New insights into pesticide occurrence and multicompartmental monitoring strategies in stream ecosystems using periphyton and suspended sediment.

Streams are susceptible to pesticide pollutants which are transported outside of the intended area of application from surrounding agricultural fields. It is essential to monitor the occurrence and levels of pesticides in aquatic ecosystems to comprehend their effects on the aquatic environment. The common sampling strategy used for monitoring pesticides in stream ecosystems is through the collection and analysis of grab water samples. However, grab water sampling may not effectively monitor pesticides due to its limited ability to capture temporal and spatial variability, potentially missing fluctuations and uneven distribution of pesticides in aquatic environments. Monitoring using periphyton and sediment sampling may offer a more comprehensive approach by accounting for accumulative processes and temporal variations. Periphyton are a collective of microorganisms that grow on hard surfaces in aquatic ecosystems. They are responsive to chemical and biological changes in the environment, and therefore have the potential to act as a cost-effective, integrated sampling tool to monitor pesticide exposures in aquatic ecosystems. The objective of this study was to assess pesticides detected through periphyton, suspended sediment, and conventional grab water sampling methods and identify the matrix that offers a more comprehensive characterization of a stream's pesticide exposure profile. Ten streams across Southern Ontario were sampled in 2021 and 2022. At each stream site, water, sediment and periphyton, colonizing both artificial and natural substrates, were collected and analyzed for the presence of ~500 pesticides. Each of the three matrices detected distinctive pesticide exposure profiles. The frequency of detection in periphyton, sediment and water matrices were related to pesticides' log Kow and log Koc (P < 0.05). In addition, periphyton bioconcentrated 22 pesticides above levels observed in the ambient water. The bioconcentration factors of pesticides in periphyton can be predicted from their log Kow (simple linear regressions, P < 0.05). The results demonstrate that sediment and periphyton accumulate pesticides in stream environments. This highlights the importance of monitoring pesticide exposure using these matrices to ensure a complete and comprehensive characterization of exposure in stream ecosystems.

An Assessment of the Toxicity of Pesticide Mixtures in Periphyton from Agricultural Streams to the Mayfly Neocloeon triangulifer.

Residual concentrations of pesticides are commonly found outside the intended area of application in Ontario's surface waters. Periphyton are a vital dietary component for grazing organisms in aquatic ecosystems but can also accumulate substantial levels of pesticides from the surrounding water. Consequently, grazing aquatic organisms are likely subjected to pesticide exposure through the consumption of pesticide-contaminated periphyton. The objectives of the present study were to determine if pesticides partition into periphyton in riverine environments across southern Ontario and, if so, to determine the toxicity of pesticides in periphyton when fed to the grazing mayfly Neocloeon triangulifer. Sites with low, medium, and high pesticide exposure based on historic water quality monitoring data were selected to incorporate a pesticide exposure gradient into the study design. Artificial substrate samplers were utilized to colonize periphyton in situ, which were then analyzed for the presence of approximately 500 pesticides. The results demonstrate that periphyton are capable of accumulating pesticides in agricultural streams. A novel 7-day toxicity test method was created to investigate the effects of pesticides partitioned into periphyton when fed to N. triangulifer. Periphyton collected from the field sites were fed to N. triangulifer and survival and biomass production recorded. Survival and biomass production significantly decreased when fed periphyton colonized in streams with catchments dominated by agricultural land use (p < 0.05). However, the relationship between pesticide concentration and survival or biomass production was not consistent. Using field-colonized periphyton allowed us to assess the dietary toxicity of environmentally relevant concentrations of pesticide mixtures; however, nutrition and taxonomic composition of the periphyton may vary between sites. Environ Toxicol Chem 2023;42:2143-2157. © 2023 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Comparison of Established and Novel Insecticides on Survival and Reproduction of Folsomia candida.

