Functional analysis of SDR112C1 associated with fenpropathrin tolerance in Tetranychus cinnabarinus (Boisduval).

Short-chain dehydrogenases/reductases (SDRs) are ubiquitously distributed across diverse organisms and play pivotal roles in the growth, as well as endogenous and exogenous metabolism of various substances, including drugs. The expression levels of SDR genes are reportedly upregulated in the fenpropathrin (FEN)-resistant (FeR) strain of Tetranychus cinnabarinus. However, the functions of these SDR genes in acaricide tolerance remain elusive. In this study, the activity of SDRs was found to be significantly higher (2.26-fold) in the FeR strain compared to the susceptible strain (SS) of T. cinnabarinus. A specific upregulated SDR gene, named SDR112C1, exhibited significant overexpression (3.13-fold) in the FeR population compared with that in the SS population. Furthermore, the expression of SDR112C1 showed a significant increase in the response to FEN induction. Additionally, knockdown of the SDR112C1 gene resulted in decreased SDR activity and reduced mite viability against FEN. Importantly, heterologous expression and in vitro incubation assays confirmed that recombinant SDR112C1 could effectively deplete FEN. Moreover, the overexpression of the SDR112C1 gene in Drosophila melanogaster significantly decreased the toxicity of FEN to transgenic fruit flies. These findings suggest that the overexpression of SDR SDR112C1 is a crucial factor contributing to FEN tolerance in T. cinnabarinus. This discovery not only enhances our understanding of SDR-mediated acaricide tolerance but also introduces a new family of detoxification enzymes to consider in practice, beyond cytochrome P450s, carboxyl/choline esterases and glutathione S-transferases.

A novel spider venom peptide from the predatory mite Neoseiulus barkeri shows lethal effect on phytophagous pests.

The long-term use of pesticides in the field, and the high fertility and adaptability of phytophagous mites have led to resistance problems; consequently, novel safe and efficient active substances are necessary to broaden the tools of pest mite control. Natural enemies of arthropods typically secrete substances with paralytic or lethal effects on their prey, and those substances are a resource for future biopesticides. In this study, two putative venom peptide genes were identified in a parasitic mite Neoseiulus barkeri transcriptome. Recombinant venom NbSP2 peptide injected into Tetranychus cinnabarinus mites was significantly more lethal than recombinant NBSP1. NbSP2 was also lethal to Spodoptera litura when injected but not when fed to third instar larvae. The interaction proteins of NbSP2 in T. cinnabarinus and S. litura were identified by affinity chromatography. Among these proteins, ATP synthase subunit ß (ATP SSß) was deduced as a potential target. Four binding sites were predicted between NBSP2 and ATP SSß of T. cinnabarinus and S. litura. In conclusion, we identified a venom peptide with activity against T. cinnabarinus and S. litura. This study provides a novel component for development of a new biological pesticide.

Uridine Diphosphate Glycosyltransferases (UGTs) Involved in the Carotenoid-Based Body Color Difference between Tetranychus cinnabarinus (Red) and Tetranychus urticae (Green).

It has long been disputed whether Tetranychus cinnabarinus and Tetranychus urticae belong to the same genus, with T. cinnabarinus regarded as a red form of T. urticae. However, it is unclear why T. urticae and T. cinnabarinus have different body colors. Since carotenoids are responsible for the color of many organisms, the carotenoid profiles of T. cinnabarinus and T. urticae were compared by HPLC. There was no difference in carotenoid type, but T. cinnabarinus contained significantly more neoxanthin, astaxanthin, α-carotene, ß-carotene, and γ-carotene, which may contribute to the deep red color. The transcriptome sequencing of both species identified 4079 differentially expressed genes (DEGs), of which 12 were related to carotenoid metabolism. RNA interference (RNAi) experiments demonstrated that silencing seven of these DEGs resulted in the different accumulation of carotenoid compounds in T. cinnabarinus and T. urticae. In addition, the body of T. urticae turned yellow after two days of feeding with UGT double-stranded RNAs and ß-UGT small interfering RNAs. In conclusion, differences in the carotenoid profiles of T. urticae and T. cinnabarinus may be responsible for the different body colors.

Research Progress of a Pesticide Polymer-Controlled Release System Based on Polysaccharides.

