Bioremediation
Some Background
• Non-discriminate disposal of waste materials
o Point source pollution issues
o Non-point source
• Leaching of materials into groundwater (recall that a very large percentage of communities obtain
fresh water from groundwater)
• Responsibility---Where does it lie and what happens to a contaminated site if a company dissolves?
• Cost of clean-up. How clean is clean and when does risk balance cost?
These issues led to a sequence of legal actions that created the branch of law known as “Environmental
Law”. The body of legislation deals with conservation, pollution control, and planning and coordination of
testing and clean-up.
Review NEPA and its implications (National Environmental Policy Act, early 1970)
o Requires each agency (government or industry) to produce environmental impact statements. These
statements are really disclosure statements that make everyone aware of the environmental
consequences of certain activities….there is no regulation…just awareness
o Review the OECD…Organization for Economic Cooperation and Development. An organization
that promotes cross border compliance with environmental requirements. (link to TOSCA)
o Review other landmark legislation (Table 16.1 from Maier et al. 1999)
So what happens to the materials that are produced or are already released into the environment and how
does microbiology come into play?
Biodegradation
o Breakdown of organic ‘contaminants’ due to microbial activity
o Basically this is mineralization
o Review the concept of microbial infallibility
o Review the concept of co-metabolism (not for energy metabolism)
The idea of Tiered Testing
o Substances are considered by structure
o Materials having simple structures likely to be easily degraded are ‘tested’ under optimized
conditions using rapid screening methods (CO2 evolution, O2 consumption, DOC reduction)
Some representative structures (from Maier et al. 1999
Point out the dangers of
optimized conditions
Point out the dangers
associated with wrong
assumptions when using
isotopes to do this kind of work
(Fig. 14.1 from Atlas and
Bartha, 1998)
o Testing leads to additional review
under more appropriate conditions
(closer to the conditions that the
material will experience). For
example: will a detergent released into
the environment, be degraded fast enough that it will not cause foaming in rivers?
o Finally the material is studied under conditions that closely duplicate field conditions.
o If a material is largely recalcitrant or a likely candidate for bioremediation artificial
ecosystems may be developed to study its decay (mesocosms).
What regulates biodegradation?
o Genetic potential
o Is there a natural analogue?
o Gene transfer or mutational event may promote degradation
o Bioavailability
o Low water solubility limits potential for uptake
o Adsorption to soil limits availability
o Phase separation limits availability
o So….only 3 options for cells
Direct utilization if target is in water phase
Direct contact of cells with substrate
Direct contact with fine droplets in the water phase
(Review the role of surfactants in promoting access to substrates)
o Contaminant structure
o Branching limits degradation
o Frequency of substitution with halogens
o Toxicity
o Some materials limit membrane integrity
o Environmental factors
o Oxygen
Generally speeds rates of decomposition. For example hydrocarbons are generally
susceptible to attack under aerobic conditions but poor so under anaerobic conditions
o Organic matter
Low concentrations of organic matter naturally means small populations of bacteria
so low natural rates of decomposition and low rates of recruitment (less probability
of an organism being on site that could degrade a substance, grow, and dominate the
community)
o Nitrogen
Some substrates are poor substrates…for example hydrocarbons are all carbon.
