Bioremediation for Marine Oil Spills

May 1991

OTA-BP-O-70

NTIS order #PB91-186197
Recommended Citation:

U.S. Congress, Office of Technology Assessment, Bioremediation for Marine Oil Spills—

Background Paper, OTA-BP-O-70 (Washington, DC: U.S. Government Printing Office, May

1991).

For sale by the Superintendent of Documents

U.S. Government Printing Office, Washington, DC 20402-9325

(order form can be found in the back of this report)
Foreword

If anything positive has resulted from the massive Exxon Valdez oil spill, it is that this

environmental calamity increased the Nation’s awareness of the shortcomings of its capability

to fight oil spills and prompted it to take steps to correct this situation. In the 2 years since this

spill occurred, Congress passed major oil pollution legislation (the Oil Pollution Act of 1990);

private industry established and provided significant funding for the Marine Spill Response

Corp.; and Federal agencies reevaluated their responsibilities, revised contingency plans, and

took steps to improve response technologies.

In addition to concern with refining the efficiency and reliability of the existing response

technologies, both the public and private sectors have sought to develop innovative new

technologies for responding to spills. bioremediation is one such technology. Although the

possibility of using the capabilities of oil-degrading microorganisms to accelerate the natural

biodegradation of oil has been discussed for years, it is only recently that some of the practical

problems associated with this idea have begun to be addressed. The Exxon Valdez spill, in

particular, gave researchers a rare opportunity to evaluate the feasibility of using bioremediation as an oil spill countermeasure.

This OTA background paper evaluates the current state of knowledge and assesses the

potential of bioremediation for responding to marine oil spills. Our basic message is a dual

one: we caution that there are still many uncertainties about the use of bioremediation as a

practical oil spill response technology; nevertheless, it could be appropriate in certain

circumstances, and further research and development of bioremediation technologies could

lead to enhancing the Nation’s capability to fight marine oil spills. The request for this study

came from Senator Ted Stevens, a member of OTA’s Technology Assessment Board, and the

senior senator from the State of Alaska, where bioremediation was tested on beaches polluted

by oil from the Exxon Valdez.

w Directo r

. . . Ill
OTA Project Staff

bioremediation for Marine Oil Spills

John Andelin, Assistant Director, OTA

Science, Information, and Natural Resources Division

Robert W. Niblock, Oceans and Environment Program Manager

William E. Westermeyer, Project Director

Contributor

Florence Poillon, Editor

Administrative Staff

Kathleen Beil

Sally Van Aller
Contents

Page

INTRODUCTION . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 1

SUMMARY AND FIlNDINGS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2

BACKGROUND . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 4

The Fate of Oil in the Marine Environment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 5

Biodegradation and the Chemical Nature of Petroleum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 7

Microbial Processes and the Degradation of Petroleum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9

Environmental Influences on Biodegradation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 10

General Advantages and Disadvantages of bioremediation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12

ALTERNATIVE bioremediation TECHNOLOGIES . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13

Nutrient Enrichment . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .13

Seeding With Naturally Occurring Microorganisms . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 14

Seeding With Genetically Engineered Microorganisms . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18

ENVIRONMENTAL AND HEALTH ISSUES . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 18

BIOREMEDIATION IN RELATION TO OTHER RESPONSE TECHNOLOGIES . . . . . . . . . 20

bioremediation ACTIVITIES IN THE PUBLIC AND PRIVATE SECTORS . . . . . . . . . 23

The Environmental Protection Agency . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 23

NETAC and Commercialization of Innovative Technologies . . . . . . . . . . . . . . . . . . . . . . . . . . . . 26

The Private Sector . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . , 27

REGULATORY ISSUES . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 28

APPENDIX A: GLOSSARY . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 29

APPENDIX B: ACKNOWLEDGMENTS . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 30

Boxes

Box Page

A. bioremediation v. Biodegradation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2

B. The Alaska bioremediation Experiments . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 15

Figures

Figure Page

l. Schematic of Physical, Chemical, and Biological Processes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 6

2. organization of the bioremediation Action Committee . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 24

Tables

Table Page

l. Major Genera of Oil-Degrading Bacteria and Fungi . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 9

2. bioremediation: Potential Advantages and Disadvantages . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 12

3. Principal Features of Alternative bioremediation Approaches . . . . . . . . . . . . . . . . . . . . . . . . . . . . 13

4. Key Research Needs . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 26
INTRODUCTION

In March 1990 the Office of Technology Assessment (OTA) published Coping With an Oiled Sea:

An Analysis of Oil Spill Response Technologies. The

study was prompted in part by the alarm and concern

that followed the Nation’s largest oil spill to date, the

n-million-gallon spill in Prince William Sound,

Alaska, caused by the grounding of the Exxon

Valdez. OTA concluded that if the damage from

such pollution incidents is to be minimized in the

future, major improvements are required in the

organization and the technologies employed to fight

oil spills. OTA evaluated the state-of-the-art and the

potential for improvement of the most widespread

technologies-mechanical containment and cleanup

methods-and assessed what appeared to be the

most promising alternatives to mechanical methodsin particular, the use of chemical dispersants and

in situ burning.

One “new” technology about which OTA had

little to say in its initial study was bioremediation—

the use of microorganisms to accelerate the degradation of oil or other environmental contaminants (box

A). Degradation of oil by microorganisms is one of

the most important long-term natural processes for

removal of oil from the marine environment.l Given

enough time-at least several years, for example, for

oil stranded on beaches-some microorganisms are

capable of at least partially cleaning environments

polluted with oil. Because bioremediation is a

potentially significant method for mitigating the

damage caused by marine oil spills,2 techniques to

accelerate and improve the efficiency of this natural

process have had a number of proponents over the

past two decades. Although considerable research

has been conducted in the last 10 years on the

development of bioremediation techniques, imporant questions remain about their effectiveness,

possible unintended side effects of their use, and

their importance in comparison with more conventional oil spill response technologies. Research

concerning these issues has been given new momentum, in large part because the Exxon Valdez accident

stimulated a general search for more effective

methods to fight oil spills. Consequently, the data

that those directly responsible for fighting oil

spills--e. g., on-scene coordinators-will need before they are willing or able to include biological

techniques in their arsenal of countermeasures are

beginning to be developed.

This study examines the potential of bioremediation technologies to clean up marine oil spills and to

minimize the damage they cause. Thus, the study

evaluates a small, but highly visible, subset of the

many possible applications of bioremediation technologies to environmental problems. Among the

other applications for which bioremediation is being

considered or is currently in use are: 1) treatment of

nontoxic liquid and solid wastes, 2) treatment of

toxic or hazardous wastes, 3) treatment of contaminated groundwater, and 4) grease decomposition.3

Although recent marine oil spills and bioremediation efforts have called attention to the potential of

bioremediation as an oil spill response technology,

some of these other applications, in particular the

treatment of hazardous waste, appear to have greater

potential. Officials at approximately 135 hazardous

waste sites, for example, are now either considering,

plannin g, or operating full-scale bioremediation

systems.4

Consideration of the applicability of bioremediation to oil spills is not new. Investigation of

microbial degradation of oil dates to at least 1942,

when the American Petroleum Institute began to

subsidize research on the topic.5 Considerable basic

knowledge about factors that affect natural biodegradation, about the kinds of hydrocarbons capable of

being degraded, and about the species and distribuINatio~ Research COunCil, oil in the Sea: Inputs, Fates, and Effects (Washington DC: National Academy ~ess, 1983. P. 2~.

me term “marine oil spills” is here defined to include spills at sea, in bays and estuaries, on beaches, and in environments such as salt marshes

in contact with the sea.

3sever~ o~s~dies ~ve ad&es~ v~ou5 ~wts of these topics. s=, for e~plq cornmer~”uf ~iotec~~~l~gy: An Znternafi”onal Analysis,

OTA-BA-218 (Washington DC: U.S. Government Printing OffIce, 1984); New Developments in Biotechnology, OTA-BA-360 (Washington DC: U.S.

Government Printing OffIce, July 1988); Coming CZean: Supe@nd ProbZems Can Be Solved, OTA-ITE-433 (Washington DC: U.S. Government

PrMng OffIce, October 1989); and Biotechnology in a GZobaZEconomy (Washington, DC: U.S. Government Printing Off&, expected publication date

mid-1991).

4u.s. Enviro~en~ Protection Agency, bioremediation Action committee, Summary of Nov. 7, 1990 meeting.

5c.E. Z@ell, “Microb~ Degradation of Oil: Present Status, Problems, and PerSptXtiWS, “ in The Microbial Degradation of Oil Pollutants

(proceedings of a workshop at Georgia State University, Atlanta, December 1972), D.G. Ahearn and S.P. Meyers (eds.), 1973.

–l–
2 ● bioremediationfor Marine Oil Spills

Box A—bioremediation v. Biodegradation

Biodegradation refers to the natural process

whereby bacteria or other microorganisms alter and

break down organic molecules into other substances, such as fatty acids and carbon dioxide.

bioremediation is the act of adding materials to

contaminated environments, such as oil spill sites,

to cause an acceleration of the natural biodegradation process.

Fertilization is the bioremediation method of

adding nutrients, such as nitrogen and phosphorus, to a contaminated environment to stimulate

the growth of indigenous microorganisms. This

approach is also termed nutrient enrichment.

Seeding refers to the addition of microorganisms to a spill site. Such microorganisms mayor

may not be accompanied by nutrients. Current

seeding efforts use naturally occurring microorganisms. Seeding with genetically engineered

microorganisms (GEMs) may also be possible,

but this approach is not now being considered for

remediating oil spills.

tion of the microorganisms involved in biodegradation had already been developed by the early 1970s.

For example, the Office of Naval Research sponsored about 15 basic and applied research projects in

the late 1960s and early 1970s on oil biodegradation

as part of the charge to the Navy at that time to take

the lead in mitigating marine oil pollution. In 1972,

a workshop on the microbial degradation of oil

pollutants was sponsored by the Office of Naval

Research, the U.S. Coast Guard, and the U.S.

Environmental Protection Agency (EPA).6 The 1980s

were a period of rapid advances in knowledge of the

genetics and molecular biology of bacterial degradation of different hydrocarbons, and of renewed

interest in the microbial ecology of pollutionstressed environments.

Much progress has been made in applying basic

knowledge of biodegradation to cleaning up terrestrial and enclosed sites polluted with oil. As long ago

as 1967, contractors employed bioremediation to

improve the quality of 800,000 gallons of oily

wastewater remaining in the bilge tanks of the

Queen Mary after it was permanently moored in

Long Beach Harbor.7 City officials approved the

discharge of this bilge water 6 weeks after treatment.

More importantly, a large number of refineries, tank

farms, and transfer stations now employ in situ

bioremediation to restore land contaminated by

accidental spills of fuel oil or other hydrocarbons.8

Much less progress has been made with respect to

the practical problems of applying bioremediation

technologies to marine oil spills, although advocates

have suggested their use in the wake of several major

spills. The problems associated with using bioremediation technologies in marine environments are

fundamentally different from those associated with

land-based applications. Although bioremediation

of oil-contaminated soil is one of the fastest growing

uses of this technology, bioremediation applications

on land have all been accomplished in closed or

semi-enclosed environments where microorganisms

have little or no competition and where conditions

can be closely controlled and monitored. The marine

environment is a dynamic, open system that is much

less susceptible to control, and many additional

variables exist to compound the difficulties of

applying bioremediation techniques. One of the

most important series of tests and the first largescale application of a bioremediation technology in

a marine setting was conducted between 1989 and

1991 by the EPA, Exxon, and the State of Alaska on

the Prince William Sound beaches fouled by oil

from the Exxon Valdez.

SUMMARY AND FINDINGS

Biodegradation is a natural process, and there is

no question that, with enough time, microorganisms

can eliminate many components of oil from the

environment. The central concern of this study is

whether bioremediation technologies can accelerate

this natural process enough to be considered practical, and, if so, whether they might find a niche as

replacements for, or adjuncts to, other oil spill

response technologies. The key findings from this

OTA study are summarized below:

. The usefulness of bioremediation for marine

oil spills is still being evaluated, and their

6D.G. AIMXUII and S.P. Meyers (eds.), The MicrobialDegradation of Oil Pollutants (proeeedi.ngs of a workshop at Georgia State hiversity, A~Q

December 1972) (Baton Rouge, LA: Center for Wetland Resources, Louisiana State University, 1973).

TApplied BiOtreament Associario@ “Case History Compendiurq” November 1989, p. 34. The compendium also Contains Oher exwples of the

use of bioremediation technologies to address environmental problems.

8T.G. Zitrides, “Bioremediat.ion Comes of Age, ” Pollution Engineering, vol. XXII, No. 5, May 1990, pp. 59-60.
Bioremediation for Marine Oil Spills . 3

●

Photo credit: Environmental Protection Agency

Prince William Sound, Alaska, site of the extensive

bioremediation experiments carried out by the

Environmental Protection Agency, Exxon, and

the State of Alaska.

ultimate importance relative to other oil spill

response technologies remains uncertain.

Recent research and field testing of bioremediation technologies on oiled beaches has produced some encouraging, if not altogether

conclusive, results. Nevertheless, technologies

other than bioremediation (especially mechanical ones) are likely to remain the mainstay of

the Nation’s response arsenal for now. In

certain non-emergency situations (e.g., for

cleaning lightly to moderately oiled beaches),

bioremediation could be employed as a primary

technology. Mechanical methods, dispersants,

and possibly in situ burning will most likely

remain more appropriate technologies for the

immediate response to spills at sea.

Potential bioremediation approaches for marine oil spills fall into three major categories:

1) stimulation of indigenous microorganisms

through addition of nutrients (fertilization),

2) introduction of special assemblages of naturally occurring oil-degrading microorganisms

(seeding), and 3) introduction of genetically

engineered microorganisms (GEMs) with special oil-degrading properties. Stimulation of

indigenous organisms by the addition of

nutrients is the approach that has been

tested most rigorously. This approach is

viewed by many researchers as the most

promising one for responding to most types

of marine spills. Recent experiments suggest

that rates of biodegradation in most marine

environments are constrained by lack of nutrients rather than by the absence of oil-degrading

microbes. The introduction of microbes might

be beneficial in areas where native organisms

grow slowly or are unable to degrade a particular hydrocarbon. However, the effectiveness of

this approach has not yet been demonstrated.

The wide availability of naturally occurring

microorganisms capable of degrading components of petroleum will likely deter consideration of GEMs for remediating marine oil spills.

