Phylogenomic and functional characterization of an evolutionary conserved cytochrome P450-based insecticide detoxification mechanism in bees.

The regulatory process for assessing the risks of pesticides to bees relies heavily on the use of the honeybee, Apis mellifera, as a model for other bee species. However, the validity of using A. mellifera as a surrogate for other Apis and non-Apis bees in pesticide risk assessment has been questioned. Related to this line of research, recent work on A. mellifera has shown that specific P450 enzymes belonging to the CYP9Q subfamily act as critically important determinants of insecticide sensitivity in this species by efficiently detoxifying certain insecticide chemotypes. However, the extent to which the presence of functional orthologs of these enzymes is conserved across the diversity of bees is unclear. Here we used a phylogenomic approach to identify > 100 putative CYP9Q functional orthologs across 75 bee species encompassing all major bee families. Functional analysis of 26 P450s from 20 representative bee species revealed that P450-mediated detoxification of certain systemic insecticides, including the neonicotinoid thiacloprid and the butenolide flupyradifurone, is conserved across all major bee pollinator families. However, our analyses also reveal that CYP9Q-related genes are not universal to all bee species, with some Megachilidae species lacking such genes. Thus, our results reveal an evolutionary conserved capacity to metabolize certain insecticides across all major bee families while identifying a small number of bee species where this function may have been lost. Furthermore, they illustrate the potential of a toxicogenomic approach to inform pesticide risk assessment for nonmanaged bee species by predicting the capability of bee pollinator species to break down synthetic insecticides.

Effects of exposure to the neonicotinoid pesticide clothianidin on mouse intestinal microbiota under unpredictable environmental stress.

Recent research has demonstrated the toxicity of neonicotinoid pesticides (NNs) in mammals through their interaction with nicotinic acetylcholine receptors (nAChRs). These effects are reported to extend to the intestinal microbiota as well. In addition, environmental stress affects the expression of nAChRs, which may alter sensitivity to NNs. In this study, we analyzed the intestinal microbiota of mice exposed to clothianidin (CLO), a type of NN, under environmental stress, and aimed to clarify the effects of such combined exposure on the intestinal microbiota. C57BL/6N male mice (9 weeks old) were subchronically administered a no-observed-adverse-effect-level (NOAEL) CLO-mixed rehydration gel for 29 days and simultaneously subjected to chronic unpredictable mild stress (CUMS). After the administration period, cecum contents were collected and analyzed by 16S rRNA sequencing for intestinal microbiota. CLO exposure alone resulted in alterations in the relative abundance of Alistipes and ASF356, which produce short-chain fatty acids. The addition of CUMS amplified these changes. On the other hand, CLO alone did not affect the relative abundance of Lactobacillus, but the abundance decreased when CUMS was added. This study revealed that the combined exposure to CLO and stress not only amplifies their individual effects on intestinal microbiota but also demonstrates combined and multifaceted toxicities.

Quantification of the tissue distribution and accumulation of the neonicotinoid pesticide clothianidin and its metabolites in maternal and fetal mice.

Neonicotinoids (NNs) are commonly used pesticides that have a selective agonistic action on insect nicotinic acetylcholine receptors. Recent evidence has shown that NNs have adverse effects in the next generation of mammals, but it remains unclear how NNs transferred from dams to fetuses are distributed and accumulated in fetal tissues. Here, we aimed to clarify the tissue distribution and accumulation properties of the NN clothianidin (CLO) and its 6 metabolites in 7 tissues and blood in both dams and fetuses of mice administered CLO for a single day or for 9 consecutive days. The results showed that the total concentrations of CLO-related compounds in the brain and kidney were higher in fetuses than in dams, whereas in the liver, heart, and blood they were lower in fetuses. The multi-day administration increased the total levels in heart and blood only in the fetuses of the single administration group. In addition, dimethyl metabolites of CLO showed fetus/dam ratios >1 in some tissues, suggesting that fetuses have higher accumulation property and are thus at higher risks of exposure to CLO-related compounds than dams. These findings revealed differences in the tissue-specific distribution patterns of CLO and its metabolites between dams and fetuses, providing new insights into the assessment of the developmental toxicity of NNs.

