Environ. Sci. Technol. 2006, 40, 1547-1555
Virus Retention and Transport in Chemically Heterogeneous Porous Media under Saturated and Unsaturated Flow Conditions J I E H A N , † Y A N J I N , * ,† A N D CLINTON S. WILLSON‡ Department of Plant and Soil Sciences, University of Delaware, Newark, Delaware 19716, and Department of Civil and Environmental Engineering, Louisiana State University, Baton Rouge, Louisiana 70803
Retention and transport of colloids and microorganisms are complex processes, especially in the vadose zone due to the more complicated water flow regime and additional interfacial reactions involved. In this study, we examined the retention and transport behavior of two bacteriophages, MS-2 and φX174, in homogeneous and chemically heterogeneous media under variably saturated conditions. Column experiments with glass beads (treated to have either hydrophilic or hydrophobic surface properties) were conducted using a phosphate-buffered saline solution at different pore water ionic strengths ranging from 0.025 to 0.163 M. In columns packed with 100% hydrophilic glass beads, retention of the viruses increased with decreasing water content and increasing ionic strength, a result similar to those reported in the literature. However, greater retention of both MS-2 and φX174 was observed in saturated columns than in unsaturated columns packed with a 1:1 mixture of hydrophilic and hydrophobic glass beads, especially at high ionic strengths. This result contradicts the common belief that viruses (and colloids in general) are subject to greater removal in unsaturated media. Our study suggests that while the mechanisms controlling colloid interfacial interactions (i.e., attachment on solid-water and air-water interfaces and film straining) on the pore scale are relevant, nonuniform wetting conditions due to heterogeneous grain surface hydrophobicity can strongly influence water flow and phase interconnection. Under these conditions, hydrodynamic effects on the mesopore scale will dominate pore-scale interfacial reactions in controlling the extent of colloid retention and movement in unsaturated media.
Introduction Retention and transport of colloids and microorganisms (sometimes referred to as biocolloids) in porous media is an area of active research. In particular, the presence of pathogenic viruses in wells and groundwater has incurred significant attention from environmental researchers because ∼70% of waterborne diseases in the United States have been associated with groundwater (1, 2). The potential sources of * Corresponding author phone: (302) 831-6962; fax: (302) 8310605; e-mail:
[email protected]. † University of Delaware. ‡ Louisiana State University. 10.1021/es051351m CCC: $33.50 Published on Web 01/20/2006
2006 American Chemical Society
groundwater pollution resulting from virus movement into the subsurface include septic tanks, landfills, land disposal of biosolids, and wastewater discharge and reuse (3, 4). Effective policy making and the establishment of disinfection regulations concerning viruses in drinking water require a thorough understanding of the fate and transport of viruses in the subsurface. Studies have shown that retention and transport of colloids and microorganisms (such as viruses and bacteria) in porous media are functions of many factors. These include solution chemistry (ionic strength, composition, and pH of the bulk solution), physical and chemical properties of the porous media, flow conditions, and properties of the colloids. Our current understanding of the processes involved in colloid retention and transport and the factors that influence them can be found in several reviews (5-8). Only the literature relevant to the specific processes and factors addressed in this study, i.e., medium chemical heterogeneity, degree of water saturation (or unsaturated flow conditions), and solution ionic strength, is briefly reviewed in this Introduction. Effect of Ionic Strength. Ionic strength of the solution can significantly influence colloid deposition and transport, as shown in numerous studies (6, 9-14). Bacteria displayed decreased sticking efficiency and adhesion onto negatively charged quartz sand as the ionic strength decreased (13, 14). This was attributed to the increased thickness of the electrical double layer around the surfaces of both the bacteria and collector grains. Redman et al. (11) suggested that enhanced bacterial deposition at high ionic strength is due to the cells’ ability to enter into the second energy minimum, which increases in depth with increasing ionic strength. In column experiments, Chu et al. (9) examined the adsorption behavior of bacteriophages MS-2 and φX174 in response to the changes of solution ionic strength and showed that the removal of MS-2 was decreased when the ionic strength was decreased from 0.163 to 0.002 M whereas φX174 was not affected. Saiers and Lenhart (10) found that the ionic strength affected the transport of silica colloids through unsaturated sand columns. As the ionic strength increased from 2 × 10-4 to 0.2 M, the dominant mechanism of colloid retention changed from film straining to air-water interface (AWI) sorption and to mineral-grain attachment. The effect