Progress 07/01/06 to 06/30/10
Outputs
OUTPUTS: Persistent organic pollutants (POPs) are intractable contaminants of significant concern because of their resistance to degradation, toxicity, and global distribution. However, currently available remedial options for POP-contaminated soils are limited and expensive. Phytoremediation uses plants to remove contaminants from soil and earlier Hatch-supported work from this laboratory has shown that Cucurbita pepo subspecies pepo (zucchini) may extract large quantities of these pollutants, including DDT/DDE, chlordane, and PCBs. In the final year of this project, the field experiments described in the preceding report were completed. In this multi-year trial, traditional breeding techniques were used to create several hybrid generations of DDE accumulating zucchini and non-accumulating squash cultivars. An analysis of the ability of various parental, F1 hybrids and F1 backcrosses to accumulate DDE will be highly useful in determining the inheritance, expression, and molecular details of the critical proteins involved in C. pepo POP phytoextraction process. In 2007, the parental generation was grown in DDE-contaminated soil and the pollutant accumulation capacity of three zucchini and three squash cultivars was determined and reported earlier. During that growing season, all 18 possible first filial (F1) hybrids were created and the seed was subsequently harvested. Those F1 hybrids were planted in 2008 and the pollutant accumulating abilities were determined. The hybrid zucchini (zucchini pollinated with squash) accumulated significantly less DDE in their tissues than did the parental zucchini generation. Similarly, the hybrid squash (squash pollinated with zucchini) accumulated significantly more DDE in their tissues than did the parental squash generation. The amount of DDE accumulated by the zucchini and squash hybrids was not significantly different. During that growing season, all possible F1 backcrosses (BC) were created; each F1 hybrid was crossed with its original parent. In 2009, the F1 BCs were grown and the analysis of those data was completed during the current reporting period. When backcrossed with the original parent, the nine zucchini F1 BC cultivars did not regain the ability to take up DDE; stem BCFs and percent phytoextraction values were equivalent to the F1 generation and still significantly less than the original parent. However, the nine squash F1 BC cultivars lost much of the DDE uptake capability of the F1 generation; stem BCFs and percent phytoextraction values were intermediate but closer to the parental squash. The inheritance patterns suggest single locus control for persistent organic pollutant (POP) uptake ability in zucchini. PARTICIPANTS: Dr. Jason C. White of the Connecticut Agricultural Experiment Station (CAES)is the primary investigator on this project. Dr.Jason Kelsey of Muhlenberg College (Allentown, PA) has assisted in the ongoing evaluation of parental, F1, and F1 backcross squash and zucchini. Dr. Om Parkash of the University of Massachusetts (Amherst,MA), along with a post-doctoral research associate and a graduate student, conducted the subtraction hybridization work and construction of the cDNA libraries for DDE-exposed and non-exposed squash and zucchini. TARGET AUDIENCES: The target audiences are as follows: environmental engineers, federal and state regulators, farmers, owners of land formerly used for farming, Brownfield site owners, and owners of sites contaminated with persistent organic pollutants. There were several efforts to deliver science based knowledge to a wider audience. First, research findings were published in peer-reviewed scientific journals and web-based publications. Results were also presented at several technical and non-technical meetings. Second,a graduate student at the University of Massachusetts worked on the molecular aspects described in the current reporting period. Third, an undergraduate from the University of New Haven (West Haven, CT) worked on this project part time during the current reporting period. Last, a high school junior (Sacred Heart Academy, Hamden CT) participated in an 80 hour mentorship program as part of this project. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.
