Subcellular Accumulation and Depuration of Zinc in Periphytic Algae during Episodic and Continuous Exposures.

As the severity of extreme precipitation events increases with global climate change, so will episodic pulses of contamination into lotic systems. Periphytic algae represents bioindicator species in most freshwater systems due to their rapid accumulation of toxicants; therefore, it is vital to understand how accumulation in this group differs across temporally variable exposure regimes. The ability to rapidly accrue contaminants has additional implications for the trophic transfer of metals to primary consumers. While dietary toxicity has been studied in algivorous consumers, techniques used to prepare contaminated periphytic algae for consumption have not been compared. This study used a modified subcellular fractionation method to compare the partitioning of zinc (Zn) in periphyton cultures exposed for various durations (cultured in the presence of Zn and 15 min, 24 h, and 48 h exposures). Three exposure groups were additionally depurated over a period of 24 h in order to compare retention of Zn, an important aspect of preparing diets used in dietary toxicity studies. The results not only provide evidence for increased retention by periphytic algae cultured in the presence of Zn but reveal relationships among treatments and subcellular partitioning that suggest time-dependent accumulation and detoxification. These relationships suggest that episodic exposure of periphytic algae to contaminants may pose a greater risk than that of chronic regimes. Based on these results, we additionally advocate for culturing periphytic algae in the presence of contamination to produce a more reliable diet for dietary exposure testing in algivorous organisms.

Metals Alter Membership but Not Diversity of a Headwater Stream Microbiome.

Metal contamination from mining or natural weathering is a common feature of surface waters in the American west. Advances in microbial analyses have created the potential for routine sampling of aquatic microbiomes as a tool to assess the quality of stream habitat. We sought to determine if microbiome diversity and membership were affected by metal contamination and identify candidate microbial taxa to be used to indicate metal stress in stream ecosystems. We evaluated microbiome membership from sediments at multiple sites within the principal drainage of an EPA superfund site near the headwaters of the Upper Arkansas River, Leadville, CO. From each sample, we extracted DNA and sequenced the 16S rRNA gene amplicon on the Illumina MiSeq platform. We used the remaining sediments to simultaneously evaluate environmental metal concentrations. We also conducted an artificial stream mesocosm experiment using sediments collected from two of the observational study sites. The mesocosm experiment had a two-by-two factorial design: (i) location (upstream or downstream of contaminating tributary), and (ii) treatment (metal exposure or control). We found no difference in diversity between upstream and downstream sites in the field. Similarly, diversity changed very little following experimental metal exposure. However, microbiome membership differed between upstream and downstream locations and experimental metal exposure changed microbiome membership in a manner that depended on origin of the sediments used in each mesocosm.IMPORTANCE Our results suggest that microbiomes can be reliable indicators of ecosystem metal stress even when surface water chemistry and other metrics used to assess ecosystem health do not indicate ecosystem stress. Results presented in this study, in combination with previously published work on this same ecosystem, are consistent with the idea that a microbial response to metals at the base of the food web may be affecting primary consumers. If effects of metals are mediated through shifts in the microbiome, then microbial metrics, as presented here, may aid in the assessment of stream ecosystem health, which currently does not include assessments of the microbiome.

Size-Dependent Sensitivity of Aquatic Insects to Metals.

Laboratory assessments of aqueous metal toxicity generally demonstrate aquatic insects tolerate relatively high concentrations of metals in aqueous exposures; however, mesocosm experiments and field biomonitoring often indicate effects at relatively low metal concentrations. One hypothesis proposed to reconcile this discrepancy is an increased sensitivity of smaller size classes of organisms. We exposed field colonized benthic communities to aqueous metals in a series of mesocosm experiments. In addition, a novel single-species test system was used to expose first instar, mid-instar, and late instar mayflies (Ephemeroptera, Baetis tricaudatus) to Zn. These experimental approaches tested the hypothesis that small invertebrate size classes are more sensitive than large, mature size classes. Mesocosm results demonstrated strong size-dependent responses of aquatic insects to metals. Smaller organisms generally displayed greater mortality than large, mature individuals, and models were improved when size was included as a predictor of mortality. Size-dependent responses of Baetis spp. occurred in mesocosm experiments and in our single-species test system. The median lethal concentration (LC50) for early instar B. tricaudatus was less than 6% of the previously reported LC50 for late instars. Together, these results suggest that aquatic insect body size is an important predictor of susceptibility to aqueous metals. Toxicity models that account for insect phenology by integrating the natural size progression of organisms have the potential to improve accuracy in predicting effects of metals in the field.