Review C:N:P ratios again
o Temperature
Generally the warmer the conditions the better the degradation rate
o pH, salinity, water availability
All play a role to some extent
Pollutants and Xenobiotics
Biomagnification and Persistence
o Principle of microbial infallibility: All NATURAL organic compounds are subject to microbial
attack and hence decay
o Co evolution of cells and substrates
o Lack of any large geologic deposits of organics
o Failure of the Principle of microbial infallibility
o Chemical synthesis produces combinations of elements that do not normally occur in nature
Organic molecules with chlorine substations
Organic molecules with nitrogen substitutions
Unusual bond sequences
Highly substituted aromatic rings
Excessive molecular sizes
All lead to molecules that persist in the environment for long periods of time
o “Dilution is the solution to pollution”
o Review ‘biodegradation’ and
‘mineralization’
Political/industrial view of
biodegradation vs. the
biological view (Figure 13.1
from Atlas and Bartha 1998,
all ‘degradation’ products are
biologically active)
Recalcitrant Molecules
Recalcitrant molecules are molecules that are
poorly metabolized by microbes and
consequently are long-lived in a wide variety
of environments (Table 14.7 from Madigan et
al. 1997). Some additional examples:
Halocarbons
• Synthetics containing carbon-halogen
bonds (mostly carbon-chlorine)
• Examples include refrigerants,
chlorobenzenes, chlorophenols
• Haloalkyl propellants
o C1-C2 alkanes having
essentially all H atoms replaced
by chlorine, fluorine, or
combinations
o Most common example is Freon
(F-11) CCL3F or (F-12) CCl2F2
o Others include cleaning solvents
as carbon tetrachloride, or
trichloroethylene
o Degradation proceeds most
efficiently under anaerobic conditions and by microbial consortia co-metabolizing the
material
o Degradability (aerobic) decreases as the number of halo-substitutes increase
o As halo-substitutes are removed anaerobically the material becomes less likely to be
degraded
Nitroaromatic compounds
• Include military explosives, solvents and pesticides
• Very poorly degraded leading to partial degradation products that tend to be recalcitrant
Polychlorinated biphenyls (PCBs)
• Biphenyls having various degrees of chlorine substitutions (Fig from Atlas and Bartha, 1998)
• Oily having great electrical resistance, high boiling point, and great chemical resistance
• Used as plasticizers, insulators, heat exchange fluids
• Extremely resistant to microbial attack
• Most of the population has some PCB bioburden and PCBs are biologically active causing liver
damage, thinning eggshells in predatory birds and a general disruption of reproductive processes.
(Figures 13.8 and 13.11 from Atlas and Barth, 1998)
Synthetic polymers
• Include polyethylene, polyvinyl chloride, and polystyrene (Fig. 13.12: Atlas and Bartha, 1998)
• 50 million metric tons of plastic produce each
year
• Not so much a biological problem directly as
material is biologically inert---indirect effects
• Large mol. wt. material is not degradable;
plasticizer leaks out leaving a recalcitrant polymer
• Review poly B-hydroxyalkanoates and
degradable plastics (Fig. 14.53 from Madigan et
al. 1997)
• Synthesized by Alcaligenes, Bacillus, and
Psuedomonas
Petroleum
• 6.1 million metric tons of petroleum
hydrocarbons enter the ocean
annually
• Degradation of this material is very
slow—a combination of physical
forces and microbial action
• Petroleum hydrocarbons are a wide
range of material (Fig. 8.9 from
Copone and Bauer, 1992)
• Particularly nasty stuff
o Gooey
o Disrupt chemoreceptors of
some marine life
o Carcinogenic components
may be biomagnified
o Even bacterial metabolism is
affected
• Degradation can be promoted by selective enrichments
o Spills effectively select for hydrocarbon degrading populations
o Natural populations of degraders grow rapidly
o N and P enrichment allow better use of C contained in the hydrocarbon (Bioremediation
figure from Atlas and Bartha, 1998)
o Degradation of hydrocarbons is a combination of physical and microbial processes (Review
Fig 8.2 from Capone and Bauer. 1992)
Bioremediation
• Object here is to use naturally occurring (or introduced) bacteria to clean-up an environment
contaminated with a pollutant
• Process can be in situ (in the habitat) or ex situ (removed from the habitat)
• Process can be
o Intrinsic: That which occur without additional support
o Promoted: That which occurs with additional support
o Oxygen addition
o Nutrient addition
o TEA injection
o Surfactant injection
• The failure of GEMs….and why.
Some examples from Maier et al. 1999 (fig. 16.28) and from Atlas and Bartha 1998 (fig. 14.6)
References:
Atlas, R.M. and R. Bartha. 1998. Microbial Ecology: Fundamentals and Applications 4th ed.
Benjamin/Cummings, Redwood City, CA.
Capone, D.G., and J.E. Bauer. 1992. Microbial processes is coastal pollution. Pgs.191-238 in [R. Mitchel,
ed.], Environmental Microbiology. Wiley, New York.
Maier, R.M., I.L. Pepper, and C.P. Gerba. 1999. Environmental Microbiology. Academic Press, San
Diego.
Madigan, M.T., J.M. Martinko, and J. Parker. 1997. Brock: Biology of Microorganisms 8th ed. Prentice-
Hall, Upper Saddle River, NJ.