Neonicotinoids have been among the most widely and abundantly used insecticides for most of the current century. The effects of these substances on nontarget terrestrial and aquatic organisms have resulted in a significant decrease in their use in many parts of the world. In response, the application of several novel classes of insecticides including diamides, ketoenols, pyridines, and butenolides has significantly increased. The hexapod subclass Collembola is an ecologically significant and widely distributed group of soil invertebrates often found in leaf litter and in surficial soils. We exposed the parthenogenic collembolan species Folsomia candida to six insecticides in a sandy loam soil for 28 days, including two neonicotinoids (thiamethoxam and clothianidin), a diamide (cyantraniliprole), a ketoenol (spirotetramat), a pyridine (flonicamid), and a butanolide (flupyradifurone) to assess the effect of each insecticide on survival and reproduction. Clothianidin, thiamethoxam, and cyantraniliprole (median effective concentration [EC50] values for reproduction: 0.19, 0.38, and 0.49 mg/kg soil, respectively) had a greater effect on survival and reproduction of F. candida than flupyradifurone, spirotetramat, and flonicamid (EC50 values for reproduction: 0.73, >3.08, and 5.20 mg/kg soil, respectively). All significant impacts found in our study were observed at concentrations below concentrations of the active ingredients that would be expected in agricultural soils. Environ Toxicol Chem 2023;42:1516-1528. © 2023 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Cyanotoxins accumulate in Lake St. Clair fish yet their fillets are safe to eat.

Consuming fish exposed to cyanobacterial harmful algal blooms (HABs) may be a major route of microcystin toxin exposure to humans. However, it remains unknown whether fish can accumulate and retain microcystins temporally in waterbodies with recurring seasonal HABs, particularly before and after a HAB event when fishing is active. We conducted a field study on Largemouth Bass, Northern Pike, Smallmouth Bass, Rock Bass, Walleye, White Bass, and Yellow Perch to assess the human health risks to microcystin toxicity via fish consumption. We collected 124 fish in 2016 and 2018 from Lake St. Clair, a large freshwater ecosystem in the North American Great Lakes that is actively fished pre- and post-HAB periods. Muscles were analyzed using the 2-methyl-3-methoxy-4-phenylbutyric acid (MMPB) Lemieux Oxidation method for total microcystins, which was used to perform a human health risk assessment for comparison against fish consumption advisory benchmarks available for Lake St. Clair. From this collection 35 fish livers were additionally extracted to confirm the presence of microcystins. Microcystins were detected in all livers at widely varying concentrations (1-1500 ng g-1 ww), suggesting HABs are an underappreciated and pervasive stressor to fish populations. Conversely, microcystin levels were consistently low in muscles (0-15 ng g-1 ww) and presented negligible risk, empirically supporting that fillets may be safely consumed before and after HAB events following fish consumption advisories.

Lake Erie fish safe to eat yet afflicted by algal hepatotoxins.

Toxic harmful algal blooms (HABs) pose serious threats to human health and instances of wildlife death have been documented across taxa. However, the extent of toxicological impacts on wildlife species is largely unresolved, raising uncertainty about the repercussions of increasingly severe HABs on the biodiversity and functioning of aquatic ecosystems. Here, we conducted a field study to assess human health risks from consuming fish caught across all stages of a HAB and to determine the pervasiveness of potentially harmful levels of the cosmopolitan toxin microcystin on fish populations. We collected 190 fish in 2015 and 2017 from Lake Erie, a large freshwater ecosystem that is highly productive for fisheries and is an epicenter of HABs and microcystin toxicity events. Fish muscles and livers were analyzed for total microcystins, which was used to conduct a human health risk assessment for comparison against fish consumption advisory benchmarks available for Lake Erie. We found microcystins pose low risks to human health from fillet consumption (mean 1.80 ng g-1 ww) but substantial risks to fish health and recruitment from liver concentrations measured well before and after seasonal bloom events (mean 460.13 ng g-1 ww). Our findings indicate HABs are a previously underappreciated but pervasive threat to fish populations.

Are There Longitudinal Effects of Forest Harvesting on Carbon Quality and Flow and Methylmercury Bioaccumulation in Primary Consumers of Temperate Stream Networks?

