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.

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.

The effects of chronological age and size on toxicity of zinc to juvenile brown trout.

A series of toxicity tests were conducted to investigate the role of chronological age on zinc tolerance in juvenile brown trout (Salmo trutta). Four different incubation temperatures were used to control the maturation of the juveniles before zinc exposures. These 96-h exposures used flow-through conditions and four chronological ages of fish with weights ranging from 0.148 to 1.432 g. Time-to-death (TTD) data were collected throughout the exposure along with the final mortality. The results indicate that chronological age does not play a predictable role in zinc tolerance for juvenile brown trout. However, a relationship between zinc tolerance and fish size was observed in all chronological age populations, which prompted us to conduct additional exploratory data analysis to quantify how much of an effect size had during this stage of development. The smallest fish (0.148-0.423 g) were shown to be less sensitive than the largest fish (0.639-1.432 g) with LC50 values of 868 and 354 µg Zn/L, respectively. The Kaplan-Meier product estimation method was used to determine survival functions from the TTD data and supports the LC50 results with a greater median TTD for smaller fish than larger juvenile fish. These results indicate that fish size or a related characteristic may be a significant determinant of susceptibility and should be considered in acute zinc toxicity tests with specific attention paid to the expected exposure scenario in the field.

Acclimation and deacclimation of brown trout (Salmo trutta) to zinc and copper singly and in combination with cadmium or copper.

Brown trout (Salmo trutta) were chronically exposed to low and high levels of zinc (Zn) alone, copper (Cu) alone, a Zn-Cu mixture, and a Zn-cadmium (Cd) mixture all starting with eyed eggs and continuing through to the fingerling stage. Exposure to the metals and metal mixtures resulted in acclimation as measured by greater median lethal concentrations (LC50) values relative to metal-naïve fry. The degree of acclimation was similar between the low and high exposures except for Cu, where acclimation was observed at the high but not the low acclimation level. The increases in tolerance relative to metal-naïve controls were usually less than a factor of 2 and never exceeded a factor of 3. Acclimation exposures did not affect hatch or survival except for the high-acclimation regime of Zn plus Cu. Acclimation came at an apparent metabolic cost because growth was decreased by most acclimation exposures. Deacclimation, as evidenced by a return of LC50 values to naïve levels, occurred after 2-5 weeks in clean water.

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.

Acute toxicity of zinc to several aquatic species native to the Rocky Mountains.

National water-quality criteria for the protection of aquatic life are based on toxicity tests, often using organisms that are easy to culture in the laboratory. Species native to the Rocky Mountains are poorly represented in data sets used to derive national water-quality criteria. To provide additional data on the toxicity of zinc, several laboratory acute-toxicity tests were conducted with a diverse assortment of fish, benthic invertebrates, and an amphibian native to the Rocky Mountains. Tests with fish were conducted using three subspecies of cutthroat trout (Colorado River cutthroat trout Oncorhynchus clarkii pleuriticus, greenback cutthroat trout O. clarkii stomias, and Rio Grande cutthroat trout O. clarkii virginalis), mountain whitefish (Prosopium williamsoni), mottled sculpin (Cottus bairdi), longnose dace (Rhinichthys cataractae), and flathead chub (Platygobio gracilis). Aquatic invertebrate tests were conducted with mayflies (Baetis tricaudatus, Drunella doddsi, Cinygmula sp. and Ephemerella sp.), a stonefly (Chloroperlidae), and a caddis fly (Lepidostoma sp.). The amphibian test was conducted with tadpoles of the boreal toad (Bufo boreas). Median lethal concentrations (LC(50)s) ranged more than three orders of magnitude from 166 µg/L for Rio Grande cutthroat trout to >67,000 µg/L for several benthic invertebrates. Of the organisms tested, vertebrates were the most sensitive, and benthic invertebrates were the most tolerant.

Copper toxicity in Bristol Bay headwaters: Part 1-Acute mortality and ambient water quality criteria in low-hardness water.

The world-class Alaskan Bristol Bay salmon fishery and vast deposits of copper (Cu) and other metals in the watershed warrant further investigation into the potential toxicity of Cu to salmonids under the low water-hardness conditions that occur in the watershed. Therefore we investigated the acute toxicity of Cu to rainbow trout (Oncorhynchus mykiss) and fathead minnows (Pimephales promelas) in low-hardness water (∼ 30 mg/L as CaCO3 ) formulated in the laboratory and collected from the Bristol Bay watershed. The median lethal concentration (LC50) for rainbow trout exposed to Cu in low-hardness laboratory water was 16 µg Cu/L (95% confidence intervals [CIs]: 12, 21; dissolved Cu, filtered to 0.45 µm). The LC50 values for fathead minnows exposed to Cu in low-hardness laboratory water or site water were 29 and 79 µg Cu/L (95% CIs: 23, 35 and 58, 125; dissolved Cu), respectively. The biotic ligand model (BLM) LC50 estimates for these bioassays were 1.3 to 2.3 times higher than the actual LC50 values. We also calculated and analyzed acute Cu water quality criteria, also known as criterion maximum concentration (CMC), using hardness-based methods and the BLM for water samples collected throughout the Bristol Bay watershed in 2007. Biotic ligand model CMCs ranged from 0.05 to 17.5 µg Cu/L and hardness-based CMCs ranged from 2.3 to 6.1 µg Cu/L for the 65 samples analyzed. Our results show the need for site-specific research and subsequent water quality guidelines in low-hardness aquatic habitats. Environ Toxicol Chem 2019;38:190-197. © 2018 SETAC.

