U.S. EPA Contaminated Site Cleanup Information (CLU-IN)

U.S. Environmental Protection Agency
U.S. EPA Technology Innovation and Field Services Division


Anaerobic Bioremediation (Direct)

Additional Information

Applications and Benefits of Groundwater Recirculation for Electron Donor Delivery and pH-Adjustment during Enhanced Anaerobic Dechlorination
Falatko, D.M., S.A. Fam, and G. Pon.
Proceedings of the Annual International Conference on Soils, Sediments, Water and Energy 16:77-89(2011)

The proper design and implementation of groundwater recirculation for in situ enhanced anaerobic dechlorination of various chlorinated organic compounds is presented with a review of the applicable concepts.

Contaminants in the Subsurface: Source Zone Assessment and Remediation
National Research Council, Committee on Source Removal of Contaminants in the Subsurface. National Academies Press, Washington, DC. ISBN: 030909447X, 383 pp, 2004

After discussing the definition of 'source zone' and the characterization thereof, this report reviews the suite of technologies available for source remediation and their ability to reach a variety of cleanup goals, from meeting regulatory standards for groundwater to reducing costs. The report proposes elements of a protocol for accomplishing source remediation that should enable project managers to decide whether and how to pursue source remediation their sites.

Adobe PDF LogoCost and Performance Report for Bioavailable Ferric Iron (BAFeIII) Assay
Environmental Security Technology Certification Program, ESTCP Project ER-0009, 43 pp, Feb 2007

This report describes the demonstration and validation at four DoD installations of a bioavailable ferric iron (BAFe[III]) assay. BAFe(III) is defined as ferric iron (Fe[III]), a form that is capable of being reduced by microorganisms that oxidize another chemical species and derive energy from the electron transfer. BAFe(III) is an important terminal electron acceptor with significant assimilative capacity in many natural environments. The overall objective of this project was to demonstrate and validate the performance of the BAFe(III) assay as an analytical technology for use in supporting bioremediation. Specific objectives were to validate the BAFe(III) assay method using a combination of confirmatory analyses and to quantify costs associated with the technology.

Dense Nonaqueous Phase Liquids (DNAPLs) Treatment Technologies Bioremediation

This CLU-IN Web page provides a discussion of bioremediation techniques for DNAPL chemicals, most of which are biologically degraded under anaerobic conditions. Included are citations and case studies.

Development of Assessment Tools for Evaluation of the Benefits of DNAPL Source Zone Treatment
Abriola, L.M., P. Goovaerts, K.D. Pennell, and F.E. Loeffler.
SERDP Project ER-1293, 173 pp, 2008

This report details the results of work that has enhanced the understanding of significant mechanisms controlling DNAPL source zone behavior and describes lessons learned that can provide improved DNAPL site management strategies. It discusses 4 important concepts: (1) partial source-zone mass removal can result in substantial local concentration and mass flux reductions; (2) potential remediation efficiency is closely linked to source-zone architecture (ganglia-to-pool ratios); (3) biostimulation and bioaugmentation approaches are feasible for treatment of DNAPL source zones; and (4) the uncertainty in mass discharge ([M/T]) estimates can be quantified through application of geostatistical methods to field measurements.

Adobe PDF LogoDevelopment of a Design Tool for Planning Aqueous Amendment Injection Systems: User's Guide
Borden, R.C. et al.
ESTCP Project ER-0626

A simple spreadsheet-based tool developed to assist in the design of injection-only systems for distributing emulsions or soluble substrate allows quick comparison of the relative costs and performance of different injection alternatives and identification of the design best suited to site-specific conditions. Emulsion Design Tool (2008)Adobe PDF Logo; Soluble Substrate Design Tool (2012) Adobe PDF Logo.

Adobe PDF LogoDraft Technical Protocol: A Treatability Test for Evaluating the Potential Applicability of the Reductive Anaerobic Biological In Situ Treatment Technology (RABITT) to Remediate Chloroethenes
Morse; J.J., B.C. Alleman; J.M. Gossett; S.H. Zinder; D.E. Fennell, Battelle Memorial Inst., Columbus, OH. AFRL-ML-TY-TR-1998-4522, NTIS: ADA352416/XAB, 94 pp, 1998.

