Insecticidal activity and underlying molecular mechanisms of a phytochemical plumbagin against Spodoptera frugiperda.

Introduction: Plumbagin is an important phytochemical and has been reported to exhibit potent larvicidal activity against several insect pests, However, the insecticidal mechanism of plumbagin against pests is still poorly understood. This study aimed to investigate the insecticidal activities of plumbagin and the underlying molecular mechanisms against a devastating agricultural pest, the fall armyworm Spodoptera frugiperda. Methods: The effects of plumbagin on S. frugiperda larval development and the activities of two detoxification enzymes were initially examined. Next, transcriptomic changes in S. frugiperda after plumbagin treatment were investigated. Furthermore, RNA-seq results were validated by qPCR. Results: Plumbagin exhibited a high larvicidal activity against the second and third instar larvae of S. frugiperda with 72 h LC50 of 0.573 and 2.676 mg/g, respectively. The activities of the two detoxification enzymes carboxylesterase and P450 were significantly increased after 1.5 mg/g plumbagin treatment. Furthermore, RNA-seq analysis provided a comprehensive overview of complex transcriptomic changes in S. frugiperda larvae in response to 1.5 mg/g plumbagin exposure, and revealed that plumbagin treatment led to aberrant expression of a large number of genes related to nutrient and energy metabolism, humoral immune response, insect cuticle protein, chitin-binding proteins, chitin synthesis and degradation, insect hormone, and xenobiotic detoxification. The qPCR results further validated the reproducibility and reliability of the transcriptomic data. Discussion: Our findings provide a valuable insight into understanding the insecticidal mechanism of the phytochemical plumbagin.

Toxicity of 6-gingerol and Cymbopogon citratus against Pediculus humanus capitis De Geer (Phthiraptera: Pediculidae): Mortality, detoxifying enzymes, and morphological ultrastructure alterations in lice.

Pediculus humanus capitis (head louse), which causes pediculosis capitis, remains a global health concern. Plant products are efficient alternative pediculicides for treating the human ectoparasite P. h. capitis which is resistant to permethrin. The study evaluates the toxicity and mechanisms of 6-gingerol and Cymbopogon citratus leaf extract on P. h. capitis. Pediculus humanus capitis adult stages were exposed to three different dosages of 6-gingerol and C. citratus crude leaf extract on filter sheets for 5, 10, and 30 min, respectively. The biochemical approach was used to assess the activity of detoxifying enzymes including acetylcholinesterase (AChE), glutathione S-transferase (GST), and oxidase. Scanning electron microscope (SEM) was used to investigate the ultrastructure of the morphological body of lice. After 30 min, 6-gingerol and C. citratus leaf extract killed P. h. capitis completely. Bioassay periods significantly affected lice mortality (P < 0.05). The LC50 values for 6-gingerol and C. citratus extract were 1.79 µg/cm2 and 25.0 µg/cm2, respectively. 6-Gingerol and C. citratus leaf extract significantly lower AChE and GST activity (P < 0.05). Cymbopogon citratus also caused morphological ultrastructure changes in P. h. capitis, including an irregularly formed head, thorax, abdominal respiratory spiracles, and belly. 6-Gingerol and C. citratus leaf extracts could be used as an alternate pediculicide to decrease P. h. capitis populations.

Induction of insulin resistance in female mice due to prolonged phenanthrene exposure: Unveiling the low-dose effect and potential mechanisms.

Phenanthrene (Phe) is a commonly occurring polycyclic aromatic hydrocarbon (PAH) found in various food sources and drinking water. Previous studies have shown that long-term exposure to Phe in male mice leads to insulin resistance in a dose-dependent manner. However, the effect of Phe on glucose homeostasis in female mice remains unknown. To address this knowledge gap, female Kunming mice were exposed to Phe through their drinking water at concentrations of 0.05, 0.5, and 5 ng/mL. After 270 d of exposure, we surprisingly discovered a low-dose effect of Phe on insulin resistance in female mice, which differed from the effect observed in male mice and showed sexual dimorphism. Specifically, insulin resistance was only observed in the 0.05 ng/mL treatment, and this low-dose effect was also reflected in the concentration of Phe in white adipose tissue (WAT). Differences in metabolic enzyme activities in the liver may potentially explain this effect. The observed sexual dimorphism in Phe exposure could be attributed to variations in estrogen (E2) level and estrogen receptor beta (ERß) expression in WAT. These findings highlight the association between environmental factors and the development of insulin resistance, emphasizing the pathogenic effect of even low doses of Phe. Moreover, sex dependent-effect should be given more attention when studying the toxic effects of environmental pollutants.

