Selenium distribution and speciation in the hyperaccumulator Astragalus bisulcatus and associated ecological partners.

The goal of this study was to investigate how plant selenium (Se) hyperaccumulation may affect ecological interactions and whether associated partners may affect Se hyperaccumulation. The Se hyperaccumulator Astragalus bisulcatus was collected in its natural seleniferous habitat, and x-ray fluorescence mapping and x-ray absorption near-edge structure spectroscopy were used to characterize Se distribution and speciation in all organs as well as in encountered microbial symbionts and herbivores. Se was present at high levels (704-4,661 mg kg(-1) dry weight) in all organs, mainly as organic C-Se-C compounds (i.e. Se bonded to two carbon atoms, e.g. methylselenocysteine). In nodule, root, and stem, up to 34% of Se was found as elemental Se, which was potentially due to microbial activity. In addition to a nitrogen-fixing symbiont, the plants harbored an endophytic fungus that produced elemental Se. Furthermore, two Se-resistant herbivorous moths were discovered on A. bisulcatus, one of which was parasitized by a wasp. Adult moths, larvae, and wasps all accumulated predominantly C-Se-C compounds. In conclusion, hyperaccumulators live in association with a variety of Se-resistant ecological partners. Among these partners, microbial endosymbionts may affect Se speciation in hyperaccumulators. Hyperaccumulators have been shown earlier to negatively affect Se-sensitive ecological partners while apparently offering a niche for Se-resistant partners. Through their positive and negative effects on different ecological partners, hyperaccumulators may influence species composition and Se cycling in seleniferous ecosystems.

Effects of selenium hyperaccumulation on plant-plant interactions: evidence for elemental allelopathy?

• Few studies have investigated plant-plant interactions involving hyperaccumulator plants. Here, we investigated the effect of selenium (Se) hyperaccumulation on neighboring plants. • Soil and litter Se concentrations were determined around the hyperaccumulators Astragalus bisulcatus and Stanleya pinnata and around the nonhyperaccumulators Medicago sativa and Helianthus pumilus. We also compared surrounding vegetative cover, species composition and Se concentration in two plant species (Artemisia ludoviciana and Symphyotrichum ericoides) growing either close to or far from Se hyperaccumulators. Then, Arabidopsis thaliana germination and growth were compared on soils collected next to the hyperaccumulators and the nonhyperaccumulators. • Soil collected around hyperaccumulators contained more Se (up to 266 mg Se kg(-1) ) than soil collected around nonhyperaccumulators. Vegetative ground cover was 10% lower around Se hyperaccumulators compared with nonhyperaccumulators. The Se concentration was higher in neighboring species A. ludoviciana and S. ericoides when growing close to, compared with far from, Se hyperaccumulators. A. thaliana showed reduced germination and growth, and higher Se accumulation, when grown on soil collected around Se hyperaccumulators compared with soil collected around nonaccumulators. • In conclusion, Se hyperaccumulators may increase the surrounding soil Se concentration (phytoenrichment). The enhanced soil Se contents around hyperaccumulators can impair the growth of Se-sensitive plant species, pointing to a possible role of Se hyperaccumulation in elemental allelopathy.

Selenium accumulation in flowers and its effects on pollination.

• Selenium (Se) hyperaccumulation has a profound effect on plant-arthropod interactions. Here, we investigated floral Se distribution and speciation in flowers and the effects of floral Se on pollen quality and plant-pollinator interactions. • Floral Se distribution and speciation were compared in Stanleya pinnata, an Se hyperaccumulator, and Brassica juncea, a comparable nonhyperaccumulator. Pollen germination was measured from plants grown with varying concentrations of Se and floral visitation was compared between plants with high and low Se. • Stanleya pinnata preferentially allocated Se to flowers, as nontoxic methyl-selenocysteine (MeSeCys). Brassica juncea had higher Se concentrations in leaves than flowers, and a lower fraction of MeSeCys. For B. juncea, high floral Se concentration impaired pollen germination; in S. pinnata Se had no effect on pollen germination. Floral visitors collected from Se-rich S. pinnata contained up to 270 µg g(-1), concentrations toxic to many herbivores. Indeed, floral visitors showed no visitation preference between high- and low-Se plants. Honey from seleniferous areas contained 0.4-1 µg Se g(-1), concentrations that could provide human health benefits. • This study is the first to shed light on the possible evolutionary cost, through decreased pollen germination in B. juncea, of Se accumulation and has implications for the management of seleniferous areas.

