including NO3-, NO2-, and Fe(III), it provides a unique opportunity to study the competitive effects of different electron acceptors on dehalogenation in a single organism. The results of batch studies showed that dehalogenation of CT by S. putrefaciens 200 was inhibited by O2, 10 mM NO3-, and 3 mM NO2-, but not by 15 mM Fe(III), 15 mM fumarate, or 15 mM trimethylamine oxide. Using measured O2, Fe(III), NO2-, and NO3- reduction rates, we developed a speculative model of electron transport to explain inhibition patterns on the basis of (i) the kinetics of electron transfer at branch points in the electron transport chain, and (ii) possible direct inhibition by nitrogen oxides. In additional experiments in which we used 20 mM lactate, 20 mM glucose, 20 mM glycerol, 20 mM pyruvate, or 20 mM formate as the electron donor, dehalogenation rates were independent of the electron donor used. The results of other experiments suggested that sufficient quantities of endogenous substrates were present to support transformation of tetrachloromethane even in the absence of an exogenous electron donor. Our results should be significant for evaluating (i) the bioremediation potential at sites contaminated with both halogenated organic compounds and nitrogen oxides, and (ii) the bioremediation potential of iron-reducing bacteria at contaminated locations containing significant amounts of iron-bearing minerals.     

On site bioremediation of hydrocarbon-contaminated Arctic tundra soils in inoculated biopiles

There is a need to develop technology to allow the remediation of soil in polar regions that have been contaminated by hydrocarbon fuel spills. Bioremediation is potentially useful for this purpose, but has not been well demonstrated in polar regions. We investigated biopiles for on-site bioremediation of soil contaminated with Arctic diesel fuel in two independent small-scale field experiments at different sites on the Arctic tundra. The results were highly consistent with one another. In biopiles at both sites, extensive hydrocarbon removal occurred after one summer. After 1 year in treatments with optimal conditions, total petroleum hydrocarbons were reduced from 196 to below 10 mg per kg of soil at one site, and from 2,109 to 195 mg per kg of soil at the other site. Addition of ammonium chloride and sodium phosphate greatly stimulated hydrocarbon removal and indicates that biodegradation was the primary mechanism by which this was achieved. Inoculation with cold-adapted, mixed microbial cultures further stimulated hydrocarbon removal during the summer immediately following inoculation. At one site, soil temperature was monitored during the summer season, and a clear plastic cover increased biopile soil temperature, measured as degree-day accumulation, by 30-49%. Our results show that on-site bioremediation of fuel-contaminated soil at Arctic tundra sites is feasible.     

Stimulation of bioprocesses by ultrasound

Ultrasound (US) has become a ubiquitous technological process in a large variety of scientific disciplines. However, little information exists on the use of ultrasound to enhance biological processes and/or processing and consequently this paper provides an overview of work reported to date on this topic. This review provides a brief introduction to ultrasound and the history of ultrasound as applied to bioprocesses. This is followed by a discussion of the influence of US on discrete enzyme systems, enzymes used in bioremediation, microbial fermentations and enzymatic hydrolysis of biopolymers. Augmentation of anaerobic digestion by US is then considered along with enhancement of enzymes in food science and technology. The use of ultrasonically stimulated enzymes in synthesis is then considered and other relevant miscellaneous topics are described. It is concluded that the precise mechanism of action of US in bio-processing remains to be elucidated though a variety of plausible suggestions are made.     

Screening Software for an Innovative In Situ Bioremediation Technology

Innovative in situ treatment technologies show promise as efficient methods for remediating the nation's waste sites. Unfortunately, due to various barriers, some innovative technologies that have been demonstrated at full scale are never transferred for commercial application. The National Research Council (NRC) has recently presented recommendations on how to overcome these barriers (NRC, 1997). User-friendly screening software, which specifically addresses each of the NRC recommendations, is presented for use by site managers to determine the appropriateness of an innovative remediation technology, in situ aerobic cometabolic bioremediation, to clean up a contaminated site with specified hydrogeologic and contaminant characteristics. The software estimates the performance and cost of the technology at the site. Software, such as the one presented, can be used to aid in the transfer and implementation of innovative remediation technologies.     

