Unravelling nicotinic receptor and ligand features underlying neonicotinoid knockdown actions on the malaria vector mosquito Anopheles gambiae.

With the spread of resistance to long-established insecticides targeting Anopheles malaria vectors, understanding the actions of compounds newly identified for vector control is essential. With new commercial vector-control products containing neonicotinoids under development, we investigate the actions of 6 neonicotinoids (imidacloprid, thiacloprid, clothianidin, dinotefuran, nitenpyram and acetamiprid) on 13 Anopheles gambiae nicotinic acetylcholine receptor (nAChR) subtypes produced by expression of combinations of the Agα1, Agα2, Agα3, Agα8 and Agß1 subunits in Xenopus laevis oocytes, the Drosophila melanogaster orthologues of which we have previously shown to be important in neonicotinoid actions. The presence of the Agα2 subunit reduces neonicotinoid affinity for the mosquito nAChRs, whereas the Agα3 subunit increases it. Crystal structures of the acetylcholine binding protein (AChBP), an established surrogate for the ligand-binding domain, with dinotefuran bound, shows a unique target site interaction through hydrogen bond formation and CH-N interaction at the tetrahydrofuran ring. This is of interest as dinotefuran is also under trial as the toxic element in baited traps. Multiple regression analyses show a correlation between the efficacy of neonicotinoids for the Agα1/Agα2/Agα8/Agß1 nAChR, their hydrophobicity and their rate of knockdown of adult female An. gambiae, providing new insights into neonicotinoid features important for malaria vector control.

Changes in the proteome of Apis mellifera acutely exposed to sublethal dosage of glyphosate and imidacloprid.

Uncontrolled use of pesticides has caused a dramatic reduction in the number of pollinators, including bees. Studies on the effects of pesticides on bees have reported effects on both metabolic and neurological levels under chronic exposure. In this study, variations in the differential expression of head and thorax-abdomen proteins in Africanized A. mellifera bees treated acutely with sublethal doses of glyphosate and imidacloprid were studied using a proteomic approach. A total of 92 proteins were detected, 49 of which were differentially expressed compared to those in the control group (47 downregulated and 2 upregulated). Protein interaction networks with differential protein expression ratios suggested that acute exposure of A. mellifera to sublethal doses of glyphosate could cause head damage, which is mainly associated with behavior and metabolism. Simultaneously, imidacloprid can cause damage associated with metabolism as well as, neuronal damage, cellular stress, and impairment of the detoxification system. Regarding the thorax-abdomen fractions, glyphosate could lead to cytoskeleton reorganization and a reduction in defense mechanisms, whereas imidacloprid could affect the coordination and impairment of the oxidative stress response.

Acute toxicity of the fungicide captan to honey bees and mixed evidence for synergism with the insecticide thiamethoxam.

Honey bees are commonly co-exposed to pesticides during crop pollination, including the fungicide captan and neonicotinoid insecticide thiamethoxam. We assessed the impact of exposure to these two pesticides individually and in combination, at a range of field-realistic doses. In laboratory assays, mortality of larvae treated with captan was 80-90% greater than controls, dose-independent, and similar to mortality from the lowest dose of thiamethoxam. There was evidence of synergism (i.e., a non-additive response) from captan-thiamethoxam co-exposure at the highest dose of thiamethoxam, but not at lower doses. In the field, we exposed whole colonies to the lowest doses used in the laboratory. Exposure to captan and thiamethoxam individually and in combination resulted in minimal impacts on population growth or colony mortality, and there was no evidence of synergism or antagonism. These results suggest captan and thiamethoxam are each acutely toxic to immature honey bees, but whole colonies can potentially compensate for detrimental effects, at least at the low doses used in our field trial, or that methodological differences of the field experiment impacted results (e.g., dilution of treatments with natural pollen). If compensation occurred, further work is needed to assess how it occurred, potentially via increased queen egg laying, and whether short-term compensation leads to long-term costs. Further work is also needed for other crop pollinators that lack the social detoxification capabilities of honey bee colonies and may be less resilient to pesticides.

A Tunable Threshold Colorimetric DNA Logic Gate for Intuitive Assessment of Chemical Contaminant Exceedance.