Forest harvesting affects dissolved organic matter (DOM) and aqueous mercury inputs as well as the food web structure in small-headwater streams, but how these upstream changes manifest downstream is unclear. To address this uncertainty, we examined DOM quality, autochthony in the caddisfly Hydropsychidae (using δ2 H), and methylmercury (MeHg) concentrations in stream water and the caddisfly along a longitudinal gradient (first- to fourth-order streams, subcatchments of 50-1900 ha) in paired partially harvested and reference catchments in central Ontario, Canada. Although measures of DOM quality (specific ultraviolet absorbance at 254 nm 2.20-11.62) and autochthony in caddisflies (4.9%-34.0%) varied among sites, no upstream-to-downstream differences in these measures were observed between the paired harvested and reference catchments. In contrast, MeHg levels in stream water (0.06-0.35 ng/L) and caddisflies (29.7-192 µg/kg dry wt) were significantly higher in the upstream sites but not the farthest downstream sites in the harvested catchments compared to the reference catchments. This suggests that while current mitigation measures used by forestry companies did not prevent elevated MeHg in water and invertebrates at smaller spatial scales (subcatchments of 50-400 ha), these upstream impacts did not manifest at larger spatial scales (subcatchments of 800-1900 ha). The present study advances our understanding of spatially cumulative impacts within harvested catchments, which is critical to help forest managers maintain healthy forest streams and their provisioning of aquatic ecosystem services. Environ Toxicol Chem 2022;41:1490-1507. © 2022 SETAC.

Fish tissue accumulation and proteomic response to microcystins is species-dependent.

Cyanotoxins including microcystins are increasing globally, escalating health risks to humans and wildlife. Freshwater fish can accumulate and retain microcystins in tissues; however, uptake and depuration studies thus far have not exposed fish to microcystins in its intracellular state (i.e., cell-bound or conserved within cyanobacteria), which is a primary route of exposure in the field, nor have they investigated sublethal molecular-level effects in tissues, limiting our knowledge of proteins responsible for microcystin toxicity pathways in pre-to-postsenescent stages of a harmful algal bloom. We address these gaps with a 2-wk study (1 wk of 'uptake' exposure to intracellular microcystins (0-40 µg L-1) produced by Microcystis aeruginosa followed by 1 wk of 'depuration' in clean water) using Rainbow Trout (Oncorhynchus mykiss) and Lake Trout (Salvelinus namaycush). Liver and muscle samples were collected throughout uptake and depuration phases for targeted microcystin quantification and nontargeted proteomics. For both species, microcystins accumulated at a higher concentration in the liver than muscle, and activated cellular responses related to oxidative stress, apoptosis, DNA repair, and carcinogenicity. However, intraspecific proteomic effects between Rainbow Trout and Lake Trout differed, and interspecific accumulation and retention of microcystins in tissues within each species also differed. We demonstrate that fish do not respond the same to cyanobacterial toxicity within and among species despite being reared in the same environment and diet.

The sustainable agriculture imperative: A perspective on the need for an agrosystem approach to meet the United Nations Sustainable Development Goals by 2030.

The development of modern, industrial agriculture and its high input-high output carbon energy model is rendering agricultural landscapes less resilient. The expected continued increase in the frequency and intensity of extreme weather events, in conjunction with declining soil health and biodiversity losses, could make food more expensive to produce. The United Nations has called for global action by establishing 17 sustainable development goals (SDGs), four of which are linked to food production and security: declining biodiversity (SDG 15), loss of ecosystem services and agroecosystem stability caused by increasing stress from food production intensification and climate change (SDG 13), declining soil health caused by agricultural practices (SDGs 2 and 6), and dependence on synthetic fertilizers and pesticides to maintain high productivity (SDG 2). To achieve these SDGs, the agriculture sector must take a leading role in reversing the many negative environmental trends apparent in today's agricultural landscapes to ensure that they will adapt and be resilient to climate change in 2030 and beyond. This will demand fundamental changes in how we practice agriculture from an environmental standpoint. Here, we present a perspective focused on the implementation of an agrosystem approach, which we define to promote regenerative agriculture, an integrative approach that provides greater resilience to a changing climate, reverses biodiversity loss, and improves soil health; honors Indigenous ways of knowing and a holistic approach to living off and learning from the land; and supports the establishment of emerging circular economies and community well-being. Integr Environ Assess Manag 2022;18:1199-1205. © 2021 SETAC.