Impacts
Information on novel techniques to remediate soils contaminated with weathered persistent organic pollutants is valuable to scientists, engineers, and regulatory agencies. The vast majority of POP-impacted land involves large areas of moderately contaminated soils. Consequently, the most common "remedial" approach involves letting the area remain undisturbed for long periods of time through a process called natural attenuation. The use of zucchini and related plants for POP phytoremediation is still in the research phase. However, work such as that from this project will serve to validate and garner support for this developing technique. In this project, experimental systems ranged from hydroponics to full scale field trials. The use of hydroponics to provide data on the phytotoxicity and uptake of POPs such as DDE is useful as little is known in this area. In addition, the use of these methods to provide treated and untreated tissues is critical to the molecular techniques and analysis being used to elucidate the genetic basis for POP phytoextraction in zucchini. Field studies demonstrating the POP extraction ability of parental and hybrid, as well as hybrid-back crossed, zucchini and squash (non-POP accumulating) cultivars will also be highly instructive in uncovering the genetics of this process. Separate ongoing formal and informal collaborations with colleagues within the Federal government (US EPA, Department of Energy, USDA), as well as with private companies(Edenspace Systems, Inc.; URS Corporation) and Universities (University of Massachusetts, Muhlenberg College) have helped to advance this remedial approach and to contribute to overall project outputs and outcomes.
Publications
- Chhikara, S.; Paulose, B.; White, J.C.; Parkash Dhankher, O. 2010. Understanding the Physiological and Molecular Mechanism of Persistent Organic Pollutant (POP) Uptake and Detoxification in Cucurbit Species (Zucchini and Squash). Environ. Sci. Technol. 44:7295-7301.
- White, J.C. 2010. Inheritance of p,p-DDE Phytoextraction Ability in Hybridized Cucurbita pepo Cultivars. Environ. Sci. Technol.44, 5164-5169.
- Slizovskiy, I.B.; White, J.C.; Kelsey, J.W. 2010. Technical Note: Evaluation of Extraction Methodologies for the Determination of Pesticide Residues in Vegetation. Int. J. Phytorem. 12:820-832.
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Progress 01/01/09 to 12/31/09
Outputs
OUTPUTS: Persistent organic pollutants (POPs) are intractable contaminants of significant concern because of their resistance to degradation, toxicity, and global distribution. However, currently available remedial options for POP-contaminated soils are limited and expensive. Phytoremediation uses plants to remove contaminants from soil and earlier Hatch-supported work has shown that Cucurbita pepo subspecies pepo (zucchini) may extract large quantities of these pollutants, including DDT/DDE, chlordane, and PCBs. Investigations this year continued to focus on two areas. First, as described in a previous report, zucchini and squash were exposed under hydroponic conditions to DDE at 0.12 mg/L for 96 hours. Although these plants are in the same species, they are in different subspecies (pepo vs ovifera) and have been shown to accumulate dramatically different levels of the pollutants when grown in soil. These tissues were frozen in liquid nitrogen and sent to a colleague at the University of Massachusetts to assist in the identification of differentially expressed genes in DDE-treated and untreated plants. Two separate cDNA libraries were created. The first compared across subspecies; here, two separate genes were found to be significantly up-regulated in zucchini exposed to DDE but not in squash. In the shoots, a phloem-loading protein was found; in the roots, the up-regulated gene was a cytochrome p450. The second cDNA library compared only zucchini; in excess of 20 genes were found to be significantly up-regulated in zucchini upon exposure to DDE. This is part of an effort to identify the molecular basis for zucchini's unique ability to accumulate weathered POPs. Second, field experiments were conducted in which traditional breeding techniques were used to create hybrid crosses of six separate zucchini and squash cultivars. In the first season, the parental generation was grown in DDE containing soil and the contaminant content of three zucchini and three squash cultivars was determined and reported earlier. During the growth of the parental generation, hybrids were created for all possible zucchini x squash