Molecular mechanisms of selenium tolerance and hyperaccumulation in Stanleya pinnata.

The molecular mechanisms responsible for selenium (Se) tolerance and hyperaccumulation were studied in the Se hyperaccumulator Stanleya pinnata (Brassicaceae) by comparing it with the related secondary Se accumulator Stanleya albescens using a combination of physiological, structural, genomic, and biochemical approaches. S. pinnata accumulated 3.6-fold more Se and was tolerant to 20 microm selenate, while S. albescens suffered reduced growth, chlorosis and necrosis, impaired photosynthesis, and high levels of reactive oxygen species. Levels of ascorbic acid, glutathione, total sulfur, and nonprotein thiols were higher in S. pinnata, suggesting that Se tolerance may in part be due to increased antioxidants and up-regulated sulfur assimilation. S. pinnata had higher selenocysteine methyltransferase protein levels and, judged from liquid chromatography-mass spectrometry, mainly accumulated the free amino acid methylselenocysteine, while S. albescens accumulated mainly the free amino acid selenocystathionine. S. albescens leaf x-ray absorption near-edge structure scans mainly detected a carbon-Se-carbon compound (presumably selenocystathionine) in addition to some selenocysteine and selenate. Thus, S. albescens may accumulate more toxic forms of Se in its leaves than S. pinnata. The species also showed different leaf Se sequestration patterns: while S. albescens showed a diffuse pattern, S. pinnata sequestered Se in localized epidermal cell clusters along leaf margins and tips, concentrated inside of epidermal cells. Transcript analyses of S. pinnata showed a constitutively higher expression of genes involved in sulfur assimilation, antioxidant activities, defense, and response to (methyl)jasmonic acid, salicylic acid, or ethylene. The levels of some of these hormones were constitutively elevated in S. pinnata compared with S. albescens, and leaf Se accumulation was slightly enhanced in both species when these hormones were supplied. Thus, defense-related phytohormones may play an important signaling role in the Se hyperaccumulation of S. pinnata, perhaps by constitutively up-regulating sulfur/Se assimilation followed by methylation of selenocysteine and the targeted sequestration of methylselenocysteine.

Effects of selenium accumulation on reproductive functions in Brassica juncea and Stanleya pinnata.

Selenium (Se) is an essential micronutrient for many organisms, but is also a toxin and environmental pollutant at elevated levels. Due to its chemical similarity to sulphur, most plants readily take up and assimilate Se. Se accumulators such as Brassica juncea can accumulate Se between 0.01% and 0.1% of dry weight (DW), and Se hyperaccumulators such as Stanleya pinnata (Brassicaeae) contain between 0.1% and 1.5% DW of Se. While Se accumulation offers the plant a variety of ecological benefits, particularly protection from herbivory, its potential costs are still unexplored. This study examines the effects of plant Se levels on reproductive functions. In B. juncea, Se concentrations >0.05-0.1% caused decreases in biomass, pollen germination, individual seed and total seed weight, number of seeds produced, and seed germination. In S. pinnata there was no negative effect of increased Se concentration on pollen germination. In cross-pollination of B. juncea plants with different Se levels, both the maternal and paternal Se level affected reproduction, but the maternal Se concentration had the most pronounced effect. Interestingly, high-Se maternal plants were most efficiently pollinated by Se-treated paternal plants. These data provide novel insights into the potential reproductive costs of Se accumulation, interactive effects of Se in pollen grains and in the pistil, and the apparent evolution of physiological tolerance mechanisms in hyperaccumulators to avoid reproductive repercussions.

