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.

Density declines, richness increases, and composition shifts in stream macroinvertebrates.

Documenting trends of stream macroinvertebrate biodiversity is challenging because biomonitoring often has limited spatial, temporal, and taxonomic scopes. We analyzed biodiversity and composition of assemblages of >500 genera, spanning 27 years, and 6131 stream sites across forested, grassland, urban, and agricultural land uses throughout the United States. In this dataset, macroinvertebrate density declined by 11% and richness increased by 12.2%, and insect density and richness declined by 23.3 and 6.8%, respectively, over 27 years. In addition, differences in richness and composition between urban and agricultural versus forested and grassland streams have increased over time. Urban and agricultural streams lost the few disturbance-sensitive taxa they once had and gained disturbance-tolerant taxa. These results suggest that current efforts to protect and restore streams are not sufficient to mitigate anthropogenic effects.

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.

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.

The Combined Effects of Road Salt and Biotic Stressors on Amphibian Sex Ratios.

Aquatic systems worldwide are threatened by the anthropogenic use of synthetic chemicals, including pesticides, pharmaceuticals, and road de-icers. Exposure to contaminants can alter the behavior, morphology, and physiology of organisms if it occurs during sensitive life stages. For instance, past studies have documented feminization of male amphibians following herbicide exposure and skewed sex ratios among amphibian populations exposed to road salt. However, many of these studies lack the complexities found within natural environments, such as competition with conspecifics or threat of predation, which are also known to influence development. Thus, it is important to understand how anthropogenic and natural stressors interact to alter animal sex ratios. Given the growing concern of secondary salinization of freshwater systems, we exposed larval wood frogs (Rana sylvatica) to either road salt (sodium chloride [NaCl]) or an alternative salt mixture (NaCl, magnesium chloride [MgCl2 ], and potassium chloride [KCl]) at 3 concentrations (200, 600, and 1000 mg Cl- /L) crossed with 3 biotic stressors (no-stressor control, competition, or predator cues) to examine their potentially interactive effects on sex. Exposure to biotic stressors and NaCl did not influence wood frog sex ratios. In contrast, tadpole exposure to the intermediate salt mixture concentration significantly reduced the proportion of female frogs. Future studies are needed to determine whether such changes in sex are widespread among sensitive species with complex life cycles, and to assess the consequences of sex ratio changes on long-term population dynamics. Environ Toxicol Chem 2021;40:231-235. © 2020 SETAC.

Cascading effects of insecticides and road salt on wetland communities.

Novel stressors introduced by human activities increasingly threaten freshwater ecosystems. The annual application of more than 2.3 billion kg of pesticide active ingredient and 22 billion kg of road salt has led to the contamination of temperate waterways. While pesticides and road salt are known to cause direct and indirect effects in aquatic communities, their possible interactive effects remain widely unknown. Using outdoor mesocosms, we created wetland communities consisting of zooplankton, phytoplankton, periphyton, and leopard frog (Rana pipiens) tadpoles. We evaluated the toxic effects of six broad-spectrum insecticides from three families (neonicotinoids: thiamethoxam, imidacloprid; organophosphates: chlorpyrifos, malathion; pyrethroids: cypermethrin, permethrin), as well as the potentially interactive effects of four of these insecticides with three concentrations of road salt (NaCl; 44, 160, 1600 Cl- mg/L). Organophosphate exposure decreased zooplankton abundance, elevated phytoplankton biomass, and reduced tadpole mass whereas exposure to neonicotinoids and pyrethroids decreased zooplankton abundance but had no significant effect on phytoplankton abundance or tadpole mass. While organophosphates decreased zooplankton abundance at all salt concentrations, effects on phytoplankton abundance and tadpole mass were dependent upon salt concentration. In contrast, while pyrethroids had no effects in the absence of salt, they decreased zooplankton and phytoplankton density under increased salt concentrations. Our results highlight the importance of multiple-stressor research under natural conditions. As human activities continue to imperil freshwater systems, it is vital to move beyond single-stressor experiments that exclude potentially interactive effects of chemical contaminants.

