Naïve and induced tolerance of 15 amphibian populations to three commonly applied insecticides.

Human impacts on ecological communities are pervasive and species must either move or adapt to changing environmental conditions. For environments polluted by contaminants, researchers have found hundreds of target pest species evolving increased tolerance, but we have substantially fewer cases of evolved tolerance in non-target species. When species do evolve increased tolerance, inducible tolerance can provide immediate protection and favor the evolution of increased tolerance over generations via genetic assimilation. Using a model larval amphibian (wood frogs, Rana sylvatica), we examined the tolerance of 15 populations from western Pennsylvania and eastern New York (USA), when first exposed to no pesticide or sublethal concentrations and subsequently exposed to lethal concentrations of three common insecticides (carbaryl, chlorpyrifos, and diazinon). We found high variation in naïve tolerance among the populations for all three insecticides. We also discovered that nearly half of the populations exhibited inducible tolerance, though the degree of inducible tolerance (magnitude of tolerance plasticity; MoTP) varied. We observed a cross-tolerance pattern of the populations between chlorpyrifos and diazinon, but no pattern of similar MoTP among the pesticides. With populations combined from two regions, increased tolerance was not associated with proximity to agricultural fields, but there were correlations between proximity to agriculture and MoTP. Collectively, these results suggests that amphibian populations possess a wide range of naïve tolerance to common pesticides, with many also being able to rapidly induce increased tolerance. Future research should examine inducible tolerance in a wide variety of other taxa and contaminants to determine the ubiquity of these responses to anthropogenic factors.

Assessing the Combined Effects of Host and Parasite Exposure to Forever Chemicals in an Amphibian-Echinostome System.

Per- and polyfluoroalkyl substances (PFAS) are environmental contaminants of growing concern due to their potential negative effects on wildlife and human health. Per- and polyfluoroalkyl substances have been shown to alter immune function in various taxa, which could influence the outcomes of host-parasite interactions. To date, studies have focused on the effects of PFAS on host susceptibility to parasites, but no studies have addressed the effects of PFAS on parasites. To address this knowledge gap, we independently manipulated exposure of larval northern leopard frogs (Rana pipiens) and parasites (flatworms) via their snail intermediate host to environmentally relevant PFAS concentrations and then conducted trials to assess host susceptibility to infection, parasite infectivity, and parasite longevity after emergence from the host. We found that PFAS exposure to only the host led to no significant change in parasite load, whereas exposure of parasites to a 10-µg/L mixture of PFAS led to a significant reduction in parasite load in hosts that were not exposed to PFAS. We found that when both host and parasite were exposed to PFAS there was no difference in parasite load. In addition, we found significant differences in parasite longevity post emergence following exposure to PFAS. Although some PFAS-exposed parasites had greater longevity, this did not necessarily translate into increased infection success, possibly because of impaired movement of the parasite. Our results indicate that exposure to PFAS can potentially impact host-parasite interactions. Environ Toxicol Chem 2024;00:1-10. © 2024 SETAC.

Strobilurin fungicide increases the susceptibility of amphibian larvae to trematode infections.

The global rise in fungal pathogens has driven the increased usage of fungicides, yet our understanding of their ecotoxicity remains largely limited to acute toxicity. While such data is critical for projecting the risk of fungicide exposure to individual species, the contamination of natural systems with fungicides also has the potential to alter species interactions within communities including host-parasite relationships. We examined the effects of the fungicide pyraclostrobin on the susceptibility of larval American bullfrogs (Rana catesbeiana) to trematode (echinostome) infections using a controlled laboratory experiment. Following a 2-wk exposure to 0, 1.0, 5.2, or 8.4 µg/L of pyraclostrobin, tadpoles were then exposed to parasites either in the 1) presence (continued/simultaneous exposure) or 2) absence (fungicide-free water) of pyraclostrobin. We found that when exposed to pyraclostrobin during parasite exposure, meta cercariae counts increased 4 to 8 times compared to control tadpoles. Additionally, parasite loads were approximately 2 times higher in tadpoles with continued fungicide exposures compared to tadpoles that were moved to fresh water following fungicide exposure. This research demonstrates that fungicides at environmentally relevant concentrations can indirectly alter host-parasite interactions, which could elevate disease risk. It also underscores the need for studies that expand beyond traditional toxicity experiments to assess the potential community and ecosystem-level implications of environmental contaminants.