Current molecular logic gates are predominantly focused on the qualitative assessment of target presence, which has certain limitations in scenarios requiring quantitative assessment, such as chemical contaminant monitoring. To bridge this gap, we have developed a novel DNA logic gate featuring a tunable threshold, specifically tailored to the limits of contaminants. At the core of this logic gate is a DNA-gold nanoparticle (AuNP) hybrid film that incorporates aptamer sequences to selectively bind to acetamiprid (ACE) and atrazine (ATR). Upon interaction with these contaminants, the film degrades, releasing AuNPs that, in the presence of Hg2+, catalyze the oxidation of TMB, resulting in a visible blue coloration on test paper. This aptamer-enabled process effectively establishes an OR logic gate, with ACE and ATR as inputs and the appearance of blue color as the output. A key innovation of our system is its tunable input threshold. By adjusting the concentration of Hg2+, we can fine-tune the color mutation points to match the input threshold to predefined limits, such as Maximum Residue Limits (MRLs). This alignment allows semiquantitative assessment of contaminant levels, providing intuitive visual feedback of contaminant exceedance. Validation experiments with spiked samples confirm its accuracy and reliability by closely matching HPLC results. Therefore, our colorimetric DNA logic gate is emerging as a promising tool for easy and semiquantitative monitoring of chemical contaminants across diverse applications.

Exposure of pregnant women to neonicotinoids in Wenzhou City, East China: A biomonitoring study.

BACKGROUND: China produces and consumes a large amount of neonicotinoids. A non-negligible exposure to neonicotinoids might occur for Chinese pregnant women, but relevant data remain limited. OBJECTIVE: To investigate the exposure to neonicotinoids by urinary biomonitoring in pregnant women from Wenzhou City, East China. METHODS: We selected 432 pregnant women in Wenzhou City in 2022. A total of eight parent neonicotinoids and four metabolites were determined in single spot urine by liquid chromatography coupled to mass spectrometry. Basic characteristics, physical activity, pre-pregnant body mass index, and intake of drinking water and food were investigated by the questionnaire. Health risk was assessed by hazard quotient (HQ) and hazard index (HI) based on human safety thresholds derived from different health endpoints. RESULTS: Neonicotinoids and their metabolites in urine had a detection frequency between 0 % and 80.1 %. At least one neonicotinoid or metabolite was detected in 93.5 % of urine samples. Except for clothianidin (51.2 %) and N-desmethyl-acetamiprid (80.1 %), the detection frequencies of other neonicotinoids and metabolites ranged from 0 % to 43.8 %. The summed concentrations of all neonicotinoids and their metabolites ranged from < LOD to 222.83 µg/g creatinine with the median concentration of 2.58 µg/g creatinine. Maternal age, educational level, occupation, household income, screen time, and pre-pregnant body mass index were associated with detection frequencies or concentrations of neonicotinoids and their metabolites. Pregnant women with higher consumption frequencies of wheat, fresh vegetable, shellfish, fresh milk, and powdered milk had higher detection frequencies of neonicotinoids and their metabolites. Both HQ and HI were less than one. CONCLUSIONS: Overall, pregnant women in Wenzhou City showed a notable frequency of exposure to at least one neonicotinoid, although the exposure frequency for each specific neonicotinoid was generally low. Several food items derived from plants and animals were potential exposure sources. A low health risk was found based on current safety thresholds.

In vivo tracing of cyromazine and three neonicotinoids in cowpea under field conditions by solid-phase microextraction combined with ultra-performance liquid chromatography-tandem mass spectrometry.