Stream Mesocosm Experiments Show Significant Differences in Sensitivity of Larval and Emerging Adults to Metals.

Evaluations of aquatic insect responses to contaminants typically use larval life stages to characterize taxa sensitivity, but the effects of contaminants to emerging terrestrial adults have received less attention. We present the results of two stream mesocosm experiments that exposed aquatic insects to mixtures of Cu and Zn. We compared responses of larvae and emerging adults in a single-species experiment with the mayfly Rhithrogena robusta and a benthic community experiment. Results showed that R. robusta larvae and emerging adults were highly tolerant of metals. In the benthic community experiment, larval and emerging adult life stages of the mayfly Baetidae were highly sensitive to metals exposure, with significant alterations in adult sex ratios. In contrast, the emergence of Chironomidae (midge) was unaffected, but larval abundance strongly decreased. Timing of adult emergence was significantly different among treatments and varied among taxa, with emergence stimulation in Chironomidae and delays in emergence in R. robusta and Simuliidae (black fly). Our results demonstrate that metal tolerance in aquatic insects is life stage dependent and that taxa sensitivity is influenced by a combination of physiology and phylogeny. Regulatory frameworks would benefit by including test results that account for effects of contaminants on metamorphosis and adult insect emergence for the development of aquatic life standards.

Indirect Effects of Iron Oxide on Stream Benthic Communities: Capturing Ecological Complexity with Controlled Mesocosm Experiments.

Ferric iron (Fe(III)) oxyhydroxides commonly precipitate at neutral pH and in highly oxygenated conditions in waterways receiving acid mine drainage, degrading stream benthic communities by smothering of habitat, primary producers, and aquatic invertebrates. Stream mesocosms were used to expose naturally colonized benthic communities to a gradient of ferric Fe (0-15 mg/L) for 14 days to estimate the effects of Fe precipitates on primary production, larval and emerging adult aquatic insects, and the macroinvertebrate community structure. Community composition was significantly altered at concentrations near or below the US Environmental Protection Agency chronic Fe criterion (1.0 mg/L). Iron exposure significantly decreased larval and emerging adult abundances of Baetidae (mayfly) and Chironomidae (Diptera); however, while Simuliidae (Diptera) larvae were not reduced by the Fe treatments, abundance of emerged adults significantly decreased. Iron substantially decreased the colonization biomass of green algae and diatoms, with estimated EC20 values well below the Fe criterion. In contrast, cyanobacteria were stimulated with increasing Fe concentration. By integrating environmentally realistic exposure conditions to native benthic communities that have complex structural and functional responses, the ability to predict the effects of Fe in the field is improved. Traditional toxicity testing methodologies were not developed to evaluate indirect effects of contaminants, and modernized approaches such as community mesocosm experiments better characterize and predict responses in aquatic ecosystems outside the laboratory. Therefore, the development of water quality standards would benefit by including mesocosm testing results.

Integrating Results of Field Biomonitoring and Mesocosm Experiments To Validate Postspill Impacts of Petroleum Hydrocarbons on Stream Benthic Communities.