This document describes a comprehensive approach for conducting a phased treatability test to determine the potential for employing RABITT at any specific site. It is not meant as a guide for designing either full or pilot-scale in situ biotreatment systems for chlorinated ethenes or any other contaminant. The protocol guides the user through a decision process in which information is collected and evaluated to determine if the technology should be given further consideration. RABITT will be screened out if it is determined that site-specific characteristics, regulatory constraints, or other logistic problems suggest that the technology will be difficult or impossible to employ, or if a competing technology clearly is superior.

Adobe PDF LogoElucidation of the Mechanisms and Environmental Relevance of cis-Dichloroethene and Vinyl Chloride Biodegradation
Cox, E.
SERDP Project ER-1557, 170 pp, 2012

Major results of this project can be summarized as follows: (1) JS666 remains the only isolated organism known to mediate aerobic oxidation of cDCE to CO2, and DNA-based molecular biological tools exist to track its presence and fate during bioaugmentation projects; (2) significant advances were made in understanding the pathway, mechanisms, and enzymes associated with aerobic oxidation of cDCE in JS666; (3) anaerobic oxidation of cDCE and/or VC under iron- or manganese-reducing conditions could not be confirmed, despite substantial efforts with materials from many sites; (4) suspected anaerobic oxidation of VC may in fact be aerobic oxidation to CO2 at extremely low levels of oxygen in the subsurface; and (5) compound-specific isotope fractionation of carbon occurs in both anaerobic and aerobic microbial degradation of ethane, allowing the use of CSIA to assess ethene degradation as a possible means to explain poor ethene mass balance in enhanced in situ bioremediation and MNA projects.

Enhanced Attenuation: A Reference Guide on Approaches to Increase the Natural Treatment Capacity of a System
Early, T., B. Borden, M. Heitkamp, B.B. Looney, D. Major, W.J. Waugh, G. Wein, T. Wiedemeier, K.M. Vangelas, K.M. Adams, and C.H. Sink. WSRC-STI-2006-00083, Revision 1, 161 pp, Aug 2006

This guide covers the following EA approaches: (1) hydraulic manipulation to reduce contaminant infiltration using low-permeability barriers, diffusion barriers, covers, encapsulation, and diversion of electron acceptors; (2) passive residual source reduction (e.g., bioventing); (3) increase in system attenuation capacity via biological processes, such as bioaugmentation, biostimulation, and wetlands development and other plant-based methods; (4) abiotic and biologically mediated abiotic attenuation methods; and (5) reactive barriers.

Adobe PDF LogoGuidance Protocol: Application of Nucleic Acid-Based Tools for Monitoring Monitored Natural Attenuation (MNA), Biostimulation, and Bioaugmentation at Chlorinated Solvent Sites
ESTCP Project ER-0518, 34 pp, 2011

This protocol summarizes the current state of the practice of molecular biological tools (MBTs), specifically nucleic-acid based tools commercially available to identify relevant Dehalococcoides bacteria. It is intended to provide a technically sound and practical approach to MBT use. This document provides recommendations regarding sampling approaches and criteria in evaluation of data for use in bioremediation decision making. See also the Project ER-0518 Final ReportAdobe PDF Logo and the ESTCP Cost and Performance ReportAdobe PDF Logo.

Adobe PDF LogoImpacts of Enhanced Reductive Bioremediation on Post-Remediation Groundwater Quality
Borden, R.C., J.M. Tillotson, G.-H.C. Ng, B.A. Bekins, and D.B. Kent.
SERDP Project ER-2131, 68 pp, 2015

This report presents results from the development of a reactive transport model, secondary water quality impacts (SWQI) database, and indicator simulations that were integrated to develop a general conceptual model of the major processes controlling SWQI production and attenuation during enhanced reductive bioremediation (ERB). The conceptual model can be used as a guide in understanding the magnitude, areal extent, and duration of SWQIs in ERB treatment zones and the natural attenuation of SWQI parameters as the dissolved solutes migrate downgradient with ambient groundwater flow. The model can assist in identifying conditions where SWQIs may pose a concern, e.g., at sites with low iron/high sulfate (H2S mobilization), high groundwater velocity, or low CH4 anaerobic oxidation rates (CH4 migration). Additional resources: Protocol for Evaluating MNA of SWQIAdobe PDF Logo