Cyanotoxins within and Outside of Microcystis aeruginosa Cause Adverse Effects in Rainbow Trout (Oncorhynchus mykiss).

The global expansion of toxic Microcystis blooms, and production of cyanotoxins including microcystins, are an increasing risk to freshwater fish. Differentiating intracellular and extracellular microcystin toxicity pathways (i.e., within and outside of cyanobacterial cells) in fish is necessary to assess the severity of risks to populations that encounter harmful algal blooms in pre-to-postsenescent stages. To address this, adult and juvenile Rainbow Trout (Oncorhynchus mykiss) were, respectively, exposed for 96 h to intracellular and extracellular microcystins (0, 20, and 100 µg L-1) produced by Microcystis aeruginosa. Fish were dissected at 24 h intervals for histopathology, targeted microcystin quantification, and nontargeted proteomics. Rainbow Trout accumulated intracellular and extracellular microcystins in all tissues within 24 h, with greater accumulation in the extracellular state. Proteomics revealed intracellular and extracellular microcystins caused sublethal toxicity by significantly dysregulating proteins linked to the cytoskeletal structure, stress responses, and DNA repair in all tissues. Pyruvate metabolism in livers, anion binding in kidneys, and myopathy in muscles were also significantly impacted. Histopathology corroborated these findings with evidence of necrosis, apoptosis, and hemorrhage at similar severity in both microcystin treatments. We demonstrate that sublethal concentrations of intracellular and extracellular microcystins cause adverse effects in Rainbow Trout after short-term exposure.

Deterministic risk assessment of firefighting water additives to terrestrial organisms.

Firefighting water additives are used to increase the rate at which fires can be extinguished. The majority of ecotoxicological research has focused on firefighting formulations containing perfluorinated compounds as additives, due to the persistence and bioaccumulative nature of the perfluorinated constituents. A number of relatively new additives have come on the market to replace the products containing perfluorinated compounds. The potential effect of these new additives on the environment has been largely unstudied. This study investigated the toxicity of six firefighting water additives: Eco-Gel™, ThermoGel 200L™, FireAde™, Fire-Brake™, Novacool Foam™, and F-500™ to terrestrial biota. Terrestrial organisms could be exposed to firefighting water additives through leaching into soil and/or runoff following a firefighting event or through direct aerial application during a forest fire. Toxicity to three plant species was assessed through seedling germination and emergence tests: Fagopyrum esculentum (buckwheat), Raphanus raphanistrum subsp. sativus (radish), and Rudbeckia hirta (black-eyed Susan). The effects of firefighting water additives on three soil invertebrates, the collembolan Folsomia candida, the earthworms Eisenia andrei, and Dendrodrilus rubidus, were also investigated using static acute tests to estimate EC50/LC50s. The concentration that resulted in a 50% reduction in survival (LC50) for the acute toxicity tests conducted with F. candida ranged from 3 (Eco-Gel) to 0.175% (Novacool) by volume. Comparatively, the acute toxicity of two firefighting water additives to D. rubidus could not be determined, as a 50% reduction in survival was not observed. A number of firefighting water additives were found to pose a hazard to terrestrial organisms based on a worst-case exposure scenario of direct application at the greatest recommended application rate for a class A fire (e.g., wood, paper). The firefighting water additive F-500 was found to pose a hazard (HQ ≥ 1) for all species tested, except for the acute test conducted with D. rubidus. Comparatively, Eco-Gel posed a hazard for only the acute and chronic tests with F. candida. This study represents the first comparative deterministic risk assessment of firefighting water additives to terrestrial ecosystems.

Optimization of an MMPB Lemieux Oxidation method for the quantitative analysis of microcystins in fish tissue by LC-QTOF MS.