In recent years, with the development of the nanomaterials discipline, many new pesticide drug-carrying systems-such as pesticide nano-metal particles, nano-metal oxides, and other drug-carrying materials-had been developed and applied to pesticide formulations. Although these new drug-loading systems are relatively friendly to the environment, the direct exposure of many metal nanoparticles to the environment will inevitably lead to potential effects. In response to these problems, organic nanomaterials have been rapidly developed due to their high-quality biodegradation and biocompatibility. Most of these organic nanomaterials were mainly polysaccharide materials, such as chitosan, carboxymethyl chitosan, sodium alginate, ß-cyclodextrin, cellulose, starch, guar gum, etc. Some of these materials could be used to carry inorganic materials to develop a temperature- or pH-sensitive pesticide drug delivery system. Herein, the pesticide drug-carrying system developed based on polysaccharide materials, such as chitosan, was referred to as the pesticide polymer drug-carrying system based on polysaccharide materials. This kind of drug-loading system could be used to protect the pesticide molecules from harsh environments, such as pH, light, temperature, etc., and was used to develop the function of a sustained release, targeted release of pesticides in the intestine of insects, and achieve the goal of precise application, reduction, and efficiency of pesticides. In this review, the recent progress in the field of polysaccharide-based polymer drug delivery systems for pesticides has been discussed, and suggestions for future development were proposed based on the current situation.

Population dynamics and molecular adaption of Tetranychus cinnabarinus to long-term thermal stress.

BACKGROUND: Global warming is a general trend in the current era. Temperature is one of the most important nonbiological factors that affects the development, life cycle and distribution of arthropods, which are a major component of agriculture pests. This study focused on life-table parameters and the molecular adaption of Tetranychus cinnabarinus under long-term thermal stress. RESULTS: The life tables of T. cinnabarinus were constructed at room temperature (26 °C) and high temperature (34 °C). Results showed that although the lifespan of the mites was shortened, the developmental periods of egg, larva and nymph stages were accelerated, and the peak egg-laying period came earlier at high temperature, which resulted in faster expansion of pest mite population. RNA-seq was used to reveal the thermal adaption mechanism according to differentially expressed genes. Combined with transcriptome data and quantitative polymerase chain reaction (qPCR) verification, MAPK, CAT, HSP20 and HSP70 were found highly expressed at 34 °C, which were associated with thermal adaption of T. cinnabarinus. RNAi analysis proved that expression of HSP20 was closely related to the survival of mites at high temperature. CONCLUSION: These results indicated that long-term high temperature treatment was beneficial to the expansion of the T. cinnabarinus population. The genes involved in heat tolerance of T. cinnabarinus such as MAPK-HSP pathway provides ideas for subsequent control measures. © 2023 Society of Chemical Industry.

Retinoid X receptor 1 is a specific lethal RNAi target disturbing chitin metabolism during hatching of Tetranychus cinnabarinus.

RNA interference (RNAi) can be developed as an alternative method of chemical pesticides for pest control. In this study, we noticed a specifically expressed gene (retinoid X receptor 1, TcRXR1) in the egg stage of T. cinnabarinus. RNAi was applied to investigate the function of TcRXR1. Results showed that with continuous feeding of dsTcRXR1, the larvae of T. cinnabarinus could still successfully develop to adult, which was in accordance with the low expression of TcRXR1 out of egg stage. High mortality of eggs was observed after eggs were treated with dsTcRXR1. To investigate the downstream genes of TcRXR1, the RNA samples after successful RNAi of TcRXR1 were analyzed by transcriptome analysis. According to function annotation of differentially expressed genes, 6 genes were selected for their potential function with the phenotype of dsTcRXR1, and among them, a chitinase gene (TcCHT-E) attained a high expression level in the late stage of egg, peaking just after the expression peak of TcRXR1. Mortality of eggs was observed under the effect of dsTcCHT-E as well as dsTcRXR1. In conclusion, TcRXR1 is a specific RNAi target for control of T. cinnabarinus, and its lethal mechanism might be disturbing chitin metabolism hatching of egg.

The mechanism of coumarin inhibits germination of ryegrass (Lolium perenne) and its application as coumarin-carbon dots nanocomposites.