Efficacy of 2 botanical aphicides, chicoric and 3,5-dicaffeoylquinic acids, on aphids susceptible and resistant to synthetic insecticides.

Dicaffeoyltartaric acid (diCT) and 3,5-dicaffeoylquinic acid (3,5-diCQ) are described for their aphicidal properties on several aphid species. Intending to valorize diCT and 3,5-diCQ as biocontrol products and because of the high adaptive capacities of aphids to xenobiotics, we sought to determine the existence of adaptation first in Myzus persicae (Sulzer) (Hemiptera: Aphididae) and then other aphids. Resistance of aphids to these biopesticides could be promoted by (i) the existence of resistance to synthetic insecticides that may confer cross-resistance and (ii) the presence of these compounds in wild plants likely which may have led to pre-existing adaptation in aphids. We assessed the resistance levels to diCT and 3,5-diCQ in 7 lab strains (including some resistant to synthetic aphicides) and 7 wild populations of M. persicae using biotests. The activities of detoxification enzymes contributing to insecticide resistance were also measured. Additionally, we followed the same method to characterize susceptibility to these caffeic derivatives in wild populations of Nasonovia ribisnigri (Mosley) (Hemiptera: Aphididae), Brevicoryne brassicae(Linnaeus) (Hemiptera: Aphididae) and, Aphis craccivora(Koch) (Hemiptera: Aphididae). Our results show variability in susceptibility to diCT between populations of M. persicae, but resistance ratios (RR) were low (RR = 3.59). We found no cross-resistance between synthetic insecticides and diCT. Carboxylesterase and glutathione-S-transferase did not seem to be involved in its detoxification. A clone of A. craccivora collected from peanut, a species rich in diCT, was not susceptible to either diCT or 3,5-diCQ, suggesting a common molecular target for these 2 molecules and the existence of a high-effect resistance mechanism. These active botanical substances remain good candidates for M. persicae biocontrol in agriculture.

Sublethal effect and detoxifying metabolism of metaflumizone and indoxacarb on the fall armyworm, Spodoptera frugiperda.

The fall armyworm (FAW), Spodoptera frugiperda (J.E. Smith) (Lepidoptera, Noctuidae), is a highly polyphagous invasive pest that damages various crops. Pesticide control is the most common and effective strategy to control FAW. In this study, we evaluated the toxicity of metaflumizone and indoxacarb against third-instar FAW larvae using the insecticide-incorporated artificial diet method under laboratory conditions. Both metaflumizone and indoxacarb exhibited substantial toxicity against FAW, with LC50 values of 2.43 and 14.66 mg/L at 72 h, respectively. The sublethal effects of metaflumizone and indoxacarb on parental and F1 generation FAW were investigated by exposing third-instar larvae to LC10 and LC30 concentrations of these insecticides. Sublethal exposure to these two insecticides significantly shortened adult longevity, extended pupal developmental times and led to reduced pupal weight, pupation rates, and adult fecundity in the treated parental generation and F1 generation at LC10 or LC30 concentrations, in comparison to the control group. The larval developmental times were shortened in the parental generation but prolonged in the F1 generation, after being treated with sublethal concentrations of metaflumizone. Furthermore, larvae exposed to LC10 or LC30 concentrations of indoxacarb exhibited elevated activity levels of cytochrome P450 monooxygenase and glutathione S-transferase, which coincides with the observed synergistic effect of piperonyl butoxide and diethyl maleate. In conclusion, the high toxicity and negative impact of metaflumizone and indoxacarb on FAW provided significant implications for the rational utilization of insecticides against this pest.

Screening and functional validation of the core detoxification genes conferring broad-spectrum response to insecticides in Spodoptera frugiperda.

