Aquatic insect accumulation of uranium at spring outflows in the Grand Canyon region as influenced by aqueous and sediment geochemistry and biological factors: implications for monitoring.

Potential adverse ecological effects of expanded uranium (U) mining within the Grand Canyon region motivated studies to better understand U exposure and risk to endemic species. This study documents U exposures and analyzes geochemical and biological factors affecting U bioaccumulation at spring-fed systems within the Grand Canyon region. The principal objective was to determine if aqueous U was broadly indicative of U accumulated by insect larvae, a dominate fauna. Analyses focused on three widely distributed taxa: Argia sp. (a predatory damselfly), Culicidae (suspension feeding mosquitos), and Limnephilus sp. (a detritivorous caddisfly). The study showed that U accumulated by aquatic insects (and periphyton) generally correlated positively with total dissolved U, although correlations were strongest when based on modeled concentrations of the U-dicarbonato complex, UO2(CO3)2-2, and UO2(OH)2. Sediment metal concentration was a redundant indicator of U bioaccumulation. Neither insect size or U in the gut content of Limnephilus sp. substantially affected correlations between aqueous U and whole-body U concentrations. However, in Limnephilus sp., the gut and its content contained large quantities of U. Estimates of the sediment burden in the gut indicated that sediment was a minor source of U mass but contributed substantially to the total insect weight. As a result, whole-body U concentration would tend to vary inversely with the sediment burden of the gut. The correlations between aqueous U and bioaccumulated U provide an initial relational baseline against which newly acquired data could be evaluated for changes in U exposure during and after mining operations.

Uranium Bioaccumulation Dynamics in the Mayfly Neocloeon triangulifer and Application to Site-Specific Prediction.

Little is known about the underlying mechanisms governing the bioaccumulation of uranium (U) in aquatic insects. We experimentally parameterized conditional rate constants for aqueous U uptake, dietary U uptake, and U elimination for the aquatic baetid mayfly Neocloeon triangulifer. Results showed that this species accumulates U from both the surrounding water and diet, with waterborne uptake prevailing. Elevated dietary U concentrations decreased feeding rates, presumably by altering food palatability or impairing the mayfly's digestive processes, or both. Nearly 90% of the accumulated U was eliminated within 24 h after the waterborne exposure ceased, reflecting the desorption of weakly bound U from the insect's integument. To examine whether the experimentally derived rate constants for N. triangulifer could be generalized to baetid mayflies, mayfly U concentrations were predicted using the water chemistry and U measured in periphyton from springs in Grand Canyon (United States) and were compared to U concentrations in spring-dwelling mayflies. Predicted and observed mayfly U concentrations were in good agreement. Under the modeled site-specific conditions, waterborne U uptake accounted for 52-93% of the bioaccumulated U. U accumulation was limited in these wild populations due to a combination of factors including low concentrations of bioavailable dissolved U species, slow U uptake rates from food, and fast U elimination.

Assessing the Dietary Bioavailability of Metals Associated with Natural Particles: Extending the Use of the Reverse Labeling Approach to Zinc.

We extend the use of a novel tracing technique to quantify the bioavailability of zinc (Zn) associated with natural particles using snails enriched with a less common Zn stable isotope. Lymnaea stagnalis is a model species that has relatively fast Zn uptake rates from the dissolved phase, enabling their rapid enrichment in 67Zn during the initial phase of labeling. Isotopically enriched snails were subsequently exposed to algae mixed with increasing amounts of metal-rich particles collected from two acid mine drainage impacted rivers. Zinc bioavailability from the natural particles was inferred from calculations of 66Zn assimilation into the snail's soft tissues. Zinc assimilation efficiency (AE) varied from 28% for the Animas River particles to 45% for the Snake River particles, indicating that particle-bound, or sorbed Zn, was bioavailable from acid mine drainage wastes. The relative binding strength of Zn sorption to the natural particles was inversely related to Zn bioavailability; a finding that would not have been possible without using the reverse labeling approach. Differences in the chemical composition of the particles suggest that their geochemical properties may influence the extent of Zn bioavailability.

Biogeochemical Controls of Uranium Bioavailability from the Dissolved Phase in Natural Freshwaters.