We integrated the results of field surveys with a mesocosm experiment to quantify the ecological impacts of a gasoline and diesel spill on a third-order stream in western Colorado (United States). The spill caused a massive fish-kill of brown trout ( Salmo trutta) and mottled sculpin ( Cottus bairdii), which extended several kilometers downstream. Despite significant decreases in petroleum hydrocarbon concentrations, subsequent surveys indicated little recovery of fish populations 4 years after the spill. Benthic macroinvertebrate communities were also affected, yet some metrics commonly used to assess stream ecological integrity (e.g., total abundance and species richness) showed no difference between impacted and reference sites. The failure of some groups to recover 2.5 years after the spill was likely a result of their comparatively slow reproduction and recolonization rates. To support our hypothesis that effects observed in the field resulted from petroleum exposure, we conducted a mesocosm experiment in which benthic macroinvertebrate communities were exposed to a simulated diesel spill. We observed significant decreases in the abundance of most macroinvertebrate groups at the lowest exposure concentration (75 mg/L diesel) and a strong concentration-dependent drift response across all groups. Our study suggests that relatively small petroleum spills can significantly affect stream communities, and these effects may persist several years after sediment concentrations return to background levels.

Quantifying Differences in Responses of Aquatic Insects to Trace Metal Exposure in Field Studies and Short-Term Stream Mesocosm Experiments.

Characterizing macroinvertebrate taxa as either sensitive or tolerant is of critical importance for investigating impacts of anthropogenic stressors in aquatic ecosystems and for inferring causality. However, our understanding of relative sensitivity of aquatic insects to metals in the field and under controlled conditions in the laboratory or mesocosm experiments is limited. In this study, we compared the response of 16 lotic macroinvertebrate families to metals in short-term (10-day) stream mesocosm experiments and in a spatially extensive field study of 154 Colorado streams. Comparisons of field and mesocosm-derived EC20 (effect concentration of 20%) values showed that aquatic insects were generally more sensitive to metals in the field. Although the ranked sensitivity to metals was similar for many families, we observed large differences between field and mesocosm responses for some groups (e.g., Baetidae and Heptageniidae). These differences most likely resulted from the inability of short-term experiments to account for factors such as dietary exposure to metals, rapid recolonization in the field, and effects of metals on sensitive life stages. Understanding mechanisms responsible for differences among field, mesocosm, and laboratory approaches would improve our ability to predict contaminant effects and establish ecologically meaningful water-quality criteria.

Influence of Metal Contamination and Sediment Deposition on Benthic Invertebrate Colonization at the North Fork Clear Creek Superfund Site, Colorado, USA.

Assessing benthic invertebrate community responses to multiple stressors is necessary to improve the success of restoration and biomonitoring projects. Results of mesocosm and field experiments were integrated to predict how benthic macroinvertebrate communities would recover following the removal of acid mine drainage from the North Fork of Clear Creek (NFCC), a U.S. EPA Superfund site in Colorado, USA. We transferred reference and metal-contaminated sediment to an upstream reference site where colonization by benthic macroinvertebrates was measured over 30 days. Additionally, a mesocosm experiment was performed to test the hypothesis that patches of metal-contaminated substrate impede recolonization downstream. Abundance in all treatments increased over time during field experiments; however, colonization was slower in treatments with metal-contaminated fine sediment. Community assemblages in treatments with metal-contaminated fine substrate were significantly different from other treatments. Patterns in the mesocosm study were consistent with results of the field experiment and showed greater separation in community structure between streams with metal-contaminated sediments and reference-coarse habitats; however, biological traits also helped explain downstream colonization. This study suggests that after water quality improvements at NFCC, fine-sediment deposition will likely reduce recovery potential for some taxa; however highly mobile taxa that avoid patches of contaminated habitats can recover quickly.

Metamorphosis Affects Metal Concentrations and Isotopic Signatures in a Mayfly (Baetis tricaudatus): Implications for the Aquatic-Terrestrial Transfer of Metals.