BACKGROUND: Fall armyworm, Spodoptera frugiperda, a formidable agricultural pest, has developed resistance to various synthetic insecticides. However, how S. frugiperda utilizes its limited energy and resources to deal with various insecticides remains largely unexplored. RESULTS: We utilized transcriptome sequencing to decipher the broad-spectrum adaptation mechanism of S. frugiperda to eight insecticides with distinct modes-of-action. Analysis of the Venn diagram revealed that 1014 upregulated genes and 778 downregulated genes were present in S. frugiperda treated with at least five different insecticides, compared to the control group. Exposure to various insecticides led to the significant upregulation of eight cytochrome P450 monooxygenases (P450s), four UDP glucosyltransferases (UGTs), two glutathione-S-transferases (GSTs) and two ATP-binding cassette transporters (ABCs). Among them, the sfCYP340AD3 and sfCYP4G74 genes were demonstrated to respond to stress from six different insecticides in S. frugiperda, as evidenced by RNA interference and toxicity bioassays. Furthermore, homology modeling and molecular docking analyses showed that sfCYP340AD3 and sfCYP4G74 possess strong binding affinities to a variety of insecticides. CONCLUSION: Collectively, these findings showed that S. frugiperda utilizes a battery of core detoxification genes to cope with the exposure of synthetic insecticides. This study also sheds light on the identification of efficient insecticidal targets gene and the development of resistance management strategies in S. frugiperda, thereby facilitating the sustainable control of this serious pest. © 2024 Society of Chemical Industry.

Involvement of a catalase gene in lignin catalysis and immune defense against pathogenic fungus in Coptotermes formosanus: a potential new target for termite control.

BACKGROUND: Detoxifying enzymes are likely involved in lignin feeding and immune defense mechanisms within termites, rendering them potential targets for biological control. However, investigations into the dual functionality of termite detoxification enzymes in vivo have not been documented. RESULTS: In this study, the complete cDNA of the catalase gene (Cfcat) derived from Coptotermes formosanus Shiraki was amplified. CFCAT comprises an open reading frame spanning 1527 bp, encoding a 508-amino acid sequence. The highest expression was observed in the epidermal tissues (including the fat body and hemolymph) followed by the foregut/salivary gland. Furthermore, we confirmed the catalase activity of the recombinant Cfcat protein. Using RNA interference (RNAi) technology, the importance of Cfcat in the lignin-feeding of C. formosanus was demonstrated, and the role of Cfcat in innate immunity was investigated. Survival assays showed that Cfcat RNAi significantly increased the susceptibility of C. formosanus to Metarhizium anisopliae. Irrespective of the infection status, Cfcat inhibition had a significant impact on multiple factors of humoral and intestinal immunity in C. formosanus. Notably, Cfcat RNAi exhibited a more pronounced immunosuppressive effect on humoral immunity than on intestinal immunity. CONCLUSION: Cfcat plays an important role in the regulation of innate immunity and lignin feeding in C. formosanus. Cfcat RNAi can weaken the immune response of termites against M. anisopliae, which may aid the biocontrol efficiency of M. anisopliae against C. formosanus. This study provides a theoretical basis and technical reference for the development of a novel biocontrol strategy targeting detoxifying enzymes of termites. © 2024 Society of Chemical Industry.

Comparative transcriptomics reveals the conservation and divergence of reproductive genes across three sympatric Tomicus bark beetles.

Three tree-killing bark beetles belonging to the genus Tomicus, Tomicus yunnanensis, Tomicus brevipilosus and Tomicus minor (Coleoptera; Curculionidae, Scolytinae), are serious wood-borers with larvae feeding on the phloem tissues of Pinus yunnanensis. The three Tomicus beetles, in some cases, coexist in a same habitat, providing a best system for exploring the conservation and divergence of reproductive genes. Here, we applied comparative transcriptomics and molecular biology approaches to characterize reproductive-related genes in three sympatric Tomicus species. Illumina sequencing of female and male reproductive systems and residual bodies generated a large number of clean reads, representing 185,920,232 sequences in T. yunnanensis, 169,153,404 in T. brevipilosus and 178,493,176 in T. minor that were assembled into 32,802, 56,912 and 33,670 unigenes, respectively. The majority of the genes had detectable expression in reproductive tissues (FPKM >1), particularly those genes in T. brevipilosus accounting for 76.61 % of the total genes. From the transcriptomes, totally 838 genes encoding 463 detoxification enzymes, 339 chemosensory membrane proteins and 36 ionotropic glutamate receptors (iGluRs) were identified, including 622 reproductive tissue-expressed genes. Of these, members of carboxylesterases (COEs), ionotropic receptors (IRs), sensory neuron membrane proteins (SNMPs) and iGluRs were highly conserved in gene numbers and sequence identities across three Tomicus species. Further, expression profiling analyses revealed a number of genes expressed in reproductive tissues and the diverse expression characteristics in these beetles. The results provide evidence for the conservation and differences of reproductive genes among three sympatric closely related beetles, helping understand their different reproductive strategies and the maximization of the reproductive success.