Selenium hyperaccumulation offers protection from cell disruptor herbivores.

BACKGROUND: Hyperaccumulation, the rare capacity of certain plant species to accumulate toxic trace elements to levels several orders of magnitude higher than other species growing on the same site, is thought to be an elemental defense mechanism against herbivores and pathogens. Previous research has shown that selenium (Se) hyperaccumulation protects plants from a variety of herbivores and pathogens. Selenium hyperaccumulating plants sequester Se in discrete locations in the leaf periphery, making them potentially more susceptible to some herbivore feeding modes than others. In this study we investigate the protective function of Se in the Se hyperaccumulators Stanleya pinnata and Astragalus bisulcatus against two cell disrupting herbivores, the western flower thrips (Frankliniella occidentalis) and the two-spotted spider mite (Tetranychus urticae). RESULTS: Astragalus bisulcatus and S. pinnata with high Se concentrations (greater than 650 mg Se kg(-1)) were less subject to thrips herbivory than plants with low Se levels (less than 150 mg Se kg(-1)). Furthermore, in plants containing elevated Se levels, leaves with higher concentrations of Se suffered less herbivory than leaves with less Se. Spider mites also preferred to feed on low-Se A. bisulcatus and S. pinnata plants rather than high-Se plants. Spider mite populations on A. bisulcatus decreased after plants were given a higher concentration of Se. Interestingly, spider mites could colonize A. bisulcatus plants containing up to 200 mg Se kg(-1) dry weight, concentrations which are toxic to many other herbivores. Selenium distribution and speciation studies using micro-focused X-ray fluorescence (µXRF) mapping and Se K-edge X-ray absorption spectroscopy revealed that the spider mites accumulated primarily methylselenocysteine, the relatively non-toxic form of Se that is also the predominant form of Se in hyperaccumulators. CONCLUSIONS: This is the first reported study investigating the protective effect of hyperaccumulated Se against cell-disrupting herbivores. The finding that Se protected the two hyperaccumulator species from both cell disruptors lends further support to the elemental defense hypothesis and increases the number of herbivores and feeding modes against which Se has shown a protective effect. Because western flower thrips and two-spotted spider mites are widespread and economically important herbivores, the results from this study also have potential applications in agriculture or horticulture, and implications for the management of Se-rich crops.

Selenium-tolerant diamondback moth disarms hyperaccumulator plant defense.

BACKGROUND: Some plants hyperaccumulate the toxic element selenium (Se) to extreme levels, up to 1% of dry weight. The function of this intriguing phenomenon is obscure. RESULTS: Here, we show that the Se in the hyperaccumulator prince's plume (Stanleya pinnata) protects it from caterpillar herbivory because of deterrence and toxicity. In its natural habitat, however, a newly discovered variety of the invasive diamondback moth (Plutella xylostella) has disarmed this elemental defense. It thrives on plants containing highly toxic Se levels and shows no oviposition or feeding deterrence, in contrast to related varieties. Interestingly, a Se-tolerant wasp (Diadegma insulare) was found to parasitize the tolerant moth. The insect's Se tolerance mechanism was revealed by X-ray absorption spectroscopy and liquid chromatography-mass spectroscopy, which showed that the Se-tolerant moth and its parasite both accumulate methylselenocysteine, the same form found in the hyperaccumulator plant, whereas related sensitive moths accumulate selenocysteine. The latter is toxic because of its nonspecific incorporation into proteins. Indeed, the Se-tolerant diamondback moth incorporated less Se into protein. Additionally, the tolerant variety sequestered Se in distinct abdominal areas, potentially involved in detoxification and larval defense to predators. CONCLUSIONS: Although Se hyperaccumulation protects plants from herbivory by some invertebrates, it can give rise to the evolution of unique Se-tolerant herbivores and thus provide a portal for Se into the local ecosystem. In a broader context, this study provides insight into the possible ecological implications of using Se-enriched crops as a source of anti-carcinogenic selenocompounds and for the remediation of Se-polluted environments.