Moreover, greater research and development

needs, regulatory hurdles, and public perception problems will remain obstacles to the

near-term use of GEMS even if they could

prove useful for degrading some recalcitrant

components of petroleum.

bioremediation technologies for beach cleanup

have thus far received the most attention.

Experiments conducted by EPA, Exxon, and

the State of Alaska on cobble beaches fouled by

oil from the Exxon Valdez indicated that the

addition of nutrients at least doubled the

natural rate of biodegradation. The efficacy of

commercial microbial products in remediating

beaches is not yet known. Limited EPA field

tests using two microbial products on heavily

weathered oil in Alaska were inconclusive.

Additional field experiments are required on

other types of beaches that involve different

oils and different climatic and marine conditions.

bioremediation has not yet been demonstrated to be an effective response to "atsea” oil spills. The necessary studies have not

been done, in part because of the difficulty of

conducting controlled experiments and monitoring in the open ocean. Limited applications

have been made in the Gulf of Mexico, but they

have not provided definitive data on effectiveness. The design and validation of open ocean

protocols to test products are necessary before

the efficacy of bioremediation at sea can be

determined or widely accepted. Even if bioremediation proves effective in some situations,

other, quicker-acting alternatives may be preferable as primary response tools.
4 ● Bioremediation for Marine Oil Spills

bioremediation may have a role in settings such

as salt marshes and sensitive ecosystems where

the use of mechanical or other approaches

might do more harm than good. Just as for open

water spills, however, appropriate protocols

need to be developed for testing and applying bioremediation technologies in these

situations, and more research is required to

prove their effectiveness.

No significant adverse impacts related to the

use of bioremediation technologies for oil spill

cleanup have been identified in recent field

applications. Effects that have been measured

have been short-lived and minor. On beaches,

in particular, bioremediation may be a less

intrusive approach than other alternatives. However, experience with bioremediation in marine

settings is limited, and it is premature to

conclude that the use of bioremediation technologies will be safe in all circumstances.

Regulatory controls to ensure the safe use of

bioremediation appear adequate, and there

appear to be no significant Federal regulatory

obstacles to the greater use of bioremediation

technologies, except those using GEMs. However, more development and testing of both

fertilization and seeding technologies are

needed before on-scene coordinators or others responsible for oil spill cleanup would be

comfortable advocating their use. Most decisionmakers prefer more traditional methods,

and usually are not willing to experiment

during a real spill. bioremediation technologies

for response to marine oil spills, although

promising, are still in the experimental phase.

One regulatory change that could help stimulate development of both bioremediation and

other oil spill response technologies is for the

Federal Government to allow occasional controlled oil spills for research and development

purposes.

If additional research confirms the effectiveness of bioremediation and leads to the development of more reliable technologies, oil spill

decisionmakers will have to be educated about

the efficacy of various techniques, the advantages and disadvantages of their use, and the

availability of materials and expert assistance.

Preliminary efforts to accomplish this have

recently been undertaken by EPA. However,

before detailed bioremediation contingency

plans can be developed, uncertainties about the

effectiveness of bioremediation must be addressed. Detailed plans, when and if necessary,

would require such information as the oildegrading capabilities of microorganisms indigenous to a particular area, the characteristics

of the oil most likely to be spilled in that area,

environmental factors constraining oil biodegradation, and the circumstances under which

the use of bioremediation technologies would

be appropriate or allowed.

EPA, through its bioremediation Action Committee and research labs, and with the assistance of the National Environmental Technology Applications Corp., is developing protocols to determine the efficacy and toxicity of

bioremediationproducts in a variety of settings.

Testing products against such protocols would

provide decisionmakers with the kind of data

needed to determine whether these products

could be used in response to marine oil spills.

A research program to expand the present

knowledge of biodegradation mechanisms, and

to improve bioremediation technologies and

the means of applying them to marine oil spills,

appears to be justi.tied. Redirecting an appreciable fraction of available marine oil spill research funds to bioremediation does not, however, seem warranted. Efforts to improve other

oil spill prevention and response technologies

are also important, and funding is limited.

Improving methods for enhancing the growth

and activity of petroleum-degrading bacteria is

important, as is the development of better

analytical techniques for measuring and monitoring effectiveness, and field validation of

laboratory work. Government and industry

should be encouraged to coordinate their research efforts and to share as much information

as possible.

BACKGROUND

Evaluating the effectiveness of bioremediation

technologies is complicated by several factors. First,

biodegradation is only one of the processes at work

removing petroleum from the marine environment;

to understand the effect of this process on oil

removal, one must know the effects of other

processes. Second, petroleum is not the simple

material many people presume it to be; rather, it

contains thousands of compounds. Some of these are
Bioremediation for Marine Oil Spills ● 5

easily biodegraded; others are relatively resistant to

biodegradation. Third, a large number-not just one

or a few-of species of microorganisms are responsible for biodegradation, and these species have

evolved many metabolic pathways to degrade oil.

Although the general mechanisms of biodegradation

are known, many details remain to be filled in.

Underlying these complications are the basic

issues of what constitutes clean and how long one is

willing to wait for results. These are both political

and scientific issues. Can a beach be considered

clean, for example, if its surface appears clean but

close examination reveals oil below the surface, or

if unsightly but relatively less harmful constituents

of oil, such as hard-to-degrade asphalt, remain on the

beach after bioremediation is used? How much of an

improvement over natural biodegradation rates must

bioremediation technologies offer before their use

would be warranted? A rate increase of a factor of 2

or more would be significant for a spill that might

otherwise persist for 5 or more years, but much less

so for a spill that might be naturally degraded in less

than a year.

The Fate of Oil in the Marine Environment

The fate of petroleum in marine ecosystems has

been intensively studied.9 Crude oil and petroleum

distillate products introduced to the marine environment are immediately subject to a variety of physical

and chemical, as well as biological, changes (figure

1).10 Abiological weathering processes include evaporation, dissolution, dispersion, photochemical oxidation, water-in-oil emulsification, adsorption onto

suspended particulate material, sinking, and sedimentation. Biological processes include ingestion

by organisms as well as microbial degradation.ll

These processes occur simultaneously and cause

important changes in the chemical composition and

physical properties of the original pollutant, which

in turn may affect the rate or effectiveness of

biodegradation.

The most important weathering process during

the first 48 hours of a spill is usually evaporation, the

process by which low- to medium-weight crude oil

components with low boiling points volatilize into

the atmosphere. Evaporation can be responsible for

the loss of one- to two-thirds of an oil spill’s mass

during this period,12 with the loss rate decreasing

rapidly over time.13 Roughly one-third of the oil

spilled from the Amoco Cadiz, for example, evaporated within the frost 3 days. Evaporative loss is

controlled by the composition of the oil, its surface

area and physical properties, wind velocity, air and

sea temperatures, sea state, and the intensity of solar

radiation. 14 The material left behind is richer in

metals (mainly nickel and vanadium), waxes, and

asphaltenes than the original oil.15 With evaporation, the specific gravity and viscosity of the original

oil also increase. For instance, after several days,

spilled crude oil may begin to resemble Bunker C

(heavy) oil in composition.lG

None of the other abiological weathering processes accounts for as significant a proportion of the

losses from a spill. For example, the dissolving, or

dissolution, of oil in the water column is a much less

important process than evaporation from the perspective of mass lost from a spill; dissolution of even

a few percent of a spill’s mass is unlikely. Dissolution is important, however, because some watersoluble fractions of crude oil (e.g., the light aromatic

compounds) are acutely toxic to various marine

organisms (including microorganisms that may be

able to degrade other fractions of oil), and their

impact on the marine environment is greater than

mass balance considerations might imply.17

%or example, see references listed in National Research Council, Oil in the Sea: Inputs, Fates, and l?~ecfs (Washington, DC: National Academy

Press, 1985), pp. 335-368; and in G.D. Floodgate, ‘‘The Fate of Petroleum in Marine Environments,’ R.M. Atlas (cd.), Petroleum Microbiology (New

York, NY: Macmillan Publishing Co., 1984), pp. 355-397.

l~atio~ Rese~ch Council, op. cit., footnote 1, p. 270.

1lJ.R. payne~d G.D. McNabb, Jr., “Weatheringof Petroleum in the Marine Environment,’ Marine Technology Society Journal, vol. 18, No. 3,1984,

p. 24.

lzNatio@ Research Council, op. cit., footnote 1, p. 276.

13Flwdgate, op. cit., footnote g, P. 362.

14paPe ~d McNabb, op. cit., footnote 11, p. 26.

ls~oodgate, op. cit., footnote 9, p. 362

115J.E. Mie&e, “oil in the Ocean: The Short- and Imng-T&m Impacts of a Spill,” Congressional Research Service, 90-356 SPR, July 24, 1990, p.

11.

ITNatio~ Resemch Council, op. cit., footnote 1, pp. 277-278.
6 ● bioremediationfor Marine Oil Spills

Figure l—Schematic of Physical, Chemical, and Biological Processes

Wind

Sea

surface

Atmospheric oxidation

(photo-oxidation)

Current

movement ~

Water-in-oil

h emulsions +

Tar

Spreading Sea surface oil slick “chocolate mousse” balls b

~ Dissolution t

I

( I

Globular Solubilizing I

dispersion

I

chemical Current movement

Oil-in. transformations ~ and ~

/

water Chemical

diffusion

I

emulsion degradation

Attachment

I

to Dispersed I

particulate+ crude oil

\

matter

\

Degradation or

\

I

Nonbuoyant assimilation by

oil residues fish I

v

“ \ !

/

Bulk

subsurface \ \ Biological /

discharge detritus

-- I

/

1

Sea floor

— /

— ~ w

Sea floor sediments — —

assimilation by bottom

— — dwelling organisms —

+

SOURCE: National Research Council, Oil in the Sea: Inputs, Fates, and Effects (Washington, DC: National Academy Press, 1985), p. 271. Adapted from

R. Burwood and G.C. Speers, “Photo-oxidation as a Factor in the Environmental Dispersal of Crude Oil,” Estuarine Coastal Marine Science,

vol. 2, 1974, pp. 117-135.

Dispersion, the breakup of oil and its transport as have had trouble being accepted in the United States.

small particles from the surface to the water column, The National Research Council has generally apis an extremely important process in the disappear- proved their use, 19 but effectiveness and, to a lesser

ance of a surface slick.18 Dispersion is controlled degree, toxicity remain concerns. Dispersed oil

largely by sea surface turbulence: the more turbu- particles are more susceptible to biological attack

lence, the more dispersion. Chemical dispersants than undispersed ones because they have a greater

have been formulated to enhance this process. Such exposed surface area. Hence, dispersants may endispersants are intended as a first-line defense hance the rate of natural biodegradation.20

against oil spills that threaten beaches and sensitive

habitats such as salt marshes and mangrove swamps. Water-in-oil emulsions, often termed “mousse,”

Although used widely in other countries, dispersants are formed when seawater, through heavy wave

lspawe and McNabb, op. cit., footnote 11, p. 30.

l~atio~ Research COunCil, Marine Board, Using OiZDispersants on the Sea (Washington, DC: National kad~y press, 1989).

%.R. Colwell and J.D. Walker, “Ecological Aspects of Microbial Degradation of Petroleum in the Marine Environment” CRC Critical Reviews

in Microbiology, September 1977, p, 430.
Bioremediation for Marine Oil Spills ● 7

action, becomes entrained with the insoluble components of oil. Such emulsions can form quickly in

turbulent conditions and may contain 30 to 80

percent water.

21 Heavier or weathered crudes with

high viscosities form the most stable mousses.

Mousse will eventually disperse in the water column

and/or be biodegraded, but may first sink or become

stranded on beaches. A water-in-oil emulsion is

more difficult for microorganisms to degrade than

oil alone.22 Mousse formation, for example, has been

suggested as a major limiting factor in petroleum

biodegradation of the Ixtoc I and Metula spills,

probably because of the low surface area of the

mousse and the low flux of oxygen and mineral

nutrients to the oil-degrading microorganisms within

i t .

2 3

Natural biodegradation is ultimately one of the

most important means by which oil is removed from

the marine environment, especially the nonvolatile

components of crude or refined petroleum (see

below). In general, it is the process whereby

microorganisms (especially bacteria, but yeasts,

fungi, and some other organisms as well) chemically

transform compounds such as petroleum hydrocarbons into simpler products. Although some products

can actually be more complex, ideally hydrocarbons

would be converted to carbon dioxide (i.e., mineralized), nontoxic water-soluble products, and new

microbial biomass. The mere disappearance of oil

(e.g., through emulsification by living cells) technically is not biodegradation if the oil has not actually

been chemically transformed by microbes.25 The

ideal may be difficult to reach, particularly in a

reasonably short time, given the recalcitrance of

some petroleum fractions to biodegradation (discussed below) and the many variables that affect its

rate and extent. Man-made bioremediation technologies are intended to improve the effectiveness of

natural biodegradation.

Biodegradation and the Chemical Nature

of Petroleum

Far from being a homogeneous substance, crude

oil is a complex mixture of thousands of different

chemical compounds. In addition, the composition

of each accumulation of oil is unique, varying in

different producing regions and even in different

unconnected zones of the same formation.26 The

composition of oil also varies with the amount of

refining. Significantly, the many compounds in oil

differ markedly in volatility, volubility, and susceptibility to biodegradation. Some compounds are

readily degraded; others stubbornly resist degradation; still others are virtually nonbiodegradable. The

biodegradation of different petroleum compounds

occurs simultaneously but at very different rates.

This leads to the sequential disappearance of individual components of petroleum over time and,

because different species of microbes preferentially

attack different compounds, to successional changes

in the degrading microbial community .27 Since

components of petroleum degrade at different rates,

it is difficult and misleading to speak in terms of an

overall biodegradation rate.

Petroleum hydrocarbons can, in general, be divided into four broad categories: saturates, aromatics, asphaltenes, and resins.28 Saturated hydrocarbons-those with only single carbon-carbon

bonds—usually constitute the largest group. Of

these, the normal or straight-chain alkane series is

the most abundant and the most quickly degraded.

Compounds with chains of up to 44 carbon atoms

can be metabolized by microorganisms, but those

having 10 to 24 carbon atoms (CIO-C24) are usually

the easiest to metabolize. Shorter chains (up to about

C12) also evaporate relatively easily. Only a few

species can use Cl

-C4 alkanes; C5-C9 alkanes are

degradable by some microorganisms but toxic to

zl~e~e, Op. cit., footnote 16, P. 12.