Ecotoxicological impact of dinotefuran insecticide and its metabolites on non-targets in agroecosystem: Harnessing nanotechnology- and bio-based management strategies to reduce its impact on non-target ecosystems.

The class of insecticides known as neonicotinoid insecticides has gained extensive application worldwide. Two characteristics of neonicotinoid pesticides are excellent insecticidal activity and a wide insecticidal spectrum for problematic insects. Neonicotinoid pesticides can also successfully manage pest insects that have developed resistance to other insecticide classes. Due to its powerful insecticidal properties and rapid plant absorption and translocation, dinotefuran, the most recent generation of neonicotinoid insecticides, has been widely used against biting and sucking insects. Dinotefuran has a wide range of potential applications and is often used globally. However, there is growing evidence that they negatively impact the biodiversity of organisms in agricultural settings as well as non-target organisms. The objective of this review is to present an updated summary of current understanding regarding the non-target effects of dinotefuran; we also enumerated nano- and bio-based mitigation and management strategies to reduce the impact of dinotefuran on non-target organisms and to pinpoint knowledge gaps. Finally, future study directions are suggested based on the limitations of the existing studies, with the goal of providing a scientific basis for risk assessment and the prudent use of these insecticides.

Transport of polyethylene and polypropylene microplastics under the action of agricultural chemicals: Role of pesticide adjuvants and neonicotinoid active ingredients.

Understanding the impact of various agricultural chemical components on the fate and transport of microplastics (MPs) in the subsurface is essential. In this study, column experiments on saturated porous media were conducted to explore the influence of the coexistence environment of pesticide adjuvants (surfactants) and active ingredients (neonicotinoids) on the transport of polyethylene (PE) and polypropylene (PP) MPs. An anionic surfactant (sodium dodecyl sulfate (SDS)), a nonionic surfactant (nonylphenol ethoxylate (NP-40)), and three neonicotinoid insecticides (acetamiprid, dinotefuran, and nitenpyram) could independently increase MP migration by 9.31%-61.01% by improving the hydrophilicity. Acetamiprid or dinotefuran reduced the adhesion work of the binary system by competing with SDS for adsorption sites, thereby inhibiting PE mobility. However, nitenpyram in the mixture was not easily adsorbed on the surface of PE MPs together with SDS because of nitenpyram's high hydrophilicity. Neonicotinoid molecules could not reduce the hydrophilic modification of SDS on PP MPs by competing for adsorption sites. Owing to their weak charge and adhesion work of nonionic surfactants (-4.80 mV and 28.45 kT for PE and -8.21 mV and 17.64 kT for PP), neonicotinoids tended to occupy the adsorption sites originally belonging to NP-40. The long molecular chain of NP-40 made it difficult for high-concentration neonicotinoids to affect the adhesion on MPs. In addition, NP-40 was harder to peel off from the MP surface than SDS, leading to a larger MP transport ability in the sand column.

Contaminants of emerging concern in water and sediment of the Venice Lagoon, Italy.

This study investigates for the first time the contamination of water and sediment of the Venice Lagoon by twenty Contaminants of Emerging Concern (CECs): three hormones, six pharmaceutical compounds (diclofenac and five antibiotics, three of which are macrolides), nine pesticides (methiocarb, oxadiazon, metaflumizone, triallate, and five neonicotinoids), one antioxidant (BHT), and one UV filter (EHMC). Water and sediment samples were collected in seven sites in four seasons, with the aim of investigating the occurrence, distribution, and possible emission sources of the selected CECs in the studied transitional environment. The most frequently detected contaminants in water were neonicotinoid insecticides (with a frequency of quantification of single contaminants ranging from 73% to 92%), and EHMC (detected in the 77% of samples), followed by BHT (42%), diclofenac (39%), and clarithromycin (35%). In sediment the highest quantification frequencies were those of BHT (54%), estrogens (ranging from 35% to 65%), and azithromycin (46%). Although this baseline study does not highlight seasonal or spatial trends, results suggested that two of the major emission sources of CECs in the Venice Lagoon could be tributary rivers from its drainage basin and treated wastewater, due to the limited removal rates of some CECs in WWTPs. These preliminary results call for further investigations to better map priority emission sources and improve the understanding of CECs environmental behavior, with the final aim of drawing up a site-specific Watch List of CECs for the Venice Lagoon and support the design of more comprehensive monitoring plans in the future.

