Cellular Autometa Matlab algorithm

13 In Situ: Groundwater Bioremediation

K. N. Timm # Springer

T. C. Hazen Lawrence Berkeley National Laboratory, Berkeley, CA, USA

[email protected]

1

Introduction ... . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2584

2

Terminology .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2584

3

Characterization and Monitoring Feasibility ... . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2585

4

Biostimulation and Bioaugmentation of Groundwater ... . . . . . . . . . . . . . . . . . . . . . . . . . 2589

5

Intrinsic Bioremediation and Modeling ... . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2592

6

Research Needs .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2594

is (ed.), Handbook of Hydrocarbon and Lipid Microbiology, DOI 10.1007/978-3-540-77587-4_191,

-Verlag Berlin Heidelberg, 2010

2584 13 In Situ: Groundwater Bioremediation

Abstract: In situ groundwater bioremediation of hydrocarbons has been used for more than

40 years. Most strategies involve biostimulation; however, recently bioaugmentation have been

used for dehalorespiration. Aquifer and contaminant profiles are critical to determining the

feasibility and strategy for in situ groundwater bioremediation. Hydraulic conductivity and

redox conditions, including concentrations of terminal electron acceptors are critical to

determine the feasibility and strategy for potential bioremediation applications. Conceptual

models followed by characterization and subsequent numerical models are critical for efficient

and cost effective bioremediation. Critical research needs in this area include better modeling

and integration of remediation strategies with natural attenuation.

1 Introduction

A patent for in situ bioremediation of groundwater contaminated with gasoline by stimulating

indigenous bacteria via nutrient injection into the terrestrial subsurface was issued to Dick

Raymond in 1974 (US Patent 3,846,290). He successfully demonstrated this technology and

began commercial applications in 1972 (Raymond et al., 1977). Clearly in situ groundwater

bioremediation has been used successfully for more than 50 years and much is understood

about where it is applicable, especially for petroleum contaminants. The really new bioreme-

diation applications that have been done in the last 20 years are in the area of solvent, PAH,

PCB, dioxin, MTBE, and metals. Bioremediation has been around for a long time, only its

application breadth in terms of types of contaminants and environments has increased in the

last 20 years. This explosive proliferation of new applications and environments in the last

20 years, especially by companies trying to establish themselves with a proprietary edge, has

lead to a large number of terms, many of which are highly redundant, in what they try to

uniquely describe. Also, the bioremediation field applications that have been reported, fre-

quently lack comprehensive field data, especially in the terrestrial subsurface. Though biore-

mediation has been used at a large number of sites these applications were nearly all done

by companies trying to do the study for (1) clients, who usually wanted to remain confidential,

(2) the least possible cost to the client and the vendor, and (3) protecting the vendors

proprietary edge for their product. This has lead to a paucity of peer-reviewed data, miss

application of terminology, and confusion as to what some terms mean. More importantly it

has also lead to many ‘‘failures’’ of in situ groundwater bioremediation due to a lack of

fundamental understanding of requirements, and limitations, in terms of hydrology, geology,

and biogeochemistry at various scales.

2 Terminology

Biological Treatment – Any treatment process that involves organisms or their products, e.g.,

enzymes.

Biotransformation – A biological treatment process that involves changing the contami-

nant, e.g., valence states of metals, chemical structure, etc.

Intrinsic Bioremediation – Unmanipulated, unstimulated, unenhanced biological reme-

diation of an environment; i.e., biological natural attenuation of contaminants in the

environment.

In Situ: Groundwater Bioremediation 13 2585

Engineered Bioremediation – Any type of manipulated or stimulated or enhanced

biological remediation of an environment.

Biostimulation – The addition of organic or inorganic compounds to cause indigenous

organisms to effect remediation of the environment, e.g., fertilizer.

Bioaugmentation – The addition of organisms to effect remediation of the environment,

e.g., contaminant-degrading bacteria injection into an aquifer.

Biosparging – Injection of air or specific gases below ground, usually into saturated

sediments (aquifer material) to increase biological rates of remediation.

