Exposure Assessment of Pesticides in Surface Waters of Ontario, Canada Reveals Low Probability of Exceeding Acute Regulatory Thresholds.

The objectives of this study were to: (1) characterize the exposure of aquatic ecosystems in Southern Ontario, Canada to pesticides between 2002 and 2016 by constructing environmental exposure distributions (EEDs), including censored data; and (2) predict the probability of exceeding acute regulatory guidelines. Surface water samples were collected over a 15-year period by Environment and Climate Change Canada. The dataset contained 167 compounds, sampled across 114 sites, with a total of 2,213 samples. There were 67,920 total observations of which 55,058 were non-detects (81%), and 12,862 detects (19%). The most commonly detected compound was atrazine, with a maximum concentration of 18,600 ngL- 1 and ~ 4% chance of exceeding an acute guideline (1,000 ngL- 1) in rivers and streams. Using Southern Ontario as a case study, this study provides insight into the risk that pesticides pose to aquatic ecosystems and the utility of EEDs that include censored data for the purpose of risk assessment.

Novel Reproduction Toxicity Test Method for Hyalella azteca Using Sexually Mature Amphipods Provides More Robust Data Than Standard Methods in Exposures to Imidacloprid.

Hyalella azteca is an epibenthic crustacean used in ecotoxicology, but there are challenges associated with standard methods using reproduction as an endpoint. A novel, 28-day reproduction toxicity test method for H. azteca was created to address these issues by initiating tests with sexually mature amphipods to eliminate the confounding effects of growth, using a sex ratio of seven females to three males to reduce reproductive variability, and conducting tests in water-only conditions to make recovery of juveniles easier and expand testing capabilities to water-soluble compounds. In the present study, we evaluated the 28-day novel method by comparing it with the 42-day standard test method in duplicate and parallel water-only, static-renewal exposures to sublethal concentrations of imidacloprid (0.5-8 µg/L). Both methods showed similar effects on survival, with survival approaching 50% in the highest test concentration (8 µg/L). However, the 42-day median effect concentrations (EC50s) for growth were more sensitive in the standard method (1.5-3.2 µg/L) compared with the 28-day EC50s generated by the novel method (>8 µg/L). Reproduction endpoints (juveniles/female) produced similar EC50s between methods, but the data were less variable in novel tests (smaller coefficients of variation); therefore, fewer replicates would be required to detect effects on reproduction compared with the standard method. In addition, novel tests generated 28 days of reproduction data compared with 14 days using standard tests and allowed survival and growth of sexes to be assessed independently. Thus, the novel method shows promise to improve the use of reproduction as an endpoint in water-only toxicity tests with H. azteca. Environ Toxicol Chem 2024;43:723-735. © 2024 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Human health and environmental risk assessment of metals in community gardens of Winnipeg, Manitoba, Canada.

Fresh produce is an important component of maintaining cognitive and physical health, particularly for children. A mechanism to increase access to fresh produce is the construction of community gardens in urban centres. While reducing barriers to nutritious food, the soil of the community garden can contain contaminants (e.g. metals) depending on the location and how the garden was constructed. This study quantified, for the first time, seven metals (As, Cd, Cr, Cu, Pb, Mn, and Ni) in soil from 83 community gardens across the City of Winnipeg in Manitoba, Canada. Concentrations of metals in soil were used to create distributions for environmental exposure and estimated daily intake, which were then used to determine exceedances of soil quality guidelines and acceptable daily intakes, respectively. Raised garden beds and gardens further from roads had typically lower concentrations of metals in surface gardens and those nearer to roads. While some concentrations of metals exceeded CCME guidelines levels for the protection of environmental health, the vast majority represent a low risk. For human health, only As posed a quantifiable risk of exceeding the USEPA acceptable daily intake via the consumption of produce from gardens, though this was < 1.2% for the whole population and < 10.2% for children aged 1 to 2 years. Overall, this study is the first to show that the concentration of the metals in soil from gardens typically poses a low risk to environmental and human health. We recommend the use of raised gardens to further mitigate risk.

Natural compounds from freshwater mussels disrupt fungal virulence determinants and influence fluconazole susceptibility in the presence of macrophages in Cryptococcus neoformans.