22K. ~~d EM. ~v, $ $Bi~ewa&tion of pe~oleu ~ the -eEnvironment ad Its E~cemen~” inA~tic Toxicology and Water QuaZity

Management, J.O. Nngau and J.S.S. Lakshminarayana (eds.) (New York NY: John Wiley & Sons, 1989), p. 221.

23R.M. A~~, 1‘BiodeW~ation of Hy&W~omin the Environmen~>’~Environ~nta/Bi~tec~ ~o/ogy, G.S. omenn (cd.), (New York NY: plenum

Press, 1988), p. 214.

~Natio~Resemch COunCil, Op. Cit., footnote 1, p. 290-

‘J.J. Cooney, “Microbial Ecology and Hydrocm%on Degradation” paper presented at the Alaska Story Symposiuq Cincinnati, OH, Sept. 17-18,

1990., p. 2.

zsNatio@ Resemch Council, op. cit., footnote 1, p. 17.

zTAths, op. cit., footnote 23, p. 212.

MJ.G. bay and R.R.Colwell, “Microbial Degradation of Hydrocarbons in the Environment” Microbiological Reviews, September 1990, p. 305.
8 ● Bioremediation for Marine Oil Spills

others. 29 Branched alkanes are usually more resistant

to biodegradation than normal alkanes but less

resistant than cycloalkanes (naphthenes)-those alkanes having carbon atoms in ringlike central

structures.30 Branched alkanes are increasingly resistant to microbial attack as the number of branches

increases. At low concentrations, cycloalkanes may

be degraded at moderate rates, but some highly

condensed cycloalkanes can persist for long periods

after a spill.31 Light oils contain 10 to 40 percent

normal alkanes, but weathered and heavier oils may

have only a fraction of a percent. Heavier alkanes

constitute 5 to 20 percent of light oils and up to 60

percent of heavier oils.32

Aromatic hydrocarbons are those characterized

by the presence of at least one benzene (or substituted benzene) ring. The low-molecular-weight aromatic hydrocarbons are subject to evaporation and,

although toxic to much marine life, are also relatively easily degraded. Light oils typically contain

between 2 and 20 percent light aromatic compounds,

whereas heavy oils contain 2 percent or less.33 As

molecular weight and complexity increase, aromatics are less readily degraded. Thus, the degradation

rate of polyaromatics is slower than that of monoaromatics. Aromatics with five or more rings are not

easily attacked and may persist in the environment

for long periods.34 High-molecular-weight aromatics

comprise 2 to 10 percent of light oils and up to 35

percent of heavy oils.35

The asphaltic fraction contains compounds that

either are not biodegradable or are degraded very

slowly. 36 One of the reasons that tar, which is high in

asphaltenes, makes an excellent road paving material is because it is slow to degrade. Tar balls, like

mousse, are difficult to degrade because their low

surface area restricts the availability of oxygen and

other nutrients. Resins include petroleum compounds containing nitrogen, sulfur, and/or oxygen as

constituents. If not highly condensed, they may be

subject to limited microbial degradation. Asphaltenes and resins are difficult to analyze and, to date,

little information is available on the biodegradability of most compounds in these groups.37 Light oils

may contain about 1 to 5 percent of both asphaltenes

and resins; heavy or weathered oils may have up to

25 percent asphaltenes and 20 percent resins.38

To summarized biodegradation rates are typically

highest for the saturates, followed by the light

aromatics, with high-molecular-weight aromatics,

asphaltenes, and resins exhibiting extremely low

rates of degradation.39 As a spill weathers, its

composition changes: the light aromatics and alkanes dissolve or evaporate rapidly and are metabolized by microorganisms. The heavier components

that are harder to degrade remain. Weathered

Prudhoe Bay oil contains about 10 percent low-molecular-weight alkanes, 45 percent high-molecularweight alkanes, 5 percent light aromatics, 20 percent

high-molecular-weight aromatics, 10 percent asphaltenes, and 10 percent resins.40

Departures from the typical pattern of biodegradation, however, have been noted by some researchers.

For example, extensive losses of asphaltenes and

resins have been observed in some cases. The

microbial degradation of these relatively recalcitrant

fractions has been ascribed to co-oxidation.41 In this

process, a normally refractory hydrocarbon may be

partially degraded in the presence of a second readily

degraded hydrocarbon. Clearly, degradation rates

depend on many factors, and generalizations are

difficult to make. One conclusion, however, seems

reasonable: no crude oil is subject to complete

biodegradation, and claims that all of a light oil or

zgAtlm, op. cit., footnote 23, p. 212.

30Cwney, op. cit., fOOmOte 25.

slA~as, op. cit., footnote 23, p. 212.

32M. HngIs, Environment CanadA personal COIIIIIItiCatiOIL wt. % 1990.

3%id.

34Natio~ Rme~ch Council, op. cit., footnote 1, p. 293.

35Fqy5, 0p. Cit., footnote 32-

36Cooney, op. cit., fOOtnOte 25, P“ 3“

37Cwney, op. cit., fm~ote 25, p. 3; and National Research Council, op. cit., foo~ote 1, P. 295-

38F~gm, op. cit., fOOtnOte 32.

3WY and Colwell, op. cit., footnote 28, p. 305.

@13ngaa, op. cit., footnote 32.

41UY and Colwell, op. cit., footnote 28, p. 306.
Bioremediation for Marine Oil Spills . 9

more than 50 percent of a heavy oil can be

biodegraded in days or weeks are highly suspect.42

Microbial Processes and the Degradation

of Petroleum

Despite the difficulty of degrading certain fractions, some hydrocarbons are among the most easily

biodegradable naturally occurring compounds. Altogether, more than 70 microbial genera are known

to contain organisms that can degrade petroleum

components (table 1). Many more as-yet-unidentified strains are likely to occur in nature.43 Moreover, these genera are distributed worldwide. All

marine and freshwater ecosystems contain some

oil-degrading bacteria. No one species of microorganism, however, is capable of degrading all the

components of a given oil. Hence, many different

species are usually required for significant overall

degradation. 44 Both the quantity and the diversity of

microbes are greater in chronically polluted areas. In

waters that have not been polluted by hydrocarbons,

hydrocarbon-degrading bacteria typically make up

less than 1 percent of the bacterial population,

whereas in most chronically polluted systems (harbors, for example) they constitute 10 percent or more

of the total population.45

Microorganisms have evolved their capability to

degrade hydrocarbon compounds over millions of

years. These compounds are a rich source of the

carbon and energy that microbes require for growth.

Before that carbon is available to microorganisms,

however, large hydrocarbon molecules must be

metabolized or broken down into simpler molecules

suitable for use as precursors of cell constituents.

The activity of microorganisms at a spill site is

governed by the organisms’ ability to produce

enzymes to catalyze metabolic reactions. This ability is, in turn, governed by their genetic composition.

Enzymes produced by microorganisms in the presence of carbon sources are responsible for attacking

the hydrocarbon molecules. Other enzymes are

utilized to break down hydrocarbons further.% Lack

of an appropriate enzyme either prevents attack or is

a barrier to complete hydrocarbon degradation.

Table l—Major Genera of Oil-Degrading Bacteria

and Fungi

Bacteria

Achrornobacter

Acinetobacter

Actinomyces

Aeromonas

Alcaligenes

Arthrobacter

Bacillus

Beneckea

Brevebacterium

Coryneforms

Erwinia

Flavobacterium

Klebsiella

Lactobaoillus

Leumthrix

Moraxella

Nocardia

Peptococcus

Pseudomonas

Sarcina

Spherotilus

Spirillum

Streptomyces

Vibrio

Xanthomyces

Fungi

Allescheria

Aspergillus

Aureobasidium

Botrytis

Candida

Cephalosporium

Cladosporium

Cunninghamella

Debaromyces

Fusarium

Gonytrichum

Hansenula

Helminthosporium

Mucor

Oidiodendrum

Paecylomyces

Phialophora

Penicillium

Rhodosporidium

Rhodotorula

Saccharomyces

Saccharomycopisis

Scopulariopsis

Sporobolomyces

Torulopsis

Trichoderma

Trichosporon

SOURCE: G.D. Floodgate, “The Fate of Petroleum in Marine Ecosystems,”

Petroleum Microbiology, R.M. Atlas (ad.) (New York, NY:

Macmillan Publishing Co., 1984), p. 373.

The complex series of steps by which biodegradation occurs constitutes a metabolic pathway. Many

different enzymes and metabolic pathways, not all of

which can be found in any single species, are

required to degrade a significant portion of the

hydrocarbons contained in petroleum. (Thus, advocates of using specially selected mixtures of microorganisms to bioremediate oil spills or of creating,

through recombinant DNA technology, genetically

engineered organisms are motivated in part by the

desire to combine all the requisite enzymes and

pathways. 47)

Knowledge of the numerous metabolic pathways

involved in the breakdown of hydrocarbons is far

from complete. Additional research characterizing

the microbiology and population dynamics of bac4~~ney, op. cit., footnote 25, P- 3<

43flmdgate, op. cit., footnote 9, P. 372.

~~e and IAWy, op. cit., footnote 22, pp. 217-243.

45Cmney, op. cit., fOOtnOte 257 P. 1.

46Applied Bio@ea@ent A~~~iatioq “me Role of Bio@~~ent of Ofi Spifls,” KW. 2, August 1990, p. 4.

dTAtks, op. cit., footnote 23, P. 213.
10 ● Bioremediation for Marine Oil Spills

terial species capable of degrading oil is critical to

understanding the biodegradation process.

Environmental Influences on Biodegradation

Environmental variables can also greatly influence the rate and extent of biodegradation. Variables

such as oxygen and nutrient availability can often be

manipulated at spill sites to enhance natural biodegradation (i.e., using bioremediation). Other variables, such as salinity, are not usually controllable.

The great extent to which a given environment can

influence biodegradation accounts for some of the

difficulty in accurately predicting the success of

bioremediation efforts. Lack of sufficient knowledge about the effect of various environmental

factors on the rate and extent of biodegradation is

another source of uncertainty.

Oxygen

Oxygen is one of the most important requirements

for microbial degradation of hydrocarbons. However, its availability is rarely a rate-limiting factor in

the biodegradation of marine oil spills. Microorganisms employ oxygen-incorporating enzymes to initiate attack on hydrocarbons. Anaerobic degradation

of certain hydrocarbons (i.e., degradation in the

absence of oxygen) also occurs, but usually at

negligible rates. Such degradation follows different

chemical paths, and its ecological significance is

generally considered minor.48 For example, studies

of sediments impacted by the Amoco Cadiz spill

found that, at best, anaerobic biodegradation is

several orders of magnitude slower than aerobic

biodegradation. 49 Oxygen is generally necessary for

the initial breakdown of hydrocarbons, and subsequent reactions may also require direct incorporation

of oxygen. Requirements can be substantial; 3 to 4

parts of dissolved oxygen are necessary to completely oxidize 1 part of hydrocarbon into carbon

dioxide and water.50

Oxygen is usually not a factor limiting the rate of

biodegradation on or near the surface of the ocean,

where it is plentiful and where oil can spread out to

provide a large, exposed surface area. Oxygen is also

generally plentiful on and just below the surface of

beaches where wave and tide action constantly assist

aeration. When oxygen is less available, however,

the rates of biodegradation decrease. Thus, oil that

has sunk to the sea floor and been covered by

sediment takes much longer to degrade. Oxygen

availability there is determined by depth in the

sediment, height of the water column, and turbulence (some oxygen may also become available as

the burrowing of bottom-dwelling organisms helps

aeration) .51 Low-energy beaches and fine-grained

sediments may also be depleted in oxygen; thus, the

rate of biodegradation may be limited in these areas.

Pools of oil are a problem because oxygen is less

available below their surfaces. Thus, it may be

preferable to remove large pools of oil on beaches,

as was done in Alaska, before attempting bioremediation.

Nutrients

Nutrients such as nitrogen, phosphorus, and iron

play a much more critical role than oxygen in

limiting the rate of biodegradation in marine waters.

Several studies have shown that an inadequate

supply of these nutrients may result in a slow rate of

biodegradation. 52 Although petroleum is rich in the

carbon required by microorganisms, it is deficient in

the mineral nutrients necessary to support microbial

growth.53 Marine and other ecosystems are often

deficient in these substances because non-oildegrading microorganisms (including phytoplankton) consume them in competition with the oildegrading species. Also, phosphorus precipitates as

calcium phosphate at the pH of seawater. Lack of

nitrogen and phosphorus is most likely to limit

biodegradation, but lack of iron or other trace

minerals may sometimes be important. Iron, for

instance, is more limited in clear offshore waters

than in sediment-rich coastal waters.54

4S~~y and Colwell, op. cit., footnote 28, P. 307.

%.M. War@ R.M. Atlas, P.D. Boehrn, and J.A. Calder, “Microbial Biodegradation and the Chemical Evolution of Amoco CUdiz Oil Pollu@@”

Ambio, vol. IX, No. 6, 1980, pp. 279.

%ee and levy, op. cit., footnote 22, p. 224.

51 CWney, op. cit., fOOmOte 25, P“ 4“

sz~ md UVY, op. cit., footnote 22, p. 223.

5sA&, op. cit., footnote 23, p. 216.

541bid.
Bioremediationfor Marine Oil Spills ● 11

Phofo credit: Exxon Corp.

Workman applying fertilizer to the cobble beaches of

Prince William Sound.

Scientists have attempted to adjust nutrient levels

(e.g., by adding nitrogen- and phosphorus-rich

fertilizers) to stimulate biodegradation of petroleum

hydrocarbons. This is the experimental bioremediation

approach used recently on about 110 miles of

beaches in Prince William Sound, Alaska. Researchers have also experimented with alternative methods

of applying nutrients. Given the necessity of keeping

nutrients in contact with oil, the method of application is itself likely to be an important factor in the

success of bioremediation.