Bioslurping – This treatment combines soil vapor extraction with removal of light non-

aqueous phase liquid contaminants from the surface of the groundwater table, thereby

enhancing biological treatment of the unsaturated zone and the groundwater, especially the

capillary fringe zone where hydrocarbons tend to smear.

Biofilters – Normally used to refer to treatment of gases by passing through a support

material containing organisms, e.g., soil, compost, trickle filter. Sometimes used to refer to

treatment of groundwater via passage through a biologically active area in the subsurface.

Biocurtain – The process of creating a subsurface area of high biological activity to contain

or remediate, usually in aquifer material.

Bioremoval – A biological treatment involving uptake of the contaminant from the

environment by an organism or its agent.

Bioimmobilization – A biological treatment process that involves sequestering the contam-

inant in the environment. No biodegradation of the contaminant, e.g., metal bioreduction.

Biomobilization – A biological treatment process that involves making the contaminant

more mobile in the environment. No biodegradation of the contaminant, but usually requires

removal of the contaminant.

Permeable Reactive Barrier (PRBs) – are often referred to as iron filing walls, reactive

barriers, funnel and gate systems, or passive treatment walls. They are constructed under-

ground to intercept groundwater flows and to provide preferential flow paths through

bioreactive materials, e.g., as groundwater moves through the bioreactive materials, contami-

nants are treated and transformed into harmless by-products.

3 Characterization and Monitoring Feasibility

The success of any bioremediation application will be highly dependent on the characteriza-

tion and monitoring that is done before and during the field deployment. For any field

remediation, the first step is to form a conceptual model of the contaminant plume in the

environment and how that environment effects that plume. The uncertainties in this concep-

tual model provide the drivers for the characterization and monitoring needs. For example,

characteristics of the aquifer will have a profound impact on the remediation strategy

(> Fig. 1). The largest part of the expense of any remediation project is the characterization

and monitoring. Hydraulic conductivities can have a severe effect on your ability to deliver

nutrients to the subsurface (> Fig. 2) and can be the most limiting part of the environment.

Fortunately, new advances in geophysics and hydraulic push technology (Geoprobe) has

enabled us to characterize sites in a fraction of the time and cost. Once we have established

the hydrology and basic geochemistry at the site and used that data to refine our conceptual

model, a base line characterization of the microbiology is essential to establish that the right

. Figure 1 Aquifer and contaminant characteristics.

2586 13 In Situ: Groundwater Bioremediation

microorganisms are present, that they can be stimulated, and that no undesirable reactions

with the stimulants or daughter products from the stimulation will occur. This usually

requires some treatability and soil compatibility studies and monitoring of microbial com-

munity structure and function to establish the base conditions prior to stimulation (Plaza

et al., 2001). For example, some metals like arsenic actually increase solubility under the same

redox potentials that precipitate Cr and U. > Table 1 provides an example list of the types of

measurements that should be performed from either treatability slurries, soil columns or in

situ sampling (Hazen, 1997). This data and the refined conceptual model provide the

functional design criteria for the remediation and can be used to develop a numerical

model to predict the remediation rates, stability, and legacy management needs, e.g., moni-

toring, especially if the remediation is an immobilization strategy.

Bioremediation strategies will be limited most by our ability to deliver the stimulus to

the environment. The permeability of the formation must be sufficient to allow perfusion of

the nutrients and/or microorganisms through the formation. The minimum average hydraulic

. Figure 2 Hydraulic conductivity.

In Situ: Groundwater Bioremediation 13 2587

conductivity for a formation is generally considered to be 10 �4

cm s �1

(Thomas and Ward,

1989). Additionally, the stimulants required must be compatible with the environment. For

example, hydrogen peroxide is an excellent source of oxygen, but it can cause precipitation of

metals in soils, and such dense microbial growth around the injection site that all soil pores are

plugged. It is also toxic to bacteria at high concentrations, >100 ppm (Thomas and Ward, 1989).