of ionic strength on colloid attachment has mainly been attributed to electrical double layer interactions, which can be explained with the Derjaguin-Landau-Verwey-Overbeek (DLVO) theory (10, 12). Effect of Water Content. Research on colloid retention and transport in the vadose zone has increased considerably in recent years because of the important role it plays in the overall understanding of colloid behavior in the subsurface. Reduced colloid mobility under unsaturated flow conditions as compared to saturated conditions has been widely reported on the basis of results from mostly laboratory-scale column and micromodel studies (15-24). These studies have mainly focused on exploring pore-scale mechanisms that are responsible for colloid retention and the various factors that influence them. Several mechanisms have been proposed in the literature to account for increased removal of colloids under unsaturated conditions. For example, film straining can occur if the diameter of a colloidal particle is greater than the thickness of the water film around the solid particles (18, 25). Colloids can also attach to the AWI, an additional interface that exists only in unsaturated media (10, 15, 17, 23, 26). Retention of colloids can also occur on the contact line between an AWI and a solid-water interface (SWI) (24, 27) or the triple-phase boundary (28). Furthermore, colloid VOL. 40, NO. 5, 2006 / ENVIRONMENTAL SCIENCE & TECHNOLOGY
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retention to the SWI may increase under unsaturated conditions, especially at high ionic strengths (10, 19, 20). Effect of Heterogeneity. Most previous studies on the retention and transport of colloids in porous media were conducted in physically and chemically homogeneous media. The effect of the heterogeneous nature of porous media on the transport of colloids (including viruses) has been largely ignored. A majority of the studies conducted in unsaturated media have mainly focused on pore-scale mechanisms of colloid retention and the effects of chemical and geochemical conditions on these mechanisms. Such studies naturally are only possible when experiments are simplified by using model porous media (e.g., chemically and physically homogeneous with a narrow particle size distribution) and under idealized flow regimes. However, as pointed out by Keller and Sirivithayapakorn (29), the already very complex water flow and colloid transport processes in unsaturated media due to flow discontinuity, interfacial processes, the wetting history, and transient effects are further complicated when the system is heterogeneous. Heterogeneity can cause greater variation in flow velocity, increase dispersion, and result in preferential flow. Gamerdinger and Kaplan (30) studied the transport and deposition behavior of hydrophilic latex particles under variably saturated conditions and found that, within a range of saturation levels, as the water content decreased, the colloid deposition rate was increased. However, when the water content was reduced to a very low level where most water was associated with solid surfaces, colloid retention decreased. They suggested that immobile water, excluded from the convective pore volume, was associated with a portion of the favorable sites for deposition, which made these sites inaccessible to the colloids. Studies on the effect of medium heterogeneity and the resulting complex flow regime on colloid transport have been very limited and mainly restricted to model simulations. For example, physical and geochemical heterogeneities were included in a two-dimensional colloid transport model by Sun et al. (31). Their modeling results demonstrate that both types of heterogeneity affect colloid transport by generating preferential flow paths but the effect of physical heterogeneity is more significant than geochemical heterogeneity. A more recent study using a two-dimensional stochastic model indicates that the effect of geochemical heterogeneity is important only when it is correlated to that of physical heterogeneity (varying conductivity) (32). While the effect of physical heterogeneity to cause preferential transport is more straightforward, the effect of chemical heterogeneity on colloid retention and transport seems subtle and perhaps more complex. The focus of this study is to evaluate the effect of the heterogeneity of the porous medium chemical properties on colloid transport and retention under saturated and unsaturated flow conditions. To isolate the influence of chemical heterogeneity from that of physical heterogeneity, experiments were conducted in uniformly packed columns (to eliminate physical heterogeneity) with glass beads having different fractions coated with a hydrophobic organic compound (to create varying degrees of surface heterogeneity). Two bacteriophages, MS-2 and φX174, were used, and experiments were run over a range of ionic strength from 0.025 to 0.163 M in a phosphatebuffered saline (PBS) solution. Two additional columns packed with chemically heterogeneous glass beads containing water at an irreducible content were scanned using highresolution synchrotron X-ray microtomography to nondestructively examine the water distribution.