IPM reduces insecticide applications by 95% while maintaining or enhancing crop yields through wild pollinator conservation.

Pest management practices in modern industrial agriculture have increasingly relied on insurance-based insecticides such as seed treatments that are poorly correlated with pest density or crop damage. This approach, combined with high invertebrate toxicity for newer products like neonicotinoids, makes it challenging to conserve beneficial insects and the services that they provide. We used a 4-y experiment using commercial-scale fields replicated across multiple sites in the midwestern United States to evaluate the consequences of adopting integrated pest management (IPM) using pest thresholds compared with standard conventional management (CM). To do so, we employed a systems approach that integrated coproduction of a regionally dominant row crop (corn) with a pollinator-dependent specialty crop (watermelon). Pest populations, pollination rates, crop yields, and system profitability were measured. Despite higher pest densities and/or damage in both crops, IPM-managed pests rarely reached economic thresholds, resulting in 95% lower insecticide use (97 versus 4 treatments in CM and IPM, respectively, across all sites, crops, and years). In IPM corn, the absence of a neonicotinoid seed treatment had no impact on yields, whereas IPM watermelon experienced a 129% increase in flower visitation rate by pollinators, resulting in 26% higher yields. The pollinator-enhancement effect under IPM management was mediated entirely by wild bees; foraging by managed honey bees was unaffected by treatments and, overall, did not correlate with crop yield. This proof-of-concept experiment mimicking on-farm practices illustrates that cropping systems in major agricultural commodities can be redesigned via IPM to exploit ecosystem services without compromising, and in some cases increasing, yields.

Associations between maternal serum neonicotinoid pesticide exposure during pregnancy and newborn telomere length: Effect modification by sampling season.

BACKGROUND: An increasing amount of evidence suggests that telomere length (TL) at birth can predict lifespan and is associated with chronic diseases later in life, but newborn TL may be affected by environmental pollutants. Neonicotinoids (NEOs) are widely used worldwide, and despite an increasing number of studies showing that they may have adverse effects on birth in mammals and even humans, few studies have examined the effect of NEO exposure on newborn TLs. OBJECTIVE: To investigate the effects of prenatal exposure to NEOs and the interactions between NEOs and sampling season on newborn TL. METHODS: We conducted a prospective cohort study of 500 mother-newborn pairs from the Guangxi Zhuang Birth Cohort. Ultraperformance liquid chromatographymass spectrometry was used to detect ten NEOs in maternal serum, and fluorescence quantitative PCR was used to estimate the newborn TL. A generalized linear model (GLM) was used to evaluate the relationships between individual NEO exposures and TLs , and quantile g-computation (Qgcomp) model and Bayesian kernel machine regression (BKMR) model were used to evaluate the combined effect of mixtures of components. RESULTS: The results of the GLM showed that compared with maternal TMX levels < LOD, maternal TMX levels < median were negatively correlated with newborn TL (-6.93%, 95% CI%: -11.92%, -1.66%), and the decrease in newborn TL was more pronounced in girls (-9.60%, 95% CI: -16.84%, -1.72%). Moreover, different kinds of maternal NEO exposure had different effects on newborn TL in different sampling seasons, and the effect was statistically significant in all seasons except in autumn. Mixed exposure analysis revealed a potential positive trend between NEOs and newborn TL, but the association was not statistically significant. CONCLUSION: Prenatal exposure to TMX may shorten newborn TL, and this effect is more pronounced among female newborns. Furthermore, the relationship between NEO exposure and TL may be modified by the sampling season.