Toxicities of Legacy and Current-Use PFAS in an Anuran: Do Larval Exposures Influence Responses to a Terrestrial Pathogen Challenge?

Legacy polyfluoroalkyl substances (PFAS) [perfluorooctanesulfonate (PFOS) and perfluorooctanoic acid (PFOA)] are being replaced by various other fluorinated compounds, such as hexafluoropropylene oxide dimer acid (GenX). These alternatives are thought to be less bioaccumulative and, therefore, less toxic than legacy PFAS. Contaminant exposures occur concurrently with exposure to natural stressors, including the fungal pathogen Batrachocytrium dendrobatidis (Bd). Despite evidence that other pollutants can increase the adverse effects of Bd on anurans, no studies have examined the interactive effects of Bd and PFAS. This study tested the growth and developmental effects of PFOS, PFOA, and GenX on gray treefrog (Hyla versicolor) tadpoles, followed by a Bd challenge after metamorphosis. Despite PFAS exposure only occurring during the larval stage, carry-over effects on growth were observed post metamorphosis. Further, PFAS interacted with Bd exposure to influence growth; Bd-exposed animals had significantly shorter SVL [snout-vent length (mm)] with significantly increased body condition, among other time-dependent effects. Our data suggest that larval exposure to PFAS can continue to impact growth in the juvenile stage after exposure has ended. Contrary to predictions, GenX affected terrestrial performance more consistently than its legacy congener, PFOA. Given the role of Bd in amphibian declines, further investigation of interactions of PFAS with Bd and other environmentally relevant pathogens is warranted.

Chronic Exposure to a PFAS Mixture Resembling AFFF-Impacted Surface Water Decreases Body Size in Northern Leopard Frogs (Rana pipiens).

Per- and polyfluoroalkyl substances (PFAS) occur in the environment as mixtures, yet mixture toxicity remains poorly understood. Aqueous film-forming foams (AFFFs) are a common source of PFAS. Our objective was to examine chronic effects of a complex PFAS mixture on amphibian growth and development. We tested toxicity of a five-chemical PFAS mixture summing to 10 µg/L and that accounts for >90% of the PFAS in AFFF-affected surface waters: perfluorooctane sulfonate (PFOS, 40%), perfluorohexane sulfonic acid (PFHxS, 30%), perflurooctanoic acid (PFOA, 12.5%), perfluorohexanoic acid (PFHxA, 12.5%), and perfluoropentanoic acid (PFPeA, 5%). We also included treatments to determine whether PFOS drove mixture toxicity and whether PFOS and mixture components act additively. We exposed Northern leopard frog (Rana pipiens) larvae through metamorphosis (∼130 d) in outdoor mesocosms. After 21 days of exposure, the larval body condition fell ∼5% relative to controls in the 4 µg/L PFOS treatment and mixtures lacking PFOS. At metamorphosis, the full 5-component 10 µg/L PFAS mixture reduced mass by 16% relative to controls. We did not observe effects on development. Our results indicate that toxicity of PFOS and other PFAS mixtures typical of AFFF sites act additively and that PFOS is not more inherently toxic than other mixture components.

Population-level variation in insecticide tolerance across three life stages of the trematode Echinostoma trivolvis.