Cryptococcus neoformans is a human fungal pathogen responsible for fatal infections, especially in patients with a depressed immune system. Overexposure to antifungal drugs due to prolonged treatment regimens and structure-similar applications in agriculture have weakened the efficacy of current antifungals in the clinic. The rapid evolution of antifungal resistance urges the discovery of new compounds that inhibit fungal virulence determinants, rather than directly killing the pathogen, as alternative strategies to overcome disease and reduce selective pressure toward resistance. Here, we evaluated the efficacy of freshwater mussel extracts (crude and clarified) against the production of well-defined virulence determinants (i.e., thermotolerance, melanin, capsule, and biofilm) and fluconazole resistance in C. neoformans. We demonstrated the extracts' influence on fungal thermotolerance, capsule production, and biofilm formation, as well as susceptibility to fluconazole in the presence of macrophages. Additionally, we measured the inhibitory activity of extracts against commercial peptidases (family representatives of cryptococcal orthologs) related to fungal virulence determinants and fluconazole resistance, and integrated these phenotypic findings with quantitative proteomics profiling. Our approach defined distinct signatures of each treatment and validated a new mechanism of anti-virulence action toward the polysaccharide capsule from a selected extract following fractionation. By understanding the mechanisms driving the antifungal activity of mussels, we may develop innovative treatment options to overcome fungal infections and promote susceptibility to fluconazole in resistant strains. IMPORTANCE: As the prevalence and severity of global fungal infections rise, along with an increasing incidence of antifungal resistance, new strategies to combat fungal pathogens and overcome resistance are urgently needed. Critically, our current methods to overcome fungal infections are limited and drive the evolution of resistance forward; however, an anti-virulence approach to disarm virulence factors of the pathogen and promote host cell clearance is promising. Here, we explore the efficacy of natural compounds derived from freshwater mussels against classical fungal virulence determinants, including thermotolerance, capsule production, stress response, and biofilm formation. We integrate our phenotypic discoveries with state-of-the-art mass spectrometry-based proteomics to identify mechanistic drivers of these antifungal properties and propose innovative avenues to reduce infection and support the treatment of resistant strains.

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.

Characterizing the Microplastic Content of Biosolids in Southern Ontario, Canada.

The application of biosolids to agricultural land has been identified as a major pathway of microplastic (MP) pollution to the environment. Very little research, however, has been done on the MP content of biosolids within Canada. Fifteen biosolid samples from different treatment processes (liquid, dewatered, pelletized, and alkali-stabilized) were collected from 11 sources across southern Ontario to quantify and characterize the MP load within them. All samples exhibited MP concentrations ranging from 188 200 (±24 161) to 512 000 (±28 571) MPs/kg dry weight and from 4122 (±231) to 453 746 (±38 194) MPs/kg wet weight. Field amendment of these biosolids can introduce up to 3.73 × 106 to 4.12 × 108 MP/ha of agricultural soil. There was no significant difference in the MP concentrations of liquid, dewatered, and pelletized samples; but a reduction in MP content was observed in alkali-stabilized biosolids. Fragments composed 57.6% of the MPs identified, while 36.7% were fibers. In addition, MPs showed an exponential increase in abundance with decreasing size. Characterization of MPs confirmed that polyester was the most abundant, while polyethylene, polypropylene, polyamide, polyacrylamide, and polyurethane were present across the majority of biosolid samples. The results of the present study provide an estimate of the potential extent of MP contamination to agricultural fields through the amendment of biosolids. Environ Toxicol Chem 2024;43:793-806. © 2023 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Terrestrial Isopods Generate Microplastics from Low-Density Polyethylene Without Effects on Survival.

