Hyperaccumulation is a unique phenomenon where large amounts of trace elements are stored in leaves, but it only occurs in less than 1% of all plant species. Plants that hyperaccumulate trace elements are studied as a means to remediate polluted substrates, a process known as phytoremediation. The legume genus Astragalus contains the majority of plants that hyperaccumulate Se. My goal was to indentify if and how soil microorganisms may influence Astragalus Se accumulation. I examined the root nodule symbiosis in Astragalus as a tractable interaction to explore the mechanism by which microorganisms mediate changes in plant Se accumulation. I also investigated plant-based Se removal (phytoextraction) through screening different species.

In the first chapter I summarized literature on how plants hyperaccumulate elements through their root systems. Mechanisms related to physical and chemical characteristics of roots are discussed. The microbial assemblage in the rhizosphere is also important in hyperaccumulation. From this basis I explore how soil microorganisms interact in the rhizosphere of Astragalus Se-hyperaccumulators. I tested three hypotheses in the following chapters; (i) soil microorganisms affect Se accumulation in plants, (ii) specifically the root nodule symbiosis has a role in Se-hyperaccumulation, and (iii) the mechanism of root nodule symbiosis affecting hyperaccumulation is through nitrogen allocation into selenoamino acids.

In Chapter 2 I investigated if Se-hyperaccumulators incurred a cost where their symbiotic interactions were disrupted because of their Se tolerance. My experiments did not support evidence of a cost to the symbiosis. I investigated organ [Se] in multiple legume species growing in field conditions, including Se-hyperaccumulators. In general, nodule [Se] were below the threshold used to define Se-hyperaccumulation by leaf concentrations, but they were still notable with some nodules having [Se] near 100 µg Se g^-1 dry weight. I also detected differences in root and nodule [Se] in the hyperaccumulator A. bisulcatus, which may point to a role in Se protecting belowground organs from herbivory. Through x-ray absorption spectra analysis I found that Se was distributed throughout the root nodule in the Se-hyperaccumulators A. bisulcatus, A. praelongus, and A. racemosus . The most abundant form of Se in the nodules was organic, C-Se-C.

Following this I conducted further investigations on root nodules. In Chapter 3 I found an effect of root nodule symbiosis in Se accumulation in A. bisulcatus, where shoot [Se] was positively correlated with shoot [N] and nodulated plants contained higher shoot [Se] than non-nodulated plants. These effects were not evident in non-accumulators. I determined that a mechanism by which root nodule symbiosis could affect A. bisulcatus [Se] was through a 10-fold increase in the selenoamino acid gamma-glutamyl-methylselenocysteine.

In Chapter 4 I studied soil microorganisms in general, without focusing on a specific group, and attempted to identify if soil inoculant source affected Astragalus Se accumulation. I found a significant increase in root (approximately +200%) and shoot (approximately +70%) [Se] in 6 Astragalus species when they were grown in soil inoculant derived from hyperaccumulators compared to inoculant derived from non-accumulators. The 6 species included three Se-hyperaccumulators and three non-accumulators. These results indicate that Se accumulation is mediated by soil microorganisms in some way.

Given the recent explosion of interest in hyperaccumulators as an environmental friendly means of remediating contaminated substrates I investigated Astragalus Se-hyperaccumulators for their ability to remove Se from contaminated biosolids produced in Pueblo, Colorado. I found that the Se-hyperaccumulator A. crotalariae performs better than the annual crop species Brassica juncea, but there is a lack of seed source and agronomic techniques to move forward at a larger scale with the Astragalus. Using the more conventional, fast-growing agronomic species B. juncea and B. napus I found that a dilution of 75% biosolids and 25% sand by volume achieved the highest Se removal potential. Using the information I had gathered in Chapter 4 I attempted to increase plant [Se] in biosolids phytoextraction trials by applying an inoculant derived from soil obtained from the hyperaccumulator A. bisulcatus. This approach did not significantly alter plant [Se] in my 13 species trial.

Finally, in Chapter 6 I synthesize the results of my dissertation into my experience conducting scientific research. I have found that often experimental results that are not consistent with research hypotheses can be the most interesting to pursue because they make you continuously wonder why the outcome occurred. I conclude with potential future outlooks for my work. Although my dissertation has focused much on the root-nodule symbiosis in Astragalus this work has broader implications for ecological theory of mutualisms, the co-evolution of mutualistic partnerships, and the utilization of rhizosphere communities in phytoremediation.