Ecotoxicological studies using single test populations may miss the inherent variation of natural systems and limit our understanding of how contaminants affect focal species. Though population-level variation in pesticide tolerance is commonly observed in host taxa, few studies have assessed population-level differences in the tolerance of parasites to different contaminants. We investigated population-level variation in insecticide tolerance of three Echinostoma trivolvis life stages (egg, miracidium, and cercaria) to three insecticides (carbaryl, chlorpyrifos, and diazinon). We tested two relevant metrics of insecticide tolerance-baseline and induced-across up to eight different parasite populations per life stage. Across all life stages, the insecticide treatments tended to reduce survival, but the magnitude of their effects often varied significantly among populations. Surprisingly, we found that exposure to chlorpyrifos increased the hatching success of echinostome eggs relative to the control treatment in three of six tested populations. We also found that cercariae shed from snails previously exposed to a sublethal concentration of chlorpyrifos had a significantly lower mortality rate when subsequently exposed to a lethal concentration of chlorpyrifos relative to individuals from snails that were not previously exposed; this suggests inducible tolerance in cercariae. We found no evidence that insecticide tolerance is correlated across parasite life stages within a population. Together the findings of our study demonstrate that single-population toxicity assays may greatly over- or underestimate the effects of pesticides on the survival of free-living parasite stages, insecticide tolerance levels may not be predictable from one parasite life stage to the next, and insecticides can have both expected and counterintuitive effects on non-target taxa.

A Critical Review of Amphibian Per- and Polyfluoroalkyl Substance Ecotoxicity Research Studies: Identification of Screening Levels in Water and Other Useful Resources for Site-Specific Ecological Risk Assessments.

With the goal of aiding risk assessors conducting site-specific risk assessments at per- and polyfluoroalkyl substance (PFAS)-contaminated sites, this critical review synthesizes information on the ecotoxicity of PFAS to amphibians in 10 amphibian species and 16 peer-reviewed publications. The studies in this review consisted of spiked-PFAS chronic toxicity experiments with perfluorooctane sulfonate (PFOS), perfluorooctanoic acid (PFOA), perfluorohexane sulfonate (PFHxS), and 6:2 fluorotelomer sulfonate (6:2 FTS) that evaluated apical endpoints typical of ecological risk-based decision making (survival, growth, and development). Body mass was the most sensitive endpoint, showing clear and biologically meaningful population level adverse effect sizes (≥20% adverse effects). From these results, we recommend chronic no observed effect concentration (NOEC) screening levels of 590 µg/L for PFOS and 130 µg/L for PFOA. At or above recommended chronic lowest observed effect concentration screening levels of 1100 µg/L PFOS and 1400 µg/L PFOA, there is an increased chance of adverse biologically relevant chronic effects. Biologically relevant adverse effects were not observed for PFHxS and 6:2 FTS, so unbounded NOECs of 1300 µg/L PFHxS and 1800 µg/L 6:2 FTS are recommended. Screening levels are also provided for the concentration of PFAS in an amphibian diet, amphibian tissue, and moss substrate. In addition, we recommend bioconcentration factors that can be useful to predict concentrations of PFAS in amphibians using concentrations in water; these values are useful for food web modeling to understand risks to vertebrate wildlife that prey on amphibians. Overall, the present study provides a guide to the wealth of ecotoxicological research on PFAS conducted by our research group and highlights the need for additional work that would improve the understanding of chemical risks to amphibians. Environ Toxicol Chem 2023;42:2078-2090. © 2023 SETAC.

Acute aquatic toxicity of two commonly used fungicides to midwestern amphibian larvae.

Fungicide usage has increased globally in response to the rise in fungal pathogens, especially in the agricultural sector. However, research examining the toxicity of fungicides is still limited for many aquatic species. In this study, we examined the acute toxicity of two widely used fungicides, chlorothalonil and pyraclostrobin, on six North American larval amphibian species across multiple families using 96-h LC50 tests. We found that pyraclostrobin was approximately 3.5x more toxic than chlorothalonil; estimated LC50 values ranged from 5-18 µg/L for pyraclostrobin and 15-50 µg/L for chlorothalonil. Comparing across amphibian groups, we found that salamanders were 3x more sensitive to pyraclostrobin than anuran species and equally as sensitive to chlorothalonil. Notably, our estimated LC50 values within the range of the expected environmental concentration for these fungicides suggesting environmental exposures could lead to direct mortality in these species. Given the widespread and increasing usage of fungicides, additional work should be conducted to assess the general risk posed by these chemicals to amphibian and their associated aquatic habitats.

