Devin K. Jones

The toxicity of Roundup Original Max® to 13 species of larval amphibians

With the increased use of glyphosate-based herbicides (marketed under several names, including Roundup and Vision), there has been a concomitant increased concern about the unintended impacts that particular formulations containing the popular surfactant polyethoxylated tallowamine (POEA) might have on amphibians. Published studies have examined a relatively small number of anuran species (primarily from Australia and eastern North America) and, surprisingly, no species of salamanders. Using a popular formulation of glyphosate (Roundup Original Max), the goal of the present study was to conduct tests of lethal concentrations estimated to kill 50% of a population after 96 h (LC50(96-h)) on a wider diversity of species from both eastern and western North America. Tests were conducted on nine species of stage 25, larval anurans from three families (Ranidae: Rana pipiens, R. clamitans, R. sylvatica, R. catesbeiana, R. cascadae; Bufonidae: Bufo americanus, B. boreas; and Hylidae: Hyla versicolor, Pseudacris crucifer) and four species of larval salamanders from two families (Ambystomatidae: Ambystoma gracile, A. maculatum, A. laterale; and Salamandridae: Notophthalmus viridescens). For the nine species of larval anurans, LC50(96-h) values ranged from 0.8- to 2.0-mg acid equivalents per liter with relatively little pattern in differential sensitivity among the species or families. The four species of larval salamanders were less sensitive than the anurans, with LC50(96-h) values ranging from 2.7- to 3.2-mg acid equivalents per liter and no substantial differences among the species of salamanders. This work substantially increases the available data on amphibian sensitivity to glyphosate formulations that include either POEA surfactants or the equally moderately to highly toxic surfactants of Roundup Original Max and should be useful for improving future risk assessments.

Very highly toxic effects of endosulfan across nine species of tadpoles: lag effects and family-level sensitivity.

Pesticides are commonly used for health and economic benefits worldwide, but increased use has led to increased contamination of aquatic habitats. To understand potential impacts on nontarget organisms in these habitats, toxicologists generally use short-term (4-d) toxicity tests on model organisms. For most pesticides, few amphibian tests have been conducted, but there is growing concern about the potential impact of pesticides to amphibian populations. For the insecticide endosulfan, previous studies have found that low concentrations can be very highly toxic to amphibians and have suggested that this mortality may exhibit important lag effects. To estimate the lethal concentration of endosulfan that would cause 50% mortality after 4 d (LC50(4-d)) across a diversity of amphibians and the presence of lag effects, LC50(4-d) experiments were conducted on nine species of tadpoles from three families (Bufonidae: Bufo americanus, B. boreas; Hylidae: Pseudacris crucifer, P. regilla, Hyla versicolor; and Ranidae: Rana pipiens, R. clamitans, R. cascadae, R. catesbeiana) and then held the animals for an additional 4 d in clean water. The LC50(4-d) values for endosulfan ranged from 1.3 to 120 ppb, which classifies endosulfan as highly toxic to very highly toxic. Moreover, holding the animals for an additional 4 d in clean water revealed significant additional mortality in three of the nine species. Leopard frogs, for example, experienced no significant death during the initial 4-d exposure to 60 ppb but 97% death after an additional 4 d in clean water. A phylogenetic pattern also appears to exist among families, with Bufonidae being least susceptible, Hylidae being moderately susceptible, and Ranidae being most susceptible. Results from the present study provide valuable data to assess the impact of endosulfan on a globally declining group of vertebrates.

Competitive stress can make the herbicide Roundup® more deadly to larval amphibians.

Toxicity assessments on nontarget organisms have largely been addressed using short-term, single-species laboratory experiments. Although extremely helpful, these experiments inherently lack many pervasive ecological stressors found in nature. Though a substantial challenge, incorporating these ecological stressors in contaminant studies would shed light on potential synergistic effects. For the worlds leading herbicide, glyphosate, we know little about how natural stressors affect the toxicity to nontarget organisms. To explore how the natural stress of competition might interact with a glyphosate-based herbicide, we used outdoor mesocosms containing three tadpole species that were exposed to a factorial combination of three glyphosate concentrations (0, 1, 2, or 3 mg acid equivalent (a.e.)/L of the commercial formulation Roundup Original MAX®) and three tadpole densities (low, medium, or high). We found that increased tadpole density caused declines in tadpole growth, but also made the herbicide significantly more lethal to one species. Whereas the median lethal concentration (LC50) values were similar across all densities for gray treefrogs (Hyla versicolor; 1.7-2.3 mg a.e./L) and green frogs (Rana clamitans; 2.2-2.6 mg a.e./L), the LC50 values for bullfrogs (R. catesbeiana) were 2.1 to 2.2 mg a.e./L at low and medium densities, but declined to 1.6 mg a.e./L at high densities. The large decrease in amphibian survival with increased herbicide concentration was associated with increases in periphyton abundance. We also found evidence that temperature stratification lead to herbicide stratification in the water column, confirming the results of a previous study and raising important questions about exposure risk in natural systems.