With concern growing regarding the impact of microplastics (MPs) on terrestrial ecosystems, it is important to assess the role invertebrates may play in the fate of MPs within these ecosystems. Commonly, MPs enter these environments through improperly discarded waste or the application of treated biosolids and/or wastewater on agricultural soils. The present study investigated whether three species of terrestrial isopod (Porcellio scaber, Porcellio laevis, and Porcellionides pruinosus) ingest plastic debris and generate MPs during exposures varying from 24 h to 14 days and whether this may have an adverse effect on their health. Test vessels were designed to expose isopods to plastic fragments in the form of polyethylene plastic foam. Isopods were exposed to plastic that was either (1) pristine, or (2) weathered in a soil and water solution prior to incorporation in test vessels. When exposed to weathered polyethylene, all three species generated MPs (minimum-maximum size values for all durations inclusive: P. laevis = 114-1673 µm, P. scaber = 99-1635 µm, P. pruinosus = 85-1113 µm) through the consumption of macroplastic fragments with no observed impact on their health. In the shorter-duration exposures, the number of MPs generated by the isopod species in the present study was highly variable between experimental vessels (minimum-maximum generated MPs for 14-day exposure: P. laevis = 25-420, P. scaber = 50-583, P. pruinosus = 48-311). However, as the exposure durations increased, there was a clear trend of increasing MP generation, indicating that the isopods continued to consume the plastic fragments as long as the surface was weathered. A significant difference in the size of generated MPs was observed as well, with smaller isopod species generating smaller MP fragments on average. The results of the present study confirm that certain species of isopod can contribute to the generation of MPs, which constitutes an additional pathway of MP exposure to soil ecosystems. Environ Toxicol Chem 2024;43:784-792. © 2023 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

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.

Lethal and Sub-lethal Implications of Sodium Chloride Exposure for Adult Unionid Mussel Species: Eurynia dilatata and Lasmigona costata.

The elevated use of salt as a de-icing agent on roads in Canada is causing an increase in the chloride concentration of freshwater ecosystems. Freshwater Unionid mussels are a group of organisms that are sensitive to increases in chloride levels. Unionids have greater diversity in North America than anywhere else on Earth, but they are also one of the most imperiled groups of organisms. This underscores the importance of understanding the effect that increasing salt exposure has on these threatened species. There are more data on the acute toxicity of chloride to Unionids than on chronic toxicity. This study investigated the effect of chronic sodium chloride exposure on the survival and filtering activity of two Unionid species (Eurynia dilatata, and Lasmigona costata) and assessed the effect on the metabolome in L. costata hemolymph. The concentration causing mortality after 28 days of exposure was similar for E. dilatata (1893 mg Cl-/L) and L. costata (1903 mg Cl-/L). Significant changes in the metabolome of the L. costata hemolymph were observed for mussels exposed to non-lethal concentrations. For example, several phosphatidylethanolamines, several hydroxyeicosatetraenoic acids, pyropheophorbide-a, and alpha-linolenic acid were significantly upregulated in the hemolymph of mussels exposed to 1000 mg Cl-/L for 28 days. While no mortality occurred in the treatment, elevated metabolites in the hemolymph are an indicator of stress.

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.

Differentiated extracts from freshwater and terrestrial mollusks inhibit virulence factor production in Cryptococcus neoformans.

The human fungal pathogen, Cryptococcus neoformans, is responsible for deadly infections among immunocompromised individuals with the evolution of antifungal resistance driving the solution to discover new compounds that inhibit fungal virulence factors rather than kill the pathogen. Recently, exploration into natural sources (e.g., plants, invertebrates, microbes) of antifungal agents has garnered attention by integrating a One Health approach for new compound discovery. Here, we explore extracts from three mollusk species (freshwater and terrestrial) and evaluate effects against the growth and virulence factor production (i.e., thermotolerance, melanin, capsule, and biofilm) in C. neoformans. We demonstrate that clarified extracts of Planorbella pilsbryi have a fungicidal effect on cryptococcal cells comparable to fluconazole. Similarly, all extracts of Cipangopaludina chinensis affect cryptococcal thermotolerance and impair biofilm and capsule production, with clarified extracts of Cepaea nemoralis also conveying the latter effect. Next, inhibitory activity of extracts against peptidases related to specific virulence factors, combined with stress assays and quantitative proteomics, defined distinct proteome signatures and proposed proteins driving the observed anti-virulence properties. Overall, this work highlights the potential of compounds derived from natural sources to inhibit virulence factor production in a clinically important fungal pathogen.

Assessing the Putative Anticryptococcal Properties of Crude and Clarified Extracts from Mollusks.

