Interactive effects of Louisiana Sweet Crude (LSC) thin oil sheens and ultraviolet light on mortality and swimming behavior of the larval Eastern oyster, Crassostrea virginica.

The Eastern oyster (Crassostrea virginica) is an important commercial bivalve species which also has numerous ecological roles including biogeochemical cycling, providing habitat for larval fish and crustaceans, and reducing the impacts of coastal storms. Oil may pose a threat to oyster larvae swimming in the water column, leading to potential negative effects on survival, growth, and development. Oil toxicity may be further enhanced by chemical changes in the presence of sunlight. This study determined the toxicity of thin oil sheens with and without ultraviolet (UV) light, then examined the latent effects of the short term exposure on longer term survival and swimming ability. Larval C. virginica were exposed to four different oil sheen thicknesses for 24 h with either no UV light or 2-h UV light. Following the exposure, larvae were transferred to clean seawater and no UV light for 96 h. The presence of a 2-h UV light exposure significantly increased oyster mortality, indicating photo-enhanced toxicity. The LC50 for a 24-h oil sheen exposure without UV was 7.26 µm (23 µg/L PAH50) while a 2 h-UV exposure lowered the sheen toxicity threshold to 2.67 µm (10 µg/L PAH50). A previous 24-h oil sheen exposure (≥0.5 µm) led to latent effects on larval oyster survival, regardless of previous UV exposure. Sublethal impacts to larval oyster swimming behavior were also observed from the previous oil sheen exposure combined with UV exposure. This study provides new data for the toxicity of thin oil sheens to a sensitive early life stage of estuarine bivalve.

Improving the design and conduct of aquatic toxicity studies with oils based on 20 years of CROSERF experience.

Laboratory toxicity testing is a key tool used in oil spill science, spill effects assessment, and mitigation strategy decisions to minimize environmental impacts. A major consideration in oil toxicity testing is how to replicate real-world spill conditions, oil types, weathering states, receptor organisms, and modifying environmental factors under laboratory conditions. Oils and petroleum-derived products are comprised of thousands of compounds with different physicochemical and toxicological properties, and this leads to challenges in conducting and interpreting oil toxicity studies. Experimental methods used to mix oils with aqueous test media have been shown to influence the aqueous-phase hydrocarbon composition and concentrations, hydrocarbon phase distribution (i.e., dissolved phase versus in oil droplets), and the stability of oil:water solutions which, in turn, influence the bioavailability and toxicity of the oil containing media. Studies have shown that differences in experimental methods can lead to divergent test results. Therefore, it is imperative to standardize the methods used to prepare oil:water solutions in order to improve the realism and comparability of laboratory tests. The CROSERF methodology, originally published in 2005, was developed as a standardized method to prepare oil:water solutions for testing and evaluating dispersants and dispersed oil. However, it was found equally applicable for use in testing oil-derived petroleum substances. The goals of the current effort were to: (1) build upon two decades of experience to update existing CROSERF guidance for conducting aquatic toxicity tests and (2) to improve the design of laboratory toxicity studies for use in hazard evaluation and development of quantitative effects models that can then be applied in spill assessment. Key experimental design considerations discussed include species selection (standard vs field collected), test substance (single compound vs whole oil), exposure regime (static vs flow-through) and duration, exposure metrics, toxicity endpoints, and quality assurance and control.

Comparative Toxicity of Two Chemical Dispersants and Dispersed Oil in Estuarine Organisms.

Chemical dispersants can be a useful tool to mitigate oil spills. This study examined potential risks to sensitive estuarine species by comparing the toxicity of two dispersants (Corexit® EC9500A and Finasol® OSR 52) individually and in chemically enhanced water-accommodated fractions (CEWAFs) of Louisiana Sweet Crude oil. Acute toxicity thresholds and sublethal biomarker responses were determined in seven species (sheepshead minnow, grass shrimp, mysid, amphipod, polychaete, hard clam, mud snail). Comparing median lethal (LC50) values for the dispersants, Finasol was generally more toxic than Corexit and had greater sublethal toxicity (impaired embryonic hatching, increased lipid peroxidation, decreased acetylcholinesterase activity). The nominal concentration-based mean LC50 for all species tested with Corexit was 150.31 mg/L compared with 43.27 mg/L with Finasol. Comparing the toxicity of the CEWAFs using the nominal concentrations (% CEWAF), Corexit-CEWAFs appeared more toxic than Finasol-CEWAFs; however, when LC50 values were calculated using measured hydrocarbon concentrations, the Finasol-CEWAFs were more toxic. There was greater dispersion efficiency leading to greater hydrocarbon concentrations measured in the Corexit-CEWAF solutions than in equivalent Finasol-CEWAF solutions. The measured concentration-based mean LC50 values for all species tested with Corexit-CEWAF were 261.96 mg/L total extractable hydrocarbons (TEH) and 2.95 mg/L total polycyclic aromatic hydrocarbons (PAH), whereas the mean LC50 values for all species tested with Finasol-CEWAF were 23.19 mg/L TEH and 0.49 mg/L total PAH. Larval life stages were generally more sensitive to dispersants and dispersed oil than adult life stages within a species. These results will help to inform management decisions regarding the use of oil-spill dispersants.

