VIRTUS, a model of virus transport in unsaturated soils.

As a result of the recently proposed mandatory groundwater disinfection requirements to inactivate viruses in potable water supplies, there has been increasing interest in virus fate and transport in the subsurface. Several models have been developed to predict the fate of viruses in groundwater, but few include transport in the unsaturated zone and all require a constant virus inactivation rate. These are serious limitations in the models, as it has been well documented that considerable virus removal occurs in the unsaturated zone and that the inactivation rate of viruses is dependent on environmental conditions. The purpose of this research was to develop a predictive model of virus fate and transport in unsaturated soils that allows the virus inactivation rate to vary on the basis of changes in soil temperature. The model was developed on the basis of the law of mass conservation of a contaminant in porous media and couples the flows of water, viruses, and heat through the soil. Model predictions were compared with measured data of virus transport in laboratory column studies and, with the exception of one point, were within the 95% confidence limits of the measured concentrations. The model should be a useful tool for anyone wishing to estimate the number of viruses entering groundwater after traveling through the soil from a contamination source. In addition, model simulations were performed to identify parameters that have a large effect on the results. This information can be used to help design experiments so that important variables are measured accurately.     

Biodegradation, sorption, and transport of 2,4-dichlorophenoxyacetic acid in saturated and unsaturated soils.

The fate of an organic contaminant in soil depends on many factors, including sorption, biodegradation, and transport. The herbicide 2,4-dichlorophenoxyacetic acid (2,4-D) was used as a model compound to illustrate the impact of these interacting factors on the fate of an organic contaminant. Batch and column experiments performed with a sandy loam soil mixture under saturated and unsaturated conditions were used to determine the effects of sorption and biodegradation on the fate and transport of 2,4-D. Sorption of 2,4-D was found to have a slight but significant effect on transport of 2,4-D under saturated conditions (retardation factor, 1.8) and unsaturated conditions (retardation factor, 3.4). Biodegradation of 2,4-D was extensive under both batch and column conditions and was found to have a significant impact on 2,4-D transport in column experiments. In batch experiments, complete mineralization of 2,4-D (100 mg kg-1) occurred over a 4-day period following a 3-day lag phase under both saturated and unsaturated conditions. The biodegradation rate parameters calculated for batch experiments were found to be significantly different from those estimated for column experiments.     

Effects of organic matter on virus transport in unsaturated flow.

The effects of natural humic material and sewage sludge organic matter (SSOM) derived from primary treated sewage sludge on virus transport by unsaturated flow through soil columns were evaluated. Bacteriophage MS-2 was applied to loamy fine sand columns 0.052 m in diameter and 1.05 m long. Virus concentrations in the influent and effluent were measured daily for 7 to 9 days. In the first experiment, virus transport through two fresh soil columns was compared with that through a column previously leached with more than four pore volumes (T) of well water. The soil water organic matter concentrations in the leachate of the fresh soil declined with time. Relative virus concentrations (C/Co) from one fresh soil column reached 0.82 in 0.9 T and then declined to 0.51 by 2.1 T. The other fresh soil column reached and maintained a steady-state relative virus concentration [(C/Co)s] of 0.47 from 1.5 to 2.5 T. The leached column reached and maintained a (C/Co)s of 0.05. Concentrations measured at 0.2-, 0.4-, 0.8-, and 1.05-m depths indicated that most virus particles were removed in the surface 0.2 m. In the second experiment, one leached column was pretreated with SSOM derived from primary treated sewage sludge and the other leached column was untreated. SSOM concentrations declined with depth. A suspension of virus and SSOM in well water was applied to both columns. Although the (C/Co)s values were similar (0.41 for the pretreated column and 0.47 for the untreated column), breakthrough was delayed for the untreated column. Both natural humic material and sewage sludge-derived SSOM increased the unsaturated-flow transport of MS-2.     

Effects of organic matter on virus transport in unsaturated flow.