F1 crosses. The seeds were harvested from the fruit and were used as the F1 generation. The hybrid zucchini (zucchini pollinated with squash) accumulated significantly less DDE in their tissues than did the parental zucchini generation. Similarly, the hybrid squash (squash pollinated with zucchini) accumulated significantly more DDE in their tissues than did the parental squash generation. The amount of DDE accumulated by the zucchini and squash hybrids was not significantly different. Finally, F1 backcrosses are currently being created to further explore the line of research. An analysis of the ability of various F1 hybrids and F1 backcrosses to accumulate DDE will be highly useful in determining the inheritance, expression, and molecular details of the critical proteins involved in C. pepo POP phytoextraction process. These findings are being disseminated to scientists, engineers, and regulators through publication in a peer reviewed journal, as well as by verbal reporting at scientific society meetings and talks to the general public. PARTICIPANTS: Dr. Jason C. White of the Connecticut Agricultural Experiment Station is the primary investigator on this project. Dr.Jason Kelsey of Muhlenberg College (Allentown, PA) has assisted in the ongoing evaluation of parental, F1, and F1 backcross squash and zucchini. Dr. Om Parkash of the University of Massachusetts (Amherst,MA)is conducting the subtraction hybridization work and construction of the cDNA libraries for DDE-exposed and non-exposed squash and zucchini. TARGET AUDIENCES: The target audiences are as follows: environmental engineers, federal and state regulators, farmers, owners of land formerly used for farming, Brownfield site owners, and owners of sites contaminated with persistent organic pollutants. There were several efforts to deliver science based knowledge to a wider audience. First, research findings were published in peer reviewed scientific journals and web-based publications. Second, an undergraduate from Muhlenberg College worked on this project in the laboratory for one week. Last, a high school junior participated in an 80 hour mentorship program as part of this project. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
New information on techniques to remediate soils contaminated with weathered POPs is valuable to scientists, engineers, and regulatory agencies. The vast majority of POP-impacted land consists of vast areas of moderately contaminated soils. Therefore, the most common approach is to simply let the area sit undisturbed through a process called natural attenuation. The use of zucchini and related plants for POP phytoremediation is still in the research phase. However, work such as that from the current year, will serve to validate and garner support for this developing technique. The use of hydroponics to provide data on the phytotoxicity and uptake of POPs such as DDE is useful as little is known in this area. In addition, the use of these methods to provide treated and untreated tissues is critical to the molecular techniques and analysis being used to elucidate the genetic basis for POP phytoextraction in zucchini. Field studies demonstrating the POP extraction ability of parental and hybrid, as well as hybrid-back crossed, zucchini and squash (non-POP accumulating) cultivars will also be highly instructive in uncovering the genetics of this process. Currently, a field test at a 10-acre PCB-contaminated Brownfield site on Long Island is being planned for the summer of 2010. This trial will be run in conjunction with scientists at Brookhaven National Laboratory and the New York State Department of Environmental Conservation. Separate ongoing formal and informal collaborations with colleagues within the Federal government (US EPA, Department of Energy, USDA), as well as with private companies (Edenspace Systems, Inc.; URS Corporation) have helped to advance this remedial approach and to contribute to overall project outputs and outcomes.
Publications
- 1. White, J.C. 2009. Optimizing planting density for DDE phytoextraction by Cucurbita pepo. Environ. Engin. Sci. 26, 369-375.
- 2. White, J.C.; Burken, J.G. 2009. Conference Review: 4th International Phytotechnologies Conference, Denver Colorado September 24-26 2007. Int. J. Phytorem. 11:413-415.
- 3. White, J.C.; Newman, L.A. 2010. Phytoremediation of soils contaminated with organic pollutants. In: Biophysico-chemical Processes in Environmental Systems; International Union of Pure and Applied Chemistry (IUPAC), John Wiley and Sons, Hoboken, NY, USA.