Copper toxicity in Bristol Bay headwaters: Part 2-Olfactory inhibition in low-hardness water.

We investigated the olfactory toxicity of copper (Cu) to rainbow trout in low-hardness (27 mg/L as CaCO3 ) water formulated in the laboratory over a 120-h period using a flow-through design. The fish's response to an alarm cue (e.g., reduction in activity) was recorded to determine the exposure concentrations and durations that inhibited olfactory detection of the cue after 3, 24, 48, and 96 h of Cu exposure and after 24 h of clean water recovery following the 96-h exposure period. Exposures were conducted with a range of Cu concentrations from 0.13 (control) to 7.14 µg Cu/L (dissolved Cu). We observed a dose-dependent response in olfactory inhibition with a 20% reduction in the probability of responding to the alarm cue, relative to controls, at 2.7 and 2.4 µg Cu/L after 24 or 96 h of exposure, respectively. Olfactory inhibition manifested between 3 and 24 h of exposure. Our 24- and 96-h 20% olfactory inhibition estimates fell between the criteria derived using the biotic ligand model (BLM; criterion maximum concentration [CMC] and criterion continuous concentration [CCC] values were 0.63 and 0.39 µg Cu/L, respectively) and water hardness-based criteria (CMC and CCC values were 3.9 and 2.9 µg Cu/L, respectively). Therefore, the hardness-based criteria do not appear to be protective and the BLM-derived criteria do appear to be protective against Cu-induced olfactory inhibition given our test water chemistry. Neither the hardness-based criteria nor the BLM-derived criteria appear to be protective against our estimated Cu behavioral avoidance response concentrations at 24- and 96-h exposures (0.54 and 0.50 µg Cu/L, respectively). Environ Toxicol Chem 2019;38:198-209. © 2018 SETAC.

Acute toxicity of aqueous copper, cadmium, and zinc to the mayfly Rhithrogena hageni.

Heptageniid mayfly nymphs have been suggested as sensitive indicators of metal contamination in streams based on biomonitoring studies, experimentation in situ, and experimentation in microcosm. Laboratory tests were conducted to evaluate the sensitivity of Rhithrogena hageni, a heptageniid mayfly, to waterborne copper, cadmium, and zinc. Tests were conducted with soft water (hardness = 40-50 mg/L) at about 12 degrees C. Toxicity endpoints were survival and moulting (%/day). Median 96 hr lethal concentrations were 0.137, 10.5, and 50.5 mg/L for copper, cadmium and zinc, respectively. The average daily moulting rate of survivors significantly decreased after exposure to these metals in solution.

Toxicity of cadmium to early life stages of brown trout (Salmo trutta) at multiple water hardnesses.

Toxicity of cadmium to early life stages of brown trout (Salmo trutta) was determined at multiple water hardnesses. Increasing water hardness decreased cadmium toxicity. Postswimup fry were much more sensitive than embryos and larvae. Chronic values from early life stage tests initiated with eyed embryos were 3.52, 6.36, and 13.6 microg Cd/L at water hardnesses of 30.6, 71.3, and 149 mg/L, respectively. In tests initiated with 30-d postswimup fry, chronic values were 1.02, 1.83, and 6.54 microg Cd/L at water hardnesses of 29.2, 67.6, and 151 mg/L, respectively. Higher chronic values from the early life stage tests compared to tests initiated with swimup fry likely are caused by acclimation during cadmium-tolerant embryo and larval stages. Growth was not affected by cadmium in the early life stage tests but was negatively affected in tests initiated with fry at water hardnesses of 29.2 and 67.6 mg/L. Concentrations of cadmium that reduced growth were higher than those that increased mortality. Median lethal concentrations for swimup fry after 96 h were 1.23, 3.90, and 10.1 microg Cd/L at water hardnesses of 29.2, 67.6, and 151 mg/L, respectively. Test results enable prediction of acute mortality of brown trout swimup fry based on cadmium concentration and water hardness.

Application of a generalized linear mixed model to analyze mixture toxicity: survival of brown trout affected by copper and zinc.

Increased concerns about the toxicity of chemical mixtures have led to greater emphasis on analyzing the interactions among the mixture components based on observed effects. The authors applied a generalized linear mixed model (GLMM) to analyze survival of brown trout (Salmo trutta) acutely exposed to metal mixtures that contained copper and zinc. Compared with dominant conventional approaches based on an assumption of concentration addition and the concentration of a chemical that causes x% effect (ECx), the GLMM approach has 2 major advantages. First, binary response variables such as survival can be modeled without any transformations, and thus sample size can be taken into consideration. Second, the importance of the chemical interaction can be tested in a simple statistical manner. Through this application, the authors investigated whether the estimated concentration of the 2 metals binding to humic acid, which is assumed to be a proxy of nonspecific biotic ligand sites, provided a better prediction of survival effects than dissolved and free-ion concentrations of metals. The results suggest that the estimated concentration of metals binding to humic acid is a better predictor of survival effects, and thus the metal competition at the ligands could be an important mechanism responsible for effects of metal mixtures. Application of the GLMM (and the generalized linear model) presents an alternative or complementary approach to analyzing mixture toxicity.