BACKGROUND: Excessive pesticide residues in agricultural products could accumulate in organisms through the food chain, causing potential harm to human health. The investigation of dissipation kinetics and residues of pesticides in crops is crucial for the scientific application of pesticides and the mitigation of their adverse effects on human health. In vivo solid-phase microextraction (in vivo SPME) has unique advantages, but the research on field plants is still lacking and the quantitative correction methods need to be further developed. RESULTS: A method combining in vivo solid-phase microextraction with ultra-performance liquid chromatography-tandem mass spectrometry (in vivo SPME-UPLC-MS/MS) was developed to monitor the presence of acetamiprid, cyromazine, thiamethoxam and imidacloprid in cowpea fruits grown in the field. The sampling rates (Rs) were determined using both in vitro SPME in homogenized cowpea samples and in vivo SPME in intact cowpea fruit samples. The in vivo-Rs values were significantly higher than the in vitro-Rs for the same analyte, which were used for in vivo SPME correction. The accuracy of this method was confirmed by comparison with a QuEChERS-based approach and subsequently applied to trace pesticide residues in field-grown cowpea fruits. The residual concentrations of each pesticide positively correlated with application doses. After 7 days of application at two different doses, all of the pesticides had residual concentrations below China's maximum residue limits. Both experimental data and predictions indicated that a safe preharvest interval for these pesticides is 7 days; however, if the European Union standards are to be met, a safe preharvest interval for cyromazine should be at least 13 days. SIGNIFICANCE: This study highlights the advantages of in vivo SPME for simultaneous analysis and tracking of multiple pesticides in crops under field conditions. This technique is environmentally friendly, minimally invasive, highly sensitive, accurate, rapid, user-friendly, cost-effective, and capable of providing precise and timely data for long-term pesticide surveillance. Consequently, it furnishes valuable insights to guide the safe utilization of pesticides in agricultural production.

Enhanced detection of acetamiprid via a gold nanoparticle-based colorimetric aptasensor integrated with a hybridization chain reaction.

This study presents a novel colorimetric aptasensor, which seamlessly integrates gold nanoparticles (AuNPs) with the amplification potential of the hybridization chain reaction (HCR) for enhanced detection of acetamiprid. The aptamer, hybridized with a partially complementary strand that is covalently linked to AuNPs, serves as the recognition element for acetamiprid. The free end sequence of the aptamer, distal from the AuNP surface, functions as the initiating strand for the HCR, triggering the amplification process. In the absence of acetamiprid, the HCR efficiently occurs, conferring robust salt tolerance to the AuNPs and maintaining their characteristic red coloration. However, in the presence of acetamiprid, the aptamer preferentially binds to its target, disrupting the double-stranded structure and leading to the dissociation of the aptamer from the AuNPs. This dissociation results in a decrease in the HCR product, subsequently diminishing the salt tolerance of AuNPs and triggering a colorimetric transition from red to gray. This integration enhances sensitivity to 3.14 nM. Additionally, carbon quantum dots (CQDs) transduce colorimetric signals to fluorescent ones, further boosting the sensitivity to 0.24 nM. The aptasensor exhibits excellent selectivity and robustness. Real-world testing on tomato, peach, and lettuce shows recoveries of 98.50% to 100.36% with low standard deviations, validating its utility for pesticide residue analysis and food safety. This study provides a powerful tool for rapid and accurate pesticide detection, crucial for food safety.

Structural and functional impacts of neonicotinoids analogues on Apis mellifera's chemosensory protein: Insights from spectroscopic and molecular modeling investigations.

In the intricate web of ecological relationships, pollinators such as the Italian honeybee (Apis mellifera) play a crucial role in maintaining biodiversity and agricultural productivity. This study focuses on the interactions between three neonicotinoid compounds and the honeybee's chemosensory protein 3 (CSP3), a key player in their olfactory system. Employing advanced spectroscopic techniques and molecular modeling, we explore the binding dynamics and conformational changes in CSP3 upon exposure to these pesticides. The research reveals that all three neonicotinoids considerably quench CSP3's fluorescence through a dynamic and static mixing mechanism, indicating a strong binding affinity, predominantly driven by hydrophobic interactions. UV-visible absorption, synchronous fluorescence, and 3D fluorescence spectra support slight changes in the microenvironment around the aromatic amino acids of CSP3. Circular dichroism spectra indicate a reduction in CSP3's α-helix content, suggesting structural alterations. Molecular docking and dynamics simulations further elucidate the binding modes and stability of these interactions, highlighting the role of specific amino acids in CSP3's binding cavity. Findings provide critical insights into molecular mechanisms by which neonicotinoids may impair honeybee chemosensory function, offering implications for designing safer pesticides and understanding the broader ecological impact of these chemicals on pollinator health.

Temporal and spatial trends of imidacloprid-related hazards in France.

