Model Systems for In Situ Bioremediation: Novel Methods for Monitoring Microbial Degradation Activity in Subsurface Environments

Abstract

Bioremediation has emerged as a promising treatment approach for restoring contaminated environments, where microorganisms degrade organic pollutants into non-harmful end products. Polycyclic Aromatic Hydrocarbons (PAHs) are a hazardous class of pollutants which have accumulated in soils as a by-product of fossil fuel use. A growing field of research is investigating ways to enhance bioremediation as strategy for removing PAHs from soil environments, where the principal limiting factors for biological degradation activity are pollutant/microbe interaction and the ability of an organism to catabolise a pollutant. A number of engineered treatment approaches aim to address these limits by improving PAH bioavailability and stimulating microbial degradation activity; such as the bioaugmentation of soils with live bacteria degrader cultures and the application of surface acting agents in liquid and microbubble form. However, progress in this field has been limited, as bioremediation processes have not been well characterised at a cellular or molecular scale. Recent advances in microbiology and genetics have allowed for the creation of fluorescent biological sensor organisms which can be engineered to produce a signal response in association with a specific cellular activity. This research presents proof of concept for application of a novel fibre optic 'Optrode' fluorescence detection system to monitor biological activity in model soil column environments. A new strain of naphthalene degrading bacteria was engineered for use in this research, from which fluorescent biosensor strains were developed using recombinant gene technology. Biosensors were detectable at levels commonly reported at bioremediation treatment sites, and were identified in model bench-scale soil columns using the Optrode system. Recent studies suggest that surfactant microbubble foams may enhance bacterial transport and pollutant availability in bioremediation scenarios, however there has been no investigation in regards to the behaviour of these substances in the soil environments. Bench scale-soil columns were developed in order to investigate and compare the fate of biosurfactant solutions, foams, biosensor bacteria and naphthalene in model subsurface environments using advection-dispersion models. Rhamnolipid biosurfactant solution was found to significantly improve hydraulic conductivity and bacterial advection in packed porous media. Naphthalene was highly immobile in surfactant solution, and model results suggest that a major aspect of naphthalene adsorption loss was due to adhesion with micellar structures in the soil column. Advection-dispersion models were used to describe the distribution of Rhamnolipid microbubble foams in soil columns, which constitutes a crucial first step in characterising the behaviour and distribution of biosurfactant microbubble foams in subsurface environments. Breakthrough models suggest that the liquid and gaseous phases of microbubble foams have different fates in the soil column, and their distribution may be better described using a two phase flow system. Best fits for advection dispersion models suggest that while solute distribution in foam is restricted to the liquid phase, bacteria interact with both the liquid and gaseous phases, so their transport through soil column environments may be enhanced even when liquid drainage occurs. This research demonstrates that a combined approach of bench-scale soil column modelling and in situ monitoring of biological activity is able to characterise real-time biodegradation processes at a fine scale. The development of a comprehensive system for investigating and optimising bioremediation is an important aspect of enhancing the progression of treatment strategies into successful field application.