Experimental evidence that host species composition alters host-pathogen dynamics in a ranavirus-amphibian assemblage.

Losses in biodiversity can alter disease risk through changes in host species composition. Host species vary in pathogen susceptibility and competence, yet how changes in diversity alter host-pathogen dynamics remains unclear in many systems, particularly with respect to generalist pathogens. Amphibians are experiencing worldwide population declines linked to generalist pathogens, such as ranavirus, and thus represent an ideal group to investigate how host species composition affects disease risk. We conducted experiments in which amphibian larvae of three native species (Pacific tree frogs, Pseudacris regilla; Cascades frogs, Rana cascadae; and Western toads, Anaxyrus boreas) were exposed to ranavirus individually (in the laboratory) or as assemblages (in outdoor mesocosms). In a laboratory experiment, we observed low survival and high viral loads in P. regilla compared to the other species, suggesting that this species was highly susceptible to the pathogen. In the mesocosm experiment, we observed 41% A. boreas mortality when alone and 98% mortality when maintained with P. regilla and R. cascadae. Our results suggest that the presence of highly susceptible species can alter disease dynamics across multiple species, potentially increasing infection risk and mortality in co-occurring species.

Population-level variation in pesticide tolerance predicts survival under field conditions in mayflies.

An increasing number of studies have found tolerance variation in populations consistently exposed to contaminants, but few studies have examined whether these laboratory-derived estimates of tolerance have survival implications in field conditions. We examined four populations of the mayfly Stenacron interpunctatum for variation in tolerance to the common agricultural insecticide clothianidin. Using laboratory bioassays, we found a 2.3× range in 96 h EC50 tolerance values to clothianidin between our four populations. We then conducted a common-garden experiment with nymphs from each population placed into the collection stream most heavily impacted by upstream agricultural activities to assess whether our laboratory tolerance estimates predict survival under field conditions. We monitored survival and growth in situ for three weeks during the spring planting season, when clothianidin is applied to croplands upstream of our study site. While growth was similar across all groups, the most tolerant population, which was native to the impacted stream, had higher survival than the more sensitive populations. This suggests that population-level variation in contaminant tolerance as measured in laboratory bioassays could have real-world survival implications for sensitive aquatic macroinvertebrates in contaminated streams.

Effects of Per- and Polyfluoroalkyl Substance Mixtures on the Susceptibility of Larval American Bullfrogs to Parasites.

Per- and polyfluoroalkyl substances (PFAS) are persistent environmental contaminants known to adversely affect health and development in many taxa. Although PFAS generally occur as mixtures in the environment, little is known about the effects of PFAS mixtures on organisms compared to single chemical exposures. Moreover, PFAS exposure in nature occurs alongside biotic factors such as parasitism. Even though host-parasite interactions are common in natural systems, there is little information about how PFAS affect these interactions. Here, we examined the effects of PFAS mixtures on the susceptibility of larval American bullfrogs (Rana catesbeiana) to echinostomes. Our PFAS treatments included perfluorooctanesulfonic acid (PFOS) at 4 and 10 ppb, two mixtures without PFOS as a component at 6 and 10 ppb total PFAS, and a mixture containing PFOS at 10 ppb total PFAS. We found that a 62-day PFAS exposure increased parasite loads by 42-100% in all treatments relative to the control. Additionally, we found that the singular exposure to PFOS increased parasite loads by ∼40% compared to a mixture containing PFOS suggesting antagonism among PFAS in mixtures. Our results highlight the need for further investigation into the effects of PFAS mixtures on organisms and how PFAS affect common ecological interactions.

