Bacterial Transport in Saturated, Unsaturated, and Air-Sparged Porous Media

A research project conducted in cooperation with the University of Arizona NIEHS Superfund Center.

For more information on the NIEHS program, click on: http://www.niehs.nih.gov/sbrp/home.htm.
For information on the University of Arizona Superfund center, click on: http://www.pharmacy.arizona.edu/centers/tox_center/superfund.htm

Need: There are over 217,000 contaminated waste sites in the United States awaiting remedial actions, with 547 designated as Superfund National Priority Lists (NPL) sites. The remedial costs for the cleanup of the contaminants emanating from solvents, petroleum products, and metals is projected to exceed $187 billion. Due to the need for less costly remedial treatments, there is a great interest in contaminant-specific treatment techniques such as bioremediation.

In-situ soil bioremediation is a promising method of aquifer remediation. In many cases pollutant biodegradation can be accelerated through bioaugmentation, or the delivery of pollutant-degrading microbes into the contaminated soils. Researchers have isolated, identified, and cultured in the laboratory microbes capable of degrading toxic pollutants such as benzene, toluene, and carbon tetrachloride. Unfortunately, most of these laboratory-grown bacteria readily stick to soil grains, limiting their use for bioaugmentation. There is little benefit to digging a well and injecting bacteria, for example, if the bacteria travel only a few centimeters from the well casing. From an economic standpoint, it would be cheaper to dig up the soil than to attempt in-situ remediation. Fortunately, there are several methods that can be used to enhance bacterial transport in soils.

Transport Studies: As a Part of the NIEHS Superfund Program(web site), The University of Arizona was designated as a Superfund Research Center. Professor Logan and graduate students Terri Camesano and Amanda DeSantis are working with The University of Arizona to investigate methods to increase the transport of bacteria in porous media. The approach used by the research team to identify factors that reduce attachment was to study bacterial retention in short soil columns. By incorporating a radiolabeled amino acid into the cells, they were able to detect low cell concentrations sorbed to soil. This made it possible to measure changes in retention, in columns only centimeters long, that would enhance the transport cells over distances of tens of meters in the field. The success of adhesion modifying treatments was quantified in terms of a sticking coefficient (the fraction of collisions of cells with soil grains that are successful) and mathematical models. Transport was judged to be successful when sticking coefficients were reduced to below 10-2 to 10-3.

It was discovered that by suspending cells in demineralized water it was possible to transport cells over much longer distances than that possible using groundwater. Increased transport resulted from an increase in the electrostatic repulsive forces between the negatively charged soil particles and bacteria. Although soils will slowly weather and increase the conductivity of the injected water, low conductivities in the soils were maintained over the brief periods necessary to inject the bacterial pulse. Similar enhancements in bacterial transport were obtained using non-ionic surfactants (such as Tween 20), although surfactants had to be added at concentrations that were too high (0.01%) to make this approach practical at actual sites.

Other factors are being investigated that can be used to increase the effectiveness of bioaugmentation-- although the success of these approaches was not anticipated from simulations using current particle transport models. Critical factors include fluid injection rate (fluid velocity), cell concentration, and whether or not the cells are motile. Current models, based on a set of equations known as clean-bed filtration theory, suggest that high fluid velocities can increase cell transport. Furthermore, these models suggest that soil retention would be larger for motile than non-motile cells and that cell concentrations do not alter overall deposition rates. However, much different results have been observed in laboratory column experiments. The transport of several motile strains of bacteria (Pseudomonas stutzeri KC, Pseudomonas fluorescens P17, and Pseudomonas putida KT2442) have been found to be much greater at low fluid velocities (<1 m/d) than at a high fluid velocities (>100 m/d).

The distance bacteria can move in the soil increases for some strains as cells are increasingly deposited on the soil, indicating there is an effect of particle concentration (or total injected cell mass). Deposited cells can either reduce the subsequent attachment rates of other cells, via a phenomenon known as blocking, or increase rates (ripening). The outcome is strain specific. A trichloroethylene (TCE) degrader Burkholderia cepacia G4 exhibited a blocking effect while Pseudomonas fluorescens P17 has been observed to promote ripening. Blocking can substantially reduce deposition rates because deposited bacterium creates a "shadow zone", or blocked surface area, that can be greater than the size of the bacterium itself. The size of this shadow zone is a function of solution chemistry and hydrodynamics (flow velocity).