Selenium hyperaccumulator plants Stanleya pinnata and Astragalus bisulcatus are colonized by Se-resistant, Se-excluding wasp and beetle seed herbivores.

Selenium (Se) hyperaccumulator plants can concentrate the toxic element Se up to 1% of shoot (DW) which is known to protect hyperaccumulator plants from generalist herbivores. There is evidence for Se-resistant insect herbivores capable of feeding upon hyperaccumulators. In this study, resistance to Se was investigated in seed chalcids and seed beetles found consuming seeds inside pods of Se-hyperaccumulator species Astragalus bisulcatus and Stanleya pinnata. Selenium accumulation, localization and speciation were determined in seeds collected from hyperaccumulators in a seleniferous habitat and in seed herbivores. Astragalus bisulcatus seeds were consumed by seed beetle larvae (Acanthoscelides fraterculus Horn, Coleoptera: Bruchidae) and seed chalcid larvae (Bruchophagus mexicanus, Hymenoptera: Eurytomidae). Stanleya pinnata seeds were consumed by an unidentified seed chalcid larva. Micro X-ray absorption near-edge structure (µXANES) and micro-X-Ray Fluorescence mapping (µXRF) demonstrated Se was mostly organic C-Se-C forms in seeds of both hyperaccumulators, and S. pinnata seeds contained ∼24% elemental Se. Liquid chromatography-mass spectrometry of Se-compounds in S. pinnata seeds detected the C-Se-C compound seleno-cystathionine while previous studies of A. bisulcatus seeds detected the C-Se-C compounds methyl-selenocysteine and γ-glutamyl-methyl-selenocysteine. Micro-XRF and µXANES revealed Se ingested from hyperaccumulator seeds redistributed throughout seed herbivore tissues, and portions of seed C-Se-C were biotransformed into selenocysteine, selenocystine, selenodiglutathione, selenate and selenite. Astragalus bisulcatus seeds contained on average 5,750 µg Se g(-1), however adult beetles and adult chalcid wasps emerging from A. bisulcatus seed pods contained 4-6 µg Se g(-1). Stanleya pinnata seeds contained 1,329 µg Se g(-1) on average; however chalcid wasp larvae and adults emerging from S. pinnata seed pods contained 9 and 47 µg Se g(-1). The results suggest Se resistant seed herbivores exclude Se, greatly reducing tissue accumulation; this explains their ability to consume high-Se seeds without suffering toxicity, allowing them to occupy the unique niche offered by Se hyperaccumulator plants.

Selenium hyperaccumulators facilitate selenium-tolerant neighbors via phytoenrichment and reduced herbivory.

BACKGROUND: Soil surrounding selenium (Se) hyperaccumulator plants was shown earlier to be enriched in Se, impairing the growth of Se-sensitive plant species. Because Se levels in neighbors of hyperaccumulators were higher and Se has been shown to protect plants from herbivory, we investigate here the potential facilitating effect of Se hyperaccumulators on Se-tolerant neighboring species in the field. RESULTS: We measured growth and herbivory of Artemisia ludoviciana and Symphyotrichum ericoides as a function of their Se concentration and proximity to hyperaccumulators Astragalus bisulcatus and Stanleya pinnata. When growing next to hyperaccumulators, A. ludoviciana and S. ericoides contained 10- to 20-fold higher Se levels (800-2,000 mg kg(-1) DW) than when growing next to nonaccumulators. The roots of both species were predominantly (70%-90%) directed toward hyperaccumulator neighbors, not toward other neighbors. Moreover, neighbors of hyperaccumulators were 2-fold bigger, showed 2-fold less herbivory damage, and harbored 3- to 4-fold fewer arthropods. When used in laboratory choice and nonchoice grasshopper herbivory experiments, Se-rich neighbors of hyperaccumulators experienced less herbivory and caused higher grasshopper Se accumulation (10-fold) and mortality (4-fold). CONCLUSIONS: Enhanced soil Se levels around hyperaccumulators can facilitate growth of Se-tolerant plant species through reduced herbivory and enhanced growth. This study is the first to show facilitation via enrichment with a nonessential element. It is interesting that Se enrichment of hyperaccumulator neighbors may affect competition in two ways, by reducing growth of Se-sensitive neighbors while facilitating Se-tolerant neighbors. Via these competitive and facilitating effects, Se hyperaccumulators may affect plant community composition and, consequently, higher trophic levels.