Modeling temperature-dependent life-cycle toxicity of thiamethoxam in Chironomus riparius using a DEB-TKTD model.

The neonicotinoid insecticide thiamethoxam (TMX) is widely used to protect crops against insect pests. Despite some desirable properties such as its low toxicity to birds and mammals, concerns have been raised about its toxicity to non-target arthropods, including freshwater insects like chironomids. Whereas multiple studies have investigated chronic effects of neonicotinoids in chironomid larvae at standardized laboratory conditions, a better understanding of their chronic toxicity under variable temperatures and exposure is needed for coherent extrapolation from the laboratory to the field. Here, we developed a quantitative mechanistic effect model for Chironomus riparius, to simulate the species' life history under dynamic temperatures and exposure concentrations of TMX. Laboratory experiments at four different temperatures (12, 15, 20, 23 °C) and TMX concentrations between 4 and 51 µg/L were used to calibrate the model. Observed concentration-dependent effects of TMX in C. riparius included slower growth, later emergence, and higher mortality rates with increasing concentrations. Furthermore, besides a typical accelerating effect on the organisms' growth and development, higher temperatures further increased the effects associated with TMX. With some data-informed modeling decisions, most prominently the inclusion of a size dependence that makes larger animals more sensitive to TMX, the model was parametrized to convincingly reproduce the data. Experiments at both a constant (20 °C) and a dynamically increasing temperature (15-23 °C) with pulsed exposure were used to validate the model. Finally, the model was used to simulate realistic exposure conditions using two reference exposure scenarios measured in Missouri and Nebraska, utilizing a moving time window (MTW) and either a constant temperature (20 °C) or the measured temperature profiles belonging to each respective scenario. Minimum exposure multiplication factors leading to a 10% effect (EP10) in the survival at pupation, i.e., the most sensitive endpoint found in this study, were 25.67 and 21.87 for the Missouri scenario and 38.58 and 44.64 for the Nebraska scenario, when using the respective temperature assumptions. While the results illustrate that the use of real temperature scenarios does not systematically modify the EPx in the same direction (making it either more or less conservative when used as a risk indicator), the advantage of this approach is that it increases the realism and thus reduces the uncertainty associated with the model predictions.

Identification of the cytochrome P450 gene AccCYP6A13 in Apis cerana cerana and its response to environmental stress.

Cytochrome P450 plays a crucial role in regulating insect growth, development, and resisting a variety of stresses. Insect metamorphosis and response to external stress are altered by deleting CYP450 genes. In this study, we identified and analyzed a novel gene of CYP450 family, AccCYP6A13, from Apis cerana cerana, and explored its role in the response of Apis cerana cerana to adverse external stressors. It was found that the expression of AccCYP6A13 was spatiotemporal specificity. The expression level increased with age and reached its highest value in the adult stage. The primarily expressiong location were legs, brain, and epidermis of honeybees. Stress conditions can affect the expression of AccCYP6A13 depending on treatment times. RNA interference experiments have shown that knocking down AccCYP6A13 reduces antioxidant activity and deactivates detoxification enzymes, resulting in oxidative damage accumulation and a decline in detoxification capability in bees, as well as inhibiting numerous antioxidant genes. Additionally, knockdown of the AccCYP6A13 gene in Apis cerana cerana resulted in increased sensitivity to pesticides and increased mortality when treated with neonicotinoid pesticides such as thiamethoxam. AccCYP6A13 overexpression in a prokaryotic system further confirmed its role in resistance to oxidative stress. To summarize, AccCYP6A13 may play an essential role in the normal development and response to environmental stress in Apis cerana cerana. Furthermore, this study contributed to the theoretical understanding of bee resistance biology.