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Progress 01/01/08 to 12/31/08
Outputs
OUTPUTS: Persistent organic pollutants (POPs) are intractable contaminants of significant concern because of their resistance to degradation, toxicity, and global distribution. However, currently available remedial options are limited and expensive. Phytoremediation uses plants to remediate contaminated soil and earlier Hatch-supported work has shown that Cucurbita pepo (zucchini) may extract large quantities of these pollutants, including DDT/DDE and chlordane. Therefore, it is important to determine if C. pepo can be developed into a technique to implement at POP-contaminated sites. Investigations this year focused on two areas. First, batch style-hydroponic studies were conducted to determine the impact of DDE (2-20 mg/L) on the transpiration and biomass of zucchini and squash. These plants, although within the same species, have been shown to differ greatly on DDE uptake ability but the relationship of this parameter to overall species-specific phytotoxicity from the contaminant is not known. Separately, zucchini and squash were exposed to DDE at 0.12 mg/L for 96 hours. This allowed a determination of DDE content in exposed and non-exposed squash and zucchini. In addition, these tissues were frozen in liquid nitrogen and sent to a colleague at the University of Massachusetts to identify differentially expressed genes in DDE treated and untreated plants. This is part of an effort to identify the molecular basis for zucchini's unique ability to accumulate weathered persistent organic pollutants. Second, field experiments were conducted in which traditional breeding techniques were used to create hybrid crosses of six separate zucchini and squash cultivars. As I said, these plants represent two different subspecies of Cucurbita pepo. In the first season, the parental generation was grown in DDE containing soil and the contaminant content of three zucchini and three squash parental cultivars was determined. The DDE content in the roots and stems of zucchini were nearly 4 and 7-fold greater than in the squash, respectively. Zucchini phytoextracted nearly 2% of the DDE; a value 12-times more than that of squash. This DDE extraction value approaches remedial levels achieved with some heavy metals in practical or real-world remediation scenarios. These data agree with previous and published findings for these plants. In addition, during the growth of the parental generation, hybrids were created for all possible zucchini x squash crosses. The hybrid seeds were harvested from the fruit and are being used as the F1 generation. The ability of the various F1 hybrids to accumulate DDE will be highly useful in determining the inheritance, expression, and molecular details of the critical proteins involved in zucchini POP phytoextraction process. In addition, F2 backcrosses are currently being created to further explore the line of research. These findings are being disseminated to scientists, engineers, and regulators through publication in a peer reviewed journal, as well as by verbal reporting at scientific society meetings and talks to the general public. PARTICIPANTS: Jason C. White of the Connecticut Agricultural Experiment Station is the chief investigator on this project. Dr. Jason Kelsey of Muhlenberg College (Allentown, PA) has assisted in the ongoing evaluation of the parental and F1 hybrids of the zucchini and squash. Dr. Om Parkash of the University of Massachussetts (Amherst, MA) is conducting the subtraction mRNA analysis of the DDE-exposed and non-exposed zucchini and squash. TARGET AUDIENCES: The target audiences are as follows: environmental engineers, federal and state regulators, farmers, owners of land formerly used for farming, Brownfield site owners, and owners of sites contaminated with persistent organic pollutants. There were several efforts to deliver science-based knowledge to a wider audience. First, research findings were published in peer reviewed scientific journals and web-based publications. Second, an undergraduate and a graduate student from the University of New Haven worked on this project for five months (part-time) and assisted in field experiments and laboratory analysis. Last, an undergraduate student from Muhlenberg College (Allentown PA) visited my laboratory for one week as an intern and assisted in both field experiments and laboratory analysis. PROJECT MODIFICATIONS: Not relevant to this project.
Impacts
New information on techniques to remediate soils contaminated with weathered POPs is valuable to scientists, engineers, and regulatory agencies. The vast majority of POP-impacted land consists of vast areas of moderately contaminated soils. Therefore, the most common approach is to simply let the area sit undisturbed through a process called natural attenuation. The use of zucchini and related plants for POP phytoremediation is still in the research phase. However, work such as that from the current year will serve to validate and garner support for this developing technique. The use of hydroponics to provide data on the phytotoxicity of POPs such as DDE is useful as little is known in this area. In addition, the use of these methods to provide treated and untreated tissues is critical to the molecular techniques and analysis being used to elucidate the genetic basis for POP phytoextraction in zucchini. Field studies demonstrating the POP extraction ability of parental and hybrid, as well as hybrid-back crossed, zucchini and squash (non-POP accumulating) cultivars will also be highly instructive in uncovering the genetics of this process. Ongoing formal and informal collaborations with colleagues within the Federal government (US EPA, Department of Energy, USDA), as well as with private companies (Edenspace Systems, Inc.; URS Corporation; Applied Phytogenetics) have helped to advance this remedial approach and to contribute to overall project outputs and outomes.