To gain insights into the risks associated with uranium (U) mining and processing, we investigated the biogeochemical controls of U bioavailability in the model freshwater species Lymnaea stagnalis (Gastropoda). Bioavailability of dissolved U(VI) was characterized in controlled laboratory experiments over a range of water hardness, pH, and in the presence of complexing ligands in the form of dissolved natural organic matter (DOM). Results show that dissolved U is bioavailable under all the geochemical conditions tested. Uranium uptake rates follow first order kinetics over a range encompassing most environmental concentrations. Uranium uptake rates in L. stagnalis ultimately demonstrate saturation uptake kinetics when exposure concentrations exceed 100 nM, suggesting uptake via a finite number of carriers or ion channels. The lack of a relationship between U uptake rate constants and Ca uptake rates suggest that U does not exclusively use Ca membrane transporters. In general, U bioavailability decreases with increasing pH, increasing Ca and Mg concentrations, and when DOM is present. Competing ions did not affect U uptake rates. Speciation modeling that includes formation constants for U ternary complexes reveals that the aqueous concentration of dicarbonato U species (UO2(CO3)2(-2)) best predicts U bioavailability to L. stagnalis, challenging the free-ion activity model postulate.

Dietary Uptake of Cu Sorbed to Hydrous Iron Oxide is Linked to Cellular Toxicity and Feeding Inhibition in a Benthic Grazer.

Whereas feeding inhibition caused by exposure to contaminants has been extensively documented, the underlying mechanism(s) are less well understood. For this study, the behavior of several key feeding processes, including ingestion rate and assimilation efficiency, that affect the dietary uptake of Cu were evaluated in the benthic grazer Lymnaea stagnalis following 4-5 h exposures to Cu adsorbed to synthetic hydrous ferric oxide (Cu-HFO). The particles were mixed with a cultured alga to create algal mats with Cu exposures spanning nearly 3 orders of magnitude at variable or constant Fe concentrations, thereby allowing first order and interactive effects of Cu and Fe to be evaluated. Results showed that Cu influx rates and ingestion rates decreased as Cu exposures of the algal mat mixture exceeded 10(4) nmol/g. Ingestion rate appeared to exert primary control on the Cu influx rate. Lysosomal destabilization rates increased directly with Cu influx rates. At the highest Cu exposure where the incidence of lysosomal membrane damage was greatest (51%), the ingestion rate was suppressed 80%. The findings suggested that feeding inhibition was a stress response emanating from excessive uptake of dietary Cu and cellular toxicity.

Novel and nontraditional use of stable isotope tracers to study metal bioavailability from natural particles.

We devised a novel tracing approach that involves enriching test organisms with a stable metal isotope of low natural abundance prior to characterizing metal bioavailability from natural inorganic particles. In addition to circumventing uncertainties associated with labeling natural particles and distinguishing background metals, the proposed "reverse labeling" technique overcomes many drawbacks inherent to using radioisotope tracers. Specifically, we chronically exposed freshwater snails ( Lymnaea stagnalis ) to synthetic water spiked with Cu that was 99.4% (65)Cu to increase the relative abundance of (65)Cu in the snail's tissues from ~32% to >80%. The isotopically enriched snails were then exposed to benthic algae mixed with Cu-bearing Fe-Al particles collected from the Animas River (Colorado), an acid mine drainage impacted river. We used (63)Cu to trace Cu uptake from the natural particles and inferred their bioavailability from calculation of Cu assimilation into tissues. Cu assimilation from these particles was 44%, indicating that 44% of the particulate Cu was absorbed by the invertebrate. This demonstrates that inorganic particulate Cu can be bioavailable. The reverse labeling approach shows great potential in various scientific areas such as environmental contamination and nutrition for addressing questions involving uptake of an element that naturally has multiple isotopes.

Dietary bioavailability of Cu adsorbed to colloidal hydrous ferric oxide.

The dietary bioavailability of copper (Cu) adsorbed to synthetic colloidal hydrous ferric oxide (HFO) was evaluated from the assimilation of (65)Cu by two benthic grazers, a gastropod and a larval mayfly. HFO was synthesized, labeled with (65)Cu to achieve a Cu/Fe ratio comparable to that determined in naturally formed HFO, and then aged. The labeled colloids were mixed with a food source (the diatom Nitzschia palea) to yield dietary (65)Cu concentrations ranging from 211 to 2204 nmol/g (dry weight). Animals were pulse fed the contaminated diet and assimilation of (65)Cu from HFO was determined following 1-3 days of depuration. Mass transfer of (65)Cu from HFO to the diatom was less than 1%, indicating that HFO was the source of (65)Cu to the grazers. Estimates of assimilation efficiency indicated that the majority of Cu ingested as HFO was assimilated (values >70%), implying that colloidal HFO potentially represents a source of dietary Cu to benthic grazers, especially where there is active formation and infiltration of these particles into benthic substrates.