Design, Synthesis, and Synergistic Activities of Eight-Membered Carbon Bridged Neonicotinoid Derivatives.

Insecticide synergists are an effective approach to increase the control efficacy and reduce active ingredient usage. In order to explore neonicotinoid-specific synergists with novel scaffolds and higher potency, a series of eight-membered carbon bridged neonicotinoid derivatives were designed and synthesized in accordance with our previous research. The synergistic effects of the target compounds on neonicotinoids in Aphis craccivora were evaluated, and the structure-activity relationships were summarized. The results indicated that most of the target compounds exhibited significant synergistic effects on imidacloprid in A. craccivora at low concentrations. In particular, compound 1 at a concentration of 1 mg/L reduced the LC50 value of imidacloprid from 0.856 mg/L to 0.170 mg/L. Meanwhile, compound 1 also increased the insecticidal activity of most neonicotinoid insecticides belonging to the Insecticide Resistance Action Committee (IRAC) 4 A subgroup against A. craccivora. The present study might be meaningful for directing the design of neonicotinoid-specific synergists.

Neonicotinoid exposure increases Varroa destructor (Mesostigmata: Varroidae) mite parasitism severity in honey bee colonies and is not mitigated by increased colony genetic diversity.

Agrochemical exposure is a major contributor to ecological declines worldwide, including the loss of crucial pollinator species. In addition to direct toxicity, field-relevant doses of pesticides can increase species' vulnerabilities to other stressors, including parasites. Experimental field demonstrations of potential interactive effects of pesticides and additional stressors are rare, as are tests of mechanisms via which pollinators tolerate pesticides. Here, we controlled honey bee colony exposure to field-relevant concentrations of 2 neonicotinoid insecticides (clothianidin and thiamethoxam) in pollen and simultaneously manipulated intracolony genetic heterogeneity. We showed that exposure increased rates of Varroa destructor (Anderson and Trueman) parasitism and that while increased genetic heterogeneity overall improved survivability, it did not reduce the negative effect size of neonicotinoid exposure. This study is, to our knowledge, the first experimental field demonstration of how neonicotinoid exposure can increase V. destructor populations in honey bees and also demonstrates that colony genetic diversity cannot mitigate the effects of neonicotinoid pesticides.

Surface-Enhanced Raman Spectroscopy Using a Silver Nanostar Substrate for Neonicotinoid Pesticides Detection.

Surface-enhanced Raman spectroscopy (SERS) has been introduced to detect pesticides at low concentrations and in complex matrices to help developing countries monitor pesticides to keep their concentrations at safe levels in food and the environment. SERS is a surface-sensitive technique that enhances the Raman signal of molecules absorbed on metal nanostructure surfaces and provides vibrational information for sample identification and quantitation. In this work, we report the use of silver nanostars (AgNs) as SERS-active elements to detect four neonicotinoid pesticides (thiacloprid, imidacloprid, thiamethoxam and nitenpyram). The SERS substrates were prepared with multiple depositions of the nanostars using a self-assembly approach to give a dense coverage of the AgNs on a glass surface, which ultimately increased the availability of the spikes needed for SERS activity. The SERS substrates developed in this work show very high sensitivity and excellent reproducibility. Our research opens an avenue for the development of portable, field-based pesticide sensors, which will be critical for the effective monitoring of these important but potentially dangerous chemicals.

Clinical Profile and Predictors of Intensive Care Admission in Neonicotinoid Poisoning in a Tertiary Care Hospital in South India.