An Environmentally Relevant Mixture of Perfluorooctanesulfonic Acid and Perfluorohexanesulfonic Acid Does Not Conform to Additivity in Northern Leopard Frogs Exposed Through Metamorphosis.

Per- and polyfluoroalkyl substances (PFAS) are chemicals associated with adverse health effects. At aqueous film-forming foam sites, they occur as mixtures, with perfluorooctanesulfonic acid (PFOS) and perfluorohexanesulfonic acid (PFHxS) commonly co-occurring in the highest concentrations. Although PFOS and PFHxS toxicities have been studied, few studies have tested their potential interaction. Using Rana pipiens, the present study compared toxicities of a 1:1 PFOS:PFHxS mixture to PFOS and PFHxS individually with the prediction that responses would be additive. Gosner stage 25 (GS 25) tadpoles were exposed through metamorphosis (GS 46) to 0.5 and 1 ppb PFOS or PFHxS alone or to a mixture of 0.5 ppb PFOS and 0.5 ppb PFHxS. Tadpoles were weighed and measured (snout-vent length [SVL]) at day 31, metamorphic climax (GS 42), and GS 46. These values were used to calculate the scaled mass index (SMI), a measure of body condition. Body burdens were quantified on day 31 and at GS 46. The PFOS and PFHxS body burdens were elevated relative to controls at GS 46. No effects were observed on survival, SVL, or mass. Single PFAS effects included a 17% reduction in SMI at day 31 (0.5 ppb PFHxS) and a 1.1-day longer metamorphic period (1 ppb PFHxS) relative to controls. Mixture results deviated from additivity-SMIs were higher than expected on day 31 and lower than expected at GS 42. In addition, time to GS 42 in the PFAS mixture exceeded expected additivity by 12 days. Results from a chronic exposure to a 1:1 PFOS:PFHxS mixture resulted in changes in body condition and length of metamorphosis that deviated from additivity. More PFAS mixture toxicity studies conducted at relevant ratios and concentrations are needed. Environ Toxicol Chem 2022;41:3007-3016. © 2022 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Pathogens and predators: examining the separate and combined effects of natural enemies on assemblage structure.

Natural enemy ecology strives to unify predator-prey and host-pathogen interactions under a common framework to gain insights into community- and ecosystem-level processes. To address this goal, ecologists need a greater emphasis on: (1) quantifying pathogen-mediated effects on community structure to enable comparisons with predator-mediated effects and (2) determining the interactive effects of combined natural enemies on communities. We conducted a mesocosm experiment to assess the individual and combined effects of predators (dragonfly larvae and adult water bugs) and a pathogen (ranavirus) on the abundance and composition of a larval amphibian assemblage. We found that our three natural enemies structured victim assemblages in unique ways, producing distinct assemblages. Additionally, we found that in combination treatments, predators mainly drove assemblage structure such that the assemblages most closely resembled their respective predator treatments. We also found that predators reduced infection prevalence in combination treatments, and that the magnitude of this effect was dependent on predator identity. Compared to virus-alone treatments, the presence of dragonflies and water bugs reduced infection prevalence by 79% and 63%, respectively. Additionally, the presence of dragonflies eliminated ranavirus infection in two species, which demonstrates the prominent role of predators in disease dynamics in this system. Overall, this work demonstrates the importance of considering natural enemies in community ecology, as each enemy can elicit a unique structural change. Additionally, this study provides a unique empirical test of the healthy herds hypothesis for multi-species assemblages and underscores the importance of advancing our understanding of multi-enemy interactions within communities.

Acute Toxicity of Eight Aqueous Film-Forming Foams to 14 Aquatic Species.