Temperature

The temperature of most seawater is between

–2 and 35oC.55 Biodegradation has been observed in

this entire temperature range, and thus in water

temperatures as different as those of Prince William

Sound and the Persian Gulf. The rates of biodegradation are fastest at the higher end of this range and

usually decrease—sometimes dramatically in very

cold climates-with decreasing temperature. One

experiment showed that a temperature drop from 25

to 5oC caused a tenfold decrease in response.56 At

low temperature, the rate of hydrocarbon metabolism by microorganisms decreases.57 Also, lighter

fractions of petroleum become less volatile, thereby

leaving the petroleum constituents that are toxic to

microbes in the water for a longer time and

depressing microbial activity. Petroleum also becomes more viscous at low temperature. Hence, less

spreading occurs and less surface area is available

for colonization by microorganisms. In temperate

regions, seasonal changes in water temperature

affect the rate of biodegradation, but the process

continues year-round.58

Other Factors

Several variables, including pressure, salinity,

and pH may also have important effects on biodegradation rates. Increasing pressure has been correlated

with decreasing rates of biodegradation; therefore,

pressure may be very important in the deep ocean.

Oil reaching great ocean depths degrades very

slowly and, although probably of little concern, is

likely to persist for a long time.59

Microorganisms are typically well adapted to

cope with the range of salinities common in the

world’s oceans. Estuaries may present a special case

because salinity values, as well as oxygen and

nutrient levels, are quite different from those in

coastal or ocean areas. However, there is little

evidence to suggest that microorganisms are adversely affected by other than hypersaline environments.60

Extremes in pH affect a microbe’s ability to

degrade hydrocarbons. However, like salinity, pH

does not fluctuate much in the oceans-it remains

between 7.6 and 8. l—and does not appear to have an

important effect on biodegradation rates in most

marine environments. In salt marshes, however, the

pH maybe as low as 5.0, and thus may slow the rate

of biodegradation in these habitats.6l

55~oodgate, op. cit., footnote 9, P. 3*1.

‘eIbid.

WU&y and Colwell, op. cit., footnote 28, p. 307.

58moodgate, op. cit., fOO~Ote 9, P. 381-

59J.R. &-hwm, J*D. Wtier, ad R*R4 Colweu, “Deep.sea Bacteria: @owth ad Utibtion of Hy&oc~m at Ambient ~d bSitu PKxXlllV,’

Applied Microbiology, vol. 28, 1974, pp. 982-986.

%e and Levy, op. cit., footnote 22, p. 225.

Glh~y and Colwell, op. cit., footnote 28, p. 308.

292-854 0 - 91 - 2
12 ● Bioremediation for Marine Oil Spills

Table 2—bioremediation: Potential Advantages

and Disadvantages

Advantages:

. Usually involves only minimal physical disruption of a site

. No significant adverse effects when used correctly

. Maybe helpful in removing some of the toxic components of oil

● Offers a simpler and more thorough solution than mechanical

technologies

● Possibly less costly than other approaches

Disadvantages:

● Of undetermined effectiveness for many types of spills

● May not be appropriate at sea

● Takes time to work

● Approach must be specifically tailored for each polluted site

. Optimization requires substantial information about spill site

and oil characteristics

SOURCE: Office of Technology Assessment, 1991.

General Advantages and Disadvantages

of bioremediation

bioremediation technologies have several attributes that, depending on the situation and type of site

may support their use in responding to some oil

spills (table 2).62 First, bioremediation usually

involves minimal physical disruption of a site. This

attribute is especially important on beaches where

other available cleanup technologies (e.g., high- and

low-pressure spraying, steam cleaning, manual scrubbing, and raking of congealed oil) may cause

additional damage to beach-dwelling biota.63 Application of oleophilic (i.e., oil seeking) fertilizers

during the 1989-90 Alaska bioremediation experiments was accomplished largely from shallow draft

boats located just off the beach. Second, bioremediation technologies appear to have no or only minor

and short-lived adverse effects when used correctly.

Although research on possible negative impacts is

continuing, there is so far little evidence to suggest

that potential problems would be significant.

Third, bioremediation may be useful in helping

remove some of the toxic components of petroleum

(e.g., low-molecular-weight aromatic hydrocarbons)

from a spill site more quickly than they might

otherwise be removed by evaporation alone.64

Fourth, bioremediation of oil spills is accomplished

on-site, and offers a simpler and more thorough

solution to polluted areas. In contrast, hot water

Photo credit: Exxon COrP.

One of the shallow draft boats used by Exxon to apply

oleophilic fertilizer to Prince William Sound beaches.

spraying of an oiled beach, for example, flushes

some surface oil back into the water, and this oil

must then be recovered by skimmers. The recovered

oil-and-water mixture must be separated, and the oil

disposed of or recycled. Also, a significant amount

of mechanical equipment and logistical capability is

required to deal with a large spill.

Because bioremediation equipment and logistics

are usually simpler and less labor intensive, costs

may be lower than for other techniques. At the same

time, the total cost of cleanup is the more important

concern, and where bioremediation is used as an

adjunct or secondary technology, total costs—as

well as total benefits--could be greater. The costs of

monitoring bioremediation must also be considered.

bioremediation technologies have several general

disadvantages. Although bioremediation may work

faster-potentially much faster—than natural biodegradation, it cannot produce significant shortterm results. If beaches are threatened by a large

offshore spill, for instance, bioremediation is probably not appropriate as an initial defensive measure.

In this circumstance, it would usually be more

appropriate to get the oil out of the water as quickly

as possible or, failing this, to disperse or burn it

before it drifts onto beaches. bioremediation takes

Gz~esea~butw ~e &SCUSSed in the Applied Biotreatment Association, ‘‘BriefmgPaperonthe Role ofbioremediation of Oil Spills,” rev. 2, August

1990.

63M.s. Foster, J*A. Tqley, ~d S.L. Dq“To Cl- or Not lb Clean: The Rationale, Methods, and Consequences of Remov@ Oti From

lkmperate Shores,” The Northwest Environwntal JournuZ, vol. 6, 1990, pp. 105-120.

~J. Gl~r, Envirorlmen~ prot~tion Agency, Risk Reduction Engineering Laboratory, personaJ co~ticatiou Feb. 20, 1991.
Bioremediation for Marine Oil Spills . 13

too much time to work as a primary response

measure for such a threat.

Second, the bioremediation approach must be

specifically tailored to each polluted site. Bioremediation technologies are not, and are unlikely soon to

become, off-the-shelf technologies that can be used

with equal effectiveness in every locale. Although

other oil spill response technologies are subject to

this same constraint, the advance knowledge needed

for bioremediation technologies is greater. Advance

knowledge of, for example, the efficiency of the

bacteria indigenous to an area in degrading oil, the

availability of rate-limiting nutrients, and the susceptibility of the particular spilled crude oil or

refined product to microbial attack is required, so

prespill planning will be important.

Finally, the public is still unfamiliar with bioremediation technologies. Although public attitudes

toward “natural” solutions to environmental problems are generally favorable, the lack of knowledge

about microorganisms and their natural role in the

environment could affect the acceptability of their

use.

65 Before bioremediation technologies are likely

to be widely used, their efficacy and safety will have

to be convincingly demonstrated and communicated

to the public.

ALTERNATIVE

bioremediatio n

TECHNOLOGIES

bioremediation technologies for responding to

marine oil spills may be divided into three discrete

categories: 1) nutrient enrichment, 2) seeding with

naturally occurring microorganisms, and 3) seeding

with genetically engineered microorganisms (GEMs)

(table 3).

Nutrient Enrichment

Of all the factors that potentially limit the rate of

petroleum biodegradation in marine environments,

lack of an adequate supply of nutrients, such as

nitrogen and phosphorus, is probably the most

important and perhaps the most easily modified.

Nutrient enrichment (sometimes called nutrition)

also has been more thoroughly studied than the other

two approaches, especially now that EPA, Exxon,

Table 3-Principal Features of Alternative

bioremediation Approaches

Nutrient enrichment:

● Intended to overcome the chief limitation on the rate of the

natural biodegradation of oil

● Most studied of the three approaches and currently seen as the

most promising approach for most types of spills

● No indication that fertilizer use causes algal blooms or other

significant adverse impacts

● In Alaska tests, fertilizer use appeared to increase biodegradation rate by at least a factor of two.

Seeding:

● Intended to take advantage of the properties of the most

efficient species of oil degrading microorganisms

● Results of field tests of seeding have thus far been inconclusive

● May not be necessary at most sites because there are few

locales where oil-degrading microbes do not exist.

● Requirements for successful seeding more demanding than

those for nutrient enrichment

● In some cases, seeding may help biodegradation get started

faster

Use of genetlcally engineered mlcroorganisms:

● Probably not needed in most cases because of wide availability

of naturally occurring microbes

● Potential use for components of petroleum not degradable by

naturally occurring microorganisms

● Development and use could face major regulatory hurdles

SOURCE: Office of Technology Assessment, 1991.

and the State of Alaska have carried out extensive

nutrient enrichment testing on beaches polluted by

oil from the Exxon Valdez. In part for these reasons,

many scientists currently view nutrient enrichment

as the most promising of the three approaches for

those oil spill situations in which bioremediation

could be appropriate.66

This approach involves the addition of those

nutrients that limit biodegradation rates (but not any

additional microorganisms) to a spill site and

conceptually is not much different than fertilizing a

lawn. The rationale behind the approach is that

oil-degrading microorganisms are usually plentiful

in marine environments and well adapted to resisting

local environmental stresses. However, when oil is

released in large quantities, microorganisms are

limited in their ability to degrade petroleum by the

lack of sufficient nutrients. The addition of nitrogen,

phosphorus, and other nutrients is intended to

overcome these deficits and allow petroleum biodegradation to proceed at the optimal rate. Experiments dating to at least 1973 have demonstrated the

potential of this approach. Researchers, for example,

have tested nutrient enrichment in nearshore areas

651bid., p. 15.

66See, for exmple, ke and Levy, op. cit., footnote 66, PP- 228-234.
14 ● Bioremediation for Marine Oil Spills

off the coast of New Jersey, in Prudhoe Bay, and in

several ponds near Barrow, Alaska. In each case, the

addition of fertilizer was found to stimulate biodegradation by naturally occurring microbial populations. 67

The recent nutrient enrichment experiments in

Alaska provided a wealth of experimental data about

bioremediation in an open environment (box B).

Since previous research findings had already demonstrated the general value of this approach, the

experiments were intended to determine for one type

of environment how much enhancement of natural

biodegradation could be expected and to evaluate

the most effective methods of application. The

results provided additional evidence that application

of nutrients could significantly enhance the natural

rate of biodegradation on and below the surface of

some beaches. As a result, Exxon was authorized by

the Coast Guard on-scene coordinator, in concurrence with the Alaska Regional Response Team,68 to

apply fertilizers to the oiled beaches in Prince

William Sound. To date, about 110 miles of

shoreline have been treated with nutrients, and a

monitoring program has been established.

Without additional research, however, it is premature to conclude that nutrient enrichment will be

effective under all conditions or that it will always

be more effective than other bioremediation approaches, other oil spill response technologies, or

merely the operation of natural processes. The

results of the Alaska experiments were influenced by

the beach characteristics (mostly rocky beaches,

well-washed by wave and tide action), the water

temperature (cold), the kind of oil (Prudhoe Bay

crude), and the type and quantity of indigenous

microorganisms in Prince William Sound.

Few detailed analyses or performance data are yet

available for different sets of circumstances. One

smaller-scale test using the same fertilizer as in

Alaska was recently conducted on beaches in

Madeira polluted by the Spanish tanker Aragon.

Results in this very different setting and with a

different type of oil were not especially encouraging.

Researchers speculated that the unsatisfactory results could have been due to differences in the type

of oil, the concentration of fertilizer used, the lower

initial bacterial activity, and/or different climatic

conditions. 69 At the same time, Exxon recently used

what it learned in Alaska to help degrade subsurface

no. 2 heating oil spilled in a wildlife refuge

bordering the Arthur Kill at Prall’s Island, New

Jersey. An innovative aspect of this application was

the use of two trenches parallel to the beach in which

to distribute fertilizer. Nutrients were dissolved with

the incoming tide and pulled down the beach with

the ebb tide, enabling a more even distribution than

point sources of fertilizer. Exxon reports that 3

months after applying fertilizers, the oil in the

treated zone had been reduced substantially relative

to that in an untreated control zone.70

Seeding With Naturally Occurring

Microorganisms

Seeding (also called inoculation) is the addition of

microorganisms to a polluted environment to promote increased rates of biodegradation. The inoculum maybe a blend of nonindigenous microbes from

various polluted environments, specially selected

and cultivated for their oil-degrading characteristics,

or it may be a mix of oil-degrading microbes selected

from the site to be remediated and mass-cultured in

the laboratory or in on-site bioreactors. Nutrients

would usually also accompany the seed culture.

The rationale for adding microorganisms to a spill

site is that indigenous microbial populations may

not include the diversity or density of oil-degraders

needed to efficiently degrade the many components

of a spill. Some companies that advocate seeding

with microorganisms also claim that commercial

bacterial blends can be custom-tailored for different

types of oil in advance of a spill, that the nutritional

needs and limitations of seed cultures are well

understood, that microbes can easily be produced in

large quantities for emergency situations, and that

6TR.M. Atlas, “bioremediation of Fossil Fuel Contamma “ ted Sites,” in press, proceedings of Battelle conference on In Situ and On-Site

BiorecZamution, March 1991. Atlas and his colleagues did some of this early work.

~~e most important members of the wka Regional Response lkam are the U.S. Environmental Protection Agency, the A.ktsh Depwtment of

Environmental Conservation the U.S. Department of the Interior, the National Oceanic and Atmospheric Administration and the U.S. Forest Service.

@M. Biscoito ~d M. Morei.rii, MUSW Municipal do Funchal, Made@ “Application of In.ipol EAP22 in Porto ?mnto,” repo~ JUIY 1990.

mP.C. Madden, ExxonResearchand Engineering Co., Ietterand KCOmpm@gs~ report onprall’s Island bioremediation to U.S. Coast Guard,

Mar. 12, 1991.
Bioremediation for Marine Oil Spills ● 15

Box B—The Alaska bioremediation Experiments

Following the March 1989 Exxon Valdez oil spill, the U.S. Environmental Protection Agency (EPA), Exxon,

and Alaska’s Department of Environmental Conservation (ADEC) undertook what is perhaps the largest and most

comprehensive series of experiments on oil spill bioremediation to date. The principal objectives of the research

initiated in May 1989 were to determine if the addition of nutrients to Alaska’s polluted beaches would enhance

the rate and extent of oil biodegradation sufficiently to support widespread use of this technology there and to

evaluate which application methods could be most effective. 1 Research begun in the summer of 1990 was designed

to evaluate the effectiveness and safety of several microbial products in cleaning Alaska’s beaches.