Neonicotinoids are the top-selling insecticides worldwide. Because of their method of use, mainly to coat seeds, neonicotinoids have been found to widely contaminate the environment. Their high toxicity has been shown to be a major concern in terms of impact on biodiversity, and the use of these insecticides has been associated with population declines of species in different countries. Despite the widespread recognition of the risk of neonicotinoids to biodiversity, their temporal and spatial use remains poorly known in many countries. Yet this information is essential to address the potential impacts of these pesticides on biodiversity and to inform measures to establish protected areas or biodiversity restoration. The present study relied a large publicly available dataset to characterise the temporal and spatial use in France of imidacloprid, the most widely used neonicotinoid worldwide, as well as analysed water contamination surveys between 2005 and 2022 to assess the contamination of the environment. The results show that imidacloprid was the main neonicotinoid used in France over the study period. This use was spatially structured, with higher use in northern and western France, particularly related to cereal and beet crops area. The water contamination survey indicated that imidacloprid has widely contaminated the environment and consequently increased the risk to biodiversity, especially in counties crossed by the Loire, Seine and Vilaine rivers. This risk increased between 2005 and 2018 due to the higher use of imidacloprid and decreased sharply after 2018 due to its ban, although it was reauthorized by derogation for sugar beet in 2021. This study is the first assessment of imidacloprid pressure on biodiversity in France and shows the spatial and temporal correlation between agricultural practices and the freshwater contamination level. These results will help to identify priority areas for mitigation and restoration measures.

Maleic anhydride-functionalized cellulose nanocrystal-stabilized high internal phase Pickering emulsion for pesticide delivery.

The salt-responsiveness of Pickering emulsions has significantly influenced their applications due to the large amount of salt on the surface of plant leaves. The present study provided a maleic anhydride-functionalized cellulose nanocrystal-stabilized high internal phase Pickering emulsion (MACNCs-HIPPEs) that was stable to high-concentration salt and used for pesticide delivery. The stability of MACNCs-HIPPEs was investigated by adjusting the oil-phase volume fraction (φ), the MACNCs concentration, NaCl dosages, and the rheological properties. The high internal phase Pickering emulsion was obtained at φ of 0.8 and MACNCs concentration of 2wt% and showed excellent salt stability (NaCl, 1200 mM) and significant storage stability (60 days). The sustained release of imidacloprid (IMI) from imidacloprid-loaded MACNCs-HIPPEs (IMI@MACNCs-HIPPEs) showed a positive correlation to the temperature (15°C, 25°C, 35°C), indicating clear thermo-responsiveness of the prepared pesticide formulation. The test of spread and retention of IMI@MACNCs-HIPPEs on the leaf surface showed a significant advantage compared with the commercial IMI water dispersible granules (CG). All the advantages mentioned above showed the excellent potential of the MACNCs-HIPPEs in delivering lipophilic pesticides.

Magnetic carboxymethyl gond katira-grafted-poly(3-aminobenzoic acid) as an antibacterial biosorbent for purification of acetamiprid-contaminated water.

The elimination of pesticides from polluted water is critical due to their harmful environmental and biological impacts. Recently, there has been interest in utilizing natural polymer-based adsorbents as an eco-friendly approach to eliminate or reduce the levels of water pollutants. In this work, we synthesized an antimicrobial and magnetic bionanocomposite consisting of carboxymethyl gond katira-grafted- poly(3-aminobenzoic acid) with iron oxide and zinc oxide NPs (CMT-g-P3ABA/ZnO/Fe3O4) through an in situ polymerization reaction and examined for its ability to adsorb the pesticide acetamiprid (AP). The bionanocomposite was characterized using several analytical techniques, including spectroscopy; XRD presented the crystalline structure of ZnO/Fe3O4 in the CMT-g-P3ABA amorphous matrix. The ZnO/Fe3O4 partially aggregated formation and exhibited polyhedral crystal shapes was depicted by electron microscopy images, vibrating sample magnetometer (45.06 emu/g), porosimetry (5.52 m2/g), and thermal (Chair yield of approximately 43.83 %) and elemental analyses. Under various conditions, including solution pH (4-9), adsorbent dosage (0.005-0.025 g), time of contact (10-30 min), and pesticide preliminary concentration (200-400 mg/L) in 10 mL of the solution. Based on this research, Adsorption data were perfectly fitted by the Freundlich isotherm model with RAP2= 0.99038, while the pseudo-second-order (PSO) model well-explained adsorption kinetics with RAP2= 0.99847. AP adsorption to the CMT-g-P3ABA/ZnO/Fe3O4 bionanocomposite was successful due to hydrophobic interactions, hydrogen bonding, and π-π stacking. Furthermore, adsorption-desorption experiments demonstrated that the bionanocomposite could be regenerated after three reuse cycles without considerable loss of pesticide removal performance. The bionanocomposite also exhibited promising antimicrobial activity in contradiction to test bacteria.