Selenium accumulation in plants--phytotechnological applications and ecological implications.

Selenium (Se) is an essential trace element for many organisms including humans, yet toxic at higher levels. Both Se deficiency and toxicity are problems worldwide. Since plants readily accumulate and volatilize Se, they may be used both as a source of dietary Se and for removing excess Se from the environment. Plant species differ in their capacity to metabolize and accumulate Se, from non-Se accumulators (< 100 mg Se/kg DW), to Se-accumulators (100-1000 mg Se/kg DW) to Se hyperaccumulators (> 1,000 mg Se/kg DW). Here we review plant mechanisms of Se metabolism in these various plant types. We also summarize results from genetic engineering that have led to enhanced plant Se accumulation, volatilization, and/or tolerance, including field studies. Before using Se-accumulating plants at a large scale we need to evaluate the ecological implications. Research so far indicates that plant Se accumulation significantly affects the plant's ecological interactions below and above ground. Selenium canprotect plants from fungal pathogens and from a variety of invertebrate and vertebrate herbivores, due to both deterrence and toxicity. However, specialist (Se-tolerant herbivores), detritivores and endophytes appear to utilize Se hyperaccumulator plants as a resource. These findings are relevant for managing phytoremediation of Se and similar elements.

Physiological functions of beneficial elements.

Aluminum (Al), cobalt (Co), sodium (Na), selenium (Se), and silicon (Si) are considered beneficial elements for plants: they are not required by all plants but can promote plant growth and may be essential for particular taxa. These beneficial elements have been reported to enhance resistance to biotic stresses such as pathogens and herbivory, and to abiotic stresses such as drought, salinity, and nutrient toxicity or deficiency. The beneficial effects of low doses of Al, Co, Na and Se have received little attention compared to toxic effects that typically occur at higher concentrations. Better understanding of the effects of beneficial elements is important to improve crop productivity and enhance plant nutritional value for a growing world population.

Selenium protects the hyperaccumulator Stanleya pinnata against black-tailed prairie dog herbivory in native seleniferous habitats.

Elemental hyperaccumulation in plants is hypothesized to represent a plant defense mechanism. The objective of this study was to determine whether selenium (Se) hyperaccumulation offers plants long-term protection from the black-tailed prairie dog (Cynomys ludovicianus). Prairie dogs are a keystone species. The hyperaccumulator Stanleya pinnata (prince's plume) co-occurs with prairie dogs in seleniferous areas in the western United States. Stanleya pinnata plants pretreated with high or low Se concentrations were planted on two prairie dog towns with different levels of herbivory pressure, and herbivory of these plants was monitored over 2 years. Throughout this study, plants with elevated Se levels suffered less herbivory and survived better than plants with low leaf Se concentrations. This study indicates that the Se in hyperaccumulator S. pinnata protects the plant in its natural habitat from herbivory by the black-tailed prairie dog. The results from this study support the hypothesis that herbivory by prairie dogs or similar small mammals has been a contributing selection pressure for the evolution of plant Se hyperaccumulation in North America. This study is the first to test the ecological significance of hyperaccumulation over a long period in a hyperaccumulator's natural habitat.

The role of selenium in protecting plants against prairie dog herbivory: implications for the evolution of selenium hyperaccumulation.