Insect metamorphosis often results in substantial chemical changes that can alter contaminant concentrations and fractionate isotopes. We exposed larval mayflies (Baetis tricaudatus) and their food (periphyton) to an aqueous zinc gradient (3-340 µg Zn/l) and measured zinc concentrations at different stages of metamorphosis: larval, subimago, and imago. We also measured changes in stable isotopes (δ15N and δ13C) in unexposed mayflies. Larval zinc concentrations were positively related to aqueous zinc, increasing 9-fold across the exposure gradient. Adult zinc concentrations were also positively related to aqueous zinc, but were 7-fold lower than larvae. This relationship varied according to adult substage and sex. Tissue concentrations in female imagoes were not related to exposure concentrations, but the converse was true for all other stage-by-sex combinations. Metamorphosis also increased δ15N by ∼0.8‰, but not δ13C. Thus, the main effects of metamorphosis on insect chemistry were large declines in zinc concentrations coupled with increased δ15N signatures. For zinc, this change was largely consistent across the aqueous exposure gradient. However, differences among sexes and stages suggest that caution is warranted when using nitrogen isotopes or metal concentrations measured in one insect stage (e.g., larvae) to assess risk to wildlife that feed on subsequent life stages (e.g., adults).

The Use of Field and Mesocosm Experiments to Quantify Effects of Physical and Chemical Stressors in Mining-Contaminated Streams.

Identifying causal relationships between acid mine drainage (AMD) and ecological responses in the field is challenging. In addition to the direct toxicological effects of elevated metals and reduced pH, mining activities influence aquatic organisms indirectly through physical alterations of habitat. The primary goal of this research was to quantify the relative importance of physical (metal-oxide deposition) and chemical (elevated metal concentrations) stressors on benthic macroinvertebrate communities. Mesocosm experiments conducted with natural assemblages of benthic macroinvertebrates established concentration-response relationships between metals and community structure. Field experiments quantified effects of metal-oxide contaminated substrate and showed significant differences in sensitivity among taxa. To predict the recovery of dominant taxa in the field, we integrated our measures of metal tolerance and substrate tolerance with estimates of drift propensity obtained from the literature. Our estimates of recovery were consistent with patterns observed at downstream recovery sites in the NFCC, which were dominated by caddisflies and baetid mayflies. We conclude that mesocosm and small-scale field experiments, particularly those conducted with natural communities, provide an ecologically realistic complement to laboratory toxicity tests. These experiments also control for the confounding variables associated with field-based approaches, thereby supporting causal relationships between AMD stressors and responses.

Ecological Effects of Biochar on the Structure and Function of Stream Benthic Communities.

The introduction of biochar, activated carbon, and other carbonaceous materials to aquatic ecosystems significantly reduces the toxicity and bioavailability of contaminants. However, previous studies have shown that these materials can have negative effects on aquatic organisms. We conducted field and mesocosm experiments to test the hypothesis that biochar altered the structure and function of stream benthic communities. After 30 d in the field, colonization by stoneflies (Plecoptera) was significantly lower in trays containing biochar compared to the results from the controls. In stream mesocosms, biochar increased macroinvertebrate drift and significantly reduced community metabolism. However, most measures of community composition showed little variation among biochar treatments, and significant responses were limited to a single stonefly species (Capnia confusa). When benthic communities were simultaneously exposed to biochar and Cu, effects were primarily associated with metal exposure. Because it is unlikely that biochar treatments would be employed in uncontaminated areas, these moderately negative effects should be considered within the context of the positive benefits associated with reduced contaminant bioavailability and toxicity. Additional research is necessary to improve our understanding of the mechanisms responsible for biochar effects on benthic communities and to identify the optimal application rates and size fractions that will maximize contaminant sorption but minimize potential negative effects.

Responses of aquatic insects to Cu and Zn in stream microcosms: understanding differences between single species tests and field responses.