BACKGROUND: As an important plant allelochemical, coumarin can effectively inhibit the germination of various seeds. However, little is known about the inhibition mechanism of coumarin on weed seed germination. Moreover, the herbicidal activity of coumarin is needed to be improved as a natural pesticide. RESULTS: Coumarin had the highest inhibition effect on the ryegrass (Lolium perenne) seed, where coumarin disturbed the hormone pathway by decreasing the content of gibberellic acid 3, resulting in the reduction of amylase activity and consumption of starch during the germination process of ryegrass seed. Moreover, coumarin induced decreased activity of catalase and subsequently led to the accumulation of hydrogen peroxide and malondialdehyde, causing oxidative stress during the germination process of ryegrass seed. Furthermore, to enhance the herbicidal activity of coumarin, carbon dots (CDs) modified with polyetherimide were prepared, characterized, and then combined with coumarin to form coumarin-carbon dots (Cm-CDs) nanocomposites. Compared with coumarin, Cm-CDs nanocomposites significantly increased the herbicidal activity of coumarin on ryegrass, which implies that Cm-CDs nanocomposites could be used as a potential formulation to improve the herbicidal activity of coumarin. CONCLUSION: This study not only reveals the mechanism of coumarin on ryegrass germination, but also develop Cm-CDs nanocomposites to enhance the herbicidal activity of coumarin. Our findings will stimulate the application of Cm-CDs nanomaterials as an effective and environmentally friendly formulation in agriculture. © 2023 Society of Chemical Industry.

Molecular Basis for the Selectivity of the Succinate Dehydrogenase Inhibitor Cyflumetofen between Pest and Predatory Mites.

Acaricides that act as inhibitors of the mitochondrial succinate dehydrogenase (SDHIs) provide excellent control of phytophagous mites but display limited toxicity to predatory mites and other beneficial organisms. However, the molecular mechanism of selectivity is not fully understood. Here, we first confirm that SDHI acaricides are over 10,000-fold more toxic to spider mites than predatory mites. Next, we show that differential penetration, pro-acaricide activation, or metabolism are most likely not the main reason for this selectivity. In contrast, the inhibition of AB-1 on the SDH target is approximately 200-fold more potent in spider mites compared to predatory mites, revealing strong target-site selectivity. Strikingly, a key motif associated with differential binding was identified and validated by gene editing in Drosophila. Our findings contribute to understanding the selectivity of SDHIs, which can be used for the rational design of selective acaricides in support of an integrated pest management.

A pH- and enzymatic-responsive nanopesticide to control pea aphids and reduce toxicity for earthworms.

Thiacloprid is a new chlorinated nicotinoid insecticide against stinging-oral pests, such as aphids. It is less toxic to bees but more toxic to earthworms. In this study, a pH- and amylase-responsive MOF (ZIF-8) was constructed for site-specific delivery of thiacloprid to control pea aphids and more safety for earthworms. Thiacloprid from α-cyclodextrin@Thiacloprid@ZIF-8 (α-CD@T@ZIF-8) could be released quickly in pea aphids, which was ascribed to disintegration of ZIF-8 at low pH values in pea aphid intestines and degradation of α-CD under the action of α-amylase. The release results showed a significant pH dependence of α-CD@T@ZIF-8, with an approximately 65 % release amount at pH = 7 and a 95 % release amount at pH = 5 for 7 d. The results of the pot experiment and biosafety showed that for α-CD@T@ZIF-8, 88 % pea aphids could be killed compared with 32 % aphids for commercially available formulation on the 7th day after application. Meanwhile the LC50 of thiacloprid OD was 0.034 µg/cm2 and the LC50 of α-CD@T@ZIF-8 was 0.564 µg/cm2 on earthworms, and it was more safety for pea and lower acute toxicity and enrichment for the earthworms. α-CD@T@ZIF-8 could be used for intelligently controlled release of other insecticides against aphids.

Preparation of Berberine@carbon Dots Nano-Formulation: Synthesis, Characterization and Herbicidal Activity against Echinochloa crus-galli and Amaranthus retroflexus Two Common Species of Weed.