Variation in the physiological response of adult worker bees of different ages (Apis mellifera L.) to pyraclostrobin stress.

The social division of labor within the honeybee colony is closely related to the age of the bees, and the age structure is essential to the development and survival of the colony. Differences in tolerance to pesticides and other external stresses among worker bees of different ages may be related to their social division of labor and corresponding physiological states. Pyraclostrobin was widely used to control the fungal diseases of nectar and pollen plants, though it was not friend to honey bees and other pollinators. This work aimed to determine the effects of field recommended concentrations of pyraclostrobin on the activities of protective and detoxifying enzymes, on the expression of genes involved in nutrient metabolism, and immune response in worker bees of different ages determined to investigate the physiological and biochemical differences in sensitivity to pyraclostrobin among different age of worker bees. The result demonstrates that the tolerance of adult worker bees to pyraclostrobin was negatively correlated with their age, and the significantly reduced survival rate of forager bees (21 day-old) with continued fungicide exposure. The activities of protective enzymes (CAT and SOD) and detoxifying enzymes (CarE, GSTs and CYP450) in different ages of adult worker bees were significantly altered, indicating the physiological response and the regulatory capacity of worker bees of different ages to fungicide stress was variation. Compared with 1 and 8 day-old worker bees, the expression of nutrient-related genes (ilp1 and ilp2) and immunity-related genes (apidaecin and defensin1) in forager bees (21 day-old) was gradually downregulated with increasing pyraclostrobin concentrations. Moreover, the expression of vitellogenin and hymenoptaecin in forager bees (21 day-old) was also decreased in high concentration treatment groups (250 and 313 mg/L). The present study confirmed the findings of the chronic toxicity of pyraclostrobin on the physiology and biochemistry of worker bees of different ages, especially to forager bees (21 day-old). These results would provide important physiological and biochemical insight for better understanding the potential risks of pyraclostrobin on honeybees and other non-target pollinators.

First report on evaluation of commercial eugenol and piperine against Aedes aegypti L (Diptera: Culicidae) larvae: Mortality, detoxifying enzyme, and histopathological changes in the midgut.

Dengue fever is a worldwide public health problem, and efforts to eradicate it have focused on controlling the dengue vector, Aedes aegypti. This study aims to assess the toxicity and effect of commercial eugenol and piperine on Ae. aegypti larvae through enzyme detoxification and histopathological changes in the midgut. Laboratory-reared Ae. aegypti larvae were treated with various concentrations of commercial eugenol and piperine and observed after 24, 48, and 72 h. Biochemical methods were used to assess detoxification enzyme activity for acetylcholinesterase, glutathione S-transferase, and oxidase, and changes in the midgut were examined using routine histological examination. In terms of larvicidal activity, piperine exceeded eugenol. Piperine and eugenol had LC50 and LC90 values of 3.057 and 5.543 µM, respectively, and 6.421 and 44.722 µM at 24 h. Piperine and eugenol reduced oxidase activity significantly (p < 0.05), but increased acetylcholinesterase and glutathione S-transferase activity significantly (p < 0.05). After being exposed to piperine and eugenol, the food bolus and peritrophic membrane ruptured, the epithelial layer was interrupted and irregular, the epithelial cells shrank and formed irregularly, and the microvilli became irregular in shape. Commercial piperine and eugenol behave as potential larvicides, with processes involving altered detoxifying enzymes, specifically decreased oxidase function and increased GST activity, as well as midgut histological abnormalities.

DIMBOA-induced gene expression, activity profiles of detoxification enzymes, multi-resistance mechanisms, and increased resistance to indoxacarb in tobacco cutworm, Spodoptera litura (Fabricius).