The Alaska bioremediation work consisted of several discrete elements, including: 1) work begun shortly after

the spill to determine if nutrient enhancement could be an appropriate technology for mitigating oil pollution of

Prince William Sound’s beaches; 2) application by Exxon of fertilizers to about 110 miles of polluted beaches, after

the initial studies suggested that fertilizers could be both effective and safe; 3) additional EPA studies to support

Exxon’s treatment program and further evaluate application techniques; 4) a long-term program to monitor treated

beaches, conducted jointly by EPA, Exxon, and ADEC; and 5) evaluation of the potential of adding microbes to

Alaska’s beaches to stimulate biodegradation.

Several types of nutrients and application methods were evaluated, including slow-release, water soluble

fertilizers in both briquette and granular forms; a water soluble fertilizer applied with a sprinkler system; and a liquid

oleophilic fertilizer,2 applied with sprayers, specially formulated to keep nutrients and oil in contact. Visual changes

between control and experimental plots were observed, and changes in oil chemistry, microbial populations, and

oil weight over time were measured. Findings pertain specifically to Prince William Sound, which has a number

of features favorable to nutrient enrichment: a high percentage of naturally occurring hydrocarbon-degrading

bacteria, low concentrations of ammonia and phosphate in seawater, highly porous beaches, and large tidal fluxes.

Although the work carried out in Alaska is important to bioremediation research and applications in other areas,

the same results cannot be expected elsewhere. The major findings follow:

Based on a synthesis of all available evidence, researchers concluded that biodegradation on beach surfaces

was accelerated as much as two- to four-fold by a single application of fertilizer; thus, the addition of

nutrients to Alaska’s beaches did significantly

stimulate the rate of biodegradation. The water

soluble fertilizer delivered by a sprinkler system

proved the most effective approach, but this

method was impractical on a large scale. The

oleophilic fertilizer and slow-release granules

were almost as effective and more practical to use.

EPA determined that the most practical approach

for this setting was to apply the oleophilic

fertilizer to beaches with surface oiling and to use

both oleophilic fertilizer and fertilizer granules

where surface and subsurface oil were found.

After several weeks, dramatic visual changes were

observed in the amount of oil on beaches treated

with fertilizer. Visual changes do not provide

quantitative data or prove that biodegradation is

occurring. However, similar changes observed in

beaches treated with the “plain” water soluble

fertilizer and those treated with oleophilic fertilizer provided evidence that visually cleaner beaches

Photo credit: Environmental Protection Agency

Visual effect of oleophilic fertilizer on the

biodegradation of surface oil in Snug Harbor, Alaska.

The clear “window” indicates where the fertilizer

was applied.

Iu.s. Environmen~ protection Agency, office of Research and Develo~men~ “InterimRerm_t: Oil SDill bioremediation Project+’ Feb.

28, 1990, 220 pp. See also P.H. Pritch&d ad C.F. Costa, “EPA Alaska Oil Spill Bioremed&tion proj~c~” Environmental S;ience and

Technology, March 1991, pp. 372-379. Much of the material in this box was reported in these two citations.

2For more on oleopwc fertilizers see A. Ladousse and B. Tramier, Societe Nationale ELF AQUIT_, “Results of 12 Yem of

Research in Spilled Oil bioremediation: Inipol EAP 22,” 1990.

Continued on next page
16 ● Bioremediation for Marine Oil Spills

Box B—The Alaska bioremediation Experiments-Continued

resulted directly from the addition of nutrients, not from the suggested rock washing effects of oleophilic

fertilizers. The oleophilic fertilizers evidently worked as intended, sequestering nutrients at the oil-water

interface where microorganisms could be effective.

. Changes in oil chemistry provided additional evidence that biodegradation was limited by lack of nutrients.

Analysis of fertilizer-treated samples and samples that had been artificially weathered to control for

evaporation indicated that many of the easily degradable constituents of petroleum in fertilizer-treated

samples had decreased substantially over 4 weeks. Some harder-to-degrade fractions of oil also appeared

to decrease. No conclusions could be drawn about other difficult-to-degrade fractions because good

measurement methods had not been devised for them.

• In early testing, statistically significant increases in the oil degrading microbial population-increases that

would correlate strongly with the rate of biodegradation-were not observed because of the high variability

in numbers of bacteria in each sample. However, later results from the joint monitoring program appeared

to indicate a sustained three- to four-fold increase in microbial activity.

• No statistically significant conclusions could be made about the rate of biodegradation from “before and

after” measurements of the weight of oil samples. Although a significant decrease in oil residue weight over

time would be evidence of degradation, this might not be a good criterion because bacterial production of

high-molecular-weight compounds can occur. Precise measurements were impossible because oil was

distributed unevenly on the beaches. Although the rate of biodegradation was probably the same in all areas,

samples from more heavily oiled areas indicated a slower rate of biodegradation than samples from lightly

oiled areas.

. The monitoring program indicated that enhanced microbial activity could be sustained for more than 30 days

from a single fertilizer application. Additional applications were found to increase microbial activity. In

particular, a second application of fertilizer after 3 to 5 weeks replenished nutrients and stimulated microbial

activity five- to ten-fold.3

. Fertilizer applications appeared to enhance biodegradation to a depth of at least 50 centimeters on treated

beaches. Researchers found increased nitrogen nutrients, sufficient dissolved oxygen, and increased

microbial numbers and activity at this depth following treatment.4

. Although evidence was not conclusive, researchers suspected that primary treatment with mechanical

methods resulted in a more even distribution of oil on the beaches and hence prepared the beaches for

optimum bioremediation.s

. Results of the 1990 research on two microbial products were inconclusive, with no statistically significant

enhancement of the rate of biodegradation over natural rates. However, the tests were conducted on oil that

had weathered and degraded naturally during the 18 months since the Exxon Valdez spill. The more easily

degraded components of the oil had already disappeared. The limited testing period-27 days-may also

have affected the results.6

SR. prince, J. Clark and J. Lindstrom, ‘‘bioremediation Monitoring Program,’ December 1990, pp. 2,85. The authors of this report

represent EXXOU EPA, and the Alaska Department of Environmental Conservation respectively.

‘%id.

5wtc~d and COS@ op. cit., footnote 1.

6A. Venos% EPA, Risk Reduction Engineering Laboratov, presentation of research results at EPA-sponsored bioremediation meeting,

Las Vegas, NV, Feb. 20, 1991.

seed cultures can be stored, ready for use, for up to studies have not been conducted in such settings, so

3 years.71 no data are available to evaluate the effectiveness of

The value of introducing nonindigenous micro- this approach. Many scientists question the necesorganisms to marine environments is still being sity of adding microbes to a spill site because most

evaluated. With some exceptions, the scientilc locales have sufficient indigenous oil-degrading

community has not been encouraging about the microbes, and in most environments biodegradation

promise of seeding marine oil spills. Controlled is limited more by lack of nutrients than by lack of

71Appfi~ Biotreatment ~sociatio~ Op. Cit., fOOtnOte % Pp. 13-14.
Bioremediation for Marine Oil Spills . 17

microbes.72 At many spill sites, a very low level of

oil is often present as ‘‘chronic” input, inducing

oil-degrading capability in naturally occurring microorganisms. Moreover, the requirements for successful seeding are more demanding than those for

nutrient enrichment. Not only would introduced

microbes have to degrade petroleum hydrocarbons

better than indigenous microbes, they would also

have to compete for survival against a mixed

population of indigenous organisms well adapted to

their environment. They would have to cope with

physical conditions (such as local water temperature, chemistry, and salinity) and predation by other

species, factors to which the native organisms are

likely to be well adapted.

The time required for introduced microbes to

begin metabolizing hydrocarbons is also important.

If a seed culture can stimulate the rapid onset of

biodegradation, it would have an advantage over

relying on indigenous microbes that may take time

to adapt. Despite some claims, seed cultures have

not yet demonstrated such an advantage over indigenous microbial communities. Seed cultures are

typically freeze-dried (and therefore dormant) and

require time before they become active.73 Seed

cultures also must be genetically stable, must not be

pathogenic, and must not produce toxic metabelites. 74

Some laboratory and small-scale experiments in

controlled environments have demonstrated that

seeding can promote biodegradation.75 However, it

is exceedingly difficult to extrapolate the results of

such tests to open water where many more variables

enter the picture. Results of experimental seeding of

oil spills in the field have thus far been inconclusive.76 As noted in box B, recent EPA tests of two

commercial products applied to contaminated beaches

in Alaska concluded that, during the period of

testing, there was no advantage from their use.77 In

a well-publicized attempt to demonstrate seeding at

sea, one company applied microorganisms to oil

from the 1990 Mega Borg spill in the Gulf of

Photo credit: National Oceanic and Atmospheric Administration

Application of a microbial product to Marrow Marsh in

response to an oil spill caused by the collision of the Greek

tanker Shinoussa with three barges in the Houston

Ship Channel.

Mexico. 78 Although the experiment aroused some

interest, the results were inconclusive and illustrated

the difficulty of conducting a controlled bioremediation experiment at sea and measuring the results.

Although there were changes observed in the seeded

oil, in the absence of controls the experiment could

not tell whether they were due to biodegradation or

bioemulsification (the process in which microbes

assist the dispersal of surface oil), or were unrelated

to the seeding. (Even if bioemulstilcation rather than

biodegradation was the process at work in this

experiment, it may be of potential interest for oil

spill response and could be investigated further.)

An attempt has been made to apply a seed culture

to a polluted salt marsh. In July 1990 the Greek

tanker Shinoussa collided with three barges in the

Houston Ship Channel, resulting in a spill of about

700,000 gallons of catalytic feed stock, a partially

refined oil. Some of this oil impacted neighboring

Marrow Marsh. Microbes were applied to experimental areas within the marsh, and control areas

were established. Visual observations made by the

scientific support coordinator who monitored the

Tz~e ~d LRVY, op. cit., footnote 22, p. 229; see also Atlas, op. cit., footnote 23, p. 218.

TSR-M. Aflas, Dep~ent of Biology, University of Louisville, p~sorMI COmm~~tiOm NOV. 29, 1990.

T4R.M. Aflas, “Stim~ated Petroleu Biode~datioq ” critical Reviews of Microbiology, VOI. 5, 1977, pp. 371-386.

TSSee, for ex~ple, lkxas General kdOff% “Combating Oil Spills Along the lkxas Coast: A Report on the Effects of Bioremediatiou” June

12, 1990; see also hxihy and Colweil, op. cit., footnote 28, p. 311.

76p.H. fitc~d ad CF. Cosa ~~EpA ~m~ oil Spfil bioremediation ~o~~’Environmenta/Sciencea~Technology, mch1991, pp. 372-379.

77E. Berkey, Natio~ Environment Twhnology Applications Corp., persoml COmmticatiO14 Feb. 15, 1991.

7SII=.. Gene~ Laud office, Mega Borg Oil Spill: An Open Water bioremediation Test, JtiY 12, 1990.
18 ● Bioremediation for Marine Oil Spills

application for the National Oceanic and Atmospheric Administration (NOAA) indicated that treated

oil changed color within a few minutes to a few

hours after treatment, but that after several days

there were no significant visual differences between

treated and untreated plots. More importantly, chemical analyses indicated “no apparent chemical

differences in petroleum hydrocarbon patterns between treated and untreated plots several days after

treatment. ’79 Not all of the monitoring data have

been analyzed yet, so a final determination of

effectiveness has not been made.

Seed cultures may be most appropriate for situations in which native organisms are either present as

slow growers or unable to degrade a particular

hydrocarbon. Especially difficult-to-degrade petroleum components, such as polynuclear aromatic

hydrocarbons, might be appropriate candidates for

seeding. 80 In other cases, if a time advantage can be

realized, there may be some utility in seeding with

a culture consisting of indigenous organisms.81

Thus, the potential environmental adaptation problems of nonindigenous organisms might be avoided.

In many cases, fertilizers would also have to be

“added.

Seeding may offer promise in environments

where conditions can be more or less controlled. In

such cases one would have to consider the proper

choice of bacteria, a suitable method of application,

and suitable site engineering. Arrangements would

have to be made for keeping cells moist and in

contact with the oil; for protecting them from excess

ultraviolet light; for providing adequate nutrients;

and for controlling temperature, pH, and salinity.

However, before claims about the utility of seeding

marine oil spills can be proved (or disproved),

additional research-verified by repeatable experiments—is required.

Seeding With Genetically Engineered

Microorganisms

Although it was not demonstrably superior to

indigenous organisms and has never been tested in

the field, the frost organism ever patented was a

microorganism genetically engineered to degrade

oil.82 The rationale for creating such organisms is

that they might possibly be designed either to be

more efficient than naturally occurring species or to

have the ability to degrade fractions of petroleum not

degradable by naturally occurring species. To be

effective, such microorganisms would have to overcome all of the problems related to seeding a spill

with nonindigenous microbes.

EPA has not yet conducted any GEM product

reviews for commercial applications, although at

least two companies are considering using genetically engineered products for remediating hazardous

waste. Since the development and use of GEMs are

still limited by scientific, economic, regulatory, and

public perception obstacles, the imminent use of

bioengineered microorganisms for environmental

cleanup is unlikely. Lack of a strong research

infrastructure, the predominance of small companies

in the bioremediation field, lack of data sharing, and

regulatory hurdles are all barriers to the commercial

use of genetically engineered organisms.83 The

development of GEMs for application to marine oil

spills does not have high priority. Many individuals,

including EPA officials, believe that we are so far

away from realizing the potential of naturally

occurring microorganisms to degrade marine oil

spills that the increased problems associated with

GEMs render them unnecessary at this time.84

ENVIRONMENTAL AND

HEALTH ISSUES

To date, no significant environmental or health

problems have been associated with the testing or

application of bioremediation technologies to ma79AJ. Me~, Mder, BioAssessment ‘lkauL National OCtiC ~d Amosphtic~“ “stratioq “Observations of An Oil Spill bioremediation

Activity in Galveston Bay, ‘I&as,” Marcm 1991.

80Atlas, “bioremediation of Fossil Fuel Contamma “ ted Sites,” op. cit., footnote 67.

81R. Colwell, OTA bioremediation Workshop, DCC. 4, 1~.

82&.e, for eqle,D,A. FrieUo, J*R. My~oie, ad J*M.-ab@,“use of ~neti~lyEnginmredMultip~smid Microorganisms for Rapid

Degradation of Fuel Hydrocarbons,”Biodeterioration of Maten”als, No. 3, 1976, pp. 205-214.