Berberine (Ber) is easy to synthesize and has a variety of biological and pharmacological activities. At present, the existing studies on berberine have focused predominantly on its antibacterial activity; its herbicidal activity is rarely reported. In addition, there are a number of preparations of berberine, which are not enough to solve its shortcomings of low solubility and biological activity and the difficult storage of berberine. Here, berberine was combined with carbon dots to obtain carbon dots-berberine (CDs-Ber) nano formulation. The fluorescence quenching results showed that the CDs-Ber nano drug delivery system was successfully constructed, and the fluorescence quenching mechanism of the two was static quenching. The bioassay results showed that CDs had no adverse effects on the growth of barnyard grass (Echinochloa crus-galli) and redroot pigweed (Amaranthus retroflexus), and had high biocompatibility. Berberine and CDs-Ber predominantly affected the root growth of barnyard grass and redroot pigweed and could enhance the growth inhibition effect on weeds, to some extent. The results of the protective enzyme system showed that both berberine and CDs-Ber could increase the activities of Superoxide dismutase (SOD), Peroxidase (POD), and Catalase (CAT) in barnyard grass, and CDs-Ber had a stronger stress effect on barnyard grass than berberine. The determination of the number of bacterial communities in the soil after the berberine and CDs-Ber treatments showed that there was no significant difference in the effects of the two, indicating that CDs-Ber would not have more negative impacts on the environment. The CDs-Ber nano formulation improved the biological activity of berberine, enhanced the herbicidal effect, and was relatively safe for soil colonies.

Preparation and performance characteristics of spider venom peptide nanocapsules.

BACKGROUND: ω-hexatoxin-Hvn1b is an insecticidal toxin produced by the Tasmanian funnel-web spider (Hadronyche venenata), that can be exploited for development of novel bioinsecticides. Due to its larger size and low membrane permeability, this toxin usually has a slower mode of action compared to conventional small molecule insecticides. Nanoscale materials have unique optical, electrical, mechanical and biological properties, and show great application prospects for pesticide delivery. RESULTS: The physical and chemical properties of nanocapsules were characterized using transmission electron microscopy, laser particle size analysis, Fourier transform infrared spectroscopy, contact angle testing and with a fluorescence spectrophotometer. The results indicated that the nanocapsules were spherical, with an average particle size of 197.70 nm, the encapsulation efficiency rate was 75.82% and the Zeta potential was -32.90 mV. Penetration experiments showed that the nanocapsules could promote protein passage through the intestinal tract of Spodoptera litura and reach the body fluid. Then we expressed ω-hexatoxin-Hvn1b by prokaryotic expression. Bioassay results showed that the oral toxicity of ω-hexatoxin-Hvn1b nanocapsules to S. litura was higher than that of the ω-hexatoxin-Hvn1b. CONCLUSION: In this paper, we reported a construction method of spider venom peptide nanocapsules based on polylactic-co-glycolic acid by multiple emulsion for delivery of protein to improve the insecticidal effect and oral activity of ω-hexatoxin-Hv1a. © 2022 Society of Chemical Industry.

Transcription factors CncC and Maf connect the molecular network between pesticide resistance and resurgence of pest mites.

Pesticide resistance and resurgence are serious problems often occurring simultaneously in the field. In our long-term study of a fenpropathrin-resistant strain of Tetranychus cinnabaribus, enhancement of detoxification and modified fecundity mechanisms were both observed. Here we investigate the network across these two mechanisms and find a key node between resistance and resurgence. We show that the ecdysone pathway is involved in regulating the fecundity of T. cinnabaribus. The concentration change of ecdysone is consistent with the fecundity curve; the concentration of ecdysone is higher in the fenpropathrin-resistant strain which has stronger fecundity. The enhancement of ecdysone is due to overexpression of two P450 genes (CYP314A1 and CYP315A1) in the ecdysone synthesis pathway. Silencing expression of these CYP genes resulted in lower concentration of ecdysone, reduced expression of vitellogenin, and reduced fecundity of T. cinnabaribus. The expression of CYP315A1 is regulated by transcription factors Cap-n-collar isoform C (CncC) and Musculoaponeurotic fibrosarcoma protein (Maf), which are involved in regulating other P450 genes functioning in detoxification of fenpropathrin in T. cinnabaribus. A similar regulation is established in citrus pest mite Panonychus citri showing that the CncC pathway regulates expression of PcCYP315A1, which affects mite fecundity. Transcription factors are activated to upregulate detoxification genes facilitating pesticide resistance, while the "one to multiple" regulation mode of transcription factors simultaneously increases expression of metabolic enzyme genes in hormone pathways and alters the physiology of pests. This is an important response of arthropods to pesticides which leads to resistance and population resurgence.