Calibrating biomonitors to ecological disturbance: a new technique for explaining metal effects in natural waters.

Bioaccumulated toxic metals in tolerant biomonitors are indicators of metal bioavailability and can be calibrated against metal-specific responses in sensitive species, thus creating a tool for defining dose-response for metals in a field setting. Dose-response curves that define metal toxicity in natural waters are rare. Demonstrating cause and effect under field conditions and integrated chemical measures of metal bioavailability from food and water is problematic. The total bioaccumulated metal concentration in any organism that is a net accumulator of the metal is informative about metal bioavailability summed across exposure routes. However, there is typically no one universal metal concentration that is indicative of toxicity, especially across species, largely because of interspecies differences in detoxification. Stressed organisms are also only present across a narrow range in the dose-response curve, limiting the use of singles species as both biomonitors and bioindicator of stress. Herein we show, in 3 field settings, that bioaccumulated Cu concentrations in a metal-tolerant, riverine biomonitor (species of the caddisfly genus Hydropsyche spp.) can be calibrated against metal-specific ecological responses across very wide ranges of contamination. Using the calibrated dose-response, we show that reduced abundance of species and individuals from particularly sensitive mayfly families (heptageniid mayflies) is more than 2-fold more sensitive to bioavailable Cu than other traditional measures of stress like EPT or total number of benthic macroinvertebrate species. We propose that this field dose-response curve be tested more widely for general application, and that calibrations against other stress responses be developed for biomonitors from lakes, estuaries, and coastal marine ecosystems.

Aquatic insect ecophysiological traits reveal phylogenetically based differences in dissolved cadmium susceptibility.

We used a phylogenetically based comparative approach to evaluate the potential for physiological studies to reveal patterns of diversity in traits related to susceptibility to an environmental stressor, the trace metal cadmium (Cd). Physiological traits related to Cd bioaccumulation, compartmentalization, and ultimately susceptibility were measured in 21 aquatic insect species representing the orders Ephemeroptera, Plecoptera, and Trichoptera. We mapped these experimentally derived physiological traits onto a phylogeny and quantified the tendency for related species to be similar (phylogenetic signal). All traits related to Cd bioaccumulation and susceptibility exhibited statistically significant phylogenetic signal, although the signal strength varied among traits. Conventional and phylogenetically based regression models were compared, revealing great variability within orders but consistent, strong differences among insect families. Uptake and elimination rate constants were positively correlated among species, but only when effects of body size and phylogeny were incorporated in the analysis. Together, uptake and elimination rates predicted dramatic Cd bioaccumulation differences among species that agreed with field-based measurements. We discovered a potential tradeoff between the ability to eliminate Cd and the ability to detoxify it across species, particularly mayflies. The best-fit regression models were driven by phylogenetic parameters (especially differences among families) rather than functional traits, suggesting that it may eventually be possible to predict a taxon's physiological performance based on its phylogenetic position, provided adequate physiological information is available for close relatives. There appears to be great potential for evolutionary physiological approaches to augment our understanding of insect responses to environmental stressors in nature.

Cadmium biodynamics in the oligochaete Lumbriculus variegatus and its implications for trophic transfer.

It has become increasingly apparent that diet can be a major source of trace metal bioaccumulation in aquatic organisms. In this study, we examined cadmium uptake, efflux, and subcellular compartmentalization dynamics in the freshwater oligochaete Lumbriculus variegatus. L. variegatus is an important component of freshwater food webs in Europe and North America and is potentially useful as a standard food source for laboratory-based trophic transfer studies. Cadmium accumulation and depuration were each followed for 10 days. Rate constants of uptake (k(u)) and efflux (k(e)) were estimated and subcellular Cd compartmentalization was followed over the course of uptake and efflux. The partitioning of Cd into operationally-defined subcellular compartments was relatively consistent throughout the 20-day experiment, with the majority of Cd accumulating in the cytosol. No major changes in Cd compartmentalization were observed over uptake or depuration, but there appeared to be some exchange between heat-stable and heat-labile cytosolic protein fractions. Cadmium accumulation from solution was strongly affected by ambient calcium concentrations, suggesting competition between Cd and Ca for uptake sites. Finally, we demonstrate the ability to manipulate the whole body calcium content of L. variegatus as a potential tool for examining calcium influences on dietary Cd dynamics. The potential for this species to be an important conduit of Cd to higher trophic levels is discussed, along with its potential as a standardized food source in metal trophic transfer studies.