Effects of salinity on oil dispersant toxicity in the grass shrimp, Palaemonetes pugio.

Chemical dispersants can be a useful tool to mitigate oil spills, but the potential risks to sensitive estuarine species should be carefully considered. To improve the decision making process, more information is needed regarding the effects of oil spill dispersants on the health of coastal ecosystems under variable environmental conditions such as salinity. The effects of salinity on the toxicity of two oil dispersants, Corexit® 9500 and Finasol® OSR 52, were examined in this study. Corexit® 9500 was the primary dispersant used during the 2010 Deepwater Horizon oil spill event, while Finasol® OSR 52 is another dispersant approved for oil spill response in the U.S., yet considerably less is known regarding its toxicity to estuarine species. The grass shrimp, Palaemonetes pugio, was used as a model estuarine species. It is a euryhaline species that tolerates salinities from brackish to full strength seawater. Adult and larval life stages were tested with each dispersant at three salinities, 5, 20, and 30 ppt. Median acute lethal toxicity thresholds and oxidative stress responses were determined. The toxicity of both dispersants was significantly influenced by salinity, with greatest toxicity observed at the lowest salinity tested. Larval shrimp were significantly more sensitive than adult shrimp to both dispersants, and both life stages were significantly more sensitive to Finasol than to Corexit. Oxidative stress in adult shrimp, as measured by increased lipid peroxidation activity, occurred with exposure to both dispersants. These data will assist environmental managers in making informed decisions regarding dispersant use in future oil spills.

Exposure of the grass shrimp, Palaemonetes pugio, to antimicrobial compounds affects associated Vibrio bacterial density and development of antibiotic resistance.

Antimicrobial compounds are widespread, emerging contaminants in the aquatic environment and may threaten ecosystem and human health. This study characterized effects of antimicrobial compounds common to human and veterinary medicine, aquaculture, and consumer personal care products [erythromycin (ERY), sulfamethoxazole (SMX), oxytetracycline (OTC), and triclosan (TCS)] in the grass shrimp Palaemonetes pugio. The effects of antimicrobial treatments on grass shrimp mortality and lipid peroxidation activity were measured. The effects of antimicrobial treatments on the bacterial community of the shrimp were then assessed by measuring Vibrio density and testing bacterial isolates for antibiotic resistance. TCS (0.33 mg/L) increased shrimp mortality by 37% and increased lipid peroxidation activity by 63%. A mixture of 0.33 mg/L TCS and 60 mg/L SMX caused a 47% increase in shrimp mortality and an 88% increase in lipid peroxidation activity. Exposure to SMX (30 mg/L or 60 mg/L) alone and to a mixture of SMX/ERY/OTC did not significantly affect shrimp survival or lipid peroxidation activity. Shrimp exposure to 0.33 mg/L TCS increased Vibrio density 350% as compared to the control whereas SMX, the SMX/TCS mixture, and the mixture of SMX/ERY/OTC decreased Vibrio density 78-94%. Increased Vibrio antibiotic resistance was observed for all shrimp antimicrobial treatments except for the mixture of SMX/ERY/OTC. Approximately 87% of grass shrimp Vibrio isolates displayed resistance to TCS in the control treatment suggesting a high level of TCS resistance in environmental Vibrio populations. The presence of TCS in coastal waters may preferentially increase the resistance and abundance of pathogenic bacteria. These results indicate the need for further study into the potential interactions between antimicrobials, aquatic organisms, and associated bacterial communities.

Interactive effects of mosquito control insecticide toxicity, hypoxia, and increased carbon dioxide on larval and juvenile eastern oysters and hard clams.