Researchers have developed numerous per- and polyfluoroalkyl substances (PFAS)-free aqueous film-forming foam (AFFF) formulations to replace PFAS-containing AFFF used for fire suppression. As part of the Department of Defense's Strategic Environmental Research and Development Program (SERDP), we examined the direct lethal effects of seven PFAS-free AFFF and a PFAS-containing AFFF on 14 aquatic species using a series of lethal concentration (LC50) tests. We assessed the LC10, LC50, and LC90 values using log-logistic and logit analyses. Across all aquatic species tested, we discovered that exposure to at least one PFAS-free AFFF was more or as toxic as exposure to the PFAS-containing AFFF. For most cases, National Foam Avio F3 Green KHC 3% and Buckeye Platinum Plus C6MILSPEC 3% were the most and least toxic formulations, respectively. Moreover, we found consistency among results from multiple experiments using the same minnow species (Pimephales promelas) and among closely related taxa (e.g., daphnids, amphibians). Lastly, the LC50 values for AFFF formulations trended lower for tested marine species as compared to those of freshwater species. These results dramatically increase the current knowledge on the potentially toxic effects of AFFF but also highlight the need for additional research and the development of new PFAS-free AFFF that are more "ecologically friendly" than those containing persistent PFAS.

Exposure to clothianidin and predators increases mortality for heptageniidae.

Neonicotinoids are a class of insecticide with global impacts on natural environments. Due to their high solubility, they are frequently found in stream ecosystems where they have the potential to impact non-target biota. While environmental concentrations are generally below lethal levels for most organisms, there are concerns that sublethal exposures can impact aquatic insects, particularly mayflies, which are highly sensitive to neonicotinoids. Because sublethal doses of neonicotinoids can reduce mobility in mayflies, exposure could indirectly increase mortality due to predation by impairing their ability to avoid initial detection or escape predators. We examined whether exposure to the neonicotinoid clothianidin at a concentration below the 96-h EC50 (7.5 µg/L), would increase the predation risk of Stenacron and Stenonema mayfly nymphs by larval southern two-lined salamanders (Eurycea cirrigera) or eastern dobsonfly nymphs (Corydalus cornutus) using a controlled laboratory experiment. For Stenacron, we found significant interactive effects between pesticide and dobsonfly exposure that increased the hazard ratio (HR). The HR assesses risk relative to a control population, in this case mayflies in similar experimental conditions but without exposure to neonicotinoids or predators. With the addition of clothianidin, the HR of mayflies exposed to a dobsonfly nymph significantly increased from 1.8 to 6.2 while the HR for those exposed to salamanders increased from 7.6 to 12.5. For Stenonema, the HR initially decreased due to dobsonfly exposure (1 to 0.3) but increased when clothianidin and dobsonflies were combined (0.3 to 1.6). Our study shows that aquatic exposure to clothianidin can increase mortality for aquatic insects through predator pressure. Such indirect effects associated with neonicotinoid exposure warrant further investigation to expand our understanding of pesticide impacts to aquatic systems.

Comparative Toxicity of Aquatic Per- and Polyfluoroalkyl Substance Exposure in Three Species of Amphibians.

Per- and polyfluoroalkyl substances (PFAS) are contaminants of concern due to their widespread occurrence in the environment, persistence, and potential to elicit a range of negative health effects. Per- and polyfluoroalkyl substances are regularly detected in surface waters, but their effects on many aquatic organisms are still poorly understood. Species with thyroid-dependent development, like amphibians, can be especially susceptible to PFAS effects on thyroid hormone regulation. We examined sublethal effects of aquatic exposure to four commonly detected PFAS on larval northern leopard frogs (Rana [Lithobates] pipiens), American toads (Anaxyrus americanus), and eastern tiger salamanders (Ambystoma tigrinum). Animals were exposed for 30 days (frogs and salamanders) or until metamorphosis (toads) to 10, 100, or 1000 µg/L of perfluorooctane sulfonate (PFOS), perfluorooctanoic acid (PFOA), perfluorohexane sulfonate (PFHxS), or 6:2 fluorotelomer sulfonate (6:2 FTS). We determined that chronic exposure to common PFAS can negatively affect amphibian body condition and development at concentrations as low as 10 µg/L. These effects were highly species dependent, with species having prolonged larval development (frogs and salamanders) being more sensitive to PFAS than more rapidly developing species (toads). Our results demonstrate that some species could experience sublethal effects at sites with surface waters highly affected by PFAS. Our results also indicate that evaluating PFAS toxicity using a single species may not be sufficient for accurate amphibian risk assessment. Future studies are needed to determine whether these differences in susceptibility can be predicted from species' life histories and whether more commonly occurring environmental levels of PFAS could affect amphibians. Environ Toxicol Chem 2022;41:1407-1415. © 2022 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Temperature affects the toxicity of pesticides to cercariae of the trematode Echinostoma trivolvis.