Spodoptera litura (Fabricius) (Lepidoptera: Noctuidae) is one of the most destructive insect pests owned strong resistance to different insecticides. Indoxacarb as a novel oxadiazine insecticide becomes the main pesticide against S. litura. DIMBOA [2,4-dihydroxy-7-methoxy-2 H-1,4-benz-oxazin-3(4 H)-one] is involved in important chemical defense processes in corn plants. However, the insects' adaptation mechanism to insecticides when exposed to defensive allelochemicals in their host plants remains unclear. Here, we assessed multi-resistance, and resistance mechanisms based on S. litura life history traits. After 18 generations of selection, indoxacarb resistance was increased by 61.95-fold (Ind-Sel) and 86.06-fold (Dim-Sel) as compared to the Lab-Sus. Also, DIMBOA-pretreated larvae developed high resistance to beta-cypermethrin, chlorpyrifos, phoxim, chlorantraniliprole, and emamectin benzoate. Meanwhile, indoxacarb (LC50) was applied to detect its impact on thirty-eight detoxification-related genes expression. The transcripts of SlituCOE073, SlituCOE009, SlituCOE074, and SlituCOE111 as well as SlGSTs5, SlGSTu1, and SlGSTe13 were considerably raised in the Ind-Sel strain. Among the twenty-three P450s, CYP6AE68, CYP321B1, CYP6B50, CYP9A39, CYP4L10, and CYP4S9v1 transcripts denoted significantly higher levels in the Ind-Sel strain, suggesting that CarEs, GSTs and P450s genes may be engaged in indoxacarb resistance. These outcomes further highlighted the importance of detoxification enzymes for S. litura gene expression and their role in responses to insecticides and pest management approaches.

Resistance risk assessment of six pyrethroids and acephate toward the resistant adult tarnished plant bug, Lygus lineolaris.

Due to rapidly developed resistance, pest management relies less on pyrethroids to control economically damaging infestations of the tarnished plant bug (TPB), Lygus lineolaris (Palisot de Beauvois) in cotton fields of Mississippi. Yet, pyrethroid resistance remains prevalent in TPB populations. This study assessed the resistance levels in adult TPB to six common pyrethroids and acephate. Resistant TBPs were collected from wild host plants in late October after harvest in the Mississippi Delta region of the United States. Based on LC50 values, the field-resistant TPBs displayed higher resistance to permethrin, esfenvalerate, and bifenthrin (approximately 30 fold) and moderate resistance to λ-cyhalothrin, ß-cyfluthrin, ζ-cypermethrin, and acephate (approximately 15 fold). Further investigations showed that the inhibitors of three detoxification enzyme, triphenyl phosphate (TPP), diethyl maleate (DEM), and piperonyl butoxide (PBO) had synergistic effects on permethrin, λ-cyhalothrin, and bifenthrin in resistant TPBs. Furthermore, elevated esterase, GST, and P450 activities were significantly expressed in field-resistant TPBs. Additionally, GST and esterase were reduced after 48 h exposure to certain pyrethroids at LC50 dose. The synergistic and biochemical assays consistently indicated that P450 and esterase were involved in pyrethroid detoxification in TPBs. This study provides valuable information for the continued use of pyrethroids and acephate in controlling TPBs in cotton fields in the Mississippi Delta region of the United States.

Environmental impacts of industrial activities on floral coverage with special emphasis on detoxification enzyme activities in Cataglyphis savignyi as pollution biomarker.

The present study investigates the environmental impact of industrial activities on floral coverage within the major industrial district of Borg El-Arab City, Egypt. Additionally, it aims to evaluate the detoxification enzymatic activity of Cataglyphis savignyi as a pollution biomarker. To achieve this objective, seasonal soil samples were collected from the studied sites to determine soil properties and heavy metal concentrations. Furthermore, a seasonal specimen of C. savignyi was collected to study the enzymatic activity of acetylcholinesterase (AChE) and glutathione-S-transferase (GST). Heavy metal contamination pollution indices were calculated, and fourteen plant species were identified at the investigated sites for four successive seasons from 2020 to 2021. The soil physicochemical parameters significantly varied in the industrial sites compared to the control site. The accumulation of heavy metal contamination in soil for investigated sites followed the order Ni > As > Pb > Hg. Calculated Cdeg and PLI for industrial 3 revealed a very high degree of contamination, attributed to increased industrial activity from the chemical and silicate factories that characterize this region. The current results highlight the inhibition of GST levels in C. savignyi at the industrial site compared to the control site. In contrast, AChE increases, which might be due to heavy metals enhancing acetylcholine activity at synapses. Consequently, the antioxidant enzymatic activities are useful as biomarkers for assessing and monitoring environmental contamination. In conclusion, this study underscores insects as potent biomarkers for heavy metal contamination, marking a significant advancement in environmental monitoring. These bioindicators offer crucial insights into the impacts of climate change and industrial pollution. The research reveals distinct plant diversity variations and higher heavy metal content in industrial sites, indicating pronounced contamination. Additionally, the study highlights altered enzyme activities in insects, emphasizing their utility as biomarkers for assessing environmental contamination. This work represents a substantial leap forward in comprehending the complex dynamics between contamination and ecological balance.