Cryptococcus neoformans is an encapsulated human fungal pathogen with a global distribution that primarily infects immunocompromised individuals. The widespread use of antifungals in clinical settings, their use in agriculture, and strain hybridization have led to increased evolution of resistance. This rising rate of resistance against antifungals is a growing concern among clinicians and scientists worldwide, and there is heightened urgency to develop novel antifungal therapies. For instance, C. neoformans produces several virulence factors, including intra- and extra-cellular enzymes (e.g., peptidases) with roles in tissue degradation, cellular regulation, and nutrient acquisition. The disruption of such peptidase activity by inhibitors perturbs fungal growth and proliferation, suggesting this may be an important strategy for combating the pathogen. Importantly, invertebrates such as mollusks produce peptidase inhibitors with biomedical applications and anti-microbial activity, but they are underexplored in terms of their usage against fungal pathogens. In this protocol, a global extraction from mollusks was performed to isolate potential peptidase inhibitors in crude and clarified extracts, and their effects against classical cryptococcal virulence factors were assessed. This method supports the prioritization of mollusks with antifungal properties and provides opportunities for the discovery of anti-virulence agents by harnessing the natural inhibitors found in mollusks.

Freshwater mussels in an impacted watershed: Influences of pollution from point and non-point sources.

The Grand River watershed in a densely populated region of Ontario supports one of the richest assemblages of freshwater mussels in Canada. However, water quality in this watershed is influenced by urban development, agriculture, and industry. Mussel populations and water chemistry in the lower Grand River and the Boston Creek tributary were evaluated to determine whether point sources of pollution such as discharges of domestic wastewater and industrial effluent, and non-point sources of pollution are affecting mussel distribution and population structure. Semi-quantitative population surveys conducted at 9 study sites identified 20 mussel species, including 3 Species at Risk. Mussel abundance (34-160 mussels/search hour) and species richness indicated that mussel populations in the lower Grand River watershed are continuing to recover from historical lows reported in the 1970s. However, changes in populations at some sites were consistent with altered water chemistry. Most notable was that the three most abundant mussel species in the Boston Creek tributary downstream of a gypsum plant discharge were significantly smaller in length than those upstream of this site. The water chemistry in this habitat was characterized by elevated conductivity (∼2000 µS/cm) and calcium (∼500 mg/L), as well as concentrations of sulfate (∼1000 mg/L) that can be toxic to freshwater mussels. In the Grand River downstream of the confluence with Boston Creek, there tended to be (p > 0.05) fewer mussels (mean 34 ± 20/search h) compared to upstream (mean 67 ± 15/search h) and this corresponded to altered water chemistry, including elevated sulfate (239 mg/L) downstream of the confluence relative to upstream (58 mg/L). These data indicate that chronic exposures to high levels of major ions is likely driving changes to mussel population structure. In addition, the discharges of wash water from a gypsum plant may be impacting sensitive biota in the main stem Grand River well beyond the immediate tributary receiving environment.

Characterizing the exposure of streams in southern Ontario to agricultural pesticides.

Aquatic communities can be exposed to pesticides transported from land. Characterizing this exposure is key to predicting potential toxic effects. In this study, samples of streamwater from 21 sites were used to characterize pesticide exposure to aquatic communities. Sites were in agricultural areas of southwestern Ontario, Canada and were sampled monthly from 2012 to 2019 from April to November. Samples were analyzed for a suite of hundreds of pesticides and pesticide degradation products and other water quality indicators (e.g., nutrients). Frequently detected pesticides included herbicides (2,4-D; bentazon; MCPP; metolachlor) and neonicotinoid insecticides (NNIs) (clothianidin; thiamethoxam) which were detected in >50% of samples collected between 2015 and 2019. Non-metric multidimensional scaling (NMDS) was used to explore connections between pesticide concentrations and upstream land use and crop type. Detectable concentrations of the NNI clothianidin and many herbicides were related to corn, soybean, and grain/cereal crops while concentrations of the NNI imidacloprid, insecticide flonicamid, and fungicide boscalid were related to greenhouse/nursery land use. Potential toxicity to aquatic communities was assessed by comparing pesticide concentrations to Pesticide Toxicity Index (PTI) values. Few samples exceeded levels where acute (1% of samples) or chronic toxicity (10.5%) would be expected. The diamide insecticide chlorantraniliprole was detected in several streamwater samples at levels that may cause toxicity to aquatic invertebrates, highlighting the need for continued toxicity research into this pesticide class. The number of pesticides detected was positively correlated with nutrient and total suspended solids levels, underscoring the multiple stressors aquatic communities are exposed to in these habitats.