Mosquito control insecticide use in the coastal zone coincides with the habitat and mariculture operations of commercially and ecologically important shellfish species. Few data are available regarding insecticide toxicity to shellfish early life stages, and potential interactions with abiotic stressors, such as low oxygen and increased CO2 (low pH), are less understood. Toxicity was assessed at 4 and 21 days for larval and juvenile stages of the Eastern oyster, Crassostrea virginica, and the hard clam, Mercenaria mercenaria, using two pyrethroids (resmethrin and permethrin), an organophosphate (naled), and a juvenile growth hormone mimic (methoprene). Acute toxicity (4-day LC50) values ranged from 1.59 to >10 mg/L. Overall, clams were more susceptible to mosquito control insecticides than oysters. Naled was the most toxic compound in oyster larvae, whereas resmethrin was the most toxic compound in clam larvae. Mortality for both species generally increased with chronic insecticide exposure (21-day LC50 values ranged from 0.60 to 9.49 mg/L). Insecticide exposure also caused sublethal effects, including decreased swimming activity after 4 days in larval oysters (4-day EC50 values of 0.60 to 2.33 mg/L) and decreased growth (shell area and weight) in juvenile clams and oysters after 21 days (detected at concentrations ranging from 0.625 to 10 mg/L). Hypoxia, hypercapnia, and a combination of hypoxia and hypercapnia caused mortality in larval clams and increased resmethrin toxicity. These data will benefit both shellfish mariculture operations and environmental resource agencies as they manage the use of mosquito control insecticides near coastal ecosystems.

Effects of chemically spiked sediments on estuarine benthic communities: a controlled mesocosm study.

Ambient sediments were collected from a reference site in the North Edisto River, SC and transferred to a laboratory facility to investigate effects of chemical contaminants on estuarine infaunal communities under controlled mesocosm conditions. Sediment contaminant slurries were prepared using dried sediments collected from the reference site and spiked with a metal (copper), a polycyclic aromatic hydrocarbon (pyrene), and a pesticide (4,4(')-dichlorodiphenyltrichloroethane) to yield nominal mean effects range-median (ERM) quotients of <0.01 (no addition), 0.1, and 1.0 and applied to control, low dose (TRT A), and high dose (TRT B) treatment groups, respectively. Sediment samples for contaminant and benthic analyses were collected at the start of the experiment, 1 month after dosing, and 3 months after dosing. Near-nominal mean ERM quotients of 0.001, 0.075, and 0.818 were measured initially after dosing and remained fairly constant throughout the experiment. Measures of benthic condition, diversity, and richness were significantly reduced in both treatment groups relative to the control 1 month after dosing and persisted in TRT B at 3 months. The results demonstrate that benthic community effects can be observed at mean ERM quotients that are about an order of magnitude lower than levels that have been shown to be associated with significant toxicity in acute laboratory bioassays with single species (e.g., amphipods) in other studies.

Effects of the antifouling compound, Irgarol 1051, on a simulated estuarine salt marsh ecosystem.

Toxicity effects of the antifouling compound, Irgarol 1051, were examined using a simulated estuarine salt marsh ecosystem. The 35 day mesocosm exposure incorporated tidal flux and contained seawater, sediments, marsh grass, and estuarine biota. Irgarol (10.0 microg/l) caused a significant reduction in phytoplankton biomass and primary productivity. HPLC pigment analysis indicated significant effects of irgarol on both phytoplankton and periphyton community composition, with decreased concentrations of pigments representative of diatom species. There was also a significant decrease in dissolved oxygen levels in the 10.0 microg/l irgarol treatment. Growth of the hard shell clam was significantly reduced in the 1.0 and 10.0 microg/l irgarol treatments. The effects observed occurred at irgarol concentrations greater than those typically measured in the environment. Prolonged exposure, the accumulation of irgarol in sediments, plant, or animal tissues, and the interaction of irgarol with other chemicals in the environment; however, could increase risk.

Toxicity of the antimicrobial compound triclosan and formation of the metabolite methyl-triclosan in estuarine systems.

Triclosan, a commonly used antimicrobial compound, has been measured in aquatic systems worldwide. This study exposed marine species to triclosan to examine effects primarily on survival and to investigate the formation of the degradation product, methyl-triclosan, in the estuarine environment. Acute toxicity was assessed using the bacterium Vibrio fischeri, the phytoplankton species Dunaliella tertiolecta, and three life stages of the grass shrimp Palaemonetes pugio. P. pugio larvae were more sensitive to triclosan than adult shrimp or embryos. Acute aqueous toxicity values (96 h LC50) were 305 microg/L for adult shrimp, 154 microg/L for larvae, and 651 microg/L for embryos. The presence of sediment decreased triclosan toxicity in adult shrimp (24 h LC50s were 620 microg/L with sediment, and 482 microg/L without sediment). The bacterium was more sensitive to triclosan than the grass shrimp, with a 15 min aqueous IC50 value of 53 microg/L and a 15 min spiked sediment IC50 value of 616 microg/kg. The phytoplankton species was the most sensitive species tested, with a 96 h EC50 value of 3.55 microg/L. Adult grass shrimp were found to accumulate methyl-triclosan after a 14-day exposure to 100 microg/L triclosan, indicating formation of this metabolite in a seawater environment and its potential to bioaccumulate in higher organisms. Triclosan was detected in limited surface water sampling of Charleston Harbor, SC at a maximum concentration of 0.001 microg/L, substantially lower than the determined toxicity values. These findings suggest triclosan poses low acute toxicity risk to estuarine organisms; however, the potential for chronic, sublethal, and metabolite effects should be investigated.

