Tham C. Hoang

The Effects of Water Quality and Age on the Acute Toxicity of Copper to the Florida Apple Snail, Pomacea paludosa

Copper (Cu)-containing compounds have been used in Florida as fungicides, herbicides, and soil amendments, resulting in elevated Cu in the aquatic ecosystem. The Florida apple snail (Pomacea paludosa), a key species in south Florida, may be adversely affected by Cu. Water-quality parameters, such as hardness, dissolved organic carbon (DOC), pH, and alkalinity, affect metal bioavailability and toxicity in aquatic organisms; however, it is uncertain to what extent these factors affect Cu toxicity in the Florida apple snail. The research presented here characterized the acute (96-hour) toxicity of Cu in water to the Florida apple snail at various life stages and under different water-quality parameters. Cu was more toxic to juvenile than adult apple snails. There was no difference between the 96-hour LC50 at pH 5.5 and 6.5; however, the 96-hour LC50 values at pH 7.5 and 8.5 were greater than at lower pHs. The decrease in Cu2+ above pH 7, as predicted by the MINTEQ model, accounted for the pH effect. Cu toxicity decreased as DOC increased from 0.2 to 30 mg/L. Unlike other aquatic organisms, hardness had no effect on Cu toxicity to the Florida apple snail, suggesting another mechanism of toxicity. Whole-body tissue analysis indicated that the lethal body burden of 120-day-old snails exposed to Cu for 4 days was 30 mg/kg Cu dry weight. Multiple regression analysis indicated that Cu toxicity was a function of organism age, DOC, and pH.

Copper uptake and depuration by juvenile and adult Florida apple snails (Pomacea paludosa)

The present study characterized copper (Cu) uptake and depuration by juvenile and adult Florida apple snails (Pomacea paludosa) from water, soil, and diet. During a 28-day uptake period, juvenile apple snails were exposed to aqueous Cu and adult apple snails were exposed to Cu-contaminated soil, water, and food. In the follow-up 14-day depuration period, both juvenile and adult apple snails were held in laboratory freshwater with background Cu concentrations <4 μg/l. For juvenile apple snails, whole body Cu concentrations increased with time and reached a plateau after 14 days. The data followed Michaelis–Menten kinetics rather than a one compartment first order kinetics model. The mean Cu bioconcentration factor (BCF) for juvenile apple snails was 1493 and the depuration half-life was 10.5–13.8 days. For adult snails, dietary uptake of Cu resulted in higher bioaccumulation factors (BAFs) compared to uptake from soil. Most of the accumulated Cu was located in soft tissue (about 60% in the viscera and 40% in the foot). The shell contained <1% of the total accumulated copper. Soft tissue is usually consumed by predators of the apple snail. Therefore, the results of the present study show that Cu transfer through the food chain to the apple snail may lead to potential risk to its predators.

Use of butterflies as nontarget insect test species and the acute toxicity and hazard of mosquito control insecticides

Honeybees are the standard insect test species used for toxicity testing of pesticides on nontarget insects for the U.S. Environmental Protection Agency (U.S. EPA) under the Federal Insecticide Fungicide and Rodenticide Act (FIFRA). Butterflies are another important insect order and a valued ecological resource in pollination. The current study conducted acute toxicity tests with naled, permethrin, and dichlorvos on fifth larval instar (caterpillars) and adults of different native Florida, USA, butterfly species to determine median lethal doses (24-h LD50), because limited acute toxicity data are available with this major insect group. Thorax- and wing-only applications of each insecticide were conducted. Based on LD50s, thorax and wing application exposures were acutely toxic to both caterpillars and adults. Permethrin was the most acutely toxic insecticide after thorax exposure to fifth instars and adult butterflies. However, no generalization on acute toxicity (sensitivity) of the insecticides could be concluded based on exposures to fifth instars versus adult butterflies or on thorax versus wing exposures of adult butterflies. A comparison of LD50s of the butterflies from this study (caterpillars and adults) with honeybee LD50s for the adult mosquito insecticides on a µg/organism or µg/g basis indicates that several butterfly species are more sensitive to these insecticides than are honeybees. A comparison of species sensitivity distributions for all three insecticides shows that permethrin had the lowest 10th percentile. Using a hazard quotient approach indicates that both permethrin and naled applications in the field may present potential acute hazards to butterflies, whereas no acute hazard of dichlorvos is apparent in butterflies. Butterflies should be considered as potential test organisms when nontarget insect testing of pesticides is suggested under FIFRA.