The effects of natural humic material and sewage sludge organic matter (SSOM) derived from primary treated sewage sludge on virus transport by unsaturated flow through soil columns were evaluated. Bacteriophage MS-2 was applied to loamy fine sand columns 0.052 m in diameter and 1.05 m long. Virus concentrations in the influent and effluent were measured daily for 7 to 9 days. In the first experiment, virus transport through two fresh soil columns was compared with that through a column previously leached with more than four pore volumes (T) of well water. The soil water organic matter concentrations in the leachate of the fresh soil declined with time. Relative virus concentrations (C/Co) from one fresh soil column reached 0.82 in 0.9 T and then declined to 0.51 by 2.1 T. The other fresh soil column reached and maintained a steady-state relative virus concentration [(C/Co)s] of 0.47 from 1.5 to 2.5 T. The leached column reached and maintained a (C/Co)s of 0.05. Concentrations measured at 0.2-, 0.4-, 0.8-, and 1.05-m depths indicated that most virus particles were removed in the surface 0.2 m. In the second experiment, one leached column was pretreated with SSOM derived from primary treated sewage sludge and the other leached column was untreated. SSOM concentrations declined with depth. A suspension of virus and SSOM in well water was applied to both columns. Although the (C/Co)s values were similar (0.41 for the pretreated column and 0.47 for the untreated column), breakthrough was delayed for the untreated column. Both natural humic material and sewage sludge-derived SSOM increased the unsaturated-flow transport of MS-2.     

Estimation of contaminant depletion in unsaturated soils using a reduced-order biodegradation model and carbon dioxide measurement

The objective of this study was to develop a reduced-order model of biodegradation in unsaturated soils that allows the estimation of contaminant depletion, using available on-line measurements. Hexadecane was chosen as a model compound for petroleum hydrocarbons. A two-compartment model was developed, decoupling the intrinsic biodegradation kinetics from limiting factors imposed by field conditions, such as oxygen transfer and contaminant bioavailability. Two new experimental protocols (one for the liquid phase and the other for the solid phase) were developed to monitor hexadecane depletion, hexadecane mineralization, total mineralization, and evolution of the degraders. Using the liquid-phase experiment, parameters of a Haldane kinetic model and yield coefficients were identified and used in the complete model of biodegradation in soil. Using the carbon dioxide production curve, a biocontact kinetic model was identified so that, despite the high sensitivity of the model outputs to variations in the parameters, hexadecane depletion could be correctly predicted with an average error on the entire time trajectory of about 8%. Moreover, the ratio between hexadecane mineralization and total mineralization remained constant after a brief transient period, indicating that hexadecane mineralization could be deduced from the total carbon dioxide measurement. Finally, the new model developed in this study allows real-time monitoring of contaminant biodegradation, using on-line carbon dioxide measurement.     

RNA recombination in brome mosaic virus, a model plus strand RNA virus.

Studies on the molecular mechanism of genetic recombination in RNA viruses have progressed at the time when experimental systems of efficient recombination crossovers were established. The system of brome mosaic virus (BMV) represents one of the most useful and most advanced tools for investigation of the molecular aspects of the mechanism of RNA-RNA recombination events. By using engineered BMV RNA components, the occurrence of both homologous and nonhomologous crosses were demonstrated among the segments of the BMV RNA genome. Studies show that the two types of crossovers require different RNA signal sequences and that both types depend upon the participation of BMV replicase proteins. Mutations in the two BMV-encoded replicase polypeptides (proteins 1a and 2a) reveal that their different regions participate in homologous and in nonhomologous crossovers. Based on all these data, it is most likely that homologous and nonhomologous recombinant crosses do occur via two different types of template switching events (copy-choice mechanism) where viral replicase complex changes RNA templates during viral RNA replication at distinct signal sequences. In this review we discuss various aspects of the mechanism of RNA recombination in BMV and we emphasize future projections of this research.     

Cucumber mosaic virus, a model for RNA virus evolution

Taxonomic relationships: Cucumber mosaic virus (CMV) is the type member of the Cucumovirus genus, in the family Bromoviridae . Additional members of the genus are Peanut stunt virus (PSV) and Tomato aspermy virus (TAV). The RNAs 3 of all members of the genus can be exchanged and still yield a viable virus, while the RNAs 1 and 2 can only be exchanged within a species.
Physical properties: The virus particles are about 29 nm in diameter, and are composed of 180 subunits (T = 3 icosahedral symmetry). The particles sediment with an s value of approximately 98. The virions contain 18% RNA, and are highly labile, relying on RNA–protein interactions for their integrity. The three genomic RNAs, designated RNA 1 (3.3 kb in length), RNA 2 (3.0 kb) and RNA 3 (2.2 kb) are packaged in individual particles; a subgenomic RNA, RNA 4 (1.0 kb), is packaged with the genomic RNA 3, making all the particles roughly equivalent in composition. In some strains an additional subgenomic RNA, RNA 4A is also encapsidated at low levels. The genomic RNAs are single stranded, plus sense RNAs with 5' cap structures, and 3' conserved regions that can be folded into tRNA-like structures.
Satellite RNAs: CMV can harbour molecular parasites known as satellite RNAs (satRNAs) that can dramatically alter the symptom phenotype induced by the virus. The CMV satRNAs do not encode any proteins but rely on the RNA for their biological activity.
Hosts: CMV infects over 1000 species of hosts, including members of 85 plant families, making it the broadest host range virus known. The virus is transmitted from host to host by aphid vectors, in a nonpersistent manner.
Useful web sites: http://mmtsb.scripps.edu/viper/1f15.html (structure); http://www.ncbi.nlm.nih.gov/ICTVdb/ICTVdB/10040001.htm (general information)     

Ion transport in a model gramicidin channel. Structure and thermodynamics.