Adobe PDF LogoIn Situ Bioremediation of Chlorinated Ethene: DNAPL Source Zones
Interstate Technology & Regulatory Council (ITRC), Bioremediation of DNAPLs Team. BioDNAPL-3, 138 pp, June 2008

This publication systematically lays out the technical and related regulatory considerations for in situ bioremediation (ISB) of chlorinated ethene DNAPL source zones, providing information related to site characterization requirements, treatment system application and design criteria, process monitoring, and process optimization. The ability of ISB technology to enhance the dissolution and desorption of nonaqueous-phase contaminants to the aqueous phase, where they can be degraded by the microbial population, depends on the spatial distribution of DNAPL mass in the subsurface (e.g., pool/ganglia ratio) and the ability to deliver amendments throughout this architecture.

In Situ Bioremediation of TCE-Contaminated Groundwater
Travis, B.J. (Los Alamos National Lab., NM); N.D. Rosenberg (Lawrence Livermore National Lab., CA). LA-UR-98-2605, NTIS: DE99001639, 22 pp, 1998

The authors have developed a biokinetics model that includes microbial competition and predation processes. Predator species can feed on the microbial species that degrade contaminants. Simulation studies show that species interactions must be considered when designing in situ bioremediation systems. This report is the final product of a two-year, Laboratory Directed Research and Development (LDRD) project the Los Alamos National Laboratory. The report is available through the DOE Information Bridge.

Adobe PDF LogoIn-Situ Bioremediation of Chlorinated Hydrocarbons: An Assessment of Projects in California
California Department of Toxic Substances Control, Office of Pollution Prevention and Technology Development.
OPPTD Document No. 1217, 163 pp, 2006.

During an evaluation of the performance of in situ bioremediation (ISB) systems at 5 sites in California, the reviewers observed several recurring issues. The project case studies illustrate the reviewers' recommendations for avoiding common ISB problems.

In-Situ Substrate Addition to Create Reactive Zones for Treatment of Chlorinated Aliphatic Hydrocarbons: ESTCP Cost and Performance Report
Environmental Security Technology Certification Program (ESTCP), CU-9920, 93 pp, Mar 2007.

Demonstrations of enhanced reductive dechlorination (ERD) were conducted at two Air Force bases—Vandenberg and Hanscom—to show the ability of this bioremediation approach to dechlorinate TCE plumes in the subsurface over a relatively short time period and to gather information for estimating long-term treatment effectiveness, life span, and costs.

Adobe PDF LogoLoading Rates and Impacts of Substrate Delivery for Enhanced Anaerobic Bioremediation
Henry, B.
ESTCP Project ER-0627, 476 pp, 2010

The author evaluated 15 case studies of different substrates used to stimulate biodegradation of chlorinated compounds: Hydrogen Release Compounds (HRC and HRC-X), vegetable oil (neat and emulsified), whey, molasses, ethanol and lactate, and mulch in permeable biowalls. This report discusses the factors that limit enhanced in situ bioremediation and describes (in Appendix B) a Substrate Design Tool developed in Microsoft Excel to assist the practitioner in evaluating a site for an application of enhanced in situ bioremediation. Substrate Design Tool; ESTCP Cost & Performance ReportAdobe PDF Logo; 2010 AddendumAdobe PDF Logo

Loading Rates and Impacts of Substrate Delivery for Enhanced Anaerobic Bioremediation: Addendum to the Principles and Practices Manual
Henry, B.
Environmental Security Technology Certification Program (ESTCP), Project ER-200627, 39 pp, Jan 2010