Effect of Agrochemical Exposure on Schistosoma mansoni Cercariae Survival and Activity.

Singular use of activity assays or staining dyes to assess pathogen agrochemical tolerance can underestimate tolerance if pesticides cause sublethal effects. We exposed Schistosoma mansoni cercariae, the aquatic life stage of this trematode that infects humans, to 4 insecticides at 5 concentrations using a 24-h time-to-death assay. We used Trypan blue dye, which stains dead tissue, and activity assays simultaneously to discriminate dead from live but paralyzed individuals. Whereas cypermethrin, deltamethrin, and dimethoate exposure did not affect cercariae at any ecologically relevant concentrations, methamidophos exposure increased survival of cercariae compared with those in the controls. This was because methamidophos-induced paralysis reduced cercarial activity and thus energy expenditures, extending the lifespan of this short-lived parasite that causes human schistosomiasis. These findings highlight that sublethal effects should be considered when pesticide effects on disease are under investigation. Environ Toxicol Chem 2020;39:1421-1428. © 2020 SETAC.

Nutrients influence the multi-trophic impacts of an invasive species unaffected by native competitors or predators.

Non-native species often lead to undesirable ecological and environmental impacts. Two hypotheses that predict establishment of non-native species are enemy release and biotic resistance. Support for these hypotheses in freshwater invasions is mixed. Experiments combined with field observations provide a complementary approach to understanding how interactions between native and non-native species lead to enemy release or biotic resistance. We tested experimentally whether these hypotheses provided insights into the invasion of the banded mystery snail (Viviparus georgianus), which has invaded the Great Lakes region and northeastern Unites States (US) from the southeastern US. Because freshwater systems vary widely in their nutrient concentrations due to natural and anthropogenic processes, we tested whether nutrient additions altered competitive and predatory interactions that regulate mechanisms of enemy release or biotic resistance. We evaluated the status of the mystery snail invasion in a 3-year field survey of Lake George (NY, US) to identify if field observations supported any experimental conclusions. The presence of the banded mystery snail led to a 14% and 27% reduction in biomass of a native competitor under low- and high-nutrient concentrations, respectively. The mystery snail also triggered a 29% biomass loss of a native snail predator, but only in low-nutrient concentrations. Field surveys indicated that the mystery snail dominated the snail community; of seven snail species, it comprised 77% of all snails. Results from the field surveys combined with experimental results indicate that neither competitors nor predators have likely suppressed the invasion of the banded mystery snail. This conclusion is consistent with competitive- and predatory-enemy release as we found no indication of biotic resistance via competition or predation from native species. Our results further highlight that the post-establishment impacts of invasive species are altered by the trophic state of freshwater ecosystems.

Local adaptation of the MHC class IIß gene in populations of wood frogs (Lithobates sylvaticus) correlates with proximity to agriculture.

Major histocompatibility complex (MHC) genes code for membrane-embedded proteins that are involved in parasite/pathogen recognition. The link between the MHC and immunity makes these genes important genetic markers to evaluate in systems where infectious disease is associated with population declines. As human impacts on wildlife populations continue to increase, it is also essential to evaluate the role of MHC and immunity in the context of anthropogenic change. Amphibians are an ideal model to test the role of the MHC in infectious disease resistance, as parasites and anthropogenic disturbances currently threaten populations worldwide. We characterized the diversity of MHC class IIß peptide binding region alleles, 13 microsatellite loci, and population-level trematode resistance in 14 populations of wood frogs (Lithobates sylvaticus) in northwestern Pennsylvania with varying geographic distances to agriculture. To assess local adaptation in the MHC IIß, we compared genetic differentiation of MHC IIß and microsatellite markers (FST). We also tested for an effect of isolation by distance on genetic differentiation of MHC IIß and microsatellite markers. In addition, we evaluated whether population-level MHC IIß diversity and common allele frequencies correlate with distance to agriculture and trematode resistance. We found no evidence for genetic structure based on microsatellite analysis nor an effect of isolation by distance on neutral and immunogenetic markers. However, we did detect structure based on the MHC IIß locus, suggesting that it is under local selection. The MHC IIß allele Lisy-DAB*1 was more common in populations living near agricultural sites. Populations with higher MHC IIß diversity showed increased resistance to trematodes. Our results suggest that wood frog populations experience immunogenetic differences at a small scale. In addition, agriculture may disturb natural associations between hosts and parasites through its influence on immunocompetence, underscoring the importance of examining the effects of environmental context on host-parasite interactions.