Impact of present and future regulations on bioremediation

Summary Innovative treatment technologies are in increasing demand to clean up the nation's existing environmental contamination. There also are mounting pressures for industry to minimize the production or generation of hazardous pollutants. Bioremediation is a viable, cost-effective treatment option for both field remediation and treatment in enclosed systems. The use of innovative treatment technologies is largely regulatory driven. Over the last two decades, at least a dozen Federal environmental statutes have been enacted and hundreds of regulations implemented to control releases of pollutants into the air, water and on land. These statutes not only have created markets for the use of treatment technologies, they also may regulate some aspect of the application of that technology. Regarding bioremediation, four statutes should be reviewed to determine if compliance is necessary before employing microorganisms in the field or in enclosed systems. This paper summarizes the Federal statutes (i.e., the Toxic Substances Control Act (TSCA); the Resource Conservation and Recovery Act (RCRA); the Comprehensive Environmental Response, Compensation and Liability Act (CERCLA); and the Federal Plant Pest Act (FPPA)), and regulations that may impact the bioremediation industry; outlines potential markets for bioremediation that are being driven by regulations; and highlights, within the regulatory framework, promising applications for the bioremediation of hazardous wastes.     

A new Klebsiella planticola strain (Cd-1) grows anaerobically at high cadmium concentrations and precipitates cadmium sulfide

Heavy metal resistance by bacteria is a topic of much importance to the bioremediation of contaminated soils and sediments. We report here the isolation of a highly cadmium-resistant Klebsiella planticola strain, Cd-1, from reducing salt marsh sediments. The strain grows in up to 15 mM CdCl(2) under a wide range of NaCl concentrations and at acidic or neutral pH. In growth medium amended with thiosulfate, it precipitated significant amounts of cadmium sulfide (CdS), as confirmed by x-absorption spectroscopy. In comparison with various other strains tested, Cd-1 is superior for precipitating CdS in cultures containing thiosulfate. Thus, our results suggest that Cd-1 is a good candidate for the accelerated bioremediation of systems contaminated by high levels of cadmium.     

MINOTAURUS: microorganism and enzyme immobilization: novel techniques and approaches for upgraded remediation of underground-, wastewater and soil

The European project MINOTAURUS aims to deliver innovative bio-processes to eliminate emerging and classic organic pollutants. These bio-processes are all based on the concept of immobilization of biocatalysts (microorganisms and enzymes) and encompass bioaugmentation, enzyme technology, rhizoremediation with halophytes, and a bioelectrochemical remediation process. The immobilization-based technologies are applied to engineered ex situ and natural systems in situ for the bioremediation of groundwater, wastewater and soil. The selection and application of modern physico-chemical, biological and ecotoxicological monitoring tools combined with a rational understanding of engineering, enzymology and microbial physiology is a pertinent approach to open the black-box of the selected technologies. Reliable process-monitoring constitutes the basis for developing and refining biodegradation kinetics models, which in turn will improve the predictability of performances to be achieved with our technologies. A key strength of MINOTAURUS is the possibility of direct implementation of our technologies at five European reference sites that are confronted with pollutants (two technologies will be tested on-site starting from the first year). We will deliver not only a set of tools, techniques, and processes, which will enhance the ability of our communities to respond to the challenges of organic pollutants but also frameworks for structuring and making evidence-based decisions for the most sustainable and appropriate bioremediation measures. The MINOTAURUS consortium includes fifteen partners from eight European countries. Eight research & education institutions, five SMEs covering the whole chain of our bioremediation approaches (production, and monitoring of biocatalysts, bioremediation and engineering), one large end-user operating wastewater treatment plants and one environmental agency work together with the support of an advisory board mainly consisting of environmental decision-makers.     