Global climate change is predicted to have significant impacts on ecological interactions such as host-parasite relationships. Increased temperatures may also interact with other anthropogenic stressors, such as chemical contaminants, to exacerbate or reduce parasite transmission. However, studies on the effects of pesticides on non-target species are typically conducted at one standard temperature, despite the toxicity of many agrochemicals being temperature-dependent. Furthermore, most studies assessing the effects of temperature on pesticide toxicity have been conducted on host organisms, limiting our understanding of how temperature affects the toxicity of pesticides to free-living parasite stages as they move through the environment in search of a host. Using the free-swimming cercariae stage of the trematode Echinostoma trivolvis, we examined how the toxicities of three different pesticides (a carbamate insecticide, strobilurin fungicide, and triazine herbicide) vary by temperature by monitoring cercarial swimming activity over time. Our three main findings were: 1) a strong main effect of temperature across all pesticide trials - higher temperatures caused cercariae to cease swimming activity earlier, likely due to an increased rate of energy expenditure, 2) atrazine, azoxystrobin, and carbaryl were directly toxic to cercariae to some degree, but not in a predictable dose-dependent manner, and 3) the temperature at which pesticide exposure occurs could affect its toxicity to cercariae. The interaction between pesticide and temperature was most evident in the azoxystrobin exposure; azoxystrobin caused cercariae to cease swimming activity earlier at 30 °C but caused cercariae to maintain swimming activity longer at 18 °C relative to their respective pesticide-free control treatments. These findings highlight the importance of conducting toxicity assays at multiple temperatures and suggest that the combined effects of pesticides and temperature on host-parasite interactions may be difficult to generalize.

Pesticides alter ecosystem respiration via phytoplankton abundance and community structure: Effects on the carbon cycle?

Freshwater systems are critical to life on earth, yet they are threatened by the increasing rate of synthetic chemical pollution. Current predictions of the effects of synthetic chemicals on freshwater ecosystems are hampered by the sheer number of chemical contaminants entering aquatic systems, the diversity of organisms inhabiting these systems, the myriad possible direct and indirect effects resulting from these combinations, and uncertainties concerning how contaminants might alter ecosystem metabolism via changes in biodiversity. To address these knowledge gaps, we conducted a mesocosm experiment that elucidated the responses of ponds composed of phytoplankton and zooplankton to standardized concentrations of 12 pesticides, nested within four pesticide classes, and two pesticide types. We show that the effects of the pesticides on algae were consistent within herbicides and insecticides and that responses of over 70 phytoplankton species and genera were consistent within broad taxonomic groups. Insecticides generated top-down effects on phytoplankton community composition and abundance, which were associated with persistent increases in ecosystem respiration. Insecticides had direct toxic effects on cladocerans, which led to competitive release of copepods. These changes in the zooplankton community led to a decrease in green algae and a modest increase in diatoms. Herbicides did not change phytoplankton composition but reduced total phytoplankton abundance. This reduction in phytoplankton led to short-term decreases in ecosystem respiration. Given that ponds release atmospheric carbon and that worldwide pesticide pollution continues to increase exponentially, scientists and policy makers should pay more attention to the ways pesticides alter the carbon cycle in ponds via changes in communities, as demonstrated by our results. Our results show that these predictions can be simplified by grouping pesticides into types and species into functional groups. Adopting this approach provides an opportunity to improve the efficiency of risk assessment and mitigation responses to global change.