The potential of mean force for Na+ and K+ ions as a function of position in the interior of a periodic poly(L,D)-alanine model for the gramicidin beta-helix is calculated with a detailed atomic model and realistic interactions. The calculated free energy barriers are 4.5 kcal/mol for Na+ and 1.0 kcal/mol for K+. A decomposition of the free energy demonstrates that the water molecules make a significant contribution to the free energy of activation. There is an increase in entropy at the transition state associated with greater fluctuations. Analysis reveals that the free energy profile of ions in the periodic channel is controlled not by the large interaction energy involving the ion but rather by the weaker water-water, water-peptide and peptide-peptide hydrogen bond interactions. The interior of the channel retains much of the solvation properties of a liquid in its interactions with the cations. Of particular importance is the flexibility of the helix, which permits it to respond to the presence of an ion in a fluidlike manner. The distortion of the helix is local (limited to a few carbonyls) because the structure is too flexible to transmit a perturbation to large distances. The plasticity of the structure (i.e., the property to deform without generating a large energy stress) appears to be an essential factor in the transport of ions, suggesting that a rigid helix model would be inappropriate.     

Adsorption and movement of metalaxyl in soils under unsaturated flow conditions

Adsorption and mobility of the fungicide metalaxyl was studied in three soil types, viz., Hessaraghatta loamy sand (HLS), Bellary clay (BC), and Ooty clay (OC) soils, under laboratory conditions. The K values obtained from the Freundlich adsorption isotherms were 0.04, 10.65 and 13.32 µg/g for HLS, BC and OC soils respectively. Mobility of metalaxyl in these soils was inversely related to K values (r = –0.97). Although, metalaxyl persisted for over 60 days in all the soils, it did not move beyond a depth of 17.5 cm in any of the soils under unsaturated flow conditions. Mobility of metalaxyl was fastest in HLS and slowest in OC soils.     

Oligomerization, transport, and Golgi retention of Punta Toro virus glycoproteins.

We have investigated the oligomerization and intracellular transport of the membrane glycoproteins of Punta Toro virus, a member of the Phlebovirus genus of the family Bunyaviridae, which is assembled by budding in the Golgi complex. By using one- or two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis, chemical cross-linking, and sucrose gradient centrifugation, we found that the majority of the G1 and G2 glycoproteins are assembled into noncovalently linked G1-G2 heterodimers. At the same time, a fraction of the G2 protein, possibly produced independently of the G1 protein, is assembled into G2 homodimers. Kinetic analysis indicates that heterodimerization occurs between newly synthesized G1 and G2 within 3 min after protein synthesis, and that the G1 and G2 glycoproteins are associated as dimeric forms both during transport and after accumulation in the Golgi complex. Analysis of a G1-truncated G2 mutant, which is also targeted to the Golgi complex, showed that these molecules also assemble into dimeric forms, which are linked by disulfide bonds. Both the G1-G2 heterodimer and the G2 homodimer were found to be able to exit from the endoplasmic reticulum. Differences in transport kinetics observed for the G1 and G2 proteins may be due to the differences in the transport efficiency between the G1-G2 heterodimer and the G2 homodimer from the endoplasmic reticulum to the Golgi complex. These and previous results (S.-Y. Chen, Y. Matsuoka, and R.W. Compans, Virology 183:351-365, 1991) suggest that Golgi retention of the G2 homodimer occurs by association with the G1-G2 heterodimer, whereas the Golgi targeting of the G1-G2 heterodimer occurs by a specific retention mechanism.     

