Negative bioenergetic responses to pesticides in damselfly larvae are more likely when it is hotter and when temperatures fluctuate.

To make more realistic predictions about the current and future effects of pesticides, we need to better understand physiological mechanisms associated with the widespread higher toxicity of many pesticides under increasing mean temperatures and daily temperature fluctuations (DTFs). One overlooked, yet insightful, mechanism are bioenergetic responses as these provide information about the balance between energy gains and costs. Therefore, we studied how the bioenergetic responses to the insecticide chlorpyrifos were affected by a higher mean temperature and a higher DTF in Ischnura elegans damselfly larvae. To quantify bioenergetic responses we measured energy availability (Ea), energy consumption (Ec) and total net energy budget (cellular energy allocation, CEA). Exposure to chlorpyrifos considerably reduced CEA values when a high mean temperature was combined with a high DTF (up to -18%). Notably, chlorpyrifos had little effect on CEA at a constant 20 °C, meaning that the bioenergetic impact of chlorpyrifos would have been underestimated if we had only tested under standard testing conditions. The chlorpyrifos-induced reductions in CEA under warming were driven by reductions in Ea (up to -16%, mainly through large reductions in sugar and fat contents) while Ec was unaffected by chlorpyrifos. Treatment groups with a lower CEA value showed a higher mortality and a lower growth rate, indicating bioenergetic responses are contributing to the higher toxicity of chlorpyrifos under warming. Our study highlights the importance of evaluating the effects of pesticides under an increase in both mean temperature and DTF to improve the ecological risk assessment of pesticides under global warming.

Shrinking Body Size and Physiology Contribute to Geographic Variation and the Higher Toxicity of Pesticides in a Warming World.

To improve current and future risk assessment of pesticides under global warming, mechanistic insights and consideration of daily temperature fluctuations (DTFs) are needed. One overlooked mechanism how both higher mean temperatures and DTFs may increase toxicity is by reducing body size (temperature-size-rule). We studied whether a higher mean temperature and DTF magnified chlorpyrifos toxicity in Ischnura elegans damselfly larvae, and whether this was mediated by temperature-induced reductions in body size and/or physiological changes. The lethal effects of chlorpyrifos were magnified at the high mean temperature (up to ∼15%) and under DTF (up to ∼33%), and especially at their combination (up to ∼46%) indicating synergisms. This highlights that not only considering DTFs, but also their interaction with higher mean temperatures is pivotal for realistic predictions of pesticide toxicity. Both higher temperatures and DTFs resulted in smaller larvae, which were more sensitive to chlorpyrifos. Notably, the DTF-induced smaller body sizes, as well as the higher oxidative damage to lipids, contributed to the higher chlorpyrifos toxicity under DTF. By integrating the temperature-size rule and size-pesticide sensitivity pattern we provide proof-of-principle for a novel, likely general mechanism contributing to geographic variation and the higher toxicity of pesticides in a warming world.

A widespread morphological antipredator mechanism reduces the sensitivity to pesticides and increases the susceptibility to warming.

Pollution and predation are two omnipresent stressors in aquatic systems that can interact in multiple ways, thereby challenging accurate assessment of the effects of pollutants in natural systems. Despite the widespread occurrence of morphological antipredator mechanisms, no studies have tested how these can affect the sensitivity of prey to pesticides. Sensitivity to pesticides is typically measured via reductions in growth rates and survival, but also reductions in heat tolerance are to be expected and are becoming increasingly important in a warming world. We investigated how autotomy, a widespread morphological antipredator mechanism where animals sacrifice a body part (here the caudal lamellae) to escape when attacked by a predator, modified the sensitivity to the insecticide chlorpyrifos in larvae of the damselfly Coenagrion puella. Exposure to chlorpyrifos reduced the growth rate and heat tolerance (measured as CTmax). A key finding was that the pesticide had a greater impact on growth rates of intact animals, i.e. those that retained their lamellae. This reduced sensitivity to chlorpyrifos in animals without lamellae can be explained by the reduced outer surface area which is expected to result in a lower uptake of the pesticide. Larvae that underwent autotomy exhibited a lower heat tolerance, which may also be explained by the reduced surface area and the associated reduction in oxygen uptake. There is a wide diversity of morphological antipredator mechanisms, suggesting that there will be more examples where these mechanisms affect the vulnerability to pollutants. Given the importance of pollution and predation as structuring forces in aquatic food webs, exploring the potential interactions between morphological antipredator mechanisms and sensitivity to pollutants will be crucial for risk assessment of pollutants in aquatic systems.

Combined effects of cadmium exposure and temperature on the annual killifish (Nothobranchius furzeri).