Field surveys of metal-contaminated streams suggest that some aquatic insects, particularly mayflies (Ephemeroptera) and stoneflies (Plecoptera), are highly sensitive to metals. However, results of single species toxicity tests indicate these organisms are quite tolerant, with LC50 values often several orders of magnitude greater than those obtained using standard test organisms (e.g., cladocerans and fathead minnows). Reconciling these differences is a critical research need, particularly since water quality criteria for metals are based primarily on results of single species toxicity tests. In this research we provide evidence based on community-level microcosm experiments to support the hypothesis that some aquatic insects are highly sensitive to metals. We present results of three experiments that quantified effects of Cu and Zn, alone and in combination, on stream insect communities. EC50 values, defined as the metal concentration that reduced abundance of aquatic insects by 50%, were several orders of magnitude lower than previously published values obtained from single species tests. We hypothesize that the short duration of laboratory toxicity tests and the failure to evaluate effects of metals on sensitive early life stages are the primary factors responsible for unrealistically high LC50 values in the literature. We also observed that Cu alone was significantly more toxic to aquatic insects than the combination of Cu and Zn, despite the fact that exposure concentrations represented theoretically similar toxicity levels. Our results suggest that water quality criteria for Zn were protective of most aquatic insects, whereas Cu was highly toxic to some species at concentrations near water quality criteria. Because of the functional significance of aquatic insects in stream ecosystems and their well-established importance as indicators of water quality, reconciling differences between field and laboratory responses and understanding the mechanisms responsible for variation in sensitivity among metals and metal mixtures is of critical importance.

Integrated Assessment of Chemical and Biological Recovery After Diversion and Treatment of Acid Mine Drainage in a Rocky Mountain Stream.

Responses of stream ecosystems to gradual reductions in metal loading following remediation or restoration activities have been well documented in the literature. However, much less is known about how these systems respond to the immediate or more rapid elimination of metal inputs. Construction of a water treatment plant on the North Fork of Clear Creek (NFCC; CO, USA), a US Environmental Protection Agency Superfund site, captured, diverted, and treated the two major point-source inputs of acid mine drainage (AMD) and provided an opportunity to investigate immediate improvements in water quality. We conducted a 9-year study that included intensive within- and among-year monitoring of receiving-stream chemistry and benthic communities before and after construction of the treatment plant. Results showed a 64%-86% decrease in metal concentrations within months at the most contaminated sites. Benthic communities responded with increased abundance and diversity, but downstream stations remained impaired relative to reference conditions, with significantly lower taxonomic richness represented by a few dominant taxa (i.e., Baetis sp., Hydropsyche sp., Simulium sp., Orthocladiinae). Elevated metal concentrations from apparent residual sources, and relatively high conductivity from contributing major ions not removed during the treatment process, are likely limiting downstream recovery. Our study demonstrates that direct AMD treatment can rapidly improve water quality and benefit aquatic life, but effectiveness is limited, in part, to the extent that inputs of metals are captured and treated. Consideration should also be given to the effects of elevated major ion concentrations from the treated effluent not removed during the lime treatment process. Continued chemical and biological monitoring will be needed to quantify the NFCC recovery trajectory and to inform future remediation strategies. Environ Toxicol Chem 2023;42:512-524. © 2022 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC. This article has been contributed to by U.S. Government employees and their work is in the public domain in the USA.

Geologic processes influence the effects of mining on aquatic ecosystems.

Geologic processes strongly influence water and sediment quality in aquatic ecosystems but rarely are geologic principles incorporated into routine biomonitoring studies. We test if elevated concentrations of metals in water and sediment are restricted to streams downstream of mines or areas that may discharge mine wastes. We surveyed 198 catchments classified as "historically mined" or "unmined," and based on mineral-deposit criteria, to determine whether water and sediment quality were influenced by naturally occurring mineralized rock, by historical mining, or by a combination of both. By accounting for different geologic sources of metals to the environment, we were able to distinguish aquatic ecosystems limited by metals derived from natural processes from those due to mining. Elevated concentrations of metals in water and sediment were not restricted to mined catchments; depauperate aquatic communities were found in unmined catchments. The type and intensity of hydrothermal alteration and the mineral deposit type were important determinants of water and sediment quality as well as the aquatic community in both mined and unmined catchments. This study distinguished the effects of different rock types and geologic sources of metals on ecosystems by incorporating basic geologic processes into reference and baseline site selection, resulting in a refined assessment. Our results indicate that biomonitoring studies should account for natural sources of metals in some geologic environments as contributors to the effect of mines on aquatic ecosystems, recognizing that in mining-impacted drainages there may have been high pre-mining background metal concentrations.