Dynamics of C,N, P and S in grassland soils: a model

We have developed a model to simulate the dynamics of C, N, P, and S in cultivated and uncultivated grassland soils. The model uses a monthly time step and can simulate the dynamics of soil organic matter over long time periods (100 to 10,000 years). It was used to simulate the impact of cultivation (100 years) on soil organic matter dynamics, nutrient mineralization, and plant production and to simulate soil formation during a 10,000 year run. The model was validated by comparing the simulated impact of cultivation on soil organic matter C, N, P, and S dynamics with observed data from sites in the northern Great Plains. The model correctly predicted that N and P are the primary limiting nutrients for plant production and simulated the response of the system to inorganic N, P, and S fertilizer. Simulation results indicate that controlling the C:P and C:S ratios of soil organic matter fractions as functions of the labile P and S levels respectively, allows the model to correctly simulate the observed changes in C:P and C:S ratios in the soil and to simulate the impact of varying the labile P and S levels on soil P and S net mineralization rates.     

Heterogeneous amino acid transport rate changes in an E. coli unsaturated fatty acid auxotroph.

Amino acid transport rates in an $\text{E. coli}$ unsaturated fatty acid auxotroph were non-uniformly affected by enrichment of membrane lipids in various unsaturated fatty acids. Proline and threonine transport rates were depressed much more than lysine and asparagine rates by trans unsaturated acids. Myristoleate and linolenate enrichment also produced non-uniform but lesser rate reductions. Although changes in the relative numoer of effective transport catalysts could account for these findings, comparisons of proline and lysine transport rates over a broad temperature range indicated that non-uniform alterations in transport catalyst reaction rates account at least partly for the activity changes associated with membrane lipid alterations.     

Ion transport across the chick ileum: a good model for transport studies.

1. The young chick (5-8 days) has been found to be an excellent preparation for the study of transepithelial intestinal ion transport. Due to the thinness of the intestinal tissue, it is not necessary to remove the serosal layers (serosal membranes, circular, and longitudinal muscles), thus circumventing the problems inherent in "stripping" the tissue. 2. The intact chick ileum had a significantly greater short-circuit current (Isc) and lower resistance than did intact adult ileum and transport parameters remained stable over the 6 hr experimental period. 3. Compared to the adult tissue, unidirectional fluxes of Na and Cl were greater in the chick ileum. Net flux of Na (absorption) was about 3 times greater in the chick ileum and the flux was equivalent to the Isc, thus this preparation appears to be characterized by electrogenic Na absorption. 4. Several ileal preparations from day old chicks were studied over an 18 hr period and these preparations were found to remain viable for this period of time with the Isc at the end of 18 hr being nearly identical to that at 2 hr. 5. Besides the advantage of not having to strip the intestinal tissue, and the long-term viability of the tissue, the chick is very inexpensive and easy to obtain and maintain.     

Processing and transport of environmental virus samples.

Poliovirus-seeded tap water, conditioned with MgCl2 and passed through virus-adsorbing filters, gave better poliovirus recovery than water identically treated but conditioned with AlCl3. Elution of several filter types with beef extract yielded higher recoveries than did elution with glycine. Seeded samples filtered through various filters and stored showed considerable virus loss in 2 days when stored at 4 degrees C, whereas those stored at -70 degrees C gave stable virus recovery up to 4 days. Additionally, the use of antifoam during the elution process reduced foaming and increased virus recovery by 28%.     

Variants of vaccinia virus hemagglutinin altered in intracellular transport.

Two classes of revertants were isolated from a vaccinia virus mutant whose hemagglutinins (HAs) accumulate on nuclear envelopes and rough endoplasmic reticulums. The HAs of one of the revertants had the same phenotype as the wild type, i.e., rapid and efficient movement to the cell surface. The HAs of the second class had biphasic transport: rapid export to the cell surface as in the wild type and slow movement to the medial cisternae of the Golgi apparatus. Biochemical and nucleotide sequence analyses showed that the HAs of all the mutants examined that have defects in transport from the rough endoplasmic reticulum to the Golgi apparatus have altered cytoplasmic domains and that the HAs of the second class of revertants lack the whole cytoplasmic domain, while the HAs of the first class of revertants have a wild-type cytoplasmic domain.     

Red queen dynamics, competition and critical points in a model of RNA virus quasispecies.

RNA viruses offer a unique opportunity for the study of evolution at the molecular level. Recent experiments involving clonal populations of RNA viruses have shown that competition among virus strains of approximately equal relative fitness can result in the eventual competitive exclusion of one of the species. As competition proceeds in time, both the winners and the losers exhibited absolute gains in fitness, consistent with the "Red Queen" hypothesis of evolution. Further experiments involving closely related evolving quasispecies revealed a highly predictable nonlinear behavior suggesting a deterministic component in the underlying quasispecies dynamics. This is apparently in contradiction with the standard view of RNA virus evolution as a highly unpredictable process. In this paper we present a simple model which allows previous hypothesis to be tested and provides an interpretation for the observed experimental results.