Sodium selenite-carbon dots nanocomposites enhance acaricidal activity of fenpropathrin: Mechanism and application.

As an essential trace element, selenium can be used to protect crops from pests, while, in nature, most crops cannot accumulate enough selenium from the soil to reach the effective dose for pest control. In this study, carbon dots modified with arginine in nano-scale was prepared and characterized, then, it was combined with sodium selenite to form selenium-carbon dots (Se-CDs). Function evaluation of Se-CDs showed that it could increase the absorption of selenium in plant leaves, promote the control efficiency of fenpropathrin, and protect plant from damage caused by Tetranychus cinnabarinus. In addition, we found that expressions of P450 genes and activity of P450 enzyme both decreased in selenium treated mites. In vivo, the acaricidal activity of fenpropathrin increased significantly when one of the P450 genes, CYP389B1, was silenced, and the recombinant protein of CYP389B1 could metabolize fenpropathrin in vitro. The results suggested that inhibiting the expression of P450 gene and repressing the detoxification of T. cinnabarinus was the molecular mechanism that how selenium promoted the acaricidal activity of fenpropathrin. The application of Se-CDs in the field will decrease the use of chemicals acaricides, reduce chemical residues, and ensure the safety of agricultural products.

Selenium mediated host plant-mite conflict: defense and adaptation.

BACKGROUND: Selenium has shown effectiveness in protecting plants from herbivores. However, some insects have evolved adaptability to selenium. RESULTS: Selenium accumulation in host plants protected them against spider mite feeding. Selenium showed toxic effects on spider mites by reducing growth and interfering with reproduction. After 40 generations on selenium-rich plants, a Tetranychus cinnabarinus strain (Tc-Se) developed adaptability to selenium, with an increased rate of population growth and enhanced ability for selenium metabolism. The high expression of two genes (GSTd07 and SPS1) in the selenium metabolism pathway might be involved in selenium metabolism in spider mites. After GSTd07 and SPS1 were silenced, the selenium adaptability decreased. Recombinant GSTd07 protein promoted the reaction between sodium selenite and glutathione (GSH) and increased the production of sodium selenite metabolites. The results indicated that GSTd07 was involved in the first step of selenium metabolism. CONCLUSION: Plants can resist spider mite feeding by accumulating selenium. Spider mites subjected to long-term selenium exposure can adapt to selenium by increasing the expression of key genes involved in selenium metabolism. These results elucidate the mechanism of the interaction between mites and host plants mediated by selenium. This study of the interaction between selenium-mediated host plants and spider mites may lead to the development of new and less toxic methods for the prevention and control of spider mites. © 2021 Society of Chemical Industry.

Identification of Lysinibacillus sphaericus Binary toxin binding proteins in a malarial mosquito cell line by proteomics: A novel approach towards improving mosquito control.

Bacterial insecticidal proteins, such as the Bin toxin from Lysinibacillus sphaericus, could be used more extensively to control insecticide resistant mosquitoes. This study was aimed at identification of mosquito cell proteins binding Bin toxin. Results showed that purified toxin was toxic to Anopheles gambiae larvae and Ag55 cultured cells. Clathrin heavy chain (an endocytosis protein) and glycolytic enzymes such as pyruvate kinase, enolase and dihydrolipoamide dehydrogenase were identified as binders of Bin toxin. The viability of Ag55 cells in the presence of endocytosis inhibitor, pitstop2, was significantly decreased upon Bin treatment, while the inhibitor chlorpromazine did not affect Bin toxicity. Bin toxin treatment decreased ATP production and mitochondrial respiration in Ag55 cells, whereas non-mitochondrial oxygen consumption significantly increased after Bin toxin treatment. These findings are steps towards understanding how Bin toxin kills mosquitoes. SIGNIFICANCE: Mosquitoes are vectors of pathogens causing human diseases such as dengue fever, yellow fever, zika virus and malaria. An insecticidal toxin from Lysinibacillus sphaericus called Binary, or Bin, toxin could be used more extensively to control insecticide resistant mosquitoes. Bin toxin enter cells in susceptible mosquitoes and induces apoptosis or autophagy. In the current research, we used the malaria mosquito Anopheles gambiae Ag55 cell line as a model. A proteomic-based approach identified proteins that interact with Bin toxin. Interacting proteins include clathrin heavy chain (endocytosis protein) and glycolysis enzymes such as pyruvate kinase, enolase and dihydrolipoamide dehydrogenase. In Ag55 cell toxicity assays, an endocytosis inhibitor, pitstop2, increased Bin toxicity. Real time assays with a Seahorse™ flux analyzer showed that Bin significantly affects mitochondrial respiration, a result consistent with cell death via apoptosis or autophagy. These research findings add insights into how an unusual binary protein exploits cellular machinery to kill mosquitoes.

