Craig L. Just

Gene Expression and Microscopic Analysis of Arabidopsis Exposed to Chloroacetanilide Herbicides and Explosive Compounds. A Phytoremediation Approach

Understanding the function of detoxifying enzymes in plants toward xenobiotics is of major importance for phytoremediation applications. In this work, Arabidopsis (Arabidopsis thaliana; ecotype Columbia) seedlings were exposed to 0.6 mm acetochlor (AOC), 2 mm metolachlor (MOC), 0.6 mm 2,4,6-trinitrotoluene (TNT), and 0.3 mm hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX). In vivo glutathione (GSH) conjugation reactions of AOC, MOC, RDX, and TNT were studied in root cells using a multiphoton microscope. In situ labeling with monochlorobimane, used as a competitive compound for conjugation reactions with GSH, confirmed that AOC and MOC are conjugated in Arabidopsis cells. Reverse transcription-PCR established the expression profile of glutathione S-transferases (GSTs) and nitroreductases enzymes. Genes selected for this study were AtGSTF2, AtGSTU1, AtGSTU24, and two isoforms of 12-oxophytodienoate reductase (OPR1 and OPR2). The five transcripts tested were induced by all treatments, but RDX resulted in low induction. The mRNA level of AtGSTU24 showed substantial increase for all chemicals (23-fold induction for AOC, 18-fold for MOC, 5-fold for RDX, and 40-fold for TNT). It appears that GSTs are also involved in the conjugation reactions with metabolites of TNT, and to a lesser extent with RDX. Results indicate that OPR2 is involved in plant metabolism of TNT (11-fold induction), and in oxidative stress when exposed to AOC (7-fold), MOC (9-fold), and RDX (2-fold). This study comprises gene expression analysis of Arabidopsis exposed to RDX and AOC, which are considered significant environmental contaminants, and demonstrates the importance of microscopy methods for phytoremediation investigations.

The effects of individual PCB congeners on the soil bacterial community structure and the abundance of biphenyl dioxygenase genes

Polychlorinated biphenyls (PCBs) are toxic environmental contaminants that represent a class of 209 congeners characterized by different degrees of chlorination and substitution patterns. Most of experimental studies about microbial degradation of PCBs have been conducted on PCB mixtures, even though evidence accumulated in bacteria and other organisms shows that exposure to different congeners may have different biological effects. Microcosm experiments were conducted using aerobic agitated soil slurries individually exposed to PCB congeners with different degrees of chlorination: PCB-3, 15, 28, and 77, and the commercial mixture Aroclor 1242. After four weeks of incubation, PCBs were analyzed by gas chromatography/mass spectrometry (GC/MS) showing different transformation extents: With the exception of PCB-15 that was not significantly transformed (7%), biodegradation rates decreased with the degree of chlorination, from 75% for PCB-3 to 22% for PCB-77 and Aroclor 1242. The bacterial abundance, as measured by colony counting and 16S rDNA quantification by real-time PCR, was lower (of about 40%) in soil microcosms exposed to the higher-chlorinated congeners, PCB-28, PCB-77, and Aroclor 1242, as compared to non-exposed soils and soils exposed to the lower-chlorinated congeners, PCB-3 and PCB-15. The relative abundance of different taxonomic groups, as determined by real-time PCR, revealed an increase of β-Proteobacteria and Actinobacteria in all microcosms exposed to PCBs, as compared with non-exposed soil. In addition, exposure to PCB-77 and Aroclor 1242 resulted in a higher abundance of α-Proteobacteria and Acidobacteria. Globally, these results suggest that exposure to PCBs (and especially to higher-chlorinated congeners and Aroclor 1242) selected bacterial groups involving most known PCB degraders, i.e., β-Proteobacteria and Acidobacteria. The quantification of biphenyl dioxygenase (BPH) genes--involved in the aerobic degradation of PCBs--using real-time PCR showed that exposure to all PCB congeners and Aroclor 1242 resulted in a marked increase of two out of the four BPH genes tested, similarly suggesting the selection of PCB-degrading bacteria. This paper showed that exposure to different PCB congeners leads to different structures of the soil bacterial community and BPH genes expression patterns.