Some plants can hyperaccumulate the element selenium (Se) up to 10,000 mg Se kg(-1) dry weight. Hyperaccumulation has been hypothesized to defend against herbivory. In laboratory studies high Se levels protect plants from invertebrate herbivores and pathogens. However, field studies and mammalian herbivore studies that link Se accumulation to herbivory protection are lacking. In this study a combination of field surveys and manipulative field studies were carried out to determine whether plant Se accumulation in the field deters herbivory by black-tailed prairie dogs (Cynomys ludovicianus). The Se hyperaccumulator Astragalus bisulcatus (two-grooved milkvetch) occurs naturally on seleniferous soils in the Western USA, often on prairie dog colonies. Field surveys have shown that this Se hyperaccumulator is relatively abundant on some prairie dog colonies and suffers less herbivory than other forb species. This protection was likely owing to Se accumulation, as judged from subsequent manipulative field experiments. When given a choice between pairs of plants of the Se hyperaccumulator Stanleya pinnata (prince's plume) that were pretreated with or without Se, prairie dogs preferred to feed on the plants with low Se; the same results were obtained for the non-hyperaccumulator Brassica juncea (Indian mustard). Plants containing as little as 38 mg Se kg(-1) DW were protected from herbivory. Taken together these results shed light on the functional significance of Se hyperaccumulation and the possible selection pressures driving its evolution. They also have implications for the use of plants in Se phytoremediation, or as Se-fortified crops.

Selenium hyperaccumulation reduces plant arthropod loads in the field.

The elemental defense hypothesis proposes that some plants hyperaccumulate toxic elements as a defense mechanism. In this study the effectiveness of selenium (Se) as an arthropod deterrent was investigated under field conditions. Arthropod loads were measured over two growing seasons in Se hyperaccumulator habitats in Colorado, USA, comparing Se hyperaccumulator species (Astragalus bisulcatus and Stanleya pinnata) with nonhyperaccumulators (Camelina microcarpa, Astragalus americanus, Descurainia pinnata, Medicago sativa, and Helianthus pumilus). The Se hyperaccumulating plant species, which contained 1000-14 000 microg Se g(-1) DW, harbored significantly fewer arthropods (c. twofold) and fewer arthropod species (c. 1.5-fold) compared with nonhyperaccumulator species that contained < 30 microg Se g(-1) DW. Arthropods collected on Se-hyperaccumulating plants contained three- to 10-fold higher Se concentrations than those found on nonhyperaccumulating species, but > 10-fold lower Se concentrations than their hyperaccumulator hosts. Several arthropod species contained > 100 microg Se g(-1) DW, indicating Se tolerance and perhaps feeding specialization. These results support the elemental defense hypothesis and suggest that invertebrate herbivory may have contributed to the evolution of Se hyperaccumulation.

Selenium accumulation protects plants from herbivory by Orthoptera via toxicity and deterrence.

To investigate whether selenium (Se) accumulation in plants provides a chemical defense against generalist insect herbivores, the feeding preference and performance of a mix of orthopteran species were investigated. The selenium hyperaccumulator Stanleya pinnata and accumulator Brassica juncea were used in herbivory studies in the laboratory, and S. pinnata was also used in a manipulative field experiment. In laboratory studies, both crickets and grasshoppers avoided plants pretreated with selenate, while those given no choice died after eating leaves with elevated Se (447 +/- 68 and 230 +/- 68 microg Se g(-1) DW, respectively). B. juncea has previously been shown to accumulate selenate, while S. pinnata hyperaccumulates methyl-selenocysteine. Thus, these findings demonstrate that both inorganic and organic forms of selenium protect plants from herbivory. Grasshoppers fed S. pinnata contained methylselenocysteine in their midgut and absorbed this form into surrounding tissues. In a manipulative field experiment, methylselenocysteine protected S. pinnata from invertebrate herbivory and increased its long-term survival rate over an entire growth season. * In native habitats of selenium hyperaccumulators, orthopterans represent a major group of insect herbivores. Protection offered by organic selenium accumulation against these herbivores may have promoted the evolution of selenium hyperaccumulation in plants.