Materials and Methods Porous Media. Glass beads (P-0230, Potters Industries Inc., Chelmsford, MA) with diameters of 0.43-0.6 mm were used 1548
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in this study. The main component of the glass beads is sodalime glass with an average density of 2.475 g cm-3. Hydrophilic Glass Beads. A procedure modified from Chu et al. (20) was used to remove metal oxides and other impurities from the glass beads. Briefly, 300 g of glass beads were soaked in 500 mL of 0.2 M citrate buffer solution containing 44.1 g L-1 sodium citrate (Na2C6H5O7‚2H2O) and 10.5 g L-1 citric acid (H2C6H5O7) in a flask and kept in an oven at 80 °C overnight. Then, 15 g of sodium dithionite (Na2S2O4) was added to the flask, and the flask was hand-shaken thoroughly several times. The solution was decanted and the procedure repeated three additional times. The beads were rinsed with tap water and deionized water several times and oven dried at 105 °C for 24 h. The glass beads cleaned following the above procedure are referred to as hydrophilic glass beads. Hydrophobic Glass Beads. The hydrophilic glass beads were coated with octadecyltrichlorosilane (OTS) following a procedure similar to that reported by Bales et al. (33). Three hundred grams of the hydrophilic glass beads were thoroughly rotated in 300 mL of solution containing 0.375 mL of OTS for 2 h and dried at 110 °C for 24 h. Fifty gram portions of the coated glass beads were washed with 150 mL of pentane two times. Two methanol rinses, two 1 M HCl rinses, and several deionized water rinses were performed until the rinse solution reached neutral pH. The beads were then oven dried at 105 °C for 24 h. The OTS-coated glass beads are referred to as hydrophobic glass beads. Contact Angle Measurement. The contact angles of the glass beads were measured using the Washburn method (34, 35). The hydrophilic and hydrophobic glass beads were separately packed into small columns with porous bottoms. When the beads were brought into contact with a water surface, the mass of water rising into the medium was measured as a function of time. According to the following relationship, the slope of the graph of time vs the square of the mass is η/(CF2γ cos θ). The contact angle θ is then calculated from the equation
cos θ )
η Μ2 CF2γ T
where η is the viscosity of water (0.001 N s m-2 at 20 °C), C is the material constant characteristic of the solid sample, F is the density of water (0.998 g cm-3 at 20 °C), γ is the surface tension of water (0.0729 J m-2 at 20 °C), M is the mass of water adsorbed on the medium, and T is the time after contact with the water surface. The measured contact angle is ∼0° and 65.9° for the hydrophilic and the hydrophobic beads, respectively. Viruses and Viral Assay. Two bacteriophages, MS-2 and φX174, were selected in this study because they have been used as surrogates for pathogenic enteric viruses in many studies and they have different surface properties arising from their different isoelectric points and surface hydrophobicities (7, 8). Bacteriophage MS-2 is an icosahedral single-stranded RNA virus with a diameter of 24-26 nm and has a low isoelectric point of 3.9 (36, as cited in ref 16). MS-2 was obtained from the American Type Culture Collection (ATCC 15597-B1) and grown on bacterial lawns of Escherichia coli (ATCC 15597). φX174 is a spherical single-stranded DNA bacteriophage with a diameter of 25-27 nm (37, 38) and an isoelectric point of 6.6 (39, 40, as cited in ref 41). φX174 was also obtained from the ATCC (ATCC 13706-B1) and grown on bacterial lawns of E. coli (ATCC 13706). MS-2 is more hydrophobic than φX174, which is one of the most hydrophilic viruses (42-44). Both viruses were assayed by the plaqueforming unit (pfu) method as described by Adam (45). Each sample was assayed in duplicate, and the average of two plate counts is reported.