Exposure routes of copper: Short term effects on survival, weight, and uptake in Florida apple snails (Pomacea paludosa)

The uptake and effects (survival, weight) of copper (Cu) on Florida apple snails (Pomacea paludosa) via exposures to copper-enriched agricultural soil-water and water-only treatments were investigated. Soils were collected from citrus sites in south Florida and flooded with laboratory freshwater for 14d. Neonate apple snails (96-h-old) were then exposed to either Cu from a soil-overlying water (i.e., flooded agricultural soils) treatment or overlying water-only (i.e., equilibrated overlying water produced from 14d flooding of agriculture soils) treatment for 14d under standard laboratory conditions. Survival, weight (dry, wet), and whole body Cu uptake were measured. Copper exposure via soil-water exposures resulted in higher mortality and whole body Cu uptake than water-only exposures, indicating Cu uptake from soils. However, snail wet and dry weights were higher in soil-water treatments than in water-only treatments. Micronutrients from soils may be consumed by snails increasing weights. Survival, apple snail dry weight, and whole body Cu concentrations were significantly correlated with soil and water Cu concentrations in soil-water treatments. Survival was significantly correlated with the concentration of Cu(CO3)2(2-) in water-only treatments. This suggests that Cu(CO3)2(2-) is toxic to apple snails. Whole body Cu concentrations were higher in surviving snails than dead snails, suggesting that apple snails have the ability to detoxify accumulated Cu (e.g., through metallothionein induction, granules).

ACUTE TOXICITY OF BINARY-METAL MIXTURES OF COPPER, ZINC, AND NICKEL TO PIMEPHALES PROMELAS: EVIDENCE OF MORE-THAN-ADDITIVE EFFECT

Metal mixture toxicity has been studied for decades. However, the results are not consistent, and thus ecological risk assessment and regulation of mixtures has been difficult. The objective of the present study was to use a systematic experimental design to characterize the toxicity of binary-metal mixture of Cu, Zn, and Ni to Pimephales promelas, typically to determine whether the effect of these binary-metal mixtures on P. promelas is more-than-additive. Standard 96-h toxicity tests were conducted with larval P. promelas based on US Environmental and Protection Agency methods to determine metal mixture effects. All experiments were conducted in synthetic moderately hard water with no addition of dissolved organic matter. Three different effect analysis approaches, the MixTox model, the Finney model, and the toxic unit method, were used for comparison. The results indicate that the toxicity of Cu+Zn, Cu+Ni, and Zn+Ni mixtures to P. promelas was more-than-additive. Among the 3 mixtures, the effect of the Cu+Ni mixture was the most profound. The results of the present study are useful for applications to models such as the metal mixture biotic ligand model. More research should be conducted to determine the mechanisms of acute and chronic toxicity of metal mixtures.

Bioconcentration and depuration of endosulfan sulfate in mosquito fish (Gambusia affinis).

Endosulfan is an insecticide which has been widely used in agriculture. The technical grade material consists of two isomers (alpha and beta). Under natural environmental conditions, endosulfan is metabolized through oxidation and the main metabolite in the environment is endosulfan sulfate. Most ecotoxicology research has been conducted with technical grade endosulfan to determine effects on non-target aquatic organisms. Little data on the effects of endosulfan sulfate on aquatic organisms are available in the literature. This study characterizes endosulfan sulfate bioconcentration and depuration in mosquito fish (Gambusia affinis). During the study, G. affinis was exposed to an environmentally relevant endosulfan sulfate concentration of 0.25 μg L(-1) for 5 weeks (uptake phase) followed by a 3-week period (depuration phase) in clean water. This study found that G. affinis bioconcentrated endosulfan sulfate. During the exposure phase, fish tissue concentrations of endosulfan sulfate increased with time up to 730 μg kg(-1) dw or 215 μg kg(-1) ww. The bioconcentration data followed Michaelis-Menten kinetics better than the one-compartment first order kinetics (1-CFOK). Using these models, the bioconcentration factors for endosulfan sulfate-exposed G. affinis were from 687 to 888 L kg(-1) in wet weight or 2263 to 2936 L kg(-1) in dry weight. During the depuration phase, endosulfan sulfate concentrations in tissue significantly decreased and the data followed first order kinetics. The half-life of endosulfan sulfate in G. affinis was about 9 d. There was no significant difference in standard length or weight between control and exposed fish. The growth data followed the von Bertalanffy growth model. However, the condition factor of exposed fish increased with time during the exposure phase.