Direct observation of negative cooperativity in a detoxification enzyme at the atomic level by Electron Paramagnetic Resonance spectroscopy and simulation.

The catalytic activity of human glutathione S-transferase A1-1 (hGSTA1-1), a homodimeric detoxification enzyme, is dependent on the conformational dynamics of a key C-terminal helix α9 in each monomer. However, the structural details of how the two monomers interact upon binding of substrates is not well understood and the structure of the ligand-free state of the hGSTA1-1 homodimer has not been resolved. Here, we used a combination of electron paramagnetic resonance (EPR) distance measurements and weighted ensemble (WE) simulations to characterize the conformational ensemble of the ligand-free state at the atomic level. EPR measurements reveal a broad distance distribution between a pair of Cu(II) labels in the ligand-free state that gradually shifts and narrows as a function of increasing ligand concentration. These shifts suggest changes in the relative positioning of the two α9 helices upon ligand binding. WE simulations generated unbiased pathways for the seconds-timescale transition between alternate states of the enzyme, leading to the generation of atomically detailed structures of the ligand-free state. Notably, the simulations provide direct observations of negative cooperativity between the monomers of hGSTA1-1, which involve the mutually exclusive docking of α9 in each monomer as a lid over the active site. We identify key interactions between residues that lead to this negative cooperativity. Negative cooperativity may be essential for interaction of hGSTA1-1 with a wide variety of toxic substrates and their subsequent neutralization. More broadly, this work demonstrates the power of integrating EPR distances with WE rare-events sampling strategy to gain mechanistic information on protein function at the atomic level.

Bottom-Up Effects of Drought-Stressed Cotton Plants on Performance and Feeding Behavior of Aphis gossypii.

Drought, a major stress for crop plants, is expected to increase in frequency due to climate change. Drought can alter crop growth and levels of secondary plant metabolites, which in turn can affect herbivores, but this latter point is still controversial. This study used three different polyethylene glycol (PEG-6000) levels (0%, 1%, and 3%) to simulate drought stress and evaluated their effects on cotton plants and the impacts on the performance of the cotton aphid Aphis gossypii. Cotton plants under drought stress showed decreased water content, above-ground biomass, and nitrogen content and increased soluble protein, soluble sugar, and tannin contents. Based on analysis of the developmental time and fecundity data from individuals and at the population level, a significantly lower fecundity and population abundance of A. gossypii were detected on cotton plants with drought stress, which supports the "plant vigor hypothesis". The poor development of A. gossypii is possibly related to lower xylem sap and phloem ingestion under drought stress. In addition, the increased tannin content of cotton plants induced by drought and lower detoxification enzyme activities of A. gossypii may have affected the responses of aphids to drought-stressed plants. Overall, the results showed that drought stress altered the physiological characteristics of the cotton plants, resulting in adverse bottom-up effects on cotton aphid performances. This implies that the adoption of drip irrigation under plastic film that can help alleviate drought stress may favor the population growth of cotton aphids.

Identification and functional study of detoxification-related genes in response to tolfenpyrad stress in Glyphodes pyloalis Walker (Lepidoptera: Pyralidae).