Ammonia also can be problematic, because it adsorbs rapidly to clays, causes pH changes in

poorly buffered environments, and can cause clays to swell, decreasing permeability around

the injection point. It is generally accepted that soil bacteria need a C:N:P ratio of 30:5:1 for

unrestricted growth (Paul and Clark, 1989). The actual injection ratio used is usually slightly

higher (a ratio of 100:10:2) (Litchfield, 1993), since these nutrients must be bioavailable, a

condition that is much more difficult to measure and control in the terrestrial subsurface. It

may also be necessary to remove light nonaqueous phase liquid (LNAPL) contaminants that

are floating on the water table or smearing the capillary fringe zone, hence bioslurping (Keet,

1995). This strategy greatly increases the biostimulation response time by lowering the highest

concentration of contaminant the organisms are forced to transform.

Recent advances in geophysics are now enabling us to determine aquifer heterogeneity,

hydraulic conductivity, amendment movement in the subsurface, changes in biogeochemistry,

and real-time monitoring of changes (> Fig. 3). These measurements can potentially save

time, expense, and increase our resolution of biogeochemical changes, hydrology, contaminant

inventory, and amendment injection pathway (Faybishenko et al., 2008; Hubbard et al., 2008).

The type of sample used for monitoring and characterization of groundwater can have a

significant impact on a bioremediation project. Hazen et al. (1991) demonstrated that deep

oligotrophic aquifers have dense attached communities of bacteria that are not reflected in the

groundwater from that aquifer. This has serious implications for the in situ bioremediation of

deep contaminated aquifers, since monitoring of groundwater is the principal method used

to characterize and control biodegradation by indigenous bacteria stimulated by nutrient

infiltration. Groundwater monitoring may not indicate community or population numbers,

or physiological activity of the sediment attached microbes, the principal biologically active

. Table 1 Characterization and monitoring parameters

Measurements Parameter

Biomass

Viable counts Plate counts, Most Probable Number (MPN), enrichments, BIOLOGTM

Direct counts Acridine Orange Direct Count (AODC), Fluorescine Isothiocyanate (FITC),

Direct Fluorescent Antibody (DFA)

Signature compounds Phospholipid Fatty Acid (PLFA), DNA, RNA,qPCR, phylochips, functional

gene arrays

Bioactivity and

bioremediation

Daughter products Cl, CO2, CH4, stable isotopic C, reduced contaminants, stable isotopic

fractionation of contaminants

Intermediary

metabolites

Epoxides, reduced contaminants

Signature compounds PLFA, ribosome probes, BIOLOGTM, phosphatase, dehydogenase,

lodophenylNitrophenyal, Tetrazolium Chloride (INT), acetylene

reduction, recalcitrant contaminants

Electron acceptors O2, NO3, SO4, Fe(III), CO2

Conservative tracers He, CH4, Cl, Br

Radiolabeled

mineralization

14C, 3H – labeled contaminants, acetate, thyamine

Sediment

Nutrients PO4, NO3, NH4, O2, total organics, SO4

Physical/chemical Porosity, lithology, cationic exchange, redox potential, pH, temperature,

moisture, heavymetals

Toxicity MicrotoxTM, Mutatox TM

2588 13 In Situ: Groundwater Bioremediation

component of these aquifers. Harvey et al. (1984) and Harvey and George (1987) have shown

that shallow, eutrophic, rapidly moving aquifers, behave quite differently, in that there are no

significant differences between groundwater and attached sediment communities. This is

reasonable because attachment in such an environment would have no significant advantage,

unlike the oligotrophic deep aquifers. Enzien et al. (1994) further underscored the need for

careful sampling when they showed significant anaerobic reductive dechlorination processes

occurring in an aquifer whose bulk groundwater was aerobic (>2 mg l �1

O2).

