Toxic explosives and propellants such as TNT, RDX, and perchlorate that contaminate military testing and training ranges are known to be biodegraded by plants and microbes in the laboratory.  Paradoxically, they are also notoriously persistent under field conditions at ranges, and they can migrate from the source and pose a hazard to humans and ecosystems.  The central hypothesis of this project was that phytoremediation can be used to improve the in situ biodegradation and containment of explosives in contaminated soils at testing and training ranges.

Previous work conducted under SERDP project ER-1317 using radiolabeled explosives demonstrated that RDX is taken up by the plant and translocated to leaves with significant mineralization to carbon dioxide in poplar tissue cultures.  TNT was shown to be taken up, though most of this compound remains in root tissue and is quickly reduced to aminodinitrotoluene and diaminonitrotoluene derivatives.  Gene expression studies have identified potential enzymes involved in metabolism of these compounds in poplar plantlet tissues, and work continues to confirm that these enzymes are active with explosive compounds.  Further work was needed to focus on phytoremediation processes specific to the soil and rhizosphere microbial community to identify other factors affecting the potential for phytoremediation at contaminated sites.

The overall objective of this project was to understand the mechanisms by which toxic energetic compounds, known to be susceptible to biodegradation, were actually detoxified in contaminated subsurface soils at DoD firing ranges by plants native to the site, either by direct uptake and transformation in plant tissues, or by microbial activity in the rhizosphere.  The specific objectives of the research were to determine: (1) whether plants significantly improved biodegradation of explosives using actual soils and plants from representative sites; (2) the respective contribution of plants and soil microbes in the process; and (3) whether the aging of explosives affected the biodegradation process.  Additionally, a field-scale implementation of phytoremediation was performed.  The specific objectives of this field study were to: (1) determine if the implementation of phytoremediation study significantly improved the biodegradation of explosives in soil through a field study; (2) determine whether plants could significantly uptake and degrade explosives in the field; and (3) compare fate and transport processes in laboratory studies using actual soils from the site of the field study with the field demonstration results.

Biodegradation experiments were conducted at the laboratory, greenhouse, and field scale.  The laboratory and greenhouse studies utilized soil materials and plant species from Eglin Air Force Base (Eglin AFB), the site of the field study.  Several biodegradation studies were performed, including the exposure of unplanted soils to TNT, RDX, and HMX and the exposure of planted soils to TNT and RDX.  Weathering and sequestration of chemicals were simulated by aging artificially contaminated soil in the laboratory prior to the start of degradation experiments.  Hybrid poplar and switchgrass was exposed to radiolabeled TNT and RDX and were then examined using phosphor imager autoradiography in order to determine pathway and transport mechanisms.  The microbial communities of native Eglin AFB soils were “finger printed” using T-RFLP.  The properties of each soil type were analyzed and the bioavailability of TNT and RDX was determined by performing sorption studies on these soils.  Gene expression in hybrid poplars exposed to TNT also was investigated.  A field study was conducted consisting of three 0.4 acre plots planted with Paspalum notatum Pensacola (Bahiagrass) with biannual sampling over 18 months.  Both soil and plant samples were taken from each plot, as well as unplanted perimeter soil samples as a control.

The biodegradation studies found that TNT was readily transformed and degraded by microbial communities in native Eglin AFB soils, while RDX and HMX remained recalcitrant under unplanted conditions.  Additional biodegradation studies found that the RDX concentration in soil rapidly decreased in the presence of Bahiagrass and hybrid poplar.  Phosphor imager autoradiography showed the majority of TNT remained in or bonded to the root tissue of hybrid poplar and switchgrass, while RDX was readily transported to the leaf or blade.  Microautoradiographs showed the translocation of RDX or its metabolites into chloroplasts or incorporation into plant structure.  T-RFLP analysis of soils found little diversity in Lakeland soil compared to the much greater diversity found in Dorovan muck.  This was expected given the higher organic and nutrient levels in Dorovan muck.  In the gene expression study, several new genes were demonstrated to be implicated in the detoxification and metabolism of TNT by hybrid poplar.  The results of the field study showed that TNT was transformed in the soil with no apparent treatment benefit in the planted areas and both RDX and HMX migrated downward through the soil before Bahiagrass could effectively treat the compounds.  There was some evidence that the application of high carbon content soil, in which the Bahiagrass was established, slowed the migration of TNT and RDX.

Phytoremediation is a cost-effective and sustainable long-term strategy for management of risk at testing and training ranges.  While phytoremediation was not effective in treating explosives contamination in the sandy soil at Eglin AFB, it is believed that the treatment may be effective in different soils or using different plant species.  The work at Eglin AFB investigated phytoremediation holistically and representatively.  This research attempted to account for all of the many factors influencing phytoremediation at Eglin AFB and used factors that are specific to the site.  The tools and methods developed in this research lay groundwork for creating standards in phytoremediation research.  Sites of interest for phytoremediation should be fully characterized from native soils to native plant species in order to devise a fully integrated approach and predict the success of the treatment technology.  This project provided new insights into the mechanisms underlying phytoremediation of explosives and propellants in the field.