83u.s0 congas, Office of ~~olo= As~ssment, Biotechnology in a G/o~z Economy (wa-o~ ~: I-J.s. Government Prhlt@ ~~,

scheduled for publication summer 1991).

~AoW. Lind=y, Director, Offlce of Environment Engineer@ ~d ~~olo~, U.S. Environment@ ~t~tion Agency, PtXsolld COInmUrd~tiO~

September 1990.
Bioremediation for Marine Oil Spills ● 19

rine oil spills. Experience with bioremediation in

marine settings is still limited, so it is premature to

conclude that its use will always be safe or that

possible risks will be acceptable in all of the

circumstances in which bioremediation might be

employed. The evidence to date, nevertheless, suggests that risks will be unimportant in most situations.

Concerns have been raised about several potential

adverse environmental effects. Among these are the

possibility that the addition of fertilizers could cause

eutrophication, leading to algal blooms and oxygen

depletion; that components of some fertilizers could

be toxic to sensitive marine species or harmful to

human health; that the introduction of nonnative

microorganisms could be pathogenic to some indigenous species; that the use of bioremediation technologies could upset ecological balances; and that

some intermediate products of bioremediation could

be harmful.

The possible adverse effects of nutrient enrichment were examined in some detail during the

1989-90 Alaska beach bioremediation experiments.85

To determine the potential for eutrophication in

Prince William Sound, researchers measured ammonia, phosphate, chlorophyll, bacterial numbers, and

primary productivity in the water column directly

offshore of fertilizer-treated beaches and in control

areas. They could find no significant difference

between measurements in control areas and those in

experimental areas.86 There were no indications that

fertilizer application stimulated algal blooms.

The possible toxicity of fertilizer components was

examined in both laboratory and field tests on a

number of marine species, including sticklebacks

fish, Pacific herring, silver salmon, mussels, oysters,

shrimp, and mysids. In the absence of tidal dilution,

certain components of the oleophilic fertilizer were

mildly toxic to oyster larvae, the most sensitive

marine species.87 However, in the view of researchers working in Alaska, such effects were transient

and limited to areas immediately adjacent to fertilized shorelines.88 The concentration of ammonia,

the only component of fertilizers shown to be

acutely toxic to marine animals, never reached toxic

levels.

The butoxyethanol constituent of the oleophilic

fertilizer is potentially harmful to some mammals.

This constituent, however, evaporated from beach

surfaces in less than 24 hours, during which time

wildlife deterrent devices were employed. Care had

to be taken, as well, by humans applying the

oleophilic fertilizer to avoid inhalation or skin

contact. Researchers were also able to show that the

oil itself did not wash off the treated beaches and

gssee, for ex~ple,U.S.Environmen~ Protection Agency, OffIce of Research and Development Interim Report: Oil Spill bioremediation Prqkt,

Feb. 28, 1990; R.C. Prince, J.R. Clark, and J.E. Lindstrom, bioremediation Monitoring Program, December 1990; and P.H. Pritchard and C.F. COS@

“EPA Alaska Oil Spill bioremediation project” Environmental Science and Technology, March 1991.

861bid.

8Tu.s. Environmental Protection Agency, “Alaskan Oil bioremediation Project: Update,” EPA/600/13-89/073, July 1990, p. 11.

gg~ce et d., op. cit., footnote 85, p. 72.
20 ● Bioremediation for Marine Oil Spills

accumulate in the tissues of marine test species. In

this environment, dilution, tidal mixing, and evaporation reduced the potential for significant impacts.

In some low-energy environments (e.g., protected

bays), greater impacts might occur. In other environments, the species present, water depth, and water

temperature are all variables to consider in estimating potential impact. The effect of any impact of

treatment, however, must be considered in view of

the damage already caused by oil.

Evidence is also lacking that introduced organisms might be pathogenic to other life forms. In a

series of experiments with North Slope crude oil, for

example, researchers failed to find any significantly

greater invertebrate mortality with bacterial seeding

(or fertilization) than occurred with crude oil alone.89

However, microorganisms to be considered as

seeding candidates must be screened carefully to

eliminate potential human or animal pathogens,

including opportunistic pathogens such as Pseudomonas spp.

The possibility that introduced microbes might

proliferate and upset the ecological balance appears

to be of less concern. If effective at all, such

organisms should die and be preyed on by protozoa

once they have utilized the oil from a spill.90 Of

greater concern is that microbes introduced from

other environments will not be able to compete as

well as native species and will die before they can do

their job effectively. EPA’s Office of Pesticides and

Toxic Substances has been developing procedures

for evaluating the toxicity of biotechnology products. In concert with EPA’s Office of Research and

Development, it is establishing tests to evaluate the

potential pathogenicity of nonindigenous microbes.91

Similar but greater concerns attend the introduction of genetically engineered organisms. Before

such organisms are likely to be introduced to the

marine environment (if they have a role to play at

all), more basic knowledge of their potential impacts

on that environment will be required, and regulatory

officials and the public will have to become more

familiar with biological mitigation techniques.

An additional concern is that although bacteria

may break down the complex hydrocarbons contained in oil, they could leave behind products of

partial biodegradation that are more toxic to marine

life than the original constituents of the oil. 92

However, in time, intermediate products of possible

concern, such as quinones and naphthalenes, are

likely to be broken down further and thus unlikely to

accumulate in the environment.93

bioremediation IN RELATION

TO OTHER RESPONSE

TECHNOLOGIES

Whether bioremediation technologies will be

considered for use as primary or secondary response

tools, or will be deemed of no use at all, will depend

on the circumstances of each oil spill. All response

technologies have the common purpose of minimizing the damage caused by a spill. How well a

technology can accomplish this goal indicates its

effectiveness. The perfect response technology has

not been developed, and numerous uncontrollable

variables may reduce effectiveness far below what it

would be under optimal circumstances. As the size

of a spill increases, for example, the difficulty of

responding to it by any means grows. Adverse sea

and weather conditions may greatly reduce the

effectiveness of any open sea response. Even technologies that are adequate in some spill situations

will be much less effective if they cannot be

deployed, operated, and maintained easily.

Before bioremediation is likely to be considered

as a response tool, it must be deemed not only

effective for its intended use, but also more effective

than traditional technologies. The effectiveness and

safety of bioremediation technologies for responding to different types of spills have not yet been

established adequately, and on-scene coordinators

and other decisionmakers generally are not familiar

with these technologies. Hence, most decisionmakers are reluctant to try bioremediation if other

techniques could be effective. More traditional

methods are preferred, and experimentation during

WR.M. Atlas and M. Busdos@ ‘hlicmbia.l Degra&ionof pdroleum hthi? ArCtiC,” Proceedings of the3rdInternational Biodegr&tion Symposium,

Kingston, RI, August 1975, p. 85. The seeding trials used an organism from the environment being treated.

%. Colwell, Maryland Biotechnology Institute, University of Maryland, personal communicatio~ February 1991.

glu.s. Environmen~ prot~tion Agency, Office of Res~h and Development, bioremediation Action Committee, ‘ ‘SummarY of June 20, 1990

Meeting,” Jtiy 1990.

~M.c. ICenni~~ Geochemical and Environmental Research Group, ‘lkxas A&M University, pfxsOd cOmmurdCatiO%@tok1990.

93R.M. Aflas, Dep~~t of Biology, University of Louisville, personal Communication February 1991.
Bioremediation for Marine Oil Spills ● 21

a real spill is not the best strategy. The acceptability

(or nonacceptability) of bioremediation technologies may take time to establish. The example of

dispersant development may provide a relevant

analogy for bioremediation in the immediate future:

Dispersants have been advocated by many and

considered for more widespread use for years, but

uncertainty about their effectiveness, as well as

continuing environmental concerns and regulatory

limitations, has considerably slowed their acceptance in the United States. Probably the most serious

setback was the fact that a few early dispersants were

toxic; however, since the frost attempts to formulate

successful dispersants, toxicity has been addressed

and, in some cases, is no longer a serious factor.

bioremediation technologies have been considered for use at sea, on beaches, and in especially

sensitive habitats such as salt marshes. It is unclear

whether bioremediation could be useful on the open

ocean. Most of the scientific community and many

oil spill professionals remain skeptical about the

utility of bioremediation at sea because rigorously

controlled and documented experiments have not

yet been done. Several companies have advocated

using bioremediation for open ocean spills, but they

have not yet produced convincing evidence that their

products work as claimed.

In addition to the previously noted problems

associated with seeding, a potentially significant

problem at sea may be the difficulty of keeping

microbes or nutrients in contact with spilled oil long

enough to stimulate degradation.94 This is afar more

difficult task than on land or beaches because wind,

waves, and currents create a dynamic, changeable

environment. As with dispersants, efficient application could also be difficult because ocean conditions

are often less than ideal, and the oil may be difficult

to locate, may be emulsified or broken into windrows, or in a major spill, may have spread over many

square miles. These same problems also limit the

effective use of booms and skimmers.

The probability is low that a response with any

type of technology would be mounted for a spill far

out to sea that does not threaten the coast. The initial

goal in responding to spills that do threaten the coast

is to prevent oil from reaching the shore. Thus,

unless seeding, nutrient enrichment, or both can be

shown to act quickly and can be applied efficiently,

spill fighters typically would prefer to get the oil out

of the water as quickly as possible or to disperse it.

If open sea bioremediation can be shown to be

effective over a longer period (i.e., weeks), it might

be useful as a response technology either after a

full-scale mechanical or dispersant effort had been

launched or if such an effort was not possible.

Conceivably, nutrients or a seed culture might also

be applied to the oil and its residues remaining in the

water after the intentional (or unintentional) burning

of oil. bioremediation is less likely to be attempted

following the use of chemical dispersants; however,

there is evidence that some microorganisms may

stimulate bioemulsification and thus cause oil to

disperse upon application. This possibility and the

possible merits and limitations associated with it

have not been investigated thoroughly.

Although bioremediation at sea has not been

convincingly demonstrated to be effective in any

type of spill situation, alternative response technologies leave much to be desired. bioremediation,

although unproven, appears relatively promising to

some. The State of Texas, for example, has been

particularly enthusiastic about its potential. Texas

has taken the position that there is little to lose by

trying it and, given the limitations of other technologies, potentially much to gain or, at least, to learn.95

The Texas State Water Commission points out that

those responsible for responding to an emergency

may not be able to wait for a definitive unambiguous

ruling from the scientific community on the effectiveness of bioremediation. The Federal Government and other States have been more cautious. The

State of Alaska, for example, has tentatively concluded that bioremediation would not be appropriate

as an emergency response tool for nearshore spills

threatening the coast.96 In any case, controlled

testing, difficult to conduct on the open ocean, will

be required to evaluate the potential of bioremediation at sea. ~~bioremediation shows promise for at

least some types of open water spills, effective

application techniques would still have to be developed for the promise of the technology to be

fulfilled.

~R.M, A~as, Dep~ent of Biology, University of Louisville, personal COmmti~tiOt4 Nov. 29, 199Q

95B.J. Wynne, Chamnzq “ lkxas Water Commission statement at OTA Workshop on bioremediation, Dec. 4, 1990.

%E. P@r, ~ka Shte ~.sc~e Coor-tor, E.zxon VaZdez Oil Spill Response. Letter to ~XaS Water COmm.iSSiOU @t. 19, 19~.
22 ● Bioremediation for Marine Oil Spills

Somewhat more is known about the potential for

bioremediation of beaches, thanks largely to the

Alaska experiments. The potential use of bioremediation methods on at least some beaches looks

promising, but, as previously noted, results from the

Alaska experiments cannot be extrapolated in toto to

other types of beaches or spill situations (especially

without more precise knowledge of the effect of

environmental and microbiological variables on the

rate and extent of biodegradation).

The greater promise of bioremediation for oiled

beaches is due in part to the fact that bioremediation

is more easily controlled and monitored onshore

than it is at sea. Application of nutrients or seed

cultures is also easier and less subject to disruption

by adverse conditions. Also, once oil reaches a

beach, there is usually more time to consider the

approach to take: certain damage has already

been done, and the emergency response required to

deal with oil seeping from a stricken tanker, for

example, is no longer quite so necessary.

Depending on circumstances, bioremediation of

beaches may be appropriate sometimes as a primary

response tool and sometimes as a secondary tool. An

important consideration is how heavily oiled a beach

is. Heavily oiled beaches may require removal of

gross amounts of oil by mechanical means before

bioremediation can be a practical finishing tool.

Some lightly oiled beaches may not require any

treatment. Moderately oiled beaches are likely the

main candidates for primary bioremediation treatment. After reviewing the Alaska bioremediation

experiments, the Alaska Department of Environmental Conservation concluded that bioremediation

was useful as a finishing or polishing tool, but that

pooled oil, tar balls, mousse, asphalt, and other

heavy concentrations of oil should be picked up with

conventional manual and mechanical techniques.97

The effectiveness of bioremediation on beaches

may also depend on the coarseness of the beach, but

the relationship between the size of oiled sediments

and the rate of biodegradation has not been evaluated thoroughly. The beaches treated in Alaska with

some success were all very coarse, consisting mostly

of cobbles and coarse sand. It is uncertain how

successful bioremediation of freer grained beaches

will be, especially when oil is trapped below the

surface. In very fine-grained sediments, lack of

oxygen below the surface may limit the rate of

biodegradation.

bioremediation, where effective, may offer a

promising option for beach cleanup because the

existing mechanical technologies can cause additional damage to beaches and beach biota.98 This

damage may be unavoidable if the goal is to

“restore” a beach as quickly as possible. Doing

nothing (i.e., letting the beach recover naturally at a

slower rate) may sometimes be preferable to using

mechanical technologies, but this is seldom politically acceptable. bioremediation offers the possibilities of being faster than simply allowing nature to

take its course unassisted and of avoiding the

negative impacts of mechanical technologies. Moreover, when beaches are inaccessible, the mechanical

equipment that can be brought into use may be

limited, but fertilizers or seed cultures can be

dispensed without the need for massive machinery.

In general, bioremediation is less costly and less

equipment- and labor-intensive than mechanical

cleanup technologies, which suggests a clear advantage for bioremediation where it can be used

effectively instead of other technologies (e.g., moderately oiled beaches). The advantage is also evident

where it can be used as a secondary technology (i.e.,

as a finishing tool), because it offers the possibility

of a more complete solution, more quickly attained

(however, the total cost of the cleanup may be

greater).

