Bioremediation

Bioremediation DefinedAny process that uses microorganisms, fungi, green plants or their enzymes to break down harmful chemicals and pollutants in order to return the environment to its original natural condition.

Bioremediation is not only about genetics and enzymology but also about physiology and ultimately ecology.de Lorenzo V: Systems biology approaches to bioremediation. Curr Opin Biotechnol 2008, 19:579-589.

Alleviating PollutionEx situ or in situ interventionNatural attenuation Example: phytoremediation (hyperaccumulators) store heavy metals in vacuolesSebertia acuminata 20% dry weight is nickel. Plants on side of freeways are taking up lead from gas exhaust

Bio-stimulationAdd nutrients (nitrate/sulfate) that cause blooms of naturally occurring microbial bioremediators.Example: bacteria that metabolize polycyclic aromatic hydrocarbons or polychlorinated biphenyls

Bio-augmentationGenetically Modified BioremediatorsAlter organisms to manufacture proteins for desired metabolismYellow poplar tree given enzyme mercuric reductase thrives in mercury soil, cadmium, TCEBacteria gene breaks down TNT is linked to jellyfish gene that glows. Bacteria spread on soil glows green near explosivesChakrabarty first patented oil eater bacterium. Combined 4 plasmids in one bacterial cell gave it the ability to degrade four components of crude oil.

Why do we even need it?We cant seem to stop pollutingInorganicsUranium, technicium, sulfur, slfuric acidExplosivesRDX, TNTPolyaromatic hydrocarbonscreosoteChlorinated hydrocarbonsTrichlorethylene, PCBs, pentachlorophenolPetroleum hydrocarbonsGas, gas additives (MTBE), deisel

From mid-1980s up to 90s numerous attempts were made to design GMO for environmental release for pollutants and heavy metals (USGS). Failures to program: bacteria doesnt behave in a predictable fashion from the lab.

Case Study 1

Case Study 1 contMost failures at bioremediation are due to failure of introduced organisms to thrive in the natural environment or a failure to access the contaminant. This could be due to: Lack of nutrientsPredation or parasitismCompetition (GEMs tend to compete poorly with indigenous populations).Immobility of introduced bacteriaContaminant concentrations below threshold for organism survivalOrganisms may feed on alternative substrates (E. coli and Pseudomonas diverge genetically from initial inoculum in field trials).

A few examples of failed bioremediation attempts:Inoculation of soil with aliphatic hydrocarbon degrading bacteria did not enhance degradation of fuel oil Venosa AD, Wrenn BA (1996) Selective enumeration of aromatic and aliphatic hydrocarbon degrading bacteria by a mostprobable number procedure. Can. J. Microbiol.42: 252258A Pseudomonas sp. shown in lab cultures to degrade 1,4-dichlorophenol failed to degrade the compound when added to surface soilsSayler GS, Ripp S (2000) Field applications of genetically engineered microorganisms for bioremediation processes. Curr Opin Biotechnol 11:286-289

Case study 2:degradation of crude oil by halophilic Archaea

Defining bioremediation by natural attenuationwhat is the environment? what is the pollutant?

are bacterial, Archaeal, and/or plant species present that degrade the pollutant of interest?

what conditions (nutrient, temperature, pH, salt etc) are necessary for that activity?

Defining bioremediation by natural attenuation:hydrocarbon degradation in the Arabian Gulf hypersaline coastWest, Ian. 2008. Qatar - sabkhas, evaporites and some other desert features: an introduction.http://www.soton.ac.uk/~imw/Qatar-Sabkhas.htmwhat is the environment? what is the pollutant?the hypersaline coast of the Arabian coastcrude oil (hydrocarbons)

Defining bioremediation by natural attenuation:hydrocarbon degradation in the Arabian Gulf hypersaline coastwhat is the environment? what is the pollutant?the hypersaline coast of the Arabian coastcrude oil (hydrocarbons)are bacterial, Archaeal, and/or plant species present that degrade the pollutant of interest? environmental samples grow on minimal mineral plates with crude oil vapor as sole carbon/energy source2 Haloferax strains1 Halobacterium strain1 Halococcus strain(Al-Mailem et al., 2010 Extremophiles)

Defining bioremediation by natural attenuation:hydrocarbon degradation in the Arabian Gulf hypersaline coastare bacterial, Archaeal, and/or plant species present that degrade the pollutant of interest?autoclaved controlHaloferax isolateHalobacterium isolateHalococcus isolateC18 hydrocarbongas-liquid chromatography to measurehydrocarbon degradation(Al-Mailem et al., 2010 Extremophiles)

Defining bioremediation by natural attenuation:hydrocarbon degradation in the Arabian Gulf hypersaline coastwhat conditions (nutrient, temperature, pH, salt etc) are necessary for that activity?increased salt increased hydrocarbon degradationHaloferax isolateHaloferax isolateHaloferax isolateHalococcus isolateHalobacterium isolate(Al-Mailem et al., 2010 Extremophiles)

How could this hydrocarbon degradation activity by Haloarchaea be improved?

General strategies for improving microbial bioremediation: stimulation or augmentation

Some existing bio-engineering toolsUniversity of Minnesota Biocatalysis/Biodegradation Database (UMBBD): enzymes, pathways, reactions, compounds from hundreds of bacterial species of interest (Gao et al., 2010)MetaRouter: tracks possible breakdowns from a starting point using all possible reactions (Pazos et al., 2005)in silico modeling of altered strains: ex. Optstrain (Pharkya et al. 2004), DESHARKY (Rodrigo et al. 2008)

How could this hydrocarbon degradation activity by Haloarchaea be improved?

How could systems level knowledge help design stimulation or augmentation strategies?need to know network topology of the pathway being added or altered as well as the influence of the environment on that pathwayunderstand coupling of pathways to better integrate the engineered or altered pathway into the rest of the host systemunderstand demands created by the new flux on resources needed for growth/survival: energy, carbon, redox balance, transcriptional and translation capacityis it possible to compensate by altering regulation by TFs etc, or by adding or deleting other pathways?understand effect of environment on pathway fluxunderstand role of noise in pathway regulationif cooperation between multiple microbial species is used, then systems analysis can evaluate impact of biodegradative flux on the multispecies consortia