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.

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.

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.

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.

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.

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.

Context-Dependent Responses of Aquatic Insects to Metals and Metal Mixtures: A Quantitative Analysis Summarizing 24 Yr of Stream Mesocosm Experiments.

Modernizing water quality criteria to predict how contaminants affect natural aquatic communities requires that we utilize data obtained across multiple lines of evidence, including laboratory, mesocosm, and field studies. We report the results of 29 mesocosm experiments conducted from 1994 to 2017 at the Colorado State University Stream Research Laboratory (Fort Collins, CO, USA). The primary goal of the present study was to quantify responses of aquatic insect communities collected from 8 different locations to different combinations of cadmium (Cd), copper (Cu), iron (Fe), and zinc (Zn). Treatments that included Cu or Fe, either alone or in combination with other metals, were especially toxic to aquatic insects. The results showed that effects of metals were context dependent and varied significantly among the 8 sites where communities were collected. In particular, effects on communities from smaller streams were significantly greater than those from larger streams. Our analyses also showed that several morphological (body size, shape, gills, degree of sclerotization) and life history (voltinism) traits were significantly correlated with sensitivity to metals. Across all taxa and experiments, aquatic insects broadly classified as small (maximum body length <8 mm) were significantly more sensitive to metals than medium or large individuals. These findings demonstrate the advantages of integrating results of mesocosm experiments with species traits to develop a mechanistic understanding of biotic and abiotic factors that influence community responses to contaminants. Environ Toxicol Chem 2019;38:2486-2496. © 2019 SETAC.

Chronic Toxicity of Ferric Iron for North American Aquatic Organisms: Derivation of a Chronic Water Quality Criterion Using Single Species and Mesocosm Data.

Iron is a common pollutant in waters near coal and hard rock mine disturbances. The current 1000 µg/L total recoverable chronic criterion for iron (Fe) for protection of aquatic life in the United States was developed using very limited data in 1976 and has not been revised since. To develop a more scientifically based criterion, several chronic laboratory toxicity experiments (> 30 days) were conducted with ferric Fe at circumneutral pH on a taxonomically diverse group of organisms including brown trout (Salmo trutta), mountain whitefish (Prosopium williamsoni), boreal toad tadpoles (Bufo boreas), the oligochaete worm Lumbriculus variegatus, the mayfly Hexagenia limbata, and the planarian Dugesia dorotocephala. Results of these tests and those of previously published toxicity data were used to derive a Final Chronic Value (FCV) of 499 µg/L by using the US Environmental Protection Agency's recommended methods based on single species toxicity tests. In addition to single species toxicity tests, ferric Fe toxicity experiments (10 days) were performed on mesocosms containing naturally colonized communities of benthic macroinvertebrates. Fourteen genera in the mesocosms occurred at sufficient densities to estimate an iron concentration resulting in 20% reduction in abundance (EC20). Three of these taxa had EC20s less than the FCV of 499 µg/L derived from single species tests: the mayfly Epeorus sp. (335 µg/L), the caddisfly Micrasema sp. (356 µg/L), and midge Tanytarsini (234 µg/L). When mesocosm results were included, the FCV was lowered to 251 µg/L. These findings support the suggestion that modernization of water quality criteria should include data generated from mesocosm experiments and other lines of evidence.

Structural and functional responses of periphyton and macroinvertebrate communities to ferric Fe, Cu, and Zn in stream mesocosms.

Two mesocosm experiments were conducted to examine effects of ferric iron (Fe) and mixtures of ferric Fe with aqueous metals (Cu, Zn) on stream benthic communities. Naturally colonized benthic communities were exposed to a gradient of ferric Fe (0, 0.4, 1.0, 2.5, 6.2, and 15.6 mg/L) that bracketed the current US Environmental Protection Agency water quality criterion value (1.0 mg/L). After 10 d of exposure to ferric Fe, total macroinvertebrate abundance, number of taxa, and abundance of all major macroinvertebrate groups (Ephemeroptera, Plecoptera, Trichoptera, and Diptera) were significantly reduced. Heptageniid mayflies and chironomids were especially sensitive to Fe oxide deposition and were significantly reduced at 0.4 and 1.0 mg/L total Fe, respectively. In a second mesocosm experiment, periphyton and macroinvertebrate communities were exposed to ferric Fe (0.60 mg/L) with or without aqueous Cu and Zn at 2 treatment levels: low (0.01 mg/L Cu + 0.1 mg/L Zn) and high (0.05 mg/L Cu + 0.5 mg/L Zn). In contrast to previous research, we observed no evidence of a protective effect of Fe on toxicity of metals. Growth rates and protein content of periphyton were significantly reduced by both ferric Fe and aqueous metals, whereas abundance of heptageniid mayflies (Cinygmula) and whole community metabolism were significantly reduced by ferric Fe alone. We hypothesize that Fe oxides inhibited algal growth and enhanced metal accumulation, leading to a reduction in the quantity and quality of food resources for grazers. Mesocosm experiments conducted using natural benthic communities provide a unique opportunity to quantify the relative importance of indirect physical effects and to develop a better understanding of the relationship between basal food resources and consumers in natural stream ecosystems. Environ Toxicol Chem 2018;37:1320-1329. © 2017 SETAC.

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.

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.

Comparison of different predictors of exposure for modeling impacts of metal mixtures on macroinvertebrates in stream microcosms.

Knowledge about which predictors of metal exposure are best to model the impacts of metal mixtures on river macroinvertebrates remains uncertain. A new predictor based on the amount of metals binding to humic acid, which is assumed to be a proxy of non-specific biotic ligand sites, has been proposed. The amount can be calculated using Windermere Humic Aqueous Model (WHAM), which we will refer to as the WHAM-HA approach. Here, we tested the hypothesis that the predictor based on the WHAM-HA approach provided a better estimate of metal effects observed in microcosm experiments than three other measures: total metal concentrations, free metal ion concentrations, and the cumulative criterion unit (CCU) which is a measure of the ratios of measured metal concentrations relative to the U.S. Environmental Protection Agency hardness adjusted criterion values. For this evaluation, we used nine macroinvertebrate metrics of abundance and richness obtained from microcosm experiments conducted with metal mixtures (Zn alone, Zn+Cd, and Zn+Cd+Cu). For each of the four predictors, we performed multiple linear regression with variables corresponding to the three metal concentrations or CCU and selected the best model based on Akaike's information criterion corrected for small sample sizes. For all of the macroinvertebrate metrics affected by metals, the WHAM-HA approach was selected as the best among the four predictors, followed by the model with total metal concentration. In most of best models, Zn and Cu or Cu alone was responsible for reductions in invertebrate metrics, even though the highest concentrations of Cd exceeded 100 times the hardness-adjusted criterion value. Either of the models with free metal ion concentration and CCU was the third ranked model. Our results suggest that the estimated amount of metals binding to humic acid is a better predictor for the effects on macroinvertebrate richness and abundance observed in microcosm experiments than total or free ion concentrations of metals and CCU.