Microcystins are toxic heptapeptides produced by cyanobacteria in marine and freshwater environments. In biological samples such as fish, microcystins can be found in the free form or covalently bound to protein phosphatases type I and II. Total microcystins in fish have been quantified in the past using the Lemieux Oxidation approach, where all toxins are oxidated to a common fragment (2-methyl-3-methoxy-4-phenylbutyric acid, MMPB) regardless of their initial amino acid configuration or form (free or protein bound). These studies have been carried out using different experimental conditions and employed different quantification strategies. The present study has further investigated the oxidation step using a systematic approach, to identify the most important factors leading to a higher, more robust MMPB generation yield from fish tissue in order to reduce the method detection limit. Field samples were quantified using an in-situ generated MMPB matrix matched calibration curve by isotope dilution with d3-MMPB via liquid chromatography coupled to time-of-flight mass spectrometry (LC-QTOF MS). This approach improves method's accuracy by taking into account of potential matrix effects that could affect the derivatization, sample prepation and instrumental analysis steps. The validated method showed 16.7% precision (RSD) and +6.7% accuracy (bias), with calculated method detection limits of 7.28 ng g-1 Performance of the method was assessed with the analysis of laboratory exposed Rainbow Trout (Oncorhynchus mykiss) to cyanobacteria as a positive control, where no microcystins were detected in the pre-exposure fish liver and fillet, low levels in the exposed fillet (65.0 ng g-1) and higher levels in the exposed liver (696 ng g-1). Finally, the method was employed for the analysis of 26 fillets (muscle) and livers of Walleye (Sander vitreus) and Yellow Perch (Perca flavescens) from Lake Erie, showing very low concentrations of microcystins in the fillet and higher concentrations in liver, up to 3720 ng g-1.

Shotgun proteomics analysis reveals sub-lethal effects in Daphnia magna exposed to cell-bound microcystins produced by Microcystis aeruginosa.

Microcystins that are cell-bound within Microcystis have demonstrated the ability to cause lethal and reproductive impairment in Daphnia, who constitute an important part of aquatic food chains and are known to feed on viable cyanobacterial cells. Recent advances in environmental toxicogenomics can be used to better understand the mechanistic effects from exposure to cell-bound microcystins in Daphnia; however, there remains a need to examine the effects of microcystins exposure as a function of dose and time in order to help elucidate the progression of (sub-)lethal effects. This study examines the effects of cell-bound microcystin exposure in Daphnia magna as a function of dose and time with shotgun proteomics in order to measure and provide insightful evidence describing functional mechanisms from, and relationships between, protein populations in response to toxic Microcystis aeruginosa. We further characterize the life-history fitness of D. magna in the presence of toxic exposure by measuring somatic growth rate. Chronic dietary exposure to cell-bound microcystins reduced the somatic growth rate of D. magna. Through proteomics analysis, we identified a significant increase in abundance of proteins related to reproductive success and development, removal of superoxide radicals, and motor activity in D. magna parents exposed to cell-bound microcystins at sub-lethal concentrations. We also identified a significant decrease in abundance of proteins related to apoptosis, metabolism, DNA damage repair, and immunity in D. magna neonates. This information will improve our understanding of the risks posed by cell-bound microcystins to cladocerans in freshwater ecosystems.

Forest management influences the effects of streamside wet areas on stream ecosystems.

Riparian zones contain areas of strong hydrological connectivity between land and stream, referred to as variable source areas (VSAs), and are considered biogeochemical control points. However, little is known about whether VSAs influence stream communities and whether this connectivity is affected by forest management. To address this, we used multiple biotic and abiotic indicators to (1) examine the influence of VSAs on riparian vegetation and stream ecosystems by comparing VSA and non-VSA reaches and (2) explore how forest management may affect the influence of VSAs on stream ecosystems. We detected some significant differences between VSA and non-VSA reaches in the riparian vegetation (greater understory and lower tree density) and stream ecosystem indicators (greater dissolved organic matter aromaticity, microbial biomass, peroxidase activity and collector-gatherer density, and lower dissolved organic carbon concentrations, algal biomass, and predatory macroinvertebrate density), which suggests that VSAs may create a more heterotrophic ecosystem locally. However, we show some evidence that forest management activities (specifically, road density) can alter the influence of VSAs and eliminate the differences observed at lower forest management intensities, and that the most hydrologically connected areas seem more sensitive to disturbance. Therefore, we suggest that the heterogeneity in hydrological connectivity along riparian zones should be considered when planning forest harvesting operations and road building (e.g., wider riparian buffers around VSAs).