A single dose of clothianidin exposure induces varying sex-specific behavioral changes in adulthood depending on the developmental stage of its administration.

Clothianidin (CLO), a neonicotinoid that is widely used in forests and agricultural areas, was recently reported to cause toxicity in mammals. Although sensitivity to chemicals varies between sexes and developmental stages, studies that comprehensively evaluate both males and females are limited. Therefore, in this study we utilized murine models to compare the sex-specific differences in behavioral effects following CLO exposure at different developmental stages. We orally administered CLO to male and female mice as a single high-dose solution (80 mg/kg) during the postnatal period (2-week-old), adolescence (6-week-old), or maturity (10-week-old), and subsequently evaluated higher brain function. The behavioral battery test consisted of open field, light/dark transition, and contextual/cued fear conditioning tests conducted at three and seven months of age. After the behavioral test, the brains were dissected and prepared for immunohistochemical staining. We observed behavioral abnormalities in anxiety, spatial memory, and cued memory only in female mice. Moreover, the immunohistochemical analysis showed a reduction in astrocytes within the hippocampus of female mice with behavioral abnormalities. The behavioral abnormalities observed in female CLO-treated mice were consistent with the typical behavioral abnormalities associated with hippocampal astrocyte dysfunction. It is therefore possible that the CLO-induced behavioral abnormalities are at least in part related to a reduction in astrocyte numbers. The results of this study highlight the differences in behavioral effects following CLO exposure between sexes and developmental stages.

Impact of imidacloprid exposure on gestational hyperglycemia: A multi-omics analysis.

Imidacloprid (IMI), a commonly utilized neonicotinoid insecticide, has been identified to adversely impact glucose homeostasis. Pregnant women are believed to be more sensitive to toxins than non-pregnant women, and the impact of IMI exposure on gestational hyperglycemia remain unclear. To explore the impact, pregnant mice fed a high-fat diet were exposed to different doses (0.06, 0.6, 6 mg/kg bw/day) of IMI by gavage. Glucose homeostasis-related parameters were measured. The glucose homeostasis influenced by IMI treatment was explored through integrating gut microbiota, metabolomic and transcriptomic analysis. Results showed that IMI-H (6 mg/kg bw/day) exposure notably restricted gestational weight gain and perturbed glucose homeostasis characterized by reduced glucose tolerance and insulin sensitivity, alongside elevated levels of fasting blood glucose and insulin. Multi-omics analysis revealed that IMI-H exposure induced significant changes in the richness and composition of the gut microbiome. The metabolite profiles of serum samples and cecal contents, and transcriptome of liver and ileum were all affected by IMI-H treatment. The altered gut microbiota, metabolites and genes exhibited significant correlations with glucose homeostasis-related parameters. These differential metabolites and genes were implicated in various metabolic pathways including bile secretion, glucagon signaling pathway, lipid metabolism, fatty acid metabolism. Significant correlations were observed between the altered gut microbiota and caecum metabolome as well as liver transcriptome. For example, the abundance of Oscillibacter was strongly correlated with gut microflora-related metabolites (Icosenoic acid, Lysosulfatide, and fluticasone) and liver differential genes (Grin3b, Lifr, and Spta1). Together, IMI exposure resulted in significant changes in microbial composition, along with alterations in certain metabolites and genes associated with metabolic process, which may promote gestational hyperglycemia.

Genotoxic effects of sub-lethal doses of nicotine and acetamiprid in neuroblasts of Drosophila melanogaster and Drosophila suzukii.