Publications
- White, J.C. 2008. Optimizing planting density for p,p-DDE phytoextraction by Cucurbita pepo. Environ. Engin. Sci. In press.
- Liste, H-H.; White, J.C. 2008. Plant hydraulic lift of soil water: Implications for crop production and land restoration. Plant Soil 313:1-17.
- White, J.C. 2008. Phytoremediation of persistent organic pollutants, in Phytoremediation, [Ed. Lee A. Newman], in Ecyclopedia of Life Support Systems (EOLSS), Developed under the Auspices of the UNESCO, EOLSS Publishers, Oxford, UK, [http://www.eolss.net].
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Progress 01/01/07 to 12/31/07
Outputs
OUTPUTS: Persistent organic pollutants (POPs) are intractable contaminants of significant concern because of their resistance to degradation, toxicity, and global distribution. However, currently available remedial options are limited and expensive. Phytoremediation uses plants to remediate contaminated soil and earlier Hatch-supported work has shown that Cucurbita pepo (zucchini) may extract large quantities of these pollutants, including DDT/DDE and chlordane. Therefore, it is important to determine if C. pepo can be developed in a technique to implement at POP-contaminated sites. Investigations this year focused on two areas. First, hydroponic studies were conducted and the amount of DDE translocated within cucurbit tissues was quantified. This information was then used to develop and validate a mathematic model that can be used predict POP movement within plants. Second, field experiments were conducted to determine the optimum planting density for C. pepo to extract weathered
DDE from soil. A preliminary greenhouse study had shown that in small pots, increasing the number of plants per given volume of soil (0.5 kg) could significantly increase DDE concentrations in the plants. Of the 4 cultivation scenarios investigated with 3 different zucchini cultivars, a density of 16 plants per square meter provided the greatest remedial value. At this density, DDE concentrations in the roots and stems were an order of magnitude above soil levels and 1.4% of the pollutant was extracted in 2.5 months. This value approximates remedial values of heavy metal phytoextraction technologies currently in real-world use. These findings are being disseminated to scientists, engineers, and regulators through publication in peer reviewed journals, as well as by verbal reporting at scientific society meetings and talks to the general public.
PARTICIPANTS: Jason C. White of the Connecticut Agricultural Experiment Station (Department of Soil and Water) is the principal investigator on this project. Dr. Martin Gent of the Connecticut Agricultural Experiment Station (Department of Forestry and Horticulture) conducted the hydroponic experiments and formulated the mathematical model. Dr. Jason Kelsey of Muhlenberg College (Allentown PA) conducted small pot experiments monitoring the uptake of DDE under various cultivation and soil amendment conditions. Dr. Barbara Zeeb of the Royal Military College of Canada (Kingston, Ontario) co-authored a book chapter with J.C. White. Mrs. Terri Arsenault of the Connecticut Agricultural Experiment Station (Department of Analytical Chemisty) provided technical assistance for sample extraction and analysis.
TARGET AUDIENCES: The target audiences are as follows: environmental engineers, federal and state regulators, farmers, owners of land formerly used for farming, brownfield site owners. There were several efforts to deliver science-based knowledge to a wider audience. First, research findings were published in peer reviewed scientific journals. Second, an undergraduate college student (Chemical Engineering major from Worcester Polytechnic Institute) worked for 3 months on this project and assisted in field studies and laboratory analysis. Lastly, findings were also discussed with a reporter from the Haleakala Times (Hawaii).