Antecedent Moisture Controls on Stream Nitrate Flux in an Agricultural Watershed

Evaluating nitrate-N fluxes from agricultural landscapes is inherently complex due to the wide range of intrinsic and dynamic controlling variables. In this study, we investigate the influence of contrasting antecedent moisture conditions on nitrate-N flux magnitude and dynamics in a single agricultural watershed on intra-annual and rainfall-event temporal scales. High temporal resolution discharge and nitrate concentration data were collected to evaluate nitrate-N flux magnitude associated with wet (2009) and dry (2012) conditions. Analysis of individual rainfall events revealed a marked and consistent difference in nitrate-N flux response attributed to wet/dry cycles. Large-magnitude dilutions (up to 10 mg N L) persisted during the wet antecedent conditions (2009), consistent with a dominant baseflow contribution and excess groundwater release in relation to precipitation volume (discharge > > precipitation). Smaller-magnitude concentrations (<7 mg N L) were observed during the drought conditions of 2012, consistent with a quickflow-dominated response to rain events and infiltration/storage of precipitation resulting in discharge < precipitation. Nitrate-N loads and yields from the watershed were much higher (up to an order of magnitude) in the wet year vs. the dry year. Our results suggest that the response of nitrate-N loading to rain events is highly dependent on intra-annual antecedent moisture conditions and subsurface hydrologic connectivity, which together dictate the dominant hydrologic pathways for stream recharge. Additionally, the results of our study indicate that continued pronounced wet/dry cycles may become more dominant as the short-term driver of future nitrate-N exports.

Development of a Synthetic PCB Mixture Resembling the Average Polychlorinated Biphenyl Profile in Chicago Air

Studies of environmental and toxic effects of polychlorinated biphenyls (PCBs) are ideally performed with PCB mixtures reflecting the composition of environmental PCB profiles to mimic actual effects and to account for complex interactions among individual PCB congeners. Unfortunately, only a few laboratory studies employing synthetic PCB mixtures have been reported, in part because of the challenges associated with the preparation of complex PCB mixtures containing many individual PCB congeners. The objective of this study was to develop a PCB mixture that resembles the average PCB profile recorded from 1996 to 2002 at a satellite station of the Integrated Atmospheric Deposition Network located at the Illinois Institute of Technology (IIT) in Chicago, Illinois, using commercial PCB mixtures. Initial simulations, using published Aroclor profiles, showed that a mixture containing 65% Aroclor 1242 and 35% Aroclor 1254 was a good approximation of the target profile. A synthetic Chicago air mixture (CAM) was prepared by mixing the respective Aroclors in this ratio, followed by GC/MS/MS analysis. Comparison of the PCB profile of the synthetic mixture with the target profile suggests that the synthetic PCB mixture is a good approximation of the average IIT Chicago air profiles (similarity coefficient cos θ = 0.82; average relative percent difference = 84%). The synthetic CAM was also a reasonable approximation of the average of 184 PCB profiles analyzed in 2007 at 37 sites throughout Chicago as part of the University of Iowa Superfund Basic Research Program (isbrp), with a cos θ of 0.70 and an average relative percent difference of 118%. While the CAM and the two Chicago air profiles contained primarily di- to pentachlorobiphenyls, higher chlorinated congeners, including congeners with seven or eight chlorine atoms, were underrepresented in the synthetic CAM. The calculated TCDD toxic equivalency quotients of the synthetic CAM (2.7 ng/mg PCB) and the IIT Chicago air profile (1.6 ng/mg PCB) were comparable, but lower by two orders of magnitude than the isbrp Chicago air profile (865 ng/mg PCB) due to surprisingly high PCB 126 levels in Chicago air. In contrast, the calculated neurotoxic equivalency quotients of the CAM (0.33 mg/mg PCB) and the two Chicago air profiles (0.44 and 0.30 mg/mg PCB, respectively) were similar. This study demonstrates the challenges and methods of creating and characterizing synthetic, environmental mixtures of PCBs.

Mathematical modeling of RDX and HMX metabolism in poplar (Populus deltoides x Populus nigra, DN34) tissue culture

ABSTRACT Three mathematical models were developed based on a fate study as an approach to define transformation pathways of hexahydro-1,3,5-trinitro-1,3,5-triazine (RDX) and octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine (HMX) within plant cells. [U-14C]RDX and [U-14C]HMX were added in Murashige and Skoog (MS) liquid media containing Populus deltoides × P. nigra (DN34) tissue cultures. Radioactivity of samples was analyzed using HPLC, a bio-oxidizer and liquid scintillation counter. Based on information collected, transformation pathways of nitramine compounds were fitted with the raw data obtained and using a modified “green liver” model. Ordinary differential equations were developed and simulations were performed with MicroMath Scientist version 2.0 (MicroMath Inc., St. Louis, MO, USA). The three models, with different sequential transformation processes, were tested in order to support the raw data (model I) and the assumptions of the modified “green liver” model (models II and III). The results showed a high correlation between the collected data and the simulated concentrations for all models. Thus, the simplest model developed (model I) is the best model description of these particular results. The results obtained suggest that the principle of parsimony should be applied. The “green liver”-based models also demonstrated a reliable approach for the investigation of degradation pathways of nitramines within plant cells.