Glyphodes pyloalis Walker (G. pyloalis) is a common destructive mulberry pest. Due to the long-term and frequent use of insecticides, it has developed tolerance to commonly used insecticides. Tolfenpyrad (TFP) is a novel pyrazole heterocyclic insecticide. In order to understand the TFP detoxification mechanism of G. pyloalis larvae, we first estimated the LC30 dose of TFP for 3rd instar G. pyloalis larvae. Next, we identified genes that were differentially expressed in 3rd instar G. pyloalis larvae treated with TFP compared to the control group by transcriptome sequencing. In total, 86,949,569 and 67,442,028 clean reads were obtained from TFP-treated and control G. pyloalis larvae, respectively. A total of 5588 differentially expressed genes (DEGs) were identified in TFP-treated and control G. pyloalis larvae, of which 3084 genes were upregulated and 2504 genes were downregulated. We analyzed the expression of 43 candidate detoxification enzyme genes associated with insecticide tolerance using qPCR. According to the spatiotemporal expression pattern of DEGs, we found that CYP6ABE1, CYP333A36 and GST-epsilon8 were highly expressed in the midgut, while CarEs14 was strongly expressed in haemolymph. Furthermore, we successfully knocked down these genes by RNA interference. After silencing CYP6ABE1 and CYP333A36, bioassay showed that the mortality rate of TFP-treated G. pyloalis larvae was significantly higher compared to the control group. This study provides a theoretical foundation for understanding the sensitivity of G. pyloalis to TFP and establish the basis for the effective and green management of this pest.

Risk Assessment of Fluxametamide Resistance and Fitness Costs in Fall Armyworm (Spodoptera frugiperda).

The fall armyworm (FAW), Spodoptera frugiperda, is one of the most devastating invasive polyphagous pests, which has attracted recent global attention by developing resistance to various insecticidal active ingredients with independent mode of action. Fluxametamide, a newly commercialized isoxazoline insecticide, is exceptionally selective towards several lepidopteran pests. The present study aimed to evaluate resistance risk in FAW to fluxametamide and the fitness costs associated with fluxametamide resistance. A field-collected and genetically mixed population of FAW was artificially selected through continuous exposure to fluxametamide. After successive selection of 10 generations, there was no obvious increase in the LC50 (RF: 2.63-fold). The realized heritability (h2) of fluxametamide resistance was estimated as h2 = 0.084 using a quantitative genetic approach. Compared with the susceptible F0 strain, the Flux-SEL (F10) strain of FAW displayed no significant cross-resistance to broflanilide, chlorantraniliprole, fipronil, indoxacarb, lambda cyhalothrin, spinetoram, and tetraniliprole, except emamectin benzoate (RF: 2.08-fold). Increased activity of glutathione S-transferase (ratio 1.94) was observed in the Flux-SEL (F10) strain of FAW, while the cytochrome P450 and carboxylesterase activities were not altered. The fluxametamide-selection significantly affected the development and reproductive traits of FAW with a lower R0, T and relative fitness (Rf = 0.353). The results alluded that the risk of fluxametamide resistance evolution in FAW is relatively lower; however, proactive implementation of resistance management approaches should be done to maintain the field efficacy of fluxametamide against FAW.

Metamifop resistance in Echinochloa glabrescens via glutathione S-transferases-involved enhanced metabolism.

BACKGROUND: Echinochloa glabrescens Munro ex Hook. f. is one of the main Echinochloa spp. seriously invading Chinese rice fields and has evolved resistance to commonly used herbicides. Previously, an E. glabrescens population (LJ-02) with suspected resistance to the acetyl-CoA carboxylase (ACCase)-inhibiting herbicide metamifop was collected. This study aimed to determine its resistance status to metamifop and investigate the internal molecular mechanisms of resistance. RESULTS: Single-dose testing confirmed that the LJ-02 population had evolved resistance to metamifop. Gene sequencing and a relative expression assay of ACCase ruled out target-site based resistance to metamifop in LJ-02. Whole-plant bioassays revealed that, compared with the susceptible population XZ-01, LJ-02 was highly resistant to metamifop and exhibited cross-resistance to fenoxaprop-P-ethyl. Pretreatment with the known glutathione S-transferase (GST) inhibitor, 4-chloro-7-nitrobenzoxadiazole (NBD-Cl), largely reversed the resistance to metamifop by approximately 81%. Liquid chromatography-tandem mass spectrometry analysis indicated that the metabolic rates of one of the major metabolites of metamifop, N-(2-fluorophenyl)-2-hydroxy-N-methylpropionamide (HPFMA), were up to 383-fold faster in LJ-02 plants than in XZ-01 plants. There were higher basal and metamifop-inducible GST activities toward 1-chloro-2,4-dinitrobenzene (CDNB) in LJ-02 than in XZ-01. Six GST genes were metamifop-induced and overexpressed in the resistant LJ-02 population. CONCLUSION: This study reports, for the first time, the occurrence of metabolic metamifop resistance in E. glabrescens worldwide. The high-level metamifop resistance in the LJ-02 population may mainly involve specific isoforms of GSTs that endow high catalytic activity and strong substrate specificity. © 2023 Society of Chemical Industry.