lincRNA_Tc13743.2-miR-133-5p-TcGSTm02 regulation pathway mediates cyflumetofen resistance in Tetranychus cinnabarinus.

Differential expression of metabolic detoxification enzymes is an important mechanism involved in pesticide/acaricide resistance of mite pests. The competing endogenous RNA hypothesis offers a new opportunity to investigate post-transcriptional regulation of those genes. In this study, 4454 long non-coding RNAs were identified in the carmine spider mite Tetranychus cinnabarinus by transcriptome sequencing. Software-based predictions indicated that a long intergenic non-coding RNA, (lincRNA)_Tc13743.2 and a detoxification enzyme gene, TcGSTm02, both contained a microRNA (miR-133-5p) response element. Over-expression of lincRNA_Tc13743.2 and TcGSTm02 were detected in a cyflumetofen-resistant T. cinnabarinus strain (CyR), whereas down-regulation of miR-133-5p was observed in this strain. Conversely, up-regulation of miR-133-5p could inhibit TcGSTm02 expression levels, and both lincRNA_Tc13743.2 and TcGSTm02 were significantly enriched in miR-133-5p biotin-avidin pull-down assays. RNA-binding protein immunoprecipitation assay showed that lincRNA_Tc13743.2 and TcGSTm02 bound to a silencing complex containing miR-133-5p. Moreover, a luciferase reporter assay based on a human cell line revealed that over-expression of lincRNA_Tc13743.2 could significantly reduce the inhibition exerted by miR-133-5p through the TcGSTm02 3'UTR. In addition, co-localization of lincRNA_Tc13743.2 and miR-133-5p was detected using fluorescence in situ hybridization, suggesting that lincRNA_Tc13743.2 interacts directly with miR-133-5p in spider mites. More importantly, silencing the expression of lincRNA_Tc13743.2 significantly reduced the expression levels of TcGSTm02 and increased the sensitivity of spider mites to cyflumetofen. Our data show that lincRNA_Tc13743.2 up-regulates TcGSTm02 expression by competing for miR-133-5p binding, demonstrating that a lincRNA_Tc13743.2-miR-133-5p-TcGSTm02 pathway mediates cyflumetofen resistance in mites.

Amidase, a novel detoxifying enzyme, is involved in cyflumetofen resistance in Tetranychus cinnabarinus (Boisduval).

Amidase is an important hydrolytic enzyme in detoxification metabolism. Amidase hydrolyzes a wide variety of nonpeptide carbon­nitrogen bonds by attacking a cyano group or carbonyl carbon. However, little is known about the relationship between amidase and insecticides. In this study, the amidase activity was significantly higher in cyflumetofen-resistant strain (CyR) than in the susceptible strain (SS) of Tetranychus cinnabarinus, and diethyl-phosphoramidate (an amidase inhibitor) significantly decreased cyflumetofen resistance in T. cinnabarinus. More importantly, an amidase gene, TcAmidase01, was identified in T. cinnabarinus, and the TcAmidase01 overexpression was detected in both two cyflumetofen-resistant strains (CyR and YN-CyR), indicating that it is involved in cyflumetofen resistance in mites. A phylogenetic analysis showed that TcAmidase01 was clustered with deaminated glutathione amidases, which possess hydrolytic activity. The recombinant TcAmidase01 protein showed amidase activity toward succinamate, and the activity could be inhibited by cyflumetofen. High-performance liquid chromatography-mass spectrometry (HPLC-MS) analysis provided evidence that recombinant TcAmidase01 could decompose cyflumetofen by hydrolysis, and the potential metabolites (2-(4-(tert-butyl) phenyl)-2-cyanoacetate and 2-(trifluoromethyl) benzoic acid) were identified. These results show that TcAmidase01 contribute to cyflumetofen-resistance in T. cinnabarinus by hydrolyzing cyflumetofen, and this is the first study to suggest that amidase has a role in insecticides resistance in arthropods.