Impacts
New information on techniques to remediate soils contaminated with weathered POPs is valuable to scientists, engineers, and regulatory agencies. The vast majority of POP-impacted land consists of vast areas of moderately contaminated soils. Therefore, the most common approach is to simply let the area sit undisturbed through a process called natural attenuation. The use of zucchini and related plants for POP phytoremediation is still in the research phase. However, work such as that from the current year will serve to validate and garner support for this developing technique. The use of hydroponics and the development of a predictive and mechanistic mathematical model demonstrate the reliability and uniqueness of the remedial system. Field studies demonstrating that the overall efficacy can be increased by common agronomic and horticultural techniques also demonstrate the value of this approach. Collaborations with colleagues within the Federal government (US EPA,
Department of Energy), as well as with private companies (Edenspace Systems, Inc.; URS Corporation) have helped to advance this remedial approach and to contribute to overall project outputs and outomes.
Publications
- Kelsey, J.W.; Colino, A.; Koberle, M.; White, J.C. 2006. Growth conditions impact DDE accumulation in Cucurbita pepo. Int. J. Phytorem. 8:261-271.
- Gent, M.P.N.; White, J.C.;.Parrish, Z.D.; Isleyen, M.; Eitzer, B.D; Mattina, M.I. 2007. Uptake and translocation of p,p -dichlorodiphenyldichloroethylene supplied in hydroponics solution to cucurbita. Environ. Tox. Chem. 12:2467-2475.
- White, J.C.; Peters, R.P.; Kelsey, J.W. 2007. Surfactants impact p,p -DDE accumulation by plant and earthworm species. Environ. Sci. Technol. 41:2922-2929.
- Gent, M.P.N.; White, J.C.; Eitzer, B.D.; Mattina, M.I. 2007. Modeling the difference among cucurbita in uptake and translocation of p,p -dichlorodiphenyldichloroethylene. Environ. Tox. Chem. 12:2476-2485.
- White, J.C. and Zeeb, B.A. 2007. Plant phylogeny and the remediation of POPs. In: Phytoremediation: Methods in Biotechnology. Ch. 16, Humana Press, Totowa, NJ.
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Progress 01/01/06 to 12/31/06
Outputs
Although this project just started in June of this year, I was able to complete one experiment that applies to this project. Field experiments were conducted to investigate the effect of a rhamnolipid biosurfactant (1000 mg/L) on DDE phytoextraction by Cucurbita subspecies. The surfactant was added as a one liter solution the day before planting. Cucurbita pepo ssp pepo accumulated large amounts of the contaminant from soil, having stem bioconcentration factors, amounts of DDE translocated, and overall percent contaminant phytoextracted that were 14, 9.9, and 5.0 times greater than C. pepo ssp ovifera, respectively. Enhanced contaminant accumulation was observed with both subspecies upon biosurfactant amendment, although treatment reduced ssp ovifera biomass by 60%. The biosurfactant had no effect on the biomass of ssp pepo, increased the average contaminant concentration by 3.6-fold, and doubled the overall amount of DDE removed from soil. Soil amendments such as
surfactants that enhance the mobility of weathered persistent organic pollutants will significantly increase the amount of contaminant phytoextracted by Cucurbita pepo.
Impacts
Persistent organic pollutants are ubiquitous contaminants of soils and sediments yet the remedial options are limited. Earlier Hatch-supported work has shown that although Cucurbita pepo may accumulate large amounts of these contaminants, pollutant release or desorption from the soil is still limiting remedial potential. Results of the experiment described in this report indicate that under certain conditions, a biosurfactant may promote contaminant release, followed by increased uptake and greater/more rapid remediation.
Publications
- White, J.C.; Parrish, Z.D.; Iseleyen, M.; Gent, M.P.N.; Iannucci-Berger, W.; Eitzer, B.D.; Mattina, M.I. 2006. Soil amendments, plant age, and intercropping impact DDE bioavailability to C. pepo. J. Environ. Qual. 35:992-1000.
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