A New Klebsiella planticola Strain (Cd-1) Grows Anaerobically at High Cadmium Concentrations and Precipitates Cadmium Sulfide

Heavy metal resistance by bacteria is a topic of much importance to the bioremediation of contaminated soils and sediments. We report here the isolation of a highly cadmium-resistant Klebsiella planticola strain, Cd-1, from reducing salt marsh sediments. The strain grows in up to 15 mM CdCl₂ under a wide range of NaCl concentrations and at acidic or neutral pH. In growth medium amended with thiosulfate, it precipitated significant amounts of cadmium sulfide (CdS), as confirmed by x-absorption spectroscopy. In comparison with various other strains tested, Cd-1 is superior for precipitating CdS in cultures containing thiosulfate. Thus, our results suggest that Cd-1 is a good candidate for the accelerated bioremediation of systems contaminated by high levels of cadmium.     

Microorganisms relevant to bioremediation

Naturally occurring microbial consortia have been utilized in a variety of bioremediation processes. Recent developments in molecular microbial ecology offer new tools that facilitate molecular analyses of microbial populations at contaminated and bioremediated sites. Information provided by such analyses aids in the evaluation of the effectiveness of bioremediation and the formulation of strategies that might accelerate bioremediation.     

合成生物学技术在环境保护中的应用

合成生物学是一个基于生物学和工程学原理的科学领域,其目的是重新设计和重组微生物,以优化或创建具有增强功能的新生物系统。该领域利用分子工具、系统生物学和遗传框架的重编程,从而构建合成途径以获得具有替代功能的微生物。传统上,合成生物学方法通常旨在开发具有成本效益的微生物细胞工厂进而从可再生资源中生产化学物质。然而,近年来合成生物学技术开始在环境保护中发挥着更直接的作用。本综述介绍了基因工程中的合成生物学工具,讨论了基于基因工程的微生物修复策略,强调了合成生物学技术可以通过响应特定污染物进行生物修复来保护环境。其中,规律间隔成簇短回文重复序列(Clustered Regularly Interspersed Short Palindromic Repeats, CRISPR)技术在基因工程细菌和古细菌的生物修复中得到了广泛应用,生物修复领域也出现了很多新的先进技术,包括生物膜工程、人工微生物群落的构建、基因驱动、酶和蛋白质工程等。有了这些新的技术和工具,生物修复将成为当今最好和最有效的污染物去除方式之一。     

Quantitative matrix comparisons in ecological and evolutionary investigations

The pair-wise statistical comparison of data matrices is a methodological problem which must be dealt with in a variety of disciplines. In this report we demonstrate an innovative approach using the Mantel test (a nonparametric, multivariate evaluation of test matrices) to quantitatively contrast observed color polymorphisms in male Poecilia reticulata collected from 41 samples sites in Trinidad against four evolutionary models: (1) response to an environmental gradient, (2) localized environmental patches, (3) isolation by distance, and (4) historical factors. To represent these models we derived pair-wise distances between study sites for the following data: (1) altitude, (2) density of predators, and (3) kilometric distances. To represent model four above, we generated an imposed asymptotic distance matrix for geographically contiguous sites, and a Gabriel connectivity matrix for stream-connected sites (e.g. those within the same watershed). We found that differences in color polymorphisms covary significantly with differences in predator densities and in altitudes, suggesting that male color polymorphisms track clinally distributed communities of visually hunting predators. These data substantiate previously published results from field and laboratory experiments. The utility of the Mantel procedure is that it permits a quantitative evaluation of ecological and evolutionary problems which have previously been difficult to approach statistically.     

Genomic tools in bioremediation

Bioremediation is a process that uses microorganisms or their enzymes to remove pollutants from the environment. Generally, bioremediation technologies can be classified as in situ or ex situ. In situ bioremediation involves treating the contaminated material at the site while ex situ involves the removal of the contaminated material to be treated elsewhere. Like so much else in biology, the ease and availability of genomic data has created a new level of understanding this system. Bioremediation capabilities of the microbial population can be analyzed; not only by physiological parameters, but also by the use of genomic tools, and efficient remediation strategies can be planned. PCR and DNA- or oligonucleotide-based microarray technology is a powerful functional genomics tool that allows researchers to view the physiology of a living cell from a comprehensive and dynamic molecular perspective. This paper explores the use of such tools in bioremediation process.     

