1. Chapter 19 - Transport Objectives Be able to explain the possible interactions of a bacterium with soil pores Be able to explain the relationship between bacterial size and soil pore size and effective transport Be able to list and understand the four factors that affect microbial transport For a given a set of conditions (bacterial shape and size, ionic strength, soil texture) be able to provide an educated prediction of whether microbial transport will occur

2. Transport of microorganisms in soil Distribution of microorganisms in nature preference is shown for attachment mountain streams sediments subsurface environments microorganisms tend to be found in patches or small colonies rather than evenly distributed on soil surfaces soil often filters out microorganisms as they move with water flow

3. Importance of understanding transport of microorganisms To determine the fate of added micoorganisms (either selected or GEM) life vs. death proliferation vs. maintenance adhesion vs. transport To determine the facilitated transport of pollutants

4. Pore spaces in microaggregates with neck diameters less than 6 um have more activity than pore spaces with larger diameters. Bacteria within the former are protected from protozoal predation.

5. 200 um microaggregate root hypha aggregrates or soil particles 2000 um macroaggregate solid pore Assume that 50% of the aggregate is pore space and the pores are 15 um in diameter, there will be 1,000,000 pores Assume the pores are 30um in diameter, there will be 150,000 pores

6. Pore Radius Sand % Hayhook % Vinton Mixture % < 1 um 1 – 10 um 10 - 60 um > 60 um 0.025 0.35 17.13 82.5 6.51 16.05 30.93 46.51 11.70 17.68 49.9 20.83 Hayhook: 10% clay, 5% silt, 85% sand Vinton Mix: 5% clay, 10% silt, 85% sand Pore size distribution for three porous media Bacteria can be no more than 5% of the average pore diameter to get effective transport. 2 um 40 um 0.05 – 0.5 um 0.5 – 3 um

7. Convective flux velocity Q = K DH A t where: z Q = volume of water moving through the column (l 3 ) K = hydraulic conductivity (l/t) DH = hydraulic head difference between inlet and outlet (l) A = cross sectional area of column (l 2 ) t = time (t) z = length of column (l) large flow limited flow Factors affecting microbial transport in soil Advection - movement with bulk fluid

8. Dispersion mechanical mixing – path tortuosity creates velocity differences depending on pore sizes molecular diffusion – random movement of very small particles in a fluid generally due to a concentration gradient. Usually not important for bacteria but might affect virus transport Factors that cause mechanical mixing

9. Adsorption loss of cells from the solution phase due to interaction with surfaces (ranges from reversible to irreversible) There are several ways a cell can approach a surface. Active movement (chemotaxis) is in response to a chemical gradient Diffusion – brownian motion allows random interactions with a surface Convective transport - due to water movement, usually several orders of magnitude > than diffusion

10. Once at the surface several different forces govern the interaction Electrostatic interactions – repulsive forces Hydrophobic interactions – attractive forces Van der Waals forces – attractive forces Electrostatic interactions Coulombs Law: F = k q 1. q 2 where: e. r 2 F = force between the particles q 1, q 2 = charged particles k = constant e = dielectric constant (depends on ionic strength and type)

11. Is F expected to be positive or negative between a bacterial cell and the soil? ? Electrostatic forces are repulsive What is the effect of increasing the ionic strength of the medium? Electrostatic repulsion is reduced.

12. C 0 = 5 x 10 7 cells/ml CEC = 0.03% Transport of Pseudomonas aeruginosa 9027 through sand deionized water 2 mM NaCl Bai et al., 1997. Appl. Environ. Microbiol. 63:1266-1273 Model prediction Experimental data

13. Modeling was performed using a one-dimensional advection- dispersion model that includes combined instantaneous and rate- limited sorption and two first-order irreversible retention terms. where: C = bacterial concentration (M V -1 ) S = sorbed phase bacterial concentration (M V -1 ) R = retardation factor T = time = fraction of instantaneous retardation P = Peclet number, = dimensionless first order cell sticking rate constants retardationfirst-order retention terms advectiondispersion

14. Hydrophobic interactions Nonpolar molecules attract each other Electrostatic repulsion is reduced and hydrophobic interactions can increase What is the effect of increasing ionic strength? van der Waals Forces Occurs between neutral molecules. Electron motion is such as to produce net electrostatic attraction at every instant. van der Waals forces are attractive

15. Polystyrene Glass

16. = electrostatic interaction = separation distance = van der Walls interaction When a cell is right next to the surface the attraction is very strong due to attractive forces creating a primary minimum (H-bonding and dipole interactions). As the two surfaces separate slightly (several nm) repulsive forces grow quickly. At slightly longer distances another, smaller minimum exists. At the secondary minimum the cell is not in actual contact with the surface and so the cells can be removed by increasing water velocity or by changing the chemistry of the system. DLVO theory – Gibbs free energy between a sphere and a flat surface

17. Reversible vs. irreversible attachment

18. Factors affecting microbial transport in soil Advection - movement with bulk fluid Dispersion mechanical mixing – path tortuosity creates velocity differences depending on pore sizes molecular diffusion – random movement of very small particles in a fluid generally due to a concentration gradient. Usually not important for bacteria but might affect virus transport Adsorption – loss of cells from the solution phase due to interaction with surfaces (ranges from reversible to irreversible) Decay – loss of cells from the solution phase due to death (irreversible)

19. Advection only Advection, dispersion Advection, dispersion, adsorption Advection, dispersion, adsorption, decay A short pulse of cells have been added to a column and this is a snapshot of the distribution of cells along the length of the column at some time later.

20. OrganismIonic strengthMineral grain size% Recovery W6 W8 W8 low high low high fine coarse fine coarse fine coarse fine coarse W6 a coccus with radius 0.75 um W8 a bacillus with dimensions 0.75 x 1.8 um Summary and Homework (predict relative recoveries) 14.5% 80.4% 2.8% 49.3% 3.9% 43.6 0.3% 4.3%

21. A series of experiments were performed in glass bead columns to determine the impact of injected cells on the permeability of the column. 500 PV of Klebsiella pneumoniae (10 8 CFU/mL) were injected. The cells were either vegetative, starved for 2 weeks, or starved for 4 weeks. MacLeod et al., 1988. Appl. Environ. Microbiol. 54:1365- 1372.

22. Differences in the DNA-derived cell distribution in glass bead cores injected with vegetative or starved cells. MacLeod et al., 1988. Appl. Environ. Microbiol. 54:1365- 1372.