Salt marshes and other sensitive environments,

even more than beaches, may be further damaged by

intrusive mechanical technologies. For these environments, bioremediation could be the only feasible

alternative to doing nothing. Little work has been

done in these settings to evaluate the effectiveness or

environmental impacts of using bioremediation.

Biodegradation of oil stranded in salt marshes is

generally limited by oxygen availability. However,

the results of one recent study of waxy crude oils99

in salt marshes suggest that nutrient enrichment may

be an effective countermeasure, provided that large

%’~id.

98M.S. Foster, J.A. Tarpley, and S.L.D- “To C!leaU or Not lb Clean: The Rationale, Methods, and Consequences of Remov@ Ofi From

‘Rznperate Shores,” The Northwest Environmental Journai, vol. 6, 1990, pp. 105-120.

~axycmde oils tend to spread, evaporate, and naturally disperse very slowly on water, and usually survive in a relatively fresh state considerably

longer than conventional oils.
Bioremediation for Marine Oil Spills ● 23

amounts of oil do not penetrate beneath the aerobic

surface layer. Where oil did penetrate the surface

layer, the researchers observed little degradation.l00

Even bioremediation activities, if they are not

carefully conducted, have the potential for being

intrusive in salt marshes. The scientific support

coordinator of the National Oceanic and Atmospheric Administration who monitored the application of microorganisms to Marrow Marsh in the

Houston Ship Channel noted, for example, that

excessive foot traffic associated with bioremediation operations caused some unnecessary damage to

marsh grass.lO1

bioremediation ACTIVITIES IN

THE PUBLIC AND PRIVATE

SECTORS

The Environmental Protection Agency

EPA is the lead Federal agency for oil spill

bioremediation research. Both the Department of

Energy and the Department of Defense are actively

engaged in research on bioremediation of hazardous

waste, but neither these nor other Federal agencies

are engaged in research on bioremediation of marine

oil spills. EPA regards biotechnologies as having

significant potential for the prevention, reduction,

and treatment of pollution, and the Agency has

placed considerable emphasis on the demonstration

and development of these technologies.102 This

coincides with an important general EPA goal of

promoting the development of new and innovative

technologies to address environmental problems.

Agency activities in support of bioremediation for

marine oil spills represent a small fraction of its

overall biotechnology activities. Nevertheless, EPA

would like to establish the technical basis for a

national bioremediation response capability for oil

spills. l03

In February 1990, EPA convened a meeting of

interested industry, academic, and government personnel “to prepare an agenda for action” for

increasing the use of biotechnology. One important

outcome of this meeting was the formation of the

bioremediation Action Committee (BAC). The

objective of the Committee is to facilitate the safe

development and use of biotechnology as a solution

to environmental problems. The BAC has now been

subdivided into six subcommittees: Oil Spill Response, Treatability Protocol Development, Research, Education, Data Identification and Collection, and Pollution Prevention (figure 2). Several of

these subcommittees have, in turn, been further

subdivided. All subcommittees report to the assistant administrator of EPA’s Office of Research and

Development (ORD), who functions as the chair of

the BAC. Many of the same industry, academic, and

governmental representatives who attended the February meeting are participants on the BAC or its

subcommittees.

The Oil Spill Response Subcommittee is concerned directly with the bioremediation of marine oil

spills. Its major goals are: 1) to evaluate scientific

and applied engineering data on the safety and

effectiveness of bioremediation technologies; 2) to

assess the information required for bioremediation

decisionmaking by Federal on-scene coordinators

and State oil spill response officials; 3) to prepare

interim guidelines on when and how to use bioremediation technologies; and 4) to investigate longerterm issues for incorporating bioremediation into the

National Spill Response Plan.l04 An eventual result

of deliberations relating to these goals could be the

design of a national bioremediation oil spill response

plan.105 However, further research and development

of more reliable technologies are required before

EPA is likely to undertake the effort required to

develop such a plan. In the meantime, the Subcommittee has prepared interim guidelines to assist

Regional Response Teams in assessing the desirabilIm. ~e ~d E.M. tivy, “Bior~e&tion: Waxy Crude Oils Stranded on bw-13nergy Shorelties, ” Proceedings: 1991 Oil Spill Conference, San

Diego, CA, Mar. 4-7, 1991.

101 M-, op. cit., footnote 79.

10ZU.S. Environmental Protection Agency, “SUUMWUY Report on the EPA-Industry Meeting on Environmental Applications of Biotechnology,”

Crystal City, VA, Feb. 22, 1990.

103u.s. Enviromn~M Protection Agency, bioremediation #Wion Committee, “S~ of June 20, 1990 Meeting,” p. 3.

l@U.S. Env~omenM fiotwtion Agency, officeofReSe=h~ dDevelopment, Bior~@tionAction Committee, “S~ of November7, 1990

Meeting,” Washingto~ DC, Nov. 7, 1990, pp. 4-7.

105F.M. Gregorio, memkr, Oil Spill Response SubCommittee, “Development of a National bioremediation Spill Response Plan,” Aug. 27, 1990.
Figure 2--Organization of the bioremediation Action

~

Full

committee

I

Pollution Oil

prevention spill response I Research T

Treatability

protocol

I I

,

EIEIECIEEESI

SOURCE: U.S. Environmental Protection Agency.

T

I 9

User Contributor

issues issues
bioremediationfor Marine Oil Spills ● 25

ity of bioremediation and in planning for its use.l06

Such guidelines will enable decisionmakers to make

quick and defensible decisions about the use of

bioremediation technologies.

The Treatability Protocol Development Subcommittee has focused its attention on providing technical advice on the development of protocols for

testing the applicability and effectiveness of bioremediation technologies in different environmental

settings. Protocols use both chemical analyses and

bioassays to evaluate a bioremediation product’s

ability to degrade a waste product or pollutant and to

ensure that the product is safe to introduce into the

environment. 107 Performance criteria included in the

protocols can provide a standard for technology

developers against which they can compare their

processes.

The development of oil spill protocols is a

high-priority EPA activity: without them it is not

possible to validate technology or process claims

made by product vendors. This activity is being

carried out largely through the National Environmental Technology Applications Corp. (NETAC)

and EPA’s ORD labs (see below). One laboratorybased protocol has already been developed and used

to evaluate products intended for use on Alaska’s

beaches. Work is in progress on the development of

an open water protocol, as well as on protocols for

sensitive marine environments (e.g., marshes). The

Subcommittee will also be involved in developing

protocols for bioremediation of hazardous wastes.

The Treatability Protocol Development Subcommittee is also addressing several policy issues

related to protocols. For example, bioremediation

companies are concerned about having products

retested that did not do well initially. Another issue

is the recourse available to companies that disagree

with test procedures. A third is the means by which

results will be reported. (Results will probably be

reported as statistically superior to, not statistically

different from, or statistically inferior to a standard;

products are unlikely to be ranked. Also, products

will be judged on different criteria, including

efficacy, shelf life, toxicity, etc. A product that does

well according to one criterion may not do well on

another.) Finally, there is the question of who pays

for product testing. EPA appears receptive to some

cost sharing with product developers, but only for

those products that have met minimum criteria. 108 A

cost sharing program involving EPA, the petroleum

industry, and product vendors might also be arranged so that a broad commercial testing program

could be established.l09

The main objective of the Research Subcommittee is to identify high priority needs for general

bioremediation research,l10 not specifically for oil

spill applications, but research advances in priority

areas could directly or indirectly benefit the latter.

The Education Subcommittee will evaluate future

needs for scientists, technicians, and engineers in

bioremediation research and applications, as well as

ways to educate the public about the use of

biotechnologies.

Although considerable bioremediation information has been generated by industry, States, and

Federal agencies, this information is not necessarily

easily accessible, nor has it been certified or

standardized for easy use by others. The Data

Identification and Collection Subcommittee will

focus its efforts on identifying data on field applications and tests of bioremediation technologies (including marine spill applications such as those in

Alaska and Texas), on providing guidance for

making data available without compromising client

or proprietary information, and on establishing

routine procedures for submission of data. The

Subcommittee has recommended that EPA’s Alternative Treatment Technology Information Center

(ATTIC) database be used as the central database for

all biological treatment technologies. Designed primarily as a retrieval network for information on

innovative technologies for treating hazardous

wastes, ATTIC should have no trouble incorporating

l~u.s. Environmen~ prot~tion Agency, Subcommittee on National bioremediation Spill Response, bioremediation ACtiOn committee, ‘ ‘rnterim

Guidelines for Preparing bioremediation Spill Response Plans” (draft), Feb. 11, 1991.

~07E.B. Bmkey, NatiO@ Environmental lkchnology Applications COW, “Presentation of the Prognxs of the Treatability Protocol Development

Subcommittee,” Washington, DC, Nov. 7, 1990.

IOSRe~ks of J. Skinner, Deputy Assistant ~“ “strator of the Office of Research and Developmen~ U.S. Environmental Protection Agency, during

Nov. 7, 1990 meeting of the bioremediation Action Committee.

109s+ L@le, Depu&D&tOr, ~lce of Envfi~enW Engineering and ‘IkchnologyDemonstratio~ U.S. Environmental Protection Ag~cy, P~on~

communication Feb. 20, 1991.

11 OU.S. Environment Protection Agency, op. cit., footnote 103.
26 ● Bioremediation for Marine Oil Spills

data on the applications of bioremediation for

marine oil spills.lll

The Pollution Prevention Subcommittee is the

newest BAC committee. Its purpose is to provide

advice to EPA on the potential of biotechnologies

for preventing pollution. An example of a pollution

prevention application for biotechnology already in

use is the treatment of oily ballast water from ships

in onshore biological treatment facilities before

releasing it to the ocean. This practice is followed,

for instance, in Prince William Sound.

A number of EPA laboratories have contributed to

the Agency’s bioremediation research effort. Prominent among these are the Environmental Research

Laboratory in Gulf Breeze, Florida; the Risk Reduction Engineering Laboratory in Cincinnati, Ohio; the

Environmental Research Laboratory in Athens, Georgia; and the Environmental Monitoring Systems

Laboratory in Las Vegas, Nevada. Table 4 indicates

some key bioremediation research needs.

Prior to the Exxon Valdez oil spill, EPA had

virtually no money for oil spill research. Shortly

after the March 1989 spill, EPA and Exxon signed

a cooperative research and development agreement

and initiated the Alaskan Oil Spill bioremediation

Project (described above). During 1989, EPA redirected about $1.6 million to the project and Exxon

contributed about $3 million. In 1990, Congress

appropriated $1 million to EPA for oil spill research,

which was applied entirely to the continuing Alaskan project. As in 1989, Exxon contributed about

twice as much. To date, about $8 million has been

devoted to the Alaskan project, and 1991 will

probably be its last year.

Congress appropriated $4 million to EPA for oil

spill research activities for fiscal year 1991. EPA

expects to spend roughly $2 million of this, not

including salaries, for bioremediation research. Beginning in 1992, the Oil Pollution Act of 1990

authorizes a maximum of approximately $21 million

annually for oil pollution research and development.

The exact amount must be approved by Congress

each year, and much less could be appropriated. EPA

has asked for $3.5 million of funds available through

OPA for fiscal year 1992, a sizable proportion of

Table 4-Key Research Needs

●

●

●

●

●

●

●

Better understanding of environmental parameters governing

the rate and extent of biodegradation in different environments

Improving methods for enhancing the growth and activity of

petroleum degrading bacteria

Development of better analytical techniques for measuring and

monitoring effectiveness

Field validation of laboratory work

Investigation of what can be done to degrade the more

recalcitrant components of petroleum, e.g., asphaltenes

Improving knowledge of the microbiology of communities of

microorganisms involved in biodegradation

Better understanding of the genetics of regulation of biodegradation

SOURCE: Office of Technology Assessment, 1991.

which will likely be devoted to bioremediation

research.

NETAC and Commercialization of

Innovative Technologies

EPA has entrusted some of the work of developing new bioremediation protocols to the National

Environmental Technology Applications Corp. NETAC

is a nonprofit corporation established in 1988

through a cooperative agreement between EPA and

the University of Pittsburgh Trust. It was created to

help commercialize innovative environmental technologies such as bioremediation.

The Exxon Valdez spill provided the opportunity

for NETAC to become involved in evaluating

bioremediation technologies. After the spill, EPA

and the Coast Guard received a number of proposals

from companies that wanted their bioremediation

products to be tried in Alaska; however, no mechanism existed to enable them to compare competing

technologies. 112 One of NETAC’S first charges,

therefore, was to recommend criteria by which

bioremediation products for cleaning up the Alaska

spill could be judged, that is, to develop a protocol

for assessing the effectiveness of beach bioremediation products. NETAC convened a panel of experts

for this task; from this panel’s recommendations,

EPA established an official procedure to judge

products for possible use in Alaska.

To encourage the submission of products that

might qualify for field testing in Alaska in 1990,

EPA published an announcement in the Commerce

11 IU.S.Environmen~~tWtion Agency, Office of Environmental Engineering andlkchnologyrkrnonstratio~ “AlternativeTreatment Whnolosy

Information Center” (brochure), June 1990.

1lZJ.M. Cogeq “Bioremedi~on Technology for Spilla,” Waste Bm.ness Western, September 1990, p. 17.
Bioremediation for Marine Oil Spills ● 27

Business Daily.113 Thirty-nine proposals were submitted. The NETAC panel evaluated these and

recommended that 11 undergo laboratory testing

specified by the protocol. Effectiveness and toxicity

tests were conducted in EPA’s Cincinnati, Ohio Risk

Reduction Engineering Laboratory. EPA, again with

NETAC’s help, selected two products judged most

appropriate for field testing in Alaska.

The process of identifying promising products

appears to OTA to be both appropriate and fair and,

in general, NETAC is performing a valuable service.

Nonetheless, a few of the bioremediation firms that

had submitted products contended that the tests

specified in the protocol were not appropriate to

assess the true effectiveness of these products. As

more bioremediation research results become available, it will be possible to refine this frost protocol,

if necessary. A NETAC expert panel is currently

identifying the kinds of studies and types of tests

needed for an open water bioremediation protocol,

and EPA’s ORD labs will again use the resulting

framework protocol to develop a complete experimental design. Although much skepticism remains

about the effectiveness of bioremediation at sea, an

open water protocol would be useful whether or not

any effective products were identified.