Neonicotinoids form a class of insecticides that are chemically related to nicotine and are widely used in crop protection. They have adverse effects on the neuronal nicotinic acetylcholine receptors (nAChRs). One of the neonicotinoids approved for control of the invasive pest Drosophila suzukii is acetamiprid. Despite concerns regarding its genotoxicity and data indicating the presence of small amounts of this substance in fruits intended for consumption, effects of its low doses on nerve cells are yet to be investigated. To determine whether the neurotoxic effects are species-specific and vary depending on the insecticide present in diet, multigenerational cultures of Drosophila melanogaster and D. suzukii were prepared, in this study, in media supplemented with different concentrations (below the LC50) of acetamiprid and nicotine. Acetamiprid, analogous to nicotine, caused damage to the DNA of neuroblasts in both species, at sublethal concentrations, along with a decrease in mobility, which remained at a similar level over subsequent generations. D. suzukii was found to be more sensitive to nicotine and acetamiprid, due to which the genotoxic effects were stronger even at lower doses of toxins. The results collectively indicated that even low concentrations of acetamiprid affect the stem cells of developing fly brain, and that long-term response to the tested insecticides is species-specific.

Developmental toxicity of the neonicotinoid pesticide clothianidin to the larvae of the crustacean Decapoda, Penaeus vannamei.

The developmental toxicity effects of neonicotinoid pesticides such as clothianidin have not been fully explored in agricultural applications. This is particularly noteworthy because such pesticides significantly impact the survival rates of invertebrates, with arthropod larvae being particularly vulnerable. This study aimed to address this research gap by specifically investigating the toxicological effects of clothianidin on the developmental stages of the larvae of the economically important aquaculture species Penaeus vannamei. In these experiments, shrimp eggs were exposed to seawater containing different concentrations of clothianidin beginning at N1, and each phase was observed and analyzed to determine its toxic impact on larval development. These results revealed that clothianidin induces an increase in deformity rates and triggers abnormal cell apoptosis. It also significantly reduced survival rates and markedly decreased body length and heart rate in the later stages of larval development (P3). Transcriptomic analysis revealed disruptions in larval DNA integrity, protein synthesis, and signal transduction caused by clothianidin. To survive prolonged exposure, larvae may attempt to maintain their viability by repairing cell structures and enhancing signal transduction mechanisms. This study offers the first empirical evidence of the toxicity of clothianidin to arthropod larvae, underscoring the impact of environmental pollution on aquatic health.

Thermal adaptation affects the temperature-dependent toxicity of the insecticide imidacloprid to soil invertebrates.

Terrestrial ectotherms are vulnerable to climate change since their biological rates depend on the ambient temperature. As temperature may interact with toxicant exposure, climate change may cause unpredictable responses to toxic stress. A population's thermal adaptation will impact its response to temperature change, but also to interactive effects from temperature and toxicants, but these effects are still not fully understood. Here, we assessed the combined effects of exposure to the insecticide imidacloprid across the temperatures 10-25 °C of two populations of the Collembola Hypogastrura viatica (Tullberg, 1872), by determining their responses in multiple life history traits. The con-specific populations differ considerably in thermal adaptations; one (arctic) is a temperature generalist, while the other (temperate) is a warm-adapted specialist. For both populations, the sub-lethal concentrations of imidacloprid became lethal with increasing temperature. Although the thermal maximum is higher for the warm-adapted population, the reduction in survival was stronger. Growth was reduced by imidacloprid in a temperature-dependent manner, but only at the adult life stage. The decrease in adult body size combined with the absence of an effect on the age at first reproduction suggests a selection on the timing of maturation. Egg production was reduced by imidacloprid in both populations, but the negative effect was only dependent on temperature in the warm-adapted population, with no effect at 10 °C, and decreases of 41 % at 15 °C, and 74 % at 20 °C. For several key traits, the population best adapted to utilize high temperatures was also the most sensitive to toxic stress at higher temperatures. It could be that by allocating more energy to faster growth, development, and reproduction at higher temperatures, the population had less energy for maintenance, making it more sensitive to toxic stress. Our findings demonstrate the need to take into account a population's thermal adaptation when assessing the interactive effects between temperature and other stressors.

Enantioselective toxicity of the neonicotinoid dinotefuran on honeybee (Apis mellifera) larvae.