Evolved tolerance to freshwater salinization in zooplankton: life-history trade-offs, cross-tolerance and reducing cascading effects.

Recent discoveries have documented evolutionary responses to freshwater salinization. We investigated if evolutionary responses to salinization exhibit life-history trade-offs or if they can mitigate ecological impacts such as cascading effects through mechanisms of tolerance and cross-tolerance. We conducted an outdoor mesocosm experiment using populations of Daphnia pulex-a ubiquitous algal grazer-that were either naive or had previously experienced selection to become more tolerant to sodium chloride (NaCl). During the initial phase of population growth, we discovered that evolved tolerance comes at the cost of slower population growth in the absence of salt. We found evolved Daphnia populations maintained a tolerance to NaCl approximately 30 generations after the initial discovery. Evolved tolerance to NaCl also conferred cross-tolerance to a high concentration of CaCl2 (3559 µS cm-1) and a moderate concentration of MgCl2 (967 µS cm-1). A higher concentration of MgCl2 (2188 µS cm-1) overwhelmed the cross-tolerance and killed all Daphnia Tolerance to NaCl did not mitigate NaCl-induced cascades leading to phytoplankton blooms, but cross-tolerance at moderate concentrations of MgCl2 and high concentrations of CaCl2 mitigated such cascading effects caused by these two salts. These discoveries highlight the important interplay between ecology and evolution in understanding the full impacts of freshwater salinization.This article is part of the theme issue 'Salt in freshwaters: causes, ecological consequences and future prospects'.

Timing and frequency of sublethal exposure modifies the induction and retention of increased insecticide tolerance in wood frogs (Lithobates sylvaticus).

Although the paradigm for increased tolerance to pesticides has been by selection on constitutive (naïve) traits, recent research has shown it can also occur through phenotypic plasticity. However, the time period in which induction can occur, the duration of induced tolerance, and the influence of multiple induction events remain unknown. We hypothesized that the induction of increased pesticide tolerance is limited to early sensitive periods, the magnitude of induced tolerance depends on the number of exposures, and the retention of induced tolerance depends on the time elapsed after an exposure and the number of exposures. To test these hypotheses, we exposed wood frog tadpoles to either a no-carbaryl control (water) or 0.5 mg/L carbaryl at 4 time periods, and later tested their tolerance to carbaryl using time-to-death assays. We discovered that tadpoles induced increased tolerance early and midway but not late in our experiment and their constitutive tolerance increased with age. We found no difference in the magnitude of induced tolerance after a single or 2 exposures. Finally, induced pesticide tolerance was reversed within 6 d but was retained only when tadpoles experienced all 4 consecutive exposures. Phenotypic plasticity provides an immediate response for sensitive amphibian larvae to early pesticide exposures and reduces phenotypic mismatches in aquatic environments contaminated by agrochemicals. Environ Toxicol Chem 2018;37:2188-2197. © 2018 SETAC.

Evolved pesticide tolerance influences susceptibility to parasites in amphibians.