Effects of the insecticide permethrin on three life stages of the grass shrimp, Palaemonetes pugio.

Toxicity of the pyrethroid insecticide permethrin was assessed using three life stages of the estuarine grass shrimp, Palaemonetes pugio. Adult and larval shrimp were tested with and without sediment. An aqueous embryo test was also conducted. Cellular stress biomarkers, glutathione, and lipid peroxidation, were assessed. Larval shrimp were the most sensitive life stage with a 96-h lethal concentration (LC(50)) value of 0.05 microg/L, compared to 0.25 microg/L for adults, and 6.4 microg/L for embryos. The presence of sediment significantly decreased toxicity of permethrin to both adult and larval shrimp. Permethrin exposure increased time to hatch in embryos and decreased swimming behavior of larvae. Lipid peroxidation levels were significantly decreased in the adult shrimp, but increased in larval shrimp exposed to permethrin. Low levels of permethrin may negatively affect grass shrimp health and survival. Permethrin use in the coastal zone should be carefully managed to avoid adverse impacts on nontarget estuarine organisms.

Preliminary risk assessment of the lipid-regulating pharmaceutical clofibric acid, for three estuarine species.

Clofibric acid is the active metabolite of several fibrate drugs prescribed to reduce blood cholesterol levels. It is persistent and widely detected in the environment. Clofibric acid toxicity was assessed using three estuarine organisms: an alga (Dunaliella tertiolecta), a crustacean (Palaemonetes pugio), and a fish (Fundulus heteroclitus). Mortality and sublethal physiological responses (protein, lipid, cholesterol, and cytochrome P450 levels) were examined. Clofibric acid did not significantly affect cell density or growth rate of D. tertiolecta (concentrations 1000 microg/L). Survival of P. pugio and F. heteroclitus were also unaffected at clofibric acid concentrations 1000 microg/L. In addition, no significant changes in the sublethal test endpoints were found. An additional chronic (17-day) exposure of F. heteroclitus to clofibric acid (10 microg/L) was conducted and found no effects on survival or sublethal endpoints. The rabbit polyclonal CYP450 4A antibody did cross react with F. heteroclitus, demonstrating that a CYP4A-like isoform is present in this teleost species and may be used in future induction studies. Clofibric acid, however, did not alter CYP4A levels in F. heteroclitus. Measured concentrations in the environment have not exceeded 10 microg/L. Therefore, the results of this study indicate a low risk of adverse effects from environmental exposure to clofibric acid for the species tested.

The effects of the contemporary-use insecticide (fipronil) in an estuarine mesocosm.

To examine the effects of environmentally realistic fipronil concentrations on estuarine ecosystems, replicated mesocosms containing intact marsh plots and seawater were exposed to three treatments of fipronil (150, 355, and 5000 ng/L) and a Control. Juvenile fish (Cyprinidon variegatus), juvenile clams (Mercenaria mercenaria), oysters (Crassostrea virginica), and grass shrimp (Palaemonetes pugio) were added prior to fipronil in an effort to quantify survival, growth, and the persistence of toxicity during the planned 28-day exposure. Results indicated that there were no fipronil-associated effects on the clams, oysters, or fish. Shrimp were sensitive to the highest two concentrations (40% survival at 355 ng/L and 0% survival at 5000 ng/L). Additionally, the highest fipronil treatment (5000 ng/L) was toxic to shrimp for 6 weeks post dose. These results suggest that fipronil may impact shrimp populations at low concentrations and further use in coastal areas should be carefully assessed.

Toxicity and bioconcentration potential of the agricultural pesticide endosulfan in phytoplankton and zooplankton.