TABLE 1. Summary of the Column Experiments and Experimental Parameters
porous medium hydrophilic glass beads
50% hydrophilic and 50% hydrophobic glass beads
degree of saturation (%)
ionic strength (mM)
100 100 16 20 100 100 100 17 19 16
25 100 25 100 25 100 163 25 100 163
Transport Experiments. Saturated and unsaturated experiments were conducted with the two viruses in PBS solution (containing Na2HPO4, NaCl, and KCl) at ionic strengths ranging from 0.025 to 0.163 M. (Detailed information on the composition of PBS buffer is given in Table S1, Supporting Information.) The columns were packed with either 100% hydrophilic beads (hydrophilic beads) or a 1:1 mixture of hydrophilic and hydrophobic beads (mixed beads). We used the same saturated and unsaturated column setups as presented in Jin et al. (19), which are briefly described here. The columns were 3.8 cm in diameter and 10.0 cm long. A stainless steel micronic mesh 0.075 mm thick and with pore sizes of 2.0-7.5 µm (GWP Inc., Berkeley, CA) was used to provide pressure control in the unsaturated columns. The same mesh was also placed at the bottom of the saturated columns to maintain conditions consistent with those of the unsaturated experiments. For the saturated experiments, influent was introduced from the bottom of the column and a fraction collector was connected to the top with Teflon tubing for sample collection. For the unsaturated experiments, a small solution-filling column with seven evenly distributed syringe needles was positioned above the transport column to supply input solution uniformly. The bottom outlet of the column was connected to a vacuum chamber with a fraction collector inside. The water content was adjusted by changing the vacuum chamber pressure and the input flow rate. Two tensiometers, used to monitor whether water was indeed distributed uniformly in the column, were installed at depths of 3.3 and 6.6 cm, respectively, on each side of the column. All experiments were conducted at 4-6 °C to minimize possible viral inactivation due to high temperature. The hydrophilic glass beads were wet packed for each experiment, whereas the mixed beads were dry packed and flushed with CO2 gas for 4 h to ensure complete air removal from the medium. Thereafter, each column was saturated and equilibrated with 20 pore volumes (PVs) of autoclaved and deaerated PBS solution at defined ionic strength (0.025, 0.100, or 0.163 M) to standardize the chemical conditions for each experiment and to establish a steadystate flow. The degree of water saturation was kept at ∼20% for all unsaturated experiments. To minimize the effect of shear stress or hydrodynamics, the experiments were designed to have similar interstitial or pore-water velocities (Table 1). A summary of the experiments and the relevant experimental parameters are listed in Table 1. Input solution containing approximately 5 × 105 pfu mL-1 of each virus (MS-2 and φX174) and 0.05 g L-1 KBr tracer in the PBS buffer was introduced into the columns as a step input. The effluent concentration of Br- tracer was determined by ion chromatography (IC) using an IonPac AS9-HC 4 mm anion-exchange column (Dionex Corp., Sunnyvale, CA) and with 9 mM sodium carbonate as the eluant at a flow rate of 1.0 mL min-1. A 3% (w/v) beef extract solution (BEX) adjusted to pH 9.5 was used to flush the columns immediately
porosity
interstitial velocity (cm/min)
dispersion coeff (cm2/s)
0.37 0.38 0.38 0.36 0.37 0.38 0.37 0.36 0.37 0.35
0.287 0.265 0.450 0.338 0.309 0.283 0.303 0.370 0.373 0.513
0.000298 0.000281 0.00174 0.00130 0.000249 0.000255 0.000224 0.002786 0.00520 0.01019
mass recovery (%) OX174 MS-2 84 103 111 101 78 76 74 92 86 83
79 67 69 13 91 37 3.2 92 71 38
after each experiment was completed to elute reversibly sorbed viruses. The unsaturated columns were resaturated with the BEX before flushing to avoid possible inactivation by unsaturated flow. BEX has been commonly used to detach viruses from various sorbents and has been shown to completely recover sorbed viruses in previous studies (19, 46, 47). The mass balance information obtained from BEX elution together with the measured virus breakthrough concentrations were used to examine whether the retained viruses were reversibly sorbed or inactivated/irreversibly sorbed.