Stream Mesocosm Experiments Show no Protective Effects of Calcium on Copper Toxicity to Macroinvertebrates.

Although the concept and modeling of metal bioavailability and toxicity have been well developed based largely on laboratory experiments with standard test species, additional evidence is required to demonstrate their applicability for macroinvertebrates typically found in natural lotic ecosystems. We conducted 10-day stream mesocosm experiments to test the hypothesis that increased water hardness (in the present study, the calcium [Ca] concentration was increased by adding CaCl2 ) would mitigate the effects of copper (Cu) on natural benthic macroinvertebrate communities. Exposure of macroinvertebrate communities to 25 µg/L Cu for 10 days in stream mesocosm experiments resulted in significant decreases in total abundance, in number of taxa, and in abundance of many macroinvertebrate taxa. However, the addition of Ca to stream mesocosms and the associated increase in water hardness up to 250 mg/L CaCO3 did not mitigate these effects of Cu on macroinvertebrate communities. The results showed that the hardness-based water quality criteria for Cu of the US Environmental Protection Agency were not protective under the conditions of relatively high hardness, low alkalinity, and circumneutral pH. In contrast, the water quality criteria based on the biotic ligand model predicted little protective effects of Ca on Cu toxicity, which is consistent with our results. Additional experiments are required to understand the influence of modifying factors on the toxicity of metals to macroinvertebrate communities. Environ Toxicol Chem 2022;41:1304-1310. © 2022 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Habitat restoration for brown trout (Salmo trutta) has limited effects on macroinvertebrate communities in a historically metal-contaminated stream.

Quantifying the success of stream remediation or restoration projects that are designed to improve water quality or habitat, respectively, is often challenging because of insufficient posttreatment monitoring, poorly defined restoration goals, and failure to consider fundamental aspects of ecological theory. We measured the effects of habitat restoration on aquatic and terrestrial prey resources in a system recovering from the long-term effects of mining pollution. The study was conducted in the Upper Arkansas River, a Rocky Mountain stream located in central Colorado, USA. Remediation of California Gulch, a United States Environmental Protection Agency (USEPA) Superfund Site that discharged metals from past mining operations into the stream, was completed in 2000, resulting in significant improvements in water quality, benthic macroinvertebrate communities, and brown trout (Salmo trutta) populations. A large-scale restoration project designed to improve habitat and increase the density and biomass of brown trout was completed in 2014. To assess the effectiveness of these habitat improvements on invertebrate communities in this system, we sampled sites for 9 years before (2010-2014) and after (2015-2018) restoration was completed. In contrast to our expectations, we observed few changes in the abundance of aquatic or terrestrial invertebrates after restoration. The most common response was an overall reduction in abundance resulting from significant instream disturbances during and immediately after restoration, followed by a gradual return to pretreatment conditions. Despite reductions in prey abundance, the number of prey items in the diet of brown trout increased significantly after restoration. We discuss several explanations for these responses, including the effects of residual metals, increased predation by brown trout, and the recalcitrance of novel communities dominated by metal-tolerant species. Our results suggest that the effectiveness of remediation and restoration differed between macroinvertebrates and fish. Benthic macroinvertebrates were more dependent on water quality improvements at the watershed scale, whereas brown trout populations responded to both improvements in water quality and reach-scale improvements in habitat. Integr Environ Assess Manag 2022;18:1047-1055. © 2021 SETAC.

Critical tissue residue approach linking accumulated metals in aquatic insects to population and community-level effects.