Cadmium ecophysiology in seven stonefly (Plecoptera) species: delineating sources and estimating susceptibility.

A major challenge in ecotoxicology lies in generating data under experimental conditions that are relevant to understanding contaminant effects in nature. Biodynamic modeling combines species-specific physiological traits to make predictions of metal bioaccumulation that fare well when tested in the field. We generated biodynamic models for seven predatory stonefly (Plecoptera) species representing the families Perlidae (5) and Perlodidae (2). Each taxon was exposed to cadmium independently via diet and via solution. Species varied approximately 2.6 fold in predicted steady-state cadmium concentrations. Diet was the predominant source of accumulated cadmium in five of the seven species and averaged 53.2 +/- 9.6% and 90.2 +/- 3.7% of net Cd accumulation in perlids and perlodids, respectively. Differences in Cd bioaccumulation between the two families were largely driven by differences in dissolved accumulation rates, which were considerably slower in perlodidsthan in perlids. We further examined the subcellular compartmentalization of Cd accumulated from independent aqueous and dietary exposures. Predicted steady-state concentrations were modified to only consider Cd accumulated in metal-sensitive subcellular compartments. These values ranged 5.3 fold. We discuss this variability within a phylogenetic context and its implications for bioassessment.

Influence of metal exposure history on the bioaccumulation and subcellular distribution of aqueous cadmium in the insect Hydropsyche californica.

The influence of metal exposure history on rates of aqueous Cd accumulation, elimination, and subcellular distribution was examined in the aquatic insect Hydropsyche californica. Specimens were obtained from a reference site and a metal-contaminated site and returned to the laboratory where they were continuously exposed to aqueous Cd (518 ng/L, nominal) for 6 d, followed by 9 d of depuration. Rates of Cd accumulation and elimination were similar in insects from the two sites. Efflux rate constants, k((e), ranged from 0.20 to 0.24/d (t 1/2 approximately 3 d). Immediately following exposure, the cytosol accounted for 40% of the body burden in insects from both sites; however, 89 +/- 2% of the cytosolic Cd was associated with metallothionein-like proteins (MTLP) in insects from the contaminated site, compared to 60 +/- 0% in insects from the reference site. The concentration of Cd bound to non-MTLPs (representing potentially Cd-sensitive proteins) was significantly greater in the insects from the reference site (134 +/- 7 ng/g) than in those from the contaminated site (42 +/- 2 ng/g). At the end of the depuration period, 90% of the accumulated Cd body burden had been eliminated, and Cd concentrations in MTLPs and non-MTLPs were similar between the sites. Results suggested that differences in exposure history had no influence on the bioaccumulation of Cd, but did affect the concentrations of Cd bound to MTLP during Cd exposure in these insects.

Linking metal bioaccumulation of aquatic insects to their distribution patterns in a mining-impacted river.

Although the differential responses of stream taxa to metal exposure have been exploited for bioassessment and monitoring, the mechanisms affecting these responses are not well understood. In this study, the subcellular partitioning of metals in operationally defined metal-sensitive and detoxified fractions were analyzed in five insect taxa. Samples were collected in two separate years along an extensive metal contamination gradient in the Clark Fork River (MT, USA) to determine if interspecific differences in the metal concentrations of metal-sensitive fractions and detoxified fractions were linked to the differences in distributions of taxa relative to the gradient. Most of the Cd, Cu, and Zn body burdens were internalized and potentially biologically active in all taxa, although all taxa appeared to detoxify metals (e.g., metal bound to cytosolic metal-binding proteins). Metal concentrations associated with metal-sensitive fractions were highest in the mayflies Epeorus albertae and Serratella tibialis, which were rare or absent from the most contaminated sites but occurred at less contaminated sites. Relatively low concentrations of Cu were common to the tolerant taxa Hydropsyche spp. and Baetis spp., which were widely distributed and dominant in the most contaminated sections of the river. This suggested that distributions of taxa along the contamination gradient were more closely related to the bioaccumulation of Cu than of other metals. Metal bioaccumulation did not appear to explain the spatial distribution of the caddisfly Arctopsyche grandis, considered to be a bioindicator of metal effects in the river. Thus, in this system the presence/ absence of most of these taxa from sites where metal exposure was elevated could be differentiated on the basis of differences in metal bioaccumulation.