Results Tracer Breakthrough Curves. Bromide (Br-) breakthrough curves (BTCs) (see Figure S1 in the Supporting Information) from all experiments were fitted with the convectiondispersion equation (CDE) (48) for a conservative solute to estimate dispersion coefficients (D) (Table 1). The estimated D values from the unsaturated experiments are consistently greater than those from the saturated experiments in both the hydrophilic and mixed media. Under the same ionic strength, they are also greater for mixed glass beads than hydrophilic beads. Virus Transport through 100% Hydrophilic Glass Beads. Breakthrough curves of MS-2 and φX174 through columns of the 100% hydrophilic glass beads at ionic strengths of 0.025 and 0.100 M are plotted in Figure 1. The φX174 breakthrough concentrations reached a plateau level quickly within the first 2-3 pore volumes under both saturated and unsaturated conditions and at both ionic strengths. MS-2 breakthrough also reached plateau concentrations at 0.025 M in all experiments. However, at an ionic strength of 0.100 M, the outflow concentration of MS-2 was much lower than under all other conditions and continued to increase during the entire duration of the unsaturated experiment while it reached a plateau quickly under saturated conditions. More MS-2 than φX174 was retained in all experiments. Transport of MS-2 decreased with increasing ionic strength and decreasing water content, while the effect of the ionic strength and water content was insignificant on the transport of φX174. As the ionic strength increased from 0.025 to 0.100 M, mass recoveries for MS-2 decreased from 79% to 67% in saturated experiments and from 69% to 13% in unsaturated experiments. On the other hand, ∼100% (100% ( 11%) of φX174 was recovered in all experiments. The mass recovery data indicate that at least 78% of MS-2 removed during transport was inactivated and at the same ionic strength inactivation was greater under unsaturated than saturated flow conditions. Virus Transport through Mixed Glass Beads. Breakthrough curves of MS-2 and φX174 through the mixed hydrophilic and hydrophobic beads at ionic strengths of 0.025, 0.100, and 0.163 M are presented in Figure 2. The BTCs for both viruses showed an initial rapid rise followed VOL. 40, NO. 5, 2006 / ENVIRONMENTAL SCIENCE & TECHNOLOGY
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FIGURE 1. Virus transport through hydrophilic glass beads at different solution ionic strengths: (a) OX174 in 0.025 M PBS; (b) MS2 in 0.025 M PBS; (c) OX174 in 0.100 M PBS; (d) MS-2 in 0.100 M PBS. by a slower increase at 0.100 and 0.163 M, while plateau concentrations were reached quickly within the first two volumes at 0.025 M. The outflow concentrations of both viruses decreased as the ionic strength was increased. MS-2 was more sensitive to the change in ionic strength than φX174. Contrary to the results reported in the literature and the general belief that more colloids are retained under unsaturated flow conditions, we found that there was less retention of MS-2 and φX174 in the unsaturated columns than in the saturated columns, especially at high ionic strengths. This “reversed trend” in breakthrough between saturated and unsaturated columns was more significant for MS-2 than for φX174. The lower mass recoveries of MS-2 and φX174 (Table 1) from the saturated columns as compared to the unsaturated columns agree well with the breakthrough concentrations, which indicates that most of the viruses removed during transport were inactivated. While there were no significant differences between the numbers of φX174 particles recovered at different ionic strengths at any given water saturation, the recovery of MS-2 decreased significantly as the ionic strength was increased.