A comprehensive overview of elements in bioremediation

Sustainable development requires the development and promotion of environmental management and a constant search for green technologies to treat a wide range of aquatic and terrestrial habitats contaminated by increasing anthropogenic activities. Bioremediation is an increasingly popular alternative to conventional methods for treating waste compounds and media with the possibility to degrade contaminants using natural microbial activity mediated by different consortia of microbial strains. Many studies about bioremediation have been reported and the scientific literature has revealed the progressive emergence of various bioremediation techniques. In this review, we discuss the various in situ and ex situ bioremediation techniques and elaborate on the anaerobic digestion technology, phytoremediation, hyperaccumulation, composting and biosorption for their effectiveness in the biotreatment, stabilization and eventually overall remediation of contaminated strata and environments. The review ends with a note on the recent advances genetic engineering and nanotechnology have had in improving bioremediation. Case studies have also been extensively revisited to support the discussions on biosorption of heavy metals, gene probes used in molecular diagnostics, bioremediation studies of contaminants in vadose soils, bioremediation of oil contaminated soils, bioremediation of contaminants from mining sites, air sparging, slurry phase bioremediation, phytoremediation studies for pollutants and heavy metal hyperaccumulators, and vermicomposting.     

Potential of preliminary test methods to predict biodegradation performance of petroleum hydrocarbons in soil

Preliminary tests at different scales such as degradation experiments (laboratory) in shaking flasks, soil columns and lysimeters as well as in situ respiration tests (field) were performed with soil from two hydrocarbon contaminated sites. Tests have been evaluated in terms of their potential to provide information on feasibility, degradation rates and residual concentration of bioremediation in the vadose zone. Sample size, costs and duration increased with experimental scale in the order shaking flasks – soil columns – lysimeter – in situ respiration tests, only time demand of respiration tests was relatively low. First-order rate constants observed in degradation experiments exhibited significant differences between both, different experimental sizes and different soils. Rates were in line with type and history of contamination at the sites, but somewhat overestimated field rates particularly in small scale experiments. All laboratory experiments allowed an estimation of residual concentrations after remediation. In situ respiration tests were found to be an appropriate pre-testing and monitoring tool for bioventing although residual concentrations cannot be predicted from in situ respiration tests. Moreover, this method does not account for potential limitations that might hamper biodegradation in the longer term but only reflects the actual degradation potential when the test is performed.     

Diversity and characterization of sulfate-reducing bacteria in groundwater at a uranium mill tailings site.

Microbially mediated reduction and immobilization of U(VI) to U(IV) plays a role in both natural attenuation and accelerated bioremediation of uranium-contaminated sites. To realize bioremediation potential and accurately predict natural attenuation, it is important to first understand the microbial diversity of such sites. In this paper, the distribution of sulfate-reducing bacteria (SRB) in contaminated groundwater associated with a uranium mill tailings disposal site at Shiprock, N.Mex., was investigated. Two culture-independent analyses were employed: sequencing of clone libraries of PCR-amplified dissimilatory sulfite reductase (DSR) gene fragments and phospholipid fatty acid (PLFA) biomarker analysis. A remarkable diversity among the DSR sequences was revealed, including sequences from delta-Proteobacteria, gram-positive organisms, and the Nitrospira division. PLFA analysis detected at least 52 different mid-chain-branched saturate PLFA and included a high proportion of 10me16:0. Desulfotomaculum and Desulfotomaculum-like sequences were the most dominant DSR genes detected. Those belonging to SRB within delta-Proteobacteria were mainly recovered from low-uranium (< or =302 ppb) samples. One Desulfotomaculum-like sequence cluster overwhelmingly dominated high-U (>1,500 ppb) sites. Logistic regression showed a significant influence of uranium concentration over the dominance of this cluster of sequences (P = 0.0001). This strong association indicates that Desulfotomaculum has remarkable tolerance and adaptation to high levels of uranium and suggests the organism's possible involvement in natural attenuation of uranium. The in situ activity level of Desulfotomaculum in uranium-contaminated environments and its comparison to the activities of other SRB and other functional groups should be an important area for future research.     