Assessing the toxicity of cell-bound microcystins on freshwater pelagic and benthic invertebrates.

Cyanobacterial harmful algal blooms dominated by Microcystis frequently produce microcystins, a family of toxins capable of inflicting harm to pelagic and benthic freshwater invertebrates. Research on the effect of microcystins on invertebrates is inconclusive; from one perspective, studies suggest invertebrates can coexist in toxic blooms; however, studies have also measured negative food-associated effects from microcystins. To test the latter perspective, we examined the reproduction, growth, and survival of laboratory-cultured Ceriodaphnia dubia, Daphnia magna, and Hexagenia spp. exposed to cell-bound microcystins through a series of life-cycle bioassays. Test organisms were exposed to a concentration gradient ranging from 0.5 µg L-1 to 300 µg L-1 microcystins, which corresponds to values typically found in freshwaters during bloom season. Lethal concentrations in C. dubia (LC50 = 5.53 µg L-1) and D. magna (LC50 = 85.72 µg L-1) exposed to microcystins were among the lowest recorded to date, and reproductive effects were observed at concentrations as low as 2.5 µg L-1. Length of D. magna was significantly impacted in microcystin treatments great than 2.5 µg L-1. No lethality or growth impairments were observed in Hexagenia. This information will improve our understanding of the risks posed by microcystins to food webs in freshwaters.

An efficient and affordable laboratory method to produce and sustain high concentrations of microcystins by Microcystis aeruginosa.

Microcystis aeruginosa is a cosmopolitan cyanobacteria that continues to jeopardize freshwater ecosystem services by releasing the hepatotoxin microcystin, which can, in some cases, cause death to aquatic fauna and even humans. Currently, our abilities to understand the mechanisms of microcystin toxicology are limited by the lack of a method for producing high concentrations, which are central to large-scale and long-term research in natural systems. Here we present an efficient and affordable laboratory method to produce high concentrations of microcystins by a toxigenic strain of M. aeruginosa. Through batch culture studies, we yielded microcystins at concentrations that are environmentally relevant to freshwaters around the world (1-300 µg L-1), maintained these concentrations without resupplying fresh medium (further reducing costs), and utilized rate equations to model the relationship between the environmental conditions in the cultures and changes occurring within the M. aeruginosa cells. Our assessment suggests that steady production of microcystins depends on the availability of carbon throughout the experiment. Hence, we recommend the use of tissue culture treated flasks with a vented cap to ensure the production of microcystins is uninterrupted. This method demonstrates that microcystins can be produced in the laboratory at concentrations relevant to freshwater ecosystems. •The method demonstrates M. aeruginosa CPCC 300 is a reliable strain of freshwater cyanobacteria that can yield microcystins at environmentally relevant concentrations.•Validation showed M. aeruginosa CPCC 300 is resilient in carbon-limited situations and may respond to stress by shifting the ratio of microcystin congeners.•Cell culture flasks with vented caps -filled no more than 50 % of the flask volume to allow for sufficient air exchange- are an excellent and cost-effective approach to maintaining cell growth and producing microcystins at a range between 300 to 1200 µg L-1.

Acute Effects of Binary Mixtures of Imidacloprid and Tebuconazole on 4 Freshwater Invertebrates.