Discussion Effect of Chemical Heterogeneity of Porous Media on Retention and Transport of MS-2 and OX174. The retention and transport of MS-2 and φX174 between saturated and unsaturated conditions in the mixed medium (Figure 2d-f) exhibit a trend completely opposite that in the hydrophilic medium (Figure 1c,d). Lower retention under saturated than unsaturated flow conditions in the homogeneous hydrophilic 1550
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columns is consistent with results reported in the literature (19, 20, 26). However, the reversed trend of greater removal in saturated columns of the chemically heterogeneous glass beads is contrary to the commonly reported colloid transport behavior in unsaturated media (9, 10, 17, 19, 21, 26). We provide the following discussion to explain these observations. To help understand these observations, we packed two columns with glass beads that have the same composition but smaller particle sizes (0.3-0.42 mm in diameter) than those used in the virus experiments at a water (doped with 10 wt % CsCl) saturation of ∼8%. These columns were imaged by synchrotron X-ray microtomography at the GeoSoilEnviroCARS (GSECARS) 13-BM-D beamline at the Advance Photon Source, Argonne National Laboratory. Sections of each column were imaged at a resolution of 10.92 µm at two energy levels: below (35.88 keV) and above (36.08 keV) the Cs X-ray absorption edge. Parts a and b of Figure 3 show one cross-section from each column obtained at the higher energy. The bright regions are locations where water is trapped. The three phases (solid, water, and air) were separated using the two images, a difference image (i.e., the image formed by subtracting the below edge image from the above edge image), and an indicator kriging algorithm (49) for image segmentation. Once the phases were separated, algorithms (50, 51) were used to quantify the system at the pore level. While the image resolution is high, it only captures the water trapped as pendular rings at the grain-grain contacts and within some of the pore bodies and throats, and not any water films. Nevertheless, the tomography images (Figure 3) show very uniform water distribution in the
FIGURE 2. Virus transport through mixed glass beads at different solution ionic strengths: (a) OX174 in 0.025 M PBS; (b) MS-2 in 0.025 M PBS; (c) OX174 in 0.100 M PBS; (d) MS-2 in 0.100 M PBS; (e) OX174 in 0.163 M PBS; (f) MS-2 in 0.163 M PBS. hydrophilic beads, whereas bigger and fewer water blobs are the main feature in the mixed beads. These blobs are formed due to the inability of the water phase to drain through some of the more hydrophobic regions. In the chemically homogeneous medium, more water was used to form films around the hydrophilic beads, allowing nearly complete drainage of the system and leaving less available to accumulate in corner ducts of the pore spaces. On the contrary, in the chemically heterogeneous medium, water formed fewer rings and was distributed as larger, but isolated, blobs, causing significant flow discontinuity. Analysis of the microtomography images indicates that the number of blobs (i.e., any water not trapped as a film) in the chemically heterogeneous medium was 4 times less than in the homogeneous medium of the same volume. The configuration of water in the hydrophilic glass beads could provide closer contact between the viruses and the interfaces (i.e.,
solid-water and air-water interfaces), resulting in increased virus retention, whereas flow bypassed water-repellent regions in the partially hydrophobic mixed beads, making a fraction of favorable attachment sites inaccessible to the virus particles. Therefore, transport of viruses through the unsaturated chemically heterogeneous medium was greatly enhanced. This phenomenon was more pronounced at high ionic strength when interactions between the viruses and the solid surface were more significant. Viral-Interfacial Interactions on the Pore Scale. Although we believe that flow dynamics played a dominant role in controlling the retention and transport of the viruses, especially in the chemically heterogeneous medium, interactions between the viruses and the various interfaces also contributed to the observed results. Both the AWI and SWI were likely involved in the interactions, although the relative importance of each mechanism could not be determined VOL. 40, NO. 5, 2006 / ENVIRONMENTAL SCIENCE & TECHNOLOGY
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FIGURE 3. Synchrotron-based microtomography images: (a, top left) above edge image in the 100% hydrophilic beads; (b, top right) above edge image in the 50% hydrophilic and 50% hydrophobic beads; (c, bottom left) water rings and blobs in the 100% hydrophilic beads; (d, bottom right) water rings and blobs in the 50% hydrophilic and 50% hydrophobic beads. from the available data. Additionally, attachment on the contact line of the AWI and SWI, which was only present in the mixed medium, also likely contributed to the overall virus retention. Effect of Film Straining. Film straining is considered as another possible mechanism of colloid removal in unsaturated systems (10, 18). We did not observe any differences in φX174 breakthrough between saturated and unsaturated columns at a low ionic strength of 0.025 M (Figures 1a and 2a). Therefore, we believe that film straining did not play a role in the retention of φX174. This conclusion also applies to the other ionic strengths tested because film straining remains relatively constant over a wide range of ionic strengths on the basis of analysis by Saiers and Lenhart (10). At similar levels of water saturation and flow velocity, the extent of film straining is controlled by the particle size (10, 18, 25). Because MS-2 and φX174 are very similar in size (24-26 and 25-27 nm, respectively), it is also reasonable to conclude that film straining had a negligible contribution to the removal of MS-2. Effect of Ionic Strength on Retention and Transport of MS-2 and OX174. The normalized breakthrough concentrations of MS-2 and φX174 decreased as the ionic strength was increased in all experiments. This may be attributed to the increased viral attachment onto the SWI in saturated columns and onto both the SWI and AWI, as well as the air-water1552