The state and fate of contaminants in all environments is highly dependent on the redox or

valence state of the environment. The redox potential of the environment will control the

direction of chemical equilibria and whether the contaminant is reduced or oxidized. This in

turn controls the possible compounds that the contaminant can form and the relative

solubility of these metals in the environment. To stimulate microbes to produce conditions

that are appropriate for remediation of specific contaminants requires a through knowledge of

the geochemistry of that environment. Since electron acceptors vary greatly as to the energy

that can be derived from their use in respiration, the most common terminal electron

acceptors (TEA) will be utilized in a set order, according to the energy that can be derived

. Figure 3 Geophysical measurements of polylactate injection for groundwater bioremediation.

In Situ: Groundwater Bioremediation 13 2589

(> Fig. 4). Thus, oxygen is the preferred TEA and first TEA to be utilized, followed by nitrate,

iron (III), sulfate, and carbon dioxide. Since dehalorespiration is not favored until the redox

potential is in methanogenic conditions, O2, NO3, Fe(III), and SO4 would have to be depleted

first. Indeed, for sites that also have PCE/TCE the iron (III) and the sulfate would have to be

depleted before sustained methanogenesis and subsequently dehalorespiration can occur. For

field applications, this means that enough electron donor would have to be added to deplete all

the oxygen and nitrate present, at a minimum. By monitoring the TEA and their daughter

products, it provides an excellent measure of the redox conditions at the site and the potential

for degradation of the contaminants of concern (Nelson et al., 1994).

4 Biostimulation and Bioaugmentation of Groundwater

All engineered bioremediation can be characterized as either biostimulation, i.e., the addition

of nutrients, or bioaugmentation, i.e., the addition of organisms, or processes that use both.

The problems with adding chemical nutrients to sediment and groundwater are fundamen-

tally different from those of adding organisms. Simple infiltration of soil and subsequently

groundwater is physically quite different in the two processes (Alfoldi, 1988). Even the smallest

bacterium has different adsorption properties from chemicals. For example, clayey soils have

very low porosity and may not physically allow bacteria to penetrate. These clays may also bind

the microbes that are added, e.g., cationic bridges involving divalent metals and the net

negative charge on the surface of the bacteria and the surface of the clay. In some soils,

. Figure 4 Critical biogeochemistry involving terminal electron acceptors and their heiarchical redox

potential relationships.

2590 13 In Situ: Groundwater Bioremediation

inorganic chemicals that are injected may precipitate metals, swell clays, change redox

potentials, and conductivity, thus having a profound effect on groundwater flow and biogeo-

chemistry of the environment. Indeed, bacterial plugging of subsurface formations has been

successfully used for enhanced oil recovery in oil reservoirs (Cusack et al., 1992).

Biostimulation is dependent on the indigenous organisms and thus requires that they be

present and that the environment be capable of being altered in a way that will have the desired

bioremediation effect (> Fig. 5). In most terrestrial subsurface environments, the indigenous

organisms have been exposed to the contaminant for extended periods of time and have

adapted or even naturally selected. Many contaminants, especially organic compounds are

naturally occurring or have natural analogs in the environment. Rarely can a terrestrial

subsurface environment be found that does not have a number of organisms already present

that can degrade or transform any contaminant present. Indeed, even pristine environments

have bacteria with an increasing number of plasmids with sediment depth in response to

increasing recalcitrance of the organics present (Fredrickson et al., 1988).

Oxygen is quite often limiting since the contaminant can be used as a carbon and energy

source by the organisms and the contaminant concentration greatly exceeds the oxygen input

needed by the organisms. Introduction of air, oxygen, or hydrogen peroxide via infiltration

galleries, tilling, sparging, or venting have proven to be extremely effective in bioremediating

petroleum contaminants and a variety of other organic compounds that are not particularly

recalcitrant (Thomas and Ward, 1992). However, if the environment has been anaerobic for

. Figure 5 Biostimulation versus bioaugmentation strategy requirements.