Receiving waters from agricultural areas can contain multiple pesticides such as the neonicotinoid imidacloprid and the fungicide tebuconazole, leading to the potential for aquatic life to be exposed to such mixtures. In the present study, the effects of tebuconazole were tested alone and in binary mixtures with imidacloprid on 4 aquatic invertebrates: Chironomus dilutus, Hyalella azteca, Lumbriculus variegatus, and Neocloeon triangulifer. Acute (96-h) median lethal concentrations (LC50s) were derived for individual compounds and used to design a binary mixture study to determine cumulative effects. The LC50s showed that imidacloprid was more potent than tebuconazole by 1 to 3 orders of magnitude for the 4 species. Lethality data from mixture experiments were analyzed using MIXTOX to determine deviations from independent action, followed by the model deviation ratio (MDR) technique to determine the biological significance and reproducibility of observed mixture effects. MIXTOX showed that the cumulative toxicities of imidacloprid-tebuconazole differed between the species: for C. dilutus there was no deviation from independent action; however, for H. azteca the mixture was antagonistic (specifically dose ratio-dependent), and for N. triangulifer it was synergistic. The MDR method showed that only observations with H. azteca significantly deviated from independent action. Because of the lack of evidence of a clear deviation from independent action and the much greater potency of imidacloprid, the weight of evidence indicates that the presence of tebuconazole is unlikely to appreciably increase the hazard from imidacloprid exposure to aquatic invertebrates. Environ Toxicol Chem 2019;00:1-17. © 2019 SETAC.

Sensitivity of multiple life stages of 2 freshwater mussel species (Unionidae) to various pesticides detected in Ontario (Canada) surface waters.

Freshwater mussels contribute important ecological functions to aquatic systems. The water filtered by mussel assemblages can improve water quality, and the mixing of sediments by burrowing mussels can improve oxygen content and release nutrients. However, nearly 70% of North American freshwater mussel species are listed as either endangered, threatened, or in decline. In Ontario, 28 species are in decline or in need of protection. Even though freshwater mussels have a heightened sensitivity to some contaminants, few studies have investigated the risks that various pesticide classes pose to one freshwater mussel species or among life stages. Lampsilis siliquoidea and Villosa iris were the focus of the present study, with the latter currently listed as of "special concern" in Canada. A potential risk to the recovery of freshwater mussel species is the presence and persistence of pesticides in Ontario surface waters. Acute (48 h) toxicity tests were performed with V. iris glochidia to determine the effect on viability (surrogate for survival) following exposure to 4 fungicides (azoxystrobin, boscalid, metalaxyl, and myclobutanil), 3 neonicotinoids (clothianidin, imidacloprid, and thiamethoxam), 2 carbamates (carbaryl and malathion), 1 organophosphate (chlorpyrifos), and 1 butenolide (flupyradifurone). Juvenile and adult L. siliquoidea were also exposed to azoxystrobin, clothianidin, imidacloprid (juvenile only), and carbaryl (adult only). Our study found in general that all life stages were insensitive to the pesticides tested, with median effect and lethal concentrations >161 µg/L. The pesticides tested likely represent a minimal risk (hazard quotients <5.4 × 10-3 ) to freshwater mussel viability and survival in acute (48 h) and subchronic (28 d) exposures, respectively, in Ontario streams where pesticide concentrations were considerably lower than those tested in the present study. Environ Toxicol Chem 2018;37:2871-2880. © 2018 SETAC.

Relative chronic sensitivity of neonicotinoid insecticides to Ceriodaphnia dubia and Daphnia magna.

Neonicotinoid insecticides are a group of plant protectants frequently detected in surface waters at low concentrations. Aquatic invertebrates therefore have the potential to be exposed chronically to low concentrations of neonicotinoids. The cladocerans Daphnia magna and Ceriodaphnia dubia are among the most commonly used invertebrate test species in aquatic toxicology. Both species are known to be acutely insensitive to neonicotinoids, and while chronic toxicity has been characterized for D. magna, little research has been conducted with C. dubia. In the present study we conducted 7-d static-renewal life cycle tests for 6 neonicotinoids (acetamiprid, clothianidin, dinotefuran, imidacloprid, thiacloprid, and thiamethoxam) with C. dubia, and a 21-d test with imidacloprid with D. magna. 7-d LC50s for C. dubia ranged from 8.42 mg L-1 for imidacloprid to > 100 mg L-1 for clothianidin; 7-d reproduction EC50s were 2.98 for thiacloprid, to > 67 mg L-1 for dinotefuran. D. magna were less sensitive than C. dubia to imidacloprid, by 4-fold for lethality and 1.5-fold for reproduction; however, acute-to-chronic ratios (ACRs) were similar. ACRs, based on 48-h acute LC50s and 7- or 21-d chronic reproduction EC10s, ranged from 5.4 for acetamiprid to 53.0 for imidacloprid (mean 36.6, CV = 51%). Chronic toxicity values for both species were orders of magnitude greater than concentrations reported in the environment, and thus hazard to these cladocerans is negligible.