High-Frequency Diel Dissolved Oxygen Stream Data Modeled for Variable Temperature and Scale

Diel dissolved oxygen (DO) concentrations and temperature were sensed at high-frequency and modeled in an eastern Iowan stream, Clear Creek, in an agricultural setting. The magnitude of the diel changes in DO and temperature were largest at the upstream (headwater) station. Inclusion of temperature change factors increased the accuracy of modeling results and yielded estimates of the reaeration rate constant, primary production rate, and respiration rate. The DO modeling of the high-frequency measurements (15-min intervals) revealed a temperature-driven nonlinear reaeration process that led to increases in nighttime DO concentrations. The DO modeling results from three sensing stations in the watershed revealed decreasing trends in primary productivity, respiration, and the reaeration rate constant with increasing drainage area. Light extinction from suspended solids was the main factor limiting net primary production. As a result, the P/R ratio also decreased with increasing drainage area. High-frequency...

Uses and Biases of Volunteer Water Quality Data

State water quality monitoring has been augmented by volunteer monitoring programs throughout the United States. Although a significant effort has been put forth by volunteers, questions remain as to whether volunteer data are accurate and can be used by regulators. In this study, typical volunteer water quality measurements from laboratory and environmental samples in Iowa were analyzed for error and bias. Volunteer measurements of nitrate+nitrite were significantly lower (about 2-fold) than concentrations determined via standard methods in both laboratory-prepared and environmental samples. Total reactive phosphorus concentrations analyzed by volunteers were similar to measurements determined via standard methods in laboratory-prepared samples and environmental samples, but were statistically lower than the actual concentration in four of the five laboratory-prepared samples. Volunteer water quality measurements were successful in identifying and classifying most of the waters which violate United States Environmental Protection Agency recommended water quality criteria for total nitrogen (66%) and for total phosphorus (52%) with the accuracy improving when accounting for error and biases in the volunteer data. An understanding of the error and bias in volunteer water quality measurements can allow regulators to incorporate volunteer water quality data into total maximum daily load planning or state water quality reporting.

Advances in Phytoremediation: Phytotransformation

Phytoremediation is the use of vegetation for in-situ treatment of contaminated soils, sediments, and water. It is applicable at sites containing organic, nutrient, or metal pollutants that can be accessed by the roots of plants and sequestered, degraded, immobilized, or metabolized in place. In the last few years, a greater understanding has been achieved regarding the uptake and metabolism of organic xenobiotic chemicals by plants, especially chlorinated solvents, petrochemicals, some pesticides, and explosives [1-8]. In addition, inorganics (nutrients, selenium and arsenic) and metals (lead, cadmium, nickel and zinc) have been successfully remediated using plants. These are toxic chemicals that contaminate a large number of hazardous waste sites. This chapter focuses on advancements in phytoremediation, especially with respect to phytotransformation, the enzymatic conversion of organic chemicals within plant tissues following plant uptake.

Sensor data as a measure of native freshwater mussel impact on nitrate formation and food digestion in continuous-flow mesocosms

Abstract Native freshwater mussels can influence the aquatic N cycle, but the mechanisms and magnitude of this effect are not fully understood. We assessed the effects of Amblema plicata and Lampsilis cardium on N transformations over 72 d in 4 continuous-flow mesocosms, with 2 replicates of 2 treatments (mesocosms with and without mussels), equipped with electronic water-chemistry sensors. We compared sensor data to discrete sample data to assess the effect of additional sensor measurements on the ability to detect mussel-related effects on NO3- formation. Analysis of 624 sensor-based data points detected a nearly 6% increase in NO3- concentration in overlying water of mesocosms with mussels relative to mesocosms without mussels (p < 0.05), whereas analysis of 36 discrete samples showed no statistical difference in NO3- between treatments. Mussels also significantly increased NO2- concentrations in the overlying water, but no significant difference in total N was observed. We used the sensor data for phytoplankton-N and NH4+ to infer that digestion times in mussels were 13 ± 6 h. The results suggest that rapid increases in phytoplankton-N levels in the overlying water can lead to decreased lag times between phytoplankton-N and NH4+ maxima. This result indicates that mussels may adjust their digestion rates in response to increased levels of food. The adjustment in digestion time suggests that mussels have a strong response to food availability that can disrupt typical circadian rhythms. Use of sensor data to measure directly and to infer mussel effects on aquatic N transformations at the mesocosm scale could be useful at larger scales in the future.

A preparation technique for analysis of explosives in plant tissues

Abstract A preparation technique for analysis of explosives compounds in plant tissue has been developed. The technique utilizes acetone extraction followed by ultrafiltration clean up. Ultrafiltration removes the large quantities of chlorophyll and polar organics that interfere with UV/Vis detection used in high‐performance liquid chromatography. The procedure was tested with 14 explosives‐related compounds spiked into reed canary grass extracts and method detection limits (MDLs) were calculated. In addition, the overall extraction efficiency for RDX in parrot feather and reed canary grass tissues was determined using 14C labeled compound. Scientists may find the technique useful for plant analysis at waste sites and for phytoremediation research.