In Situ: Groundwater Bioremediation 13 2591

extended periods of time and the contaminant has a high carbon content, it is likely that

denitrification has reduced the overall nitrogen content of the environment making this

nutrient limiting. Nitrogen has been successfully introduced into the terrestrial subsurface

for biostimulation using ammonia, nitrate, urea, and nitrous oxide (USEPA, 1989). Phospho-

rus is naturally quite low in most environments and, in terrestrial subsurface environments,

even if phosphorus concentrations are high it may be in a mineral form that is biologically

unavailable, e.g., apatite. Several inorganic and organic forms of phosphate have been success-

fully used to biostimulate contaminated environments (USEPA, 1989). In environments

where the contaminant is not a good carbon or energy source and other sources of carbon

or energy are absent or unavailable, it will be necessary to add an additional source of carbon

(Horvath, 1972). An additional source of organic carbon will also be required if the total

organic carbon concentration in the environment falls below 1 ppm and the contaminant

cleanup levels have still not been met. Methane, methanol, acetate, molasses, sugars, agricul-

tural compost, phenol, and toluene have all been added as secondary carbon supplements to

the terrestrial subsurface to stimulate bioremediation (National Research Council, 1993).

Bioaugmentation may provide significant advantages over biostimulation for (1) environ-

ments where the indigenous bacteria have not had time to adapt to the contaminant, (2)

particularly recalcitrant contaminants that only a very limited number of organisms are

capable of transforming or degrading, (3) environments that do not allow a critical biomass

to establish and maintain itself, (4) applications where the desired goal is to plug the forma-

tion for contaminant containment, e.g., biocurtain, and (5) controlled environments where

2592 13 In Situ: Groundwater Bioremediation

specific inocula of high rate degraders will greatly enhance the process, e.g., permeable reactive

barriers. Like biostimulation, a major factor effecting the use of bioaugmentation in the

terrestrial subsurface is hydraulic conductivity. The 10 �4

cm s �1

limit for biostimulation

will need to be an order of magnitude higher for bioaugmentation and may need to be higher

yet, depending on the size and adherence properties of the organism being applied (Baker and

Herson, 1990; Ginn et al., 2002). Studies have shown the less adherent strains of some

contaminant-degraders can be produced, allowing better formation penetration (DeFlaun

et al., 1994; Johnson et al., 2001). However, the ability to rapidly clog a formation is a

significant advantage of bioaugmentation in applications where containment is a primary

goal. The oil industry has been using this strategy to plug fluid loss zones and enhance oil

recovery for a number of years (Cusack et al., 1992).

A number of novel organisms have been successfully injected into the subsurface for in

situ bioremediation of PCBs, chlorinated solvents, PAHs, and creosote (National Research

Council, 1993). Bioaugmentation suffers the dilemma of being indistinguishable from bios-

timulation in many environments, since nutrients are often injected with the organisms and

since dead organisms are an excellent source of nutrients for most indigenous organisms. For

many applications it is difficult, if not impossible, to determine if the added organisms

provided a significant advantage over nutrient stimulation alone. Given the problems and

high cost of producing the organisms for inoculation and delivery problems, bioaugmentation

applications will probably remain limited. For example, if dehalorespiration was the strategy

and the site had a hydraulic conductivity of only 10 �8

cm s �1

with very high nitrate and sulfate

levels and high pH it may not be cost effective to use dehalorespiration at this site. These issues

also suggest why bioaugmentation has not lived up to its hope. Though bioaugmentation

promises ‘‘designer biodegraders,’’ it has not proven to be better then biostimulation in

repeated field trials over the last 2 decades. Indeed, there is only one bacterium that has

demonstrated that it can perform better then biostimulation in situ on most occasions,

Dehalococcoides ethenogenes for dehalorespiration of chlorinated solvents. Several products

are commercially available and have been widely used that are proprietary strains of this

organism (e.g., Regenesis and Geosyntec). We suspect the reason that this microbe has been

successful is that it is a strict anaerobe, chlorinated solvent dehalorespiration requires established

methanogenic redox potentials, and the organism is very small irregular coccus (0.5 m) so it can

penetrate the subsurface more easily (Loffler et al., 2000). Patchy distributions of this organism

in nature are also common, so bioaugmentation may provide a couple of advantages.