Background and objectives: Neonicotinoids are a newer class of pesticides that are believed to cause predominantly mild toxicity in humans. This study aimed to describe the clinical features of neonicotinoid poisoning and identify predictors of severe toxicity. Materials and methods: This retrospective study included all patients with neonicotinoid poisoning admitted to a Tertiary Care Center in India over an 18-year period. Clinical and laboratory features were compared against outcomes to identify predictors of the need for intensive care admission. Results: Twenty-eight patients were included in the study of which 28.6% had severe disease requiring ICU admission. A higher respiratory rate, blood lactate level, SOFA, and qSOFA scores as well as a lower Glasgow coma score at presentation predicted ICU admission. First-generation compounds and imidacloprid consumption were associated with longer ICU stays and a longer duration of invasive ventilation. Conclusion: Neonicotinoid compounds can cause significant toxicity with oral ingestion. Imidacloprid and other first-generation compounds were associated with more severe toxicity requiring intensive care. Simple clinical parameters assessed at presentation can be used to predict severe disease and the need for ICU care. Larger, prospective studies are required to confirm these findings. How to cite this article: Sanga L, Jacob A, Jayakaran JAJ, Iyadurai R. Clinical Profile and Predictors of Intensive Care Admission in Neonicotinoid Poisoning in a Tertiary Care Hospital in South India. Indian J Crit Care Med 2024;28(1):66-69.

Aiming for a Better Tomorrow.

How to cite this article: Kharbanda M. Aiming for a Better Tomorrow. Indian J Crit Care Med 2024;28(1):11-12.

Soil microbial gene expression in an agricultural ecosystem varies with time and neonicotinoid seed treatments.

Neonicotinoids, a class of systemic insecticides, have been widely used for decades against various insect pests. Previous studies have reported non-target effects of neonicotinoids on some beneficial macro- and micro-organisms. Considering the crucial role the soil microbiota plays in sustaining soil fertility, it is critical to understand how neonicotinoid exposure affects the microbial taxonomic composition and gene expression. However, most studies to date have evaluated soil microbial taxonomic compositions or assessed microbial functions based on soil biochemical analysis. In this study, we have applied a metatranscriptomic approach to quantify the variability in soil microbial gene expression in a 2 year soybean/corn crop rotation in Quebec, Canada. We identified weak and temporally inconsistent effects of neonicotinoid application on soil microbial gene expression, as well as a strong temporal variation in soil microbial gene expression among months and years. Neonicotinoid seed treatment altered the expression of a small number of microbial genes, including genes associated with heat shock proteins, regulatory functions, metabolic processes and DNA repair. These changes in gene expression varied during the growing season and between years. Overall, the composition of soil microbial expressed genes seems to be more resilient and less affected by neonicotinoid application than soil microbial taxonomic composition. Our study is among the first to document the effects of neonicotinoid seed treatment on microbial gene expression and highlights the strong temporal variability of soil microbial gene expression and its responses to neonicotinoid seed treatments.

Colony size buffers interactions between neonicotinoid exposure and cold stress in bumblebees.

Social bees are critical for supporting biodiversity, ecosystem function and crop yields globally. Colony size is a key ecological trait predicted to drive sensitivity to environmental stressors and may be especially important for species with annual cycles of sociality, such as bumblebees. However, there is limited empirical evidence assessing the effect of colony size on sensitivity to environmental stressors or the mechanisms underlying these effects. Here, we examine the relationship between colony size and sensitivity to environmental stressors in bumblebees. We exposed colonies at different developmental stages briefly (2 days) to a common neonicotinoid (imidacloprid) and cold stress, while quantifying behaviour of individuals. Combined imidacloprid and cold exposure had stronger effects on both thermoregulatory behaviour and long-term colony growth in small colonies. We find that imidacloprid's effects on behaviour are mediated by body temperature and spatial location within the nest, suggesting that social thermoregulation provides a buffering effect in large colonies. Finally, we demonstrate qualitatively similar effects in size-manipulated microcolonies, suggesting that group size per se, rather than colony age, drives these patterns. Our results provide evidence that colony size is critical in driving sensitivity to stressors and may help elucidate mechanisms underlying the complex and context-specific impacts of pesticide exposure.

Land-use-associated stressors interact to reduce bumblebee health at the individual and colony level.

