Biodegredation and Bioremediation Course Overview Slides

Define contaminant

Biological, chemical, physical, or radiological substance (normally absent in the environment) which, in sufficient concentration, can adversely affect living organisms through air, water, soil, and/or foodbiological, chemical, physical, or radiological substance (normally absent in the environment) which, in sufficient concentration, can adversely affect living organisms through air, water, soil, and/or food.

Define remediation

The development and implementation of strategies to clean up (remediate) the environment by removing the hazardous contamination disposed in properties since the beginning of the industrial revolutionThe development and implementation of strategies to clean up (remediate) the environment by removing the hazardous contamination disposed in properties since the beginning of the industrial revolution.

What does Gibbs free energy (ΔG) tell us about a reaction?

ΔG < 0: energetically favorable; ΔG > 0: requires energy input or coupling

How can an unfavorable biochemical reaction proceed?

Couple it to a sufficiently favorable reaction, such as ATP hydrolysis, so the overall ΔG is negative.

How do enzymes accelerate biochemical reactions?

Lower activation energy without changing the overall ΔG of the reaction

Which molecules can readily cross the lipid bilayer?

Small hydrophobic molecules, including O₂, CO₂, and benzene; ions require transport mechanisms.

What are hydrolysis and fermentation in sediment organic matter degradation?

Hydrolysis breaks polymers into smaller compounds; fermentation converts organics into products such as H₂, acetate, and CO₂

Why don’t viable plate counts equal direct microbial counts?

Many cells cannot grow under laboratory conditions; some are dead or slow-growing.

Why doesn’t detecting a 16S rRNA gene prove microbial activity?

DNA indicates presence, not necessarily growth, gene expression, or metabolic activity.

What are the five activity-assessment approaches?

mRNA, MICRO-FISH, BrdU, ribosome fingerprinting, and SIP

Why can mRNA indicate microbial activity?

Short-lived mRNA reflects recent gene expression

What does MICRO-FISH combine?

Radiolabeled substrate uptake with fluorescence-based microbial identification.

What does BrdU incorporation indicate?

New DNA synthesis and microbial growth activity.

How can rRNA/DNA ratios inform microbial activity?

Higher ratios can suggest increased ribosomal content and growth, but are indirect indicators.

How does DNA-SIP identify microorganisms assimilating a substrate?

¹³C-substrate → labeled DNA → density separation → PCR/sequencing.

Why is ¹³C used in SIP?

It is a stable isotope that can label microbial DNA/RNA without radioactive decay.

What is a limitation of SIP?

Cross-feeding can label secondary consumers; incubation can alter communities.

Define mRNA analysis.

Measures messenger RNA transcripts to identify genes being expressed, indicating potential microbial activity.

Define MICRO-FISH analysis.

Combines microautoradiography of radiolabeled substrate uptake with fluorescent probes identifying microbial cells.

Define BrdU incorporation analysis.

Uses a thymidine analogue incorporated into newly synthesized DNA to identify growing microorganisms.

Define ribosome fingerprinting (16S RNA/DNA analysis).

Compares ribosomal RNA and DNA community profiles to identify potentially active microbial groups.

Define stable isotope probing (SIP).

Tracks stable isotope incorporation (e.g., ¹³C) into microbial DNA/RNA to identify organisms assimilating labeled substrates.

What does mRNA analysis tell us?

Which genes are being actively expressed.

What does MICRO-FISH tell us?

Which identified microorganisms are taking up a particular substrate.

What does BrdU incorporation tell us?

Which microorganisms are synthesizing new DNA (indicating growth).

Ribosome fingerprinting

Which microbial groups show RNA/DNA patterns associated with potential activity.

What does Stable isotope probing (SIP) tell us?

Which microorganisms are incorporating a labeled substrate into their DNA or RNA.

Define the Oxford Nanopore MinION.

A portable DNA sequencing device that uses nanopores to sequence DNA and characterize microbial communities.

How does nanopore sequencing work?

DNA passes through a protein nanopore, altering electrical current. The signal is decoded into a DNA sequence.

What are the main practical advantages of the MinION?

Fast, relatively low-cost, portable, usable in the field, and requires limited specialized training.

What is microbial infallibility?

No natural compound is totally resistant to biodegradation provided that environmental conditions are favorable. Microbes have evolved that are capable of degrading most if not all naturally produced chemicals.

Why is it difficult to biodegrade xenobiotic (anthropogenic) chemicals?

Foreign chemical structures (e.g. chlorine in DDT) and production in large quantities; load greater than degradation rate.

Define xenobiotics.

Anthropogenic chemicals foreign to the biosphere.