Bioaugmentation may also have a very significant advantage when genetically engineered

microorganisms (GEMs) are used. It is possible that a GEM could be constructed with unique

combinations of enzymes to facilitate a sequential biotransformation or biodegradation of a

contaminant. This would be particularly helpful for contaminants that are extremely recalci-

trant, e.g., PCBs, or under limited conditions, e.g., tetrachloroethylene and carbon tetrachlo-

ride can only be biodegraded anaerobically. In addition, this GEM could be modified with

unique survival or adherence properties that would make it better suited to the environment

where it was to be applied.

5 Intrinsic Bioremediation and Modeling

Intrinsic bioremediation is developing rapidly as an important alternative for many contami-

nated environments. This strategy of natural attenuation by thorough characterization,

In Situ: Groundwater Bioremediation 13 2593

treatability studies, risk assessment, modeling, and verification monitoring of contaminated

environments was first proposed by John Wilson of EPA’s Kerr Lab in the early 1990s. Wilson

organized the first Symposium on Intrinsic Bioremediation in August, 1994, and develop-

ment and regulatory acceptance has been exponential ever since. Certainly, much of this

rapid deployment of intrinsic bioremediation has been due to the crushing financial burden

that environmental cleanup represents and our need to use more risk-based cleanup goals for

the thousands of new contaminated sites identified every year. Intrinsic bioremediation as

a strategy carries with it a burden of proof of: (1) risk to health and the environment, and

(2) a model that will accurately predict the unengineered bioremediation of the environment.

Thus applications of intrinsic bioremediation have been confined to environments with few

risk receptors, containing contaminants with relatively low toxicity, e.g., petroleum in fairly

homogeneous, confined, and predictable subsurface environments. The EPA reported that in

1995 intrinsic bioremediation was already in use at 29,038 leaking underground petroleum

storage tank (LUST) sites in 33 states (Tremblay et al., 1995). This represents 28% of the

103,479 LUST sites being remediated in 1995 and an increase of more than 100% since 1993.

Intrinsic bioremediation has also been implemented at a creosote-contaminated methano-

genic aquifer in Florida (Bekins et al., 1993) and in three TCE-contaminated, reducing

aquifers (Major et al., 1994; Martin and Imbrigotta, 1994; Wilson et al., 1994).

The coupling of intrinsic bioremediation to engineered bioremediation could be the best

overall solution. Nearly all engineered bioremediation projects could substantially reduce costs

by stopping the biostimulation or bioaugmentation process early and allowing intrinsic

bioremediation to finish the cleanup process. The only projects that would not benefit from

such a strategy would be those where immediate risk to health and the environment demanded

an emergency response. Intrinsic bioremediation has the same requirements for treatability,

modeling, characterization, and modeling as engineered bioremediation discussed above. The

only difference is that a greater emphasis is put on risk assessment, predictive modeling, and

verification monitoring. Once an intrinsic bioremediation project has been started, verifica-

tion monitoring of the predictive model is initially quite rigorous. Afterwards, if the model

holds true, monitoring frequency and numbers of parameters gradually decline until the site is

cleaned up.

Modeling of the bioremediation process has become increasingly important in determin-

ing the fate and effect of contaminants and predicting the outcome of different amendment

scenarios. The models will only be as good as the data they receive from the characterization

studies and the treatability studies. However, models can also be used to suggest treatability

studies that should be performed from a minimum of characterization data. The simple

kinetic models using Monod or Michaelis-Menten functions of 15 years ago are completely

inadequate for current bioremediation applications in the terrestrial subsurface. One and two-

dimensional models of aerobic biodegradation of organic contaminants in ground water did

not appear until quite recently (Molz et al., 1986; Widdowson et al., 1987). These models used

advective and dispersive transport coupled with an assumption of microcolonies. Widdowson

et al. (1988) later added nitrate respiration as an option to their model. Perhaps the best

documented and most widely used model for bioremediation has been the BIOPLUME model

(Borden and Bedient, 1986). This model, now in its forth version, uses a series of simultaneous

equations to simulate growth, decay, and transport of microorganisms, oxygen, and hydro-

carbons. Rifai et al. (1987) later modified this model (BIOPLUME II) to incorporate the

USGS two-dimensional method of characteristic model (Konikow and Bredehoeft, 1978).