Acute oral toxicity and risks of four classes of systemic insecticide to the Common Eastern Bumblebee (Bombus impatiens).

The Common Eastern Bumblebee (Bombus impatiens) is native to North America with an expanding range across Eastern Canada and the USA. This species is commercially produced primarily for greenhouse crop pollination and is a common and abundant component of the wild bumblebee fauna in agricultural, suburban and urban landscapes. However, there is a dearth of pesticide toxicity information about North American bumblebees. The present study determined the acute oral lethal toxicity (48-h LD50) of: the butenolide, flupyradifurone (>1.7 µg/bee); the diamide, cyantraniliprole (>0.54 µg/bee); the neonicotinoid, thiamethoxam (0.0012 µg/bee); and the sulfoximine, sulfoxaflor (0.0177 µg/bee). Compared with published honey bee (Apis mellifera) LD50 values, the present study shows that sulfoxaflor and thiamethoxam are 8.3× and 3.3× more acutely toxic to B. impatiens, whereas flupyradifurone is more acutely toxic to A. mellifera. The current rule of thumb for toxicity extrapolation beyond the honey bee as a model species, termed 10× safety factor, may be sufficient for bumblebee acute oral toxicity. A comparison of five risk assessment equations suggested that the Standard Risk Approach (SRA) and Fixed Dose Risk Approach (FDRA) provide more nuanced levels of risk evaluation compared to the Exposure Toxicity Ratio (ETR), Hazard Quotient (HQ), and Risk Quotient (RQ), primarily because the SRA and FDRA take into account real world variability in pollen and nectar pesticide residues and the chances that bees may be exposed to them.

Salt-Laden Winter Runoff and Freshwater Mussels; Assessing the Effect on Early Life Stages in the Laboratory and Wild Mussel Populations in Receiving Waters.

The widespread use of road salt for winter road maintenance has led to an increase in the salinity of surface water in many seasonally cold areas. Freshwater mussels have a heightened sensitivity to salt, which is a concern, because many Canadian mussel species at risk have ranges limited to southern Ontario, Canada's most road-dense region. This study examined the effect of winter road runoff on freshwater mussels. The impact of two bridges that span mussel habitat in the Thames River watershed (Ontario, Canada), the second most species-rich watershed for mussels in Canada, were studied. During a winter melt event, bridge runoff, as well as creek surface water surrounding the bridges were collected. Chloride concentrations in samples from bridge deck and tile drains varied (99-8250 mg/L). In general, survival of Lampsilis fasciola glochidia exposed to those samples reflected chloride levels (e.g. 84% at 99 mg/L; 0% at 8250 mg/L), although potassium (60 mg/L) may have at least contributed to toxicity in one sample. Serial dilution exposures with the two most toxic runoff samples revealed 48-h glochidia EC50s of 44% (McGregor Creek Tile Drain) and 26% (Baptiste Creek Deck Drain). During the melt event, the chloride concentrations in creek surface waters downstream of the bridges ranged from 69 to 179 mg Cl-/L; effects on glochidia (viability 77-91%) exposed to those waters was minimal. There were no live mussels surrounding one bridge (Baptiste Creek), likely due to poor habitat. At the other targeted bridge (McGregor Creek), fewer mussels were found close (< 100 m up- or downstream) to the bridge than further (> 200 m) away. However, other contributing factors, including agriculture, were present at both study areas.

Prioritization of Pesticides for Assessment of Risk to Aquatic Ecosystems in Canada and Identification of Knowledge Gaps.