Because ecosystems throughout the globe are contaminated with pesticides, there is a need to understand how natural populations cope with pesticides and the implications for ecological interactions. From an evolutionary perspective, there is evidence that pesticide tolerance can be achieved via two mechanisms: selection for constitutive tolerance over multiple generations or by inducing tolerance within a single generation via phenotypic plasticity. While both mechanisms can allow organisms to persist in contaminated environments, they might result in different performance trade-offs including population susceptibility to parasites. We have identified 15 wood frog populations that exist along a gradient from close to agriculture and high, constitutive pesticide tolerance to far from agriculture and inducible pesticide tolerance. Using these populations, we investigated the relationship between evolutionary responses to the common insecticide carbaryl and host susceptibility to the trematode Echinoparyphium lineage 3 and ranavirus using laboratory exposure assays. For Echinoparyphium, we discovered that wood frog populations living closer to agriculture with high, constitutive tolerance experienced lower loads than populations living far from agriculture with inducible pesticide tolerance. For ranavirus, we found no relationship between the mechanism of evolved pesticide tolerance and survival, but populations living closer to agriculture with high, constitutive tolerance experienced higher viral loads than populations far from agriculture with inducible tolerance. Land use and mechanisms of evolved pesticide tolerance were associated with susceptibility to parasites, but the direction of the relationship is dependent on the type of parasite, underscoring the complexity between land use and disease outcomes. Collectively, our results demonstrate that evolved pesticide tolerance can indirectly influence host-parasite interactions and underscores the importance of including evolutionary processes in ecotoxicological studies.

Inducible Tolerance to Agrochemicals Was Paved by Evolutionary Responses to Predators.

Recent research has reported increased tolerance to agrochemicals in target and nontarget organisms following acute physiological changes induced through phenotypic plasticity. Moreover, the most inducible populations are those from more pristine locations, far from agrochemical use. We asked why do populations with no known history of pesticide exposure have the ability to induce adaptive responses to novel agrochemicals? We hypothesized that increased pesticide tolerance results from a generalized stressor response in organisms, and would be induced following sublethal exposure to natural and anthropogenic stressors. We exposed larval wood frogs (Lithobates sylvaticus) to one of seven natural or anthropogenic stressors (predator cue (Anax spp.), 0.5 or 1.0 mg carbaryl/L, road salt (200 or 1000 mg Cl-/L), ethanol-vehicle control, or no-stressor control) and subsequently tested their tolerance to a lethal carbaryl concentration using time-to-death assays. We observed induced carbaryl tolerance in tadpoles exposed to 0.5 mg/L carbaryl and also in tadpoles exposed to predator cues. Our results suggest that the ability to induce pesticide tolerance likely arose through evolved antipredator responses. Given that antipredator responses are widespread among species, many animals might possess inducible pesticide tolerance, buffering them from agrochemical exposure.

Salinization triggers a trophic cascade in experimental freshwater communities with varying food-chain length.

The application of road deicing salts in northern regions worldwide is changing the chemical environment of freshwater ecosystems. Chloride levels in many lakes, streams, and wetlands exceed the chronic and acute thresholds established by the United States and Canada for the protection of freshwater biota. Few studies have identified the impacts of deicing salts in stream and wetland communities and none have examined impacts in lake communities. We tested how relevant concentrations of road salt (15, 100, 250, 500, and 1000 mg Cl- /L) interacted with experimental communities containing two or three trophic levels (i.e., no fish vs. predatory fish). We hypothesized that road salt and fish would have a negative synergistic effect on zooplankton, which would then induce a trophic cascade. We tested this hypothesis in outdoor mesocosms containing filamentous algae, periphyton, phytoplankton, zooplankton, several macroinvertebrate species, and fish. We found that the presence of fish and high salt had a negative synergistic effect on the zooplankton community, which in turn caused an increase in phytoplankton. Contributing to the magnitude of this trophic cascade was a direct positive effect of high salinity on phytoplankton abundance. Cascading effects were limited with respect to impacts on the benthic food web. Periphyton and snail grazers were unaffected by the salt-induced trophic cascade, but the biomass of filamentous algae decreased as a result of competition with phytoplankton for light or nutrients. We also found direct negative effects of high salinity on the biomass of filamentous algae and amphipods (Hyalella azteca) and the mortality of banded mystery snails (Viviparus georgianus) and fingernail clams (Sphaerium simile). Clam mortality was dependent on the presence of fish, suggesting a non-consumptive interactive effect with salt. Our results indicate that globally increasing concentrations of road salt can alter community structure via both direct and indirect effects.

Effect of Simultaneous Amphibian Exposure to Pesticides and an Emerging Fungal Pathogen, Batrachochytrium dendrobatidis.