The cytochrome P450 CYP389C16 contributes to the cross-resistance between cyflumetofen and pyridaben in Tetranychus cinnabarinus (Boisduval).

BACKGROUND: Acaricide resistance is a serious problem in spider mites. Cyflumetofen is a new complex II inhibitor, whereas pyridaben acts at complex I and has been used for decades. Although cross-resistance between cyflumetofen and pyridaben has been observed in Tetranychus cinnabarinus, the specific mechanisms at play have not yet been investigated. RESULTS: Investigation into the cross-resistance mechanisms identified five P450s, among which CYP389C16 was evaluated as the most likely candidate conferring cross-resistance. Knockdown of CYP389C16 expression via RNA interference diminished the level of cross-resistance in the cyflumetofen-resistant strain. In addition, recombinant CYP389C16 (40 pmol) effectively metabolized 25.0 ± 0.7% of cyflumetofen, 39.7 ± 1.0% of pyridaben, and 69.3 ± 3.3% of AB-1 (active de-esterified metabolite of cyflumetofen) within 2 h. In addition, hydroxylation metabolite of AB-1 was identified by HPLC-MS/MS. CONCLUSIONS: The study reveals that overexpressed CYP389C16 is involved in the cross-resistance between cyflumetofen and pyridaben in T. cinnabarinus. © 2019 Society of Chemical Industry.

Functional analysis of UGT201D3 associated with abamectin resistance in Tetranychus cinnabarinus (Boisduval).

Uridine diphosphate (UDP)-glycosyltransferases (UGTs) are widely distributed within living organisms and share roles in biotransformation of various lipophilic endo- and xenobiotics with activated UDP sugars. In this study, it was found that the activity of UGTs in abamectin-resistant (AbR) strain was significantly higher (2.35-fold) than that in susceptible strain (SS) of Tetranychus cinnabarinus. Further analysis showed that 5-nitrouracil, the inhibitor of UGTs, could enhance the lethal effect of abamectin on mites. From the previous microarray results, we found an UGT gene (UGT201D3) overexpressed in AbR strain. Quantitative PCR analysis showed that UGT201D3 was highly expressed and more inducible with abamectin exposure in the AbR strain. After silencing the transcription of UGT201D3, the activity of UGTs was decreased and the susceptibility to abamectin was increased in AbR strain whereas it was not in SS. Furthermore, UGT201D3 gene was then successfully expressed in Escherichia coli. The recombinant UGT201D3 exhibited α-naphthol activity (2.81 ± 0.43 nmol/mg protein/min), and the enzyme activity could be inhibited by abamectin (inhibitory concentration at 50%: 57.50 ± 3.54 µmol/L). High-performance liquid chromatography analysis demonstrated that the recombinant UGT201D3 could effectively deplete abamectin (15.77% ± 3.72%) incubating with 150 µg protein for 6 h. These results provided direct evidence that UGT201D3 was involved in abamectin resistance in T. cinnabarinus.

The Transcription Factor MafB Regulates the Susceptibility of Bactrocera dorsalis to Abamectin via GSTz2.

Pesticide resistance is a serious problem that poses a major challenge to pest control. One of the most potent resistance mechanisms is the overexpression of genes coding for detoxification enzymes. The expression of detoxification genes is regulated by a series of transcription factors. Previous studies have revealed that the increased expression of detoxification genes contributes to the insecticide tolerance of Bactrocera dorsalis. Our objective was thus to identify the transcription factors involved in this process. Temporal expression profiles showed that the transcription factor MafB and detoxification genes were expressed highly in the fat body. Further analysis showed that the expression of MafB, GSTz2, and CYP473A3 was induced by abamectin. Disruption of the MafB transcription factor through RNA interference decreased the transcript levels of GSTz2 and CYP473A3 and increased the susceptibility to abamectin significantly. Direct silencing of the expression of GSTz2 also increased susceptibility to abamectin, while CYP473A3 did not. In conclusion, these results suggest that the expression of GSTz2 and CYP473A3 was regulated by the transcription factor MafB, and the up-regulation of GSTz2 via MafB decreased the susceptibility of B. dorsalis to abamectin.