Predator- and competitor-induced responses in amphibian populations that evolved different levels of pesticide tolerance.

Exposure to agrochemicals can drive rapid phenotypic and genetic changes in exposed populations. For instance, amphibian populations living far from agriculture (a proxy for agrochemical exposure) exhibit low pesticide tolerance, but they can be induced to possess high tolerance following a sublethal pesticide exposure. In contrast, amphibian populations close to agriculture exhibit high, constitutive tolerance to pesticides. A recent study has demonstrated that induced pesticide tolerance appears to have arisen from plastic responses to predator cues. As a result, we might expect that selection for constitutive pesticide tolerance in populations near agriculture (i.e., genetic assimilation) will lead to the evolution of constitutive responses to natural stressors. Using 15 wood frog (Rana sylvatica) populations from across an agricultural gradient, we conducted an outdoor mesocosm experiment to examine morphological (mass, body length, and tail depth) and behavioral responses (number of tadpoles observed and overall activity) of tadpoles exposed to three stressor environments (no-stressor, competitors, or predator cues). We discovered widespread differences in tadpole traits among populations and stressor environments, but no population-by-environment interaction. Subsequent linear models revealed that population distance to agriculture (DTA) was occasionally correlated with tadpole traits in a given environment and with magnitudes of plasticity, but none of the correlations were significant after Bonferroni adjustment. The magnitudes of predator and competitor plasticity were never correlated with the magnitude of pesticide-induced plasticity that we documented in a companion study. These results suggest that while predator-induced plasticity appears to have laid the foundation for the evolution of pesticide-induced plasticity and its subsequent genetic assimilation, inspection of population-level differences in plastic responses show that the evolution of pesticide-induced plasticity has not had a reciprocal effect on the evolved plastic responses to natural stressors.

Relative acute toxicity of three per- and polyfluoroalkyl substances on nine species of larval amphibians.

Per- and polyfluoroalkyl substances (PFAS) are widespread, persistent environmental pollutants known to elicit a wide range of negative effects on wildlife species. There is scarce information regarding the toxicity of PFAS on amphibians, but amphibians may be highly susceptible because of their permeable skin and dependence on fresh water. Acute toxicity studies are a first step toward understanding responses to PFAS exposure, providing benchmarks for species-specific tolerances, informing ecological risk assessment (ERA), and designing chronic toxicity studies. We conducted standardized 96-h lethal concentration (LC50) toxicity tests for perfluorooctanesulfonic acid (PFOS) and perfluorooctanoic acid (PFOA) with 9 widely distributed amphibian species native to eastern and central North America. We also conducted LC50 tests with perfluorohexane sulfonate (PFHxS) for 2 species and determined whether toxicity of PFOS and PFOA varied between life stages for 3 species. Acute toxicity varied among PFAS and species and between developmental stages within species. Across all species, toxicity of PFOS was more than 8× higher than PFOA. Salamanders in the genus Ambystoma were generally more sensitive to PFOS than were anurans (frogs and a toad). Toxicity of PFOA was highest for small-mouthed salamanders and gray tree frogs and lowest for Jefferson salamanders, American bullfrogs, green frogs, and wood frogs. Although only 2 species were exposed to PFHxS, survival was lower for green frogs than for American bullfrogs. Toxicity of PFAS also varied between developmental stages of larvae. Gray tree frogs were more sensitive at later developmental stages, and small-mouthed salamanders were more sensitive at earlier developmental stages. Our study is one of the first to report species-, developmental stage-, and compound-specific differences in sensitivity to PFAS across a wide range of amphibian species. The benchmarks for toxicity we determined can inform conservation and remediation efforts, guide chronic toxicity studies, and help predict influences on amphibian communities, thereby informing future ERAs for PFAS. Integr Environ Assess Manag 2021;17:684-689. © 2021 SETAC.