Pesticides can enter aquatic environments via direct application, via overspray or drift during application, or by runoff or leaching from fields during rain events, where they can have unintended effects on non-target aquatic biota. As such, Fisheries and Oceans Canada identified a need to prioritize current-use pesticides based on potential risks towards fish, their prey species, and habitats in Canada. A literature review was conducted to: (1) Identify current-use pesticides of concern for Canadian marine and freshwater environments based on use and environmental presence in Canada, (2) Outline current knowledge on the biological effects of the pesticides of concern, and (3) Identify general data gaps specific to biological effects of pesticides on aquatic species. Prioritization was based upon recent sales data, measured concentrations in Canadian aquatic environments between 2000 and 2020, and inherent toxicity as represented by aquatic guideline values. Prioritization identified 55 pesticides for further research nationally. Based on rank, a sub-group of seven were chosen as the top-priority pesticides, including three herbicides (atrazine, diquat, and S-metolachlor), three insecticides (chlorpyrifos, clothianidin, and permethrin), and one fungicide (chlorothalonil). A number of knowledge gaps became apparent through this process, including gaps in our understanding of sub-lethal toxicity, environmental fate, species sensitivity distributions, and/or surface water concentrations for each of the active ingredients reviewed. More generally, we identified a need for more baseline fish and fish habitat data, ongoing environmental monitoring, development of marine and sediment-toxicity benchmarks, improved study design including sufficiently low method detection limits, and collaboration around accessible data reporting and management.

Ecotoxicology of Glyphosate, Its Formulants, and Environmental Degradation Products.

The chemical and biological properties of glyphosate are key to understanding its fate in the environment and potential risks to non-target organisms. Glyphosate is polar and water soluble and therefore does not bioaccumulate, biomagnify, or accumulate to high levels in the environment. It sorbs strongly to particles in soil and sediments and this reduces bioavailability so that exposures to non-target organisms in the environment are acute and decrease with half-lives in the order of hours to a few days. The target site for glyphosate is not known to be expressed in animals, which reduces the probability of toxicity and small risks. Technical glyphosate (acid or salts) is of low to moderate toxicity; however, when mixed with some formulants such as polyoxyethylene amines (POEAs), toxicity to aquatic animals increases about 15-fold on average. However, glyphosate and the formulants have different fates in the environment and they do not necessarily co-occur. Therefore, toxicity tests on formulated products in scenarios where they would not be used are unrealistic and of limited use for assessment of risk. Concentrations of glyphosate in surface water are generally low with minimal risk to aquatic organisms, including plants. Toxicity and risks to non-target terrestrial organisms other than plants treated directly are low and risks to terrestrial invertebrates and microbial processes in soil are very small. Formulations containing POEAs are not labeled for use over water but, because POEA rapidly partitions into sediment, risks to aquatic organisms from accidental over-sprays are reduced in shallow water bodies. We conclude that use of formulations of glyphosate under good agricultural practices presents a de minimis risk of direct and indirect adverse effects in non-target organisms.

Clothianidin alters leukocyte profiles and elevates measures of oxidative stress in tadpoles of the amphibian, Rana pipiens.

Neonicotinoid pesticide use is widespread and highly debated, as evidenced by recent attention received from the public, academics and pesticide regulatory agencies. However, relatively little is known about the physiological effects of neonicotinoid insecticides on aquatic vertebrates. Amphibians (larval stages in particular) are excellent vertebrate bioindicators in aquatic systems due to their risk of exposure and sensitivity to environmental stressors. Previous work with wood frog (Rana sylvatica) tadpoles exposed to formulated products containing thiamethoxam or clothianidin in outdoor mesocosms found significant shifts in leukocyte profiles, suggesting the tadpoles were physiologically stressed. The main objective of the present study was to characterize this stress response further using complementary measures of stress after exposure to clothianidin on northern leopard frogs (Rana pipiens) during their aquatic larval stages. Laboratory static-renewal exposures were conducted over eight weeks with the technical product clothianidin at 0, 0.23, 1, 10 and 100 µg/L, and diquat dibromide at 532 µg/L was used as a positive control. We assessed tadpole leukocyte profiles and measures of oxidative stress as these sub-lethal alterations could affect amphibian fitness. We found changes in several types of leukocytes at 1 and 10 µg/L, suggesting that these tadpoles exhibited signs of mild physiological stress. Clothianidin also induced an oxidative stress response at 0.23, 1 and 100 µg/L. However, we found no differences in survival, growth, development time or hepatosomatic index in frogs exposed to clothianidin. Our study indicates that tadpoles chronically exposed to clothianidin have increased stress responses, but in the absence of concentration-response relationships and effects on whole-organism endpoints, the implications on the overall health and fitness of these changes are unclear.