Acute toxicity and risk assessment of permethrin, naled, and dichlorvos to larval butterflies via ingestion of contaminated foliage

Three Florida native larval butterflies (Junonia coenia, Anartia jatrophae, Eumaeus atala) were used in the present study to determine the acute toxicity, hazard, and risk of a 24h ingestion of leaves contaminated with the adult mosquito control insecticides permethrin, naled, and dichlorvos to late 4th and early 5th in-star caterpillars. Based on 24-h LD50s for ingestion, naled was more acutely toxic than permethrin and dichlorvos to caterpillars. Hazard quotients using the ratio of the highest doses and the 90th percentile doses from field measurements in host plant foliage following actual mosquito control applications to the toxicological benchmarks from laboratory toxicity tests indicate potential high acute hazard for naled compared to permethrin and dichlorvos. Based on probabilistic ecological risk methods, naled exposure doses in the environment also presented a higher acute risk to caterpillars than permethrin and dichlorvos. The acute toxicity laboratory results and ecological risk assessment are based only on dietary ingestion and single chemical doses. It does not include other typical exposure scenarios that may occur in the environment. It is thus plausible to state that the ecological risk assessment presented here underestimates the potential risks in the field to caterpillars. However, one assumption that is scientifically feasible and certainly real from the results - if the environmental exposure doses of mosquito control operations are similar or higher to those presented here in leaves from the field, after applications, there will likely be significant mortalities and other adverse effects on caterpillar populations.

Bioaccumulation and toxicity of copper in outdoor freshwater microcosms.

This study characterizes the effects of copper (Cu) on Florida apple snails (Pomacea paludosa) and mosquito fish (Gambusia affinis) using a replicated outdoor microcosm design. Soils used in this study were collected from two Cu-enriched citrus agricultural sites in South Florida (Agler property (AGLR) in St. Lucie County and Sunrise Boys property (SRB) in Palm Beach County) and a reference site (Equus property) in St. Lucie County. The study included a 5-week aging phase, an 11 month exposure phase, and a 3 month post-treatment (exposure) phase. The aging phase was initiated by flooding agricultural soils with rainwater in 4 m(3) fiberglass microcosm tanks. Introducing juvenile apple snails (≤7 d old) and mosquito fish (2-3 cm) into the microcosm tanks initiated the exposure phase. Survival, growth, and reproduction of apple snails and fish, and Cu uptake in apple snails, fish, and periphyton were determined in this study. Water chemistry (e.g., dissolved Cu concentration, dissolved organic carbon and dissolved oxygen concentrations, pH, hardness, alkalinity, etc.) was measured daily or weekly during the study. Initial soil Cu concentrations in Equus, SRB, and AGLR microcosms were 7, 55, and 99 mg/kg dw, respectively. Dissolved Cu concentrations in Equus, SRB and AGLR microcosms at the beginning of the study were 3, 82, and 43 μg/L, respectively and decreased to low saturation levels of about ≤9 μg/L Cu after the first 3 months of the study. The decrease of dissolved Cu concentrations was likely due to the dilution of rainwater. Snail and fish mortality appeared to be higher in SRB microcosms than in Equus and AGLR microcosms. There was no significant difference in growth of the snails between treatments. Snail growth data followed the von Bertalanffy Model. The maximum shell length, shell height, and shell width of the snails calculated by the von Bertalanffy Model (L(∞)) were 2.76, 2.05, and 2.18 cm, respectively. The maximum wet weight was 9.38 g. Growth rate (k) of the snails increased in order of shell height (0.459), shell length (0.550), and shell weight (0.598). There was no reproduction in the snails in any treatments including the reference during the exposure phase. However, Cu did not affect reproduction of fish during this period. Copper concentrations in periphyton from Equus, SRB, and AGLR microcosms ranged from 2 to 62, 31 to 371, and 13 to 478 mg/kg, respectively. Copper concentrations in fish at the beginning, days 30 and 150 of the study ranged from 3.19 to 7.53 mg/kg and were not significantly different from the different treatments. Average Cu concentrations in the soft tissue of dead snails from SRB and AGLR microcosms were 4602 mg/kg dw (ranged from 2913 to 8370 mg/kg dw) and 2824 mg/kg dw (ranged from 2118 to 3600 mg/kg dw), respectively. The Cu concentrations in the soft tissue of dead snails found in this study were higher than the tissue Cu concentrations in live aquatic organisms reported in the literature. These high Cu concentrations in edible apple snail soft tissue might pose a risk to Florida apple snail predators, including the snail kite. The post-exposure phase, with snails exposed to only water (i.e., no soils) showed depuration of copper from apple snails and reproduction in all treatments.