Improvements and advances in the enhanced in situ bioremediation of chlorinated solvents have been made since the Tri-Services released Principles and Practices of Enhanced Anaerobic Bioremediation of Chlorinated Solvents in August 2004. This addendum to the 2004 document provides a description of a demonstration study conducted to evaluate substrate loading rates, including a summary of limiting factors and challenges to applying enhanced in situ bioremediation. The demonstration study involved the evaluation of 15 case studies for system design, operation, and performance. Quantitative and qualitative performance objectives were developed to evaluate the case studies and to identify limiting factors for enhanced in situ bioremediation. Supporting information for the case studies can be found in the 2010 Final Technology Demonstration Report. This addendum also summarizes advances made in the field of enhanced in situ bioremediation of chlorinated solvents over the last six years and provides resources and references that can be used to identify and mitigate the limiting factors and challenges that practitioners face when applying the technology.

Adobe PDF LogoManual for Biological Remediation Techniques
International Centre for Soil and Contaminated Sites, 81 pp, 2006

Provides an initial overview of selected organic contaminants, describes their susceptibility to microbial degradation in soil and groundwater, and reviews their treatment potential by land farming, biobeds, bioreactors, bioslurping, bioventing, biosparging, bioscreen, bioaugmentation, and monitored natural attenuation. Monitoring of bioremediation progress is also discussed.

Natural Attenuation and Biodegradation of Contaminants
US Geological Survey Toxic Substances Hydrology Program Bibliography

Provides a bibliographic reference for resources related to bioremediation, including scientific journal publications, conference presentations, agency reports, and others.

Adobe PDF LogoOperation and Analysis of the BEHIVS System at Edwards Air Force Base
McCarty, P.L., S.M. Gorelick, M.N. Goltz, G.D. Hopkins, and F.-J. Eisenberg.
Strategic Environmental Research and Development Program (SERDP). 109 pp, 2003

This report summarizes the results of operation of the bioenhanced in-well vapor stripping (BEHIVS) system at Edwards AFB in 2001, numerical modeling analysis of the results, study conclusions, and recommendations for application of the BEHIVS system at other sites.

Adobe PDF LogoOverview of In Situ Bioremediation of Chlorinated Ethene DNAPL Source Zones
The Interstate Technology & Regulatory Council (ITRC) Bioremediation of DNAPLs Team.
BioDNAPL-1, 89 pp, 2005.

This document presents a technological overview of in situ bioremediation and some of the issues to consider when selecting and designing an in situ bioremediation system for remediation of chlorinated ethene DNAPL source zones. The document provides an overview of chlorinated ethene DNAPLs and in situ bioremediation, technical considerations for in situ bioremediation of chlorinated ethene DNAPL source zones, the state of in situ bioremediation technology applications, and information on defining and measuring system performance of in situ bioremediation applications for chlorinated ethene DNAPL sources.

Petroleum Bioventing
van Eyk, J. and A.A. Balkema, Rotterdam ; Brookfield, VT. ISBN: 9054106867. 302 pp, 1997

This book investigates the composition and the behavior of petroleum in soil, soil properties and soil processes, their interaction with bacterial processes, possibilities for optimizing the removal of petroleum hydrocarbons from soil by bacteria and it explains the phenomenon of recalcitrance.

Principles and Practices of Bioventing
Leeson, A. and R. Hinchee Battelle Memorial Institute. EPA 540-R-95-534a & b [2 vols.] 1995

The manual provides details on bioventing principles; site characterization; field treatability studies; system design, installation, and operation; process monitoring; site closure; and optional technologies to combine with bioventing if warranted. Volume 1 describes the basic principles of bioventing. Volume 2 focuses on bioventing design and process monitoring.

Adobe PDF LogoProcedures for Conducting Bioventing Pilot Tests and Long-Term Monitoring of Bioventing Systems
Downey, D., R. Miller, & T. Dragoo, Parsons Denver, CO. NTIS: ADA423587, 80 pp 2004

This report replaces AFCEE's 1992 'Test Plan and Technical Protocol for a Field Treatability Test for Bioventing' and identifies an updated approach for conducting bioventing pilot tests and monitoring the long-term progress of bioventing systems.