The threat of neonicotinoids to insect pollinators, particularly honeybees (Apis mellifera), is a global concern, but the risk of chiral neonicotinoids to insect larvae remains poorly understood. In the current study, we evaluated the acute and chronic toxicity of dinotefuran enantiomers to honeybee larvae in vitro and explored the mechanism of toxicity. The results showed that the acute median lethal dose (LD50) of S-dinotefuran to honeybee larvae was 30.0 µg/larva after oral exposure for 72 h, which was more toxic than rac-dinotefuran (92.7 µg/larva) and R-dinotefuran (183.6 µg/larva). Although the acute toxicity of the three forms of dinotefuran to larvae was lower than that to adults, chronic exposure significantly reduced larval survival, larval weight, and weight of newly emerged adults. Analysis of gene expression and hormone titer indicated that dinotefuran affects larval growth and development by interfering with nutrient digestion and absorption and the molting system. Analysis of hemolymph metabolome further revealed that disturbances in the neuroactive ligand-receptor interaction pathway and energy metabolism are the key mechanisms of dinotefuran toxicity to bee larvae. In addition, melatonin and vitellogenin are used by larvae to cope with dinotefuran-induced oxidative stress. Our results contribute to a comprehensive understanding of dinotefuran damage to bees and provide new insights into the mechanism of enantioselective toxicity of insecticides to insect larvae.

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.

Effects of imidacloprid combined with validamycin on the population fitness and symbiotic of Nilaparvata lugens (Hemiptera: Delphacidae).

Using a high-efficiency insecticide in combination with fungicides that have different mechanisms of action is a conventional method in the current management of brown planthopper (BPH) resistance. In this study, we investigate the separate and combined effects of the low-toxicity fungicide validamycin and the non-cross-resistant insecticide imidacloprid on the fitness and symbiosis of BPH. These research results indicate that when the proportion of active ingredients in validamycin is combined with imidacloprid at a ratio of 1:30, the toxicity ratio and co-toxicity coefficient are 1.34 and 691.73, respectively, suggesting that the combination has a synergistic effect on the control of BPH. The number of yeast-like symbiotic (YLS) and dominant symbiotic (Noda) in the imidacloprid + validamycin groups were significantly lower than the other three treatment groups (validamycin, imidacloprid, and water). The results of the study on population fitness show that the lifespan of the BPH population in validamycin, imidacloprid, and imidacloprid + validamycin was shortened. Notably, the BPH populations in the imidacloprid + validamycin groups were significantly lower than other groups in terms of average generation cycle, intrinsic growth rate, net reproduction rate, finite rate of increase, and fitness. The Real-time quantitative PCR showed that validamycin and imidacloprid + validamycin can significantly inhibit the expression of the farnesyl diphosphate farnesyl transferase gene (EC2.5.1.21) and uricase gene (EC1.7.3.3), with imidacloprid + validamycin demonstrating the most pronounced inhibitory effect. Our research results can provide insights and approaches for delaying resistance and integrated management of BPH.

Various functions of detoxification enzymes against insecticides in Nilaparvata lugens selected by toxicity assays and RNAi methods.

The brown planthopper (BPH), Nilaparvata lugens is a devastating agricultural pest of rice, and they have developed resistance to many pesticides. In this study, we assessed the response of BPH nymphs to nitenpyram, imidacloprid, and etofenprox using contact and dietary bioassays, and investigated the underlying functional diversities of BPH glutathione-S-transferase (GST), carboxylesterase (CarE) and cytochrome P450 monooxygenase (P450) against these insecticides. Both contact and ingestion toxicity of nitenpyram to BPH were significantly higher than either imidacloprid or etofenprox. Under the LC50 concentration of each insecticide, they triggered a distinct response for GST, CarE, and P450 activities, and each insecticide induced at least one detoxification enzyme activity. These insecticides almost inhibited the expression of all tested GST, CarE, and P450 genes in contact bioassays but induced the transcriptional levels of these genes in dietary bioassays. Silencing of NlGSTD2 expression had the greatest effect on BPH sensitivity to nitenpyram in contact test and imidacloprid in dietary test. The sensitivities of BPH to insecticide increased the most in the contact test was etofenprox after silencing of NlCE, while the dietary test was nitenpyram. Knockdown of NlCYP408A1 resulted in BPH sensitivities to insecticide increasing the most in the contact test was nitenpyram, while the dietary test was imidacloprid. Taken together, these findings reveal that NlGSTD2, NlCE, and NlCYP408A1 play an indispensable role in the detoxification of the contact and ingestion toxicities of different types of insecticides to BPH, which is of great significance for the development of new strategies for the sucking pest control.