Relative to the new Interim Guidelines published

by the Oil Spill Response Subcommittee, NETAC is

also preparing to assist Regional Response Teams in

planning for the possible use of bioremediation

technologies. Specifically, NETAC has begun to:

1) systematically compile information on commercial bioremediation products, 2) collect samples for

preliminary laboratory evaluation of bioremediation

products, 3) define a national bioremediation product evaluation facility to test commercial products,

and 4) develop the capability to provide technical

assistance to the States and to regional or national

response teams.114

The Private Sector

The bioremediation industry is a young industry

seeking to develop markets for its products and

expertise. The industry is composed primarily of

companies with fewer than 100 employees, and not

many of these companies have been in existence for

more than 5 years. Of the companies that have

developed bioremediation products, few specialize

in products for marine oil spills. If a market for such

products were to develop, however, many companies would be interested. More than 50 companies

Claimin g bioremediation expertise have expressed

interest in supplying products or personnel in

response to the Persian Gulf oil spill.115 Only a

handful of these products have undergone testing to

evaluate their effectiveness on marine spills (see

above), and none has yet developed a reputation

among experts as an effective response to such

spills.

The Applied Biotreatment Association (ABTA)

was established in 1989 to promote the interests of

the bioremediation industry. The organization has

55 members and consists of about equal numbers of

corporate and adjunct associates. Corporate members are biotreatrnent companies, and adjunct members include State biotechnology centers, equipment

companies, and university professors. In addition to

bioremediation companies, ABTA now includes

among its members two large oil companies. ABTA

recently produced a briefing paper on the role of

bioremediation in oil spills.ll6 Several of its members actively participate in EPA’s bioremediation

Action Committee.

For the most part, the oil industry has taken a

wait-and-see attitude toward bioremediation. Few

companies are doing research on the bioremediation

of marine oil spills, preferring instead to let EPA

take the lead. Recently, the Petroleum Environmental Research Forum (PERF), an oil industry

group that sponsors research on environmental

problems of concern to its members, proposed a

mass balance study to evaluate the potential of

nutrient enrichment and seeding on open water.l 17

This study is expected to begin in mid-1991.

Although both the American Petroleum Institute and

the new Marine Spill Response Corp. believe that

1lSU.S. Environmen~ Protection Agency, “EPA Seeks Biological Methods for Potential Application to Cleanup of Alaskan Oil Spill,’ Commerce

Business Daily, Feb. 12, 1990.

I loNatio~ Environmen~ ‘lkcImology Applications COW., ‘‘NETAC’S Role in Supporting bioremediation Product Selection for Oil Spills, ” Feb.

11, 1991,

115K. Devine, Appfi~ Biomatment Association telephone Communication Jan. 28, 1991.

llGApplied BiOtreament &+SociatiO~ “The Role of bioremediation of Oil Spi~S, ” fall 1990.

IITJ. Salanirrq Shell Development Co., personal communication Jan. 29, 1991.
28 ● Bioremediation for Marine Oil Spills

bioremediation warrants further research, neither is

planning a significant research program of its

own.

ll8 The Exxon Corp. so far is an important

exception to the general absence of oil industry

activity. Exxon, by virtue of its Alaska and New

Jersey experiences, has more familiarity with fertilization techniques than other oil companies and, at

least for certain types of environment, has concluded

that their use is merited.

REGULATORY ISSUES

Several provisions of the Federal Water Pollution

Control Act (the Clean Water Act (CWA)), as

recently amended by the Oil Pollution Act of 1990

(OPA), affect or potentially affect the use of

bioremediation products for marine oil spills. The

CWA specified development of a National Contingency Plan (NCP) for the removal of oil and

hazardous substances. The OPA calls for the revision of parts of this plan. It requires EPA to prepare

a list of dispersants, other chemicals, and other spill

mitigating devices and substances that may be used

to carry out the plan, and to identify the waters in

which they may be used and the quantities that can

be used safely.119 Thus, before a bioremediation

product could be considered for use in response to an

oil spill, it would, at minimum, have to be on this list.

Inclusion on the list implies that certain minimum

safety standards have been met but does not necessarily imply that the product is either effective or

nontoxic for specific applications. In general, additional testing would be required to evaluate further

both efficacy and toxicity.

Subpart J of the National Contingency Plan

governs the use of biological additives (as well as

dispersants and other chemical agents) for marine oil

spills. It identifies several options that can be used to

obtain authorization for the application of a chemical or biological agent to combat a spill. Section

300.910e provides for preauthorization of the use of

regulated agents through an advance planning process. Thus, on-scene coordinators (OSCs) are authorized to use biological additives that have been

preapproved by Regional Response Teams. EPA

encourages preplanning and believes that the deliberations of Regional Response Teams provide the

best forum for considering authorizations.120

If a preauthorized plan has not been established or

is not applicable to the specific circumstances of a

spill, the OSC must obtain the concurrence of the

EPA representative to the Regional Response Team,

the affected State(s), and impractical, the Department

of the Interior and Department of Commerce natural

resource trustees, before using biological or other

agents. An exception occurs when the OSC determines that quick action is necessary to prevent or

substantially reduce a hazard to human life; in that

case, the OSC may unilaterally authorize the use of

any product, including those not on the NCP product

schedule. Continued use of the product once the

threat has been mitigated is subject to normal

concurrence procedures.121

Another statute that may have a bearing on

bioremediation technologies is the Toxic Substances

Control Act (TSCA). The TSCA is intended to

regulate the manufacture of substances that may

pose a risk to human health or the environment. The

Act applies to chemical substances generally, which

EPA has interpreted to include microorganisms.

Currently, however, manufacturers of microbial

products are not required to satisfy Section 5 of

TSCA and notify EPA regarding the use of naturally

occurring microorganisms, and EPA has no plans to

require notification. 122 Genetically engineered organisms, however, would be subject to notification and

review. EPA has not had the opportunity to conduct

product reviews of genetically engineered organisms, but a few companies are considering their

use. EPA expects to publish a draft biotechnology

rule in the Federal Register by late 1991.

lls~e-e spilI Rmpome Corp. (MSRC), formerly the Petroleum Industry Response Gganizatioq was established after thel?xxon VaZdez oil spill

and is intended to give the private sector an improved capability to respond to major oil spills around the country. MSRC has a sizable research budget.

llgofl pollution Act of 1990, s~tion 4201(b). This schedule maybe obtained from EPA’s Emergency Response Division. The list currently COnti

30 bioremediation products and includes both chemical fertilizers and microbial products.

lmJ. (--gham, U.S. Environmental Protection Agency, presentation to the Treatability Protocol Subcommittee of the bioremediation ActiolI

committee, Oct. 15, 1990.

lzl~id.

IZZE. wews~, U.S.EnvkowenM protection Agency, Office for Pesticides and Toxic Substances, pmSOMI communication ~. *6, 1991.
Appendix A

Glossary

Aerobic bacteria: Any bacteria requiring free oxygen for

growth and cell division.

Aliphatic compound: Any organic compound of hydrogen and carbon characterized by a linear or branched

chain of carbon atoms. Three subgroups of such

compounds are alkanes, alkenes, and alkynes.

Anaerobic bacteria: Any bacteria that can grow and

divide in the partial or complete absence of oxygen.

Aromatic: Organic cyclic compounds that contain one or

more benzene rings. These can be monocyclic, bicyclic, or polycyclic hydrocarbons and their substituted

derivatives. In aromatic ring structures, every ring

carbon atom possesses one double bond.

Assay: Qualitative or (more usually) quantitative determination of the components of a material or system.

Biodegradation: The natural process whereby bacteria or

other microorganisms chemically alter and break down

organic molecules.

bioremediation: A treatment technology that uses biological activity to reduce the concentration or toxicity

of contaminants: materials are added to contaminated

environments to accelerate natural biodegradation.

Catalyst: A substance that alters the rate of a chemical

reaction and may be recovered essentially unaltered in

form or amount at the end of the reaction.

Cometabolism: The process by which compounds in

petroleum may be enzymatically attacked by microorganisms without furnishing carbon for cell growth

and division.

Culture: The growth of cells or microorganisms in a

controlled artificial environment.

Dispersant: Solvents and agents for reducing surface

tension used to remove oil slicks from the water

surface.

Emulsification: The process of intimately dispersing one

liquid in a second immiscible liquid (e.g., mayonnaise

is an example of an emulsified product).

Enzyme: Any of a group of catalytic proteins that are

produced by cells and that mediate or promote the

chemical processes of life without themselves being

altered or destroyed.

Fraction: One of the portions of a chemical mixture

separated by chemical or physical means from the

remainder.

Gas chromatography: A separation technique involving

passage of a gaseous moving phase through a column

containing a fixed liquid phase; it is used principally as

a quantitative analytical technique for compounds that

are volatile or can be converted to volatile forms.

Gravimetric analysis: A technique of quantitative analytical chemistry in which a desired constituent is

efficiently recovered and weighed.

Hydrocarbon: One of a very large and diverse group of

chemical compounds composed only of carbon and

hydrogen; the largest source of hydrocarbons is

petroleum crude oil.

Inoculum: A small amount of material (either liquid or

solid) containing bacteria removed from a culture in

order to start a new culture.

Inorganic: Pertaining to, or composed of, chemical

compounds that are not organic, that is, contain no

carbon-hydrogen bonds. Examples include chemicals

with no carbon and those with carbon in non-hydrogenlinked forms.

Metabolism: The physical and chemical processes by

which foodstuffs are synthesized into complex elements, complex substances are transformed into simple ones, and energy is made available for use by an

organism; thus all biochemical reactions of a cell or

tissue, both synthetic and degradative, are included.

Metabolize: A product of metabolism.

Microorganism: Microscopic organisms including bacteria, yeasts, filamentous fungi, algae, and protozoa.

Mineralization: The biological process of complete

breakdown of organic compounds, whereby organic

materials are converted to inorganic products (e.g., the

conversion of hydrocarbons to carbon dioxide and

water).

Oleophilic: Oil seeking (e.g., nutrients that stick to or

dissolve in oil).

Organic: Chemical compounds based on carbon that also

contain hydrogen, with or without oxygen, nitrogen,

and other elements.

Pathogen: An organism that causes disease (e.g., some

bacteria or viruses).

Saturated hydrocarbon: A saturated carbon-hydrogen

compound with all carbon bonds filled; that is, there

are no double or triple bonds, as in olefins or

acetylenes.

Soluble: Capable of being dissolved.

Surface-active agent: A compound that reduces the

surface tension of liquids, or reduces interracial

tension between two liquids or a liquid and a solid; also

known as surfactant, wetting agent, or detergent.

Volatile: Readily dissipating by evaporation.

–29–
Appendix B

Acknowledgments

We are grateful to the many individuals and organizations in the United States and abroad who shared their special

knowledge, expertise, and information with us in the preparation of this study. We are especially indebted to those who

participated in our bioremediation workshop and to those who critiqued our first draft.

Partcipants, OTA Workshop on bioremediation for Marine Oil Spills

December 4,1990

James Blackburn

Energy, Environment and Resources Center

University of Tennessee

Al Bourquin

ECOVA Corp.

Peter Chapman

Environmental Research Laboratory

U.S. Environmental Protection Agency

Rita Colwell

Maryland Biotechnology Institute

University of Maryland

Joseph J. Cooney

Environmental Sciences program

University of Massachusetts

John Glaser

Risk Reduction Engineering Laboratory

U.S. Environmental Protection Agency

Jack R. Gould

American Petroleum Institute

Frank M. Gregorio

Enviroflow

Steven Hinton

Exxon Corporate Research

Stephen Lingle

Office of Environmental Engineering and

Technology Demonstration

U.S. Environmental Protection Agency

Margaret Mellon

National Wildlife Federation

James Mielke

Science Policy Research Division

Congressional Research Service

Clayton Page

Environmental Remediation, Inc.

Robert G. RokuI

BP America

John Teal

Woods Hole Oceanographic Institution

Edward Tennyson

Minerals Management Service

B.J. Wynne

Texas Water Commission

Franz K. Hiebert

Alpha Environmental, Inc.

Additional Reviewers and Other Contributors

Ronald M. Atlas

University of Louisville

Kay Austin

U.S. Environmental Protection Agency

Thomas L. Baugh

Office of Research and Development

U.S. Environmental Protection Agency

Edgar Berkey

National Environmental Technology Applications Corp.

Rosina Bierbaum

Office of Technology Assessment

Neil A. Challis

International Tanker Owners Pollution Federation

Jessica M. Cogen

National Environmental Technology Applications Corp.

Katherine Devine

Applied Biotreatment Association

F. Rainer Engelhardt

Marine Spill Response Corp.

Mervin Fingas

Technology Development and Tewchnical Services Branch

Environment Canada

-30 -
Appendix Acknowledgments .31

Mary Gade

Office of Solid Waste and Emergency Response

U.S. Environmental Protection Agency

Robert Hiltabrand

Research and Development Center

U.S. Coast Guard

Rebecca Hoff

Hazardous Materials Response Branch

National Oceanic and Atmospheric Administration

Kurt Jakobson

U.S. Environmental Protection Agency

Peter Johnson

Office of Technology Assessment

David Kennedy

Hazardous Materials Response Branch

National Oceanic and Atmospheric Administration

Mahlon C. Kennicutt

Texas A&M University

Howard Levenson

Office of Technology Assessment

Morris A. Levin

Maryland Biotechnology Institute

Alfred W. Lindsey

Office of Research and Development

U.S. Environmental Protection Agency

Henry Longest

Office of Solid Waste and Emergency Response

U.S. Environmental Protection Agency

Patrick C. Madden

Exxon Research and Engineering

Alan Mearns

Hazardous Materials Response Branch

National Oceanic and Atmospheric Administration

Tom Mersky

National Environmental Technology Applications Corp.

James G. Mueller

Southern Bio Products, Inc.

Kevin O’Connor

Office of Technology Assessment

Daniel Owen

International Tanker Owners Pollution Federation

James Payne

Science Applications International Corp.

Roger Prince

Exxon Research and Engineering

Hap Pritchard

Environmental Research Laboratory

U.S. Environmental Protection Agency

Patrick Rocques

Texas Water Commission

Pamela Russell-Harris

Office of Solid Waste and Emergency Response

U.S. Environmental Protection Agency

Joseph Salanitro

Shell Development Company

Gary Shigenaka

Hazardous Materials Response Branch

National Oceanic and Atmospheric Administration

Albert Venosa

Risk Reduction Engineering Laboratory

U.S. Environmental Protection Agency

Oskar Zaborsky

National Academy of Sciences

Raymond A. Zilinskas

Maryland Biotechnology Institute

The University of Maryland