Agricultural pesticide runoff in southeastern coastal regions of the United States is a critical issue. Bioconcentration of pesticides by phytoplankton and zooplankton at the base of the aquatic food web may increase the persistence of pesticides in aquatic ecosystems and cause effects at higher trophic levels. This study examined the toxicity of a widely used agricultural pesticide, endosulfan, to Pseudokirchneriella subcapitatum (freshwater green alga) and Daphnia magna (freshwater cladoceran). We then investigated the potential of both plankton species to sequester endosulfan from their surrounding media. We also assessed the degree to which endosulfan is accumulated by D. magna via food (endosulfan-contaminated P. subcapitatum). A 96-h growth rate EC50 of 427.80 microg/L endosulfan was determined for P. subcapitatum, whereas a 24-h immobilization EC50 of 366.33 microg/L endosulfan was determined for D. magna. The 5-h EC50s for filtration and ingestion in D. magna were 165.57 microg/L and 166.44 microg/L, respectively. An average bioconcentration factor (BCF) of 2,682 was determined for P. subcapitatum exposed to 100 microg/L endosulfan for 16 h. An average BCF of 3,278 was determined for D. magna in a 100 microg/L endosulfan water-only exposure. There was negligible uptake of endosulfan by D. magna feeding on contaminated algae in clean water (BCF approximately 0). Different proportions of parent isomers (endosulfan I and II) and the primary degradation product (endosulfan sulfate) were detected among treatments. Endosulfan was rapidly accumulated and concentrated from water by P. subcapitatum and D. magna neonates. Endosulfan contained in phytoplankton, however, was not bioaccumulated by zooplankton. These findings may prove useful in assessing ecosystem risk, because uptake from the water column appears to be the dominant route for bioconcentration of endosulfan by zooplankton.

Toxicity of pesticides to aquatic microorganisms: a review.

Microorganisms contribute significantly to primary production, nutrient cycling, and decomposition in estuarine eco-systems; therefore, detrimental effects of pesticides on microbial species may have subsequent impacts on higher trophic levels. Pesticides may affect estuarine microorganisms via spills, runoff, and drift. Both the structure and the function of microbial communities may be impaired by pesticide toxicity. Pesticides may also be metabolized or bioaccumulated by microorganisms. Mechanisms of toxicity vary, depending on the type of pesticide and the microbial species exposed. Herbicides are generally most toxic to phototrophic microorganisms, exhibiting toxicity by disrupting photosynthesis. Atrazine is the most widely used and most extensively studied herbicide. Toxic effects of organophosphate and organochlorine insecticides on microbial species have also been demonstrated, although their mechanisms of toxicity in such nontarget species remain unclear. There is a great deal of variability in the toxicity of even a single pesticide among microbial species. When attempting to predict the toxicity of pesticides in estuarine ecosystems, effects of pesticide mixtures and interactions with nutrients should be considered. The toxicity of pesticides to aquatic microorganisms, especially bacteria and protozoa, is an area of research requiring further study.

Use of Metabolic Inhibitors to Characterize Ecological Interactions in an Estuarine Microbial Food Web.

Understanding microbial food web dynamics is complicated by the multitude of competitive or interdependent trophic interactions involved in material and energy flow. Metabolic inhibitors can be used to gain information on the relative importance of trophic pathways by uncoupling selected microbial components and examining the net effect on ecosystem structure and function. A eukaryotic growth inhibitor (cycloheximide), a prokaryotic growth inhibitor (antibiotic mixture), and an inhibitor of photosynthesis (DCMU) were used to examine the trophodynamics of microbial communities from the tidal creek in North Inlet, a salt marsh estuary near Georgetown, South Carolina. Natural microbial communities were collected in the spring, summer, and fall after colonization onto polyurethane foam substrates deployed in the tidal creek. Bacterial abundance and productivity, heterotrophic ciliate and flagellate abundance, and phototrophic productivity, biomass, and biovolume were measured at five time points after inhibitor additions. The trophic responses of the estuarine microbial food web to metabolic inhibitors varied with season. In the summer, a close interdependency among phototrophs, bacteria, and protozoa was indicated, and the important influence of microzooplanktonic nutrient recycling was evident (i.e., a positive feedback loop). In the fall, phototroph and bacteria interactions were competitive rather than interdependent, and grazer nutrient regeneration did not appear to be an important regulatory factor for bacterial or phototrophic activities. The results indicate a seasonal shift in microbial food web structure and function in North Inlet, from a summer community characterized by microbial loop dynamics to a more linear trophic system in the fall. This study stresses the important role of microbial loops in driving primary and secondary production in estuaries such as North Inlet that are tidally dominated by fluctuations in nutrient supply and a summer phytoplankton bloom.