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solid contact line in the mixed medium, in unsaturated columns. The observed ionic strength effect is likely the result of electrostatic interactions between the viruses and the interfaces, a well-known behavior that can be described by the DLVO theory. At the experimental pH of 7.5, both the AWI and SWI were negatively charged. Approach of the negatively charged viruses to either interface would result in a repulsive electrical double layer force between them, and the potential energy barrier is a function of ionic strength. The magnitude of the potential energy barrier decreases with an increase of the ionic strength as a result of the shrinking electrical double layer thickness. In turn, closer proximity between the viruses and interfaces resulting from the increase in ionic strength allowed short-range attractive forces to become effective, which further promoted the capture and attachment of the viruses onto the AWI and SWI. Effect of Hydrophobic Interactions on Virus Sorption/ Inactivation. Results from the saturated experiments indicate that the presence of hydrophobic beads increased the retention of both MS-2 and φX174. Because attachment to the SWI was the only retention mechanism in saturated media, the increased viral removal in the mixed medium over the hydrophilic medium can be attributed to hydrophobic interactions between viral particles and the hydrophobic beads. When two hydrophobic surfaces approach each other, water molecules around the hydrophobic surfaces
are displaced into the bulk solution to reduce the total free energy of the system because the orientation of water molecules in contact with a hydrophobic surface is entropically unfavorable (52). Moreover, the attractive hydrophobic force operating on hydrophobic particles or grain surfaces comes into play at a greater separation distance where the double layer force is insignificant. A surface force measurement by atomic force microscopy showed the existence of large non-DLVO attractive forces between glass spheres and an OTS hydrophobized silica plate, which was attributed to hydrophobic interactions (53). The findings from the present study are consistent with previous reports, i.e., interactions of the relatively more hydrophobic MS-2 with the hydrophobic medium are the strongest, while interactions of the relatively more hydrophilic φX174 with the hydrophilic beads are the weakest. As discussed previously, decreased retention of MS-2 and φX174 in the unsaturated mixed medium may be partially attributed to the reduced contact of viruses with solid surfaces due to nonuniform distribution of water and preferential flow generated by the water-repellent beads. However, it is also possible that virus interactions with the AWI were not as significant as the interactions with the hydrophobic surfaces because the equivalent contact angle of an AWI is less than that of the hydrophobic beads (i.e., the AWI is less hydrophobic). Furthermore, the significantly different behaviors MS-2 and φX174 displayed are also an indication that hydrophobic interactions played an important role. Less than 5% of MS-2 moved through the unsaturated hydrophilic glass beads, while φX174 was not retained at all at a 0.100 M ionic strength. This difference cannot be explained by the repulsive electrical double layer interaction experienced by the viruses because MS-2 is more negatively charged than φX174 at the experimental pH. Rather, this was probably caused by increased attachment of MS-2 to both the AWI and surfaces of the hydrophobic glass beads because MS-2 is more hydrophobic than φX174. The difference in retention between MS-2 and φX174 was more significant at higher ionic strength, which indicates that hydrophobic interaction also depends on the ionic strength. Adding salts changes the original arrangement of water molecules on grain surfaces, which can result in changes in the orientation of the first several layers of water when the solution ionic strength is varied. Therefore, the magnitude of the hydrophobic force can be affected. The exact relationship between the magnitude of the hydrophobic force and the ionic strength needs further examination. Interaction of the Ionic Strength, Grain Surface Property, and Water Content. As discussed in previous sections, effects of the various factors examined in this study on virus retention and transport are interactive and very complex. We believe that the combined effect of unsaturated flow conditions and the chemical heterogeneity of the grain surfaces, which produced preferential flow, was mainly responsible for the observations. In a recent paper, Saiers and Lenhart (10) developed a relationship between the colloid-retention capacity of a porous medium and the degree of water saturation, and analyzed colloid retention by the AWI and SWI at different ionic strengths. Their calculations revealed interesting and complex interactions between water content and ionic strength with respect to their effects on colloid retention. Analysis by Saiers and Lenhart indicated that the dominant mechanism responsible for colloid retention, attachment to either the AWI or the SWI, changed with ionic strength, and that a transition from water-saturated to unsaturated conditions could diminish the overall capacity of a porous medium to remove colloids from pore water as the ionic strength was increased beyond a critical value, which was reported to be