Amphibian declines have been linked to numerous factors, including pesticide use and the fungal pathogen Batrachochytrium dendrobatidis (Bd). Moreover, research has suggested a link between amphibian sensitivity to Bd and pesticide exposure. We simultaneously exposed postmetamorphic American toads (Anaxyrus americanus), western toads (A. boreas), spring peepers (Pseudacris crucifer), Pacific treefrogs (P. regilla), leopard frogs (Lithobates pipiens), and Cascades frogs (Rana cascadae) to a factorial combination of two pathogen treatments (Bd+, Bd-) and four pesticide treatments (control, ethanol vehicle, herbicide mixture, and insecticide mixture) for 14 d to quantify survival and infection load. We found no interactive effects of pesticides and Bd on anuran survival and no effects of pesticides on infection load. Mortality following Bd exposure increased in spring peepers and American toads and was dependent upon snout-vent length in western toads, American toads, and Pacific treefrogs. Previous studies reported effects of early sublethal pesticide exposure on amphibian Bd sensitivity and infection load at later life stages, but we found simultaneous exposure to sublethal pesticide concentrations and Bd had no such effect on postmetamorphic juvenile anurans. Future research investigating complex interactions between pesticides and Bd should employ a variety of pesticide formulations and Bd strains and follow the effects of exposure throughout ontogeny.

Investigation of road salts and biotic stressors on freshwater wetland communities.

The application of road deicing salts has led to the salinization of freshwater ecosystems in northern regions worldwide. Increased chloride concentrations in lakes, streams, ponds, and wetlands may negatively affect freshwater biota, potentially threatening ecosystem services. In an effort to reduce the effects of road salt, operators have increased the use of salt alternatives, yet we lack an understanding of how these deicers affect aquatic communities. We examined the direct and indirect effects of the most commonly used road salt (NaCl) and a proprietary salt mixture (NaCl, KCl, MgCl2), at three environmentally relevant concentrations (150, 470, and 780 mg Cl-/L) on freshwater wetland communities in combination with one of three biotic stressors (control, predator cues, and competitors). The communities contained periphyton, phytoplankton, zooplankton, and two tadpole species (American toads, Anaxyrus americanus; wood frogs, Lithobates sylvaticus). Overall, we found the two road salts did not interact with the natural stressors. Both salts decreased pH and reduced zooplankton abundance. The strong decrease in zooplankton abundance in the highest NaCl concentration caused a trophic cascade that resulted in increased phytoplankton abundance. The highest NaCl concentration also reduced toad activity. For the biotic stressors, predatory stress decreased whereas competitive stress increased the activity of both tadpole species. Wood frog survival, time to metamorphosis, and mass at metamorphosis all decreased under competitive stress whereas toad time to metamorphosis increased and mass at metamorphosis decreased. Road salts and biotic stressors can both affect freshwater communities, but their effects are not interactive.

Combined effects of road salt and an insecticide on wetland communities.

As the numbers of chemical contaminants in freshwater ecosystems increase, it is important to understand whether contaminants interact in ecologically important ways. The present study investigated the independent and interactive effects of 2 contaminants that frequently co-occur in freshwater environments among higher latitudes, including a commonly applied insecticide (carbaryl) and road salt (NaCl). The hypothesis was that the addition of either contaminant would result in a decline in zooplankton, an algal bloom, and the subsequent decline of both periphyton and periphyton consumers. Another hypothesis was that combining the contaminants would result in synergistic effects on community responses. Outdoor mesocosms were used with communities that included phytoplankton, periphyton, zooplankton, amphipods, clams, snails, and tadpoles. Communities were exposed to 4 environmentally relevant concentrations of salt (27 mg Cl- L-1 , 77 mg Cl- L-1 , 277 mg Cl- L-1 , and 727 mg Cl- L-1 ) fully crossed with 4 carbaryl treatments (ethanol, 0 µg L-1 , 5 µg L-1 , and 50 µg L-1 ) over 57 d. Contaminants induced declines in rotifer and cladoceran zooplankton, but only carbaryl induced an algal bloom. Consumers exhibited both positive and negative responses to contaminants, which were likely the result of both indirect community interactions and direct toxicity. In contrast to the hypothesis, no synergistic effects were found, although copepod densities declined when high concentrations of both chemicals were combined. The results suggest that low concentrations of salt and carbaryl are likely to have mostly independent effects on aquatic communities. Environ Toxicol Chem 2017;36:771-779. © 2016 SETAC.