The original model was used to simulate PAH biodegradation at a Texas Superfund site

2594 13 In Situ: Groundwater Bioremediation

(Borden and Bedient, 1986). BIOPLUME II has been used to model biodegradation of aviation

fuel at the US Coast Guard Station at Traverse City, Michigan (Rifai et al., 1988), and to

characterize benzene biodegradation over 3 years in another shallow aquifer (Chiang et al.,

1989; Choi et al., 2009). Travis and Rosenberg (1994) used a numerical simulation model to

successfully predict aerobic bioremediation of chlorinated solvents in the groundwater and

vadose zone using methane biostimulation at the US DOE’s Savannah River Site near Aiken,

South Carolina. Their model also used a series of simultaneous equations for microbial

growth, nutrient limitations, and contaminant, microbe, and nutrient transport. The model

predicted the amount of TCE that was biodegraded during a 14-month, full-scale demonstra-

tion, and was validated by five other methods (Hazen et al., 1994). Models like these are

becoming increasingly important as our need to understand the terrestrial subsurface ‘‘black

box’’ of bioremediation increases in response to increased emphasis on intrinsic bioremedia-

tion as a solution. These types of models, along with rigorous treatability studies, are required

for intrinsic bioremediation to be acceptable, particularly as a solution for bioremediation of

terrestrial subsurface environments.

6 Research Needs

There are a large number of ex situ and in situ bioremediation methods currently available.

Ex situ methods have been around longer and are better understood, and they are easier to

contain, monitor, and control. However, in situ bioremediation has several advantages over

ex situ techniques. In situ treatment is useful for contaminants that are widely dispersed in

the environment, present in dilute concentrations, or otherwise inaccessible (e.g., due to the

presence of buildings or structures). This approach can be less costly and less disruptive than

ex situ treatments because no pumping or excavation is required. Moreover, exposure of site

workers to hazardous contaminants during in situ treatment is minimal. Broadly, bioremedi-

ation strategies can be further divided into natural attenuation, biostimulation, and bioaug-

mentation strategies. Bioaugmentation being the most aggressive, since organisms are added

to the contaminated environment. Biostimulation can be aggressive or passive, in that electron

donors, electron acceptors, and trace nutrients can be injected into the environment to

stimulate indigenous organisms to increase biomass or activity to affect the contaminant.

Passive biostimulation techniques include simple infiltration galleries. Natural attenuation

relies on the intrinsic bioremediation capabilities of that environment. Environments high in

organic carbon and energy sources, low contaminant concentrations, and without significant

nutrient deficiencies may be able to degrade or transform the contaminants of concern

without any intervention. Ideally, the most cost effective and efficient approach to treat

most large contaminant plumes is to use more aggressive approaches, e.g., bioaugmentation

or even excavation and removal, at the source, grading into natural attenuation at the leading

edge, or over time as the contaminant concentration declines. There are only a few bioaug-

mentation candidates for in situ groundwater bioremediation (Dehalococcoides ethenogenes);

however, it is technically possible to use bacteriophage as vectors to provide indigenous

bacteria with increases or new degradation capacity. The size of bacteriophages and

their specificity overcomes the inherent problem particle injection in the subsurface and

the minimizing nontarget effects. Much more research is needed in this area. Rarely is a single

remediation approach completely effective or cost efficient. Indeed, combining aggressive

physical and chemical treatment techniques like chemical oxidation/reduction, thermal

In Situ: Groundwater Bioremediation 13 2595

desorption with bioremediation can provide advantages to some types of contaminants and

allows bioremediation to be an effective polishing or sentinel strategy for the cleanup. Much

more modeling at all scales (Lee and Schwartz, 2007) using a systems biology approach is

needed to find the fastest, most efficient, and lowest life-cycle cost solution for contaminated

groundwater.

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