Toxicity and Physiological Effects of Nine Lamiaceae Essential Oils and Their Major Compounds on Reticulitermes dabieshanensis.

The volatile metabolites of Salvia sclarea, Rosmarinus officinalis, Thymus serpyllum, Mentha spicata, Melissa officinalis, Origanum majorana, Mentha piperita, Ocimum basilicum and Lavandula angustifolia were determined by gas chromatography-mass spectrometry. The vapor insecticidal properties of the analyzed essential oils and their compounds were screened using Reticulitermes dabieshanensis workers. The most effective oils were S. sclarea (major constituent linalyl acetate, 65.93%), R. officinalis (1,8-cineole, 45.56%), T. serpyllum (thymol, 33.59%), M. spicata (carvone, 58.68%), M. officinalis (citronellal, 36.99%), O. majorana (1,8-cineole, 62.29%), M. piperita (menthol, 46.04%), O. basilicum (eugenol, 71.08%) and L. angustifolia (linalool, 39.58%), which exhibited LC50 values ranging from 0.036 to 1.670 µL/L. The lowest LC50 values were recorded for eugenol (0.060 µL/L), followed by thymol (0.062 µL/L), carvone (0.074 µL/L), menthol (0.242 µL/L), linalool (0.250 µL/L), citronellal (0.330 µL/L), linalyl acetate (0.712 µL/L) and 1,8-cineole (1.478 µL/L). The increased activity of esterases (ESTs) and glutathione S-transferase (GST) were observed but only alongside the decreased activity of acetylcholinesterase (AChE) in eight main components. Our results indicate that S. sclarea, R. officinalis, T. serpyllum, M. spicata, M. officinalis, O. marjorana, M. piperita, O. basilicum and L. angustifolia essential oils (EOs) and their compounds, linalyl acetate, 1,8-cineole, thymol, carvone, citronellal, menthol, eugenol and linalool could be developed as control agents against termites.

The dynamic changes of genes revealed that persistently overexpressed genes drive the evolution of cyflumetofen resistance in Tetranychus cinnabarinus.

Changes in gene expression are associated with the evolution of pesticide resistance in arthropods. In this study, transcriptome sequencing was performed in 3 different resistance levels (low, L; medium, M; and high, H) of cyflumetofen-resistant strain (YN-CyR). A total of 1 685 genes, including 97 detoxification enzyme genes, were upregulated in all 3 stages, of which 192 genes, including 11 detoxification enzyme genes, showed a continuous increase in expression level with resistance development (L to H). RNA interference experiments showed that overexpression of 7 genes (CYP392A1, TcGSTd05, CCE06, CYP389A1, TcGSTz01, CCE59, and CYP389C2) is involved in the development of cyflumetofen resistance in Tetranychus cinnabarinus. The recombinant CYP392A1 can effectively metabolize cyflumetofen, while CCE06 can bind and sequester cyflumetofen in vitro. We compared 2 methods for rapid screening of resistance molecular markers, including short-term induction and 1-time high-dose selection. Two detoxification enzyme genes were upregulated in the field susceptible strain (YN-S) by induction with 20% lethal concentration (LC20 ) of cyflumetofen. However, 16 detoxification enzyme genes were upregulated by 1-time selection with LC80 of cyflumetofen. Interestingly, the 16 genes were overexpressed in all 3 resistance stages. These results indicated that 1 685 genes that were upregulated at the L stage constituted the basis of cyflumetofen resistance, of which 192 genes in which upregulation continued to increase were the main driving force for the development of resistance. Moreover, the 1-time high-dose selection is an efficient way to rapidly obtain the resistance-related genes that can aid in the development of resistance markers and resistance management in mites.