Influence of water quality on zinc toxicity to the Florida apple snail (Pomacea paludosa) and sensitivity of freshwater snails to zinc

The present study characterized the influence of water-quality characteristics on zinc (Zn) toxicity to the Florida apple snail (Pomacea paludosa) and the sensitivity of freshwater snails to Zn. Standard 96-h renewal acute toxicity tests were conducted with Zn and juvenile P. paludosa under 3 conditions of pH and alkalinity, water hardness, and dissolved organic carbon (DOC). Median lethal effect concentrations (96-h LC50s), no-observed- effect concentrations, lowest-observed-effect concentrations, LC10s, and LC20s were determined for each test. The results showed that Zn toxicity to P. paludosa decreased linearly with increasing hardness, pH, and DOC. A multiple linear regression model based on pH, hardness, and DOC was able to explain 99% of the observed variability in LC50s. These results are useful for the development of a biotic ligand model (BLM) for P. paludosa and Zn. Zinc acute toxicity data were collected from the literature for 12 freshwater snail species in a wide range of water-quality characteristics for species sensitivity distribution analysis. The results showed that P. paludosa is the second most sensitive to Zn. The present study also suggested that aqueous ZnCO3 and ZnHCO3 (-) can be bioavailable to P. paludosa. Therefore, bioavailability models (e.g., BLM) should take these Zn species into consideration for bioavailability when applied to snails.

Chronic toxicity of binary-metal mixtures of cadmium and zinc to Daphnia magna

The present study characterized the chronic effect of binary-metal mixtures of cadmium (Cd) and zinc (Zn) on Daphnia magna. The titration design was chosen to characterize the 21-d chronic effects of the binary-metal mixtures on survival, growth, reproduction, and metal accumulation in D. magna. Using this design, increasing concentrations of Zn (10, 20, 40, 80, 120, 160, and 200 μg/L) were titrated against a constant concentration of 1.5 μg/L Cd. The results demonstrated that Cd was highly toxic to D. magna. In a mixture with Cd and Zn, sublethal concentrations of 10 and 20 μg/L Zn were insufficient to protect D. magna from chronic Cd toxicity, whereas mixtures containing 40, 80, and 120 μg/L Zn provided strong protective effects to D. magna at all endpoints and resulted in less-than-additive effects. At higher Zn concentrations, such as 160 and 200 μg/L, Zn appeared to contribute to the toxicity. The less-than-additive effects observed in the Cd-Zn mixture can be explained by the decrease in body Cd concentration when the Zn concentration was increased in the exposure media. Embryos analyzed for morphological alterations in the Cd-Zn mixtures demonstrated severe developmental defects. The effect of Cd on undeveloped embryos while both Zn and Cd are present in the organisms raises a question of whether the competitive binding mechanism of Zn and Cd is still happening at the cellular level in the organisms. The results of the present study are useful for development of the biotic ligand model and environmental quality guidelines for metal mixtures. Environ Toxicol Chem 2017;36:2739-2749.