Evolved pesticide tolerance in amphibians: Predicting mechanisms based on pesticide novelty and mode of action.

We examined 10 wood frog populations distributed along an agricultural gradient for their tolerance to six pesticides (carbaryl, malathion, cypermethrin, permethrin, imidacloprid, and thiamethoxam) that differed in date of first registration (pesticide novelty) and mode-of-action (MOA). Our goals were to assess whether: 1) tolerance was correlated with distance to agriculture for each pesticide, 2) pesticide novelty predicted the likelihood of evolved tolerance, and 3) populations display cross-tolerance between pesticides that share and differ in MOA. Wood frog populations located close to agriculture were more tolerant to carbaryl and malathion than populations far from agriculture. Moreover, the strength of the relationship between distance to agriculture and tolerance was stronger for older pesticides compared to newer pesticides. Finally, we found evidence for cross-tolerance between carbaryl and malathion (two pesticides that share MOA). This study provides one of the most comprehensive approaches for understanding patterns of evolved tolerance in non-pest species.

The contribution of phenotypic plasticity to the evolution of insecticide tolerance in amphibian populations.

Understanding population responses to rapid environmental changes caused by anthropogenic activities, such as pesticides, is a research frontier. Genetic assimilation (GA), a process initiated by phenotypic plasticity, is one mechanism potentially influencing evolutionary responses to novel environments. While theoretical and laboratory research suggests that GA has the potential to influence evolutionary trajectories, few studies have assessed its role in the evolution of wild populations experiencing novel environments. Using the insecticide, carbaryl, and 15 wood frog populations distributed across an agricultural gradient, we tested whether GA contributed to the evolution of pesticide tolerance. First, we investigated the evidence for evolved tolerance to carbaryl and discovered that population-level patterns of tolerance were consistent with evolutionary responses to pesticides; wood frog populations living closer to agriculture were more tolerant than populations living far from agriculture. Next, we tested the potential role of GA in the evolution of pesticide tolerance by assessing whether patterns of tolerance were consistent with theoretical predictions. We found that populations close to agriculture displayed constitutive tolerance to carbaryl whereas populations far from agriculture had low naïve tolerance but high magnitudes of induced tolerance. These results suggest GA could play a role in evolutionary responses to novel environments in nature.

Here today, gone tomorrow: Short-term retention of pesticide-induced tolerance in amphibians.

Pesticide use has led to the ubiquitous contamination of natural habitats, which has inadvertently increased pesticide tolerance in target and nontarget species. Historically, increased pesticide tolerance has been attributed to natural selection for tolerance among individuals of affected populations. Recent research, however, has discovered that pesticide tolerance can be increased through phenotypic plasticity. Although induced pesticide tolerance may benefit organisms experiencing contaminated systems, little is known about its occurrence in vertebrates, its retention through ontogeny, or potential life history tradeoffs. Using time-to-death assays at 2 distinct developmental windows, the authors discovered that gray treefrog (Hyla versicolor) tadpoles exposed to sublethal concentrations (0 mg a.i./L, 0.5 mg a.i./L, and 1.0 mg a.i./L) of the insecticide Sevin® (carbaryl) early in life increased their pesticide tolerance to a lethal carbaryl concentration 5 d after sublethal exposure. However, this increased tolerance was not retained later in ontogeny (23 d post-sublethal exposure). Moreover, no indication was found of pesticide-induced treefrogs experiencing life-history tradeoffs in terms of survival to metamorphosis, mass, or snout-vent length. Gray treefrogs are only the second vertebrate species and the second amphibian family to exhibit pesticide-induced tolerance after sublethal exposure. The authors' data suggest that the ability to induce increased pesticide tolerance may play a critical role in amphibian survival in contaminated ecosystems. However, future work is needed to test the occurrence of inducible pesticide tolerance among numerous amphibian populations worldwide.