Investigation of clearance rate as an endpoint in toxicity testing with freshwater mussels (Unionidae).

The implementation of ecologically relevant sub-lethal endpoints in toxicity testing with freshwater mussels can provide valuable information during risk assessment, especially since these organisms are often exposed to low levels of contaminants. This study examined how to optimize quantifying the filtering capacity or clearance rate (CR) of mussels after exposure to a reference toxicant, sodium chloride (NaCl). CR was defined as the number of algal cells an individual mussel can remove from the overlying water by filtration over time and was determined using spectrophotometric absorbance and direct microscopic examination. Optimization included consideration of the following factors: concentration of algae mixture at test initiation, duration of CR assay, and statistical power. Experimental vessels contained either juvenile (ten, ~ 4 months old) or adult (one, ~ 2.5 years old) Lampsilis siliquoidea. To detect a 10% change in filtering capacity, the optimized adult CR assay was run for 48 h with 2.7 × 107 cells/mL of algae added at test initiation and a minimum of 6 replicates per treatment. The optimized juvenile mussel CR assay was run for 48 h with 1.77 × 107 cells/mL of algae added at test initiation; however, 13 replicates would be required to detect a 10% change to satisfy each method. To reduce the number of juvenile mussels used in testing, a minimum of 4 replicates per treatment was recommended to detect a 25% change in CR. After exposure to a reference toxicant (NaCl), EC50s from the optimized CR assay were compared to two other mussel toxicity endpoints: survival and burial (ability of mussels to bury in clean sand). CR by direct microscopic examination was slightly more sensitive than survival and burial in juveniles and only slightly more sensitive than survival in adults. No significant differences (p > 0.05) were detected between the EC/LC50 values determined from CR and the less labour-intensive survival and burial endpoints. The present study suggests the CR for juvenile and adult L. siliquoidea remained largely unaffected in mussels that survived a 7-day NaCl exposure.

Chronic toxicity of 6 neonicotinoid insecticides to Chironomus dilutus and Neocloeon triangulifer.

Neonicotinoid insecticides are frequently detected in surface waters near agricultural areas, leading to a potential for chronic exposure to sensitive aquatic species. The midge Chironomus dilutus and the mayfly Neocloeon triangulifer have been shown to be acutely sensitive to neonicotinoids. Previous studies have established chronic effects of some neonicotinoids on C. dilutus, but reproduction has not been studied. Toxic effects have not been assessed using N. triangulifer. We present the results of chronic, static-renewal tests for 6 neonicotinoids (acetamiprid, clothianidin, dinotefuran, imidacloprid, thiacloprid, and thiamethoxam) with C. dilutus (≤56-d in length) and N. triangulifer (≤32-d in length). Emergence was generally the most sensitive endpoint for both species across all neonicotinoids. Effect concentrations, 10% (EC10s; emergence) were 0.03 to 1.1 µg L-1 for acetamiprid, clothianidin, imidacloprid, and thiacloprid. Dinotefuran and thiamethoxam were less potent, with EC10s (C. dilutus) or median effect concentrations (EC50s; N. triangulifer) of 2.2 to 11.2 µg L-1 . Hazard was assessed through comparison of neonicotinoid environmental concentrations from agricultural surface waters in Ontario (Canada) with either the 5th percentile hazard concentration (for imidacloprid) or species-specific EC10s from the present study (for all remaining neonicotinoids). The resulting hazard quotients (HQs) indicated little to no hazard (HQ <1) in terms of chronic toxicity for acetamiprid, dinotefuran, thiacloprid, or thiamethoxam. A moderate hazard (HQ >1) was found for emergence of N. triangulifer for clothianidin, and a high hazard (HQ = 74) was found for imidacloprid. Environ Toxicol Chem 2018;37:2727-2739. © 2018 SETAC.