In agricultural landscapes, bees face a variety of stressors, including insecticides and poor-quality food. Although both stressors individually have been shown to affect bumblebee health negatively, few studies have focused on stressor interactions, a scenario expected in intensively used agricultural landscapes. Using the bumblebee Bombus terrestris, a key pollinator in agricultural landscapes, we conducted a fully factorial laboratory experiment starting at nest initiation. We assessed the effects of food quality and insecticides, alone and in interaction, on health traits at various levels, some of which have been rarely studied. Pollen with a diluted nutrient content (low quality) reduced ovary size and delayed colony development. Wing asymmetry, indicating developmental stress, was increased during insecticide exposure and interactions with poor food, whereas both stressors reduced body size. Both stressors and their interaction changed the workers' chemical profile and reduced worker interactions and the immune response. Our findings suggest that insecticides combined with nutritional stress reduce bumblebee health at the individual and colony levels, thus possibly affecting colony performance, such as development and reproduction, and the stability of plant-pollinator networks. The synergistic effects highlight the need of combining stressors in risk assessments and when studying the complex effects of anthropogenic stressors on health outcomes.

Expression of subunits of an insecticide target receptor varies across tissues, life stages, castes, and species of social bees.

Global losses of insects jeopardize ecosystem stability and crop pollination. Robust evidence indicates that insecticides have contributed to these losses. Notably, insecticides targeting nicotinic acetylcholine receptors (nAChRs) have neurotoxic effects on beneficial insects. Because each nAChR consists of five subunits, the alternative arrangements of subunits could create a multitude of receptors differing in structure and function. Therefore, understanding whether the use of subunits varies is essential for evaluating and predicting the effects of insecticides targeting such receptors. To better understand how the use and composition of nAChRs differ within and between insect pollinators, we analysed RNA-seq gene expression data from tissues and castes of Apis mellifera honey bees and life stages and castes of the Bombus terrestris bumble bees. We reveal that all analysed tissues express nAChRs and that relative expression levels of nAChR subunits vary widely across almost all comparisons. Our work thus shows fine-tuned spatial and temporal expression of nAChRs. Given that coexpression of subunits underpins the compositional diversity of functional receptors and that the affinities of insecticides depend on nAChR composition, our findings provide a likely mechanism for the various damaging effects of nAChR-targeting insecticides on insects. Furthermore, our results indicate that the appraisal of insecticide risks should carefully consider variation in molecular targets.

Toxic temperatures: Bee behaviours exhibit divergent pesticide toxicity relationships with warming.

Climate change and agricultural intensification are exposing insect pollinators to temperature extremes and increasing pesticide usage. Yet, we lack good quantification of how temperature modulates the sublethal effects of pesticides on behaviours vital for fitness and pollination performance. Consequently, we are uncertain if warming decreases or increases the severity of different pesticide impacts, and whether separate behaviours vary in the direction of response. Quantifying these interactive effects is vital in forecasting pesticide risk across climate regions and informing pesticide application strategies and pollinator conservation. This multi-stressor study investigated the responses of six functional behaviours of bumblebees when exposed to either a neonicotinoid (imidacloprid) or a sulfoximine (sulfoxaflor) across a standardised low, mid, and high temperature. We found the neonicotinoid had a significant effect on five of the six behaviours, with a greater effect at the lower temperature(s) when measuring responsiveness, the likelihood of movement, walking rate, and food consumption rate. In contrast, the neonicotinoid had a greater impact on flight distance at the higher temperature. Our findings show that different organismal functions can exhibit divergent thermal responses, with some pesticide-affected behaviours showing greater impact as temperatures dropped, and others as temperatures rose. We must therefore account for environmental context when determining pesticide risk. Moreover, we found evidence of synergistic effects, with just a 3°C increase causing a sudden drop in flight performance, despite seeing no effect of pesticide at the two lower temperatures. Our findings highlight the importance of multi-stressor studies to quantify threats to insects, which will help to improve dynamic evaluations of population tipping points and spatiotemporal risks to biodiversity across different climate regions.