Adobe PDF LogoReductive Anaerobic Biological In-Situ Treatment Technology (RABITT) Treatability Test. Interim Report
Environmental Security Technology Certification Program (ESTCP), Arlington, VA, 16 pp, 2001.

This document presents a summary of the reductive anaerobic biological in situ treatment technology (RABITT) protocol. It also provides the results of treatability tests (both field and microcosm studies) of the RABITT protocol at Cape Canaveral Air Station, FL, Naval Air Station Alameda, CA, Ft. Lewis, WA, and Marine Corps Base, Camp Lejeune, NC.

Adobe PDF LogoA Review of Biofouling Controls for Enhanced In Situ Bioremediation of Groundwater
Environmental Security & Technology Certification Program (ESTCP), Project ER-0429, 55 pp, 2005

The objective of this report is to review well rehabilitation and biofouling controls that are potentially relevant to enhanced in situ bioremediation applications and to identify promising biofouling controls for comparative field evaluation and validation under Environmental Security and Technology Certification Program (ESTCP) Project ER-0429. The report presents a summary of biofouling causes and mechanisms; a discussion of the differences between well rehabilitation and preventative biofouling control, including a review of case studies where biofouling controls have been used in groundwater remediation applications; identification, evaluation and scoring of promising biofouling control options for further field evaluation/validation; and conclusions for preventive biofouling controls.

Soil Bioventing: Principles and Practice
Leeson; A., R.E. Hinchee, and et al. 1997. CRC/Lewis Publishers, Boca Raton, FL.

This book explains in practical terms how to carry out a bioventing program. The book discusses physical and microbial processes affecting bioventing, site characterization activities for implementation of bioventing, system design, performance monitoring, and process evaluation. Case histories of early bioventing studies are discussed as well.

Adobe PDF LogoStandardized Procedures for Use of Nucleic Acid-Based Tools: Recommendations for Groundwater Sampling and Analysis Using qPCR
Lebron, C., P. Dennis, C. Acheson, N. Barros, D. Major, E. Petrovskis, F. Loeffler, K. Ritalahti, C. Yeager, E. Edwards, J. Hatt, and D. Ogles. SERDP Project ER-1561, 12 pp, 2014

SERDP project ER-1561 focused on identifying and minimizing the causes of variability during quantitative real-time polymerase chain reaction (qPCR) enumeration of genes of interest in groundwater, with the goal of developing of the knowledge needed to standardize methods for collecting, preserving, transporting, storing, and processing environmental samples for qPCR analysis. This document summarizes the project conclusions and recommends procedures for using qPCR analyses that will provide data of sufficient accuracy and reproducibility to allow site management decisions regarding bioremediation or MNA. Further details are available in the ER-1561 Final ReportAdobe PDF Logo (Lebron et al. 2014, 220 pages).

Adobe PDF LogoStrategies for Monitoring the Performance of DNAPL Source Zone Remedies
Interstate Technology and Regulatory Council (ITRC) Dense Nonaqueous-Phase Liquids Team. DNAPLs-5, 206 pp., Aug 2004

This document is intended for regulators and others interested in learning about approaches to performance monitoring while implementing various in situ technologies for the treatment of DNAPLs. In this document, we present a number of ways in which the success or failure in treating a DNAPL source zone has been measured. Because the vast majority of experience in DNAPL source zone remediation has been in unconsolidated geologies, such as sands and silts, many of the conclusions, recommendations, and lessons learned presented in this document do not necessarily transfer to performance assessment in fractured bedrock, karst, or other consolidated geologies.

Adobe PDF LogoA Systematic Approach to In Situ Bioremediation in Groundwater, Including Decision Trees for In Situ Bioremediation of Nitrates, Carbon Tetrachloride, and Perchlorate
Interstate Technology and Regulatory Council. ITRC ISB-8, 158 pp, 2002

This document provides guidance for the systematic characterization, evaluation, and appropriate design and testing of in situ bioremediation for any biotreatable contaminant. It includes information on considerations for a bioremediation program, including site background, hydrogeology/geochemistry, contaminant fate and transport, and limitations. The document also provides information on systematic approaches to in situ bioremediation of nitrate, carbon tetrachloride, and perchlorate.

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