What can organic compounds often be classified as?

• (Bio)degradable

• Persistent

• Recalcitrant

What is persistence a function of?

The compound, environment and time.

A compound is recalcitrant if…

It accumulates in the environment.

An environmental hazard is a function of?

Discharge and exposure.

Define biodegradation.

Biologically mediated simplification of a chemical’s structure, ultimately resulting in mineralization. The term is used, however, in different circumstances to indicate different extent of degradation, ranging from loss of compound to complete mineralization.

Define mineralization.

Conversion of an organic compound into its inorganic constituents. Aerobic is complete oxidation to CO₂ and anaerobic products are CH₄ and/or CO₂, H₂.

Define biotransformation.

Biologically mediated alteration of molecular structure of compound, can either increase or decrease the complexity of the compound, partial metabolism (oxidation, reduction, polymerization…).

Define co-metabolism.

Transformation of a chemical without it providing carbon and/or energy for the organism (another compound needed as a carbon/energy source).

How do chemical properties influence biodegradation?

Structure, molecular size, solubility, halogen substitution, and toxicity affect bioavailability and microbial degradation.

How do environmental conditions influence biodegradation?

Oxygen availability, temperature, pH, nutrients, dissolved carbon, and surfaces affect microbial activity.

How do biological factors influence biodegradation?

Availability of degradative enzymes, appropriate microorganisms, and microbial competition affect degradation potential.

Why doesn't contaminant disappearance necessarily indicate biodegradation?

Physical transport, sorption, volatilization, or abiotic transformation may also reduce measured concentrations.

Define bioremediation.

Managed or spontaneous processes using biological activity to degrade contaminants.

What properties can make persistent pollutants environmentally hazardous?

Toxicity, mutagenicity, carcinogenicity, teratogenicity, and bioaccumulation.

Define Oxidation-Reduction Potential (E_h)

Measure of the tendency of a solution to donate or receive electrons.

What are degradation mechanisms and pathways dependent on?

Chemical properties & structures, environmental conditions (redox, presence/absence of O₂,..), microbial community.

Define catabolism.

Breakdown of compounds to release energy

Define anabolism

Use of energy to synthesize cellular components.

What are central and peripheral metabolic pathways?

Peripheral pathways convert diverse complex compounds into intermediates that enter central metabolism for further breakdown and energy generation.

Distinguish an electron donor from an electron acceptor.

Donor: Loses electrons and is oxidized. Acceptor: Gains electrons and is reduced.

What is the general hierarchy of electron acceptors as conditions become more reducing?

O₂ → NO₃⁻ → Mn(IV) → Fe(III) → SO_4²⁻ → CO₂ (methanogenesis). Processes can overlap.

Describe anaerobic reductive dechlorination of PCE.

PCE accepts electrons as chlorine is removed: PCE → TCE → DCE → VC → ethene. Hydrogen can serve as the electron donor.

What are the eight major methods for measuring biodegradation?

Sum parameters, contaminant disappearance, CO₂ production, ¹⁴C labeling, metabolite formation, electron balance, CSIA, and microbial growth/activity measurements.

Define BOD.

Biological oxygen demand, measure of O₂ consumption required by microbial oxidation of readily degradable organic compounds.

Define COD.

Chemical oxygen demand, measure of oxidizable material.

What is the limitation of sum parameters (BOD, COD, TOC)?

They measure bulk properties but cannot determine the fate of an individual contaminant.

Why doesn’t decreasing contaminant concentration prove biodegradation?

Concentrations can decrease through nonbiological processes; disappearance alone does not establish transformation or mineralization.

What is the purpose of a sterile or killed control?

To distinguish biologically mediated contaminant loss from abiotic losses or transformations.

Define respirometry in the context of biodegradation.

Measuring respiratory activity, such as CO₂ production, to assess microbial mineralization of organic material.

What is the limitation of measuring total CO₂ production?

It cannot identify which specific organic compound was mineralized.

How can ¹⁴C-labeled compounds demonstrate mineralization?

Production of ¹⁴CO₂ from a ¹⁴C-labeled contaminant directly demonstrates mineralization of labeled carbon.

Why measure metabolite formation during biodegradation?

Identifies transformation products and provides evidence of specific degradation pathways.

What is an electron balance, and why is it useful?

Compares electrons released during substrate oxidation with electrons accepted during respiration, helping evaluate whether observed transformations are consistent with biodegradation.

What is compound-specific isotope analysis (CSIA)?

Measurement of stable isotope ratios of individual compounds to investigate transformation processes and isotope fractionation.

Why does the residual contaminant often become enriched in heavier isotopes during biodegradation?