between 0.1 and 0.2 M. Although we believe that preferential transport was the primary reason for the reversed trend between saturated and unsaturated virus transport observed in this study, the analysis by Saiers and Lenhart offers an additional possible mechanism. Results from this study clearly show that the presence of an air phase in unsaturated porous media does not always impede colloid transport. This is contrary to what the current literature commonly suggests. In the chemically heterogeneous porous medium where a fraction of the glass beads were coated with a hydrophobic organic material, nonuniform wetting resulted in heterogeneous water distribution, which induced preferential flow (fewer and larger flow paths) under unsaturated flow conditions. The additional attachment of virus particles to the AWI was not able to counter the enhanced virus transport due to preferential flow, which resulted in bypassing of favorable attachment sites on the hydrophobic bead surfaces. Furthermore, interactions of ionic strength, degree of water saturation, and grain surface properties resulted in very complex interactive effects on the virus attachment kinetics to both the AWI and SWI. We also found that virus retention increased with increasing ionic strength, regardless of whether the retention was caused by electrostatic or hydrophobic interactions. It is important to note that, at low ionic strength, which is more prevalent in natural systems, the differences between saturated and unsaturated virus transport were not significant. In addition, under the experimental conditions employed in this study, we observed that hydrophobic interactions dominated over other forces in both range and magnitude. Our current understanding of colloid retention and transport in porous media is largely derived from studies using homogeneous media, and most mechanistic information has been obtained through pore-scale micromodel studies. The current study shows that it is critical to extend the range of physicochemical conditions in future investigations to advance our knowledge of colloid retention and movement in the subsurface. Organic materials are present in almost all subsurface media, especially in the vadose zone. Viruses are often released into the subsurface through media such as septic tank liquids that contain high concentrations of organic carbon. The organic materials can be present as films coated on surfaces of solid grains, which alters the surface charge and aggregation behavior of metal oxides and layered silicate minerals. These reactions can greatly affect virus-particle interactions at the SWI in saturated media and therefore influence virus mobility. This problem is accentuated in unsaturated systems, where heterogeneous coating of hydrophobic organic materials also causes nonuniform wetting and creates complex water flow patterns, which can overshadow any pore-scale mechanisms that control colloid retention and transport in homogeneous media. The reported behavior can be expected to occur in, for example, water-repellent soils. Because water flow in unsaturated systems is expected to be more complex with greater velocity variations and dispersion, especially when the medium is physically and/or chemically heterogeneous, it is reasonable to assume that, under those conditions, hydrodynamic effects on the mesopore scale will dominate pore-scale interfacial reactions in controlling the extent of colloid movement. Therefore, future studies on colloid retention and transport, especially in variably saturated media, need to take both physical and chemical heterogeneity into consideration.
Acknowledgments This project was supported by National Research Initiative Competitive Grant No. 2001-35107-01235 from the USDA Cooperative State Research, Education, and Extension Services (J.H. and Y.J.) and by the National Science Foundation VOL. 40, NO. 5, 2006 / ENVIRONMENTAL SCIENCE & TECHNOLOGY
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under Grant No. EAR-0207788 (C.S.W.). We thank Dr. Markus Flury of Washington State University for his assistance in measuring the contact angles of the glass beads. Analysis of the microtomography images was performed using the Casper system at Louisiana State University’s high-performance computing center. Use of the Advanced Photon Source was supported by the U.S. Department of Energy, Office of Science, Office of Basic Energy Sciences, under Contract No. W-31-109-Eng-38. We appreciate the constructive reviews provided by the three anonymous reviewers.
Supporting Information Available Detailed information on the ionic strength and composition of the phosphate buffer solution and bromide breakthrough curves from columns packed with hydrophilic beads and mixed beads. This material is available free of charge via the Internet at http://pubs.acs.org.
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Received for review July 12, 2005. Revised manuscript received December 13, 2005. Accepted December 20, 2005. ES051351M
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