Whole body Zn concentrations in individuals (n = 825) from three aquatic insect taxa (mayflies Rhithrogena spp. and Drunella spp. and the caddisfly Arctopsyche grandis) were used to predict effects on populations and communities (n = 149 samples). Both mayflies accumulated significantly more Zn than the caddisfly. The presence/absence of Drunella spp. most reliably distinguished sites with low and high Zn concentrations; however, population densities of mayflies were more sensitive to increases in accumulated Zn. Critical tissue residues (634 µg/g Zn for Drunella spp. and 267 µg/g Zn for Rhithrogena spp.) caused a 20% reduction in maximum (90th quantile) mayfly densities. These critical tissue residues were associated with exposure to 7.0 and 3.9 µg/L dissolved Zn for Drunella spp. and Rhithrogena spp., respectively. A threshold in a measure of taxonomic completeness (observed/expected) was observed at 5.4 µg/L dissolved Zn. Dissolved Zn concentrations associated with critical tissue residues in mayflies were also associated with adverse effects in the aquatic community as a whole. These effects on populations and communities occurred at Zn concentrations below the U.S. EPA hardness-adjusted continuous chronic criterion.

Before-After Control-Impact field surveys and novel experimental approaches provide valuable insights for characterizing stream recovery from acid mine drainage.

Mineral extraction has resulted in widespread stream impairment due to habitat degradation and water quality impacts from acid mine drainage (AMD). The North Fork of Clear Creek (NFCC), Colorado, USA was historically impaired by AMD from two major point-source inputs, with some stream segments devoid of aquatic life prior to remediation. In the summer of 2017, the North Clear Creek Water Treatment Plant (NCCWTP) began AMD water treatment. To predict and characterize the biological recovery of NFCC to improvements in water quality, we conducted stream mesocosm and field experiments, as well as biomonitoring of benthic communities using a Before-After Control-Impact (BACI) study design. The NFCC stream community responded rapidly to improved water quality. Benthic algal biomass increased at impacted sites and macroinvertebrate surveys showed significant increases in abundance, taxa richness, and emerging adult aquatic insects. However, the dominant taxa colonizing downstream segments of NFCC differed considerably from those predicted based on previous field and experimental results. We hypothesize that this discrepancy is the result of differences in metal exposure regimes observed between our field and mesocosm approaches (i.e., fluctuating vs stable), colonization attributes (i.e., open vs closed system), and spatiotemporal differences in metal sensitivity due to macroinvertebrate phenology. We expect continued biological recovery in NFCC, but habitat impairment and residual sources of metals will continue to impair aquatic life until those stressors abate. Applying a combination of controlled experimental and BACI field approaches to predict and evaluate AMD-remediation projects in the future will improve the ability to understand the physical, chemical, and biological mechanisms influencing stream recovery.

Validation of Bioavailability-Based Toxicity Models for Metals.

Regulatory jurisdictions worldwide are increasingly incorporating bioavailability-based toxicity models into development of protective values (PVALs) for freshwater and saltwater aquatic life (e.g., water quality criteria, standards, and/or guidelines) for metals. Use of such models for regulatory purposes should be contingent on their ability to meet performance criteria as specified through a model-validation process. Model validation generally involves an assessment of a model's appropriateness, relevance, and accuracy. We review existing guidance for validation of bioavailability-based toxicity models, recommend questions that should be addressed in model-validation studies, discuss model study type and design considerations, present several new ways to evaluate model performance in validation studies, and suggest a framework for use of model validation in PVAL development. We conclude that model validation should be rigorous but flexible enough to fit the user's purpose. Although a model can never be fully validated to a level of zero uncertainty, it can be sufficiently validated to fit a specific purpose. Therefore, support (or lack of support) for a model should be presented in such a way that users can choose their own level of acceptability. We recommend that models be validated using experimental designs and endpoints consistent with the data sets that were used to parameterize and calibrate the model and validated across a broad range of geographically and ecologically relevant water types. Environ Toxicol Chem 2019;39:101-117. © 2019 SETAC.