Bonds involving lighter isotopes often react faster, leaving residual contaminant enriched in heavier isotopes.

How can microbial growth be measured?

Changes in cell counts, protein, DNA, or ATP; growth can indicate substrate utilization.

Does microbial transformation of a contaminant always result in microbial growth?

No. A compound can be transformed without being used as a carbon or energy source, such as during cometabolism.

Define the acclimation period in biodegradation.

Initial lag period before observable degradation, during which microbial populations or metabolic capabilities may adapt.

What processes may explain an acclimation period?

Population growth, mutation/gene transfer, enzyme induction, substrate preference, toxicity, or predation.

Define an enrichment culture.

A culture with selective conditions favoring the growth of particular microorganisms or metabolic groups, often reducing community diversity.

What is BOD_5?

The amount of dissolved oxygen consumed by microorganisms during a five-day incubation.

Define groundwater.

Water below the water table in saturated pore spaces/fractures

Define aquifer.

A water-bearing formation capable of transmitting usable groundwater.

Distinguish LNAPLs from DNAPLs.

LNAPLs: Less dense than water; accumulate near the water table.

DNAPLs: Denser than water; sink and may pool above low-permeability layers.

Why are NAPLs considered persistent contaminant sources?

They gradually dissolve into groundwater, continuously supplying contaminants to the dissolved plume.

Define contaminant rebound.

An increase in contaminant concentrations after treatment, often caused by residual NAPL dissolution or contaminant release from low-permeability zones.

Why does hydraulic conductivity affect groundwater remediation?

It controls groundwater flow and amendment delivery. Low-permeability zones limit transport and can retain contaminants.

Distinguish in situ from ex situ remediation.

In situ: Treats contamination in its original location. Ex situ: Removes contaminated material for treatment elsewhere.

Define natural recovery.

Natural recovery: Contaminants decrease through natural physical, chemical, or biological processes.

Define biostimulation.

Addition of electron donors, acceptors, nutrients, or co-substrates to stimulate indigenous microorganisms.

Define bioaugmentation.

Addition of microorganisms with specialized contaminant-degrading capabilities to a contaminated environment.

Distinguish oxidative and reductive biostimulation.

Oxidative: Supply electron acceptors (e.g., O₂) to promote oxidation of organic contaminants.

Reductive: Supply electron donors (e.g., H₂) to promote reduction of contaminants such as PCE.

Define pump-and-treat remediation.

Groundwater is extracted through wells, treated aboveground (e.g., activated carbon), and discharged or reinjected. It can provide hydraulic containment.

What are the major limitations of pump-and-treat?

Long treatment times, high costs, incomplete contaminant removal, low-permeability zones, and rebound from residual contamination.

Define soil vapor extraction (SVE).

Vacuum extraction of volatile contaminants from the unsaturated (vadose) zone, followed by vapor treatment.

Define bioventing.

Supplying low-flow air/O₂ to contaminated vadose-zone soils to stimulate aerobic biodegradation.

Define biosparging.

Injecting air/O₂ below the water table to stimulate aerobic biodegradation in the saturated zone.

How does groundwater recirculation support bioremediation?

Groundwater is extracted, amended with electron donors/acceptors or nutrients, and reinjected to distribute amendments through the treatment zone.

Define a permeable reactive barrier (PRB).

A permeable treatment zone installed across groundwater flow that allows contaminated water to pass through reactive material.

How do organic PRBs differ from zero-valent iron PRBs?

Organic PRBs: Provide substrates supporting microbial degradation.

ZVI PRBs: Promote abiotic chemical reduction, including dechlorination.

Why is lactate added during anaerobic reductive dechlorination?

Fermenting microorganisms convert lactate into products including H₂, which serves as an electron donor for organohalide respiration.

What parameters should be monitored to evaluate groundwater bioremediation?

Parent contaminants, daughter products, electron donors/acceptors, redox conditions, pH, nutrients, microbial activity, and relevant groundwater conditions.

Why can DNAPLs be difficult to remediate in fractured rock?

They can migrate rapidly to great depths through fractures, creating inaccessible sources and unexpected contaminant distributions.

Why use slow-release electron donors in anaerobic bioremediation?

They provide a sustained electron supply over time, reducing the need for repeated injections.

Define biofouling in remediation systems.

Microbial growth and biomass accumulation that can obstruct pores, wells, or treatment media and reduce flow.

How can in situ thermal treatment improve remediation?

Heating increases contaminant volatility, solubility, and diffusion, facilitating removal of NAPLs and concentrated contamination.

Define organohalides.

Organic compounds containing carbon–halogen bonds (C–F, C–Cl, C–Br, or C–I).