New Molecular-level Approaches: Ongoing research is aimed at examining the importance of cell motility and factors that affect the extent of blocking for a number of different bacteria in two ways. First, researchers are examining how deposition rates (and blocking) are altered by the addition of surface modifying chemical additives as a function of cell concentrations and fluid velocities. Second, the factors that inhibit adhesion (increase repulsion) and promote blocking are being probed at nano-scales using an atomic force microscope (AFM). This instrument can be used to generate 3-D images of surface topography and maps of surface elasticity. By studying the response of the tip to the treated and untreated cells, it is possible to precisely measure electrostatic forces that are thought to govern cell attachment to surfaces. The researchers are studying the effects of various chemical treatments on the morphologies of Burkholderia cepacia G4 and Pseudomonas stutzeri KC as well as the repulsive response of the AFM silica tip to see if it can be correlated to bacterial sticking coefficients in a sandy soil.

It is hoped that these experiments will lead to a better understanding of the factors that control cell adhesion to soil particles as well as to other surfaces. Ultimately, this research will contribute to the increased success of in situ soil remediation by improving the dispersion of laboratory cultivated bacteria and increasing the overall efficiency of bioaugmentation processes.

Ms. Terri Camesano (tac162@psu.edu), shown working on the Atomic Force Microscope (AFM), is a Ph.D. candidate in Environmental Engineering. She has shown that cell motility can increase the transport of one bacterial strain at low groundwater velocities. Using the AFM, she is examining the topography and surfaces forces of bacteria in order to improve bacterial transport for bioaugmentation.
Ms. Amanda DeSantis (aad112@psu.edu) is examining the generality of the finding that motile bacterial transport is enhanced at low groundwater velocities compared to non-motile cells. She is running soil column experiments to calculate bacterial adehsion to soil grains and to determine the effect of blocking on microbe transport.
Mr. Karl Shellenberger (kxs175@psu.edu) is using the AFM to examine surface forces important in controlling cell attachment. It has been hypothesized by others that localized regions of positive charge create favorable adhesion sites on mineral surfaces. He is using the AFM to scan surfaces to test this hypothesis by measuring surface-tip interactions over large regions of the glass surface.

For more information please contact:

Bruce Logan, Ph.D.
Phone: 814-863-7908, Fax: 814-863-7304, Email: blogan@psu.edu

To learn more about this area of research please refer to the following sources:

Rogers, B. and B.E. Logan. 2000. Bacterial transport in NAPL-contaminated porous media. J. Environ. Engrg. 126(7): In press.

Camesano, T.A. M.J. Natan, and B.E. Logan. 2000. Observation of changes in bacterial cell morphology using tapping mode atomic force microscopy. Langmuir 16(10):4563-4572.

Unice, K.M., and B.E. Logan. 2000. The insignificant role of hydrodynamic dispersion on bacterial transport. J. Environ. Engin. 126(6): 491-500.

Camesano, T.A., K.M. Unice and B.E. Logan. 1999. Modeling dynamic blocking of colloids in porous media using intracolumn deposition patterns and breakthrough curves. Colloids Surf. A. Physicochem. Engin. Aspects. 160(3):291-307.

Fang, Y. and B.E. Logan. 1999. Bacterial transport in gas sparged porous media. J. Environ. Engng. 125(7):668-673.

Logan, B.E., T.A. Camesano, A.A. DeSantis, and J.C. Baygents. 1999. Comment on "A method for calculating bacterial deposition coefficients using the fraction of bacteria recovered from laboratory columns" by Bolster et al. Environ. Sci. Technol. In press.

Jewett, D.G., B.E. Logan, R.G. Arnold, and R.C. Bales. 1999. Transport of Pseudomonas fluorescens strain P17 through porous media as a function of water content. J. Contam. Hydrol. 36(1-2):73-89.

Li, Q. and B.E. Logan. 1999. Enhancing bacterial transport for bioaugmentation of aquifers using low ionic strength solutions and surfactants. Wat. Res., 33(4):1090-1100.

Camesano, T.A. and B.E. Logan. 1998. Influence of fluid velocity and cell concentration on the transport of motile and non-motile bacteria in porous media. Environ. Sci. Technol. 32:1699-1708.

Gross, M.J., Albinger, O., Jewett, D.G., Logan, B.E., Bales, R.C., and R.G. Arnold. 1995. Measurement of Bacterial Collision Efficiencies in Porous Media. Wat. Res. 29:1151-1158.

Gross, M.J. and B.E. Logan 1995. Influence of Different Chemical Treatments on Transport of Alcaligenes paradoxus in Porous Media. Appl. Environ. Microbiol. 61:1750-1756.

Johnson, W.P., Martin, M.J., Gross, M.J., and B.E. Logan. 1996. Facilitation of Bacterial Transport through Porous Media by Changes in Solution and Surface Properties. Colloid Surf. A: Physiocochem. Eng. Aspects 107:263-271.