In situ bioremediation of contaminated aquifers and subsurface soils

Bioremediation, the use of microorganisms to detoxify and degrade hazardous wastes, is an emerging in situ treatment technology for the remediation of contaminated aquifers and subsurface soils. This technology depends upon the alteration of the physical/chemical conditions in the subsurface environment to optimize microbiological activity. As such, successful bioremediation depends not only upon an understanding of microbial degradation processes, but also upon an understanding of the complex interactions that occur between the contaminants, the subsurface environment, and the indigenous microbial populations at each site. At present, these interactions are poorly understood. Site‐specific evaluation and design therefore are essential for bioremediation. In this paper, we review microbiological, hydrological, and geochemical factors that should be considered in evaluating the appropriateness of bioremediation for hazardous waste‐contaminated aquifers and subsurface soils.     

Benzene Degradation at a Site Amended with Nitrate or Chlorate

Aromatic hydrocarbons are widespread in nature and often contribute to the pollution of soils, sediments, and groundwater. The contamination of soil with mobile aromatic compounds, generally termed BTEX (benzene, toluene, ethylbenzene, xylene) is observed at many industrial sites, especially those associated with the petrochemical industry. In situ bioremediation of sites that are contaminated with BTEX can be applied both aerobically and anaerobically. The use of anaerobic in situ bioremediation is advantageous because supply of oxygen is not needed. Nevertheless, anaerobic in situ bioremediation is less commonly used for BTEX contaminated sites. This paper describes push-pull experiments in order to stimulate the degradation of benzene by the addition of nitrate or chlorate. Deuterated benzene was subjected with nitrate-amended groundwater to the aquifer, and the mineralization was traced by the enrichment of deuterium in the groundwater. Nitrate can be used as electron acceptor, and the addition of nitrate at a site in The Netherlands resulted in partial degradation of benzene. This was demonstrated by comparing various push-pull experiments, benzene concentration measurements, stable isotope analyses of benzene and water, and modeling. Chlorate can be used for the in situ production of oxygen, followed by degradation of benzene with oxygen as electron acceptor. The addition of chlorate at the site resulted in the complete removal of benzene demonstrating a complete degradation within 4 weeks. A pull phase was not needed during this run.     

The development of multiplex simple sequence repeat (SSR) markers to complement distinctness,uniformity and stability testing of rape (Brassica napus L.) varieties

To assess the potential of multiplex SSR markers for testing distinctness, uniformity and stability of rape (Brassica napus L.) varieties, we developed three multiplex SSR sets composed of five markers each. These were used to measure the extent of diversity within and between a set of ten varieties using a fluorescence-based semi-automated detection technology. Also, we evaluated the significance of any correlation between SSRs, pedigree and five of the morphological characters currently used for statutory distinctness, uniformity and stability testing of rape varieties. An assignment test was allowed to identify 99% of the plants examined, with the correct variety based on the analysis of 48 individual plants for each variety. Principal coordinate analysis confirmed that a high degree of separation between varieties could be achieved. Varieties were separated in three groups corresponding to winter, spring and forage types. These results suggested that it should be possible to select a set of markers for obtaining a suitable separation. Diversity within varieties varied considerably, according to the variety and the locus examined. No significant correlation was found between SSR and morphological data. However, genetic distances measured by SSRs were correlated to pedigree. These results suggested that SSRs could be used for pre-screening or grouping of existing and candidate varieties, allowing the number of varieties that need to be grown for comparison to be reduced. Multiplex SSR sets gave high-throughput reproducible results, thus reducing the costs of SSR assessment. Multiplex SSR sets are a promising way forward for complementing the current variety testing system in B. napus.