Freshwater organisms are increasingly exposed to combinations of stressors. However, because it is time-consuming and costly, research on the interaction of stressors, such as compound toxicity and global warming on vertebrates, is scarce. Studies on multigenerational effects of these combined stressors are almost nonexistent. In the present study, we tested the combined effects of 4 °C warming and cadmium (Cd) exposure on life-history traits, biomarkers, bioaccumulation, and multigenerational tolerance in the turquoise killifish, Nothobranchius furzeri. The extremely short life cycle of this vertebrate model allows for assessment of sublethal and multigenerational effects within 4 mo. The applied Cd concentrations had only limited effects on the measured endpoints, which suggests that N. furzeri is more resistant to Cd than fathead minnow and rainbow trout. In contrast, the temperature increase of 4 °C was stressful: it delayed female maturation and lowered adult mass and fecundity. Finally, indications of synergistic effects were found on peak fecundity and embryonic survival. Overall, these results indicate the importance of studying chronic and multigenerational effects of combined stressors. Environ Toxicol Chem 2018;37:2361-2371. © 2018 SETAC.

Strong differences between two congeneric species in sensitivity to pesticides in a warming world.

To predict the impact of pesticides in a warming world we need to know how species differ in the interaction pathways between pesticides and warming. Trait-based approaches have been successful in identifying the 'pace of life' and body size as predictors of sensitivity to pesticides among distantly related species. However, it remains to be tested whether these traits allow predicting differences in sensitivity to pesticides between closely related species, and in the strength of the interaction pathways between pesticides and warming. We tested the effects of multiple pulses of chlorpyrifos (allowing accumulation) under warming on key life history traits, heat tolerance (CTmax) and physiology of two congeneric damselfly species: the fast-paced (fast growth and development, high metabolic rate), small Ischnura pumilio and the slow-paced, large I. elegans. Chlorpyrifos reduced survival and growth, but contrary to current trait-based predictions I. pumilio was 8× less sensitive than I. elegans. The lower sensitivity of I. pumilio could be explained by a higher fat content, and higher activities of acetylcholinesterase and of detoxifying and anti-oxidant enzymes. While for I. pumilio the effect of chlorpyrifos was small and did not depend on temperature, for I. elegans the impact was higher at 20°C compared to 24°C. This matches the higher pesticide accumulation in the water after multiple pulses at 20°C than at 24°C. The expected reduction in heat tolerance after pesticide exposure was present in I. elegans but not in I. pumilio. Our results demonstrate that closely related species can have very different sensitivities to a pesticide resulting in species-specific support for the "toxicant-induced climate change sensitivity" and the "climate-induced toxicant sensitivity" interaction pathways. Our results highlight that trait-based approaches can be strengthened by integrating physiological traits.

Food level and sex shape predator-induced physiological stress: immune defence and antioxidant defence.

Despite the potential impact on prey fitness and predator-prey interactions, most studies of predation risk ignore physiological responses and their dependence upon food level and sex. Therefore, we reared male and female larvae of the damselfly Lestes viridis under predator stress (dragonfly larvae) at high and low food levels, and subsequently scored for important variables of insect immune defence (i.e. phenoloxidase) and antioxidant defence [i.e. superoxide dismutase, and catalase (CAT)]. Under predation risk, larvae did not decrease growth rate or immune defence, and only slightly reduced food intake in the high food treatment, probably because of time stress, i.e. little time available to complete the larval development. However, larvae facing predator stress did show an upregulation of antioxidant enzymes. This upregulation was dependent upon food level for CAT and both food level and sex for SOD, consistent with energetic constraints and sex differences in the link between longevity and adult fitness. Our results illustrate that predator stress can influence life history, behavioural and physiological responses differentially and in a context-dependent way. This implies that non-consumptive physiological effects of predators on their prey show independent yet similar complexities in behavioural and life history response variables. In general, our results advocate that mechanistic studies on predator-prey interactions may benefit from including physiological variables.

Water turbidity affects predator-prey interactions in a fish-damselfly system.

Community structure may differ dramatically between clear-water and turbid lakes. These differences have been attributed to differences in the cascading effect of fish on prey populations, owing to the reduced efficiency of fish predation in the presence of macrophytes. However, recent theoretical ideas suggest that water turbidity may shape predator-prey interactions, and it is predicted that prey will relax its antipredation behaviour in turbid water (H1). As a result, the nature of predator-prey interactions is expected to shift from both direct and indirect in clear water to dominantly direct in turbid water (H2). We tested these ideas in a fish-damselfly predator-prey system. In a first behavioural experiment, we looked at antipredation behaviour of damselfly larvae isolated from habitats that differ in turbidity, in the presence of fish in clear and turbid water. As predicted in H1, the larvae were more active in turbid than in clear water. In a complementary enclosure experiment, we reared larvae in a clear-water pond and a turbid pond, respectively, and manipulated the origin of the larvae (clear-water, turbid pond), fish presence (absent, present), and vegetation density (sparse, abundant). In both ponds, fish had a direct negative effect on survival of the larvae, which was mitigated in the presence of vegetation. In the fish treatment, the change in average body mass tended to be higher in the turbid pond than in the clear-water pond, suggesting indirect effects of fish were mitigated in the turbid pond. This was supported by a negative effect of fish on the effective growth rate of larvae in the clear pond, but not in the turbid pond. These results are compatible with the idea that predator-prey relationships are mainly governed by direct effects in turbid water, and by direct and indirect effects in clear water.