Roundup® and amphibians: The importance of concentration, application time, and stratification

The widespread use of pesticides raises the possibility that non-target organisms might also be affected. To assess this, the traditional approach has been to conduct short-term laboratory experiments spanning a range of lethal concentrations and some longer-duration experiments at sublethal concentrations. While this approach has been very useful, less attention has been paid to the timing of exposure and the impacts of multiple, small exposures versus single, large exposures. We examined the role of application amount, timing, and frequency using outdoor mesocosm communities containing larval amphibians (Rana sylvatica and Bufo americanus) and using a commercial formulation of the herbicide glyphosate (Roundup Original MAX(R)). Consistent with past studies, exposures of up to 3 mg acid equivalent (a.e.)/L caused substantial amphibian death. However, the amount of death was considerably higher when the herbicide was applied earlier in the experiment than later in the experiment. Single, large applications (at different times) had larger effects on tadpole mortality and growth than multiple, small applications (of the same total amount). The results may reflect an acclimation to the herbicide over time. In treatments with high tadpole mortality, there was no resulting increase in periphyton, suggesting that the reduction in tadpole herbivory might have been offset by direct negative impacts of the herbicide. We also discovered that temperature stratification caused herbicide stratification, with higher concentrations near the surface. Such stratification has important implications to the habitat choices of ectotherms that might prefer surface waters for thermoregulation or prefer deeper waters to avoid predators. Collectively, the present study demonstrates the importance of examining multiple applications times and frequencies to understand the impacts of pesticides on organisms.

Phylogeny meets ecotoxicology: evolutionary patterns of sensitivity to a common insecticide

Pesticides commonly occur in aquatic systems and pose a substantial challenge to the conservation of many taxa. Ecotoxicology has traditionally met this challenge by focusing on short‐term, single‐species tests and conducting risk assessments based on the most sensitive species tested. Rarely have ecotoxicology data been examined from an evolutionary perspective, and to our knowledge, there has never been a phylogenetic analysis of sensitivity, despite the fact that doing so would provide insights into patterns of sensitivity among species and identify which clades are the most sensitive to a particular pesticide. We examined phylogenetic patterns of pesticide sensitivity in amphibians, a group of conservation concern owing to global population declines. Using the insecticide endosulfan, we combined previously published results across seven species of tadpoles and added eight additional species from the families Bufonidae, Hylidae, and Ranidae. We found significant phylogenetic signal in the sensitivity to the insecticide and in the existence of time lag effects on tadpole mortality. Bufonids were less sensitive than hylids, which were less sensitive than the ranids. Moreover, mortality time lags were common in ranids, occasional in hylids, and rare in bufonids. These results highlight the importance of an evolutionary perspective and offer important insights for conservation.

Exploring the effects of individual traits and within-colony variation on task differentiation and collective behavior in a desert social spider

Social animals are extraordinarily diverse and ecologically abundant. In understanding the success of complex animal societies, task differentiation has been identified as a central mechanism underlying the emergence and performance of adaptive collective behaviors. In this study, we explore how individual differences in behavior and body size determine task allocation in the social spider Stegodyphus dumicola. We found that individuals with high body condition indices were less likely to participate in prey capture, and individuals’ tendency to engage in prey capture was not associated with either their behavioral traits or body size. No traits were associated with individuals’ propensity to participation in web repair, but small individuals were more likely to engage in standard web-building. We also discovered consistent, differences among colonies in their collective behavior (i.e., colony-level personality). At the colony level, within-colony variation in behavior (aggressiveness) and body size were positively associated with aggressive foraging behavior. Together, our findings reveal a subtly complex relationship between individual variation and collective behavior in this species. We close by comparing the relationship between individual variation and social organization in nine species of social spider. We conclude that intraspecific variation is a major force behind the social organization of multiple independently derived lineages of social spider.

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.

Induced tolerance from a sublethal insecticide leads to cross-tolerance to other insecticides.

As global pesticide use increases, the ability to rapidly respond to pesticides by increasing tolerance has important implications for the persistence of nontarget organisms. A recent study of larval amphibians discovered that increased tolerance can be induced by an early exposure to low concentrations of a pesticide. Since natural systems are often exposed to a variety of pesticides that vary in mode of action, we need to know whether the induction of increased tolerance to one pesticide confers increased tolerance to other pesticides. Using larval wood frogs (Lithobates sylvaticus), we investigated whether induction of increased tolerance to the insecticide carbaryl (AChE-inhibitor) can induce increased tolerance to other insecticides that have the same mode of action (chlorpyrifos, malathion) or a different mode of action (Na(+)channel-interfering insecticides; permethrin, cypermethrin). We found that embryonic exposure to sublethal concentrations of carbaryl induced higher tolerance to carbaryl and increased cross-tolerance to malathion and cypermethrin but not to chlorpyrifos or permethrin. In one case, the embryonic exposure to carbaryl induced tolerance in a nonlinear pattern (hormesis). These results demonstrate that that the newly discovered phenomenon of induced tolerance also provides induced cross-tolerance that is not restricted to pesticides with the same mode of action.

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.

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.